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WO2014194242A2 - Flt3 inhibitors and uses thereof - Google Patents

Flt3 inhibitors and uses thereof Download PDF

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Publication number
WO2014194242A2
WO2014194242A2 PCT/US2014/040317 US2014040317W WO2014194242A2 WO 2014194242 A2 WO2014194242 A2 WO 2014194242A2 US 2014040317 W US2014040317 W US 2014040317W WO 2014194242 A2 WO2014194242 A2 WO 2014194242A2
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WO
WIPO (PCT)
Prior art keywords
ring
compound
nitrogen
sulfur
oxygen
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PCT/US2014/040317
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French (fr)
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WO2014194242A3 (en
Inventor
Divya Chaudhary
Rosana Kapeller-Libermann
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Nimbus Iris, Inc.
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Publication of WO2014194242A2 publication Critical patent/WO2014194242A2/en
Publication of WO2014194242A3 publication Critical patent/WO2014194242A3/en

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    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K31/00—Medicinal preparations containing organic active ingredients
    • A61K31/33—Heterocyclic compounds
    • A61K31/395—Heterocyclic compounds having nitrogen as a ring hetero atom, e.g. guanethidine or rifamycins
    • A61K31/495—Heterocyclic compounds having nitrogen as a ring hetero atom, e.g. guanethidine or rifamycins having six-membered rings with two or more nitrogen atoms as the only ring heteroatoms, e.g. piperazine or tetrazines
    • A61K31/505—Pyrimidines; Hydrogenated pyrimidines, e.g. trimethoprim
    • A61K31/519—Pyrimidines; Hydrogenated pyrimidines, e.g. trimethoprim ortho- or peri-condensed with heterocyclic rings

Definitions

  • the present invention relates to compounds and methods useful for inhibiting Fms- like tyrosine kinase 3 (“FLT3"), also known as Cluster of differentiation antigen 135 (CD 135) and fetal liver kinase 2 (Flk2).
  • FLT3 tyrosine kinase 3
  • CD 135 Cluster of differentiation antigen 135
  • Flk2 fetal liver kinase 2
  • the invention also provides pharmaceutically acceptable compositions comprising compounds of the present invention and methods of using said compositions in the treatment of various disorders.
  • Protein kinases constitute a large family of structurally related enzymes that are responsible for the control of a variety of signal transduction processes within the cell. Protein kinases are thought to have evolved from a common ancestral gene due to the conservation of their structure and catalytic function. Almost all kinases contain a similar 250-300 amino acid catalytic domain. The kinases may be categorized into families by the substrates they phosphorylate (e.g., protein-tyrosine, protein-serine/threonine, lipids, etc.).
  • protein kinases mediate intracellular signaling by effecting a phosphoryl transfer from a nucleoside triphosphate to a protein acceptor that is involved in a signaling pathway. These phosphorylation events act as molecular on/off switches that can modulate or regulate the target protein biological function. These phosphorylation events are ultimately triggered in response to a variety of extracellular and other stimuli.
  • Examples of such stimuli include environmental and chemical stress signals (e.g., osmotic shock, heat shock, ultraviolet radiation, bacterial endotoxin, and H 2 0 2 ), cytokines (e.g., interleukin-1 (IL-1), interleukin-8 (IL- 8) and tumor necrosis factor a (TNF-a)), and growth factors (e.g., granulocyte macrophage- colony-stimulating factor (GM-CSF), and fibroblast growth factor (FGF)).
  • An extracellular stimulus may affect one or more cellular responses related to cell growth, migration, differentiation, secretion of hormones, activation of transcription factors, muscle contraction, glucose metabolism, control of protein synthesis, and regulation of the cell cycle.
  • kinase-mediated events include, but are not limited to, autoimmune diseases, inflammatory diseases, bone diseases, metabolic diseases, neurological and neurodegenerative diseases, cancer, cardiovascular diseases, allergies and asthma, Alzheimer's disease, and hormone-related diseases. Accordingly, there remains a need to find protein kinase inhibitors useful as therapeutic agents.
  • Compounds of the present invention are useful for treating a variety of diseases, disorders or conditions, associated with regulation of signaling pathways implicating FLT3 kinases. Such diseases, disorders, or conditions include those described herein.
  • Compounds provided by this invention are also useful for the study of FLT3 enzymes in biological and pathological phenomena; the study of intracellular signal transduction pathways occurring in bodily tissues; and the comparative evaluation of new FLT3 inhibitors or other regulators of kinases, signaling pathways, and cytokine levels in vitro or in vivo.
  • the binding pocket of FLT3 contains a plurality of hydration sites, each of which is occupied by a single molecule of water. Each of these water molecules has a stability rating associated with it.
  • stability rating refers to a numerical calculation which incorporates the enthalpy, entropy, and free energy values associated with each water molecule. This stability rating allows for a measurable determination of the relative stability of water molecules that occupy hydration sites in the binding pocket of FLT3.
  • provided compounds displace or disrupt one or more unstable water molecules.
  • a provided compound displaces or disrupts at least two unstable water molecules.
  • the present invention provides a method of inhibiting FLT3 kinase comprising contacting s a I:
  • Ring A is a 3-7 membered saturated or partially unsaturated carbocyclic ring or a 4-7 membered saturated or partially unsaturated heterocyclic ring having 1-3 heteroatoms independently selected from nitrogen, oxygen, or sulfur;
  • n 0-4;
  • each R 1 is independently -R, halogen, -CN, -N0 2 , -OR, -CH 2 OR, - SR, -N(R) 2 , -S0 2 R, -S0 2 N(R) 2 , -SOR, -C(0)R, -C0 2 R, -C(0)N(R) 2 , -C(0)N(R)-OR, - NRC(0)OR, -NRC(0)N(R) 2 , Cy, or -NRS0 2 R; or R 1 is selected from one of the following formulas:
  • R 1 groups are taken together with their intervening atoms to form an optionally substituted 4-7 membered fused, spiro-fused, or bridged bicyclic ring having 0-2 heteroatoms independently selected from nitrogen, oxygen, or sulfur;
  • each Cy is an optionally substituted ring selected from a 3-7 membered saturated or partially unsaturated carbocyclic ring or a 4-7 membered saturated or partially unsaturated heterocyclic ring having 1-3 heteroatoms independently selected from nitrogen, oxygen, or sulfur;
  • each R is independently hydrogen, or an optionally substituted group selected from Ci_ 6 aliphatic, phenyl, 4-7 membered saturated or partially unsaturated heterocyclic having 1-2 heteroatoms independently selected from nitrogen, oxygen, or sulfur, or 5-6 membered heteroaryl ring having 1-4 heteroatoms independently selected from nitrogen, oxygen, or sulfur, or:
  • Ring B is a 4-8 membered partially unsaturated carbocyclic fused ring; a 4-7 membered partially unsaturated heterocyclic fused ring having 1-2 heteroatoms selected from nitrogen, oxygen, or sulfur; a benzo fused ring; or a 5-6 membered heteroaromatic fused ring having 1-3 heteroatoms independently selected from nitrogen, oxygen, and sulfur wherein said Ring B may be optionally substituted by one or more oxo, thiono, or imino groups; m is 0-4;
  • p 0-2;
  • W is N or -C(R 3 )-;
  • R z is R, CN, N0 2 , halogen, -C(0)N(R) 2 , -C(0)OR, -C(0)R, -N(R) 2 , -NH-[Ar], - N(R)C(0)OR, -NRC(0)N(R) 2 , -OR, or -S0 2 N(R) 2 ;
  • R 3 is hydrogen, halogen, -CN, Ci_ 4 aliphatic, Ci_ 4 haloaliphatic, -OR, -C(0)R, or -C(0)N(R) 2 ;
  • [Ar] is an optionally substituted phenyl or heteroaromatic ring;
  • L 1 is a covalent bond or a Ci_ 6 bivalent hydrocarbon chain wherein one or two methylene units of the chain are optionally and independently replaced by -NR-, -N(R)C(0)-, -C(0)N(R)- , -N(R)S0 2 -, -S0 2 N(R)-, -0-, -C(O)-, -OC(O)-, -C(0)0-, -S-, -SO- or -S0 2 -;
  • each L 2 is independently a covalent bond or a Ci_ 6 bivalent hydrocarbon chain wherein one or two methylene units of the chain are optionally and independently replaced by -NR-, - N(R)C(0)-, -C(0)N(R)-, -N(R)S0 2 -, -S0 2 N(R)-, -0-, -C(O)-, -OC(O)-, -C(0)0-, -S-, -SO- or -S0 2 -;
  • each R 4 is independently halogen, -CN, -N0 2 , -OR,
  • two -L 2 (R 4 ) P -R 4 groups are taken together with their intervening atoms to form an optionally substituted 4-7 membered fused, spiro-fused, or bridged bicyclic ring having 0-2 heteroatoms independently selected from nitrogen, oxygen, or sulfur.
  • aliphatic or "aliphatic group”, as used herein, means a straight-chain (i.e., unbranched) or branched, substituted or unsubstituted hydrocarbon chain that is completely saturated or that contains one or more units of unsaturation, or a monocyclic hydrocarbon or bicyclic hydrocarbon that is completely saturated or that contains one or more units of unsaturation, but which is not aromatic (also referred to herein as "carbocycle,” “cycloaliphatic” or “cycloalkyl”), that has a single point of attachment to the rest of the molecule.
  • aliphatic groups contain 1-6 aliphatic carbon atoms.
  • aliphatic groups contain 1-5 aliphatic carbon atoms. In other embodiments, aliphatic groups contain 1-4 aliphatic carbon atoms. In still other embodiments, aliphatic groups contain 1-3 aliphatic carbon atoms, and in yet other embodiments, aliphatic groups contain 1-2 aliphatic carbon atoms.
  • cycloaliphatic (or “carbocycle” or “cycloalkyl”) refers to a monocyclic C3-C6 hydrocarbon that is completely saturated or that contains one or more units of unsaturation, but which is not aromatic, that has a single point of attachment to the rest of the molecule.
  • Suitable aliphatic groups include, but are not limited to, linear or branched, substituted or unsubstituted alkyl, alkenyl, alkynyl groups and hybrids thereof such as (cycloalkyl)alkyl, (cycloalkenyl)alkyl or (cycloalkyl)alkenyl.
  • bridged bicyclic refers to any bicyclic ring system, i.e. carbocyclic or heterocyclic, saturated or partially unsaturated, having at least one bridge.
  • a "bridge” is an unbranched chain of atoms or an atom or a valence bond connecting two bridgeheads, where a "bridgehead” is any skeletal atom of the ring system which is bonded to three or more skeletal atoms (excluding hydrogen).
  • a bridged bicyclic group has 7-12 ring members and 0-4 heteroatoms independently selected from nitrogen, oxygen, or sulfur.
  • bridged bicyclic groups are well known in the art and include those groups set forth below where each group is attached to the rest of the molecule at any substitutable carbon or nitrogen atom. Unless otherwise specified, a bridged bicyclic group is optionally substituted with one or more substituents as set forth for aliphatic groups. Additionally or alternatively, any substitutable nitrogen of a bridged bicyclic group is optionally substituted. Exemplary bridged bicyclics include:
  • lower alkyl refers to a Ci_ 4 straight or branched alkyl group.
  • exemplary lower alkyl groups are methyl, ethyl, propyl, isopropyl, butyl, isobutyl, and tert-butyl.
  • lower haloalkyl refers to a Ci_ 4 straight or branched alkyl group that is substituted with one or more halogen atoms.
  • heteroatom means one or more of oxygen, sulfur, nitrogen, phosphorus, or silicon (including, any oxidized form of nitrogen, sulfur, phosphorus, or silicon; the quaternized form of any basic nitrogen or; a substitutable nitrogen of a heterocyclic ring, for example N (as in 3,4-dihydro-2H-pyrrolyl), ⁇ (as in pyrrolidinyl) or NR + (as in N-substituted pyrrolidinyl)).
  • bivalent Ci_ 8 (or Ci_ 6 ) saturated or unsaturated, straight or branched, hydrocarbon chain” refers to bivalent alkylene, alkenylene, and alkynylene chains that are straight or branched as defined herein.
  • alkylene refers to a bivalent alkyl group.
  • An "alkylene chain” is a polymethylene group, i.e., -(CH 2 ) n -, wherein n is a positive integer, preferably from 1 to 6, from 1 to 4, from 1 to 3, from 1 to 2, or from 2 to 3.
  • a substituted alkylene chain is a polymethylene group in which one or more methylene hydrogen atoms are replaced with a substituent. Suitable substituents include those described below for a substituted aliphatic group.
  • alkenylene refers to a bivalent alkenyl group.
  • a substituted alkenylene chain is a polymethylene group containing at least one double bond in which one or more hydrogen atoms are replaced with a substituent. Suitable substituents include those described below for a substituted aliphatic group.
  • cyclopropylenyl refers to a bivalent cyclopropyl group of the following structure:
  • halogen means F, CI, Br, or I.
  • aryl used alone or as part of a larger moiety as in “aralkyl,” “aralkoxy,” or “aryloxyalkyl,” refers to monocyclic or bicyclic ring systems having a total of five to fourteen ring members, wherein at least one ring in the system is aromatic and wherein each ring in the system contains 3 to 7 ring members.
  • aryl may be used interchangeably with the term “aryl ring.”
  • aryl refers to an aromatic ring system which includes, but not limited to, phenyl, biphenyl, naphthyl, anthracyl and the like, which may bear one or more substituents.
  • aryl is a group in which an aromatic ring is fused to one or more non-aromatic rings, such as indanyl, phthalimidyl, naphthimidyl, phenanthridinyl, or tetrahydronaphthyl, and the like.
  • heteroaryl and “heteroar-,” used alone or as part of a larger moiety, e.g., “heteroaralkyl,” or “heteroaralkoxy,” refer to groups having 5 to 10 ring atoms, preferably 5, 6, or 9 ring atoms; having 6, 10, or 14 ⁇ electrons shared in a cyclic array; and having, in addition to carbon atoms, from one to five heteroatoms.
  • heteroatom refers to nitrogen, oxygen, or sulfur, and includes any oxidized form of nitrogen or sulfur, and any quatemized form of a basic nitrogen.
  • Heteroaryl groups include, without limitation, thienyl, furanyl, pyrrolyl, imidazolyl, pyrazolyl, triazolyl, tetrazolyl, oxazolyl, isoxazolyl, oxadiazolyl, thiazolyl, isothiazolyl, thiadiazolyl, pyridyl, pyridazinyl, pyrimidinyl, pyrazinyl, indolizinyl, purinyl, naphthyridinyl, and pteridinyl.
  • heteroaryl and “heteroar-”, as used herein, also include groups in which a heteroaromatic ring is fused to one or more aryl, cycloaliphatic, or heterocyclyl rings, where the radical or point of attachment is on the heteroaromatic ring.
  • Nonlimiting examples include indolyl, isoindolyl, benzothienyl, benzofuranyl, dibenzofuranyl, indazolyl, benzimidazolyl, benzthiazolyl, quinolyl, isoquinolyl, cinnolinyl, phthalazinyl, quinazolinyl, quinoxalinyl, 4H-quinolizinyl, carbazolyl, acridinyl, phenazinyl, phenothiazinyl, phenoxazinyl, tetrahydroquinolinyl, tetrahydroisoquinolinyl, and pyrido[2,3-b]-l,4-oxazin- 3(4H)-one.
  • heteroaryl group may be mono- or bicyclic.
  • heteroaryl may be used interchangeably with the terms “heteroaryl ring,” “heteroaryl group,” or “heteroaromatic,” any of which terms include rings that are optionally substituted.
  • heteroarylkyl refers to an alkyl group substituted by a heteroaryl, wherein the alkyl and heteroaryl portions independently are optionally substituted.
  • heterocycle As used herein, the terms “heterocycle,” “heterocyclyl,” “heterocyclic radical,” and “heterocyclic ring” are used interchangeably and refer to a stable 5- to 7-membered monocyclic or 7-10-membered bicyclic heterocyclic moiety that is either saturated or partially unsaturated, and having, in addition to carbon atoms, one or more, preferably one to four, heteroatoms, as defined above.
  • nitrogen includes a substituted nitrogen.
  • the nitrogen may be N (as in 3,4- dihydro-2H-pyrrolyl), ⁇ (as in pyrrolidinyl), or (as in N-substituted pyrrolidinyl).
  • a heterocyclic ring can be attached to its pendant group at any heteroatom or carbon atom that results in a stable structure and any of the ring atoms can be optionally substituted.
  • saturated or partially unsaturated heterocyclic radicals include, without limitation, tetrahydrofuranyl, tetrahydrothiophenyl pyrrolidinyl, piperidinyl, pyrrolinyl, tetrahydroquinolinyl, tetrahydroisoquinolinyl, decahydroquinolinyl, oxazolidinyl, piperazinyl, dioxanyl, dioxolanyl, diazepinyl, oxazepinyl, thiazepinyl, morpholinyl, and quinuclidinyl.
  • heterocycle refers to an alkyl group substituted by a heterocyclyl, wherein the alkyl and heterocyclyl portions independently are optionally substituted.
  • partially unsaturated refers to a ring moiety that includes at least one double or triple bond.
  • partially unsaturated is intended to encompass rings having multiple sites of unsaturation, but is not intended to include aryl or heteroaryl moieties, as herein defined.
  • compounds of the invention may contain "optionally substituted” moieties.
  • substituted whether preceded by the term “optionally” or not, means that one or more hydrogens of the designated moiety are replaced with a suitable substituent.
  • an "optionally substituted” group may have a suitable substituent at each substitutable position of the group, and when more than one position in any given structure may be substituted with more than one substituent selected from a specified group, the substituent may be either the same or different at every position.
  • Combinations of substituents envisioned by this invention are preferably those that result in the formation of stable or chemically feasible compounds.
  • stable refers to compounds that are not substantially altered when subjected to conditions to allow for their production, detection, and, in certain embodiments, their recovery, purification, and use for one or more of the purposes disclosed herein.
  • Suitable monovalent substituents on R° are independently halogen, -(CH 2 ) 0 2 R*, -(haloR*), -(CH 2 ) 0 2 OH, -(CH 2 ) 0 2 OR*, -(CH 2 ) 0 2 CH(OR*) 2 ; -O(haloR'), -CN, -N 3 , -(CH 2 ) 0 2 C(0)R*, -(CH 2 ) 0 2 C(0)OH, -(CH 2 ) 0 2 C(0)OR*, - (CH 2 )o 2 SR*, -(CH 2 )o 2 SH, -(CH 2 ) 0 2 NH 2 , -(CH 2 ) 0 2 NHR*, -(CH 2 ) 0 2 NR* 2 , -N0 2 , -SiR*
  • Suitable divalent substituents that are bound to vicinal substitutable carbons of an "optionally substituted” group include: -0(CR 2 ) 2 - 3 0-, wherein each independent occurrence of R is selected from hydrogen, Ci_6 aliphatic which may be substituted as defined below, or an unsubstituted 5-6-membered saturated, partially unsaturated, or aryl ring having 0-4 heteroatoms independently selected from nitrogen, oxygen, or sulfur.
  • Suitable substituents on the aliphatic group of R include halogen, - R*, -(haloR*), -OH, -OR*, -O(haloR'), -CN, -C(0)OH, -C(0)OR*, -NH 2 , -NHR*, -NR* 2 , or
  • each R* is unsubstituted or where preceded by "halo” is substituted only with one or more halogens, and is independently Ci_ 4 aliphatic, -CH 2 Ph, -O(CH 2 ) 0 iPh, or a 5-6- membered saturated, partially unsaturated, or aryl ring having 0-4 heteroatoms independently selected from nitrogen, oxygen, or sulfur.
  • Suitable substituents on a substitutable nitrogen of an "optionally substituted" group include -R ⁇ , -NR ⁇ 2 , -C(0)R ⁇ , -C(0)OR ⁇ , -C(0)C(0)R ⁇ , C(0)CH 2 C(0)R ⁇ , -S(0) 2 R ⁇ , -S(0) 2 NR ⁇ 2 , -C(S)NR ⁇ 2 , -C(NH)NR ⁇ 2 , or -N(R ⁇ )S(0) 2 R ⁇ ; wherein each R ⁇ is independently hydrogen, Ci_6 aliphatic which may be substituted as defined below, unsubstituted -OPh, or an unsubstituted 5-6-membered saturated, partially unsaturated, or aryl ring having 0-4 heteroatoms independently selected from nitrogen, oxygen, or sulfur, or, notwithstanding the definition above, two independent occurrences of R ⁇ , taken together with their intervening atom(s) form
  • Suitable substituents on the aliphatic group of R ⁇ are independently halogen, - R*, -(haloR*), -OH, -OR*, -O(haloR'), -CN, -C(0)OH, -C(0)OR*, -NH 2 , -NHR*, -NR* 2 , or -N0 2 , wherein each R* is unsubstituted or where preceded by "halo" is substituted only with one or more halogens, and is independently Ci_ 4 aliphatic, -CH 2 Ph, -O(CH 2 ) 0 iPh, or a 5-6- membered saturated, partially unsaturated, or aryl ring having 0-4 heteroatoms independently selected from nitrogen, oxygen, or sulfur.
  • the term "pharmaceutically acceptable salt” refers to those salts which are, within the scope of sound medical judgment, suitable for use in contact with the tissues of humans and lower animals without undue toxicity, irritation, allergic response and the like, and are commensurate with a reasonable benefit/risk ratio.
  • Pharmaceutically acceptable salts are well known in the art. For example, S. M. Berge et al., describe pharmaceutically acceptable salts in detail in J. Pharmaceutical Sciences, 1977, 66, 1-19, incorporated herein by reference.
  • Pharmaceutically acceptable salts of the compounds of this invention include those derived from suitable inorganic and organic acids and bases.
  • Examples of pharmaceutically acceptable, nontoxic acid addition salts are salts of an amino group formed with inorganic acids such as hydrochloric acid, hydrobromic acid, phosphoric acid, sulfuric acid and perchloric acid or with organic acids such as acetic acid, oxalic acid, maleic acid, tartaric acid, citric acid, succinic acid or malonic acid or by using other methods used in the art such as ion exchange.
  • inorganic acids such as hydrochloric acid, hydrobromic acid, phosphoric acid, sulfuric acid and perchloric acid
  • organic acids such as acetic acid, oxalic acid, maleic acid, tartaric acid, citric acid, succinic acid or malonic acid or by using other methods used in the art such as ion exchange.
  • salts include adipate, alginate, ascorbate, aspartate, benzenesulfonate, benzoate, bisulfate, borate, butyrate, camphorate, camphorsulfonate, citrate, cyclopentanepropionate, digluconate, dodecylsulfate, ethanesulfonate, formate, fumarate, glucoheptonate, glycerophosphate, gluconate, hemisulfate, heptanoate, hexanoate, hydroiodide, 2-hydroxy-ethanesulfonate, lactobionate, lactate, laurate, lauryl sulfate, malate, maleate, malonate, methanesulfonate, 2-naphthalenesulfonate, nicotinate, nitrate, oleate, oxalate, palmitate, pamoate, pectinate,
  • Salts derived from appropriate bases include alkali metal, alkaline earth metal, ammonium and salts.
  • Representative alkali or alkaline earth metal salts include sodium, lithium, potassium, calcium, magnesium, and the like.
  • Further pharmaceutically acceptable salts include, when appropriate, nontoxic ammonium, quaternary ammonium, and amine cations formed using counterions such as halide, hydroxide, carboxylate, sulfate, phosphate, nitrate, loweralkyl sulfonate and aryl sulfonate.
  • structures depicted herein are also meant to include all isomeric (e.g., enantiomeric, diastereomeric, and geometric (or conformational)) forms of the structure; for example, the R and S configurations for each asymmetric center, Z and E double bond isomers, and Z and E conformational isomers. Therefore, single stereochemical isomers as well as enantiomeric, diastereomeric, and geometric (or conformational) mixtures of the present compounds are within the scope of the invention. Unless otherwise stated, all tautomeric forms of the compounds of the invention are within the scope of the invention.
  • structures depicted herein are also meant to include compounds that differ only in the presence of one or more isotopically enriched atoms.
  • compounds having the present structures including the replacement of hydrogen by deuterium or tritium, or the replacement of a carbon by a 13 C- or 14 C-enriched carbon are within the scope of this invention.
  • Such compounds are useful, for example, as analytical tools, as probes in biological assays, or as therapeutic agents in accordance with the present invention.
  • an R 1 group of a provided compound comprises one or more deuterium atoms.
  • Ring B of a provided compound may be substituted with one or more deuterium atoms.
  • an inhibitor is defined as a compound that binds to and /or inhibits FLT3 with measurable affinity.
  • an inhibitor has an IC 50 and/or binding constant of less than about 50 ⁇ , less than about 1 ⁇ , less than about 500 nM, less than about 100 nM, less than about 10 nM, or less than about 1 nM.
  • a compound of the present invention may be tethered to a detectable moiety. It will be appreciated that such compounds are useful as imaging agents.
  • a detectable moiety may be attached to a provided compound via a suitable substituent.
  • suitable substituent refers to a moiety that is capable of covalent attachment to a detectable moiety.
  • moieties are well known to one of ordinary skill in the art and include groups containing, e.g., a carboxylate moiety, an amino moiety, a thiol moiety, or a hydroxyl moiety, to name but a few.
  • moieties may be directly attached to a provided compound or via a tethering group, such as a bivalent saturated or unsaturated hydrocarbon chain.
  • such moieties may be attached via click chemistry.
  • such moieties may be attached via a 1,3-cycloaddition of an azide with an alkyne, optionally in the presence of a copper catalyst.
  • Methods of using click chemistry are known in the art and include those described by Rostovtsev et al. , Angew. Chem. Int. Ed. 2002, 41, 2596-99 and Sun et al, Bioconjugate Chem., 2006, 17, 52-57.
  • detecttable moiety is used interchangeably with the term “label” and relates to any moiety capable of being detected, e.g., primary labels and secondary
  • Primary labels such as radioisotopes (e.g., tritium, P, P, S, or C), mass-tags, and fluorescent labels are signal generating reporter groups which can be detected without further modifications.
  • Detectable moieties also include luminescent and phosphorescent groups.
  • secondary label refers to moieties such as biotin and various protein antigens that require the presence of a second intermediate for production of a detectable signal.
  • the secondary intermediate may include streptavidin-enzyme conjugates.
  • antigen labels secondary intermediates may include antibody-enzyme conjugates.
  • fluorescent label refers to moieties that absorb light energy at a defined excitation wavelength and emit light energy at a different wavelength.
  • fluorescent labels include, but are not limited to: Alexa Fluor dyes (Alexa Fluor 350, Alexa Fluor 488, Alexa Fluor 532, Alexa Fluor 546, Alexa Fluor 568, Alexa Fluor 594, Alexa Fluor 633, Alexa Fluor 660 and Alexa Fluor 680), AMCA, AMCA-S, BODIPY dyes (BODIPY FL, BODIPY R6G, BODIPY TMR, BODIPY TR, BODIPY 530/550, BODIPY 558/568, BODIPY 564/570, BODIPY 576/589, BODIPY 581/591, BODIPY 630/650, BODIPY 650/665), Carboxyrhodamine 6G, carboxy
  • mass-tag refers to any moiety that is capable of being uniquely detected by virtue of its mass using mass spectrometry (MS) detection techniques.
  • mass-tags include electrophore release tags such as N-[3-[4'-[(p- Methoxytetrafluorobenzyl)oxy]phenyl]-3-methylglyceronyl]isonipecotic Acid, 4 ' -[2,3 ,5 ,6- Tetrafluoro-4-(pentafluorophenoxyl)]methyl acetophenone, and their derivatives.
  • mass-tags include, but are not limited to, nucleotides, dideoxynucleotides, oligonucleotides of varying length and base composition, oligopeptides, oligosaccharides, and other synthetic polymers of varying length and monomer composition.
  • nucleotides dideoxynucleotides
  • oligonucleotides of varying length and base composition oligopeptides, oligosaccharides
  • other synthetic polymers of varying length and monomer composition.
  • a large variety of organic molecules, both neutral and charged (biomolecules or synthetic compounds) of an appropriate mass range (100-2000 Daltons) may also be used as mass-tags.
  • measurable affinity and “measurably inhibit,” as used herein, means a measurable change in a FLT3 protein kinase activity between a sample comprising a compound of the present invention, or composition thereof, and a FLT3 protein kinase, and an equivalent sample comprising an FLT3 protein kinase, in the absence of said compound, or composition thereof.
  • the present invention provides a method of inhibiting FLT3 kinase, comprising contacting said kinase with a compound of formula I:
  • Ring A is a 3-7 membered saturated or partially unsaturated carbocyclic ring or a 4-7 membered saturated or partially unsaturated heterocyclic ring having 1-3 heteroatoms independently selected from nitrogen, oxygen, or sulfur;
  • n 0-4;
  • each R 1 is independently -R, halogen, -CN, -N0 2 , -OR, -CH 2 OR, - SR, -N(R) 2 , -S0 2 R, -S0 2 N(R) 2 , -SOR, -C(0)R, -C0 2 R, -C(0)N(R) 2 , -C(0)N(R)-OR, - NRC(0)OR, -NRC(0)N(R) 2 , Cy, or -NRS0 2 R; or R 1 is selected from one of the following formulas:
  • R 1 groups are taken together with their intervening atoms to form an optionally substituted 4-7 membered fused, spiro-fused, or bridged bicyclic ring having 0-2 heteroatoms independently selected from nitrogen, oxygen, or sulfur;
  • each Cy is an optionally substituted monocyclic or bicyclic ring selected from a 3-7 membered saturated or partially unsaturated carbocyclic ring or a 4-7 membered saturated or partially unsaturated heterocyclic monocyclic or bicyclic ring having 1-3 heteroatoms independently selected from nitrogen, oxygen, or sulfur;
  • each R is independently hydrogen, or an optionally substituted group selected from Ci_ 6 aliphatic, phenyl, 4-7 membered saturated or partially unsaturated heterocyclic having 1-2 heteroatoms independently selected from nitrogen, oxygen, or sulfur, or 5-6 membered heteroaryl ring having 1-4 heteroatoms independently selected from nitrogen, oxygen, or sulfur, or:
  • Ring B is a 4-8 membered partially unsaturated carbocyclic fused ring; a 4-7 membered partially unsaturated heterocyclic fused ring having 1-2 heteroatoms selected from nitrogen, oxygen, or sulfur; a benzo fused ring; or a 5-6 membered heteroaromatic fused ring having 1-3 heteroatoms independently selected from nitrogen, oxygen, and sulfur wherein said Ring B may be optionally substituted by one or more oxo, thiono, or imino groups;
  • n 0-4;
  • p 0-2;
  • W is N or -C(R 3 )-;
  • R z is R, CN, N0 2 , halogen, -C(0)N(R) 2 , -C(0)OR, -C(0)R, -N(R) 2 , -NH-[Ar], - N(R)C(0)OR, -NRC(0)N(R) 2 , -OR, or -S0 2 N(R) 2 ;
  • R 3 is hydrogen, halogen, -CN, Ci_ 4 aliphatic, Ci_ 4 haloaliphatic, -OR, -C(0)R, or -C(0)N(R) 2 ;
  • [Ar] is an optionally substituted phenyl or heteroaromatic ring;
  • L 1 is a covalent bond or a Ci_ 6 bivalent hydrocarbon chain wherein one or two methylene units of the chain are optionally and independently replaced by -NR-, -N(R)C(0)-, -C(0)N(R)- , -N(R)S0 2 -, -S0 2 N(R)-, -0-, -C(O)-, -OC(O)-, -C(0)0-, -S-, -SO- or -S0 2 -;
  • each L 2 is independently a covalent bond or a Ci_ 6 bivalent hydrocarbon chain wherein one or two methylene units of the chain are optionally and independently replaced by -NR-, - N(R)C(0)-, -C(0)N(R)-, -N(R)S0 2 -, -S0 2 N(R)-, -0-, -C(O)-, -OC(O)-, -C(0)0-, -S-, -SO- or -S0 2 -; each R 4 is independently halogen, -CN, -N0 2 , -OR,
  • two -L 2 (R 4 ) P -R 4 groups are taken together with their intervening atoms to form an optionally substituted 4-7 membered fused, spiro-fused, or bridged bicyclic ring having 0-2 heteroatoms independently selected from nitrogen, oxygen, or sulfur.
  • the Ring A group of formula I is a 3-7 membered saturated or partially unsaturated carbocyclic ring or a 4-7 membered saturated or partially unsaturated heterocyclic ring having 1-3 heteroatoms independently selected from nitrogen, oxygen, or sulfur.
  • Ring A is a 3-7 membered saturated or partially unsaturated carbocyclic ring.
  • Ring A is a 4-7 membered saturated or partially unsaturated heterocyclic ring having 1-3 heteroatoms independently selected from nitrogen, oxygen, or sulfur.
  • Ring A is a 3-7 membered saturated carbocyclic ring. In certain embodiments, Ring A is cyclopentyl or cyclohexyl. In some embodiments, Ring A is cyclohexyl.
  • Ring A when Ring A is a disubstituted cycloalkyl ring, said ring can have cis or trans relative stereochemistry.
  • Ring A is a trans- 1 ,4-disubstituted cycloalkyl ring.
  • Ring A a trans- 1 ,4-disubstituted cyclohexyl ring.
  • Ring A is a 4-7 membered saturated heterocyclic ring having 1-3 heteroatoms independently selected from nitrogen, oxygen, or sulfur. In certain embodiments, Ring A is a 5-6 membered saturated heterocyclic ring having 1-3 heteroatoms independently selected from nitrogen, oxygen, or sulfur. In certain embodiments, Ring A is piperidinyl, piperazinyl, morpholinyl, thiomorpholinyl, tetrahydropyranyl, or tetrahydrofuranyl. In some embodiments, when Ring A is a 4-7 membered saturated heterocyclic ring, L 1 is a covalent bond. In some embodiments, when Ring A is a 4-7 membered saturated heterocyclic ring, L 1 is not a covalent bond.
  • n group of formula I is 0-4. In some embodiments, n is 0. In some embodiments, n is 1-4. In certain embodiments, n is 1. In some embodiments, n is 2.
  • each R 1 group of formula I is independently -R, halogen, -CN, -N0 2 , -OR, -CH 2 OR, -SR, -N(R) 2 , -S0 2 R, -S0 2 N(R) 2 , -SOR, -C(0)R, -C0 2 R, -C(0)N(R) 2 , -C(0)N(R)-OR, -NRC(0)R, -NRC(0)N(R) 2 , Cy, or - NRS0 2 R; or R 1 is selecte from one of the following formulas:
  • R 1 groups are taken together with their intervening atoms to form an optionally substituted 4-7 membered fused, spiro-fused, or bridged bicyclic ring having 0-2 heteroatoms independently selected from nitrogen, oxygen, or sulfur.
  • R 1 is R, -OR, -N(R) 2 , -C0 2 R, -C(0)N(R) 2 , -C(0)N(R)- OR, -S0 2 N(R) 2 , Cy, or -NRC(0)OR.
  • R 1 is -C(0)NH 2 , -C(0)NHCH 3 , - C(0)NH-OH, -CH 3 , -CH 2 CH 3 , -S0 2 t-butyl, -OH, -C(0)OH, -NH 2 , -NHCH 3 , -N(CH 3 ) 2 , - -NHC(0)CH 3 , or -CH 2 phenyl.
  • Rl is selected from one of H 2 ) -4 R j n certain embodiments, R 1 is
  • R 1 is -N(R) 2 .
  • Exemplary R 1 groups include those depicted in Table 1. In some embodiments R 1 is R only where R is not hydrogen.
  • the present invention provides a method utilizing a compound of formula I wherein two R 1 groups are taken together with their intervening atoms to form an optionally substituted 4-7 membered fused, spiro-fused, or bridged bicyclic ring having 0-2 heteroatoms independently selected from nitrogen, oxygen, or sulfur.
  • two R 1 groups on adjacent carbon atoms are taken together to form an optionally substituted 4-7 membered ring fused to Ring A.
  • two R 1 groups on the same carbon atom are taken together to form an optionally substituted 4-7 membered spiro-fused ring.
  • two R 1 groups on non-adjacent carbon atoms are taken together to form an optionally substituted bridged bicyclic ring with Ring A.
  • Cy is an optionally substituted monocyclic or bicyclic ring selected from a 3-7 membered saturated or partially unsaturated carbocyclic ring or a 4-7 membered saturated or partially unsaturated heterocyclic monocyclic or bicyclic ring having 1-3 heteroatoms independently selected from nitrogen, oxygen, or sulfur.
  • Cy is a 3-7 membered saturated carbocyclic ring. In certain embodiments, Cy is a 4-7 membered saturated heterocyclic ring containing 1-2 heteroatoms independently selected from nitrogen, oxygen or sulfur. In certain embodiments Cy is a spirobicyclic 7-membered ring. In some embodiments, Cy is 2-oxa-6-azaspiro[3.3]heptane. In certain embodiments, Cy is morpholinyl, pyrrolidinyl, azetidinyl, piperidinyl or piperazinyl.
  • an R 1 substituent on a saturated carbon of Ring A forms a chiral center.
  • that chiral center is in the (R) configuration.
  • that chiral center is in the (S) configuration.
  • the L 1 group of formula I is a covalent bond or a Ci_ 6 bivalent hydrocarbon chain wherein one or two methylene units of the chain are optionally and independently replaced by -NR-, -N(R)C(0)-, -C(0)N(R)-, -N(R)S0 2 -, -S0 2 N(R)-, -0-, -C(O)-, -OC(O)-, -C(0)0-, -S-, -SO- or -S0 2 -.
  • L 1 is a covalent bond.
  • L 1 is a Ci_ 6 bivalent hydrocarbon chain wherein one or two methylene units of the chain are optionally and independently replaced by -NR-, -N(R)C(0)-, -C(0)N(R)-, -N(R)S0 2 - , -S0 2 N(R)-, -0-, -C(O)-, -OC(O)-, -C(0)0-, -S-, -SO- or -S0 2 -.
  • L 1 is -NH- (i.e., a Ci bivalent hydrocarbon chain wherein the methylene unit is replaced by -NH-), -0-, -CH 2 0-, -OCH 2 -, -NHC(O)-, -CH 2 NH-, or -NHCH 2 -.
  • L 1 is -0-.
  • L 1 is -NR-.
  • L 1 is -OCH 2 -.
  • L 1 is -NRCH 2 -.
  • Exemplary L 1 groups include those depicted in Table 1.
  • the Ring B group of formula I is a 4-8 membered partially unsaturated carbocyclic fused ring; a 4-7 membered partially unsaturated heterocyclic fused ring having 1-2 heteroatoms selected from nitrogen, oxygen, or sulfur; a benzo fused ring; or a 5-6 membered heteroaromatic fused ring having 1-3 heteroatoms independently selected from nitrogen, oxygen, and sulfur wherein said Ring B may be optionally substituted by one or more oxo, thiono, or imino groups.
  • Ring B is a 4-8 membered partially unsaturated carbocyclic fused ring. In other embodiments, Ring B is a 4-7 membered partially unsaturated azacyclic fused ring having one or two nitrogens. In some embodiments, Ring B is a cyclohexo- or cyclopento-fused ring. In other embodiments, Ring B is a piperidino-fused ring. In some embodiments, Ring B is a tetrahydropyrano-fused ring. In some embodiments, Ring B is a pyrrolidino-fused ring. In some embodiments, Ring B is a benzo fused ring.
  • Ring B is a 5-6 membered heteroaromatic fused ring having 1-3 heteroatoms independently selected from nitrogen, oxygen and sulfur. In certain embodiments, Ring B is a 6 membered heteroaromatic fused ring having 1-2 nitrogens. In certain embodiments, Ring B is a pyrido fused ring, a pyrimidino fused ring, pyridazino or pyrazino fused ring. In certain embodiments, Ring B is a triazino fused ring. In certain embodiments, Ring B is a 5 membered heteroaromatic fused ring containing 1-2 heteroatoms independently selected from oxygen, nitrogen and sulfur.
  • Ring B is a pyrrolo fused ring, a thiopheno fused ring, a furano fused ring, a thiazolofused ring, an isothiazolo fused ring, an imidazolo fused ring, a pyrazolo fused ring, an oxazolo fused ring, an isoxazolo fused ring.
  • Ring B is a 5-6 membered heteroaromatic ring substituted with one or more oxo, thiono or imino groups.
  • a substituent on a saturated carbon of Ring B forms a chiral center.
  • that chiral center is in the (R) configuration.
  • that chiral center is in the (S) configuration.
  • the m group of formula I is 0-4. In some embodiments, m is 0. In some embodiments, m is 1-4. In some embodiments, m is 1.
  • each L 2 is independently a covalent bond or a Ci_ 6 bivalent hydrocarbon chain wherein one or two methylene units of the chain are optionally and independently replaced by -NR-, -N(R)C(0)-, -C(0)N(R)-, -N(R)S0 2 -, -S0 2 N(R)-, -0-, -C(O)-, -OC(O)-, -C(0)0-, -S-, -SO- or -S0 2 -.
  • each L 2 is independently a covalent bond.
  • each L 2 is a Ci_3 bivalent hydrocarbon chain wherein one or two methylene units of the chain are optionally and independently replaced by -C(0)N(R)-, -0-, -C(O)-, -S-, -SO- or - SO 2 -.
  • L 2 is methylene.
  • L 2 is -CH 2 -C(0)-.
  • L 2 is a C 2 hydrocarbon chain substituted with a hydroxyl group (-CH 2 CH(OH)-).
  • each R 4 is independently halogen, -CN, -N0 2 , -OR, -SR, -N(R) 2 , -S0 2 R, -S0 2 N(R) 2 , -SOR, -C(0)R, -C0 2 R, -C(0)N(R) 2 , -NRC(0)R, - NRC(0)N(R) 2 , -C(0)N(R)OR, -N(R)C(0)OR, -N(R)S(0) 2 N(R) 2 , -NRS0 2 R, or an optionally substituted group selected from Ci_ 6 aliphatic, phenyl, 4-7 membered saturated or partially unsaturated heterocyclic having 1-2 heteroatoms independently selected from nitrogen, oxygen, or sulfur, or 5-6 membered heteroaryl ring having 1-4 heteroatoms independently selected from nitrogen, oxygen, or sulfur, or: two -L 2 (R 4 ) P -R 4 groups
  • each R 4 is independently - CN, -OR, -SR, -SOR, -S0 2 R, -C(0)N(R) 2 , -NRC(0)R, or an optionally substituted group selected from Ci_ 6 aliphatic, phenyl, 4-7 membered saturated or partially unsaturated heterocyclic having 1-2 heteroatoms independently selected from nitrogen, oxygen, or sulfur, or 5-6 membered heteroaryl ring having 1-4 heteroatoms independently selected from nitrogen, oxygen, or sulfur.
  • each R 4 is independently - CN, -OR, -SR, -SOR, -S0 2 R, -C(0)N(R) 2 , -NRC(0)R, or an optionally substituted group selected from Ci_ 6 aliphatic, phenyl, 4-7 membered saturated or partially unsaturated heterocyclic having 1-2 heteroatoms independently selected from nitrogen, oxygen, or sulfur, or 5-6 membered heteroaryl ring having 1-4 heteroatoms independently selected from nitrogen, oxygen, or
  • R 4 is an optionally substituted group selected from Ci_ 6 aliphatic, 4-7 membered saturated or partially unsaturated heterocyclic having 1-2 heteroatoms independently selected from nitrogen, oxygen, or sulfur, or 5-6 membered heteroaryl ring having 1-4 heteroatoms independently selected from nitrogen, oxygen, or sulfur.
  • R 4 is hydroxyl.
  • R 4 is -C(0)N(R) 2 .
  • the present invention provides a method utilizing a compound of formula I wherein two -L 2 (R 4 ) P -R 4 groups are taken together with their intervening atoms to form an optionally substituted 4-7 membered fused, spiro-fused, or bridged bicyclic ring having 0-2 heteroatoms independently selected from nitrogen, oxygen, or sulfur.
  • two -L 2 -R 4 groups on adjacent carbon atoms are taken together to form an optionally substituted 4-7 membered ring fused to Ring B.
  • two -L 2 (R 4 ) P -R 4 groups on the same carbon atom are taken together to form an optionally substituted 4-7 membered spiro-fused ring.
  • two -L 2 (R 4 ) P -R 4 groups on non-adjacent carbon atoms are taken together to form an optionally substituted bridged bicyclic ring with Ring B.
  • any one or more -L 2 (R 4 ) P -R 4 groups are independently selected from deuterium, an unsubstituted alkyl group, a -C0 2 R group, and an unsubstituted heterocyclyl group. In some embodiments, any one or more -L 2 (R 4 ) P -R 4 groups are not independently selected from deuterium, an unsubstituted alkyl group a -C0 2 R group, and an unsubstituted heterocyclyl group.
  • an -L 2 (R 4 ) P -R 4 substituent on a saturated carbon of Ring B forms a chiral center.
  • that chiral center is in the (R) configuration.
  • that chiral center is in the (S) configuration.
  • the R z group of formula I is -R, -CN, -N0 2 , halogen, - C(0)N(R) 2 , -C(0)OR, -C(0)R, -N(R) 2 , -NH-[Ar], -N(R)C(0)OR, -NRC(0)N(R) 2 , -OR, or - S0 2 N(R) 2 .
  • R z is hydrogen.
  • R z is CN, halogen, -N(R) 2 or -C(0)N(R) 2 .
  • R z is -NH-[Ar].
  • Exemplary R z groups include those depicted in Table 1.
  • [Ar] is an optionally substituted phenyl or heteroaromatic ring. In some embodiments, [Ar] is an optionally substituted phenyl or 5-6 membered heteroaromatic ring. In some embodiments, [Ar] is an optionally substituted phenyl ring. In some embodiments, [Ar] is an optionally substituted heteroaromatic ring. In some embodiments, [Ar] is an optionally substituted 5-6 membered heteroaromatic ring. In some embodiments, [Ar] is an optionally substituted 5-membered heteroaromatic ring. In some embodiments, [Ar] is an optionally substituted 6-membered heteroaromatic ring. In some embodiments, [Ar] is an optionally substituted pyrazole ring.
  • p is 0-2. In some embodiments p is 0. In some embodiments p is 1. In certain embodiments, p is 2.
  • a provided method employs a compound of formula I, wherein Ring B is a cyclopento fused ring, and W is N, thereby forming a compound of formula II: or a pharmaceutically acceptable salt thereof, wherein each of Ring A, L 1 , L 2 , R z , R 1 , R 4 , m, n, and p is as defined above and described in embodiments herein, both singly and in combination.
  • a provided method employs a compound of formula II, wherein R 1 is one of the following formulas:
  • Ring A, L 1 , L 2 , R, R z , R 1 , R 4 , m, n, and p is as defined above and described in embodiments herein, both singly and in combination.
  • a provided method employs a compound of formula II, wherein m is 1, thereby forming a
  • a provided method employs a compound of formula III, wherein Ring A is cyclohexyl, thereby forming a compound of formula IV:
  • each of, L 1 , L 2 , R z , R 1 , R 4 , n, and p is as defined above and described in embodiments herein, both singly and in combination.
  • a provided method employs a compound of formula III, wherein n is 1 and the cyclohexyl ring has trans stereochemistry, thereby forming a compound of formula V:
  • a provided present invention provides a compound of formula V, wherein R z is -N(R) 2 , thereby forming a compound of formula VI:
  • a provided method employss a compound of formula VII, wherein W 1 1 , W2% X 1 1 , Y 1 , Y2 and Z 1 1 are each independently hydrogen or deuterium:
  • each of L 1 , L 2 , R 1 , R z , and R 4 is as defined above for formula I and described in embodiments herein, both singly and in combination.
  • a provided method employs a compound of formula VIII, wherein W 1 1 , W2% X1 1 , Z1 1 and Z 2 are each independently hydrogen or deuterium:
  • each of L 1 , L 2 , R 1 , R z , and R 4 are defined above for formula I and described in embodiments herein, both singly and in combination.
  • a provided method employs a compound of formula I, wherein Ring B is cyclohexo, W is N, and R z is hydrogen, thereby forming a compound of formula IX:
  • a provided method employs a compound of formula IX, wherein m is 1 , and L 2 is attached a to the thiophene ring, thereby forming a compound of formula X:
  • Ring A, L 1 , L 2 , R 1 , R 4 , n, and p is as defined above and described in embodiments herein, both singly and in combination.
  • a provided method employs a compound of formula IX, wherein m is 1 , and L 2 is attached ⁇ to the thiophene ring, thereby forming a compound of formula XI:
  • Ring A, L 1 , L 2 , R 1 , R 4 , n, and p is as defined above and described in embodiments herein, both singly and in combination.
  • a provided method employs a compound of formula III, wherein R z is hydrogen, and L 2 is C 2 alkylene, thereby forming a compound of formula XII:
  • a provided method employs a compound of formula XII, wherein one instance of R4 is -C(0)NR 2 , thereby forming a compound of formula XIII:
  • Ring A, L 1 , R, R 1 , R 4 , n, and p is as defined above and described in embodiments herein, both singly and in combination.
  • a provided method employs a compound of formula XIII, wherein Ring A is 4-substituted cyclohexyl, thereby forming a compound of formula XIV:
  • each of L 1 , R, R 1 , R 4 , n, and p is as defined above and described in embodiments herein, both singly and in combination.
  • a provided method employs a compound of formula XIV, wherein n is 1, and R 1 is -NR 2 , thereby forming a compound of formula XV:
  • a provided method employs a compound of formula XV, wherein the stereochemistry of the substituent on the cyclopento ring is (R), and the relative stereochemistry on the cyclohexyl rin is trans thereby forming a compound of formula XVI:
  • each of L 1 , R, R 4 , and p is as defined above and described in embodiments herein, both singly and in combination.
  • a provided method employs a compound of formula XVI wherein each R 4 is independently hydrogen, fluoro or -OR.
  • a provided method employs a compound of formula XVI wherein L 1 is -0-. In certain embodiments a provided method employs a compound of formula XVI wherein L 1 is -NH-.
  • a provided method employs a compound of formula III, wherein R z is hydrogen, and L 2 is Ci alkylene, thereby forming a compound of formula XVII:
  • Ring A, L 1 , R 1 , R 4 , n, and p is as defined above and described in embodiments herein, both singly and in combination.
  • a provided method employs a compound of formula XVII, wherein one instance of R4 is -C(0)NR 2 , thereby forming a compound of formula XVIII:
  • Ring A, L 1 , R, R 1 , R 4 , n, and p is as defined above and described in embodiments herein, both singly and in combination.
  • a provided method employs a compound of formula XVIII, wherein Ring A is 4-substituted c clohexyl, thereby forming a compound of formula XIX:
  • each of L 1 , R, R 1 , R 4 , n, and p is as defined above and described in embodiments herein, both singly and in combination.
  • a provided method employs a compound of formula XIX, wherein n is 1, and R 1 is -NR 2 , thereby forming a compound of formula XX:
  • a provided method employs a compound of formula XX, wherein the stereochemistry of the substituent on the cyclopento ring is (R), and the relative stereochemistry on the cyclohexyl ring is trans thereby forming a compound of formula XXI:
  • each of L 1 , R, R 4 , and p is as defined above and described in embodiments herein, both singly and in combination.
  • a provided method employs a compound of formula XXI wherein each R 4 is independently hydrogen, fluoro or -OR.
  • a provided method employs a compound of formula XXI wherein L 1 is -0-. In certain embodiments a provided method employs a compound of formula XXI wherein L 1 is -NH-.
  • a provided method employs a compound of formula I, wherein Ring B is piperidino, m is 1 , and R z is hydrogen, thereby forming a compound of formula XXII:
  • Ring A, L 1 , L 2 , R 1 , R 4 , n, and p is as defined above and described in embodiments herein, both singly and in combination.
  • a provided method employs a compound of formula XXII, wherein L 2 is a bond and p is 0, thereby forming a compound of formula XXIII: or a pharmaceutically acceptable salt thereof, wherein each of Ring A, L 1 , R 1 , R 4 , n, and p is as defined above and described in embodiments herein, both singly and in combination.
  • a provided method employs a compound of formula XXIII wherein R 4 is -S(0) 2 R, -C(0)R, or -C(0)N(R) 2 .
  • a provided method employs a compound of formula II, wherein L 1 is -0-, thereby forming a compound of formula XXIV:
  • Ring A, L 2 , R z , R 1 , R 4 , m, n, and p is as defined above and described in embodiments herein, both singly and in combination.
  • a provided method employs a compound of formula XXIV, wherein m is 1 , thereby forming a compound of formula XXV:
  • Ring A, L 2 , R z , R 1 , R 4 , n, and p is as defined above and described in embodiments herein, both singly and in combination.
  • a provided method employs a compound of formula XXV, wherein Ring A is cyclohexyl, thereby forming a compound of formula XXVI:
  • a provided method employs a compound of formula I, wherein R z is -NH-[Ar], thereby forming a compound of formula XXVII:
  • a provided method employs a compound of formula XXVII, wherein n is 1 and Ring A is 1 ,4-trans-substituted cyclohexyl, thereby forming a compound of formula XXVIII:
  • [Ar] is an optionally substituted phenyl or heteroaromatic ring, and each of Ring A, Ring B, L 1 , L 2 , R 1 , R 4 , m, and p is as defined above and described in embodiments herein, both singly and in combination.
  • [Ar] is an optionally substituted phenyl or heteroaromatic ring.
  • [Ar] is optionally substituted phenyl.
  • [Ar] is an optionally substituted heteroaromatic ring.
  • [Ar] is an optionally substituted 5-membered heteroaromatic ring.
  • [Ar] is an optionally substituted 6-membered heteroaromatic ring.
  • [Ar] is an optionally substituted pyrazole ring.
  • a provided method employs a compound of formula
  • a provided method employs a compound of formula XXVIII, wherein Ring B is cyclohexo, thereby forming a compound of formula XXIX:
  • a provided method employs a compound of formula XXVIII, wherein Ring B is a partially unsaturated tetrahydropyrano-fused ring, thereby forming a compound of one of formulae XXX-a, XXX-b, XXX-c, or XXX-d:
  • a provided method employs a compound of formula XXVIII, wherein Ring B is a partially unsaturated piperidino-fused ring, thereby forming compound of one of formulae XXXI-a, XXXI-b, XXXI-c, or XXXI-d:
  • a provided method employs a compound of formula
  • Ring B is a partially unsaturated pyrrolidino-fused ring, thereby forming a compound of one of formulae XXXII-a, XXXII-b, or XXXII-c:
  • a provided method employs a compound of formula I, wherein n is 1 , Ring A is trans-substituted cyclohexyl, and Ring B is a partially unsaturated tetrahydropyrano-fused ring, thereby forming a compound of one of formulae XXXIII-a, XXXIII-b, XXXIII-c, -d:
  • a provided method employs a compound of formula I, wherein n is 1 , Ring A is trans-substituted cyclohexyl, and Ring B is a partially unsaturated piperidino-fused ring, thereby forming a compound of one of formulae XXXIV-a, XXXIV-b, XXXIV-c, or XXXIV-
  • each of L 1 , L 2 , R 1 , R 4 , R z , m, and p is as defined above and described in embodiments herein, both singly and in combination.
  • a provided method employs a compound of formula I, wherein n is 1 , Ring A is trans-substituted cyclohexyl, and Ring B is a partially unsaturated pyrrolidino-fused ring, thereby forming a compound of one of formulae XXXV-a, XXXV-b, or XXXV-
  • a provided method employs a compound of formula XXXIII-a, XXXIII-b, XXXIII-c, or XXXIII-d, wherein R z is hydrogen, thereby forming a compound of one of d:
  • each of L 1 , L 2 , R 1 , R 4 , m, and p is as defined above and described in embodiments herein, both singly and in combination.
  • a provided method employs a compound of formula XXXIV-a, XXXIV-b, XXXIV-c, or XXXIV-d, wherein R z is hydrogen, thereby forming a compound of one of -d:
  • a provided method employs a compound of formula XXXV- a, XXXV-b, or XXXV-c, wherein R z is hydrogen, thereby forming a compound of one of formulae -a, XXXVIII-b, -c:
  • a provided method employs a compound of formula I, wherein W is N, and R z is hydrogen, thereby forming a compound of formula I-a:
  • Ring A, Ring B, L 1 , L 2 , W, R z , R 1 , R 4 , m, n, and p is as defined above and described in embodiments herein, both singly and in combination.
  • a provided method employs a compound of formula I, wherein Ring B is a benzo fused ring, thereby forming a compound of formula XXXIX:
  • Ring A, L 1 , L 2 , R z , R 1 , R 4 , W, m, n, and p is as defined above and described in embodiments herein, both singly and in
  • a provided method employs a compound of formula
  • R 1 is one of the following formulas:
  • a provided method employs a compound of formula
  • Ring A, L 1 , L 2 , R z , R 1 , R 4 , W, n, and p is as defined above and described in embodiments herein, both singly and in combination.
  • a provided method employs a compound of formula XXXX, wherein Ring A is cyclohexyl, thereby forming a compound of formula XXXXI:
  • each of L 1 , L 2 , R z , R 1 , R 4 , W, n, and p is as defined above and described in embodiments herein, both singly and in combination.
  • a provided method employs a compound of formula
  • each of L 1 , L 2 , R z , R 1 , R 4 , W, and p is as defined above and described in embodiments herein, both singly and in combination.
  • a provided method employs a compound of formula XXXX, wherein R z is hydrogen, and L 2 is Ci alkylene, thereby forming a compound of formula
  • Ring A, L 1 , R 1 , R 4 , W, n, and p is as defined above and described in embodiments herein, both singly and in combination.
  • a provided method employs a compound of formula XXXX, wherein L 2 is a covalent bond, thereby forming a compound of formula XXXXIV:
  • Ring A, L 1 , R 1 , R 4 , W, and n is as defined above and described in embodiments herein, both singly and in combination.
  • a provided method employs a compound of formula I, wherein Ring B is a 6 membered heteroaromatic fused ring having 1-3 nitrogen atoms.
  • a provided method employs a compound of formula I or a pharmaceutically acceptable salt thereof, wherein Ring B is a 6-membered heteroaromatic fused ring having 1-2 nitrogen atoms wherein Ring B is substituted with one or more oxo, thiono, or imino groups.
  • the method employs a compound of formula I, wherein Ring B is a 5 membered heteroaryl fused ring having 1-3 heteroatoms independently selected from nitrogen, oxygen, and sulfur.
  • a provided method employs a compound of formula I or a pharmaceutically acceptable salt thereof, wherein Ring B is a 5 -membered heteroaryl fused ring having 1 heteroatom selected from nitrogen, oxygen, and sulfur, wherein Ring B is substituted with one or more oxo, thiono, or imino groups.
  • the method employs a compound set forth in Table 1, above, or a pharmaceutically acceptable salt thereof.
  • a water molecule displaced or disrupted by an inhibitor compound, or pendant moiety of an inhibitor compound is an unstable water molecule.
  • the method employs a complex comprising FLT3 and an inhibitor, wherein at least one unstable water of FLT3 is displaced or disrupted by the inhibitor. In some embodiments, at least two unstable waters selected are displaced or disrupted by the inhibitor.
  • the compounds of this invention may be prepared or isolated in general by synthetic and/or semi-synthetic methods known to those skilled in the art for analogous compounds and by methods described in detail in the Examples, herein.
  • compositions are provided.
  • the invention provides a composition comprising a compound of this invention or a pharmaceutically acceptable derivative thereof and a pharmaceutically acceptable carrier, adjuvant, or vehicle.
  • the amount of compound in compositions of this invention is such that is effective to measurably inhibit a FLT3 protein kinase, or a mutant thereof, in a biological sample or in a patient.
  • the amount of compound in compositions of this invention is such that is effective to measurably inhibit a FLT3 protein kinase, or a mutant thereof, in a biological sample or in a patient.
  • a composition of this invention is formulated for administration to a patient in need of such composition.
  • a composition of this invention is formulated for oral administration to a patient.
  • patient means an animal, preferably a mammal, and most preferably a human.
  • compositions of this invention refers to a nontoxic carrier, adjuvant, or vehicle that does not destroy the pharmacological activity of the compound with which it is formulated.
  • Pharmaceutically acceptable carriers, adjuvants or vehicles that may be used in the compositions of this invention include, but are not limited to, ion exchangers, alumina, aluminum stearate, lecithin, serum proteins, such as human serum albumin, buffer substances such as phosphates, glycine, sorbic acid, potassium sorbate, partial glyceride mixtures of saturated vegetable fatty acids, water, salts or electrolytes, such as protamine sulfate, disodium hydrogen phosphate, potassium hydrogen phosphate, sodium chloride, zinc salts, colloidal silica, magnesium trisilicate, polyvinyl pyrrolidone, cellulose-based substances, polyethylene glycol, sodium carboxymethylcellulose, polyacrylates, waxes, polyethylene-polyoxypropy
  • a "pharmaceutically acceptable derivative” means any non-toxic salt, ester, salt of an ester or other derivative of a compound of this invention that, upon administration to a recipient, is capable of providing, either directly or indirectly, a compound of this invention or an inhibitorily active metabolite or residue thereof.
  • the term "inhibitorily active metabolite or residue thereof means that a metabolite or residue thereof is also an inhibitor of a FLT3 protein kinase, or a mutant thereof.
  • compositions of the present invention may be administered orally, parenterally, by inhalation spray, topically, rectally, nasally, buccally, vaginally or via an implanted reservoir.
  • parenteral as used herein includes subcutaneous, intravenous, intramuscular, intraarticular, intra-synovial, intrasternal, intrathecal, intrahepatic, intralesional and intracranial injection or infusion techniques.
  • the compositions are administered orally, intraperitoneally or intravenously.
  • Sterile injectable forms of the compositions of this invention may be aqueous or oleaginous suspension. These suspensions may be formulated according to techniques known in the art using suitable dispersing or wetting agents and suspending agents.
  • the sterile injectable preparation may also be a sterile injectable solution or suspension in a nontoxic parenterally acceptable diluent or solvent, for example as a solution in 1,3-butanediol.
  • a nontoxic parenterally acceptable diluent or solvent for example as a solution in 1,3-butanediol.
  • acceptable vehicles and solvents that may be employed are water, Ringer's solution and isotonic sodium chloride solution.
  • sterile, fixed oils are conventionally employed as a solvent or suspending medium.
  • any bland fixed oil may be employed including synthetic mono- or di-glycerides.
  • Fatty acids such as oleic acid and its glyceride derivatives are useful in the preparation of injectables, as are natural pharmaceutically-acceptable oils, such as olive oil or castor oil, especially in their polyoxyethylated versions.
  • These oil solutions or suspensions may also contain a long-chain alcohol diluent or dispersant, such as carboxymethyl cellulose or similar dispersing agents that are commonly used in the formulation of pharmaceutically acceptable dosage forms including emulsions and suspensions.
  • Other commonly used surfactants such as Tweens, Spans and other emulsifying agents or bioavailability enhancers which are commonly used in the manufacture of pharmaceutically acceptable solid, liquid, or other dosage forms may also be used for the purposes of formulation.
  • compositions of this invention may be orally administered in any orally acceptable dosage form including, but not limited to, capsules, tablets, aqueous suspensions or solutions.
  • carriers commonly used include lactose and corn starch.
  • Lubricating agents such as magnesium stearate, are also typically added.
  • useful diluents include lactose and dried cornstarch.
  • aqueous suspensions are required for oral use, the active ingredient is combined with emulsifying and suspending agents. If desired, certain sweetening, flavoring or coloring agents may also be added.
  • compositions of this invention may be administered in the form of suppositories for rectal administration.
  • suppositories for rectal administration.
  • suppositories can be prepared by mixing the agent with a suitable non-irritating excipient that is solid at room temperature but liquid at rectal temperature and therefore will melt in the rectum to release the drug.
  • suitable non-irritating excipient include cocoa butter, beeswax and polyethylene glycols.
  • compositions of this invention may also be administered topically, especially when the target of treatment includes areas or organs readily accessible by topical application, including diseases of the eye, the skin, or the lower intestinal tract. Suitable topical formulations are readily prepared for each of these areas or organs.
  • Topical application for the lower intestinal tract can be effected in a rectal suppository formulation (see above) or in a suitable enema formulation. Topically-transdermal patches may also be used.
  • compositions may be formulated in a suitable ointment containing the active component suspended or dissolved in one or more carriers.
  • Carriers for topical administration of compounds of this invention include, but are not limited to, mineral oil, liquid petrolatum, white petrolatum, propylene glycol, polyoxyethylene, polyoxypropylene compound, emulsifying wax and water.
  • provided pharmaceutically acceptable compositions can be formulated in a suitable lotion or cream containing the active components suspended or dissolved in one or more pharmaceutically acceptable carriers.
  • Suitable carriers include, but are not limited to, mineral oil, sorbitan monostearate, polysorbate 60, cetyl esters wax, cetearyl alcohol, 2-octyldodecanol, benzyl alcohol and water.
  • compositions may be formulated as micronized suspensions in isotonic, pH adjusted sterile saline, or, preferably, as solutions in isotonic, pH adjusted sterile saline, either with or without a preservative such as benzylalkonium chloride.
  • the pharmaceutically acceptable compositions may be formulated in an ointment such as petrolatum.
  • Pharmaceutically acceptable compositions of this invention may also be administered by nasal aerosol or inhalation.
  • compositions are prepared according to techniques well- known in the art of pharmaceutical formulation and may be prepared as solutions in saline, employing benzyl alcohol or other suitable preservatives, absorption promoters to enhance bioavailability, fluorocarbons, and/or other conventional solubilizing or dispersing agents.
  • compositions of this invention are formulated for oral administration. Such formulations may be administered with or without food. In some embodiments, pharmaceutically acceptable compositions of this invention are administered without food. In other embodiments, pharmaceutically acceptable compositions of this invention are administered with food.
  • compositions of the present invention that may be combined with the carrier materials to produce a composition in a single dosage form will vary depending upon the host treated, the particular mode of administration.
  • provided compositions should be formulated so that a dosage of between 0.01 - 100 mg/kg body weight/day of the inhibitor can be administered to a patient receiving these compositions.
  • a specific dosage and treatment regimen for any particular patient will depend upon a variety of factors, including the activity of the specific compound employed, the age, body weight, general health, sex, diet, time of administration, rate of excretion, drug combination, and the judgment of the treating physician and the severity of the particular disease being treated.
  • the amount of a compound of the present invention in the composition will also depend upon the particular compound in the composition.
  • Compounds and compositions described herein are generally useful for the inhibition of kinase activity of one or more enzymes.
  • the kinase inhibited by the compounds and methods of the invention is FLT3
  • FLT3 is a receptor tyrosine kinase (RTK) that is reported to be mutated in 25-30% of acute myeloid leukemia (AML) cases.
  • RTK receptor tyrosine kinase
  • AML acute myeloid leukemia
  • Sanz et al. "FLT3 inhibition as a targeted therapy for acute myeloid leukemia," Current Opinion Oncol. (2009) 21 :594-600.
  • ITD internal tandem duplication
  • FLT3 fins-like tyrosine 3
  • FLT3 is thus a recognized molecular target for the development of new therapies for AML. Sanz id at 596; Qi Chao et al, "Identification of (AC220), a Uniquely Potent, Selective and Efficiacious FMS-Like Tyrosine Kinase-3 (FLT3) Inhibitor," J. Med. Chem. (2009) 52:7808-7816.
  • FLT3 inhibitors including tandutinib and AC220
  • sunitinib a multitargeted kinase inhibitor
  • D835Y Another clinically relevant mutation of FLT3 is D835Y, which results in constitutive activity of FLT3.
  • D835Y is often present as a secondary mutation to ITD, resulting in further drug resistance to existing treatments.
  • Moore et al. Selective FLT3 inhibition of FLT- ITD+ acute myeloid leukaemia resulting in secondary D835Y mutation: a model for emerging clinical resistance systems" Leukemia (2012) 26(7): 1462-70.
  • the activity of a compound utilized in this invention as an inhibitor of FLT3, or a mutant thereof may be assayed in vitro, in vivo or in a cell line.
  • In vitro assays include assays that determine inhibition of either the phosphorylation activity and/or the subsequent functional consequences, or ATPase activity of activated FLT3, or a mutant thereof. Alternate in vitro assays quantitate the ability of the inhibitor to bind to FLT3. Inhibitor binding may be measured by radiolabeling the inhibitor prior to binding, isolating the inhibitor/FLT3 complex and determining the amount of radiolabel bound.
  • inhibitor binding may be determined by running a competition experiment where new inhibitors are incubated with FLT3 bound to known radioligands.
  • Representative in vitro and in vivo assays useful in assaying a FLT3 inhibitor include those described and disclosed in, e.g., , each of which is herein incorporated by reference in its entirety. Detailed conditions for assaying a compound utilized in this invention as an inhibitor of FLT3, or a mutant thereof, are set forth in the Examples below.
  • treatment refers to reversing, alleviating, delaying the onset of, or inhibiting the progress of a disease or disorder, or one or more symptoms thereof, as described herein.
  • treatment may be administered after one or more symptoms have developed.
  • treatment may be administered in the absence of symptoms.
  • treatment may be administered to a susceptible individual prior to the onset of symptoms (e.g., in light of a history of symptoms and/or in light of genetic or other susceptibility factors). Treatment may also be continued after symptoms have resolved, for example to prevent or delay their recurrence.
  • the present invention provides a method for treating a FLT3 -mediated disorder comprising the step of administering to a patient in need thereof a compound of the present invention, or pharmaceutically acceptable composition thereof.
  • FLT3 -mediated disorders, diseases, and/or conditions as used herein means any disease or other deleterious condition in which FLT3 or a mutant thereof is known to play a role. Accordingly, another embodiment of the present invention relates to treating or lessening the severity of one or more diseases in which FLT3, or a mutant thereof, is known to play a role.
  • Such FLT3 -mediated disorders include but are not limited to acute myeloid leukemia (also known as acute myelogenous leukemia), solid tumors, gliomas, myelodysplasia syndrome, renal cell carcinoma, glioblastoma, prostate cancer, melanoma, acute lymphoblastic leukemia, myeloproliferative disorder, nasopharyngeal carcinoma, breast tumors, thrombocytopenia, polycythemia vera, myelofibrosis, chronic myelocytic leukemia, lung tumors, colorectal tumors, and autoimmune diseases.
  • acute myeloid leukemia also known as acute myelogenous leukemia
  • solid tumors include gliomas, myelodysplasia syndrome, renal cell carcinoma, glioblastoma, prostate cancer, melanoma, acute lymphoblastic leukemia, myeloproliferative disorder, nasopharyngeal carcinoma, breast tumors,
  • the disorder is selected from gliomas, myelodysplasia syndrome, acute lymphoblastic leukemia, myeloproliferative disorder, nasopharyngeal carcinoma, polycythemia vera, and myelofibrosis.
  • the present invention provides a method for treating one or more disorders, diseases, and/or conditions wherein the disorder, disease, or condition is a cancer, a neurodegenative disorder, a viral disease, an autoimmune disease, an inflammatory disorder, a hereditary disorder, a hormone-related disease, a metabolic disorder, conditions associated with organ transplantation, immunodeficiency disorders, a destructive bone disorder, a proliferative disorder, an infectious disease, a condition associated with cell death, thrombin- induced platelet aggregation, liver disease, pathologic immune conditions involving T cell activation, a cardiovascular disorder, or a CNS disorder.
  • the disorder, disease, or condition is a cancer, a neurodegenative disorder, a viral disease, an autoimmune disease, an inflammatory disorder, a hereditary disorder, a hormone-related disease, a metabolic disorder, conditions associated with organ transplantation, immunodeficiency disorders, a destructive bone disorder, a proliferative disorder, an infectious disease, a condition associated with cell death, thro
  • a human patient is treated with a compound of the current invention and a pharmaceutically acceptable carrier, adjuvant, or vehicle, wherein said compound is present in an amount to measurably inhibit FLT3 kinase activity or modulate CDK8 scaffold activity as it relates to cyclinC, MED 12, and MED13.
  • Compounds of the current invention are useful in the treatment of a proliferative disease selected from a benign or malignant tumor, solid tumor, carcinoma of the brain, kidney, liver, adrenal gland, bladder, breast, stomach, gastric tumors, ovaries, colon, rectum, prostate, pancreas, lung, vagina, cervix, testis, genitourinary tract, esophagus, larynx, skin, bone or thyroid, sarcoma, glioblastomas, neuroblastomas, multiple myeloma, gastrointestinal cancer, especially colon carcinoma or colorectal adenoma, a tumor of the neck and head, an epidermal hyperproliferation, psoriasis, prostate hyperplasia, a neoplasia, a neoplasia of epithelial character, adenoma, adenocarcinoma, keratoacanthoma, epidermoid carcinoma, large cell carcinoma, non-
  • the present invention provides a method of treating acute myeloid leukemia (AML) comprising administering to a patient in need thereof, a therapeutically effective amount of a compound of the invention, or pharmaceutical composition thereof.
  • AML acute myeloid leukemia
  • the AML is associated with a length mutation in FLT3.
  • the AML is associated with an internal tandem duplication, insertion or deletion of the FLT3 gene (FLT3-ITD).
  • the AML is associated with a point mutation or deletion at residues 835 or 836 of FLT3.
  • the AML is associated with a D835Y point mutation in FLT3.
  • the AML is associated with both an internal tandem duplication and a D835Y mutation in FLT3.
  • the AML is resistant to one or more other FLT3 inhibitors.
  • the AML is resistant to one or more drugs selected from quizartinib (AC220), a staurosporine derivative (e.g. midostaurin (PKC412) or lestaurtinib), sorafenib, and sunitinib.
  • the proliferative disease which can be treated according to the methods of this invention is an IL-6 driven disorder.
  • the IL-6 driven disorder is Smoldering of indolent multiple myeloma.
  • Compounds of the invention are also useful in the treatment of inflammatory or allergic conditions of the skin, for example psoriasis, contact dermatitis, atopic dermatitis, alopecia areata, erythema multiforma, dermatitis herpetiformis, scleroderma, vitiligo, hypersensitivity angiitis, urticaria, bullous pemphigoid, lupus erythematosus, systemic lupus erythematosus, pemphigus vulgaris, pemphigus foliaceus, paraneoplastic pemphigus, epidermolysis bullosa acquisita, acne vulgaris, and other inflammatory or allergic conditions of the skin.
  • Compounds of the invention may also be used for the treatment of other diseases or conditions, such as diseases or conditions having an inflammatory component, for example, treatment of diseases and conditions of the eye such as ocular allergy, conjunctivitis, keratoconjunctivitis sicca, and vernal conjunctivitis, diseases affecting the nose including allergic rhinitis, and inflammatory disease in which autoimmune reactions are implicated or having an autoimmune component or etiology, including autoimmune hematological disorders (e.g.
  • hemolytic anemia aplastic anemia, pure red cell anemia and idiopathic thrombocytopenia
  • systemic lupus erythematosus rheumatoid arthritis, polychondritis, scleroderma, Wegener granulamatosis, dermatomyositis, chronic active hepatitis, myasthenia gravis, Steven-Johnson syndrome, idiopathic sprue, autoimmune inflammatory bowel disease (e.g.
  • ulcerative colitis and Crohn's disease irritable bowel syndrome, celiac disease, periodontitis, hyaline membrane disease, kidney disease, glomerular disease, alcoholic liver disease, multiple sclerosis, endocrine opthalmopathy, Grave's disease, sarcoidosis, alveolitis, chronic hypersensitivity pneumonitis, multiple sclerosis, primary biliary cirrhosis, uveitis (anterior and posterior), Sjogren's syndrome, keratoconjunctivitis sicca and vernal keratoconjunctivitis, interstitial lung fibrosis, psoriatic arthritis, systemic juvenile idiopathic arthritis, cryopyrin-associated periodic syndrome, nephritis, vasculitis, diverticulitis, interstitial cystitis, glomerulonephritis (with and without nephrotic syndrome, e.g.
  • idiopathic nephrotic syndrome or minal change nephropathy chronic granulomatous disease, endometriosis, leptospiriosis renal disease, glaucoma, retinal disease, ageing, headache, pain, complex regional pain syndrome, cardiac hypertrophy, musclewasting, catabolic disorders, obesity, fetal growth retardation, hypercholesterolemia, heart disease, chronic heart failure, mesothelioma, anhidrotic ecodermal dysplasia, Behcet's disease, incontinentia pigmenti, Paget' s disease, pancreatitis, hereditary periodic fever syndrome, asthma (allergic and non-allergic, mild, moderate, severe, bronchitic, and exercise-induced), acute lung injury, acute respiratory distress syndrome, eosinophilia, hypersensitivities, anaphylaxis, nasal sinusitis, ocular allergy, silica induced diseases, COPD (reduction of damage, airways inflammation, bronchial hyperreactivity, remodeling or
  • the inflammatory disease which can be treated according to the methods of this invention is selected from acute and chronic gout, chronic gouty arthritis, psoriasis, psoriatic arthritis, rheumatoid arthritis, Juvenile rheumatoid arthritis, Systemic jubenile idiopathic arthritis (SJIA), Cryopyrin Associated Periodic Syndrome (CAPS), and osteoarthritis.
  • the inflammatory disease which can be treated according to the methods of this invention is a TH17 mediated disease.
  • the TH17 mediated disease is selected from Systemic lupus erythematosus, Multiple sclerosis, and inflammatory bowel disease (including Crohn's disease or ulcerative colitis).
  • the inflammatory disease which can be treated according to the methods of this invention is selected from Sjogren's syndrome, allergic disorders, osteoarthritis, conditions of the eye such as ocular allergy, conjunctivitis, keratoconjunctivitis sicca and vernal conjunctivitis, and diseases affecting the nose such as allergic rhinitis.
  • the invention provides the use of a compound according to the definitions herein, or a pharmaceutically acceptable salt, or a hydrate or solvate thereof for the preparation of a medicament for the treatment of a proliferative disease, an inflammatory disease, an obstructive respiratory disease, a cardiovascular disease, a metabolic disease, a neurological disease, a neurodegenerative disease, a viral disease, or a disorder commonly occurring in connection with transplantation.
  • additional therapeutic agents which are normally administered to treat that condition, may be administered in combination with compounds and compositions of this invention.
  • additional therapeutic agents that are normally administered to treat a particular disease, or condition are known as "appropriate for the disease, or condition, being treated.”
  • a provided combination, or composition thereof is administered in combination with another therapeutic agent.
  • MS Multiple
  • combination therapies of the present invention are administered in combination with a monoclonal antibody or an siR A therapeutic.
  • Those additional agents may be administered separately from a provided combination therapy, as part of a multiple dosage regimen.
  • those agents may be part of a single dosage form, mixed together with a compound of this invention in a single composition. If administered as part of a multiple dosage regime, the two active agents may be submitted simultaneously, sequentially or within a period of time from one another normally within five hours from one another.
  • the term “combination,” “combined,” and related terms refers to the simultaneous or sequential administration of therapeutic agents in accordance with this invention.
  • a combination of the present invention may be administered with another therapeutic agent simultaneously or sequentially in separate unit dosage forms or together in a single unit dosage form.
  • the amount of additional therapeutic agent present in the compositions of this invention will be no more than the amount that would normally be administered in a composition comprising that therapeutic agent as the only active agent.
  • the amount of additional therapeutic agent in the presently disclosed compositions will range from about 50% to 100% of the amount normally present in a composition comprising that agent as the only therapeutically active agent.
  • the present invention provides a composition comprising a compound of formula I and one or more additional therapeutic agents.
  • the therapeutic agent may be administered together with a compound of formula I, or may be administered prior to or following administration of a compound of formula I. Suitable therapeutic agents are described in further detail below.
  • a compound of formula I may be administered up to 5 minutes, 10 minutes, 15 minutes, 30 minutes, 1 hour, 2 hours, 3 hours, 4 hours, 5, hours, 6 hours, 7 hours, 8 hours, 9 hours, 10 hours, 11 hours, 12 hours, 13 hours, 14 hours, 15 hours, 16 hours, 17 hours, or 18 hours before the therapeutic agent.
  • a compound of formula I may be administered up to 5 minutes, 10 minutes, 15 minutes, 30 minutes, 1 hour, 2 hours, 3 hours, 4 hours, 5, hours, 6 hours, 7 hours, 8 hours, 9 hours, 10 hours, 11 hours, 12 hours, 13 hours, 14 hours, 15 hours, 16 hours, 17 hours, or 18 hours following the therapeutic agent.
  • the present invention provides a method of treating an inflammatory disease, disorder or condition by administering to a patient in need thereof a compound of formula I and one or more additional therapeutic agents.
  • additional therapeutic agents may be small molecules or recombinant biologic agents and include, for example, acetaminophen, non-steroidal anti-inflammatory drugs (NSAIDS) such as aspirin, ibuprofen, naproxen, etodolac (Lodine®) and celecoxib, colchicine (Colcrys®), corticosteroids such as prednisone, prednisolone, methylprednisolone, hydrocortisone, and the like, probenecid, allopurinol, febuxostat (Uloric®), sulfasalazine (Azulfidine®), antimalarials such as hydroxychloroquine (Plaquenil®) and chloroquine (Aralen®), methotrex
  • NSAIDS non
  • the present invention provides a method of treating rheumatoid arthritis comprising administering to a patient in need thereof a compound of formula I and one or more additional therapeutic agents selected from non-steroidal antiinflammatory drugs (NSAIDS) such as aspirin, ibuprofen, naproxen, etodolac (Lodine®) and celecoxib, corticosteroids such as prednisone, prednisolone, methylprednisolone, hydrocortisone, and the like, sulfasalazine (Azulfidine®), antimalarials such as hydroxychloroquine (Plaquenil®) and chloroquine (Aralen®), methotrexate (Rheumatrex®), gold salts such as gold thioglucose (Solganal®), gold thiomalate (Myochrysine®) and auranofm (Ridaura®), D-penicill
  • NSAIDS non-
  • the present invention provides a method of treating osteoarthritis comprising administering to a patient in need thereof a compound of formula I and one or more additional therapeutic agents selected from acetaminophen, non-steroidal antiinflammatory drugs (NSAIDS) such as aspirin, ibuprofen, naproxen, etodolac (Lodine®) and celecoxib, diclofenac, cortisone, hyaluronic acid (Synvisc® or Hyalgan®) and monoclonal antibodies such as tanezumab.
  • NSAIDS non-steroidal antiinflammatory drugs
  • the present invention provides a method of treating lupus comprising administering to a patient in need thereof a compound of formula I and one or more additional therapeutic agents selected from acetaminophen, non-steroidal anti-inflammatory drugs (NSAIDS) such as aspirin, ibuprofen, naproxen, etodolac (Lodine®) and celecoxib, corticosteroids such as prednisone, prednisolone, methylprednisolone, hydrocortisone, and the like, antimalarials such as hydroxychloroquine (Plaquenil®) and chloroquine (Aralen®), cyclophosphamide (Cytoxan®), methotrexate (Rheumatrex®), azathioprine (Imuran®) and anticoagulants such as heparin (Calcinparine® or Liquaemin®) and warfarin (Coumadin®).
  • NSAIDS non-steroidal anti
  • the present invention provides a method of treating inflammatory bowel disease comprising administering to a patient in need thereof a compound of formula I and one or more additional therapeutic agents selected from mesalamine (Asacol®) sulfasalazine (Azulfidine®), antidiarrheals such as diphenoxylate (Lomotil®) and loperamide (Imodium®), bile acid binding agents such as cholestyramine, alosetron (Lotronex®), lubiprostone (Amitiza®), laxatives such as Milk of Magnesia, polyethylene glycol (MiraLax®), Dulcolax®, Correctol® and Senokot® and anticholinergics or antispasmodics such as dicyclomine (Bentyl®), anti-TNF therapies, steroids, and antibiotics such as Flagyl or ciprofloxacin.
  • mesalamine Asacol®
  • Amulfidine® antidiarrheals
  • the present invention provides a method of treating asthma comprising administering to a patient in need thereof a compound of formula I and one or more additional therapeutic agents selected from Singulair®, beta-2 agonists such as albuterol (Ventolin® HFA, Proventil® HFA), levalbuterol (Xopenex®), metaproterenol (Alupent®), pirbuterol acetate (Maxair®), terbutaline sulfate (Brethaire®), salmeterol xinafoate (Serevent®) and formoterol (Foradil®), anticholinergic agents such as ipratropium bromide (Atrovent®) and tiotropium (Spiriva®), inhaled corticosteroids such as prednisone, prednisolone, beclomethasone dipropionate (Beclovent®, Qvar®, and Vanceril®), triamcinolone acetonide (A)
  • the present invention provides a method of treating COPD comprising administering to a patient in need thereof a compound of formula I and one or more additional therapeutic agents selected from beta-2 agonists such as albuterol (Ventolin® HFA, Proventil® HFA), levalbuterol (Xopenex®), metaproterenol (Alupent®), pirbuterol acetate (Maxair®), terbutaline sulfate (Brethaire®), salmeterol xinafoate (Serevent®) and formoterol (Foradil®), anticholinergic agents such as ipratropium bromide (Atrovent®) and tiotropium (Spiriva®), methylxanthines such as theophylline (Theo-Dur®, Theolair®, Slo-bid®, Uniphyl®, Theo-24®) and aminophylline, inhaled corticosteroids such as prednisone, pre
  • the present invention provides a method of treating a hematological malignancy comprising administering to a patient in need thereof a compound of formula I and one or more additional therapeutic agents selected from rituximab (Rituxan®), cyclophosphamide (Cytoxan®), doxorubicin (Hydrodaunorubicin®), vincristine (Oncovin®), prednisone, a hedgehog signaling inhibitor, a BTK inhibitor, a JAK/pan-JAK inhibitor, a TYK2 inhibitor, a PI3K inhibitor, a SYK inhibitor, and combinations thereof.
  • the present invention provides a method of treating a solid tumor comprising administering to a patient in need thereof a compound of formula I and one or more additional therapeutic agents selected from rituximab (Rituxan®), cyclophosphamide (Cytoxan®), doxorubicin (Hydrodaunorubicin®), vincristine (Oncovin®), prednisone, a hedgehog signaling inhibitor, a BTK inhibitor, a JAK/pan-JAK inhibitor, a TYK2 inhibitor, a PI3K inhibitor, a SYK inhibitor, and combinations thereof.
  • additional therapeutic agents selected from rituximab (Rituxan®), cyclophosphamide (Cytoxan®), doxorubicin (Hydrodaunorubicin®), vincristine (Oncovin®), prednisone, a hedgehog signaling inhibitor, a BTK inhibitor, a JAK/pan-JAK inhibitor,
  • the present invention provides a method of treating a hematological malignancy comprising administering to a patient in need thereof a compound of formula I and a Hedgehog (Hh) signaling pathway inhibitor.
  • the hematological malignancy is DLBCL (Ramirez et al "Defining causative factors contributing in the activation of hedgehog signaling in diffuse large B-cell lymphoma" Leuk. Res. (2012), published online July 17, and incorporated herein by reference in its entirety).
  • the present invention provides a method of treating diffuse large B-cell lymphoma (DLBCL) comprising administering to a patient in need thereof a compound of formula I and one or more additional therapeutic agents selected from rituximab (Rituxan®), cyclophosphamide (Cytoxan®), doxorubicin (Hydrodaunorubicin®), vincristine (Oncovin®), prednisone, a hedgehog signaling inhibitor, and combinations thereof.
  • rituximab Renuxan®
  • Cytoxan® cyclophosphamide
  • doxorubicin Hydrodaunorubicin®
  • vincristine Oncovin®
  • prednisone a hedgehog signaling inhibitor
  • the present invention provides a method of treating multiple myeloma comprising administering to a patient in need thereof a compound of formula I and one or more additional therapeutic agents selected from bortezomib (Velcade®), and dexamethasone (Decadron®), a hedgehog signaling inhibitor, a BTK inhibitor, a JAK/pan-JAK inhibitor, a TYK2 inhibitor, a PI3K inhibitor, a SYK inhibitor in combination with lenalidomide (Revlimid®).
  • additional therapeutic agents selected from bortezomib (Velcade®), and dexamethasone (Decadron®), a hedgehog signaling inhibitor, a BTK inhibitor, a JAK/pan-JAK inhibitor, a TYK2 inhibitor, a PI3K inhibitor, a SYK inhibitor in combination with lenalidomide (Revlimid®).
  • the present invention provides a method of treating Waldenstrom's macroglobulinemia comprising administering to a patient in need thereof a compound of formula I and one or more additional therapeutic agents selected from chlorambucil (Leukeran®), cyclophosphamide (Cytoxan®, Neosar®), fludarabine (Fhidara®), eSadribme (Leustatm®), riiuximab (Rituxan®), a hedgehog signaling inhibitor, a BTK inhibitor, a JAK/pan-JAK inhibitor, a TYK2 inhibitor, a PI3K inhibitor, and a SYK inhibitor.
  • additional therapeutic agents selected from chlorambucil (Leukeran®), cyclophosphamide (Cytoxan®, Neosar®), fludarabine (Fhidara®), eSadribme (Leustatm®), riiuximab (Rituxan®), a hedgehog signaling inhibitor, a BTK inhibitor
  • the present invention provides a method of treating or lessening the severity of a disease comprising adm nostiring to a patient in need thereof a compotmd of formula I and a BTK inhibitor, wherein the disease is selected from inflammatory bowel disease, arthritis, systemic lupus erythematosus (SLE), vasculitis, idiopathic thrombocytopenic purpura (ITP), rheumatoid arthritis, psoriatic arthritis, osteoarthritis, Still's disease, juvenile arthritis, diabetes, myasthenia gravis, Hashimoto's thyroiditis, Ord's thyroiditis, Graves' disease, autoimmune thyroiditis, Sjogren's syndrome, multiple sclerosis, systemic sclerosis, Lyme neuroborreliosis, Guillain-Barre syndrome, acute disseminated encephalomyelitis, Addison's disease, opsoclonus-myoclonus syndrome,
  • the present invention provides a method of treating or lessening the severity of a disease comprising administering to a patient in need thereof a compound of formula I and a ⁇ 3 inhibitor, wherein the disease is selected from a cancer, a neurodegenative disorder, an angiogenic disorder, a viral disease, an autoimmune disease, an inflammatory disorder, a hormone-related disease, conditions associated with organ transplantation, immunodeficiency disorders, a destructive bone disorder, a proliferative disorder, an infectious disease, a conditio associated with cell death, thrombin-induced platelet aggregation, chronic myelogenous leukemia (C L), chronic lymphocytic leukemia (CLL), liver disease, pathologic immune conditions involving T cell activation, a cardiovascular disorder, and a CNS disorder.
  • the disease is selected from a cancer, a neurodegenative disorder, an angiogenic disorder, a viral disease, an autoimmune disease, an inflammatory disorder, a hormone-related disease, conditions associated with organ transplant
  • the present invention provides a method of treating or lessening the severity of a disease comprising administering to a patient in need thereof a compound of formula I and a PD inhibitor, wherein the disease is selected from benign or malignant tumor, carcinoma or solid tumor of the brain, kidney (e.g., renal cell carcinoma (RCC)), liver, adrenal gland, bladder, breast, stomach, gastric tumors, ovaries, colon, rectum, prostate, pancreas, lung, vagina, endometrium, cervix, testis, genitourinary tract, esophagus, larynx, skin, bone or thyroid, sarcoma, glioblastomas, neuroblastomas, multiple myeloma or gastrointestinal cancer, especially colon carcinoma or colorectal adenoma or a tumor of the neck and head, an epidermal hyperproliferation, psoriasis, prostate hyperplasia, a neoplasia, a ne
  • a PD inhibitor
  • hemolytic anemia aplastic anemia, pure red cell anemia and idiopathic thrombocytopenia
  • systemic lupus erythematosus rheumatoid arthritis, polychondritis, sclerodoma, Wegener granulamatosis, dermatomyositis, chronic active hepatitis, myasthenia gravis, Steven-Johnson syndrome, idiopathic sprue, autoimmune inflammatory bowel disease (e.g.
  • ulcerative colitis and Crohn's disease endocrine opthalmopathy
  • Grave's disease sarcoidosis, alveolitis, chronic hypersensitivity pneumonitis, multiple sclerosis, primary biliary cirrhosis, uveitis (anterior and posterior), keratoconjunctivitis sicca and vernal keratoconjunctivitis, interstitial lung fibrosis, psoriatic arthritis and glomerulonephritis (with and without nephrotic syndrome, e.g.
  • idiopathic nephrotic syndrome or minal change nephropathy, restenosis, cardiomegaly, atherosclerosis, myocardial infarction, ischemic stroke and congestive heart failure, Alzheimer's disease, Parkinson's disease, amyotrophic lateral sclerosis, Huntington's disease, and cerebral ischemia, and neurodegenerative disease caused by traumatic injury, glutamate neurotoxicity and hypoxia.
  • the compounds and compositions, according to the method of the present invention may be administered using any amount and any route of administration effective for treating or lessening the severity of a cancer, an autoimmune disorder, a proliferative disorder, an inflammatory disorder, a neurodegenerative or neurological disorder, schizophrenia, a bone- related disorder, liver disease, or a cardiac disorder.
  • the exact amount required will vary from subject to subject, depending on the species, age, and general condition of the subject, the severity of the infection, the particular agent, its mode of administration, and the like.
  • Compounds of the invention are preferably formulated in dosage unit form for ease of administration and uniformity of dosage.
  • the expression "dosage unit form" as used herein refers to a physically discrete unit of agent appropriate for the patient to be treated.
  • the specific effective dose level for any particular patient or organism will depend upon a variety of factors including the disorder being treated and the severity of the disorder; the activity of the specific compound employed; the specific composition employed; the age, body weight, general health, sex and diet of the patient; the time of administration, route of administration, and rate of excretion of the specific compound employed; the duration of the treatment; drugs used in combination or coincidental with the specific compound employed, and like factors well known in the medical arts.
  • patient means an animal, preferably a mammal, and most preferably a human.
  • compositions of this invention can be administered to humans and other animals orally, rectally, parenterally, intracisternally, intravaginally, intraperitoneally, topically (as by powders, ointments, or drops), bucally, as an oral or nasal spray, or the like, depending on the severity of the infection being treated.
  • the compounds of the invention may be administered orally or parenterally at dosage levels of about 0.01 mg/kg to about 50 mg/kg and preferably from about 1 mg/kg to about 25 mg/kg, of subject body weight per day, one or more times a day, to obtain the desired therapeutic effect.
  • Liquid dosage forms for oral administration include, but are not limited to, pharmaceutically acceptable emulsions, microemulsions, solutions, suspensions, syrups and elixirs.
  • the liquid dosage forms may contain inert diluents commonly used in the art such as, for example, water or other solvents, solubilizing agents and emulsifiers such as ethyl alcohol, isopropyl alcohol, ethyl carbonate, ethyl acetate, benzyl alcohol, benzyl benzoate, propylene glycol, 1,3-butylene glycol, dimethylformamide, oils (in particular, cottonseed, groundnut, corn, germ, olive, castor, and sesame oils), glycerol, tetrahydrofurfuryl alcohol, polyethylene glycols and fatty acid esters of sorbitan, and mixtures thereof.
  • the oral compositions can also include adj
  • Injectable preparations for example, sterile injectable aqueous or oleaginous suspensions may be formulated according to the known art using suitable dispersing or wetting agents and suspending agents.
  • the sterile injectable preparation may also be a sterile injectable solution, suspension or emulsion in a nontoxic parenterally acceptable diluent or solvent, for example, as a solution in 1,3-butanediol.
  • the acceptable vehicles and solvents that may be employed are water, Ringer's solution, U.S. P. and isotonic sodium chloride solution.
  • sterile, fixed oils are conventionally employed as a solvent or suspending medium.
  • any bland fixed oil can be employed including synthetic mono- or diglycerides.
  • fatty acids such as oleic acid are used in the preparation of injectables.
  • Injectable formulations can be sterilized, for example, by filtration through a bacterial-retaining filter, or by incorporating sterilizing agents in the form of sterile solid compositions which can be dissolved or dispersed in sterile water or other sterile injectable medium prior to use.
  • a compound of the present invention In order to prolong the effect of a compound of the present invention, it is often desirable to slow the absorption of the compound from subcutaneous or intramuscular injection. This may be accomplished by the use of a liquid suspension of crystalline or amorphous material with poor water solubility. The rate of absorption of the compound then depends upon its rate of dissolution that, in turn, may depend upon crystal size and crystalline form. Alternatively, delayed absorption of a parenterally administered compound form is accomplished by dissolving or suspending the compound in an oil vehicle. Injectable depot forms are made by forming microencapsule matrices of the compound in biodegradable polymers such as polylactide- polyglycolide.
  • the rate of compound release can be controlled.
  • biodegradable polymers include poly(orthoesters) and poly(anhydrides).
  • Depot injectable formulations are also prepared by entrapping the compound in liposomes or microemulsions that are compatible with body tissues.
  • compositions for rectal or vaginal administration are preferably suppositories which can be prepared by mixing the compounds of this invention with suitable non-irritating excipients or carriers such as cocoa butter, polyethylene glycol or a suppository wax which are solid at ambient temperature but liquid at body temperature and therefore melt in the rectum or vaginal cavity and release the active compound.
  • suitable non-irritating excipients or carriers such as cocoa butter, polyethylene glycol or a suppository wax which are solid at ambient temperature but liquid at body temperature and therefore melt in the rectum or vaginal cavity and release the active compound.
  • Solid dosage forms for oral administration include capsules, tablets, pills, powders, and granules.
  • the active compound is mixed with at least one inert, pharmaceutically acceptable excipient or carrier such as sodium citrate or dicalcium phosphate and/or a) fillers or extenders such as starches, lactose, sucrose, glucose, mannitol, and silicic acid, b) binders such as, for example, carboxymethylcellulose, alginates, gelatin, polyvinylpyrrolidinone, sucrose, and acacia, c) humectants such as glycerol, d) disintegrating agents such as agar—agar, calcium carbonate, potato or tapioca starch, alginic acid, certain silicates, and sodium carbonate, e) solution retarding agents such as paraffin, f) absorption accelerators such as quaternary ammonium compounds, g) wetting agents such as, for example, cetyl alcohol
  • Solid compositions of a similar type may also be employed as fillers in soft and hard- filled gelatin capsules using such excipients as lactose or milk sugar as well as high molecular weight polyethylene glycols and the like.
  • the solid dosage forms of tablets, dragees, capsules, pills, and granules can be prepared with coatings and shells such as enteric coatings and other coatings well known in the pharmaceutical formulating art. They may optionally contain opacifying agents and can also be of a composition that they release the active ingredient(s) only, or preferentially, in a certain part of the intestinal tract, optionally, in a delayed manner. Examples of embedding compositions that can be used include polymeric substances and waxes. Solid compositions of a similar type may also be employed as fillers in soft and hard-filled gelatin capsules using such excipients as lactose or milk sugar as well as high molecular weight polethylene glycols and the like.
  • the active compounds can also be in micro-encapsulated form with one or more excipients as noted above.
  • the solid dosage forms of tablets, dragees, capsules, pills, and granules can be prepared with coatings and shells such as enteric coatings, release controlling coatings and other coatings well known in the pharmaceutical formulating art.
  • the active compound may be admixed with at least one inert diluent such as sucrose, lactose or starch.
  • Such dosage forms may also comprise, as is normal practice, additional substances other than inert diluents, e.g., tableting lubricants and other tableting aids such a magnesium stearate and microcrystalline cellulose.
  • the dosage forms may also comprise buffering agents. They may optionally contain opacifying agents and can also be of a composition that they release the active ingredient(s) only, or preferentially, in a certain part of the intestinal tract, optionally, in a delayed manner. Examples of embedding compositions that can be used include polymeric substances and waxes.
  • Dosage forms for topical or transdermal administration of a compound of this invention include ointments, pastes, creams, lotions, gels, powders, solutions, sprays, inhalants or patches.
  • the active component is admixed under sterile conditions with a pharmaceutically acceptable carrier and any needed preservatives or buffers as may be required.
  • Ophthalmic formulation, ear drops, and eye drops are also contemplated as being within the scope of this invention. Additionally, the present invention contemplates the use of transdermal patches, which have the added advantage of providing controlled delivery of a compound to the body. Such dosage forms can be made by dissolving or dispensing the compound in the proper medium. Absorption enhancers can also be used to increase the flux of the compound across the skin. The rate can be controlled by either providing a rate controlling membrane or by dispersing the compound in a polymer matrix or gel.
  • the invention relates to a method of inhibiting protein kinase activity in a biological sample comprising the step of contacting said biological sample with a compound of this invention, or a composition comprising said compound.
  • the invention relates to a method of inhibiting FLT3, or a mutant thereof, activity in a biological sample comprising the step of contacting said biological sample with a compound of this invention, or a composition comprising said compound.
  • the invention relates to a method of irreversibly inhibiting FLT3, or a mutant thereof, activity in a biological sample comprising the step of contacting said biological sample with a compound of this invention, or a composition comprising said compound.
  • the invention provides a method of simultaneously inhibiting both FLT3 and one or more other protein kinases selected from IRAK-1, IRAK-2, and IRAK-4, CDK8, CLK1, CLK2, or mutants thereof.
  • biological sample includes, without limitation, cell cultures or extracts thereof; biopsied material obtained from a mammal or extracts thereof; and blood, saliva, urine, feces, semen, tears, or other body fluids or extracts thereof.
  • Inhibition of protein kinase, or a protein kinase selected from FLT3, or a mutant thereof, activity in a biological sample is useful for a variety of purposes that are known to one of skill in the art. Examples of such purposes include, but are not limited to, blood transfusion, organ-transplantation, biological specimen storage, and biological assays.
  • Another embodiment of the present invention relates to a method of inhibiting protein kinase activity in a patient comprising the step of administering to said patient a compound of the present invention, or a composition comprising said compound.
  • the invention relates to a method of inhibiting activity of FLT3, or a mutant thereof, in a patient comprising the step of administering to said patient a compound of the present invention, or a composition comprising said compound.
  • the invention relates to a method of reversibly or irreversibly inhibiting one or more of FLT3, or a mutant thereof, activity in a patient comprising the step of administering to said patient a compound of the present invention, or a composition comprising said compound.
  • the present invention provides a method for treating a disorder mediated by FLT3, or a mutant thereof, in a patient in need thereof, comprising the step of administering to said patient a compound according to the present invention or pharmaceutically acceptable composition thereof.
  • a disorder mediated by FLT3, or a mutant thereof in a patient in need thereof, comprising the step of administering to said patient a compound according to the present invention or pharmaceutically acceptable composition thereof.
  • additional therapeutic agents that are normally administered to treat that condition may also be present in the compositions of this invention.
  • additional therapeutic agents that are normally administered to treat a particular disease, or condition are known as "appropriate for the disease, or condition, being treated.”
  • a compound of the current invention may also be used to advantage in combination with other therapeutic compounds.
  • the other therapeutic compounds are antiproliferative compounds.
  • antiproliferative compounds include, but are not limited to aromatase inhibitors; antiestrogens; topoisomerase I inhibitors; topoisomerase II inhibitors; microtubule active compounds; alkylating compounds; histone deacetylase inhibitors; compounds which induce cell differentiation processes; cyclooxygenase inhibitors; MMP inhibitors; mTOR inhibitors; antineoplastic antimetabolites; platin compounds; compounds targeting/decreasing a protein or lipid kinase activity and further anti-angiogenic compounds; compounds which target, decrease or inhibit the activity of a protein or lipid phosphatase; gonadorelin agonists; anti-androgens; methionine aminopeptidase inhibitors; matrix metalloproteinase inhibitors; bisphosphonates; biological response modifiers; antiproliferative
  • aromatase inhibitor as used herein relates to a compound which inhibits estrogen production, for instance, the conversion of the substrates androstenedione and testosterone to estrone and estradiol, respectively.
  • the term includes, but is not limited to steroids, especially atamestane, exemestane and formestane and, in particular, non-steroids, especially aminoglutethimide, roglethimide, pyridoglutethimide, trilostane, testolactone, ketokonazole, vorozole, fadrozole, anastrozole and letrozole.
  • Exemestane is marketed under the trade name AromasinTM.
  • Formestane is marketed under the trade name LentaronTM.
  • Fadrozole is marketed under the trade name AfemaTM.
  • Anastrozole is marketed under the trade name ArimidexTM.
  • Letrozole is marketed under the trade names FemaraTM or FemarTM.
  • Aminoglutethimide is marketed under the trade name OrimetenTM.
  • a combination of the invention comprising a chemotherapeutic agent which is an aromatase inhibitor is particularly useful for the treatment of hormone receptor positive tumors, such as breast tumors.
  • antiestrogen as used herein relates to a compound which antagonizes the effect of estrogens at the estrogen receptor level.
  • the term includes, but is not limited to tamoxifen, fulvestrant, raloxifene and raloxifene hydrochloride.
  • Tamoxifen is marketed under the trade name NolvadexTM.
  • Raloxifene hydrochloride is marketed under the trade name EvistaTM.
  • Fulvestrant can be administered under the trade name FaslodexTM.
  • a combination of the invention comprising a chemotherapeutic agent which is an antiestrogen is particularly useful for the treatment of estrogen receptor positive tumors, such as breast tumors.
  • anti-androgen as used herein relates to any substance which is capable of inhibiting the biological effects of androgenic hormones and includes, but is not limited to, bicalutamide (CasodexTM).
  • gonadorelin agonist as used herein includes, but is not limited to abarelix, goserelin and goserelin acetate. Goserelin can be administered under the trade name ZoladexTM.
  • topoisomerase I inhibitor includes, but is not limited to topotecan, gimatecan, irinotecan, camptothecian and its analogues, 9-nitrocamptothecin and the macromolecular camptothecin conjugate PNU-166148.
  • Irinotecan can be administered, e.g. in the form as it is marketed, e.g. under the trademark CamptosarTM.
  • Topotecan is marketed under the trade name HycamptinTM.
  • topoisomerase II inhibitor includes, but is not limited to the anthracyclines such as doxorubicin (including liposomal formulation, such as CaelyxTM), daunorubicin, epirubicin, idarubicin and nemorubicin, the anthraquinones mitoxantrone and losoxantrone, and the podophillotoxines etoposide and teniposide.
  • Etoposide is marketed under the trade name EtopophosTM.
  • Teniposide is marketed under the trade name VM 26-Bristol
  • Doxorubicin is marketed under the trade name AcriblastinTM or AdriamycinTM.
  • Epirubicin is marketed under the trade name FarmorubicinTM.
  • Idarubicin is marketed, under the trade name ZavedosTM.
  • Mitoxantrone is marketed under the trade name Novantron.
  • microtubule active agent relates to microtubule stabilizing, microtubule destabilizing compounds and microtublin polymerization inhibitors including, but not limited to taxanes, such as paclitaxel and docetaxel; vinca alkaloids, such as vinblastine or vinblastine sulfate, vincristine or vincristine sulfate, and vinorelbine; discodermolides; cochicine and epothilones and derivatives thereof.
  • Paclitaxel is marketed under the trade name TaxolTM.
  • Docetaxel is marketed under the trade name TaxotereTM.
  • Vinblastine sulfate is marketed under the trade name Vinblastin R.PTM.
  • Vincristine sulfate is marketed under the trade name FarmistinTM.
  • alkylating agent includes, but is not limited to, cyclophosphamide, ifosfamide, melphalan or nitrosourea (BCNU or Gliadel). Cyclophosphamide is marketed under the trade name CyclostinTM. Ifosfamide is marketed under the trade name HoloxanTM.
  • histone deacetylase inhibitors or "HDAC inhibitors” relates to compounds which inhibit the histone deacetylase and which possess antiproliferative activity. This includes, but is not limited to, suberoylanilide hydroxamic acid (SAHA).
  • SAHA suberoylanilide hydroxamic acid
  • antimetabolite includes, but is not limited to, 5-fluorouracil or 5-FU, capecitabine, gemcitabine, DNA demethylating compounds, such as 5-azacytidine and decitabine, methotrexate and edatrexate, and folic acid antagonists such as pemetrexed.
  • Capecitabine is marketed under the trade name XelodaTM.
  • Gemcitabine is marketed under the trade name GemzarTM.
  • platinum compound as used herein includes, but is not limited to, carboplatin, cis-platin, cisplatinum and oxaliplatin.
  • Carboplatin can be administered, e.g., in the form as it is marketed, e.g. under the trademark CarboplatTM.
  • Oxaliplatin can be administered, e.g., in the form as it is marketed, e.g. under the trademark EloxatinTM.
  • the term "compounds targeting/decreasing a protein or lipid kinase activity; or a protein or lipid phosphatase activity; or further anti-angiogenic compounds” as used herein includes, but is not limited to, protein tyrosine kinase and/or serine and/or threonine kinase inhibitors or lipid kinase inhibitors, such as a) compounds targeting, decreasing or inhibiting the activity of the platelet-derived growth factor-receptors (PDGFR), such as compounds which target, decrease or inhibit the activity of PDGFR, especially compounds which inhibit the PDGF receptor, such as an N-phenyl-2-pyrimidine-amine derivative, such as imatinib, SU101, SU6668 and GFB-111; b) compounds targeting, decreasing or inhibiting the activity of the fibroblast growth factor-receptors (FGFR); c) compounds targeting, decreasing or inhibiting the activity of the insulin-like growth factor receptor I (PDGFR),
  • BCR-Abl kinase and mutants, such as compounds which target decrease or inhibit the activity of c-Abl family members and their gene fusion products, such as an N-phenyl-2-pyrimidine-amine derivative, such as imatinib or nilotinib (AMN107); PD 180970; AG957; NSC 680410; PD 173955 from ParkeDavis; or dasatinib (BMS-354825); j) compounds targeting, decreasing or inhibiting the activity of members of the protein kinase C (PKC) and Raf family of serine/threonine kinases, members of the MEK, SRC, JAK/pan-JAK, FAK, PDK1, PKB/Akt, Ras/MAPK, PI3K, SYK, TYK2, BTK and TEC family, and/or members of the cyclin-dependent kinase family (CDK) including staurosporine derivatives, such as midostaurin;
  • c-Met receptor compounds which target, decrease or inhibit the activity of c-Met, especially compounds which inhibit the kinase activity of c-Met receptor, or antibodies that target the extracellular domain of c-Met or bind to HGF
  • PI3K inhibitor includes, but is not limited to compounds having inhibitory activity against one or more enzymes in the phosphatidylinositol-3 -kinase family, including, but not limited to ⁇ , ⁇ , ⁇ , ⁇ , PI3K-C2a, PBK-C2P, ⁇ - C2y, Vps34, pl lO-a, ⁇ 110- ⁇ , ⁇ 110- ⁇ , ⁇ 110- ⁇ , ⁇ 85- ⁇ , ⁇ 85- ⁇ , ⁇ 55- ⁇ , ⁇ 150, plOl, and ⁇ 87.
  • ⁇ inhibitors useful in this invention include but are not limited to ATU-027, SF- 1126, DS-7423, PBI-05204, GSK-2126458, ZSTK-474, buparlisib, pictrelisib, PF-4691502, BYL-719, dactolisib, XL-147, XL-765, and idelalisib.
  • BTK inhibitor includes, but is not limited to compounds having inhibitory activity against Bruton's Tyrosine Kinase (BTK), including, but not limited to AVL-292 and ibrutinib.
  • SYK inhibitor includes, but is not limited to compounds having inhibitory activity against spleen tyrosine kinase (SYK), including but not limited to PRT-062070, Pv-343, R-333, Excellair, PRT-062607, and fostamatinib
  • BTK inhibitory compounds and conditions treatable by such compounds in combination with compounds of this invention can be found in WO2008039218 and WO2011090760, the entirety of which are incorporated herein by reference.
  • PBK inhibitory compounds and conditions treatable by such compounds in combination with compounds of this invention can be found in WO2004019973, WO2004089925, WO2007016176, US8138347, WO2002088112, WO2007084786, WO2007129161, WO2006122806, WO2005113554, and WO2007044729 the entirety of which are incorporated herein by reference.
  • JAK inhibitory compounds and conditions treatable by such compounds in combination with compounds of this invention can be found in WO2009114512, WO2008109943, WO2007053452, WO2000142246, and WO2007070514, the entirety of which are incorporated herein by reference.
  • Further anti-angiogenic compounds include compounds having another mechanism for their activity, e.g. unrelated to protein or lipid kinase inhibition e.g. thalidomide (ThalomidTM) and TNP-470.
  • ThilomidTM thalidomide
  • TNP-470 TNP-470.
  • proteasome inhibitors useful for use in combination with compounds of the invention include, but are not limited to bortezomib, disulfiram, epigallocatechin-3-gallate (EGCG), salinosporamide A, carfilzomib, ONX-0912, CEP- 18770, and MLN9708.
  • Compounds which target, decrease or inhibit the activity of a protein or lipid phosphatase are e.g. inhibitors of phosphatase 1, phosphatase 2A, or CDC25, such as okadaic acid or a derivative thereof.
  • Compounds which induce cell differentiation processes include, but are not limited to, retinoic acid, ⁇ - ⁇ - or ⁇ - tocopherol or a- ⁇ - or ⁇ -tocotrienol.
  • cyclooxygenase inhibitor as used herein includes, but is not limited to, Cox- 2 inhibitors, 5-alkyl substituted 2-arylaminophenylacetic acid and derivatives, such as celecoxib (CelebrexTM), rofecoxib (VioxxTM), etoricoxib, valdecoxib or a 5-alkyl-2- arylaminophenylacetic acid, such as 5-methyl-2-(2'-chloro-6'-fluoroanilino)phenyl acetic acid, lumiracoxib.
  • Cox- 2 inhibitors such as celecoxib (CelebrexTM), rofecoxib (VioxxTM), etoricoxib, valdecoxib or a 5-alkyl-2- arylaminophenylacetic acid, such as 5-methyl-2-(2'-chloro-6'-fluoroanilino)phenyl acetic acid, lumiracoxib.
  • bisphosphonates includes, but is not limited to, etridonic, clodronic, tiludronic, pamidronic, alendronic, ibandronic, risedronic and zoledronic acid.
  • Etridonic acid is marketed under the trade name DidronelTM.
  • Clodronic acid is marketed under the trade name BonefosTM.
  • Tiludronic acid is marketed under the trade name SkelidTM.
  • Pamidronic acid is marketed under the trade name ArediaTM.
  • Alendronic acid is marketed under the trade name FosamaxTM.
  • Ibandronic acid is marketed under the trade name BondranatTM.
  • Risedronic acid is marketed under the trade name ActonelTM.
  • Zoledronic acid is marketed under the trade name ZometaTM.
  • mTOR inhibitors relates to compounds which inhibit the mammalian target of rapamycin (mTOR) and which possess antiproliferative activity such as sirolimus (Rapamune®), everolimus (CerticanTM), CCI-779 and ABT578.
  • heparanase inhibitor refers to compounds which target, decrease or inhibit heparin sulfate degradation.
  • the term includes, but is not limited to, PI-88.
  • biological response modifier refers to a lymphokine or interferons.
  • inhibitor of Ras oncogenic isoforms such as H-Ras, K-Ras, or N-Ras, as used herein refers to compounds which target, decrease or inhibit the oncogenic activity of Ras; for example, a "farnesyl transferase inhibitor” such as L-744832, DK8G557 or Rl 15777 (ZarnestraTM).
  • telomerase inhibitor refers to compounds which target, decrease or inhibit the activity of telomerase.
  • Compounds which target, decrease or inhibit the activity of telomerase are especially compounds which inhibit the telomerase receptor, such as telomestatin.
  • methionine ammopeptidase inhibitor refers to compounds which target, decrease or inhibit the activity of methionine ammopeptidase.
  • Compounds which target, decrease or inhibit the activity of methionine ammopeptidase include, but are not limited to, bengamide or a derivative thereof.
  • proteasome inhibitor refers to compounds which target, decrease or inhibit the activity of the proteasome.
  • Compounds which target, decrease or inhibit the activity of the proteasome include, but are not limited to, Bortezomib (VelcadeTM) and MLN 341.
  • matrix metalloproteinase inhibitor or (“MMP” inhibitor) as used herein includes, but is not limited to, collagen peptidomimetic and nonpeptidomimetic inhibitors, tetracycline derivatives, e.g. hydroxamate peptidomimetic inhibitor batimastat and its orally bioavailable analogue marimastat (BB-2516), prinomastat (AG3340), metastat (NSC 683551) BMS-279251 , BAY 12-9566, TAA211 , MMI270B or AAJ996.
  • MMP matrix metalloproteinase inhibitor
  • FMS-like tyrosine kinase inhibitors which are compounds targeting, decreasing or inhibiting the activity of FMS-like tyrosine kinase receptors (Flt-3R); interferon, ⁇ - ⁇ -D-arabinofuransylcytosine (ara-c) and bisulfan; and ALK inhibitors, which are compounds which target, decrease or inhibit anaplastic lymphoma kinase.
  • FMS-like tyrosine kinase receptors are especially compounds, proteins or antibodies which inhibit members of the Flt-3R receptor kinase family, such as PKC412, midostaurin, a staurosporine derivative, SU11248 and MLN518.
  • HSP90 inhibitors includes, but is not limited to, compounds targeting, decreasing or inhibiting the intrinsic ATPase activity of HSP90; degrading, targeting, decreasing or inhibiting the HSP90 client proteins via the ubiquitin proteosome pathway.
  • Compounds targeting, decreasing or inhibiting the intrinsic ATPase activity of HSP90 are especially compounds, proteins or antibodies which inhibit the ATPase activity of HSP90, such as 17-allylamino, 17-demethoxygeldanamycin (17AAG), a geldanamycin derivative; other geldanamycin related compounds; radicicol and HDAC inhibitors.
  • antiproliferative antibodies includes, but is not limited to, trastuzumab (HerceptinTM), Trastuzumab-DMl , erbitux, bevacizumab (AvastinTM), rituximab (Rituxan®), PR064553 (anti-CD40) and 2C4 Antibody.
  • antibodies is meant intact monoclonal antibodies, polyclonal antibodies, multispecific antibodies formed from at least 2 intact antibodies, and antibodies fragments so long as they exhibit the desired biological activity.
  • compounds of the current invention can be used in combination with standard leukemia therapies, especially in combination with therapies used for the treatment of AML.
  • compounds of the current invention can be administered in combination with, for example, farnesyl transferase inhibitors and/or other drugs useful for the treatment of AML, such as Daunorubicin, Adriamycin, Ara-C, VP- 16, Teniposide, Mitoxantrone, Idarubicin, Carboplatinum and PKC412.
  • the present invention provides a method of treating AML associated with an ITD and/or D835Y mutation, comprising administering a compound of the present invention together with a one or more additional FLT3 inhibitors.
  • the other FLT3 inhibitors are selected from quizartinib (AC220), a staurosporine derivative (e.g.
  • the other FLT3 inhibitors are selected from quizartinib, midostaurin, lestaurtinib, sorafenib, and sunitinib.
  • HDAC histone deacetylase
  • SAHA suberoylanilide hydroxamic acid
  • HDAC inhibitors include MS275, SAHA, FK228 (formerly FR901228), Trichostatin A and compounds disclosed in US 6,552,065 including, but not limited to, N-hydroxy-3-[4-[[[2-(2-methyl-lH-indol-3-yl)-ethyl]- amino]methyl]phenyl]-2E-2-propenamide, or a pharmaceutically acceptable salt thereof and N- hydroxy-3-[4-[(2-hydroxyethyl) ⁇ 2-(lH-indol-3-yl)ethyl]-amino]methyl]phenyl]-2E-2- propenamide, or a pharmaceutically acceptable salt thereof, especially the lactate salt.
  • Somatostatin receptor antagonists as used herein refer to compounds which target, treat or inhibit the somatostatin receptor such as octreotide, and SOM230.
  • Tumor cell damaging approaches refer to approaches such as ionizing radiation.
  • the term "ionizing radiation” referred to above and hereinafter means ionizing radiation that occurs as either electromagnetic rays (such as X- rays and gamma rays) or particles (such as alpha and beta particles). Ionizing radiation is provided in, but not limited to, radiation therapy and is known in the art. See Hellman, Principles of Radiation Therapy, Cancer, in Principles and Practice of Oncology, Devita et al, Eds., 4 th Edition, Vol. 1 , pp. 248-275 (1993).
  • EDG binders and ribonucleotide reductase inhibitors.
  • EDG binders refers to a class of immunosuppressants that modulates lymphocyte recirculation, such as FTY720.
  • ribonucleotide reductase inhibitors refers to pyrimidine or purine nucleoside analogs including, but not limited to, fludarabine and/or cytosine arabinoside (ara-C), 6-thioguanine, 5-fluorouracil, cladribine, 6-mercaptopurine (especially in combination with ara-C against ALL) and/or pentostatin.
  • Ribonucleotide reductase inhibitors are especially hydroxyurea or 2-hydroxy-lH-isoindole-l ,3-dione derivatives.
  • VEGF vascular endothelial growth factor
  • l-(4-chloroanilino)-4-(4-pyridylmethyl)phthalazine or a pharmaceutically acceptable salt thereof l-(4-chloroanilino)-4-(4-pyridylmethyl)phthalazine succinate
  • AngiostatinTM EndostatinTM
  • anthranilic acid amides ZD4190; ZD6474; SU5416; SU6668
  • bevacizumab or anti-VEGF antibodies or anti-VEGF receptor antibodies, such as rhuMAb and RHUFab
  • VEGF aptamer such as Macugon
  • FLT-4 inhibitors, FLT-3 inhibitors VEGFR-2 IgGI antibody
  • Angiozyme RI 4610)
  • Bevacizumab AvastinTM
  • Photodynamic therapy refers to therapy which uses certain chemicals known as photosensitizing compounds to treat or prevent cancers.
  • Examples of photodynamic therapy include treatment with compounds, such as VisudyneTM and porfimer sodium.
  • Angiostatic steroids refers to compounds which block or inhibit angiogenesis, such as, e.g., anecortave, triamcinolone, hydrocortisone, 11-a-epihydrocotisol, cortexolone, 17a-hydroxyprogesterone, corticosterone, desoxycorticosterone, testosterone, estrone and dexamethasone.
  • angiogenesis such as, e.g., anecortave, triamcinolone, hydrocortisone, 11-a-epihydrocotisol, cortexolone, 17a-hydroxyprogesterone, corticosterone, desoxycorticosterone, testosterone, estrone and dexamethasone.
  • Implants containing corticosteroids refers to compounds, such as fiuocinolone and dexamethasone.
  • chemotherapeutic compounds include, but are not limited to, plant alkaloids, hormonal compounds and antagonists; biological response modifiers, preferably lymphokines or interferons; antisense oligonucleotides or oligonucleotide derivatives; shRNA or siRNA; or miscellaneous compounds or compounds with other or unknown mechanism of action.
  • the compounds of the invention are also useful as co-therapeutic compounds for use in combination with other drug substances such as anti-inflammatory, bronchodilatory or antihistamine drug substances, particularly in the treatment of obstructive or inflammatory airways diseases such as those mentioned hereinbefore, for example as potentiators of therapeutic activity of such drugs or as a means of reducing required dosaging or potential side effects of such drugs.
  • a compound of the invention may be mixed with the other drug substance in a fixed pharmaceutical composition or it may be administered separately, before, simultaneously with or after the other drug substance.
  • the invention includes a combination of a compound of the invention as hereinbefore described with an antiinflammatory, bronchodilatory, antihistamine or anti-tussive drug substance, said compound of the invention and said drug substance being in the same or different pharmaceutical composition.
  • Suitable anti-inflammatory drugs include steroids, in particular glucocorticosteroids such as budesonide, beclamethasone dipropionate, fluticasone propionate, ciclesonide or mometasone furoate; non-steroidal glucocorticoid receptor agonists; LTB4 antagonists such LY293111, CGS025019C, CP-195543, SC-53228, BIIL 284, ONO 4057, SB 209247; LTD4 antagonists such as montelukast and zafirlukast; PDE4 inhibitors such cilomilast (Arifio® GlaxoSmithKline), Rofiumilast (Byk Gulden), V-l 1294A (Napp), BAY 19-8004 (Bayer), SCH- 351591 (Schering- Plough), Arofylline (Almirall Prodesfarma), PD 189659 / PD 168787 (Parke-
  • Suitable bronchodilatory drugs include anticholinergic or antimuscarinic compounds, in particular ipratropium bromide, oxitropium bromide, tiotropium salts and CHF 4226 (Chiesi), and glycopyrrolate.
  • Suitable antihistamine drug substances include cetirizine hydrochloride, acetaminophen, clemastine fumarate, promethazine, loratidine, desloratidine, diphenhydramine and fexofenadine hydrochloride, activastine, astemizole, azelastine, ebastine, epinastine, mizolastine and tefenadine.
  • chemokine receptors e.g. CCR-1 , CCR-2, CCR-3, CCR-4, CCR-5, CCR-6, CCR-7, CCR-8, CCR-9 and CCR10
  • CXCR1 , CXCR2, CXCR3, CXCR4, CXCR5, particularly CCR-5 antagonists such as Schering-Plough antagonists SC-351125, SCH- 55700 and SCH-D
  • Takeda antagonists such as N-[[4-[[[[6,7-dihydro-2-(4-methylphenyl)- 5H-benzo-cyclohepten-8-yl]carbonyl]amino]phenyl]-methyl]tetrahydro-N,N-dimethyl-2H- pyran-4- aminium chloride (TAK-770).
  • a compound of the current invention may also be used in combination with known therapeutic processes, for example, the administration of hormones or radiation.
  • a provided compound is used as a radiosensitizer, especially for the treatment of tumors which exhibit poor sensitivity to radiotherapy.
  • a compound of the current invention can be administered alone or in combination with one or more other therapeutic compounds, possible combination therapy taking the form of fixed combinations or the administration of a compound of the invention and one or more other therapeutic compounds being staggered or given independently of one another, or the combined administration of fixed combinations and one or more other therapeutic compounds.
  • a compound of the current invention can besides or in addition be administered especially for tumor therapy in combination with chemotherapy, radiotherapy, immunotherapy, phototherapy, surgical intervention, or a combination of these. Long-term therapy is equally possible as is adjuvant therapy in the context of other treatment strategies, as described above. Other possible treatments are therapy to maintain the patient's status after tumor regression, or even chemopreventive therapy, for example in patients at risk.
  • Those additional agents may be administered separately from an inventive compound-containing composition, as part of a multiple dosage regimen.
  • those agents may be part of a single dosage form, mixed together with a compound of this invention in a single composition. If administered as part of a multiple dosage regime, the two active agents may be submitted simultaneously, sequentially or within a period of time from one another normally within five hours from one another.
  • the term “combination,” “combined,” and related terms refers to the simultaneous or sequential administration of therapeutic agents in accordance with this invention.
  • a compound of the present invention may be administered with another therapeutic agent simultaneously or sequentially in separate unit dosage forms or together in a single unit dosage form.
  • the present invention provides a single unit dosage form comprising a compound of the current invention, an additional therapeutic agent, and a pharmaceutically acceptable carrier, adjuvant, or vehicle.
  • compositions of this invention should be formulated so that a dosage of between 0.01 - 100 mg/kg body weight/day of an inventive compound can be administered.
  • compositions which comprise an additional therapeutic agent that additional therapeutic agent and the compound of this invention may act synergistically. Therefore, the amount of additional therapeutic agent in such compositions will be less than that required in a monotherapy utilizing only that therapeutic agent. In such compositions a dosage of between 0.01 - 1,000 ⁇ g/kg body weight/day of the additional therapeutic agent can be administered.
  • the amount of additional therapeutic agent present in the compositions of this invention will be no more than the amount that would normally be administered in a composition comprising that therapeutic agent as the only active agent.
  • the amount of additional therapeutic agent in the presently disclosed compositions will range from about 50% to 100% of the amount normally present in a composition comprising that agent as the only therapeutically active agent.
  • the compounds of this invention, or pharmaceutical compositions thereof may also be incorporated into compositions for coating an implantable medical device, such as prostheses, artificial valves, vascular grafts, stents and catheters.
  • vascular stents for example, have been used to overcome restenosis (re -narrowing of the vessel wall after injury).
  • Implantable devices coated with a compound of this invention are another embodiment of the present invention.
  • the ee of 1.8 (98.5%>) and of 1.9 (100%) were determined by analytical chiral SFC under the following conditions: Column: phenomenex Lux 5u Cellulose-3, 4.6*250 mm, 5 ⁇ ; mobile phase: 90% C0 2 and 10% MeOH (0.01 DEA); flow rate: 4 mL/min; UV detection at 254 ⁇ .
  • Example 2 Synthesis of (12S)-3-[[(lr,4r)-4-(morpholin-4-yl)cyclohexyl]oxy]-8- thia-4,6-diazatricyclo[7.4.0.0[2,7]]trideca-l(9),2(7),3,5-tetraene-12-carboxylic acid. (1-1)
  • Example 4 Synthesis of 12-N-[4-(morpholin-4-yl)cyclohexyl]-10-N-[4-(piperidin- 4-yl)phenyl]-7-thia-9,ll-diazatricyclo[6.4.0.0[2,6]]dodeca-l(8),2(6),9,ll-tetraene-10,12- diamine hydrochloride (1-7).
  • Compound 5.1 was prepared in a manner analogous to intermediate 3.4, substituting l-N,l-N-dimethylcyclohexane-l,4,dimaine hydrochloride for 4-(morpholin-4-yl)cyclohexan-l- amine dihydrochloride. Isolated 5.1 (300 mg, 79%) as a yellow solid. MS (ES): m/z 252 (M+H) + .
  • Example 6 Synthesis of 12-N-[4-(dimethylamino)cyclohexyl]-10-N-[4-(piperidin- 4-yl)phenyl]-7-thia-9,ll-diazatricyclo[6.4.0.0[2,6]] dodeca-l(8),2(6),9,ll-tetraene-10,12- diamine hydrochloride (1-2).
  • Example 7 Synthesis of 12-N-[4-(dimethylamino)cyclohexyl]-10-N-[4-(4- methylpiperazin-l-yl)phenyl]-7-thia-9,ll-diazatricyclo[6.4.0.0[2,6]]dodeca-l(8),2(6),9,ll- tetraene-10,12-diamine (1-3).
  • Compound 1-3 was prepared from 5.1 and 7.1 in a manner analogous to the synthesis of 4.5 from 3.4 and 4.4. Isolated 20.5 mg (8%) of a white solid. MS (ES): m/z 506 (M+H) + .
  • Example 8 Synthesis of N4-((lr,4r)-4-(dimethylamino)cyclohexyl)-N2-(4-(l- methylpiperidin-4-yl)phenyl)-6,7-dihydro-5H-cyclopenta [4,5] thieno [2,3-d] pyrimidine-2,4- diamine (1-4).
  • Example 9 Synthesis of 10-N-[l-(propan-2-yl)-lH-pyrazol-4-yl]-12-N-[trans-4- (morpholin-4-yl)cyclohexyl]-7-thia-9,ll-diazatricyclo[6.4.0.0[2,6]]dodeca-l(8),2(6),9,ll- tetraene-10,12-diamine (1-9).
  • Example 10 Synthesis of 2-[(3S)-12-[[4-(dimethylamino)cyclohexyl]oxy]-10-[(l- methyl-lH-pyrazol-4-yl)amino]-7-thia-9, ll-diazatricyclo[6.4.0.0[2,6]]dodeca-l(8),2(6),9, 1 l-tetraen-3-yl] acetamide (1-23).
  • Example 11 Synthesis of 2-[(3R)-12-[[4-(dimethylamino)cyclohexyl]oxy]-10-[(l- methyl-lH-pyrazol-4-yl)amino]-7-thia-9, ll-diazatricyclo[6.4.0.0[2,6]]dodeca-l(8),2(6),9, 1 l-tetraen-3-yl] acetamide (I- 10).
  • Example 12 Synthesis of 2-[(3R)-10-(phenylamino)-12-[[(lr,4r)-4- (dimethylamino)cyclohexyl] oxy] -7-thia-9, 11-diazatricyclo [6.4.0.0 [2,6]] dodeca- 1 (8),2(6),9, ll-tetraen-3-yl] acetamide (1-6).
  • Example 13 Synthesis of 12-N-[4-(dimethylamino)cyclohexyl]-10-N-(l-methyl- lH-pyrazol-4-yl)-7-thia-9,ll-diazatricyclo[6.4.0.0[2,6]]dodeca-l(8),2(6),9,ll-tetraene-10,12- diamine (1-12).
  • Example 15 Synthesis of 12-[[4-(dimethylamino)cyclohexyl]oxy]-N-(l-methyl- lH-pyrazol-4-yl)-7-thia-9,ll-diazatricyclo[6.4.0.0[2,6]]dodeca-l(8),2(6),9,ll-tetraen-10- amine (1-14).
  • Example 16 Synthesis of 10-N-(l-ethyl-lH-pyrazol-4-yl)-12-N-[trans-4- (morpholin-4-yl)cyclohexyl]-7-thia-9,ll-diazatricyclo[6.4.0.0[2,6]]dodeca-l(8),2(6),9,ll- tetraene-10,12-diamine (1-8)
  • Compound 1-8 was prepared from 3.4 in a manner analogous to the synthesis of 1-9, substituting 1 -ethyl- lH-pyrazol-4-amine for l-(propan-2-yl)-lH-pyrazol-4-amine.
  • Example 17 Synthesis of 5-N-(l-methyl-lH-pyrazol-4-yl)-3-N-[trans-4- (morpholin-4-yl)cyclohexyl] - 1 l-oxa-8-thia-4,6-diazatricyclo [7.4.0.0 A [2,7] ] trideca- l(9),2(7),3,5-tetraene-3,5-diamine (1-13)
  • Example 18 Synthesis of 12-N-[4-(dimethylamino)cyclohexyl]-10-N-phenyl-7- thia-9,ll-di (1-15)
  • Example 19 Synthesis of 10-N-(l-methyl-lH-pyrazol-4-yl)-12-N-[4-(morpholin- 4-yl)cyclohexyl]-7-thia-9,ll-diazatricyclo [6.4.0.0 [2,6]]dodeca-l(8),2(6),9,ll-tetraene-10,12- diamine (1-16)
  • Compound 1-16 was prepared from 3.5 in a manner analogous to the synthesis of 4.5 from 4.4 and 3.4. Isolated 55.3 mg (47%) of 1-16 as a light yellow solid. MS (ES): m/z 454 (M+H) + .
  • Example 20 Synthesis of 12-N-[trans-4-(morpholin-4-yl)cyclohexyl]-10-N-(l,2- thiazol-4-yl)-7-thia-9,ll-diazatricyclo[6.4.0.0[2,6]]dodeca-l(8),2(6),9,ll-tetraene
  • Compound 1-18 was prepared from 3.4 in a manner analogous to the synthesis of 1-9, substituting 1 ,2-thiazol-4-amine hydrochloride for l-(propan-2-yl)-lH-pyrazol-4-amine. Isolated 116.9 mg (67%) of a light brown solid.
  • Example 21 Synthesis of N-(l-methyl-lH-pyrazol-4-yl)-12-[[4-(morpholin-4- yl)cyclohexyl] oxy] -7-thia-9,l 1-diazatricyclo [6.4.0.0 [2,6] ] dodeca- 1 (8),2(6),9, 11-tetraen- 10- amine. (1-19)
  • Example 22 Synthesis of 12-[[4-(dimethylamino)cyclohexyl]oxy]-N-phenyl-7- thia-9,ll-diazatri -l(8),2(6),9,ll-tetraen-10-amine. (1-20)
  • Example 23 Synthesis of 12-N-[trans-4-(morpholin-4-yl)cyclohexyl]-10-N-(l,2- oxazol-4-yl)-7-thia-9,ll-diazatricyclo[6.4.0.0[2,6]]dodeca-l(8),2(6),9,ll-tetraene-10,12- diamine (1-21) isopr
  • Compound 1-9 was prepared from 3.4 in a manner analogous to the synthesis of 1-9 from 3.4, substituting 1 ,2-oxazol-4-amine hydrochloride for l-(propan-2-yl)-lH-pyrazol-4- amine. Isolated 46.6 mg (28%) of a light brown solid.
  • Example 26 N-((lr,4r)-4-morpholinocyclohexyl)benzo[4,5]thieno[2,3- d]pyrimidin-4-amine (1-65).
  • Example 28 Synthesis of 2-(((lr,4r)-4-(benzo[4,5]thieno[2,3-d]pyrimidin-4- lamino)cyclohexyl)(methyl)amino)-l-(pyrrolidin-l-yl)ethanone. (1-67)
  • Example 29 Synthesis of (lr,4r)-4-(benzo[4,5]thieno[2,3-d]pyrimidin-4-yloxy)- ⁇ , ⁇ -dimethylcyclohexanamine. (1-68)
  • Example 36 Synthesis of (S)-2-hydroxy-3-((R)-4-(((lr,4R)-4-(pyrrolidin-l- yl)cyclohexyl)oxy)-6,7-dihydro-5H-cyclopenta [4,5] thieno [2,3-d] pyrimidin-5- yl)propanamide (1-41) and
  • Example 37 Synthesis of (R)-2-hydroxy-3-((R)-4-(((lr,4R)-4- (pyr rolidin- l-yl)cyclohexyl)oxy)-6,7-dihydro-5H-cyclopenta [4,5] thieno [2,3-d] pyrimidin-5- yl)propanamide (1-24)
  • Compound 1-26 was prepared in a manner analogous to Compound 1-25, substituting tert-butyl ((lr,4r)-4-hydroxycyclohexyl)(ethyl)carbamate for tert-butyl ((lr,4r)-4- hydroxycyclohexyl)(methyl)carbamate in the first step. Isolated 47.3 mg (22% overall yield) as a white solid. MS (ES): m/z 405 [M+H] + .
  • Example 40 Synthesis of (2S)-3-[(3R)-12-[[4-(dimethylamino)cyclohexyl]oxy]-7- thia-9,11-diazatricyclo [6.4.0.0 [2,6] ] dodeca-l(8),2(6),9,l l-tetraen-3-yl]-2- hydroxypropanamide formate (1-27)
  • Example 41 Synthesis of 3-[(3R)-12-[[4-(dimethylamino)cyclohexyl]oxy]-7-thia- 9,ll-diazatricyclo[6.4.0.0[2,6]]dodeca-l(8),2(6),9,ll-tetraen-3-yl]-2-hydroxypropanamide
  • Example 42 Synthesis of (2S)-3-[(3R)-12-[(4-aminocyclohexyl)oxy]-7-thia-9,ll- iazatricyclo [6.4.0.0 [2,6] ] dodeca-l(8),2(6),9,l l-tetraen-3-yl] -2-hydroxypropanamide (1-29)
  • Compound 1-29 was prepared in a manner analogous to Compound 1-25, substituting tert-butyl ((lr,4r)-4-hydroxycyclohexyl)carbamate for tert-butyl ((lr,4r)-4- hydroxycyclohexyl)(methyl)carbamate in the first step. Isolated 47.3 mg (22% overall yield) as a white solid. Isolated 30.9 mg (10% overall yield) as a white solid. MS (ES): m/z 377 [M+H] + .
  • Example 43 Synthesis of 3-[[4-(morpholin-4-yl)cyclohexyl]amino]-8-thia-4,6- diazatricyclo[7.4.0.0[2,7]]trideca-l(9),2,4,6,10,12-hexaene-12-carboxamide (1-30)
  • the crude product (50 mg) was purified by prep-HPLC with the following conditions (Waters): column: Xbridge Prep C 18, 5um, 19*50 mm; mobile phase: Water with 0.05% NH 4 HCO 3 and CH 3 CN (10% CH 3 CN up to 25% in 10 min, up to 95% in 1.5 min, down to 10% in 1.5 min); flow rate: 20 mL/min; UV detection at 254/220 nm. After collection and evaporation in vacuo, 1-30 (19 mg, 50%) was obtained as a white solid. MS: (ES, m/z) 412 [M+H] + .
  • Intermediate 44.1 was prepared from 2-oxocyclopentanane-l-carboxylate in a manner analogous to the synthesis of 34.1 from 1.1. Isolated 260 g (69%) of intermediate 44.1 as a white solid. Example 45: Synthesis of intermediate 45.1.
  • Example 46 Synthesis of 2-cyano-3-((R)-4-(((lr,4R)-4- morpholinocyclohexyl)oxy)-6,7-dihydro-5H-cyclopenta [4,5] thieno [2,3-d] pyrimidin-5- yl)propanamide (1-31)
  • Example 48 (2S)-2-hydroxy-3-[(3R)-12-[(4-[methyl[2-oxo-2-(pyrrolidin-l- yl)ethyl] amino] cyclohexyl)oxy]-7-thia-9,l 1-diazatricyclo [6.4.0.0 A [2,6] ] dodeca- l(8),2 6),9,ll-tetraen-3-yl]propanamide (1-33)
  • step 1 step 2
  • Example 51 Synthesis of (S)-2-hydroxy-3-((R)-4-(((lr,4R)-4- morpholinocyclohexyl)oxy)-6,7-dihydro-5H-cyclopenta [4,5] thieno [2,3-d] pyrimidin-5- yl)propanamide (1-34) and
  • Example 52 Synthesis of (R)-2-hydroxy-3-((R)-4-(((lr,4R)-4- morpholinocyclohexyl)oxy)-6,7-dihydro-5H-cyclopenta [4,5] thieno [2,3-d] pyrimidin-5- yl)propanamide (1-44)
  • Example 53 Synthesis of 2,2-difluoro-3-[(3R)-12-[[4-(morpholin-4- yl)cyclohexyl]oxy]-7-thia-9,ll-diazatricyclo[6.4.0.0[2,6]]dodeca-l(8),2(6),9,ll-tetraen-3- yl]propanamide (1-35)
  • Example 55 Synthesis of 3-[(35)-12-[[4-(dimethylamino)cyclohexyl]oxy]-7- thia-9,ll-diazatricyclo[6.4.0.0[2,6]]dodeca-l(8),2(6),9,ll-tetraen-3-yl]propanamide. -36)
  • Example 56 Synthesis of 3-[(3R)-12-[[4-(dimethylamino)cyclohexyl]oxy]-7-thia- 9,ll-diazatricyclo[6.4.0.0[2,6]]dodeca-l(8),2(6),9,ll-tetraen-3-yl]-2,2-difluoropropanamide -37)
  • Example 59 Synthesis of 2-(((lR,4r)-4-(((R)-5-(2-hydroxyethyl)-6,7-dihydro-5H- cyclopenta[4,5]thieno[2,3-d]pyrimidin-4-yl)oxy)cyclohexyl)(methyl)amino)-l-(pyrrolidin-l- yl)ethanone. (1-48)
  • Example 60 Synthesis of 3-((S)-4-(((lr,4S)-4-(methyl(2-oxo-2-(pyrrolidin-l- yl)ethyl)amino)cyclohexyl)oxy)-6,7-dihydr o-5H-cyclopenta [4,5] thieno [2,3-d] pyrimidin-5- yl)propanamide (1-40)
  • Example 62 Synthesis of 2-[(12R)-3-[[4-(dimethylamino)cyclohexyl]oxy]-8-thia- -diazatricyclo[7.4.0.0[2,7]]trideca-l(9),2(7),3,5-tetraen-12-yl]ethan-l-ol formate. (1-43)
  • Example 63 Synthesis of (S)-l-((R)-4-(((lr,4R)-4-morpholinocyclohexyl)oxy)- 6,7-dihydro-5H-cyclopenta [4,5] thieno [2,3-d] pyrimidin-5-yl)propan-2-ol (1-45)
  • Example 64 Synthesis of 2-[(4-[[(3R)-3-[(2R)-2-hydroxypropyl]- diazatricyclo [6.4.0.0 [2,6] ] dodeca-l(8),2(6),9,l l-tetraen-12- yl]oxy]cyclohexyl)(methyl)amino]-l-(pyrrolidin-l-yl)ethan-l-one. (1-46)
  • Example 65 Synthesis of 3-[[4-(dimethylamino)cyclohexyl]oxy]-8-thia-4,6, 12- triazatricyclo[7.4.0.0[2,7]]trideca-l(9),2(7),3,5-tetraene-12-sulfonamide. (1-47)
  • step 4 step 5
  • step 6 step 7
  • step 8 step 9
  • step 10 step 1 1
  • Example 66 Synthesis of (12S)-3-[[4-(morpholin-4-yl)cyclohexyl]oxy]-8-thia-4,6- diazatricyclo[7.4.0.0[2,7]]trideca-l(9),2(7),3,5-tetraene-12-carboxamide. (1-54)
  • the pH value of the solution was adjusted to 12 with 1 M aqueous NaOH solution and extracted with 3 x 50 mL of dichloromethane. The combined organic layers were washed with brine, dried over sodium sulfate and concentrated in vacuo. The residue was applied onto a silica gel column with DCM/MeOH (80: 1 to 50:1) to give the desired 67.2 (2.0 g) as a white solid.
  • Example 68 Synthesis of (12S)-3-[[4-(4,4-difluoropiperidin-l-yl)cyclohexyl]oxy]- 8-thia-4,6-diazatricyclo[7.4.0.0[2,7]]trideca-l(9),2(7),3,5-tetraene-12-carboxamide (I-51)
  • Example 69 Synthesis of (12S)-3-[[4-(dimethylamino)cyclohexyl]oxy]-8-thia-4,6- diazatricyclo[7.4.0.0[2,7]]trideca-l(9),2(7),3,5-tetraene-12-carboxamide. (1-49)
  • Example 70 Synthesis of 2-[(3R)-12-[[4-(4,4-difluoropiperidin-l- yl)cyclohexyl] oxy] -7-thia-9, 11-diazatricyclo [6.4.0.0 [2,6] ] dodeca-l(8),2(6),9, 1 l-tetraen-3- yl]acetamide. (1-50)
  • Example 71 Synthesis of 2-[(3R)-12-[[4-(piperidin-l-yl)cyclohexyl]oxy]-7-thia-9, -diazatricyclo [6.4.0.0 [2,6] ] dodeca-l(8),2(6),9, 1 l-tetraen-3-yl] acetamide. (1-52)
  • Example 73 Synthesis of 2-[(3R)-12-[(4-aminocyclohexyl)oxy]-7-thia-9,ll- diazatricyclo[6.4.0.0[2,6]]dodeca-l(12),2(6),8,10-tetraen-3-yl]acetamide (1-53)
  • Example 74 Synthesis of 2-[(3R)-12-[[4-(piperazin-l-yl)cyclohexyl]oxy]-7-thia- -diazatricyclo[6.4.0.0[2,6]]dodeca-l(12),2(6),8,10-tetra -3-yl]acetamide (I-55)
  • step 1 step 2
  • Example 75 Synthesis of 5-[[(3R)-12-[[4-(dimethylamino)cyclohexyl]oxy]-7-thia- 9,ll-diazatricyclo[6.4.0.0[2,6]]dodeca-l(8),2(6),9,ll-tetraen-3-yl]methyl]-l,3-oxazolidine- 2,4-dione (1-56)
  • the crude product (25 mg) was purified by preparative HPLC with the following conditions (Waters): column: Xbridge Prep CI 8, 5um, 19*50mm; mobile phase: water with 0.05% NH 4 HCO 3 and CH 3 CN (10% CH 3 CN up to 35%) in 10 min, up to 95% in 2.5 min, down to 10%> in 2.5 min); flow rate: 20mL/min; UV detection at 254/220 nm. After concentration of the fractions under reduced pressure, the desired 1-56 (19.8 mg, 74%) was obtained as a white solid.
  • LCMS (ES, m/z) 431 [M+H] + .
  • Example 76 Synthesis of 2-((R)-4-(((lr,4R)-4-morpholinocyclohexyl)oxy)-6,7- dihydro-5H-cyclopenta [4,5] thieno [2,3-d] pyrimidin-5-yl)acetamide (1-57)

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Abstract

The present invention provides methods of using compounds of formula I: or compositions thereof for the inhibition of FLT3, and the treatment of FLT3-mediated disorders.

Description

FLT3 INHIBITORS AND USES THEREOF
TECHNICAL FIELD OF THE INVENTION
[0001] The present invention relates to compounds and methods useful for inhibiting Fms- like tyrosine kinase 3 ("FLT3"), also known as Cluster of differentiation antigen 135 (CD 135) and fetal liver kinase 2 (Flk2). The invention also provides pharmaceutically acceptable compositions comprising compounds of the present invention and methods of using said compositions in the treatment of various disorders.
BACKGROUND OF THE INVENTION
[0002] The search for new therapeutic agents has been greatly aided in recent years by a better understanding of the structure of enzymes and other biomolecules associated with diseases. One important class of enzymes that has been the subject of extensive study is the protein kinase family.
[0003] Protein kinases constitute a large family of structurally related enzymes that are responsible for the control of a variety of signal transduction processes within the cell. Protein kinases are thought to have evolved from a common ancestral gene due to the conservation of their structure and catalytic function. Almost all kinases contain a similar 250-300 amino acid catalytic domain. The kinases may be categorized into families by the substrates they phosphorylate (e.g., protein-tyrosine, protein-serine/threonine, lipids, etc.).
[0004] In general, protein kinases mediate intracellular signaling by effecting a phosphoryl transfer from a nucleoside triphosphate to a protein acceptor that is involved in a signaling pathway. These phosphorylation events act as molecular on/off switches that can modulate or regulate the target protein biological function. These phosphorylation events are ultimately triggered in response to a variety of extracellular and other stimuli. Examples of such stimuli include environmental and chemical stress signals (e.g., osmotic shock, heat shock, ultraviolet radiation, bacterial endotoxin, and H202), cytokines (e.g., interleukin-1 (IL-1), interleukin-8 (IL- 8) and tumor necrosis factor a (TNF-a)), and growth factors (e.g., granulocyte macrophage- colony-stimulating factor (GM-CSF), and fibroblast growth factor (FGF)). An extracellular stimulus may affect one or more cellular responses related to cell growth, migration, differentiation, secretion of hormones, activation of transcription factors, muscle contraction, glucose metabolism, control of protein synthesis, and regulation of the cell cycle.
[0005] Many diseases are associated with abnormal cellular responses triggered by kinase- mediated events. These diseases include, but are not limited to, autoimmune diseases, inflammatory diseases, bone diseases, metabolic diseases, neurological and neurodegenerative diseases, cancer, cardiovascular diseases, allergies and asthma, Alzheimer's disease, and hormone-related diseases. Accordingly, there remains a need to find protein kinase inhibitors useful as therapeutic agents.
SUMMARY OF THE INVENTION
[0006] It has now been found that compounds of this invention, and pharmaceutically acceptable compositions thereof, are effective as inhibitors of FLT3 kinases. Such compounds have the general formula I:
Figure imgf000003_0001
I
or a pharmaceutically acceptable salt thereof, wherein each variable is as defined and described herein.
[0007] Compounds of the present invention, and pharmaceutically acceptable compositions thereof, are useful for treating a variety of diseases, disorders or conditions, associated with regulation of signaling pathways implicating FLT3 kinases. Such diseases, disorders, or conditions include those described herein.
[0008] Compounds provided by this invention are also useful for the study of FLT3 enzymes in biological and pathological phenomena; the study of intracellular signal transduction pathways occurring in bodily tissues; and the comparative evaluation of new FLT3 inhibitors or other regulators of kinases, signaling pathways, and cytokine levels in vitro or in vivo. DETAILED DESCRIPTION OF CERTAIN EMBODIMENTS
1. General Description of Certain Embodiments of the Invention:
[0009] Compounds of the present invention, and compositions thereof, are useful as inhibitors of FLT3 protein kinase.
[0010] The binding pocket of FLT3 contains a plurality of hydration sites, each of which is occupied by a single molecule of water. Each of these water molecules has a stability rating associated with it. As used herein, the term "stability rating" refers to a numerical calculation which incorporates the enthalpy, entropy, and free energy values associated with each water molecule. This stability rating allows for a measurable determination of the relative stability of water molecules that occupy hydration sites in the binding pocket of FLT3.
[0011] Water molecules occupying hydration sites in the binding pocket of FLT3 having a stability rating of >2.5 kcal/mol are referred to as "unstable waters."
[0012] Without wishing to be bound by any particular theory, it is believed that displacement or disruption of an unstable water molecule (i.e., a water molecule having a stability rating of >2.5 kcal/mol), , or replacement of a stable water (i.e., a water molecule having a stability rating of <1 kcal/mol), by an inhibitor results in tighter binding of that inhibitor. Accordingly, inhibitors designed to displace one or more unstable water molecules (i.e., those unstable water molecules not displaced by any known inhibitor) will be a tighter binder and, therefore, more potent inhibitor as compared to an inhibitor that does not displace unstable water molecules.
[0013] It was surprisingly found that provided compounds displace or disrupt one or more unstable water molecules. In some embodiments, a provided compound displaces or disrupts at least two unstable water molecules.
[0014] In certain embodiments, the present invention provides a method of inhibiting FLT3 kinase comprising contacting s a I:
Figure imgf000004_0001
I
or a pharmaceutically acceptable salt thereof, wherein: Ring A is a 3-7 membered saturated or partially unsaturated carbocyclic ring or a 4-7 membered saturated or partially unsaturated heterocyclic ring having 1-3 heteroatoms independently selected from nitrogen, oxygen, or sulfur;
n is 0-4;
each R1 is independently -R, halogen, -CN, -N02, -OR, -CH2OR, - SR, -N(R)2, -S02R, -S02N(R)2, -SOR, -C(0)R, -C02R, -C(0)N(R)2, -C(0)N(R)-OR, - NRC(0)OR, -NRC(0)N(R)2, Cy, or -NRS02R; or R1 is selected from one of the following formulas:
Figure imgf000005_0001
two R1 groups are taken together with their intervening atoms to form an optionally substituted 4-7 membered fused, spiro-fused, or bridged bicyclic ring having 0-2 heteroatoms independently selected from nitrogen, oxygen, or sulfur;
each Cy is an optionally substituted ring selected from a 3-7 membered saturated or partially unsaturated carbocyclic ring or a 4-7 membered saturated or partially unsaturated heterocyclic ring having 1-3 heteroatoms independently selected from nitrogen, oxygen, or sulfur;
each R is independently hydrogen, or an optionally substituted group selected from Ci_6 aliphatic, phenyl, 4-7 membered saturated or partially unsaturated heterocyclic having 1-2 heteroatoms independently selected from nitrogen, oxygen, or sulfur, or 5-6 membered heteroaryl ring having 1-4 heteroatoms independently selected from nitrogen, oxygen, or sulfur, or:
two R groups on the same nitrogen are taken together with their intervening atoms to form a 4-7 membered saturated, partially unsaturated, or heteroaryl ring having 0-3 heteroatoms, in addition to the nitrogen, independently selected from nitrogen, oxygen, or sulfur;
Ring B is a 4-8 membered partially unsaturated carbocyclic fused ring; a 4-7 membered partially unsaturated heterocyclic fused ring having 1-2 heteroatoms selected from nitrogen, oxygen, or sulfur; a benzo fused ring; or a 5-6 membered heteroaromatic fused ring having 1-3 heteroatoms independently selected from nitrogen, oxygen, and sulfur wherein said Ring B may be optionally substituted by one or more oxo, thiono, or imino groups; m is 0-4;
p is 0-2;
W is N or -C(R3)-;
Rz is R, CN, N02, halogen, -C(0)N(R)2, -C(0)OR, -C(0)R, -N(R)2, -NH-[Ar], - N(R)C(0)OR, -NRC(0)N(R)2, -OR, or -S02N(R)2;
R3 is hydrogen, halogen, -CN, Ci_4 aliphatic, Ci_4 haloaliphatic, -OR, -C(0)R, or -C(0)N(R)2; [Ar] is an optionally substituted phenyl or heteroaromatic ring;
L1 is a covalent bond or a Ci_6 bivalent hydrocarbon chain wherein one or two methylene units of the chain are optionally and independently replaced by -NR-, -N(R)C(0)-, -C(0)N(R)- , -N(R)S02-, -S02N(R)-, -0-, -C(O)-, -OC(O)-, -C(0)0-, -S-, -SO- or -S02-;
each L2 is independently a covalent bond or a Ci_6 bivalent hydrocarbon chain wherein one or two methylene units of the chain are optionally and independently replaced by -NR-, - N(R)C(0)-, -C(0)N(R)-, -N(R)S02-, -S02N(R)-, -0-, -C(O)-, -OC(O)-, -C(0)0-, -S-, -SO- or -S02-;
each R4 is independently halogen, -CN, -N02, -OR,
SR, -N(R)2, -S02R, -S02N(R)2, -SOR, -C(0)R, -C02R, -C(0)N(R)2, -NRC(0)R, - NRC(0)N(R)2, -C(0)N(R)OR, -N(R)C(0)OR, -N(R)S(0)2N(R)2, -NRS02R, or an optionally substituted group selected from Ci_6 aliphatic, phenyl, 4-7 membered saturated or partially unsaturated heterocyclic having 1-2 heteroatoms independently selected from nitrogen, oxygen, or sulfur, or 5-6 membered heteroaryl ring having 1-4 heteroatoms independently selected from nitrogen, oxygen, or sulfur, or:
two -L2(R4)P-R4 groups are taken together with their intervening atoms to form an optionally substituted 4-7 membered fused, spiro-fused, or bridged bicyclic ring having 0-2 heteroatoms independently selected from nitrogen, oxygen, or sulfur.
2. Compounds and Definitions:
[0015] Compounds of the present invention include those described generally herein, and are further illustrated by the classes, subclasses, and species disclosed herein. As used herein, the following definitions shall apply unless otherwise indicated. For purposes of this invention, the chemical elements are identified in accordance with the Periodic Table of the Elements, CAS version, Handbook of Chemistry and Physics, 75th Ed. Additionally, general principles of organic chemistry are described in "Organic Chemistry", Thomas Sorrell, University Science Books, Sausalito: 1999, and "March's Advanced Organic Chemistry", 5th Ed., Ed.: Smith, M.B. and March, J., John Wiley & Sons, New York: 2001, the entire contents of which are hereby incorporated by reference.
[0016] The term "aliphatic" or "aliphatic group", as used herein, means a straight-chain (i.e., unbranched) or branched, substituted or unsubstituted hydrocarbon chain that is completely saturated or that contains one or more units of unsaturation, or a monocyclic hydrocarbon or bicyclic hydrocarbon that is completely saturated or that contains one or more units of unsaturation, but which is not aromatic (also referred to herein as "carbocycle," "cycloaliphatic" or "cycloalkyl"), that has a single point of attachment to the rest of the molecule. Unless otherwise specified, aliphatic groups contain 1-6 aliphatic carbon atoms. In some embodiments, aliphatic groups contain 1-5 aliphatic carbon atoms. In other embodiments, aliphatic groups contain 1-4 aliphatic carbon atoms. In still other embodiments, aliphatic groups contain 1-3 aliphatic carbon atoms, and in yet other embodiments, aliphatic groups contain 1-2 aliphatic carbon atoms. In some embodiments, "cycloaliphatic" (or "carbocycle" or "cycloalkyl") refers to a monocyclic C3-C6 hydrocarbon that is completely saturated or that contains one or more units of unsaturation, but which is not aromatic, that has a single point of attachment to the rest of the molecule. Suitable aliphatic groups include, but are not limited to, linear or branched, substituted or unsubstituted alkyl, alkenyl, alkynyl groups and hybrids thereof such as (cycloalkyl)alkyl, (cycloalkenyl)alkyl or (cycloalkyl)alkenyl.
[0017] As used herein, the term "bridged bicyclic" refers to any bicyclic ring system, i.e. carbocyclic or heterocyclic, saturated or partially unsaturated, having at least one bridge. As defined by IUPAC, a "bridge" is an unbranched chain of atoms or an atom or a valence bond connecting two bridgeheads, where a "bridgehead" is any skeletal atom of the ring system which is bonded to three or more skeletal atoms (excluding hydrogen). In some embodiments, a bridged bicyclic group has 7-12 ring members and 0-4 heteroatoms independently selected from nitrogen, oxygen, or sulfur. Such bridged bicyclic groups are well known in the art and include those groups set forth below where each group is attached to the rest of the molecule at any substitutable carbon or nitrogen atom. Unless otherwise specified, a bridged bicyclic group is optionally substituted with one or more substituents as set forth for aliphatic groups. Additionally or alternatively, any substitutable nitrogen of a bridged bicyclic group is optionally substituted. Exemplary bridged bicyclics include:
Figure imgf000008_0001
[0018] The term "lower alkyl" refers to a Ci_4 straight or branched alkyl group. Exemplary lower alkyl groups are methyl, ethyl, propyl, isopropyl, butyl, isobutyl, and tert-butyl.
[0019] The term "lower haloalkyl" refers to a Ci_4 straight or branched alkyl group that is substituted with one or more halogen atoms.
[0020] The term "heteroatom" means one or more of oxygen, sulfur, nitrogen, phosphorus, or silicon (including, any oxidized form of nitrogen, sulfur, phosphorus, or silicon; the quaternized form of any basic nitrogen or; a substitutable nitrogen of a heterocyclic ring, for example N (as in 3,4-dihydro-2H-pyrrolyl), ΝΗ (as in pyrrolidinyl) or NR+ (as in N-substituted pyrrolidinyl)).
[0021] The term "unsaturated," as used herein, means that a moiety has one or more units of unsaturation.
[0022] As used herein, the term "bivalent Ci_8 (or Ci_6) saturated or unsaturated, straight or branched, hydrocarbon chain", refers to bivalent alkylene, alkenylene, and alkynylene chains that are straight or branched as defined herein. [0023] The term "alkylene" refers to a bivalent alkyl group. An "alkylene chain" is a polymethylene group, i.e., -(CH2)n-, wherein n is a positive integer, preferably from 1 to 6, from 1 to 4, from 1 to 3, from 1 to 2, or from 2 to 3. A substituted alkylene chain is a polymethylene group in which one or more methylene hydrogen atoms are replaced with a substituent. Suitable substituents include those described below for a substituted aliphatic group.
[0024] The term "alkenylene" refers to a bivalent alkenyl group. A substituted alkenylene chain is a polymethylene group containing at least one double bond in which one or more hydrogen atoms are replaced with a substituent. Suitable substituents include those described below for a substituted aliphatic group.
[0025] As used herein, the term "cyclopropylenyl" refers to a bivalent cyclopropyl group of the following structure:
Figure imgf000009_0001
[0026] The term "halogen" means F, CI, Br, or I.
[0027] The term "aryl" used alone or as part of a larger moiety as in "aralkyl," "aralkoxy," or "aryloxyalkyl," refers to monocyclic or bicyclic ring systems having a total of five to fourteen ring members, wherein at least one ring in the system is aromatic and wherein each ring in the system contains 3 to 7 ring members. The term "aryl" may be used interchangeably with the term "aryl ring." In certain embodiments of the present invention, "aryl" refers to an aromatic ring system which includes, but not limited to, phenyl, biphenyl, naphthyl, anthracyl and the like, which may bear one or more substituents. Also included within the scope of the term "aryl," as it is used herein, is a group in which an aromatic ring is fused to one or more non-aromatic rings, such as indanyl, phthalimidyl, naphthimidyl, phenanthridinyl, or tetrahydronaphthyl, and the like.
[0028] The terms "heteroaryl" and "heteroar-," used alone or as part of a larger moiety, e.g., "heteroaralkyl," or "heteroaralkoxy," refer to groups having 5 to 10 ring atoms, preferably 5, 6, or 9 ring atoms; having 6, 10, or 14 π electrons shared in a cyclic array; and having, in addition to carbon atoms, from one to five heteroatoms. The term "heteroatom" refers to nitrogen, oxygen, or sulfur, and includes any oxidized form of nitrogen or sulfur, and any quatemized form of a basic nitrogen. Heteroaryl groups include, without limitation, thienyl, furanyl, pyrrolyl, imidazolyl, pyrazolyl, triazolyl, tetrazolyl, oxazolyl, isoxazolyl, oxadiazolyl, thiazolyl, isothiazolyl, thiadiazolyl, pyridyl, pyridazinyl, pyrimidinyl, pyrazinyl, indolizinyl, purinyl, naphthyridinyl, and pteridinyl. The terms "heteroaryl" and "heteroar-", as used herein, also include groups in which a heteroaromatic ring is fused to one or more aryl, cycloaliphatic, or heterocyclyl rings, where the radical or point of attachment is on the heteroaromatic ring. Nonlimiting examples include indolyl, isoindolyl, benzothienyl, benzofuranyl, dibenzofuranyl, indazolyl, benzimidazolyl, benzthiazolyl, quinolyl, isoquinolyl, cinnolinyl, phthalazinyl, quinazolinyl, quinoxalinyl, 4H-quinolizinyl, carbazolyl, acridinyl, phenazinyl, phenothiazinyl, phenoxazinyl, tetrahydroquinolinyl, tetrahydroisoquinolinyl, and pyrido[2,3-b]-l,4-oxazin- 3(4H)-one. A heteroaryl group may be mono- or bicyclic. The term "heteroaryl" may be used interchangeably with the terms "heteroaryl ring," "heteroaryl group," or "heteroaromatic," any of which terms include rings that are optionally substituted. The term "heteroaralkyl" refers to an alkyl group substituted by a heteroaryl, wherein the alkyl and heteroaryl portions independently are optionally substituted.
[0029] As used herein, the terms "heterocycle," "heterocyclyl," "heterocyclic radical," and "heterocyclic ring" are used interchangeably and refer to a stable 5- to 7-membered monocyclic or 7-10-membered bicyclic heterocyclic moiety that is either saturated or partially unsaturated, and having, in addition to carbon atoms, one or more, preferably one to four, heteroatoms, as defined above. When used in reference to a ring atom of a heterocycle, the term "nitrogen" includes a substituted nitrogen. As an example, in a saturated or partially unsaturated ring having 0-3 heteroatoms selected from oxygen, sulfur or nitrogen, the nitrogen may be N (as in 3,4- dihydro-2H-pyrrolyl), ΝΗ (as in pyrrolidinyl), or (as in N-substituted pyrrolidinyl).
[0030] A heterocyclic ring can be attached to its pendant group at any heteroatom or carbon atom that results in a stable structure and any of the ring atoms can be optionally substituted. Examples of such saturated or partially unsaturated heterocyclic radicals include, without limitation, tetrahydrofuranyl, tetrahydrothiophenyl pyrrolidinyl, piperidinyl, pyrrolinyl, tetrahydroquinolinyl, tetrahydroisoquinolinyl, decahydroquinolinyl, oxazolidinyl, piperazinyl, dioxanyl, dioxolanyl, diazepinyl, oxazepinyl, thiazepinyl, morpholinyl, and quinuclidinyl. The terms "heterocycle," "heterocyclyl," "heterocyclyl ring," "heterocyclic group," "heterocyclic moiety," and "heterocyclic radical," are used interchangeably herein, and also include groups in which a heterocyclyl ring is fused to one or more aryl, heteroaryl, or cycloaliphatic rings, such as indolinyl, 3H-indolyl, chromanyl, phenanthridinyl, or tetrahydroquinolinyl. A heterocyclyl group may be mono- or bicyclic. The term "heterocyclylalkyl" refers to an alkyl group substituted by a heterocyclyl, wherein the alkyl and heterocyclyl portions independently are optionally substituted.
[0031] As used herein, the term "partially unsaturated" refers to a ring moiety that includes at least one double or triple bond. The term "partially unsaturated" is intended to encompass rings having multiple sites of unsaturation, but is not intended to include aryl or heteroaryl moieties, as herein defined.
[0032] As described herein, compounds of the invention may contain "optionally substituted" moieties. In general, the term "substituted," whether preceded by the term "optionally" or not, means that one or more hydrogens of the designated moiety are replaced with a suitable substituent. Unless otherwise indicated, an "optionally substituted" group may have a suitable substituent at each substitutable position of the group, and when more than one position in any given structure may be substituted with more than one substituent selected from a specified group, the substituent may be either the same or different at every position. Combinations of substituents envisioned by this invention are preferably those that result in the formation of stable or chemically feasible compounds. The term "stable," as used herein, refers to compounds that are not substantially altered when subjected to conditions to allow for their production, detection, and, in certain embodiments, their recovery, purification, and use for one or more of the purposes disclosed herein.
[0033] Suitable monovalent substituents on a substitutable carbon atom of an "optionally substituted" group are independently halogen; -(CH2)o-4R°; -(CH2)0^OR°; -O(CH2)0_4R°, -O- (CH2)0 4C(0)OR°; -(CH2)0 4CH(OR°)2; -(CH2)0^SR°; -(CH2)0^Ph, which may be substituted with R°;
Figure imgf000011_0001
which may be substituted with R°; -CH=CHPh, which may be substituted with R°; -(CH2)o-40(CH2)0-i-pyridyl which may be substituted with R°; -N02; -CN; -N3; -(CH2)o 4N(R°)2; -(CH2)0^N(Ro)C(O)R°; -N(R°)C(S)R°; -(CH2)0 4N(R°)C(0)NR°2; -N(R°)C(S)NR°2; -(CH2)0^N(Ro)C(O)OR°;
N(R°)N(R°)C(0)R°; -N(R°)N(R°)C(0)NR°2; -N(R°)N(R°)C(0)OR°; -(CH2)0 4C(0)R°; - C(S)R°; -(CH2)o 4C(0)OR°; -(CH2)0 4C(0)SR°; -(CH2)0^C(O)OSiR°3; -(CH2)0^OC(O)R°; - OC(0)(CH2)o 4SR- SC(S)SR°; -(CH2)0 4SC(0)R°; -(CH2)0 4C(0)NR°2; -C(S)NR°2; - C(S)SR°; -SC(S)SR°, -(CH2)0 4OC(0)NR°2; -C(0)N(OR°)R°; -C(0)C(0)R°; - C(0)CH2C(0)R°; -C(NOR°)R°; -(CH2)0 4SSR°; -(CH2)0^S(O)2R°; -(CH2)0 4S(0)2OR°; - (CH2)o 4OS(0)2R°; -S(0)2NR°2; -(CH2)0 4S(0)R°; -N(R°)S(0)2NR°2; -N(R°)S(0)2R°; - N(OR°)R°; -C(NH)NR°2; -P(0)2R°; -P(0)R°2; -OP(0)R°2; -OP(0)(OR°)2; SiR°3;
Figure imgf000012_0001
straight or branched alkylene)0-N(R°)2; or -(Ci_4 straight or branched alkylene)C(0)0-N(R°)2, wherein each R° may be substituted as defined below and is independently hydrogen, Ci_ 6 aliphatic, -CH2Ph, -O(CH2)0-iPh, -CH2-(5-6 membered heteroaryl ring), or a 5-6-membered saturated, partially unsaturated, or aryl ring having 0-4 heteroatoms independently selected from nitrogen, oxygen, or sulfur, or, notwithstanding the definition above, two independent occurrences of R°, taken together with their intervening atom(s), form a 3-12-membered saturated, partially unsaturated, or aryl mono- or bicyclic ring having 0-4 heteroatoms independently selected from nitrogen, oxygen, or sulfur, which may be substituted as defined below.
[0034] Suitable monovalent substituents on R° (or the ring formed by taking two independent occurrences of R° together with their intervening atoms), are independently halogen, -(CH2)0 2R*, -(haloR*), -(CH2)0 2OH, -(CH2)0 2OR*, -(CH2)0 2CH(OR*)2; -O(haloR'), -CN, -N3, -(CH2)0 2C(0)R*, -(CH2)0 2C(0)OH, -(CH2)0 2C(0)OR*, - (CH2)o 2SR*, -(CH2)o 2SH, -(CH2)0 2NH2, -(CH2)0 2NHR*, -(CH2)0 2NR*2, -N02, -SiR*3, - OSiR*3, -C(0)SR* -(Ci_4 straight or branched alkylene)C(0)OR*, or -SSR* wherein each R* is unsubstituted or where preceded by "halo" is substituted only with one or more halogens, and is independently selected from
Figure imgf000012_0002
aliphatic, -CH2Ph, -O(CH2)0-iPh, or a 5-6-membered saturated, partially unsaturated, or aryl ring having 0-4 heteroatoms independently selected from nitrogen, oxygen, or sulfur. Suitable divalent substituents on a saturated carbon atom of R° include =0 and =S.
[0035] Suitable divalent substituents on a saturated carbon atom of an "optionally substituted" group include the following: =0, =S, = NR* 2, =NNHC(0)R*, =NNHC(0)OR*, =NNHS(0)2R*, =NR*, =NOR*, -0(C(R* 2))2 30- or -S(C(R* 2))2_3S-, wherein each independent occurrence of R is selected from hydrogen, Ci_6 aliphatic which may be substituted as defined below, or an unsubstituted 5-6-membered saturated, partially unsaturated, or aryl ring having 0- 4 heteroatoms independently selected from nitrogen, oxygen, or sulfur. Suitable divalent substituents that are bound to vicinal substitutable carbons of an "optionally substituted" group include: -0(CR 2)2-30-, wherein each independent occurrence of R is selected from hydrogen, Ci_6 aliphatic which may be substituted as defined below, or an unsubstituted 5-6-membered saturated, partially unsaturated, or aryl ring having 0-4 heteroatoms independently selected from nitrogen, oxygen, or sulfur.
[0036] Suitable substituents on the aliphatic group of R include halogen, - R*, -(haloR*), -OH, -OR*, -O(haloR'), -CN, -C(0)OH, -C(0)OR*, -NH2, -NHR*, -NR*2, or
-N02, wherein each R* is unsubstituted or where preceded by "halo" is substituted only with one or more halogens, and is independently Ci_4 aliphatic, -CH2Ph, -O(CH2)0 iPh, or a 5-6- membered saturated, partially unsaturated, or aryl ring having 0-4 heteroatoms independently selected from nitrogen, oxygen, or sulfur.
[0037] Suitable substituents on a substitutable nitrogen of an "optionally substituted" group include -R†, -NR† 2, -C(0)R†, -C(0)OR†, -C(0)C(0)R†, C(0)CH2C(0)R†, -S(0)2R†, -S(0)2NR† 2, -C(S)NR† 2, -C(NH)NR† 2, or -N(R†)S(0)2R†; wherein each R† is independently hydrogen, Ci_6 aliphatic which may be substituted as defined below, unsubstituted -OPh, or an unsubstituted 5-6-membered saturated, partially unsaturated, or aryl ring having 0-4 heteroatoms independently selected from nitrogen, oxygen, or sulfur, or, notwithstanding the definition above, two independent occurrences of R^, taken together with their intervening atom(s) form an unsubstituted 3-12-membered saturated, partially unsaturated, or aryl mono- or bicyclic ring having 0-4 heteroatoms independently selected from nitrogen, oxygen, or sulfur.
[0038] Suitable substituents on the aliphatic group of R† are independently halogen, - R*, -(haloR*), -OH, -OR*, -O(haloR'), -CN, -C(0)OH, -C(0)OR*, -NH2, -NHR*, -NR*2, or -N02, wherein each R* is unsubstituted or where preceded by "halo" is substituted only with one or more halogens, and is independently Ci_4 aliphatic, -CH2Ph, -O(CH2)0 iPh, or a 5-6- membered saturated, partially unsaturated, or aryl ring having 0-4 heteroatoms independently selected from nitrogen, oxygen, or sulfur.
[0039] As used herein, the term "pharmaceutically acceptable salt" refers to those salts which are, within the scope of sound medical judgment, suitable for use in contact with the tissues of humans and lower animals without undue toxicity, irritation, allergic response and the like, and are commensurate with a reasonable benefit/risk ratio. Pharmaceutically acceptable salts are well known in the art. For example, S. M. Berge et al., describe pharmaceutically acceptable salts in detail in J. Pharmaceutical Sciences, 1977, 66, 1-19, incorporated herein by reference. Pharmaceutically acceptable salts of the compounds of this invention include those derived from suitable inorganic and organic acids and bases. Examples of pharmaceutically acceptable, nontoxic acid addition salts are salts of an amino group formed with inorganic acids such as hydrochloric acid, hydrobromic acid, phosphoric acid, sulfuric acid and perchloric acid or with organic acids such as acetic acid, oxalic acid, maleic acid, tartaric acid, citric acid, succinic acid or malonic acid or by using other methods used in the art such as ion exchange. Other pharmaceutically acceptable salts include adipate, alginate, ascorbate, aspartate, benzenesulfonate, benzoate, bisulfate, borate, butyrate, camphorate, camphorsulfonate, citrate, cyclopentanepropionate, digluconate, dodecylsulfate, ethanesulfonate, formate, fumarate, glucoheptonate, glycerophosphate, gluconate, hemisulfate, heptanoate, hexanoate, hydroiodide, 2-hydroxy-ethanesulfonate, lactobionate, lactate, laurate, lauryl sulfate, malate, maleate, malonate, methanesulfonate, 2-naphthalenesulfonate, nicotinate, nitrate, oleate, oxalate, palmitate, pamoate, pectinate, persulfate, 3-phenylpropionate, phosphate, pivalate, propionate, stearate, succinate, sulfate, tartrate, thiocyanate, p-toluenesulfonate, undecanoate, valerate salts, and the like.
[0040] Salts derived from appropriate bases include alkali metal, alkaline earth metal, ammonium and
Figure imgf000014_0001
salts. Representative alkali or alkaline earth metal salts include sodium, lithium, potassium, calcium, magnesium, and the like. Further pharmaceutically acceptable salts include, when appropriate, nontoxic ammonium, quaternary ammonium, and amine cations formed using counterions such as halide, hydroxide, carboxylate, sulfate, phosphate, nitrate, loweralkyl sulfonate and aryl sulfonate.
[0041] Unless otherwise stated, structures depicted herein are also meant to include all isomeric (e.g., enantiomeric, diastereomeric, and geometric (or conformational)) forms of the structure; for example, the R and S configurations for each asymmetric center, Z and E double bond isomers, and Z and E conformational isomers. Therefore, single stereochemical isomers as well as enantiomeric, diastereomeric, and geometric (or conformational) mixtures of the present compounds are within the scope of the invention. Unless otherwise stated, all tautomeric forms of the compounds of the invention are within the scope of the invention. Additionally, unless otherwise stated, structures depicted herein are also meant to include compounds that differ only in the presence of one or more isotopically enriched atoms. For example, compounds having the present structures including the replacement of hydrogen by deuterium or tritium, or the replacement of a carbon by a 13C- or 14C-enriched carbon are within the scope of this invention. Such compounds are useful, for example, as analytical tools, as probes in biological assays, or as therapeutic agents in accordance with the present invention. In certain embodiments, an R1 group of a provided compound comprises one or more deuterium atoms. In certain embodiments, Ring B of a provided compound may be substituted with one or more deuterium atoms.
[0042] As used herein, the term "inhibitor" is defined as a compound that binds to and /or inhibits FLT3 with measurable affinity. In certain embodiments, an inhibitor has an IC50 and/or binding constant of less than about 50 μΜ, less than about 1 μΜ, less than about 500 nM, less than about 100 nM, less than about 10 nM, or less than about 1 nM.
[0043] A compound of the present invention may be tethered to a detectable moiety. It will be appreciated that such compounds are useful as imaging agents. One of ordinary skill in the art will recognize that a detectable moiety may be attached to a provided compound via a suitable substituent. As used herein, the term "suitable substituent" refers to a moiety that is capable of covalent attachment to a detectable moiety. Such moieties are well known to one of ordinary skill in the art and include groups containing, e.g., a carboxylate moiety, an amino moiety, a thiol moiety, or a hydroxyl moiety, to name but a few. It will be appreciated that such moieties may be directly attached to a provided compound or via a tethering group, such as a bivalent saturated or unsaturated hydrocarbon chain. In some embodiments, such moieties may be attached via click chemistry. In some embodiments, such moieties may be attached via a 1,3-cycloaddition of an azide with an alkyne, optionally in the presence of a copper catalyst. Methods of using click chemistry are known in the art and include those described by Rostovtsev et al. , Angew. Chem. Int. Ed. 2002, 41, 2596-99 and Sun et al, Bioconjugate Chem., 2006, 17, 52-57.
[0044] As used herein, the term "detectable moiety" is used interchangeably with the term "label" and relates to any moiety capable of being detected, e.g., primary labels and secondary
32 33 35 14
labels. Primary labels, such as radioisotopes (e.g., tritium, P, P, S, or C), mass-tags, and fluorescent labels are signal generating reporter groups which can be detected without further modifications. Detectable moieties also include luminescent and phosphorescent groups.
[0045] The term "secondary label" as used herein refers to moieties such as biotin and various protein antigens that require the presence of a second intermediate for production of a detectable signal. For biotin, the secondary intermediate may include streptavidin-enzyme conjugates. For antigen labels, secondary intermediates may include antibody-enzyme conjugates. Some fluorescent groups act as secondary labels because they transfer energy to another group in the process of nonradiative fluorescent resonance energy transfer (FRET), and the second group produces the detected signal.
[0046] The terms "fluorescent label", "fluorescent dye", and "fluorophore" as used herein refer to moieties that absorb light energy at a defined excitation wavelength and emit light energy at a different wavelength. Examples of fluorescent labels include, but are not limited to: Alexa Fluor dyes (Alexa Fluor 350, Alexa Fluor 488, Alexa Fluor 532, Alexa Fluor 546, Alexa Fluor 568, Alexa Fluor 594, Alexa Fluor 633, Alexa Fluor 660 and Alexa Fluor 680), AMCA, AMCA-S, BODIPY dyes (BODIPY FL, BODIPY R6G, BODIPY TMR, BODIPY TR, BODIPY 530/550, BODIPY 558/568, BODIPY 564/570, BODIPY 576/589, BODIPY 581/591, BODIPY 630/650, BODIPY 650/665), Carboxyrhodamine 6G, carboxy-X-rhodamine (ROX), Cascade Blue, Cascade Yellow, Coumarin 343, Cyanine dyes (Cy3, Cy5, Cy3.5, Cy5.5), Dansyl, Dapoxyl, Dialkylaminocoumarin, 4',5'-Dichloro-2',7'-dimethoxy-fluorescein, DM-NERF, Eosin, Erythrosin, Fluorescein, FAM, Hydroxycoumarin, IRDyes (IRD40, IRD 700, IRD 800), JOE, Lissamine rhodamine B, Marina Blue, Methoxy coumarin, Naphtho fluorescein, Oregon Green 488, Oregon Green 500, Oregon Green 514, Pacific Blue, PyMPO, Pyrene, Rhodamine B, Rhodamine 6G, Rhodamine Green, Rhodamine Red, Rhodol Green, 2',4',5',7'-Tetra- bromosulfone-fluorescein, Tetramethyl-rhodamine (TMR), Carboxytetramethylrhodamine (TAMRA), Texas Red, Texas Red-X.
[0047] The term "mass-tag" as used herein refers to any moiety that is capable of being uniquely detected by virtue of its mass using mass spectrometry (MS) detection techniques. Examples of mass-tags include electrophore release tags such as N-[3-[4'-[(p- Methoxytetrafluorobenzyl)oxy]phenyl]-3-methylglyceronyl]isonipecotic Acid, 4 ' -[2,3 ,5 ,6- Tetrafluoro-4-(pentafluorophenoxyl)]methyl acetophenone, and their derivatives. The synthesis and utility of these mass-tags is described in United States Patents 4,650,750, 4,709,016, 5,360,8191, 5,516,931, 5,602,273, 5,604,104, 5,610,020, and 5,650,270. Other examples of mass-tags include, but are not limited to, nucleotides, dideoxynucleotides, oligonucleotides of varying length and base composition, oligopeptides, oligosaccharides, and other synthetic polymers of varying length and monomer composition. A large variety of organic molecules, both neutral and charged (biomolecules or synthetic compounds) of an appropriate mass range (100-2000 Daltons) may also be used as mass-tags. [0048] The terms "measurable affinity" and "measurably inhibit," as used herein, means a measurable change in a FLT3 protein kinase activity between a sample comprising a compound of the present invention, or composition thereof, and a FLT3 protein kinase, and an equivalent sample comprising an FLT3 protein kinase, in the absence of said compound, or composition thereof.
3. Description of Exemplary Embodiments:
[0049] As described above, in certain embodiments, the present invention provides a method of inhibiting FLT3 kinase, comprising contacting said kinase with a compound of formula I:
Figure imgf000017_0001
I
or a pharmaceutically acceptable salt thereof, wherein:
Ring A is a 3-7 membered saturated or partially unsaturated carbocyclic ring or a 4-7 membered saturated or partially unsaturated heterocyclic ring having 1-3 heteroatoms independently selected from nitrogen, oxygen, or sulfur;
n is 0-4;
each R1 is independently -R, halogen, -CN, -N02, -OR, -CH2OR, - SR, -N(R)2, -S02R, -S02N(R)2, -SOR, -C(0)R, -C02R, -C(0)N(R)2, -C(0)N(R)-OR, - NRC(0)OR, -NRC(0)N(R)2, Cy, or -NRS02R; or R1 is selected from one of the following formulas:
Figure imgf000017_0002
two R1 groups are taken together with their intervening atoms to form an optionally substituted 4-7 membered fused, spiro-fused, or bridged bicyclic ring having 0-2 heteroatoms independently selected from nitrogen, oxygen, or sulfur;
each Cy is an optionally substituted monocyclic or bicyclic ring selected from a 3-7 membered saturated or partially unsaturated carbocyclic ring or a 4-7 membered saturated or partially unsaturated heterocyclic monocyclic or bicyclic ring having 1-3 heteroatoms independently selected from nitrogen, oxygen, or sulfur;
each R is independently hydrogen, or an optionally substituted group selected from Ci_6 aliphatic, phenyl, 4-7 membered saturated or partially unsaturated heterocyclic having 1-2 heteroatoms independently selected from nitrogen, oxygen, or sulfur, or 5-6 membered heteroaryl ring having 1-4 heteroatoms independently selected from nitrogen, oxygen, or sulfur, or:
two R groups on the same nitrogen are taken together with their intervening atoms to form a 4-7 membered saturated, partially unsaturated, or heteroaryl ring having 0-3 heteroatoms, in addition to the nitrogen, independently selected from nitrogen, oxygen, or sulfur;
Ring B is a 4-8 membered partially unsaturated carbocyclic fused ring; a 4-7 membered partially unsaturated heterocyclic fused ring having 1-2 heteroatoms selected from nitrogen, oxygen, or sulfur; a benzo fused ring; or a 5-6 membered heteroaromatic fused ring having 1-3 heteroatoms independently selected from nitrogen, oxygen, and sulfur wherein said Ring B may be optionally substituted by one or more oxo, thiono, or imino groups;
m is 0-4;
p is 0-2;
W is N or -C(R3)-;
Rz is R, CN, N02, halogen, -C(0)N(R)2, -C(0)OR, -C(0)R, -N(R)2, -NH-[Ar], - N(R)C(0)OR, -NRC(0)N(R)2, -OR, or -S02N(R)2;
R3 is hydrogen, halogen, -CN, Ci_4 aliphatic, Ci_4 haloaliphatic, -OR, -C(0)R, or -C(0)N(R)2; [Ar] is an optionally substituted phenyl or heteroaromatic ring;
L1 is a covalent bond or a Ci_6 bivalent hydrocarbon chain wherein one or two methylene units of the chain are optionally and independently replaced by -NR-, -N(R)C(0)-, -C(0)N(R)- , -N(R)S02-, -S02N(R)-, -0-, -C(O)-, -OC(O)-, -C(0)0-, -S-, -SO- or -S02-;
each L2 is independently a covalent bond or a Ci_6 bivalent hydrocarbon chain wherein one or two methylene units of the chain are optionally and independently replaced by -NR-, - N(R)C(0)-, -C(0)N(R)-, -N(R)S02-, -S02N(R)-, -0-, -C(O)-, -OC(O)-, -C(0)0-, -S-, -SO- or -S02-; each R4 is independently halogen, -CN, -N02, -OR,
SR, -N(R)2, -S02R, -S02N(R)2, -SOR, -C(0)R, -C02R, -C(0)N(R)2, -NRC(0)R, - NRC(0)N(R)2, -C(0)N(R)OR, -N(R)C(0)OR, -N(R)S(0)2N(R)2, -NRS02R, or an optionally substituted group selected from Ci_6 aliphatic, phenyl, 4-7 membered saturated or partially unsaturated heterocyclic having 1-2 heteroatoms independently selected from nitrogen, oxygen, or sulfur, or 5-6 membered heteroaryl ring having 1-4 heteroatoms independently selected from nitrogen, oxygen, or sulfur, or:
two -L2(R4)P-R4 groups are taken together with their intervening atoms to form an optionally substituted 4-7 membered fused, spiro-fused, or bridged bicyclic ring having 0-2 heteroatoms independently selected from nitrogen, oxygen, or sulfur.
[0050] As defined generally above, the Ring A group of formula I is a 3-7 membered saturated or partially unsaturated carbocyclic ring or a 4-7 membered saturated or partially unsaturated heterocyclic ring having 1-3 heteroatoms independently selected from nitrogen, oxygen, or sulfur. In some embodiments, Ring A is a 3-7 membered saturated or partially unsaturated carbocyclic ring. In certain embodiments, Ring A is a 4-7 membered saturated or partially unsaturated heterocyclic ring having 1-3 heteroatoms independently selected from nitrogen, oxygen, or sulfur.
[0051] In some embodiments, Ring A is a 3-7 membered saturated carbocyclic ring. In certain embodiments, Ring A is cyclopentyl or cyclohexyl. In some embodiments, Ring A is cyclohexyl.
[0052] One of skill in the art will appreciate that a when Ring A is a disubstituted cycloalkyl ring, said ring can have cis or trans relative stereochemistry. In some embodiments, Ring A is a trans- 1 ,4-disubstituted cycloalkyl ring. In some embodiments, Ring A a trans- 1 ,4-disubstituted cyclohexyl ring.
[0053] In certain embodiments, Ring A is a 4-7 membered saturated heterocyclic ring having 1-3 heteroatoms independently selected from nitrogen, oxygen, or sulfur. In certain embodiments, Ring A is a 5-6 membered saturated heterocyclic ring having 1-3 heteroatoms independently selected from nitrogen, oxygen, or sulfur. In certain embodiments, Ring A is piperidinyl, piperazinyl, morpholinyl, thiomorpholinyl, tetrahydropyranyl, or tetrahydrofuranyl. In some embodiments, when Ring A is a 4-7 membered saturated heterocyclic ring, L1 is a covalent bond. In some embodiments, when Ring A is a 4-7 membered saturated heterocyclic ring, L1 is not a covalent bond.
[0054] As defined generally above, the n group of formula I is 0-4. In some embodiments, n is 0. In some embodiments, n is 1-4. In certain embodiments, n is 1. In some embodiments, n is 2.
[0055] As defined generally above, each R1 group of formula I is independently -R, halogen, -CN, -N02, -OR, -CH2OR, -SR, -N(R)2, -S02R, -S02N(R)2, -SOR, -C(0)R, -C02R, -C(0)N(R)2, -C(0)N(R)-OR, -NRC(0)R, -NRC(0)N(R)2, Cy, or - NRS02R; or R1 is selecte from one of the following formulas:
Figure imgf000020_0001
two R1 groups are taken together with their intervening atoms to form an optionally substituted 4-7 membered fused, spiro-fused, or bridged bicyclic ring having 0-2 heteroatoms independently selected from nitrogen, oxygen, or sulfur.
[0056] . In certain embodiments, R1 is R, -OR, -N(R)2, -C02R, -C(0)N(R)2, -C(0)N(R)- OR, -S02N(R)2, Cy, or -NRC(0)OR. In some embodiments, R1 is -C(0)NH2, -C(0)NHCH3, - C(0)NH-OH, -CH3, -CH2CH3, -S02t-butyl, -OH, -C(0)OH, -NH2, -NHCH3, -N(CH3)2, - -NHC(0)CH3, or -CH2phenyl. In certain embodiments, Rl is selected from one of
Figure imgf000020_0002
H2) -4 R jn certain embodiments, R1 is
Cy. In certain embodiments, R1 is -N(R)2. Exemplary R1 groups include those depicted in Table 1. In some embodiments R1 is R only where R is not hydrogen.
[0057] In some embodiments, the present invention provides a method utilizing a compound of formula I wherein two R1 groups are taken together with their intervening atoms to form an optionally substituted 4-7 membered fused, spiro-fused, or bridged bicyclic ring having 0-2 heteroatoms independently selected from nitrogen, oxygen, or sulfur. In certain embodiments, two R1 groups on adjacent carbon atoms are taken together to form an optionally substituted 4-7 membered ring fused to Ring A. In other embodiments, two R1 groups on the same carbon atom are taken together to form an optionally substituted 4-7 membered spiro-fused ring. In other embodiments, two R1 groups on non-adjacent carbon atoms are taken together to form an optionally substituted bridged bicyclic ring with Ring A.
[0058] As defined generally above, Cy is an optionally substituted monocyclic or bicyclic ring selected from a 3-7 membered saturated or partially unsaturated carbocyclic ring or a 4-7 membered saturated or partially unsaturated heterocyclic monocyclic or bicyclic ring having 1-3 heteroatoms independently selected from nitrogen, oxygen, or sulfur.
[0059] In some embodiments, Cy is a 3-7 membered saturated carbocyclic ring. In certain embodiments, Cy is a 4-7 membered saturated heterocyclic ring containing 1-2 heteroatoms independently selected from nitrogen, oxygen or sulfur. In certain embodiments Cy is a spirobicyclic 7-membered ring. In some embodiments, Cy is 2-oxa-6-azaspiro[3.3]heptane. In certain embodiments, Cy is morpholinyl, pyrrolidinyl, azetidinyl, piperidinyl or piperazinyl.
[0060] One of ordinary skill in the art will appreciate that an R1 substituent on a saturated carbon of Ring A forms a chiral center. In some embodiments, that chiral center is in the (R) configuration. In other embodiments, that chiral center is in the (S) configuration.
[0061] As defined generally above, the L1 group of formula I is a covalent bond or a Ci_6 bivalent hydrocarbon chain wherein one or two methylene units of the chain are optionally and independently replaced by -NR-, -N(R)C(0)-, -C(0)N(R)-, -N(R)S02-, -S02N(R)-, -0-, -C(O)-, -OC(O)-, -C(0)0-, -S-, -SO- or -S02-. In some embodiments, L1 is a covalent bond. In other embodiments, L1 is a Ci_6 bivalent hydrocarbon chain wherein one or two methylene units of the chain are optionally and independently replaced by -NR-, -N(R)C(0)-, -C(0)N(R)-, -N(R)S02- , -S02N(R)-, -0-, -C(O)-, -OC(O)-, -C(0)0-, -S-, -SO- or -S02-.
[0062] In some embodiments, L1 is -NH- (i.e., a Ci bivalent hydrocarbon chain wherein the methylene unit is replaced by -NH-), -0-, -CH20-, -OCH2-, -NHC(O)-, -CH2NH-, or -NHCH2-. In some embodiments, L1 is -0-. In some embodiments, L1 is -NR-. In some embodiments, L1 is -OCH2-. In some embodiments, L1 is -NRCH2-. Exemplary L1 groups include those depicted in Table 1.
[0063] As defined generally above, the Ring B group of formula I is a 4-8 membered partially unsaturated carbocyclic fused ring; a 4-7 membered partially unsaturated heterocyclic fused ring having 1-2 heteroatoms selected from nitrogen, oxygen, or sulfur; a benzo fused ring; or a 5-6 membered heteroaromatic fused ring having 1-3 heteroatoms independently selected from nitrogen, oxygen, and sulfur wherein said Ring B may be optionally substituted by one or more oxo, thiono, or imino groups.
[0064] . In some embodiments, Ring B is a 4-8 membered partially unsaturated carbocyclic fused ring. In other embodiments, Ring B is a 4-7 membered partially unsaturated azacyclic fused ring having one or two nitrogens. In some embodiments, Ring B is a cyclohexo- or cyclopento-fused ring. In other embodiments, Ring B is a piperidino-fused ring. In some embodiments, Ring B is a tetrahydropyrano-fused ring. In some embodiments, Ring B is a pyrrolidino-fused ring. In some embodiments, Ring B is a benzo fused ring. In certain embodiments, Ring B is a 5-6 membered heteroaromatic fused ring having 1-3 heteroatoms independently selected from nitrogen, oxygen and sulfur. In certain embodiments, Ring B is a 6 membered heteroaromatic fused ring having 1-2 nitrogens. In certain embodiments, Ring B is a pyrido fused ring, a pyrimidino fused ring, pyridazino or pyrazino fused ring. In certain embodiments, Ring B is a triazino fused ring. In certain embodiments, Ring B is a 5 membered heteroaromatic fused ring containing 1-2 heteroatoms independently selected from oxygen, nitrogen and sulfur. In some embodiments, Ring B is a pyrrolo fused ring, a thiopheno fused ring, a furano fused ring, a thiazolofused ring, an isothiazolo fused ring, an imidazolo fused ring, a pyrazolo fused ring, an oxazolo fused ring, an isoxazolo fused ring. In certain embodiments Ring B is a 5-6 membered heteroaromatic ring substituted with one or more oxo, thiono or imino groups.
[0065] One of ordinary skill in the art will appreciate that a substituent on a saturated carbon of Ring B forms a chiral center. In some embodiments, that chiral center is in the (R) configuration. In other embodiments, that chiral center is in the (S) configuration.
[0066] As defined generally above, the m group of formula I is 0-4. In some embodiments, m is 0. In some embodiments, m is 1-4. In some embodiments, m is 1.
[0067] As defined generally above, each L2 is independently a covalent bond or a Ci_6 bivalent hydrocarbon chain wherein one or two methylene units of the chain are optionally and independently replaced by -NR-, -N(R)C(0)-, -C(0)N(R)-, -N(R)S02-, -S02N(R)-, -0-, -C(O)-, -OC(O)-, -C(0)0-, -S-, -SO- or -S02-.
[0068] In certain embodiments each L2 is independently a covalent bond. In some embodiments each L2 is a Ci_3 bivalent hydrocarbon chain wherein one or two methylene units of the chain are optionally and independently replaced by -C(0)N(R)-, -0-, -C(O)-, -S-, -SO- or - SO2-. In certain embodiments, L2 is methylene. In certain embodiments, L2 is -CH2-C(0)-. In certain embodiments, L2 is a C2 hydrocarbon chain substituted with a hydroxyl group (-CH2CH(OH)-).
[0069] As defined generally above, each R4 is independently halogen, -CN, -N02, -OR, - SR, -N(R)2, -S02R, -S02N(R)2, -SOR, -C(0)R, -C02R, -C(0)N(R)2, -NRC(0)R, - NRC(0)N(R)2, -C(0)N(R)OR, -N(R)C(0)OR, -N(R)S(0)2N(R)2, -NRS02R, or an optionally substituted group selected from Ci_6 aliphatic, phenyl, 4-7 membered saturated or partially unsaturated heterocyclic having 1-2 heteroatoms independently selected from nitrogen, oxygen, or sulfur, or 5-6 membered heteroaryl ring having 1-4 heteroatoms independently selected from nitrogen, oxygen, or sulfur, or: two -L2(R4)P-R4 groups are taken together with their intervening atoms to form an optionally substituted 4-7 membered fused, spiro-fused, or bridged bicyclic ring having 0-2 heteroatoms independently selected from nitrogen, oxygen, or sulfur.
[0070] In some embodiments, each R4 is independently - CN, -OR, -SR, -SOR, -S02R, -C(0)N(R)2, -NRC(0)R, or an optionally substituted group selected from Ci_6 aliphatic, phenyl, 4-7 membered saturated or partially unsaturated heterocyclic having 1-2 heteroatoms independently selected from nitrogen, oxygen, or sulfur, or 5-6 membered heteroaryl ring having 1-4 heteroatoms independently selected from nitrogen, oxygen, or sulfur. In certain embodiments, each R4 is independently -
CN, -OR, -SR, -SOR, -S02R, -C(0)N(R)2, or -NRC(0)R. In certain embodiments R4 is an optionally substituted group selected from Ci_6 aliphatic, 4-7 membered saturated or partially unsaturated heterocyclic having 1-2 heteroatoms independently selected from nitrogen, oxygen, or sulfur, or 5-6 membered heteroaryl ring having 1-4 heteroatoms independently selected from nitrogen, oxygen, or sulfur. In certain embodiments R4 is hydroxyl. In certain embodiments R4 is -C(0)N(R)2.
[0071] In some embodiments, the present invention provides a method utilizing a compound of formula I wherein two -L2(R4)P-R4 groups are taken together with their intervening atoms to form an optionally substituted 4-7 membered fused, spiro-fused, or bridged bicyclic ring having 0-2 heteroatoms independently selected from nitrogen, oxygen, or sulfur. In certain embodiments, two -L2-R4 groups on adjacent carbon atoms are taken together to form an optionally substituted 4-7 membered ring fused to Ring B. In other embodiments, two -L2(R4)P-R4 groups on the same carbon atom are taken together to form an optionally substituted 4-7 membered spiro-fused ring. In other embodiments, two -L2(R4)P-R4 groups on non-adjacent carbon atoms are taken together to form an optionally substituted bridged bicyclic ring with Ring B.
[0072] In some embodiments, any one or more -L2(R4)P-R4 groups are independently selected from deuterium, an unsubstituted alkyl group, a -C02R group, and an unsubstituted heterocyclyl group. In some embodiments, any one or more -L2(R4)P-R4 groups are not independently selected from deuterium, an unsubstituted alkyl group a -C02R group, and an unsubstituted heterocyclyl group.
[0073] One of ordinary skill in the art will appreciate that an -L2(R4)P-R4 substituent on a saturated carbon of Ring B forms a chiral center. In some embodiments, that chiral center is in the (R) configuration. In other embodiments, that chiral center is in the (S) configuration.
[0074] As defined generally above, the Rz group of formula I is -R, -CN, -N02, halogen, - C(0)N(R)2, -C(0)OR, -C(0)R, -N(R)2, -NH-[Ar], -N(R)C(0)OR, -NRC(0)N(R)2, -OR, or - S02N(R)2. In some embodiments, Rz is hydrogen. In other embodiments, Rz is CN, halogen, -N(R)2 or -C(0)N(R)2. In some embodiments, Rz is -NH-[Ar]. Exemplary Rz groups include those depicted in Table 1.
[0075] As defined generally above, [Ar] is an optionally substituted phenyl or heteroaromatic ring. In some embodiments, [Ar] is an optionally substituted phenyl or 5-6 membered heteroaromatic ring. In some embodiments, [Ar] is an optionally substituted phenyl ring. In some embodiments, [Ar] is an optionally substituted heteroaromatic ring. In some embodiments, [Ar] is an optionally substituted 5-6 membered heteroaromatic ring. In some embodiments, [Ar] is an optionally substituted 5-membered heteroaromatic ring. In some embodiments, [Ar] is an optionally substituted 6-membered heteroaromatic ring. In some embodiments, [Ar] is an optionally substituted pyrazole ring.
[0076] As defined generally above, p is 0-2. In some embodiments p is 0. In some embodiments p is 1. In certain embodiments, p is 2.
[0077] In certain embodiments, a provided method employs a compound of formula I, wherein Ring B is a cyclopento fused ring, and W is N, thereby forming a compound of formula II:
Figure imgf000025_0001
or a pharmaceutically acceptable salt thereof, wherein each of Ring A, L1, L2, Rz, R1, R4, m, n, and p is as defined above and described in embodiments herein, both singly and in combination.
[0078] In certain embodiments, a provided method employs a compound of formula II, wherein R1 is one of the following formulas:
Figure imgf000025_0002
thereby forming a compound of formula Il-a or Il-
Figure imgf000025_0003
Il-a Il-b
or a pharmaceutically acceptable salt thereof, wherein each of Ring A, L1, L2, R, Rz, R1, R4, m, n, and p is as defined above and described in embodiments herein, both singly and in combination.
[0079] In certain embodiments, a provided method employs a compound of formula II, wherein m is 1, thereby forming a
Figure imgf000025_0004
III
or a pharmaceutically acceptable salt thereof, wherein each of Ring A, L1, L2, Rz, R1, R4, n, and p is as defined above and described in embodiments herein, both singly and in combination. [0080] In certain embodiments, a provided method employs a compound of formula III, wherein Ring A is cyclohexyl, thereby forming a compound of formula IV:
Figure imgf000026_0001
IV
or a pharmaceutically acceptable salt thereof, wherein each of, L1, L2, Rz, R1, R4, n, and p is as defined above and described in embodiments herein, both singly and in combination.
[0081] In certain embodiments, a provided method employs a compound of formula III, wherein n is 1 and the cyclohexyl ring has trans stereochemistry, thereby forming a compound of formula V:
Figure imgf000026_0002
V
or a pharmaceutically acceptable salt thereof, wherein each of, L1, L2, Rz, R1, R4, and p is as defined above and described in embodiments herein, both singly and in combination.
[0082] In certain embodiments, a provided present invention provides a compound of formula V, wherein Rz is -N(R)2, thereby forming a compound of formula VI:
Figure imgf000026_0003
VI
or a pharmaceutically acceptable salt thereof, wherein each of, L1, L2, R, R1, R4, and p is as defined above and described in embodiments herein, both singly and in combination. [0083] In certain embodiments, a provided method employss a compound of formula VII, wherein W 11, W2% X 11,
Figure imgf000027_0001
Y 1 , Y2 and Z 11 are each independently hydrogen or deuterium:
Figure imgf000027_0002
VII
or a pharmaceutically acceptable salt thereof, wherein each of L1, L2, R1, Rz, and R4 is as defined above for formula I and described in embodiments herein, both singly and in combination.
[0084] In certain embodiments, a provided method employs a compound of formula VIII, wherein W 11, W2% X11,
Figure imgf000027_0003
Z11 and Z 2 are each independently hydrogen or deuterium:
Figure imgf000027_0004
VIII
or a pharmaceutically acceptable salt thereof, wherein each of L1, L2, R1, Rz, and R4 are defined above for formula I and described in embodiments herein, both singly and in combination.
[0085] In certain embodiments, a provided method employs a compound of formula I, wherein Ring B is cyclohexo, W is N, and Rz is hydrogen, thereby forming a compound of formula IX:
Figure imgf000027_0005
IX
or a pharmaceutically acceptable salt thereof, wherein each of Ring A, L1, L2, R1, R4, m, n, and p is as defined above and described in embodiments herein, both singly and in combination. [0086] In certain embodiments, a provided method employs a compound of formula IX, wherein m is 1 , and L2 is attached a to the thiophene ring, thereby forming a compound of formula X:
Figure imgf000028_0001
X
or a pharmaceutically acceptable salt thereof, wherein each of Ring A, L1, L2, R1, R4, n, and p is as defined above and described in embodiments herein, both singly and in combination.
[0087] In certain embodiments, a provided method employs a compound of formula IX, wherein m is 1 , and L2 is attached β to the thiophene ring, thereby forming a compound of formula XI:
Figure imgf000028_0002
XI
or a pharmaceutically acceptable salt thereof, wherein each of Ring A, L1, L2, R1, R4, n, and p is as defined above and described in embodiments herein, both singly and in combination.
[0088] In certain embodiments, a provided method employs a compound of formula III, wherein Rz is hydrogen, and L2 is C2 alkylene, thereby forming a compound of formula XII:
Figure imgf000028_0003
or a pharmaceutically acceptable salt thereof, wherein each of Ring A, L1, R1, R4, n, and p is as defined above and described in embodiments herein, both singly and in combination. [0089] In certain embodiments, a provided method employs a compound of formula XII, wherein one instance of R4 is -C(0)NR2 , thereby forming a compound of formula XIII:
Figure imgf000029_0001
XIII
or a pharmaceutically acceptable salt thereof, wherein each of Ring A, L1, R, R1, R4, n, and p is as defined above and described in embodiments herein, both singly and in combination.
[0090] In certain embodiments, a provided method employs a compound of formula XIII, wherein Ring A is 4-substituted cyclohexyl, thereby forming a compound of formula XIV:
Figure imgf000029_0002
XIV
or a pharmaceutically acceptable salt thereof, wherein each of L1, R, R1, R4, n, and p is as defined above and described in embodiments herein, both singly and in combination.
[0091] In certain embodiments, a provided method employs a compound of formula XIV, wherein n is 1, and R1 is -NR2, thereby forming a compound of formula XV:
Figure imgf000029_0003
XV
or a pharmaceutically acceptable salt thereof, wherein each of L1, R, R4, and p is as defined above and described in embodiments herein, both singly and in combination. [0092] In certain embodiments, a provided method employs a compound of formula XV, wherein the stereochemistry of the substituent on the cyclopento ring is (R), and the relative stereochemistry on the cyclohexyl rin is trans thereby forming a compound of formula XVI:
Figure imgf000030_0001
XVI
or a pharmaceutically acceptable salt thereof, wherein each of L1, R, R4, and p is as defined above and described in embodiments herein, both singly and in combination.
[0093] In certain embodiments, a provided method employs a compound of formula XVI wherein each R4 is independently hydrogen, fluoro or -OR.
[0094] In certain embodiments a provided method employs a compound of formula XVI wherein L1 is -0-. In certain embodiments a provided method employs a compound of formula XVI wherein L1 is -NH-.
[0095] In certain embodiments, a provided method employs a compound of formula III, wherein Rz is hydrogen, and L2 is Ci alkylene, thereby forming a compound of formula XVII:
Figure imgf000030_0002
XVII
or a pharmaceutically acceptable salt thereof, wherein each of Ring A, L1, R1, R4, n, and p is as defined above and described in embodiments herein, both singly and in combination.
[0096] In certain embodiments, a provided method employs a compound of formula XVII, wherein one instance of R4 is -C(0)NR2 , thereby forming a compound of formula XVIII:
Figure imgf000031_0001
or a pharmaceutically acceptable salt thereof, wherein each of Ring A, L1, R, R1, R4, n, and p is as defined above and described in embodiments herein, both singly and in combination.
[0097] In certain embodiments, a provided method employs a compound of formula XVIII, wherein Ring A is 4-substituted c clohexyl, thereby forming a compound of formula XIX:
Figure imgf000031_0002
XIX
or a pharmaceutically acceptable salt thereof, wherein each of L1, R, R1, R4, n, and p is as defined above and described in embodiments herein, both singly and in combination.
[0098] In certain embodiments, a provided method employs a compound of formula XIX, wherein n is 1, and R1 is -NR2, thereby forming a compound of formula XX:
Figure imgf000031_0003
or a pharmaceutically acceptable salt thereof, wherein each of L1, R, R4, and p is as defined above and described in embodiments herein, both singly and in combination. [0099] In certain embodiments, a provided method employs a compound of formula XX, wherein the stereochemistry of the substituent on the cyclopento ring is (R), and the relative stereochemistry on the cyclohexyl ring is trans thereby forming a compound of formula XXI:
Figure imgf000032_0001
XXI
or a pharmaceutically acceptable salt thereof, wherein each of L1, R, R4, and p is as defined above and described in embodiments herein, both singly and in combination.
[00100] In certain embodiments, a provided method employs a compound of formula XXI wherein each R4 is independently hydrogen, fluoro or -OR.
[00101] In certain embodiments a provided method employs a compound of formula XXI wherein L1 is -0-. In certain embodiments a provided method employs a compound of formula XXI wherein L1 is -NH-.
[00102] In certain embodiments, a provided method employs a compound of formula I, wherein Ring B is piperidino, m is 1 , and Rz is hydrogen, thereby forming a compound of formula XXII:
Figure imgf000032_0002
XXII
or a pharmaceutically acceptable salt thereof, wherein each of Ring A, L1, L2, R1, R4, n, and p is as defined above and described in embodiments herein, both singly and in combination.
[00103] In certain embodiments, a provided method employs a compound of formula XXII, wherein L2 is a bond and p is 0, thereby forming a compound of formula XXIII:
Figure imgf000033_0001
or a pharmaceutically acceptable salt thereof, wherein each of Ring A, L1, R1, R4, n, and p is as defined above and described in embodiments herein, both singly and in combination.
[00104] In certain embodiments a provided method employs a compound of formula XXIII wherein R4 is -S(0)2R, -C(0)R, or -C(0)N(R)2.
[00105] In certain embodiments, a provided method employs a compound of formula II, wherein L1 is -0-, thereby forming a compound of formula XXIV:
Figure imgf000033_0002
XXIV
or a pharmaceutically acceptable salt thereof, wherein each of Ring A, L2, Rz, R1, R4, m, n, and p is as defined above and described in embodiments herein, both singly and in combination.
[00106] In certain embodiments, a provided method employs a compound of formula XXIV, wherein m is 1 , thereby forming a compound of formula XXV:
Figure imgf000033_0003
XXV
or a pharmaceutically acceptable salt thereof, wherein each of Ring A, L2, Rz, R1, R4, n, and p is as defined above and described in embodiments herein, both singly and in combination.
[00107] In certain embodiments, a provided method employs a compound of formula XXV, wherein Ring A is cyclohexyl, thereby forming a compound of formula XXVI:
Figure imgf000034_0001
XXVI
or a pharmaceutically acceptable salt thereof, wherein each of, L2, Rz, R1, R4, n, and p is as defined above and described in embodiments herein, both singly and in combination.
[00108] In certain embodiments, a provided method employs a compound of formula I, wherein Rz is -NH-[Ar], thereby forming a compound of formula XXVII:
Figure imgf000034_0002
XXVII
or a pharmaceutically acceptable salt thereof, wherein [Ar] is an optionally substituted phenyl or heteroaromatic ring, and each of Ring A, Ring B, L2, Rz, R1, R4, m, n, and p is as defined above and described in embodiments herein, both singly and in combination.
[00109] In certain embodiments, a provided method employs a compound of formula XXVII, wherein n is 1 and Ring A is 1 ,4-trans-substituted cyclohexyl, thereby forming a compound of formula XXVIII:
Figure imgf000034_0003
XXVIII
or a pharmaceutically acceptable salt thereof, wherein [Ar] is an optionally substituted phenyl or heteroaromatic ring, and each of Ring A, Ring B, L1, L2, R1, R4, m, and p is as defined above and described in embodiments herein, both singly and in combination. [00110] As described generally above, [Ar] is an optionally substituted phenyl or heteroaromatic ring. In some embodiments, [Ar] is optionally substituted phenyl. In some embodiments, [Ar] is an optionally substituted heteroaromatic ring. In some embodiments, [Ar] is an optionally substituted 5-membered heteroaromatic ring. In some embodiments, [Ar] is an optionally substituted 6-membered heteroaromatic ring. In some embodiments, [Ar] is an optionally substituted pyrazole ring.
[00111] In certain embodiments, a provided method employs a compound of formula
XXVIII, wherein Ring B is cyclopento, thereby forming a compound of formula XXIX:
Figure imgf000035_0001
XXIX
or a pharmaceutically acceptable salt thereof, wherein each of [Ar], L1, L2, R1, R4, m, and p is as defined above and described in embodiments herein, both singly and in combination.
[00112] In certain embodiments, a provided method employs a compound of formula XXVIII, wherein Ring B is cyclohexo, thereby forming a compound of formula XXIX:
Figure imgf000035_0002
XXIX
or a pharmaceutically acceptable salt thereof, wherein each of [Ar], L1, L2, R1, R4, m, and p is as defined above and described in embodiments herein, both singly and in combination.
[00113] In certain embodiments, a provided method employs a compound of formula XXVIII, wherein Ring B is a partially unsaturated tetrahydropyrano-fused ring, thereby forming a compound of one of formulae XXX-a, XXX-b, XXX-c, or XXX-d:
Figure imgf000036_0001
Figure imgf000036_0002
XXX-c XXX-d
or a pharmaceutically acceptable salt thereof, wherein each of [Ar], L1, L2, R1, R4, m, and defined above and described in embodiments herein, both singly and in combination.
[00114] In certain embodiments, a provided method employs a compound of formula XXVIII, wherein Ring B is a partially unsaturated piperidino-fused ring, thereby forming compound of one of formulae XXXI-a, XXXI-b, XXXI-c, or XXXI-d:
Figure imgf000036_0003
XXXI-c XXXI-d or a pharmaceutically acceptable salt thereof, wherein each of [Ar], L1, L2, R1, R4, m, and p is as defined above and described in embodiments herein, both singly and in combination.
[00115] In certain embodiments, a provided method employs a compound of formula
XXVIII, wherein Ring B is a partially unsaturated pyrrolidino-fused ring, thereby forming a compound of one of formulae XXXII-a, XXXII-b, or XXXII-c:
Figure imgf000037_0001
XXXII-a XXXII-b XXXII-c or a pharmaceutically acceptable salt thereof, wherein each of [Ar], L1, L2, R1, R4, m, and p is as defined above and described in embodiments herein, both singly and in combination.
[00116] In certain embodiments, a provided method employs a compound of formula I, wherein n is 1 , Ring A is trans-substituted cyclohexyl, and Ring B is a partially unsaturated tetrahydropyrano-fused ring, thereby forming a compound of one of formulae XXXIII-a, XXXIII-b, XXXIII-c, -d:
Figure imgf000037_0002
XXXIII-c XXXIII-d
or a pharmaceutically acceptable salt thereof, wherein each of L1, L2, R1, R4, Rz, m, and p is as defined above and described in embodiments herein, both singly and in combination. [00117] In certain embodiments, a provided method employs a compound of formula I, wherein n is 1 , Ring A is trans-substituted cyclohexyl, and Ring B is a partially unsaturated piperidino-fused ring, thereby forming a compound of one of formulae XXXIV-a, XXXIV-b, XXXIV-c, or XXXIV-
Figure imgf000038_0001
XXXIV-c XXXIV-d
or a pharmaceutically acceptable salt thereof, wherein each of L1, L2, R1, R4, Rz, m, and p is as defined above and described in embodiments herein, both singly and in combination.
[00118] In certain embodiments, a provided method employs a compound of formula I, wherein n is 1 , Ring A is trans-substituted cyclohexyl, and Ring B is a partially unsaturated pyrrolidino-fused ring, thereby forming a compound of one of formulae XXXV-a, XXXV-b, or XXXV-
Figure imgf000038_0002
XXXV-a XXXV-b XXXV-c or a pharmaceutically acceptable salt thereof, wherein each of L1, L2, R1, R4,RZ, m, and p is as defined above and described in embodiments herein, both singly and in combination. [00119] In certain embodiments, a provided method employs a compound of formula XXXIII-a, XXXIII-b, XXXIII-c, or XXXIII-d, wherein Rz is hydrogen, thereby forming a compound of one of d:
Figure imgf000039_0001
XXXVI-c XXXVI-d
or a pharmaceutically acceptable salt thereof, wherein each of L1, L2, R1, R4, m, and p is as defined above and described in embodiments herein, both singly and in combination.
[00120] In certain embodiments, a provided method employs a compound of formula XXXIV-a, XXXIV-b, XXXIV-c, or XXXIV-d, wherein Rz is hydrogen, thereby forming a compound of one of -d:
Figure imgf000039_0002
XXXVII-c XXXVII-d or a pharmaceutically acceptable salt thereof, wherein each of L1, L2, R1, R4, m, and p is as defined above and described in embodiments herein, both singly and in combination.
[00121] In certain embodiments, a provided method employs a compound of formula XXXV- a, XXXV-b, or XXXV-c, wherein Rz is hydrogen, thereby forming a compound of one of formulae -a, XXXVIII-b, -c:
Figure imgf000040_0001
XXXVIII-a XXXVIII-b XXXVIII-c or a pharmaceutically acceptable salt thereof, wherein each of L1, L2, R1, R4, m, and p is as defined above and described in embodiments herein, both singly and in combination.
[00122] In certain embodiments, a provided method employs a compound of formula I, wherein W is N, and Rz is hydrogen, thereby forming a compound of formula I-a:
Figure imgf000040_0002
I-a
or a pharmaceutically acceptable salt thereof, wherein each of Ring A, Ring B, L1, L2, W, Rz, R1, R4 , m, n, and p is as defined above and described in embodiments herein, both singly and in combination.
[00123] In certain embodiments, a provided method employs a compound of formula I, wherein Ring B is a benzo fused ring, thereby forming a compound of formula XXXIX:
Figure imgf000040_0003
XXXIX
or a pharmaceutically acceptable salt thereof, wherein each of Ring A, L1, L2, Rz, R1, R4, W, m, n, and p is as defined above and described in embodiments herein, both singly and in
combination.
[00124] In certain embodiments, a provided method employs a compound of formula
XXXIX, wherein R1 is one of the following formulas:
Figure imgf000041_0001
thereby forming a compound of formula XXXIX-a -b:
Figure imgf000041_0002
XXXIX-a XXXIX-b or a pharmaceutically acceptable salt thereof, wherein each of Ring A, L1, L2, Rz, R4, W, m, n, and p is as defined above and described in embodiments herein, both singly and in combination.
[00125] In certain embodiments, a provided method employs a compound of formula
XXXIX, wherein m is 1, and -L2(R4)P-R4 is connected para to Q, thereby forming a compound of formula XXXX:
Figure imgf000041_0003
XXXX
or a pharmaceutically acceptable salt thereof, wherein each of Ring A, L1, L2, Rz, R1, R4, W, n, and p is as defined above and described in embodiments herein, both singly and in combination.
[00126] In certain embodiments, a provided method employs a compound of formula XXXX, wherein Ring A is cyclohexyl, thereby forming a compound of formula XXXXI:
Figure imgf000042_0001
XXXXI
or a pharmaceutically acceptable salt thereof, wherein each of L1, L2, Rz, R1, R4, W, n, and p is as defined above and described in embodiments herein, both singly and in combination.
[00127] In certain embodiments, a provided method employs a compound of formula
XXXXI, wherein n is 1 and the cyclohexyl ring has trans stereochemistry, thereby forming a compound of formula XXXXII:
Figure imgf000042_0002
XXXXII
or a pharmaceutically acceptable salt thereof, wherein each of L1, L2, Rz, R1, R4, W, and p is as defined above and described in embodiments herein, both singly and in combination.
[00128] In certain embodiments, a provided method employs a compound of formula XXXX, wherein Rz is hydrogen, and L2 is Ci alkylene, thereby forming a compound of formula
XXXXIII:
Figure imgf000042_0003
XXXXIII
or a pharmaceutically acceptable salt thereof, wherein each of Ring A, L1, R1, R4, W, n, and p is as defined above and described in embodiments herein, both singly and in combination.
[00129] In certain embodiments, a provided method employs a compound of formula XXXX, wherein L2 is a covalent bond, thereby forming a compound of formula XXXXIV:
Figure imgf000043_0001
XXXXIV
or a pharmaceutically acceptable salt thereof, wherein each of Ring A, L1, R1, R4, W, and n is as defined above and described in embodiments herein, both singly and in combination.
[00130] In certain embodiments, a provided method employs a compound of formula I, wherein Ring B is a 6 membered heteroaromatic fused ring having 1-3 nitrogen atoms.
[00131] In certain embodiments, a provided method employs a compound of formula I or a pharmaceutically acceptable salt thereof, wherein Ring B is a 6-membered heteroaromatic fused ring having 1-2 nitrogen atoms wherein Ring B is substituted with one or more oxo, thiono, or imino groups.
[00132] In certain embodiments, the method employs a compound of formula I, wherein Ring B is a 5 membered heteroaryl fused ring having 1-3 heteroatoms independently selected from nitrogen, oxygen, and sulfur.
[00133] In certain embodiments, a provided method employs a compound of formula I or a pharmaceutically acceptable salt thereof, wherein Ring B is a 5 -membered heteroaryl fused ring having 1 heteroatom selected from nitrogen, oxygen, and sulfur, wherein Ring B is substituted with one or more oxo, thiono, or imino groups.
[00134] Exemplary compounds of the invention are set forth in Table 1, below.
Table 1. Exemplary Compounds
Figure imgf000044_0001
Figure imgf000045_0001
Figure imgf000046_0001
Figure imgf000047_0001
Figure imgf000048_0001
Figure imgf000049_0001
Figure imgf000050_0001
Figure imgf000051_0001
Figure imgf000052_0001
Figure imgf000053_0001
Figure imgf000054_0001
Figure imgf000055_0001
Figure imgf000056_0001
[00135] In some embodiments, the method employs a compound set forth in Table 1, above, or a pharmaceutically acceptable salt thereof.
[00136] Without wishing to be bound by any particular theory, it is believed that proximity of an inhibitor compound, or pendant moiety of an inhibitor compound, to the water of interest facilitates displacement or disruption of that water by the inhibitor compound, or pendant moiety of an inhibitor compound. In some embodiments, a water molecule displaced or disrupted by an inhibitor compound, or pendant moiety of an inhibitor compound, is an unstable water molecule.
[00137] In certain embodiments, the method employs a complex comprising FLT3 and an inhibitor, wherein at least one unstable water of FLT3 is displaced or disrupted by the inhibitor. In some embodiments, at least two unstable waters selected are displaced or disrupted by the inhibitor.
4. General Methods of Providing the Present Compounds [00138] The compounds of this invention may be prepared or isolated in general by synthetic and/or semi-synthetic methods known to those skilled in the art for analogous compounds and by methods described in detail in the Examples, herein.
5. Uses, Formulation and Administration
Pharmaceutically acceptable compositions
[00139] According to another embodiment, the invention provides a composition comprising a compound of this invention or a pharmaceutically acceptable derivative thereof and a pharmaceutically acceptable carrier, adjuvant, or vehicle. The amount of compound in compositions of this invention is such that is effective to measurably inhibit a FLT3 protein kinase, or a mutant thereof, in a biological sample or in a patient. In certain embodiments, the amount of compound in compositions of this invention is such that is effective to measurably inhibit a FLT3 protein kinase, or a mutant thereof, in a biological sample or in a patient. In certain embodiments, a composition of this invention is formulated for administration to a patient in need of such composition. In some embodiments, a composition of this invention is formulated for oral administration to a patient.
[00140] The term "patient," as used herein, means an animal, preferably a mammal, and most preferably a human.
[00141] The term "pharmaceutically acceptable carrier, adjuvant, or vehicle" refers to a nontoxic carrier, adjuvant, or vehicle that does not destroy the pharmacological activity of the compound with which it is formulated. Pharmaceutically acceptable carriers, adjuvants or vehicles that may be used in the compositions of this invention include, but are not limited to, ion exchangers, alumina, aluminum stearate, lecithin, serum proteins, such as human serum albumin, buffer substances such as phosphates, glycine, sorbic acid, potassium sorbate, partial glyceride mixtures of saturated vegetable fatty acids, water, salts or electrolytes, such as protamine sulfate, disodium hydrogen phosphate, potassium hydrogen phosphate, sodium chloride, zinc salts, colloidal silica, magnesium trisilicate, polyvinyl pyrrolidone, cellulose-based substances, polyethylene glycol, sodium carboxymethylcellulose, polyacrylates, waxes, polyethylene-polyoxypropylene-block polymers, polyethylene glycol and wool fat.
[00142] A "pharmaceutically acceptable derivative" means any non-toxic salt, ester, salt of an ester or other derivative of a compound of this invention that, upon administration to a recipient, is capable of providing, either directly or indirectly, a compound of this invention or an inhibitorily active metabolite or residue thereof.
[00143] As used herein, the term "inhibitorily active metabolite or residue thereof means that a metabolite or residue thereof is also an inhibitor of a FLT3 protein kinase, or a mutant thereof.
[00144] Compositions of the present invention may be administered orally, parenterally, by inhalation spray, topically, rectally, nasally, buccally, vaginally or via an implanted reservoir. The term "parenteral" as used herein includes subcutaneous, intravenous, intramuscular, intraarticular, intra-synovial, intrasternal, intrathecal, intrahepatic, intralesional and intracranial injection or infusion techniques. Preferably, the compositions are administered orally, intraperitoneally or intravenously. Sterile injectable forms of the compositions of this invention may be aqueous or oleaginous suspension. These suspensions may be formulated according to techniques known in the art using suitable dispersing or wetting agents and suspending agents. The sterile injectable preparation may also be a sterile injectable solution or suspension in a nontoxic parenterally acceptable diluent or solvent, for example as a solution in 1,3-butanediol. Among the acceptable vehicles and solvents that may be employed are water, Ringer's solution and isotonic sodium chloride solution. In addition, sterile, fixed oils are conventionally employed as a solvent or suspending medium.
[00145] For this purpose, any bland fixed oil may be employed including synthetic mono- or di-glycerides. Fatty acids, such as oleic acid and its glyceride derivatives are useful in the preparation of injectables, as are natural pharmaceutically-acceptable oils, such as olive oil or castor oil, especially in their polyoxyethylated versions. These oil solutions or suspensions may also contain a long-chain alcohol diluent or dispersant, such as carboxymethyl cellulose or similar dispersing agents that are commonly used in the formulation of pharmaceutically acceptable dosage forms including emulsions and suspensions. Other commonly used surfactants, such as Tweens, Spans and other emulsifying agents or bioavailability enhancers which are commonly used in the manufacture of pharmaceutically acceptable solid, liquid, or other dosage forms may also be used for the purposes of formulation.
[00146] Pharmaceutically acceptable compositions of this invention may be orally administered in any orally acceptable dosage form including, but not limited to, capsules, tablets, aqueous suspensions or solutions. In the case of tablets for oral use, carriers commonly used include lactose and corn starch. Lubricating agents, such as magnesium stearate, are also typically added. For oral administration in a capsule form, useful diluents include lactose and dried cornstarch. When aqueous suspensions are required for oral use, the active ingredient is combined with emulsifying and suspending agents. If desired, certain sweetening, flavoring or coloring agents may also be added.
[00147] Alternatively, pharmaceutically acceptable compositions of this invention may be administered in the form of suppositories for rectal administration. These can be prepared by mixing the agent with a suitable non-irritating excipient that is solid at room temperature but liquid at rectal temperature and therefore will melt in the rectum to release the drug. Such materials include cocoa butter, beeswax and polyethylene glycols.
[00148] Pharmaceutically acceptable compositions of this invention may also be administered topically, especially when the target of treatment includes areas or organs readily accessible by topical application, including diseases of the eye, the skin, or the lower intestinal tract. Suitable topical formulations are readily prepared for each of these areas or organs.
[00149] Topical application for the lower intestinal tract can be effected in a rectal suppository formulation (see above) or in a suitable enema formulation. Topically-transdermal patches may also be used.
[00150] For topical applications, provided pharmaceutically acceptable compositions may be formulated in a suitable ointment containing the active component suspended or dissolved in one or more carriers. Carriers for topical administration of compounds of this invention include, but are not limited to, mineral oil, liquid petrolatum, white petrolatum, propylene glycol, polyoxyethylene, polyoxypropylene compound, emulsifying wax and water. Alternatively, provided pharmaceutically acceptable compositions can be formulated in a suitable lotion or cream containing the active components suspended or dissolved in one or more pharmaceutically acceptable carriers. Suitable carriers include, but are not limited to, mineral oil, sorbitan monostearate, polysorbate 60, cetyl esters wax, cetearyl alcohol, 2-octyldodecanol, benzyl alcohol and water.
[00151] For ophthalmic use, provided pharmaceutically acceptable compositions may be formulated as micronized suspensions in isotonic, pH adjusted sterile saline, or, preferably, as solutions in isotonic, pH adjusted sterile saline, either with or without a preservative such as benzylalkonium chloride. Alternatively, for ophthalmic uses, the pharmaceutically acceptable compositions may be formulated in an ointment such as petrolatum. [00152] Pharmaceutically acceptable compositions of this invention may also be administered by nasal aerosol or inhalation. Such compositions are prepared according to techniques well- known in the art of pharmaceutical formulation and may be prepared as solutions in saline, employing benzyl alcohol or other suitable preservatives, absorption promoters to enhance bioavailability, fluorocarbons, and/or other conventional solubilizing or dispersing agents.
[00153] Most preferably, pharmaceutically acceptable compositions of this invention are formulated for oral administration. Such formulations may be administered with or without food. In some embodiments, pharmaceutically acceptable compositions of this invention are administered without food. In other embodiments, pharmaceutically acceptable compositions of this invention are administered with food.
[00154] The amount of compounds of the present invention that may be combined with the carrier materials to produce a composition in a single dosage form will vary depending upon the host treated, the particular mode of administration. Preferably, provided compositions should be formulated so that a dosage of between 0.01 - 100 mg/kg body weight/day of the inhibitor can be administered to a patient receiving these compositions.
[00155] It should also be understood that a specific dosage and treatment regimen for any particular patient will depend upon a variety of factors, including the activity of the specific compound employed, the age, body weight, general health, sex, diet, time of administration, rate of excretion, drug combination, and the judgment of the treating physician and the severity of the particular disease being treated. The amount of a compound of the present invention in the composition will also depend upon the particular compound in the composition.
Uses of Compounds and Pharmaceutically Acceptable Compositions
[00156] Compounds and compositions described herein are generally useful for the inhibition of kinase activity of one or more enzymes. In some embodiments the kinase inhibited by the compounds and methods of the invention is FLT3
[00157] FLT3 is a receptor tyrosine kinase (RTK) that is reported to be mutated in 25-30% of acute myeloid leukemia (AML) cases. Sanz et al., "FLT3 inhibition as a targeted therapy for acute myeloid leukemia," Current Opinion Oncol. (2009) 21 :594-600. Specifically, a mutation in the internal tandem duplication (ITD) of the fins-like tyrosine 3 (FLT3) gene is reported to be the second most common genetic change associated with cytogenetically normal AML. This mutation is indicated to be an important prognostic factor for this class of patients as mutations of the ITD are associated with poor disease prognosis. Sanz, Id. FLT3 is thus a recognized molecular target for the development of new therapies for AML. Sanz id at 596; Qi Chao et al, "Identification of (AC220), a Uniquely Potent, Selective and Efficiacious FMS-Like Tyrosine Kinase-3 (FLT3) Inhibitor," J. Med. Chem. (2009) 52:7808-7816. Currently, there are a number of selective FLT3 inhibitors being investigated as treatments for AML (including tandutinib and AC220), and sunitinib, a multitargeted kinase inhibitor (including FLT3), has already been approved for sale. Another clinically relevant mutation of FLT3 is D835Y, which results in constitutive activity of FLT3. D835Y is often present as a secondary mutation to ITD, resulting in further drug resistance to existing treatments. Moore et al. "Selective FLT3 inhibition of FLT- ITD+ acute myeloid leukaemia resulting in secondary D835Y mutation: a model for emerging clinical resistance systems" Leukemia (2012) 26(7): 1462-70.
[00158] The activity of a compound utilized in this invention as an inhibitor of FLT3, or a mutant thereof, may be assayed in vitro, in vivo or in a cell line. In vitro assays include assays that determine inhibition of either the phosphorylation activity and/or the subsequent functional consequences, or ATPase activity of activated FLT3, or a mutant thereof. Alternate in vitro assays quantitate the ability of the inhibitor to bind to FLT3. Inhibitor binding may be measured by radiolabeling the inhibitor prior to binding, isolating the inhibitor/FLT3 complex and determining the amount of radiolabel bound. Alternatively, inhibitor binding may be determined by running a competition experiment where new inhibitors are incubated with FLT3 bound to known radioligands. Representative in vitro and in vivo assays useful in assaying a FLT3 inhibitor include those described and disclosed in, e.g., , each of which is herein incorporated by reference in its entirety. Detailed conditions for assaying a compound utilized in this invention as an inhibitor of FLT3, or a mutant thereof, are set forth in the Examples below.
[00159] As used herein, the terms "treatment," "treat," and "treating" refer to reversing, alleviating, delaying the onset of, or inhibiting the progress of a disease or disorder, or one or more symptoms thereof, as described herein. In some embodiments, treatment may be administered after one or more symptoms have developed. In other embodiments, treatment may be administered in the absence of symptoms. For example, treatment may be administered to a susceptible individual prior to the onset of symptoms (e.g., in light of a history of symptoms and/or in light of genetic or other susceptibility factors). Treatment may also be continued after symptoms have resolved, for example to prevent or delay their recurrence.
[00160] Provided compounds are inhibitors of FLT3 and are therefore useful for treating one or more disorders associated with activity of FLT3 or mutants thereof. Thus, in certain embodiments, the present invention provides a method for treating a FLT3 -mediated disorder comprising the step of administering to a patient in need thereof a compound of the present invention, or pharmaceutically acceptable composition thereof.
[00161] As used herein, the term "FLT3 -mediated" disorders, diseases, and/or conditions as used herein means any disease or other deleterious condition in which FLT3 or a mutant thereof is known to play a role. Accordingly, another embodiment of the present invention relates to treating or lessening the severity of one or more diseases in which FLT3, or a mutant thereof, is known to play a role. Such FLT3 -mediated disorders include but are not limited to acute myeloid leukemia (also known as acute myelogenous leukemia), solid tumors, gliomas, myelodysplasia syndrome, renal cell carcinoma, glioblastoma, prostate cancer, melanoma, acute lymphoblastic leukemia, myeloproliferative disorder, nasopharyngeal carcinoma, breast tumors, thrombocytopenia, polycythemia vera, myelofibrosis, chronic myelocytic leukemia, lung tumors, colorectal tumors, and autoimmune diseases.
[00162] In some embodiments, the disorder is selected from gliomas, myelodysplasia syndrome, acute lymphoblastic leukemia, myeloproliferative disorder, nasopharyngeal carcinoma, polycythemia vera, and myelofibrosis.
[00163] In some embodiments, the present invention provides a method for treating one or more disorders, diseases, and/or conditions wherein the disorder, disease, or condition is a cancer, a neurodegenative disorder, a viral disease, an autoimmune disease, an inflammatory disorder, a hereditary disorder, a hormone-related disease, a metabolic disorder, conditions associated with organ transplantation, immunodeficiency disorders, a destructive bone disorder, a proliferative disorder, an infectious disease, a condition associated with cell death, thrombin- induced platelet aggregation, liver disease, pathologic immune conditions involving T cell activation, a cardiovascular disorder, or a CNS disorder.
[00164] In one embodiment, a human patient is treated with a compound of the current invention and a pharmaceutically acceptable carrier, adjuvant, or vehicle, wherein said compound is present in an amount to measurably inhibit FLT3 kinase activity or modulate CDK8 scaffold activity as it relates to cyclinC, MED 12, and MED13.
[00165] Compounds of the current invention are useful in the treatment of a proliferative disease selected from a benign or malignant tumor, solid tumor, carcinoma of the brain, kidney, liver, adrenal gland, bladder, breast, stomach, gastric tumors, ovaries, colon, rectum, prostate, pancreas, lung, vagina, cervix, testis, genitourinary tract, esophagus, larynx, skin, bone or thyroid, sarcoma, glioblastomas, neuroblastomas, multiple myeloma, gastrointestinal cancer, especially colon carcinoma or colorectal adenoma, a tumor of the neck and head, an epidermal hyperproliferation, psoriasis, prostate hyperplasia, a neoplasia, a neoplasia of epithelial character, adenoma, adenocarcinoma, keratoacanthoma, epidermoid carcinoma, large cell carcinoma, non- small-cell lung carcinoma, lymphomas, Hodgkins and Non-Hodgkins, a mammary carcinoma, follicular carcinoma, undifferentiated carcinoma, papillary carcinoma, seminoma, melanoma, an IL-6 driven disorder, Smoldering of indolent multiple myeloma, or hematological malignancies (including leukemia, diffuse large B-cell lymphoma (DLBCL), ABC DLBCL, chronic lymphocytic leukemia (CLL), chronic lymphocytic lymphoma, primary effusion lymphoma, Burkitt lymphoma/leukemia, acute lymphocytic leukemia, B-cell prolymphocytic leukemia, lymphoplasmacytic lymphoma, Waldenstrom's macroglobulinemia (WM), splenic marginal zone lymphoma, multiple myeloma, plasmacytoma, intravascular large B-cell lymphoma, and acute myeloid leukemia (AML)).
[00166] In some embodiments, the present invention provides a method of treating acute myeloid leukemia (AML) comprising administering to a patient in need thereof, a therapeutically effective amount of a compound of the invention, or pharmaceutical composition thereof. In some embodiments, the AML is associated with a length mutation in FLT3. In some embodiments, the AML is associated with an internal tandem duplication, insertion or deletion of the FLT3 gene (FLT3-ITD). In some embodiments, the AML is associated with a point mutation or deletion at residues 835 or 836 of FLT3. In some embodiments, the AML is associated with a D835Y point mutation in FLT3. In some embodiments, the AML is associated with both an internal tandem duplication and a D835Y mutation in FLT3. In some embodiments, the AML is resistant to one or more other FLT3 inhibitors. In some embodiments, the AML is resistant to one or more drugs selected from quizartinib (AC220), a staurosporine derivative (e.g. midostaurin (PKC412) or lestaurtinib), sorafenib, and sunitinib. [00167] In some embodiments the proliferative disease which can be treated according to the methods of this invention is an IL-6 driven disorder. In some embodiments the IL-6 driven disorder is Smoldering of indolent multiple myeloma.
[00168] Compounds of the invention are also useful in the treatment of inflammatory or allergic conditions of the skin, for example psoriasis, contact dermatitis, atopic dermatitis, alopecia areata, erythema multiforma, dermatitis herpetiformis, scleroderma, vitiligo, hypersensitivity angiitis, urticaria, bullous pemphigoid, lupus erythematosus, systemic lupus erythematosus, pemphigus vulgaris, pemphigus foliaceus, paraneoplastic pemphigus, epidermolysis bullosa acquisita, acne vulgaris, and other inflammatory or allergic conditions of the skin.
[00169] Compounds of the invention may also be used for the treatment of other diseases or conditions, such as diseases or conditions having an inflammatory component, for example, treatment of diseases and conditions of the eye such as ocular allergy, conjunctivitis, keratoconjunctivitis sicca, and vernal conjunctivitis, diseases affecting the nose including allergic rhinitis, and inflammatory disease in which autoimmune reactions are implicated or having an autoimmune component or etiology, including autoimmune hematological disorders (e.g. hemolytic anemia, aplastic anemia, pure red cell anemia and idiopathic thrombocytopenia), systemic lupus erythematosus, rheumatoid arthritis, polychondritis, scleroderma, Wegener granulamatosis, dermatomyositis, chronic active hepatitis, myasthenia gravis, Steven-Johnson syndrome, idiopathic sprue, autoimmune inflammatory bowel disease (e.g. ulcerative colitis and Crohn's disease), irritable bowel syndrome, celiac disease, periodontitis, hyaline membrane disease, kidney disease, glomerular disease, alcoholic liver disease, multiple sclerosis, endocrine opthalmopathy, Grave's disease, sarcoidosis, alveolitis, chronic hypersensitivity pneumonitis, multiple sclerosis, primary biliary cirrhosis, uveitis (anterior and posterior), Sjogren's syndrome, keratoconjunctivitis sicca and vernal keratoconjunctivitis, interstitial lung fibrosis, psoriatic arthritis, systemic juvenile idiopathic arthritis, cryopyrin-associated periodic syndrome, nephritis, vasculitis, diverticulitis, interstitial cystitis, glomerulonephritis (with and without nephrotic syndrome, e.g. including idiopathic nephrotic syndrome or minal change nephropathy), chronic granulomatous disease, endometriosis, leptospiriosis renal disease, glaucoma, retinal disease, ageing, headache, pain, complex regional pain syndrome, cardiac hypertrophy, musclewasting, catabolic disorders, obesity, fetal growth retardation, hypercholesterolemia, heart disease, chronic heart failure, mesothelioma, anhidrotic ecodermal dysplasia, Behcet's disease, incontinentia pigmenti, Paget' s disease, pancreatitis, hereditary periodic fever syndrome, asthma (allergic and non-allergic, mild, moderate, severe, bronchitic, and exercise-induced), acute lung injury, acute respiratory distress syndrome, eosinophilia, hypersensitivities, anaphylaxis, nasal sinusitis, ocular allergy, silica induced diseases, COPD (reduction of damage, airways inflammation, bronchial hyperreactivity, remodeling or disease progression), pulmonary disease, cystic fibrosis, acid-induced lung injury, pulmonary hypertension, polyneuropathy, cataracts, muscle inflammation in conjunction with systemic sclerosis, inclusion body myositis, myasthenia gravis, thyroiditis, Addison's disease, lichen planus, Type 1 diabetes, or Type 2 diabetes, appendicitis, atopic dermatitis, asthma, allergy, blepharitis, bronchiolitis, bronchitis, bursitis, cervicitis, cholangitis, cholecystitis, chronic graft rejection, colitis, conjunctivitis, Crohn's disease, cystitis, dacryoadenitis, dermatitis, dermatomyositis, encephalitis, endocarditis, endometritis, enteritis, enterocolitis, epicondylitis, epididymitis, fasciitis, fibrositis, gastritis, gastroenteritis, Henoch-Schonlein purpura, hepatitis, hidradenitis suppurativa, immunoglobulin A nephropathy, interstitial lung disease, laryngitis, mastitis, meningitis, myelitis myocarditis, myositis, nephritis, oophoritis, orchitis, osteitis, otitis, pancreatitis, parotitis, pericarditis, peritonitis, pharyngitis, pleuritis, phlebitis, pneumonitis, pneumonia, polymyositis, proctitis, prostatitis, pyelonephritis, rhinitis, salpingitis, sinusitis, stomatitis, synovitis, tendonitis, tonsillitis, ulcerative colitis, uveitis, vaginitis, vasculitis, or vulvitis.
[00170] In some embodiments the inflammatory disease which can be treated according to the methods of this invention is selected from acute and chronic gout, chronic gouty arthritis, psoriasis, psoriatic arthritis, rheumatoid arthritis, Juvenile rheumatoid arthritis, Systemic jubenile idiopathic arthritis (SJIA), Cryopyrin Associated Periodic Syndrome (CAPS), and osteoarthritis.
[00171] In some embodiments the inflammatory disease which can be treated according to the methods of this invention is a TH17 mediated disease. In some embodiments the TH17 mediated disease is selected from Systemic lupus erythematosus, Multiple sclerosis, and inflammatory bowel disease (including Crohn's disease or ulcerative colitis).
[00172] In some embodiments the inflammatory disease which can be treated according to the methods of this invention is selected from Sjogren's syndrome, allergic disorders, osteoarthritis, conditions of the eye such as ocular allergy, conjunctivitis, keratoconjunctivitis sicca and vernal conjunctivitis, and diseases affecting the nose such as allergic rhinitis. [00173] Furthermore, the invention provides the use of a compound according to the definitions herein, or a pharmaceutically acceptable salt, or a hydrate or solvate thereof for the preparation of a medicament for the treatment of a proliferative disease, an inflammatory disease, an obstructive respiratory disease, a cardiovascular disease, a metabolic disease, a neurological disease, a neurodegenerative disease, a viral disease, or a disorder commonly occurring in connection with transplantation.
Combination Therapies
[00174] Depending upon the particular condition, or disease, to be treated, additional therapeutic agents, which are normally administered to treat that condition, may be administered in combination with compounds and compositions of this invention. As used herein, additional therapeutic agents that are normally administered to treat a particular disease, or condition, are known as "appropriate for the disease, or condition, being treated."
[00175] In certain embodiments, a provided combination, or composition thereof, is administered in combination with another therapeutic agent.
[00176] Examples of agents the combinations of this invention may also be combined with include, without limitation: treatments for Alzheimer's Disease such as Aricept® and Excelon®; treatments for HIV such as ritonavir; treatments for Parkinson's Disease such as L- DOPA/carbidopa, entacapone, ropinrole, pramipexole, bromocriptine, pergolide, trihexephendyl, and amantadine; agents for treating Multiple Sclerosis (MS) such as beta interferon (e.g., Avonex® and Rebif®), Copaxone®, and mitoxantrone; treatments for asthma such as albuterol and Singulair®; agents for treating schizophrenia such as zyprexa, risperdal, seroquel, and haloperidol; anti-inflammatory agents such as corticosteroids, TNF blockers, IL-1 RA, azathioprine, cyclophosphamide, and sulfasalazine; immunomodulatory and immunosuppressive agents such as cyclosporin, tacrolimus, rapamycin, mycophenolate mofetil, interferons, corticosteroids, cyclophophamide, azathioprine, and sulfasalazine; neurotrophic factors such as acetylcholinesterase inhibitors, MAO inhibitors, interferons, anti-convulsants, ion channel blockers, riluzole, and anti-Parkinsonian agents; agents for treating cardiovascular disease such as beta-blockers, ACE inhibitors, diuretics, nitrates, calcium channel blockers, and statins; agents for treating liver disease such as corticosteroids, cholestyramine, interferons, and anti-viral agents; agents for treating blood disorders such as corticosteroids, anti-leukemic agents, and growth factors; agents that prolong or improve pharmacokinetics such as cytochrome P450 inhibitors (i.e., inhibitors of metabolic breakdown) and CYP3A4 inhibitors (e.g., ketokenozole and ritonavir), and agents for treating immunodeficiency disorders such as gamma globulin.
[00177] In certain embodiments, combination therapies of the present invention, or a pharmaceutically acceptable composition thereof, are administered in combination with a monoclonal antibody or an siR A therapeutic.
[00178] Those additional agents may be administered separately from a provided combination therapy, as part of a multiple dosage regimen. Alternatively, those agents may be part of a single dosage form, mixed together with a compound of this invention in a single composition. If administered as part of a multiple dosage regime, the two active agents may be submitted simultaneously, sequentially or within a period of time from one another normally within five hours from one another.
[00179] As used herein, the term "combination," "combined," and related terms refers to the simultaneous or sequential administration of therapeutic agents in accordance with this invention. For example, a combination of the present invention may be administered with another therapeutic agent simultaneously or sequentially in separate unit dosage forms or together in a single unit dosage form.
[00180] The amount of additional therapeutic agent present in the compositions of this invention will be no more than the amount that would normally be administered in a composition comprising that therapeutic agent as the only active agent. Preferably the amount of additional therapeutic agent in the presently disclosed compositions will range from about 50% to 100% of the amount normally present in a composition comprising that agent as the only therapeutically active agent.
[00181] In one embodiment, the present invention provides a composition comprising a compound of formula I and one or more additional therapeutic agents. The therapeutic agent may be administered together with a compound of formula I, or may be administered prior to or following administration of a compound of formula I. Suitable therapeutic agents are described in further detail below. In certain embodiments, a compound of formula I may be administered up to 5 minutes, 10 minutes, 15 minutes, 30 minutes, 1 hour, 2 hours, 3 hours, 4 hours, 5, hours, 6 hours, 7 hours, 8 hours, 9 hours, 10 hours, 11 hours, 12 hours, 13 hours, 14 hours, 15 hours, 16 hours, 17 hours, or 18 hours before the therapeutic agent. In other embodiments, a compound of formula I may be administered up to 5 minutes, 10 minutes, 15 minutes, 30 minutes, 1 hour, 2 hours, 3 hours, 4 hours, 5, hours, 6 hours, 7 hours, 8 hours, 9 hours, 10 hours, 11 hours, 12 hours, 13 hours, 14 hours, 15 hours, 16 hours, 17 hours, or 18 hours following the therapeutic agent.
[00182] In another embodiment, the present invention provides a method of treating an inflammatory disease, disorder or condition by administering to a patient in need thereof a compound of formula I and one or more additional therapeutic agents. Such additional therapeutic agents may be small molecules or recombinant biologic agents and include, for example, acetaminophen, non-steroidal anti-inflammatory drugs (NSAIDS) such as aspirin, ibuprofen, naproxen, etodolac (Lodine®) and celecoxib, colchicine (Colcrys®), corticosteroids such as prednisone, prednisolone, methylprednisolone, hydrocortisone, and the like, probenecid, allopurinol, febuxostat (Uloric®), sulfasalazine (Azulfidine®), antimalarials such as hydroxychloroquine (Plaquenil®) and chloroquine (Aralen®), methotrexate (Rheumatrex®), gold salts such as gold thioglucose (Solganal®), gold thiomalate (Myochrysine®) and auranofin (Ridaura®), D-penicillamine (Depen® or Cuprimine®), azathioprine (Imuran®), cyclophosphamide (Cytoxan®), chlorambucil (Leukeran®), cyclosporine (Sandimmune®), leflunomide (Arava®) and "anti-TNF" agents such as etanercept (Enbrel®), infliximab (Remicade®), golimumab (Simponi®), certolizumab pegol (Cimzia®) and adalimumab (Humira®), "anti-IL-1" agents such as anakinra (Kineret®) and rilonacept (Arcalyst®), canakinumab (Ilaris®), anti-Jak inhibitors such as tofacitinib, antibodies such as rituximab (Rituxan®), "anti-T-cell" agents such as abatacept (Orencia®), "anti-IL-6" agents such as tocilizumab (Actemra®), diclofenac, cortisone, hyaluronic acid (Synvisc® or Hyalgan®), monoclonal antibodies such as tanezumab, anticoagulants such as heparin (Calcinparine® or Liquaemin®) and warfarin (Coumadin®), antidiarrheals such as diphenoxylate (Lomotil®) and loperamide (Imodium®), bile acid binding agents such as cholestyramine, alosetron (Lotronex®), lubiprostone (Amitiza®), laxatives such as Milk of Magnesia, polyethylene glycol (MiraLax®), Dulcolax®, Correctol® and Senokot®, anticholinergics or antispasmodics such as dicyclomine (Bentyl®), Singulair®, beta-2 agonists such as albuterol (Ventolin® HFA, Proventil® HFA), levalbuterol (Xopenex®), metaproterenol (Alupent®), pirbuterol acetate (Maxair®), terbutaline sulfate (Brethaire®), salmeterol xinafoate (Serevent®) and formoterol (Foradil®), anticholinergic agents such as ipratropium bromide (Atrovent®) and tiotropium (Spiriva®), inhaled corticosteroids such as beclomethasone dipropionate (Beclovent®, Qvar®, and Vanceril®), triamcinolone acetonide (Azmacort®), mometasone (Asthmanex®), budesonide (Pulmocort®), and flunisolide (Aerobid®), Afviar®, Symbicort®, Dulera®, cromolyn sodium (Intal®), methylxanthines such as theophylline (Theo-Dur®, Theolair®, Slo-bid®, Uniphyl®, Theo-24®) and aminophylline, IgE antibodies such as omalizumab (Xolair®), nucleoside reverse transcriptase inhibitors such as zidovudine (Retrovir®), abacavir (Ziagen®), abacavir/lamivudine (Epzicom®), abacavir/lamivudine/zidovudine (Trizivir®), didanosine (Videx®), emtricitabine (Emtriva®), lamivudine (Epivir®), lamivudine/zidovudine (Combivir®), stavudine (Zerit®), and zalcitabine (Hivid®), non-nucleoside reverse transcriptase inhibitors such as delavirdine (Rescriptor®), efavirenz (Sustiva®), nevairapine (Viramune®) and etravirine (Intelence®), nucleotide reverse transcriptase inhibitors such as tenofovir (Viread®), protease inhibitors such as amprenavir (Agenerase®), atazanavir (Reyataz®), darunavir (Prezista®), fosamprenavir (Lexiva®), indinavir (Crixivan®), lopinavir and ritonavir (Kaletra®), nelfinavir (Viracept®), ritonavir (Norvir®), saquinavir (Fortovase® or Invirase®), and tipranavir (Aptivus®), entry inhibitors such as enfuvirtide (Fuzeon®) and maraviroc (Selzentry®), integrase inhibitors such as raltegravir (Isentress®), doxorubicin (Hydrodaunorubicin®), vincristine (Oncovin®), bortezomib (Velcade®), and dexamethasone (Decadron ®) in combination with lenalidomide (Revlimid ®), or any combination(s) thereof.
[00183] In another embodiment, the present invention provides a method of treating rheumatoid arthritis comprising administering to a patient in need thereof a compound of formula I and one or more additional therapeutic agents selected from non-steroidal antiinflammatory drugs (NSAIDS) such as aspirin, ibuprofen, naproxen, etodolac (Lodine®) and celecoxib, corticosteroids such as prednisone, prednisolone, methylprednisolone, hydrocortisone, and the like, sulfasalazine (Azulfidine®), antimalarials such as hydroxychloroquine (Plaquenil®) and chloroquine (Aralen®), methotrexate (Rheumatrex®), gold salts such as gold thioglucose (Solganal®), gold thiomalate (Myochrysine®) and auranofm (Ridaura®), D-penicillamine (Depen® or Cuprimine®), azathioprine (Imuran®), cyclophosphamide (Cytoxan®), chlorambucil (Leukeran®), cyclosporine (Sandimmune®), leflunomide (Arava®) and "anti- TNF" agents such as etanercept (Enbrel®), infliximab (Remicade®), golimumab (Simponi®), certolizumab pegol (Cimzia®) and adalimumab (Humira®), "anti-IL-1" agents such as anakinra (Kineret®) and rilonacept (Arcalyst®), antibodies such as rituximab (Rituxan®), "anti-T-cell" agents such as abatacept (Orencia®) and "anti-IL-6" agents such as tocilizumab (Actemra®). [00184] In some embodiments, the present invention provides a method of treating osteoarthritis comprising administering to a patient in need thereof a compound of formula I and one or more additional therapeutic agents selected from acetaminophen, non-steroidal antiinflammatory drugs (NSAIDS) such as aspirin, ibuprofen, naproxen, etodolac (Lodine®) and celecoxib, diclofenac, cortisone, hyaluronic acid (Synvisc® or Hyalgan®) and monoclonal antibodies such as tanezumab.
[00185] In some embodiments, the present invention provides a method of treating lupus comprising administering to a patient in need thereof a compound of formula I and one or more additional therapeutic agents selected from acetaminophen, non-steroidal anti-inflammatory drugs (NSAIDS) such as aspirin, ibuprofen, naproxen, etodolac (Lodine®) and celecoxib, corticosteroids such as prednisone, prednisolone, methylprednisolone, hydrocortisone, and the like, antimalarials such as hydroxychloroquine (Plaquenil®) and chloroquine (Aralen®), cyclophosphamide (Cytoxan®), methotrexate (Rheumatrex®), azathioprine (Imuran®) and anticoagulants such as heparin (Calcinparine® or Liquaemin®) and warfarin (Coumadin®).
[00186] In some embodiments, the present invention provides a method of treating inflammatory bowel disease comprising administering to a patient in need thereof a compound of formula I and one or more additional therapeutic agents selected from mesalamine (Asacol®) sulfasalazine (Azulfidine®), antidiarrheals such as diphenoxylate (Lomotil®) and loperamide (Imodium®), bile acid binding agents such as cholestyramine, alosetron (Lotronex®), lubiprostone (Amitiza®), laxatives such as Milk of Magnesia, polyethylene glycol (MiraLax®), Dulcolax®, Correctol® and Senokot® and anticholinergics or antispasmodics such as dicyclomine (Bentyl®), anti-TNF therapies, steroids, and antibiotics such as Flagyl or ciprofloxacin.
[00187] In some embodiments, the present invention provides a method of treating asthma comprising administering to a patient in need thereof a compound of formula I and one or more additional therapeutic agents selected from Singulair®, beta-2 agonists such as albuterol (Ventolin® HFA, Proventil® HFA), levalbuterol (Xopenex®), metaproterenol (Alupent®), pirbuterol acetate (Maxair®), terbutaline sulfate (Brethaire®), salmeterol xinafoate (Serevent®) and formoterol (Foradil®), anticholinergic agents such as ipratropium bromide (Atrovent®) and tiotropium (Spiriva®), inhaled corticosteroids such as prednisone, prednisolone, beclomethasone dipropionate (Beclovent®, Qvar®, and Vanceril®), triamcinolone acetonide (Azmacort®), mometasone (Asthmanex®), budesonide (Pulmocort®), flunisolide (Aerobid®), Afviar®, Symbicort®, and Dulera®, cromolyn sodium (Intal®), methylxanthines such as theophylline (Theo-Dur®, Theolair®, Slo-bid®, Uniphyl®, Theo-24®) and aminophylline, and IgE antibodies such as omalizumab (Xolair®).
[00188] In some embodiments, the present invention provides a method of treating COPD comprising administering to a patient in need thereof a compound of formula I and one or more additional therapeutic agents selected from beta-2 agonists such as albuterol (Ventolin® HFA, Proventil® HFA), levalbuterol (Xopenex®), metaproterenol (Alupent®), pirbuterol acetate (Maxair®), terbutaline sulfate (Brethaire®), salmeterol xinafoate (Serevent®) and formoterol (Foradil®), anticholinergic agents such as ipratropium bromide (Atrovent®) and tiotropium (Spiriva®), methylxanthines such as theophylline (Theo-Dur®, Theolair®, Slo-bid®, Uniphyl®, Theo-24®) and aminophylline, inhaled corticosteroids such as prednisone, prednisolone, beclomethasone dipropionate (Beclovent®, Qvar®, and Vanceril®), triamcinolone acetonide (Azmacort®), mometasone (Asthmanex®), budesonide (Pulmocort®), flunisolide (Aerobid®), Afviar®, Symbicort®, and Dulera®,
[00189] In another embodiment, the present invention provides a method of treating a hematological malignancy comprising administering to a patient in need thereof a compound of formula I and one or more additional therapeutic agents selected from rituximab (Rituxan®), cyclophosphamide (Cytoxan®), doxorubicin (Hydrodaunorubicin®), vincristine (Oncovin®), prednisone, a hedgehog signaling inhibitor, a BTK inhibitor, a JAK/pan-JAK inhibitor, a TYK2 inhibitor, a PI3K inhibitor, a SYK inhibitor, and combinations thereof.
[00190] In another embodiment, the present invention provides a method of treating a solid tumor comprising administering to a patient in need thereof a compound of formula I and one or more additional therapeutic agents selected from rituximab (Rituxan®), cyclophosphamide (Cytoxan®), doxorubicin (Hydrodaunorubicin®), vincristine (Oncovin®), prednisone, a hedgehog signaling inhibitor, a BTK inhibitor, a JAK/pan-JAK inhibitor, a TYK2 inhibitor, a PI3K inhibitor, a SYK inhibitor, and combinations thereof.
[00191] In another embodiment, the present invention provides a method of treating a hematological malignancy comprising administering to a patient in need thereof a compound of formula I and a Hedgehog (Hh) signaling pathway inhibitor. In some embodiments, the hematological malignancy is DLBCL (Ramirez et al "Defining causative factors contributing in the activation of hedgehog signaling in diffuse large B-cell lymphoma" Leuk. Res. (2012), published online July 17, and incorporated herein by reference in its entirety).
[00192] In another embodiment, the present invention provides a method of treating diffuse large B-cell lymphoma (DLBCL) comprising administering to a patient in need thereof a compound of formula I and one or more additional therapeutic agents selected from rituximab (Rituxan®), cyclophosphamide (Cytoxan®), doxorubicin (Hydrodaunorubicin®), vincristine (Oncovin®), prednisone, a hedgehog signaling inhibitor, and combinations thereof.
[00193] In another embodiment, the present invention provides a method of treating multiple myeloma comprising administering to a patient in need thereof a compound of formula I and one or more additional therapeutic agents selected from bortezomib (Velcade®), and dexamethasone (Decadron®), a hedgehog signaling inhibitor, a BTK inhibitor, a JAK/pan-JAK inhibitor, a TYK2 inhibitor, a PI3K inhibitor, a SYK inhibitor in combination with lenalidomide (Revlimid®).
[00194] In another embodiment, the present invention provides a method of treating Waldenstrom's macroglobulinemia comprising administering to a patient in need thereof a compound of formula I and one or more additional therapeutic agents selected from chlorambucil (Leukeran®), cyclophosphamide (Cytoxan®, Neosar®), fludarabine (Fhidara®), eSadribme (Leustatm®), riiuximab (Rituxan®), a hedgehog signaling inhibitor, a BTK inhibitor, a JAK/pan-JAK inhibitor, a TYK2 inhibitor, a PI3K inhibitor, and a SYK inhibitor.
[00195] In another embodiment, the present invention provides a method of treating or lessening the severity of a disease comprising adm nistering to a patient in need thereof a compotmd of formula I and a BTK inhibitor, wherein the disease is selected from inflammatory bowel disease, arthritis, systemic lupus erythematosus (SLE), vasculitis, idiopathic thrombocytopenic purpura (ITP), rheumatoid arthritis, psoriatic arthritis, osteoarthritis, Still's disease, juvenile arthritis, diabetes, myasthenia gravis, Hashimoto's thyroiditis, Ord's thyroiditis, Graves' disease, autoimmune thyroiditis, Sjogren's syndrome, multiple sclerosis, systemic sclerosis, Lyme neuroborreliosis, Guillain-Barre syndrome, acute disseminated encephalomyelitis, Addison's disease, opsoclonus-myoclonus syndrome, ankylosing spondylosis, antiphospholipid antibody syndrome, aplastic anemia, autoimmune hepatitis, autoimmune gastritis, pernicious anemia, celiac disease, Goodpasture's syndrome, idiopathic thrombocytopenic purpura, optic neuritis, scleroderma, primary biliary cirrhosis, Reiter's syndrome, Takayasu's arteritis, temporal arteritis, warm autoimmune hemolytic anemia, Wegener's granulomatosis, psoriasis, alopecia universalis, Behcet's disease, chronic fatigue, dysautonomia, membranous glomerulonephropathy, endometriosis, interstitial cystitis, pemphigus vulgaris, bullous pemphigoid, neuromyotonia, scleroderma, vulvodynia, a hyperproliferative disease, rejection of transplanted organs or tissues, Acquired Immunodeficiency Syndrome (AIDS, also known as HIV), type 1 diabetes, graft versus host disease, transplantation, transfusion, anaphylaxis, allergies (e.g., allergies to plant pollens, latex, drugs, foods, insect poisons, animal hair, animal dander, dust mites, or cockroach calyx), type I hypersensitivity, allergic conjunctivitis, allergic rhinitis, and atopic dermatitis, asthma, appendicitis, atopic dermatitis, asthma, allergy, blepharitis, bronchiolitis, bronchitis, bursitis, cervicitis, cholangitis, cholecystitis, chronic graft rejection, colitis, conjunctivitis, Crohn's disease, cystitis, dacryoadenitis, dermatitis, dermatomyositis, encephalitis, endocarditis, endometritis, enteritis, enterocolitis, epicondylitis, epididymitis, fasciitis, fibrositis, gastritis, gastroenteritis, Henoch-Schonlein purpura, hepatitis, hidradenitis suppurativa, immunoglobulin A nephropathy, interstitial lung disease, laryngitis, mastitis, meningitis, myelitis myocarditis, myositis, nephritis, oophoritis, orchitis, osteitis, otitis, pancreatitis, parotitis, pericarditis, peritonitis, pharyngitis, pleuritis, phlebitis, pneumonitis, pneumonia, polymyositis, proctitis, prostatitis, pyelonephritis, rhinitis, salpingitis, sinusitis, stomatitis, synovitis, tendonitis, tonsillitis, ulcerative colitis, uveitis, vaginitis, vasculitis, or vulvitis, B-cell proliferative disorder, e.g., diffuse large B cell lymphoma, follicular lymphoma, chronic lymphocytic lymphoma, chronic lymphocytic leukemia, acute lymphocytic leukemia, B-cell prolymphocytic leukemia, lymphoplasmacytic lymphoma/Waldenstrom macroglobulinemia, splenic marginal zone lymphoma, multiple myeloma (also known as plasma cell myeloma), non-Hodgkin's lymphoma, Hodgkin's lymphoma, plasmacytoma, extranodal marginal zone B cell lymphoma, nodal marginal zone B cell lymphoma, mantle cell lymphoma, mediastinal (thymic) large B cell lymphoma, intravascular large B cell lymphoma, primary effusion lymphoma, Burkitt lymphoma/leukemia, or lymphomatoid granulomatosis, breast cancer, prostate cancer, or cancer of the mast cells (e.g., mastocytoma, mast cell leukemia, mast cell sarcoma, systemic mastocytosis), bone cancer, colorectal cancer, pancreatic cancer, diseases of the bone and joints including, without limitation, rheumatoid arthritis, seronegative spondyloarthropathies (including ankylosing spondylitis, psoriatic arthritis and Reiter's disease), Behcet's disease, Sjogren's syndrome, systemic sclerosis, osteoporosis, bone cancer, bone metastasis, a thromboembolic disorder, (e.g., myocardial infarct, angina pectoris, reocclusion after angioplasty, restenosis after angioplasty, reocclusion after aortocoronary bypass, restenosis after aortocoronary bypass, stroke, transitory ischemia, a peripheral arterial occlusive disorder, pulmonary embolism, deep venous thrombosis), inflammatory pelvic disease, urethritis, skin sunburn, sinusitis, pneumonitis, encephalitis, meningitis, myocarditis, nephritis, osteomyelitis, myositis, hepatitis, gastritis, enteritis, dermatitis, gingivitis, appendicitis, pancreatitis, cholocystitus, agammaglobulinemia, psoriasis, allergy, Crohn's disease, irritable bowel syndrome, ulcerative colitis, Sjogren's disease, tissue graft rejection, hyperacute rejection of transplanted organs, asthma, allergic rhinitis, chronic obstructive pulmonary disease (COPD), autoimmune polyglandular disease (also known as autoimmune polyglandular syndrome), autoimmune alopecia, pernicious anemia, glomerulonephritis, dermatomyositis, multiple sclerosis, scleroderma, vasculitis, autoimmune hemolytic and thrombocytopenic states, Goodpasture's syndrome, atherosclerosis, Addison's disease, Parkinson's disease, Alzheimer's disease, diabetes, septic shock, systemic lupus erythematosus (SLE), rheumatoid arthritis, psoriatic arthritis, juvenile arthritis, osteoarthritis, chronic idiopathic thrombocytopenic purpura, Waldenstrom macroglobulinemia, myasthenia gravis, Hashimoto's thyroiditis, atopic dermatitis, degenerative joint disease, vitiligo, autoimmune hypopituitarism, Guillain-Barre syndrome, Behcet's disease, scleraderma, mycosis fungoides, acute inflammatory responses (such as acute respiratory distress syndrome and ischemia/reperfusion injury), and Graves' disease.
[00196] In another embodiment, the present invention provides a method of treating or lessening the severity of a disease comprising administering to a patient in need thereof a compound of formula I and a ΡΪ3 inhibitor, wherein the disease is selected from a cancer, a neurodegenative disorder, an angiogenic disorder, a viral disease, an autoimmune disease, an inflammatory disorder, a hormone-related disease, conditions associated with organ transplantation, immunodeficiency disorders, a destructive bone disorder, a proliferative disorder, an infectious disease, a conditio associated with cell death, thrombin-induced platelet aggregation, chronic myelogenous leukemia (C L), chronic lymphocytic leukemia (CLL), liver disease, pathologic immune conditions involving T cell activation, a cardiovascular disorder, and a CNS disorder. [00197] in another embodiment, the present invention provides a method of treating or lessening the severity of a disease comprising administering to a patient in need thereof a compound of formula I and a PD inhibitor, wherein the disease is selected from benign or malignant tumor, carcinoma or solid tumor of the brain, kidney (e.g., renal cell carcinoma (RCC)), liver, adrenal gland, bladder, breast, stomach, gastric tumors, ovaries, colon, rectum, prostate, pancreas, lung, vagina, endometrium, cervix, testis, genitourinary tract, esophagus, larynx, skin, bone or thyroid, sarcoma, glioblastomas, neuroblastomas, multiple myeloma or gastrointestinal cancer, especially colon carcinoma or colorectal adenoma or a tumor of the neck and head, an epidermal hyperproliferation, psoriasis, prostate hyperplasia, a neoplasia, a neoplasia of epithelial character, adenoma, adenocarcinoma, keratoacanthoma, epidermoid carcinoma, large cell carcinoma, non-small-cell lung carcinoma, lymphomas, (including, for example, non-Hodgkin's Lymphoma (NHL) and Hodgkin's lymphoma (also termed Hodgkin's or Hodgkin's disease)), a mammary carcinoma, follicular carcinoma, undifferentiated carcinoma, papillary carcinoma, seminoma, melanoma, or a leukemia, diseases include Cowden syndrome, Lhermitte-Dudos disease and Bannayan-Zonana syndrome, or diseases in which the PI3K/PKB pathway is aberrantly activated, asthma of whatever type or genesis including both intrinsic (non-allergic) asthma and extrinsic (allergic) asthma, mild asthma, moderate asthma, severe asthma, bronchitic asthma, exercise-induced asthma, occupational asthma and asthma induced following bacterial infection, acute lung injury (ALI), adult/acute respiratory distress syndrome (ARDS), chronic obstructive pulmonary, airways or lung disease (COPD, COAD or COLD), including chronic bronchitis or dyspnea associated therewith, emphysema, as well as exacerbation of airways hyperreactivity consequent to other drug therapy, in particular other inhaled drug therapy, bronchitis of whatever type or genesis including, but not limited to, acute, arachidic, catarrhal, croupus, chronic or phthinoid bronchitis, pneumoconiosis (an inflammatory, commonly occupational, disease of the lungs, frequently accompanied by airways obstruction, whether chronic or acute, and occasioned by repeated inhalation of dusts) of whatever type or genesis, including, for example, aluminosis, anthracosis, asbestosis, chalicosis, ptilosis, siderosis, silicosis, tabacosis and byssinosis, Loffler's syndrome, eosinophilic, pneumonia, parasitic (in particular metazoan) infestation (including tropical eosinophilia), bronchopulmonary aspergillosis, polyarteritis nodosa (including Churg-Strauss syndrome), eosinophilic granuloma and eosinophil-related disorders affecting the airways occasioned by drug-reaction, psoriasis, contact dermatitis, atopic dermatitis, alopecia areata, erythema multiforma, dermatitis herpetiformis, scleroderma, vitiligo, hypersensitivity angiitis, urticaria, bullous pemphigoid, lupus erythematosus, pemphisus, epidermolysis bullosa acquisita, conjunctivitis, keratoconjunctivitis sicca, and vernal conjunctivitis, diseases affecting the nose including allergic rhinitis, and inflammatory disease in which autoimmune reactions are implicated or having an autoimmune component or etiology, including autoimmune hematological disorders (e.g. hemolytic anemia, aplastic anemia, pure red cell anemia and idiopathic thrombocytopenia), systemic lupus erythematosus, rheumatoid arthritis, polychondritis, sclerodoma, Wegener granulamatosis, dermatomyositis, chronic active hepatitis, myasthenia gravis, Steven-Johnson syndrome, idiopathic sprue, autoimmune inflammatory bowel disease (e.g. ulcerative colitis and Crohn's disease), endocrine opthalmopathy, Grave's disease, sarcoidosis, alveolitis, chronic hypersensitivity pneumonitis, multiple sclerosis, primary biliary cirrhosis, uveitis (anterior and posterior), keratoconjunctivitis sicca and vernal keratoconjunctivitis, interstitial lung fibrosis, psoriatic arthritis and glomerulonephritis (with and without nephrotic syndrome, e.g. including idiopathic nephrotic syndrome or minal change nephropathy, restenosis, cardiomegaly, atherosclerosis, myocardial infarction, ischemic stroke and congestive heart failure, Alzheimer's disease, Parkinson's disease, amyotrophic lateral sclerosis, Huntington's disease, and cerebral ischemia, and neurodegenerative disease caused by traumatic injury, glutamate neurotoxicity and hypoxia.
[00198] The compounds and compositions, according to the method of the present invention, may be administered using any amount and any route of administration effective for treating or lessening the severity of a cancer, an autoimmune disorder, a proliferative disorder, an inflammatory disorder, a neurodegenerative or neurological disorder, schizophrenia, a bone- related disorder, liver disease, or a cardiac disorder. The exact amount required will vary from subject to subject, depending on the species, age, and general condition of the subject, the severity of the infection, the particular agent, its mode of administration, and the like. Compounds of the invention are preferably formulated in dosage unit form for ease of administration and uniformity of dosage. The expression "dosage unit form" as used herein refers to a physically discrete unit of agent appropriate for the patient to be treated. It will be understood, however, that the total daily usage of the compounds and compositions of the present invention will be decided by the attending physician within the scope of sound medical judgment. The specific effective dose level for any particular patient or organism will depend upon a variety of factors including the disorder being treated and the severity of the disorder; the activity of the specific compound employed; the specific composition employed; the age, body weight, general health, sex and diet of the patient; the time of administration, route of administration, and rate of excretion of the specific compound employed; the duration of the treatment; drugs used in combination or coincidental with the specific compound employed, and like factors well known in the medical arts. The term "patient", as used herein, means an animal, preferably a mammal, and most preferably a human.
[00199] Pharmaceutically acceptable compositions of this invention can be administered to humans and other animals orally, rectally, parenterally, intracisternally, intravaginally, intraperitoneally, topically (as by powders, ointments, or drops), bucally, as an oral or nasal spray, or the like, depending on the severity of the infection being treated. In certain embodiments, the compounds of the invention may be administered orally or parenterally at dosage levels of about 0.01 mg/kg to about 50 mg/kg and preferably from about 1 mg/kg to about 25 mg/kg, of subject body weight per day, one or more times a day, to obtain the desired therapeutic effect.
[00200] Liquid dosage forms for oral administration include, but are not limited to, pharmaceutically acceptable emulsions, microemulsions, solutions, suspensions, syrups and elixirs. In addition to the active compounds, the liquid dosage forms may contain inert diluents commonly used in the art such as, for example, water or other solvents, solubilizing agents and emulsifiers such as ethyl alcohol, isopropyl alcohol, ethyl carbonate, ethyl acetate, benzyl alcohol, benzyl benzoate, propylene glycol, 1,3-butylene glycol, dimethylformamide, oils (in particular, cottonseed, groundnut, corn, germ, olive, castor, and sesame oils), glycerol, tetrahydrofurfuryl alcohol, polyethylene glycols and fatty acid esters of sorbitan, and mixtures thereof. Besides inert diluents, the oral compositions can also include adjuvants such as wetting agents, emulsifying and suspending agents, sweetening, flavoring, and perfuming agents.
[00201] Injectable preparations, for example, sterile injectable aqueous or oleaginous suspensions may be formulated according to the known art using suitable dispersing or wetting agents and suspending agents. The sterile injectable preparation may also be a sterile injectable solution, suspension or emulsion in a nontoxic parenterally acceptable diluent or solvent, for example, as a solution in 1,3-butanediol. Among the acceptable vehicles and solvents that may be employed are water, Ringer's solution, U.S. P. and isotonic sodium chloride solution. In addition, sterile, fixed oils are conventionally employed as a solvent or suspending medium. For this purpose any bland fixed oil can be employed including synthetic mono- or diglycerides. In addition, fatty acids such as oleic acid are used in the preparation of injectables.
[00202] Injectable formulations can be sterilized, for example, by filtration through a bacterial-retaining filter, or by incorporating sterilizing agents in the form of sterile solid compositions which can be dissolved or dispersed in sterile water or other sterile injectable medium prior to use.
[00203] In order to prolong the effect of a compound of the present invention, it is often desirable to slow the absorption of the compound from subcutaneous or intramuscular injection. This may be accomplished by the use of a liquid suspension of crystalline or amorphous material with poor water solubility. The rate of absorption of the compound then depends upon its rate of dissolution that, in turn, may depend upon crystal size and crystalline form. Alternatively, delayed absorption of a parenterally administered compound form is accomplished by dissolving or suspending the compound in an oil vehicle. Injectable depot forms are made by forming microencapsule matrices of the compound in biodegradable polymers such as polylactide- polyglycolide. Depending upon the ratio of compound to polymer and the nature of the particular polymer employed, the rate of compound release can be controlled. Examples of other biodegradable polymers include poly(orthoesters) and poly(anhydrides). Depot injectable formulations are also prepared by entrapping the compound in liposomes or microemulsions that are compatible with body tissues.
[00204] Compositions for rectal or vaginal administration are preferably suppositories which can be prepared by mixing the compounds of this invention with suitable non-irritating excipients or carriers such as cocoa butter, polyethylene glycol or a suppository wax which are solid at ambient temperature but liquid at body temperature and therefore melt in the rectum or vaginal cavity and release the active compound.
[00205] Solid dosage forms for oral administration include capsules, tablets, pills, powders, and granules. In such solid dosage forms, the active compound is mixed with at least one inert, pharmaceutically acceptable excipient or carrier such as sodium citrate or dicalcium phosphate and/or a) fillers or extenders such as starches, lactose, sucrose, glucose, mannitol, and silicic acid, b) binders such as, for example, carboxymethylcellulose, alginates, gelatin, polyvinylpyrrolidinone, sucrose, and acacia, c) humectants such as glycerol, d) disintegrating agents such as agar—agar, calcium carbonate, potato or tapioca starch, alginic acid, certain silicates, and sodium carbonate, e) solution retarding agents such as paraffin, f) absorption accelerators such as quaternary ammonium compounds, g) wetting agents such as, for example, cetyl alcohol and glycerol monostearate, h) absorbents such as kaolin and bentonite clay, and i) lubricants such as talc, calcium stearate, magnesium stearate, solid polyethylene glycols, sodium lauryl sulfate, and mixtures thereof. In the case of capsules, tablets and pills, the dosage form may also comprise buffering agents.
[00206] Solid compositions of a similar type may also be employed as fillers in soft and hard- filled gelatin capsules using such excipients as lactose or milk sugar as well as high molecular weight polyethylene glycols and the like. The solid dosage forms of tablets, dragees, capsules, pills, and granules can be prepared with coatings and shells such as enteric coatings and other coatings well known in the pharmaceutical formulating art. They may optionally contain opacifying agents and can also be of a composition that they release the active ingredient(s) only, or preferentially, in a certain part of the intestinal tract, optionally, in a delayed manner. Examples of embedding compositions that can be used include polymeric substances and waxes. Solid compositions of a similar type may also be employed as fillers in soft and hard-filled gelatin capsules using such excipients as lactose or milk sugar as well as high molecular weight polethylene glycols and the like.
[00207] The active compounds can also be in micro-encapsulated form with one or more excipients as noted above. The solid dosage forms of tablets, dragees, capsules, pills, and granules can be prepared with coatings and shells such as enteric coatings, release controlling coatings and other coatings well known in the pharmaceutical formulating art. In such solid dosage forms the active compound may be admixed with at least one inert diluent such as sucrose, lactose or starch. Such dosage forms may also comprise, as is normal practice, additional substances other than inert diluents, e.g., tableting lubricants and other tableting aids such a magnesium stearate and microcrystalline cellulose. In the case of capsules, tablets and pills, the dosage forms may also comprise buffering agents. They may optionally contain opacifying agents and can also be of a composition that they release the active ingredient(s) only, or preferentially, in a certain part of the intestinal tract, optionally, in a delayed manner. Examples of embedding compositions that can be used include polymeric substances and waxes. [00208] Dosage forms for topical or transdermal administration of a compound of this invention include ointments, pastes, creams, lotions, gels, powders, solutions, sprays, inhalants or patches. The active component is admixed under sterile conditions with a pharmaceutically acceptable carrier and any needed preservatives or buffers as may be required. Ophthalmic formulation, ear drops, and eye drops are also contemplated as being within the scope of this invention. Additionally, the present invention contemplates the use of transdermal patches, which have the added advantage of providing controlled delivery of a compound to the body. Such dosage forms can be made by dissolving or dispensing the compound in the proper medium. Absorption enhancers can also be used to increase the flux of the compound across the skin. The rate can be controlled by either providing a rate controlling membrane or by dispersing the compound in a polymer matrix or gel.
[00209] According to one embodiment, the invention relates to a method of inhibiting protein kinase activity in a biological sample comprising the step of contacting said biological sample with a compound of this invention, or a composition comprising said compound.
[00210] According to another embodiment, the invention relates to a method of inhibiting FLT3, or a mutant thereof, activity in a biological sample comprising the step of contacting said biological sample with a compound of this invention, or a composition comprising said compound. In certain embodiments, the invention relates to a method of irreversibly inhibiting FLT3, or a mutant thereof, activity in a biological sample comprising the step of contacting said biological sample with a compound of this invention, or a composition comprising said compound. In another embodiment, the invention provides a method of simultaneously inhibiting both FLT3 and one or more other protein kinases selected from IRAK-1, IRAK-2, and IRAK-4, CDK8, CLK1, CLK2, or mutants thereof.
[00211] The term "biological sample", as used herein, includes, without limitation, cell cultures or extracts thereof; biopsied material obtained from a mammal or extracts thereof; and blood, saliva, urine, feces, semen, tears, or other body fluids or extracts thereof.
[00212] Inhibition of protein kinase, or a protein kinase selected from FLT3, or a mutant thereof, activity in a biological sample is useful for a variety of purposes that are known to one of skill in the art. Examples of such purposes include, but are not limited to, blood transfusion, organ-transplantation, biological specimen storage, and biological assays. [00213] Another embodiment of the present invention relates to a method of inhibiting protein kinase activity in a patient comprising the step of administering to said patient a compound of the present invention, or a composition comprising said compound.
[00214] According to another embodiment, the invention relates to a method of inhibiting activity of FLT3, or a mutant thereof, in a patient comprising the step of administering to said patient a compound of the present invention, or a composition comprising said compound. According to certain embodiments, the invention relates to a method of reversibly or irreversibly inhibiting one or more of FLT3, or a mutant thereof, activity in a patient comprising the step of administering to said patient a compound of the present invention, or a composition comprising said compound. In other embodiments, the present invention provides a method for treating a disorder mediated by FLT3, or a mutant thereof, in a patient in need thereof, comprising the step of administering to said patient a compound according to the present invention or pharmaceutically acceptable composition thereof. Such disorders are described in detail herein.
[00215] Depending upon the particular condition, or disease, to be treated, additional therapeutic agents that are normally administered to treat that condition, may also be present in the compositions of this invention. As used herein, additional therapeutic agents that are normally administered to treat a particular disease, or condition, are known as "appropriate for the disease, or condition, being treated."
[00216] A compound of the current invention may also be used to advantage in combination with other therapeutic compounds. In some embodiments, the other therapeutic compounds are antiproliferative compounds. Such antiproliferative compounds include, but are not limited to aromatase inhibitors; antiestrogens; topoisomerase I inhibitors; topoisomerase II inhibitors; microtubule active compounds; alkylating compounds; histone deacetylase inhibitors; compounds which induce cell differentiation processes; cyclooxygenase inhibitors; MMP inhibitors; mTOR inhibitors; antineoplastic antimetabolites; platin compounds; compounds targeting/decreasing a protein or lipid kinase activity and further anti-angiogenic compounds; compounds which target, decrease or inhibit the activity of a protein or lipid phosphatase; gonadorelin agonists; anti-androgens; methionine aminopeptidase inhibitors; matrix metalloproteinase inhibitors; bisphosphonates; biological response modifiers; antiproliferative antibodies; heparanase inhibitors; inhibitors of Ras oncogenic isoforms; telomerase inhibitors; proteasome inhibitors; compounds used in the treatment of hematologic malignancies; compounds which target, decrease or inhibit the activity of Flt-3; Hsp90 inhibitors such as 17- AAG (17-allylaminogeldanamycin, NSC330507), 17-DMAG (17-dimethylaminoethylamino-17- demethoxy-geldanamycin, NSC707545), IPI-504, CNFIOIO, CNF2024, CNFIOIO from Conforma Therapeutics; temozolomide (Temodal®); kinesin spindle protein inhibitors, such as SB715992 or SB743921 from GlaxoSmithKline, or pentamidine/chlorpromazine from CombinatoRx; MEK inhibitors such as ARRY142886 from Array BioPharma, AZD6244 from AstraZeneca, PD 181461 from Pfizer and leucovorin. The term "aromatase inhibitor" as used herein relates to a compound which inhibits estrogen production, for instance, the conversion of the substrates androstenedione and testosterone to estrone and estradiol, respectively. The term includes, but is not limited to steroids, especially atamestane, exemestane and formestane and, in particular, non-steroids, especially aminoglutethimide, roglethimide, pyridoglutethimide, trilostane, testolactone, ketokonazole, vorozole, fadrozole, anastrozole and letrozole. Exemestane is marketed under the trade name Aromasin™. Formestane is marketed under the trade name Lentaron™. Fadrozole is marketed under the trade name Afema™. Anastrozole is marketed under the trade name Arimidex™. Letrozole is marketed under the trade names Femara™ or Femar™. Aminoglutethimide is marketed under the trade name Orimeten™. A combination of the invention comprising a chemotherapeutic agent which is an aromatase inhibitor is particularly useful for the treatment of hormone receptor positive tumors, such as breast tumors.
[00217] The term "antiestrogen" as used herein relates to a compound which antagonizes the effect of estrogens at the estrogen receptor level. The term includes, but is not limited to tamoxifen, fulvestrant, raloxifene and raloxifene hydrochloride. Tamoxifen is marketed under the trade name Nolvadex™. Raloxifene hydrochloride is marketed under the trade name Evista™. Fulvestrant can be administered under the trade name Faslodex™. A combination of the invention comprising a chemotherapeutic agent which is an antiestrogen is particularly useful for the treatment of estrogen receptor positive tumors, such as breast tumors.
[00218] The term "anti-androgen" as used herein relates to any substance which is capable of inhibiting the biological effects of androgenic hormones and includes, but is not limited to, bicalutamide (Casodex™). The term "gonadorelin agonist" as used herein includes, but is not limited to abarelix, goserelin and goserelin acetate. Goserelin can be administered under the trade name Zoladex™. [00219] The term "topoisomerase I inhibitor" as used herein includes, but is not limited to topotecan, gimatecan, irinotecan, camptothecian and its analogues, 9-nitrocamptothecin and the macromolecular camptothecin conjugate PNU-166148. Irinotecan can be administered, e.g. in the form as it is marketed, e.g. under the trademark Camptosar™. Topotecan is marketed under the trade name Hycamptin™.
[00220] The term "topoisomerase II inhibitor" as used herein includes, but is not limited to the anthracyclines such as doxorubicin (including liposomal formulation, such as Caelyx™), daunorubicin, epirubicin, idarubicin and nemorubicin, the anthraquinones mitoxantrone and losoxantrone, and the podophillotoxines etoposide and teniposide. Etoposide is marketed under the trade name Etopophos™. Teniposide is marketed under the trade name VM 26-Bristol Doxorubicin is marketed under the trade name Acriblastin™ or Adriamycin™. Epirubicin is marketed under the trade name Farmorubicin™. Idarubicin is marketed, under the trade name Zavedos™. Mitoxantrone is marketed under the trade name Novantron.
[00221] The term "microtubule active agent" relates to microtubule stabilizing, microtubule destabilizing compounds and microtublin polymerization inhibitors including, but not limited to taxanes, such as paclitaxel and docetaxel; vinca alkaloids, such as vinblastine or vinblastine sulfate, vincristine or vincristine sulfate, and vinorelbine; discodermolides; cochicine and epothilones and derivatives thereof. Paclitaxel is marketed under the trade name Taxol™. Docetaxel is marketed under the trade name Taxotere™. Vinblastine sulfate is marketed under the trade name Vinblastin R.P™. Vincristine sulfate is marketed under the trade name Farmistin™.
[00222] The term "alkylating agent" as used herein includes, but is not limited to, cyclophosphamide, ifosfamide, melphalan or nitrosourea (BCNU or Gliadel). Cyclophosphamide is marketed under the trade name Cyclostin™. Ifosfamide is marketed under the trade name Holoxan™.
[00223] The term "histone deacetylase inhibitors" or "HDAC inhibitors" relates to compounds which inhibit the histone deacetylase and which possess antiproliferative activity. This includes, but is not limited to, suberoylanilide hydroxamic acid (SAHA).
[00224] The term "antineoplastic antimetabolite" includes, but is not limited to, 5-fluorouracil or 5-FU, capecitabine, gemcitabine, DNA demethylating compounds, such as 5-azacytidine and decitabine, methotrexate and edatrexate, and folic acid antagonists such as pemetrexed. Capecitabine is marketed under the trade name Xeloda™. Gemcitabine is marketed under the trade name Gemzar™.
[00225] The term "platin compound" as used herein includes, but is not limited to, carboplatin, cis-platin, cisplatinum and oxaliplatin. Carboplatin can be administered, e.g., in the form as it is marketed, e.g. under the trademark Carboplat™. Oxaliplatin can be administered, e.g., in the form as it is marketed, e.g. under the trademark Eloxatin™.
[00226] The term "compounds targeting/decreasing a protein or lipid kinase activity; or a protein or lipid phosphatase activity; or further anti-angiogenic compounds" as used herein includes, but is not limited to, protein tyrosine kinase and/or serine and/or threonine kinase inhibitors or lipid kinase inhibitors, such as a) compounds targeting, decreasing or inhibiting the activity of the platelet-derived growth factor-receptors (PDGFR), such as compounds which target, decrease or inhibit the activity of PDGFR, especially compounds which inhibit the PDGF receptor, such as an N-phenyl-2-pyrimidine-amine derivative, such as imatinib, SU101, SU6668 and GFB-111; b) compounds targeting, decreasing or inhibiting the activity of the fibroblast growth factor-receptors (FGFR); c) compounds targeting, decreasing or inhibiting the activity of the insulin-like growth factor receptor I (IGF-IR), such as compounds which target, decrease or inhibit the activity of IGF-IR, especially compounds which inhibit the kinase activity of IGF-I receptor, or antibodies that target the extracellular domain of IGF-I receptor or its growth factors; d) compounds targeting, decreasing or inhibiting the activity of the Trk receptor tyrosine kinase family, or ephrin B4 inhibitors; e) compounds targeting, decreasing or inhibiting the activity of the Axl receptor tyrosine kinase family; f) compounds targeting, decreasing or inhibiting the activity of the Ret receptor tyrosine kinase; g) compounds targeting, decreasing or inhibiting the activity of the Kit/SCFR receptor tyrosine kinase, such as imatinib; h) compounds targeting, decreasing or inhibiting the activity of the C-kit receptor tyrosine kinases, which are part of the PDGFR family, such as compounds which target, decrease or inhibit the activity of the c-Kit receptor tyrosine kinase family, especially compounds which inhibit the c-Kit receptor, such as imatinib; i) compounds targeting, decreasing or inhibiting the activity of members of the c-Abl family, their gene-fusion products (e.g. BCR-Abl kinase) and mutants, such as compounds which target decrease or inhibit the activity of c-Abl family members and their gene fusion products, such as an N-phenyl-2-pyrimidine-amine derivative, such as imatinib or nilotinib (AMN107); PD 180970; AG957; NSC 680410; PD 173955 from ParkeDavis; or dasatinib (BMS-354825); j) compounds targeting, decreasing or inhibiting the activity of members of the protein kinase C (PKC) and Raf family of serine/threonine kinases, members of the MEK, SRC, JAK/pan-JAK, FAK, PDK1, PKB/Akt, Ras/MAPK, PI3K, SYK, TYK2, BTK and TEC family, and/or members of the cyclin-dependent kinase family (CDK) including staurosporine derivatives, such as midostaurin; examples of further compounds include UCN-01, safmgol, BAY 43-9006, Bryostatin 1, Perifosine; llmofosine; RO 318220 and RO 320432; GO 6976; lsis 3521; LY333531/LY379196; isochinoline compounds; FTIs; PD 184352 or QAN697 (a P13K inhibitor) or AT7519 (CDK inhibitor); k) compounds targeting, decreasing or inhibiting the activity of protein-tyrosine kinase inhibitors, such as compounds which target, decrease or inhibit the activity of protein-tyrosine kinase inhibitors include imatinib mesylate (Gleevec™) or tyrphostin such as Tyrphostin A23/RG-50810; AG 99; Tyrphostin AG 213; Tyrphostin AG 1748; Tyrphostin AG 490; Tyrphostin B44; Tyrphostin B44 (+) enantiomer; Tyrphostin AG 555; AG 494; Tyrphostin AG 556, AG957 and adaphostin (4-{[(2,5- dihydroxyphenyl)methyl] amino} -benzoic acid adamantyl ester; NSC 680410, adaphostin); 1) compounds targeting, decreasing or inhibiting the activity of the epidermal growth factor family of receptor tyrosine kinases (EGFRi ErbB2, ErbB3, ErbB4 as homo- or heterodimers) and their mutants, such as compounds which target, decrease or inhibit the activity of the epidermal growth factor receptor family are especially compounds, proteins or antibodies which inhibit members of the EGF receptor tyrosine kinase family, such as EGF receptor, ErbB2, ErbB3 and ErbB4 or bind to EGF or EGF related ligands, CP 358774, ZD 1839, ZM 105180; trastuzumab (Herceptin™), cetuximab (Erbitux™), Iressa, Tarceva, OSI-774, Cl-1033, EKB-569, GW-2016, El .l, E2.4, E2.5, E6.2, E6.4, E2. l l, E6.3 or E7.6.3, and 7H-pyrrolo-[2,3-d]pyrimidine derivatives; m) compounds targeting, decreasing or inhibiting the activity of the c-Met receptor, such as compounds which target, decrease or inhibit the activity of c-Met, especially compounds which inhibit the kinase activity of c-Met receptor, or antibodies that target the extracellular domain of c-Met or bind to HGF, n) compounds targeting, decreasing or inhibiting the kinase activity of one or more JAK family members (JAK1/JAK2/JAK3/TYK2 and/or pan-JAK), including but not limited to PRT-062070, SB- 1578, baricitinib, pacritinib, momelotinib, VX-509, AZD-1480, TG-101348, tofacitinib, and ruxolitinib; o) compounds targeting, decreasing or inhibiting the kinase activity of PI3 kinase (PI3K) including but not limited to ATU-027, SF- 1126, DS-7423, PBI-05204, GSK-2126458, ZSTK-474, buparlisib, pictrelisib, PF-4691502, BYL-719, dactolisib, XL- 147, XL-765, and idelalisib; and; and q) compounds targeting, decreasing or inhibiting the signaling effects of hedgehog protein (Hh) or smoothened receptor (SMO) pathways, including but not limited to cyclopamine, vismodegib, itraconazole, erismodegib, and IPI-926 (saridegib).
[00227] The term "PI3K inhibitor" as used herein includes, but is not limited to compounds having inhibitory activity against one or more enzymes in the phosphatidylinositol-3 -kinase family, including, but not limited to ΡΒΚα, ΡΒΚγ, ΡΒΚδ, ΡΒΚβ, PI3K-C2a, PBK-C2P, ΡΒΚ- C2y, Vps34, pl lO-a, ρ110-β, ρ110-γ, ρ110-δ, ρ85-α, ρ85-β, ρ55-γ, ρ150, plOl, and ρ87. Examples of ΡΒΚ inhibitors useful in this invention include but are not limited to ATU-027, SF- 1126, DS-7423, PBI-05204, GSK-2126458, ZSTK-474, buparlisib, pictrelisib, PF-4691502, BYL-719, dactolisib, XL-147, XL-765, and idelalisib.
[00228] The term "BTK inhibitor" as used herein includes, but is not limited to compounds having inhibitory activity against Bruton's Tyrosine Kinase (BTK), including, but not limited to AVL-292 and ibrutinib.
[00229] The term "SYK inhibitor" as used herein includes, but is not limited to compounds having inhibitory activity against spleen tyrosine kinase (SYK), including but not limited to PRT-062070, Pv-343, R-333, Excellair, PRT-062607, and fostamatinib
[00230] Further examples of BTK inhibitory compounds, and conditions treatable by such compounds in combination with compounds of this invention can be found in WO2008039218 and WO2011090760, the entirety of which are incorporated herein by reference.
[00231] Further examples of SYK inhibitory compounds, and conditions treatable by such compounds in combination with compounds of this invention can be found in WO2003063794,
WO2005007623, and WO2006078846, the entirety of which are incorporated herein by reference.
[00232] Further examples of PBK inhibitory compounds, and conditions treatable by such compounds in combination with compounds of this invention can be found in WO2004019973, WO2004089925, WO2007016176, US8138347, WO2002088112, WO2007084786, WO2007129161, WO2006122806, WO2005113554, and WO2007044729 the entirety of which are incorporated herein by reference.
[00233] Further examples of JAK inhibitory compounds, and conditions treatable by such compounds in combination with compounds of this invention can be found in WO2009114512, WO2008109943, WO2007053452, WO2000142246, and WO2007070514, the entirety of which are incorporated herein by reference.
[00234] Further anti-angiogenic compounds include compounds having another mechanism for their activity, e.g. unrelated to protein or lipid kinase inhibition e.g. thalidomide (Thalomid™) and TNP-470.
[00235] Examples of proteasome inhibitors useful for use in combination with compounds of the invention include, but are not limited to bortezomib, disulfiram, epigallocatechin-3-gallate (EGCG), salinosporamide A, carfilzomib, ONX-0912, CEP- 18770, and MLN9708.
[00236] Compounds which target, decrease or inhibit the activity of a protein or lipid phosphatase are e.g. inhibitors of phosphatase 1, phosphatase 2A, or CDC25, such as okadaic acid or a derivative thereof.
[00237] Compounds which induce cell differentiation processes include, but are not limited to, retinoic acid, α- γ- or δ- tocopherol or a- γ- or δ-tocotrienol.
[00238] The term cyclooxygenase inhibitor as used herein includes, but is not limited to, Cox- 2 inhibitors, 5-alkyl substituted 2-arylaminophenylacetic acid and derivatives, such as celecoxib (Celebrex™), rofecoxib (Vioxx™), etoricoxib, valdecoxib or a 5-alkyl-2- arylaminophenylacetic acid, such as 5-methyl-2-(2'-chloro-6'-fluoroanilino)phenyl acetic acid, lumiracoxib.
[00239] The term "bisphosphonates" as used herein includes, but is not limited to, etridonic, clodronic, tiludronic, pamidronic, alendronic, ibandronic, risedronic and zoledronic acid. Etridonic acid is marketed under the trade name Didronel™. Clodronic acid is marketed under the trade name Bonefos™. Tiludronic acid is marketed under the trade name Skelid™. Pamidronic acid is marketed under the trade name Aredia™. Alendronic acid is marketed under the trade name Fosamax™. Ibandronic acid is marketed under the trade name Bondranat™. Risedronic acid is marketed under the trade name Actonel™. Zoledronic acid is marketed under the trade name Zometa™. The term "mTOR inhibitors" relates to compounds which inhibit the mammalian target of rapamycin (mTOR) and which possess antiproliferative activity such as sirolimus (Rapamune®), everolimus (Certican™), CCI-779 and ABT578.
[00240] The term "heparanase inhibitor" as used herein refers to compounds which target, decrease or inhibit heparin sulfate degradation. The term includes, but is not limited to, PI-88. The term "biological response modifier" as used herein refers to a lymphokine or interferons. [00241] The term "inhibitor of Ras oncogenic isoforms", such as H-Ras, K-Ras, or N-Ras, as used herein refers to compounds which target, decrease or inhibit the oncogenic activity of Ras; for example, a "farnesyl transferase inhibitor" such as L-744832, DK8G557 or Rl 15777 (Zarnestra™). The term "telomerase inhibitor" as used herein refers to compounds which target, decrease or inhibit the activity of telomerase. Compounds which target, decrease or inhibit the activity of telomerase are especially compounds which inhibit the telomerase receptor, such as telomestatin.
[00242] The term "methionine ammopeptidase inhibitor" as used herein refers to compounds which target, decrease or inhibit the activity of methionine ammopeptidase. Compounds which target, decrease or inhibit the activity of methionine ammopeptidase include, but are not limited to, bengamide or a derivative thereof.
[00243] The term "proteasome inhibitor" as used herein refers to compounds which target, decrease or inhibit the activity of the proteasome. Compounds which target, decrease or inhibit the activity of the proteasome include, but are not limited to, Bortezomib (Velcade™) and MLN 341.
[00244] The term "matrix metalloproteinase inhibitor" or ("MMP" inhibitor) as used herein includes, but is not limited to, collagen peptidomimetic and nonpeptidomimetic inhibitors, tetracycline derivatives, e.g. hydroxamate peptidomimetic inhibitor batimastat and its orally bioavailable analogue marimastat (BB-2516), prinomastat (AG3340), metastat (NSC 683551) BMS-279251 , BAY 12-9566, TAA211 , MMI270B or AAJ996.
[00245] The term "compounds used in the treatment of hematologic malignancies" as used herein includes, but is not limited to, FMS-like tyrosine kinase inhibitors, which are compounds targeting, decreasing or inhibiting the activity of FMS-like tyrosine kinase receptors (Flt-3R); interferon, Ι-β-D-arabinofuransylcytosine (ara-c) and bisulfan; and ALK inhibitors, which are compounds which target, decrease or inhibit anaplastic lymphoma kinase.
[00246] Compounds which target, decrease or inhibit the activity of FMS-like tyrosine kinase receptors (Flt-3R) are especially compounds, proteins or antibodies which inhibit members of the Flt-3R receptor kinase family, such as PKC412, midostaurin, a staurosporine derivative, SU11248 and MLN518.
[00247] The term "HSP90 inhibitors" as used herein includes, but is not limited to, compounds targeting, decreasing or inhibiting the intrinsic ATPase activity of HSP90; degrading, targeting, decreasing or inhibiting the HSP90 client proteins via the ubiquitin proteosome pathway. Compounds targeting, decreasing or inhibiting the intrinsic ATPase activity of HSP90 are especially compounds, proteins or antibodies which inhibit the ATPase activity of HSP90, such as 17-allylamino, 17-demethoxygeldanamycin (17AAG), a geldanamycin derivative; other geldanamycin related compounds; radicicol and HDAC inhibitors.
[00248] The term "antiproliferative antibodies" as used herein includes, but is not limited to, trastuzumab (Herceptin™), Trastuzumab-DMl , erbitux, bevacizumab (Avastin™), rituximab (Rituxan®), PR064553 (anti-CD40) and 2C4 Antibody. By antibodies is meant intact monoclonal antibodies, polyclonal antibodies, multispecific antibodies formed from at least 2 intact antibodies, and antibodies fragments so long as they exhibit the desired biological activity.
[00249] For the treatment of acute myeloid leukemia (AML), compounds of the current invention can be used in combination with standard leukemia therapies, especially in combination with therapies used for the treatment of AML. In particular, compounds of the current invention can be administered in combination with, for example, farnesyl transferase inhibitors and/or other drugs useful for the treatment of AML, such as Daunorubicin, Adriamycin, Ara-C, VP- 16, Teniposide, Mitoxantrone, Idarubicin, Carboplatinum and PKC412. In some embodiments, the present invention provides a method of treating AML associated with an ITD and/or D835Y mutation, comprising administering a compound of the present invention together with a one or more additional FLT3 inhibitors. In some embodiments, the other FLT3 inhibitors are selected from quizartinib (AC220), a staurosporine derivative (e.g. midostaurin or lestaurtinib), sorafenib, tandutinib, LY-2401401 , LS-104, EB-10, famitinib, NOV-1 10302, NMS-P948, AST-487, G-749, SB-1317, S-209, SC-1 10219, AKN-028, fedratinib, tozasertib, and sunitinib. In some embodiments, the other FLT3 inhibitors are selected from quizartinib, midostaurin, lestaurtinib, sorafenib, and sunitinib.
[00250] Other anti-leukemic compounds include, for example, Ara-C, a pyrimidine analog, which is the 2 -alpha-hydroxy ribose (arabinoside) derivative of deoxycytidine. Also included is the purine analog of hypoxanthine, 6-mercaptopurine (6-MP) and fludarabine phosphate. Compounds which target, decrease or inhibit activity of histone deacetylase (HDAC) inhibitors such as sodium butyrate and suberoylanilide hydroxamic acid (SAHA) inhibit the activity of the enzymes known as histone deacetylases. Specific HDAC inhibitors include MS275, SAHA, FK228 (formerly FR901228), Trichostatin A and compounds disclosed in US 6,552,065 including, but not limited to, N-hydroxy-3-[4-[[[2-(2-methyl-lH-indol-3-yl)-ethyl]- amino]methyl]phenyl]-2E-2-propenamide, or a pharmaceutically acceptable salt thereof and N- hydroxy-3-[4-[(2-hydroxyethyl){2-(lH-indol-3-yl)ethyl]-amino]methyl]phenyl]-2E-2- propenamide, or a pharmaceutically acceptable salt thereof, especially the lactate salt. Somatostatin receptor antagonists as used herein refer to compounds which target, treat or inhibit the somatostatin receptor such as octreotide, and SOM230. Tumor cell damaging approaches refer to approaches such as ionizing radiation. The term "ionizing radiation" referred to above and hereinafter means ionizing radiation that occurs as either electromagnetic rays (such as X- rays and gamma rays) or particles (such as alpha and beta particles). Ionizing radiation is provided in, but not limited to, radiation therapy and is known in the art. See Hellman, Principles of Radiation Therapy, Cancer, in Principles and Practice of Oncology, Devita et al, Eds., 4th Edition, Vol. 1 , pp. 248-275 (1993).
[00251] Also included are EDG binders and ribonucleotide reductase inhibitors. The term "EDG binders" as used herein refers to a class of immunosuppressants that modulates lymphocyte recirculation, such as FTY720. The term "ribonucleotide reductase inhibitors" refers to pyrimidine or purine nucleoside analogs including, but not limited to, fludarabine and/or cytosine arabinoside (ara-C), 6-thioguanine, 5-fluorouracil, cladribine, 6-mercaptopurine (especially in combination with ara-C against ALL) and/or pentostatin. Ribonucleotide reductase inhibitors are especially hydroxyurea or 2-hydroxy-lH-isoindole-l ,3-dione derivatives.
[00252] Also included are in particular those compounds, proteins or monoclonal antibodies of VEGF such as l-(4-chloroanilino)-4-(4-pyridylmethyl)phthalazine or a pharmaceutically acceptable salt thereof, l-(4-chloroanilino)-4-(4-pyridylmethyl)phthalazine succinate; Angiostatin™; Endostatin™; anthranilic acid amides; ZD4190; ZD6474; SU5416; SU6668; bevacizumab; or anti-VEGF antibodies or anti-VEGF receptor antibodies, such as rhuMAb and RHUFab, VEGF aptamer such as Macugon; FLT-4 inhibitors, FLT-3 inhibitors, VEGFR-2 IgGI antibody, Angiozyme (RPI 4610) and Bevacizumab (Avastin™).
[00253] Photodynamic therapy as used herein refers to therapy which uses certain chemicals known as photosensitizing compounds to treat or prevent cancers. Examples of photodynamic therapy include treatment with compounds, such as Visudyne™ and porfimer sodium.
[00254] Angiostatic steroids as used herein refers to compounds which block or inhibit angiogenesis, such as, e.g., anecortave, triamcinolone, hydrocortisone, 11-a-epihydrocotisol, cortexolone, 17a-hydroxyprogesterone, corticosterone, desoxycorticosterone, testosterone, estrone and dexamethasone.
[00255] Implants containing corticosteroids refers to compounds, such as fiuocinolone and dexamethasone.
[00256] Other chemotherapeutic compounds include, but are not limited to, plant alkaloids, hormonal compounds and antagonists; biological response modifiers, preferably lymphokines or interferons; antisense oligonucleotides or oligonucleotide derivatives; shRNA or siRNA; or miscellaneous compounds or compounds with other or unknown mechanism of action.
[00257] The compounds of the invention are also useful as co-therapeutic compounds for use in combination with other drug substances such as anti-inflammatory, bronchodilatory or antihistamine drug substances, particularly in the treatment of obstructive or inflammatory airways diseases such as those mentioned hereinbefore, for example as potentiators of therapeutic activity of such drugs or as a means of reducing required dosaging or potential side effects of such drugs. A compound of the invention may be mixed with the other drug substance in a fixed pharmaceutical composition or it may be administered separately, before, simultaneously with or after the other drug substance. Accordingly the invention includes a combination of a compound of the invention as hereinbefore described with an antiinflammatory, bronchodilatory, antihistamine or anti-tussive drug substance, said compound of the invention and said drug substance being in the same or different pharmaceutical composition.
[00258] Suitable anti-inflammatory drugs include steroids, in particular glucocorticosteroids such as budesonide, beclamethasone dipropionate, fluticasone propionate, ciclesonide or mometasone furoate; non-steroidal glucocorticoid receptor agonists; LTB4 antagonists such LY293111, CGS025019C, CP-195543, SC-53228, BIIL 284, ONO 4057, SB 209247; LTD4 antagonists such as montelukast and zafirlukast; PDE4 inhibitors such cilomilast (Arifio® GlaxoSmithKline), Rofiumilast (Byk Gulden), V-l 1294A (Napp), BAY 19-8004 (Bayer), SCH- 351591 (Schering- Plough), Arofylline (Almirall Prodesfarma), PD 189659 / PD 168787 (Parke- Davis), AWD-12- 281 (Asta Medica), CDC-801 (Celgene), SelCID(TM) CC- 10004 (Celgene), VM554/UM565 (Vernalis), T-440 (Tanabe), KW-4490 (Kyowa Hakko Kogyo); A2a agonists; A2b antagonists; and beta-2 adrenoceptor agonists such as albuterol (salbutamol), metaproterenol, terbutaline, salmeterol fenoterol, procaterol, and especially, formoterol and pharmaceutically acceptable salts thereof. Suitable bronchodilatory drugs include anticholinergic or antimuscarinic compounds, in particular ipratropium bromide, oxitropium bromide, tiotropium salts and CHF 4226 (Chiesi), and glycopyrrolate.
[00259] Suitable antihistamine drug substances include cetirizine hydrochloride, acetaminophen, clemastine fumarate, promethazine, loratidine, desloratidine, diphenhydramine and fexofenadine hydrochloride, activastine, astemizole, azelastine, ebastine, epinastine, mizolastine and tefenadine.
[00260] Other useful combinations of compounds of the invention with anti-inflammatory drugs are those with antagonists of chemokine receptors, e.g. CCR-1 , CCR-2, CCR-3, CCR-4, CCR-5, CCR-6, CCR-7, CCR-8, CCR-9 and CCR10, CXCR1 , CXCR2, CXCR3, CXCR4, CXCR5, particularly CCR-5 antagonists such as Schering-Plough antagonists SC-351125, SCH- 55700 and SCH-D, and Takeda antagonists such as N-[[4-[[[6,7-dihydro-2-(4-methylphenyl)- 5H-benzo-cyclohepten-8-yl]carbonyl]amino]phenyl]-methyl]tetrahydro-N,N-dimethyl-2H- pyran-4- aminium chloride (TAK-770).
[00261] The structure of the active compounds identified by code numbers, generic or trade names may be taken from the actual edition of the standard compendium "The Merck Index" or from databases, e.g. Patents International (e.g. IMS World Publications).
[00262] A compound of the current invention may also be used in combination with known therapeutic processes, for example, the administration of hormones or radiation. In certain embodiments, a provided compound is used as a radiosensitizer, especially for the treatment of tumors which exhibit poor sensitivity to radiotherapy.
[00263] A compound of the current invention can be administered alone or in combination with one or more other therapeutic compounds, possible combination therapy taking the form of fixed combinations or the administration of a compound of the invention and one or more other therapeutic compounds being staggered or given independently of one another, or the combined administration of fixed combinations and one or more other therapeutic compounds. A compound of the current invention can besides or in addition be administered especially for tumor therapy in combination with chemotherapy, radiotherapy, immunotherapy, phototherapy, surgical intervention, or a combination of these. Long-term therapy is equally possible as is adjuvant therapy in the context of other treatment strategies, as described above. Other possible treatments are therapy to maintain the patient's status after tumor regression, or even chemopreventive therapy, for example in patients at risk. [00264] Those additional agents may be administered separately from an inventive compound-containing composition, as part of a multiple dosage regimen. Alternatively, those agents may be part of a single dosage form, mixed together with a compound of this invention in a single composition. If administered as part of a multiple dosage regime, the two active agents may be submitted simultaneously, sequentially or within a period of time from one another normally within five hours from one another.
[00265] As used herein, the term "combination," "combined," and related terms refers to the simultaneous or sequential administration of therapeutic agents in accordance with this invention. For example, a compound of the present invention may be administered with another therapeutic agent simultaneously or sequentially in separate unit dosage forms or together in a single unit dosage form. Accordingly, the present invention provides a single unit dosage form comprising a compound of the current invention, an additional therapeutic agent, and a pharmaceutically acceptable carrier, adjuvant, or vehicle.
[00266] The amount of both an inventive compound and additional therapeutic agent (in those compositions which comprise an additional therapeutic agent as described above) that may be combined with the carrier materials to produce a single dosage form will vary depending upon the host treated and the particular mode of administration. Preferably, compositions of this invention should be formulated so that a dosage of between 0.01 - 100 mg/kg body weight/day of an inventive compound can be administered.
[00267] In those compositions which comprise an additional therapeutic agent, that additional therapeutic agent and the compound of this invention may act synergistically. Therefore, the amount of additional therapeutic agent in such compositions will be less than that required in a monotherapy utilizing only that therapeutic agent. In such compositions a dosage of between 0.01 - 1,000 μg/kg body weight/day of the additional therapeutic agent can be administered.
[00268] The amount of additional therapeutic agent present in the compositions of this invention will be no more than the amount that would normally be administered in a composition comprising that therapeutic agent as the only active agent. Preferably the amount of additional therapeutic agent in the presently disclosed compositions will range from about 50% to 100% of the amount normally present in a composition comprising that agent as the only therapeutically active agent. [00269] The compounds of this invention, or pharmaceutical compositions thereof, may also be incorporated into compositions for coating an implantable medical device, such as prostheses, artificial valves, vascular grafts, stents and catheters. Vascular stents, for example, have been used to overcome restenosis (re -narrowing of the vessel wall after injury). However, patients using stents or other implantable devices risk clot formation or platelet activation. These unwanted effects may be prevented or mitigated by pre-coating the device with a pharmaceutically acceptable composition comprising a kinase inhibitor. Implantable devices coated with a compound of this invention are another embodiment of the present invention.
EXEMPLIFICATION
[00270] As depicted in the Examples below, in certain exemplary embodiments, compounds are prepared according to the following general procedures. It will be appreciated that, although the general methods depict the synthesis of certain compounds of the present invention, the following general methods, and other methods known to one of ordinary skill in the art, can be applied to all compounds and subclasses and species of each of these compounds, as described herein.
[00271] Example 1: Synthesis of Intermediate 1.9
Figure imgf000094_0001
[00272] Synthesis of compound 1.2. Into a 10-L 4-necked round-bottom flask was placed a solution of 1.1 (550 g, 3.23 mol, 1.00 equiv) in ethanol (2200 mL) at room temperature. This was followed by the addition of NCCH2COOEt (440 g, 1.21 equiv), Et2NH (291.5 g, 1.23 equiv) and S (126.5 g, 1.22 equiv) in portions at room temperature. The solution was stirred overnight at room temperature and then concentrated under vacuum. The resulting solution was diluted with 5000 mL of EA and was washed with 2x1000 mL of brine. The mixture was dried over anhydrous sodium sulfate and concentrated under vacuum. The residue was applied onto a silica gel column with ethyl acetate/petroleum ether (1 : 10). This resulted in 430 g (45%) of 1.2 as an orange oil.
[00273] Synthesis of compound 1.3. To a solution of triphosgene (2.205 g, 7.43 mmol, 1.0 equiv) in 80 mL of anhydrous DCM was added a solution of 1.2 (4.455 g, 14.98 mmol, 2.00 equiv) in DCM (20 mL) dropwise with stirring at 0 °C, followed by addition of TEA (3.8 g, 37.43 mmol, 5.0 equiv) via syringe under nitrogen. The resulting solution was stirred for 1 h at room temperature. To the mixture was added (2,4-dimethoxyphenyl)methanamine (5.01 g, 29.96 mmol, 4.00 equiv) and the resulting solution was allowed to react, with stirring, for an additional 1 h at ambient temperature. The solids were filtered out, washed with 2 x 100 mL of DCM and the filtrate was concentrated under vacuum. The residue was applied onto a silica gel column with ethyl acetate/petroleum ether (1 :5) to give 1.3 (4.5 g, 61%) as a yellow solid.
[00274] Synthesis of compound 1.4. Sodium hydride (2.2 g, 55.00 mmol, 3.00 equiv, 60%) was treated with 1.3 (9.0 g, 18.35 mmol, 1.00 equiv) in 100 mL of dioxane overnight at 100 °C under nitrogen. After cooling, the reaction was then quenched with water and the pH value of the solution was adjusted to 4 with 4 M hydrochloric acid. The solids were collected by filtration and dried in an oven (45 °C) to yield 6.2 g (81 >) of 1.4 as an off-white solid.
[00275] Synthesis of compound 1.5 A solution of 1.4 (6.0 g, 14.41 mmol, 1.00 equiv), ethanol (10 mL) and 4-methylbenzene-l -sulfonic acid (800 mg, 4.65 mmol, 0.32 equiv) in toluene (1 10 mL) was stirred overnight at 120 °C. After cooling, the reaction was quenched with aqueous saturated sodium bicarbonate and extracted with 2 x 200 mL of ethyl acetate. The combined organic layers were dried over sodium sulfate and concentrated under vacuum. The residue was applied onto a silica gel column with ethyl acetate/petroleum ether (1/2) to give 1.5 (6.0 g, 94%) as a yellow solid.
[00276] Synthesis of compound 1.6. To a solution of 1.5 (6.0 g, 13.4 mmol, 1.00 equiv) in 50 mL of trifluoroacetic acid was stirred for 4.5 h at 50 °C in an oil bath under nitrogen. After completion of the reaction, the resulting mixture was concentrated under vacuum to give 1.6 (4.5 g, crude) as a white solid. [00277] Synthesis of compound 1.7. Into a 250-mL round-bottom flask was placed 1.6 (4.0 g, 13.59 mmol, 1.00 equiv) in POCl3 (70 mL) under nitrogen and the resulting mixture was stirred overnight at 105 °C in an oil bath. The resulting mixture was concentrated under vacuum and the residue was diluted with 150 mL of EtO Ac. The pH value of the solution was adjusted to 7-8 with saturated sodium bicarbonate and extracted with 2 x 150 mL of ethyl acetate. The organic layers were washed with brine, dried over anhydrous sodium sulfate and concentrated under vacuum. The residue was applied onto a silica gel column with ethyl acetate/petroleum ether (1 : 10) to give the desired 1.7 (1.95 g, 43%) as a light yellow solid.
[00278] Synthesis of compound 1.8 and 1.9. The enantiomers of 1.7 (2.3 g) were separated by chiral-SFC under the following conditions: Column: Phenomenex Lux 5u Cellulose-3, 5*25 cm, 5 μιη; mobile phase: 75% C02 and 25% MeOH (0.01 DEA); flow rate: 200 g/min; UV detection at 220 μιη. The first fraction to elute (tR = 3.5 min) were collected and evaporated to remove solvent under reduced pressure to give 900 mg of 1.8. The second fraction to elute (tR = 4.25 min) was collected and evaporated to remove solvent under reduced pressure to give 900 mg of compound 1.9. The ee of 1.8 (98.5%>) and of 1.9 (100%) were determined by analytical chiral SFC under the following conditions: Column: phenomenex Lux 5u Cellulose-3, 4.6*250 mm, 5 μιη; mobile phase: 90% C02 and 10% MeOH (0.01 DEA); flow rate: 4 mL/min; UV detection at 254 μιη.
[00279] Example 2: Synthesis of (12S)-3-[[(lr,4r)-4-(morpholin-4-yl)cyclohexyl]oxy]-8- thia-4,6-diazatricyclo[7.4.0.0[2,7]]trideca-l(9),2(7),3,5-tetraene-12-carboxylic acid. (1-1)
Figure imgf000096_0001
[00280] Synthesis of compound 2.1. To a solution of trans-4-morpholinocyclohexanol (122.3 mg, 0.66 mmol, 1.1 equiv) in 5 niL of distilled THF was added NaHMDS (2 M in THF, 0.33 mL, 1.1 equiv) dropwise via a syringe at 0 °C under nitrogen. Then 1.9 (200 mg, 0.6 mmol, 1.0 equiv) in 3 mL of THF was added at this temperature and stirred for 30 min. After the reaction was complete, the reaction mixture was diluted with saturated aqueous NH4C1 and extracted with DCM, washed with brine, dried and concentrated in vacuo. The residue was purified by chromatography on silica gel with DCM/MeOH/NH4OH (80: 1 :0.01 to 50:1 :0.01) to give the desired product 2.1 (140 mg) as a light yellow oil.
[00281] Synthesis of compound 2.2. A mixture of compound 2.1 (140 mg, 0.292 mmol, 1.00 equiv), 1 -methyl- lH-pyrazol-4-amine (42.5 mg, 0.437 mmol, 1.5 equiv), Pd2dba3 (14.3 mg, 0.015 mmol, 0.05 equiv), Xantphos (18.1 mg, 0.030 mmol, 0.10 equiv), Cs2C03 (286 mg, 0.876 mmol, 3.0 equiv) in 8 mL of dioxane was degassed three times with nitrogen. The resulting mixture was stirred for 2 h at 100 °C. The reaction mixture was concentrated under vacuum and the residue was diluted with water and extracted with DCM. The combined organic layers were washed with brine, dried and concentrated in vacuo. Purification by chromatography on silica gel with DCM/MeOH/NH4OH (80: 1 to 30: 1 :0.01) to give the desired 2.2 (130 mg, 90% purity) as a yellow semi-solid.
[00282] Synthesis of compound 2.3. To the compound 2.2 (130 mg, 90% purity) dissolved in a mixture of THF/MeOH/water (3:3: 1.5 mL) was added LiOH FL (40 mg) at room temperature followed by stirring for 4 h at this temperature. The resulting solution was concentrated under reduced pressure. The residue was diluted with 3 mL of water, acidified with 1 M hydrochloric acid to pH 5 and extracted with CHC13/IPA (v/v: 3: 1) four times. The combined organic layers were dried and evaporated in vacuo to give 100 mg crude of 2.3 as a yellow solid.
[00283] Synthesis of Compound 1-1. To a mixture of 2.3 (60 mg, 0.12 mmol, 1.00 equiv) in distilled DMF (5 mL) was added NH4C1 (19.08 mg, 0.36 mmol, 3.08 equiv), HATU (54.7 mg, 0.14 mmol, 1.23 equiv) and DIEA (33.4 mg, 0.26 mmol, 2.21 equiv) followed by stirring for 3 h at room temperature under nitrogen. The resulting solution was diluted with 5 mL of H20 and extracted with 3 x 20 mL of DCM and concentrated under vacuum. The crude product (56 mg) was purified by preparative HPLC under the following conditions (Waters): Column: XBridge Shield RP18 OBD 5 μιη, 19* 150 mm; mobile phase: water with 0.01% NH4HC03 and acetonitrile (10%-35% in 10 min); flow rate: 15 ml/min; UV detection at 254 nm. This resulted in 12.5 mg (21%) of product 1-1 as a white solid. MS (ES): m/z 512 (M+H)+. 1H-NMR (400 MHz, CD3OD): δ 8.90 (s, 1H), 7.57 (s, 1H), 5.22-5.10 (m, 1H), 3.90 (s, 3H), 3.75-3.50 (m, 5H), 3.02-2.95 (m, 2H), 2.90-2.80 (m, 1H), 2.70-2.58 (m, 5H), 2.50-2.41 (m, 3H), 2.25-2.08 (m, 5H), 1.70-1.56 (m, 2H), 1.54-1.38 (m, 2H).
[00284] Example 3: Synthesis of Intermediate 3.4.
Figure imgf000098_0001
3.4
[00285] Synthesis of compound 3.1. Compound 3.1 was prepared in a manner analogous to compound 1.2, substituting cyclopentanone for 1.1. Isolated 3.1 (14.7 g, 35%) as a light yellow solid. MS (ES): m/z 212 (M+H)+. 1H NMR (400 MHz, CDC13): δ 5.84 (2H, br s), 4.26 (2H, q), 2.86-2.82 (2H, m), 2.76-2.72 (2H, m), 2.36-2.29 (2H, m), 1.34 (3H, t).
[00286] Synthesis of compound 3.2. 3.1 (500 mg, 2.37 mmol, 1.00 equiv) was treated with urea (2.1 g, 34.97 mmol, 15.00 equiv) at 180 °C for 2 h in a sand bath. After completion, the reaction temperature was cooled down to room temperature naturally and diluted with water. The pH value of the solution was adjusted to 14 with 6 M aqueous sodium hydroxide solution. The formed solids were filtered out and the filtrate was adjusted to pH 4 with 2 M hydrochloric acid. The isolated solid was collected and purified by recrystallization with water. The solid was dried in an oven under reduced pressure to give 3.2 (0.2 g, 41 >) as a pale solid.
[00287] Synthesis of compound 3.3. To a solution of 3.2 (3 g, 14.41 mmol, 1.00 equiv) in POCI3 (25 mL) was added N,N-dimethylbenzene (2 mL) and the resulting solution was stirred for 2 h at 120 °C in an oil bath under nitrogen. After removal of excess amounts of POCI3 under reduced pressure, the residue was poured into cooled aqueous sodium carbonate solution and extracted with 3x100 mL of ethyl acetate. The combined organic layers were dried and concentrated under vacuum. The residue was applied onto a silica gel column with ethyl acetate/petroleum ether (1 :10) to afford 3.3 (3.1 g, 88%) as a white solid. MS (ES): m/z 245 (M+H)+.
[00288] Synthesis of compound 3.4. A solution of 3.4 (100 mg, 0.41 mmol, 1.00 equiv), 4- (morpholin-4-yl)cyclohexan-l -amine dihydrochloride (143.8 mg, 0.56 mmol, 1.37 equiv) and potassium carbonate (338 mg, 2.45 mmol, 5.99 equiv) in CH3CN (40 mL) in a 100 mL round- bottom flask was heated to reflux overnight in an oil bath. The resulting mixture was concentrated under vacuum. The residue was applied onto a silica gel column with dichloromethane/methanol (30: 1). This resulted in 230 mg (96%) of 3.4 as a white solid.
[00289] Example 4: Synthesis of 12-N-[4-(morpholin-4-yl)cyclohexyl]-10-N-[4-(piperidin- 4-yl)phenyl]-7-thia-9,ll-diazatricyclo[6.4.0.0[2,6]]dodeca-l(8),2(6),9,ll-tetraene-10,12- diamine hydrochloride (1-7).
Figure imgf000099_0001
4.5 1-7
[00290] Synthesis of compound 4.2. A solution of 4-(piperidin-4-yl)aniline hydrochloride (950 mg, 4.47 mmol, 1.00 equiv) and l,3-dihydro-2-benzofuran-l,3-dione (667.2 mg, 4.50 mmol, 1.01 equiv) in acetic acid (100 mL) was heated to reflux for 3 h. The resulting mixture was concentrated under vacuum to give 4.2 (1.49 g, 97%>) as a white solid. [00291] Synthesis of compound 4.3. A solution of 4.2 (1.49 g, 4.33 mmol, 1.00 equiv), 4- dimethylaminopyridine (109 mg, 0.89 mmol, 0.21 equiv), triethylamine (1.805 g, 17.84 mmol, 4.12 equiv) and di-tert-butyl dicarbonate (1.462 g, 6.70 mmol, 1.55 equiv) in dichloromethane (100 mL) was stirred for 3 h at room temperature under nitrogen. The resulting mixture was washed with H20 and extracted with DCM. The combined organic layers were washed with 1M HC1 and brine and dried over anhydrous sodium sulfate. After concentration under vacuum the residue was purified by chromatography on silica gel with EtOAc/PE (1 :30 to 1 : 10) to give 4.3 (1.44 g, 82%) as a white solid.
[00292] Synthesis of compound 4.4. In a 250-mL round-bottom flask a solution of 4.3 (1.433 g, 3.53 mmol, 1.00 equiv) and ΝΗ2ΝΗ2·Η20 (1.84 g, 36.71 mmol, 10.41 equiv) in 80 mL of ethanol was stirred for 4 h at 50 °C in an oil bath. The solids were filtered out. The filtrate was concentrated under vacuum and the residue was applied onto a silica gel column with ethyl acetate/petroleum ether (1 :2) to give 4.4 (446 mg, 46%) as a white solid.
[00293] Synthesis of compound 4.5. In a 50-mL round-bottom flask purged and maintained with an inert atmosphere of nitrogen, a solution of 3.4 (90 mg, 0.23 mmol, 1.00 equiv), Pd2(dba)3 »CHCl3 (12 mg, 0.01 mmol, 0.05 equiv), Xantphos (13 mg, 0.02 mmol, 0.10 equiv), 4.4 (94 mg, 0.34 mmol, 1.48 equiv) and (tert-butoxy)sodium (88 mg, 0.92 mmol, 4.00 equiv) in 1,4- dioxane (10 mL) was stirred overnight at 100 °C in an oil bath. The solids were filtered out. The resulting mixture was concentrated under vacuum and the residue was applied onto a silica gel column with dichloromethane/methanol (30: 1) to give 4.5 (120 mg, 83%>) as a yellow solid.
[00294] Synthesis of Compound 1-7. To a solution of 4.5 (120 mg, 0.19 mmol, 1.00 equiv) in dichloromethane (20 mL) was added 12 M hydrochloric acid (0.2 mL) followed by stirring for 1 h at 0 °C in a water/ice bath. The resulting mixture was concentrated under vacuum. Compound 1-7 (43.8 mg, 41%) was obtained by precipitation in MeOH/Et20 as an off-white solid. MS (ES): m/z 533 (M+H)+. 1H NMR (300 MHz, CD3OD): δ 7.55-7.44 (m, 4H), 4.27-3.92 (m, 5H), 3.61- 3.51 (m, 4H), 3.35-2.95 (m, 10H), 2.62-2.53 (m, 2H), 2.40-2.26 (m, 4H), 2.14-1.98 (m, 4H), 1.84-1.63 (m, 4H).
[00295] Example 5: Synthesis of Intermediate 5.1.
Figure imgf000101_0001
[00296] Compound 5.1 was prepared in a manner analogous to intermediate 3.4, substituting l-N,l-N-dimethylcyclohexane-l,4,dimaine hydrochloride for 4-(morpholin-4-yl)cyclohexan-l- amine dihydrochloride. Isolated 5.1 (300 mg, 79%) as a yellow solid. MS (ES): m/z 252 (M+H)+.
[00297] Example 6: Synthesis of 12-N-[4-(dimethylamino)cyclohexyl]-10-N-[4-(piperidin- 4-yl)phenyl]-7-thia-9,ll-diazatricyclo[6.4.0.0[2,6]] dodeca-l(8),2(6),9,ll-tetraene-10,12- diamine hydrochloride (1-2).
Figure imgf000101_0002
1 1 Synthesis of Compound 1-2. Compound 1-2 was prepared from 5.1 and 4.4 in a manner analogous to the synthesis of Compound 1-7 from 3.4 and 4.4. Compound 1-2 (88.2 mg, 53%) was obtained as an off-white solid. MS (ES): m/z 491 (M+H)+. 1H NMR (400 MHz, CD3OD): δ 7.53 (d, J = 8.4 Hz, 2H), 7.45 (d, J = 8.4 Hz, 2H), 4.27-4.22 (m, 1H), 3.57-3.50 (m, 2H), 3.37-2.92 (m, 14H), 2.63-2.55 (m, 2H), 2.30-2.28 (m, 4H), 2.15-2.08 (m, 4H), 1.65-1.78 (m, 4H).
[00299] Example 7: Synthesis of 12-N-[4-(dimethylamino)cyclohexyl]-10-N-[4-(4- methylpiperazin-l-yl)phenyl]-7-thia-9,ll-diazatricyclo[6.4.0.0[2,6]]dodeca-l(8),2(6),9,ll- tetraene-10,12-diamine (1-3).
Figure imgf000102_0001
[00300] Compound 1-3 was prepared from 5.1 and 7.1 in a manner analogous to the synthesis of 4.5 from 3.4 and 4.4. Isolated 20.5 mg (8%) of a white solid. MS (ES): m/z 506 (M+H)+. 1H NMR (400 MHz, d6-DMSO): δ 8.82 (s, 1H), 7.58 (d, J = 8.8 Hz, 2H), 6.84 (d, J = 8.8 Hz, 2H), 5.76 (d, J = 7.6 Hz, 1H), 4.05-3.88 (m, 1H), 3.03 (t, J = 4.8 Hz, 4H), 2.98 (t, J = 6.4 Hz, 2H), 2.81 (t, J= 6.4 Hz, 2H), 2.45 (t, J= 4.8 Hz, 4H), 2.45-2.31 (m, 2H), 2.28-2.10 (m, 10H), 2.02 (d, (t, J= 12 Hz, 2H), 1.86 (d, (t, J= 12 Hz, 2H), 1.52-1.38 (m, 2H), 1.35-1.20 (m, 2H).
[00301] Example 8: Synthesis of N4-((lr,4r)-4-(dimethylamino)cyclohexyl)-N2-(4-(l- methylpiperidin-4-yl)phenyl)-6,7-dihydro-5H-cyclopenta [4,5] thieno [2,3-d] pyrimidine-2,4- diamine (1-4).
Figure imgf000102_0002
[00302] Compound 1-4 was prepared from 5.1 in a manner analogous to the synthesis of 4.5 from 3.4 and 4.4, substituting 4-(l-methylpiperidin-4-yl)aniline for 4.4. Isolated 31.4 mg of a white solid in 15% yield. MS (ES): m/z 505 (M+H)+. 1H NMR (300 MHz, CDC13): δ 7.59 (d, 2H), 7.16 (d, 2H), 6.82 (s, 1H), 4.76 (d, 1H), 4.10-3.90 (m, 1H), 3.05 (d, 2H), 2.98-2.85 (m, 4H), 2.5-2.22 (m, 14H), 2.21-2.00 (m, 5H), 1.95-1.85 (m, 5H), 1.60-1.38 (m, 2H), 1.35-1.15 (m, 2H).
[00303] Example 9: Synthesis of 10-N-[l-(propan-2-yl)-lH-pyrazol-4-yl]-12-N-[trans-4- (morpholin-4-yl)cyclohexyl]-7-thia-9,ll-diazatricyclo[6.4.0.0[2,6]]dodeca-l(8),2(6),9,ll- tetraene-10,12-diamine (1-9).
Figure imgf000103_0001
3.4 I-9
[00304] A suspension of 3.4 (120 mg, 0.31 mmol, 1.00 equiv), l-(propan-2-yl)-lH-pyrazol-4- amine (46.5 mg, 0.372 mmol, 1.20 equiv) and hydrochloric acid (0.5 mL, 4 M in hexane) in dry isopropanol (5 mL) was heated in the microwave at 140 °C for 1.5 h. After cooling to rt, the resulting mixture was concentrated under vacuum. The residue was applied onto a silica gel column with dichloromethane/methanol/ammonia (20: 1 :0.1) to give 66.1 mg (45%) of 1-9 as a white solid. MS (ES): m/z 482 (M+H)+. 1H NMR (300 MHz, CD3OD): δ 7.84 (s, 1H), 7.56 (s, 1H), 4.49-4.40 (m, 1H), 4.04-3.95 (m, 1H), 3.71-3.68 (m, 4H), 2.94 (t, 2H), 2.83 (t, 2H), 2.78- 2.62 (m, 4H), 2.50-2.45 (m, 2H), 2.39-2.25 (m, 1H), 2.25-2.14 (m, 2H), 2.11-1.99 (m, 2H) , 1.47 (d, 6H), 1.46-1.32 (m, 4H).
[00305] Example 10: Synthesis of 2-[(3S)-12-[[4-(dimethylamino)cyclohexyl]oxy]-10-[(l- methyl-lH-pyrazol-4-yl)amino]-7-thia-9, ll-diazatricyclo[6.4.0.0[2,6]]dodeca-l(8),2(6),9, 1 l-tetraen-3-yl] acetamide (1-23).
Figure imgf000103_0002
[00306] Synthesis of compound 10.1. Compound 10.1 was prepared from 1.8 by reacting with sodium hydride and dimethylaminocyclohexanol in THF. Isolated 0.55 g of a light yellow oil in 46% yield.
[00307] Synthesis of compound 10.2. Compound 10.2 was prepared according to the method for the preparation of compound 4.5. Purification by chromatography on silica gel column with DCM/MeOH (10: 1 to 2: 1) gave 10.2 (180 mg, 62%) as a grey solid.
[00308] Synthesis of Compound 1-23. Compound 1-23 was prepared from 10.2 in a manner analogous to the synthesis of 1-1 from 2.3. Isolated 35.9 mg of a white solid in 20%> yield. MS (ES): m/z 470 (M+H)+. 1H NMR (300 MHz, CD3OD): δ 7.89 (1H, br s), 7.59 (lH,s), 5.19 (1H, m), 3.89 (3H, s), 3.66 (1H, m), 2.83-2.99 (4H, m), 2.70 (7H, m), 2.43 ( 2H, m), 2.14-2.39 (4H, m), 1.76-1.94 (4H, m).
[00309] Example 11: Synthesis of 2-[(3R)-12-[[4-(dimethylamino)cyclohexyl]oxy]-10-[(l- methyl-lH-pyrazol-4-yl)amino]-7-thia-9, ll-diazatricyclo[6.4.0.0[2,6]]dodeca-l(8),2(6),9, 1 l-tetraen-3-yl] acetamide (I- 10).
Figure imgf000104_0001
1-10
Compound 1-10 was prepared from 1.9 and dimethylaminocyclohexanol in a manner analogous to the synthesis of 1-23 from 1.8. Isolated 62.2 mg of a white solid in 5% overall yield from 1.9. MS (ES): m/z 470 (M+H)+. 1H-NMR (300MHz, CD3OD): δ 7.90 ( 1H, br s), 7.56 (lH,s), 5.22- 5.12 (1H, m), 3.92 (3H, s), 3.78-3.62 (1H, m), 3.08-2.80 (3H, m), 2.75-2.50 (2H, m), 2.44 (6H, s), 2.41-2.26 (2H, m), 2.25-2.05 (4H, m), 1.70-1.46 (4H, m).
[00310] Example 12: Synthesis of 2-[(3R)-10-(phenylamino)-12-[[(lr,4r)-4- (dimethylamino)cyclohexyl] oxy] -7-thia-9, 11-diazatricyclo [6.4.0.0 [2,6]] dodeca- 1 (8),2(6),9, ll-tetraen-3-yl] acetamide (1-6).
Figure imgf000104_0002
[00311] Compound 1-6 was prepared from 11.1 in a manner analogous to the synthesis of 1-10, substituting aniline for 1 -methyl- lH-pyrazol-4-amine. Isolated 27.8 mg of a white solid in 13% overall yield from 11.1. MS (ES): m/z 466 (M+H)+. 1H-NMR (400 MHz, CD3OD): δ 7.70 (2H, d), 7.32 (2H, t), 7.00 (1H, t), 5.18-5.05 (1H, m), 3.65-3.50 (1H, m), 2.95-2.85 (2H, m), 2.82-2.71 (2H, m), 2.60-2.46 (7H, s), 2.38-2.28 (2H, m), 2.15-1.98 (4H, m), 1.62-1.40 (4H, m).
[00312] Example 13: Synthesis of 12-N-[4-(dimethylamino)cyclohexyl]-10-N-(l-methyl- lH-pyrazol-4-yl)-7-thia-9,ll-diazatricyclo[6.4.0.0[2,6]]dodeca-l(8),2(6),9,ll-tetraene-10,12- diamine (1-12).
Figure imgf000105_0001
5.1 1-12
[00313] Compound 1-12 was prepared from 5.1 in a manner analogous to the synthesis of 4.5 from 4.4 and 3.4, substituting 1 -methyl- lH-pyrazol-4-amine for 4.4. Isolated 85.3 mg (36%) of a white solid. MS (ES): m/z 412 (M+H)+. 1H-NMR (300 MHz, CDC13): δ 7.86 (1H, s), 7.45 (1H, s), 6.65 (1H, s), 4.75 (1H, d), 4.04-3.93 (1H, m), 3.88 (3H, s), 3.01-2.82 (4H, m), 2.53-2.44 (2H, m), 2.38 (7H, s), 2.30-2.26 (2H, d), 2.06-2.02 (2H, d), 1.54-1.41 (2H, m), 1.32-1.18 (2H, m).
[00314] Example 14: Synthesis of Intermediate 14.1
Figure imgf000105_0002
3.3 14.1
[00315] Sodium hydride (132 mg, 3.30 mmol, 5.06 equiv, 60% dispersion in mineral oil) was treated with trans-4-(dimethylamino)cyclohexan-l-ol (113 mg, 0.79 mmol, 1.21 equiv) in 10 mL of distilled THF at room temperature for 1 h under nitrogen. Then a solution of 3.3 (160 mg, 0.65 mmol, 1.00 equiv) in 5 mL of THF was added via syringe and the resulting solution was allowed to react, with stirring, for an additional 5 h while the temperature was maintained at 60 °C in an oil bath. After cooling, the reaction was then quenched with saturated aqueous NH4C1 and extracted with 5 x 50 mL of ethyl acetate. The combined organic layers were dried over sodium sulfate and concentrated under reduced pressure. The residue was purified via preparative TLC with dichloromethane/methanol/NH4OH (200: 10: 1) to afford the desired 14.1 (180 mg, 78%) as a light yellow solid.
[00316] Example 15: Synthesis of 12-[[4-(dimethylamino)cyclohexyl]oxy]-N-(l-methyl- lH-pyrazol-4-yl)-7-thia-9,ll-diazatricyclo[6.4.0.0[2,6]]dodeca-l(8),2(6),9,ll-tetraen-10- amine (1-14).
Figure imgf000106_0001
14.1 1-14
[00317] Compound 1-14 was prepared from 14.1 in a manner analogous to the synthesis of 4.5 from 4.4 and 3.4, substituting 1 -methyl- lH-pyrazol-4-amine for 4.4. Isolated 84.3 mg (48%) of a white solid. MS (ES): m/z 413 (M+H)+. 1H NMR (300 MHz, DMSO) δ 9.34 (s, 1H), 7.87 (m, 1H), 7.46 (s, 1H), 5.11-5.09 (m, 1H), 3.81 (s, 3H), 2.86-2.84 (m, 4H), 2.42-2.37 (m, 2H), 2.20 (s, 9H), 1.90-1.85 (m, 2H), 1.56-1.39 (m, 4H).
[00318] Example 16: Synthesis of 10-N-(l-ethyl-lH-pyrazol-4-yl)-12-N-[trans-4- (morpholin-4-yl)cyclohexyl]-7-thia-9,ll-diazatricyclo[6.4.0.0[2,6]]dodeca-l(8),2(6),9,ll- tetraene-10,12-diamine (1-8)
Figure imgf000106_0002
3.4 1-8
[00319] Compound 1-8 was prepared from 3.4 in a manner analogous to the synthesis of 1-9, substituting 1 -ethyl- lH-pyrazol-4-amine for l-(propan-2-yl)-lH-pyrazol-4-amine. MS (ES): m/z 468 [M+H]+; 1H NMR (300 MHz, CD3OD): δ 7.88 (1H, s), 7.58 (1H, s), 4.15 (2H, q), 4.15-4.00 (1H, m), 3.80-3.72 (4H, m), 3.00 (2H, t), 2.89 (2H, t), 2.70-2.60 (4H, m), 2.50 (2H, quintet), 2.40-2.29 (1H, m), 2.25-2.18 (2H, m), 2.15-2.08 (2H, m), 1.55-1.35 (7H, m).
[00320] Example 17: Synthesis of 5-N-(l-methyl-lH-pyrazol-4-yl)-3-N-[trans-4- (morpholin-4-yl)cyclohexyl] - 1 l-oxa-8-thia-4,6-diazatricyclo [7.4.0.0 A [2,7] ] trideca- l(9),2(7),3,5-tetraene-3,5-diamine (1-13)
Figure imgf000107_0001
17.1 17.2
Figure imgf000107_0002
17.3 17.4
Figure imgf000107_0003
17.5 1-13
[00321] Synthesis of compound 17.2. Compound 17.2 was prepared in a manner analogous to the synthesis of 1.2, substituting dihydro-2H-pyran-4(3H)-one for 1.1. Isolated 19.5 g (86%) of 17.2 as a yellow solid. MS (ES): m/z 228 (M+H)+.
[00322] Synthesis of compound 17.3. A solution of 17.2 (10.0 g, 44.00 mmol, 1.00 equiv) in dry DCM (150 mL) was cooled down to -60 °C under nitrogen. Chlorosulfonylisocyanate (9.34 g, 65.99 mmol, 1.50 equiv) was added at a rate such that the internal temperature remained at -60 to -55 °C. After completion of the addition, the reaction mixture was allowed to warm to ambient temperature. After 17.1 was consumed, the resulting mixture was concentrated under vacuum. The solid residue was transferred back to 500 mL of flask by water (250 mL) and heated for 70 °C for 1 h. Then the pH value of the mixture was adjusted to -13 with 10 M aqueous sodium hydroxide. The resulting mixture was heated at 80 °C for another 1 h. After cooling, the reaction mixture was acidified with concentrated hydrochloric acid to pH 1 and stirred overnight. The formed solids were collected by filtration and the filter cake was washed with water and dried in an oven at 50 °C for 24 h to afford 9.0 g (91%) of 17.2 as a brown solid. MS (ES): m/z 225 (M+H)+. [00323] Synthesis of compound 17.4. 17.4 was prepared from 17.3 in a manner analogous to the synthesis of 3.3 from 3.2. Isolated 4.0 g of a white solid. MS (ES): m/z 261 and 263 (M+H)+.
[00324] Synthesis of compound 17.5. Compound 17.5 was prepared from 17.4 in a manner analogous to the synthesis of intermediate 3.4 from 3.3. Isolated 140 mg (89%) of a white solid. MS (ES): m/z 409 and 411 (M+H)+.
[00325] Synthesis of Compound 1-13. Compound 1-13 was prepared from 17.5 in a manner analogous to the synthesis of 1-9, substituting 1 -methyl- lH-pyrazol-4-amine for l-(propan-2-yl)- lH-pyrazol-4-amine. MS (ES): m/z 470 (M+H)+; 1H-NMR (300 MHz, d DMSO) δ 8.94 (1H, brs), 7.80 (1H, brs), 7.45 (1H, s), 5.68 (1H, brs), 4.66 (2H, s), 4.05-3.85 (3H, m), 3.91 (3H, s), 3.57 (4H, brs), 2.95 (2H, brs), 2.40-2.28 (1H, m), 2.07 (2H, d), 1.90 (2H, d), 1.50-1.29 (4H, m).
[00326] Example 18: Synthesis of 12-N-[4-(dimethylamino)cyclohexyl]-10-N-phenyl-7- thia-9,ll-di (1-15)
Figure imgf000108_0001
[00327] Compound 1-15 was prepared from 5.1 in a manner analogous to the synthesis of 4.5 from 4.4 and 3.4. Isolated 20 mg (14%) of 1-15 as a grey solid. MS (ES): m/z 408 (M+H)+. 1H NMR (300 MHz, CD3OD) δ 6.67 (2H, d), 7.26 (2H, t), 6.94 (1H, t), 4.10-4.02 (1H, m), 3.01-2.92 (2H, m), 2.91-2.87 (2H, m), 2.55-2.45 (2H, m), 2.34 (7H, m), 2.23 (2H, brs), 2.03 (2H, brs), 1.46-1.42 (4H, m).
[00328] Example 19: Synthesis of 10-N-(l-methyl-lH-pyrazol-4-yl)-12-N-[4-(morpholin- 4-yl)cyclohexyl]-7-thia-9,ll-diazatricyclo [6.4.0.0 [2,6]]dodeca-l(8),2(6),9,ll-tetraene-10,12- diamine (1-16)
Figure imgf000109_0001
3.4 1-16
[00329] Compound 1-16 was prepared from 3.5 in a manner analogous to the synthesis of 4.5 from 4.4 and 3.4. Isolated 55.3 mg (47%) of 1-16 as a light yellow solid. MS (ES): m/z 454 (M+H)+. 1H NMR (300 MHz, CDC13): δ 7.84 (IH, s), 7.60-7.46 (IH, s), 6.91-6.52 (IH, s), 4.76- 4.73 (IH, d, J = 9.0 Hz), 4.04-3.95 (IH, m), 3.89 (3H, s), 3.78 (4H, s), 2.94-2.89 (4H, t, J = 7.5 Hz), 2.63 (4H, s), 2.53-2.44 (2H, m), 2.32-2.23 (3H, m), 2.15-2.04 (2H, m), 1.55-1.45 (2H, m), 1.31-1.19 (2H, m).
[00330] Example 20: Synthesis of 12-N-[trans-4-(morpholin-4-yl)cyclohexyl]-10-N-(l,2- thiazol-4-yl)-7-thia-9,ll-diazatricyclo[6.4.0.0[2,6]]dodeca-l(8),2(6),9,ll-tetraene
diamine 1-18
Figure imgf000109_0002
3.4 1-18
[00331] Compound 1-18 was prepared from 3.4 in a manner analogous to the synthesis of 1-9, substituting 1 ,2-thiazol-4-amine hydrochloride for l-(propan-2-yl)-lH-pyrazol-4-amine. Isolated 116.9 mg (67%) of a light brown solid. MS (ES): m/z 457 (M+H)+; 1H-NMR: (300 MHz, CDC13) δ 9.70 (IH, s), 8.77 (IH, s), 8.65 (IH, s), 5.84 (lH,d), 4.08-3.90 (m, IH), 3.57 (4H, brs), 3.03 (2H,t), 2.85 (2H, t), 2.49 4H, brs), 2.48-2.32 (2H, m), 2.28-2.10 (IH, m), 2.04 (2H, d), 1.90 (2H, d), 1.60-1.20 (4H, m).
[00332] Example 21: Synthesis of N-(l-methyl-lH-pyrazol-4-yl)-12-[[4-(morpholin-4- yl)cyclohexyl] oxy] -7-thia-9,l 1-diazatricyclo [6.4.0.0 [2,6] ] dodeca- 1 (8),2(6),9, 11-tetraen- 10- amine. (1-19)
Figure imgf000110_0001
[00333] Compound 1-19 was prepared from 3.3 in a manner analogous to the synthesis of 2.2 from 1.9. Isolated 32.9 mg of a white solid in 19% yield. MS (ES): m/z 455 (M+H)+. 1H NMR (300 MHz, CD3OD): δ 7.90 (s, 1H), 7.56 (s, 1H), 5.18-5.02 (m, 1H), 3.88 (s, 3H), 3.75-3.72 (m, 4H), 3.08-2.89 (m, 4H), 2.66-2.63 (m, 4H), 2.54-2.32 (m, 5H), 2.12-2.04 (m, 2H), 1.63-1.55 (m, 4H).
[00334] Example 22: Synthesis of 12-[[4-(dimethylamino)cyclohexyl]oxy]-N-phenyl-7- thia-9,ll-diazatri -l(8),2(6),9,ll-tetraen-10-amine. (1-20)
Figure imgf000110_0002
14.1 I-20
[00335] Compound 1-20 was prepared from 14.1 in a manner analogous to the synthesis of 2.2 from 2.1, substituting aniline for 1 -methyl- lH-pyrazol-4-amine. Isolated 51.1 mg (28%) of an off-white solid. MS (ES): m/z 408 (M+H)+. 1H NMR (400 MHz, CD3OD): δ 7.70 (d, J = 8.4 Hz, 2H), 7.30 (t, J = 7.6 Hz, 2H), 6.99 (t, J = 7.6 Hz, 1H), 5.19-5.13(m, 1H), 2.94-2.91 (m, 4H), 2.48-2.33 (m, 11H), 2.10-2.07 (m, 2H), 1.63-1.51 (m, 4H).
[00336] Example 23: Synthesis of 12-N-[trans-4-(morpholin-4-yl)cyclohexyl]-10-N-(l,2- oxazol-4-yl)-7-thia-9,ll-diazatricyclo[6.4.0.0[2,6]]dodeca-l(8),2(6),9,ll-tetraene-10,12- diamine (1-21) isopr
dioxa
micr
Figure imgf000111_0001
3.4 1-21
[00337] Compound 1-9 was prepared from 3.4 in a manner analogous to the synthesis of 1-9 from 3.4, substituting 1 ,2-oxazol-4-amine hydrochloride for l-(propan-2-yl)-lH-pyrazol-4- amine. Isolated 46.6 mg (28%) of a light brown solid. MS (ES): m/z 457 (M+H)+; 1H NMR (300 MHz, CDCls) δ 9.16 (1H, s), 8.97 (1H, s), 8.57 (1H, s), 5.84 (1H, d), 4.02-3.88 (m, 1H), 3.58- 3.54 (4H, m), 2.97 (2H, t), 2.84 (2H, t), 2.49 (4H, brs), 2.46-2.30 (2H, m), 2.25-2.10 (1H, m), 2.04 (2H, d), 1.90 (2H, d), 1.60-1.18 (4H, m).
[00338] Example 24: Synthesis of N4-((lr,4r)-4-morpholinocyclohexyl)-N2-phenyl-6,7- dihydro-5H-cyclopenta [4,5] thieno [2,3-d] pyrimidine-2,4-diamine. (1-22)
Figure imgf000111_0002
3.5 1-22
[00339] In a 50-mL round-bottom flask a solution of 3.4 (100 mg, 0.25 mmol, 1.00 equiv), aniline (118 mg, 4.99 equiv) and TMSC1 (276 mg, 2.54 mmol, 12.35 equiv) in n-butanol (20 mL) was stirred overnight at 90 °C in an oil bath. The reaction mixture was cooled to room temperature. The solids were collected by filtration and washed with 2x10 mL of ether. The solid was dried in an oven under reduced pressure. This resulted in 46.9 mg (38%) of 1-22 as an off- white solid. MS (ES): m/z = 450 [M-0.97HC1+H]+. [00340] Example 25: Synthesis of Intermediate
DDQ / dioxane
Figure imgf000112_0001
25.4 25.5
[00341] Synthesis of compound 25.2. Compound 25.2 was prepared according to the method for the synthesis of 1.2, substituting cyclohexanone for 1.1. Isolated 18 g (80%) of the desired 25.1 as a yellow solid.
[00342] Synthesis of compound 25.3. Into a 100-mL round-bottom flask containing a solution of 25.2 (4.06 g, 18.02 mmol, 1.00 equiv) in 100 mL of formamide was stirred for 8 h at 160 °C in an oil bath under nitrogen. The reaction mixture was cooled to 0°C and diluted with water. The solids were filtered and precipitated with ethyl acetate to provide 25.3 (3.0 g,81%) as a light yellow solid.
[00343] Synthesis of compound 25.4. To a solution of 25.3 (2.06 g, 9.99 mmol, 1.00 equiv) in 1,4-dioxane (30 mL) was added DDQ (5.68 g, 25.02 mmol, 2.50 equiv) at room temperature under nitrogen and the resulting solution was stirred overnight at 90 °C in an oil bath. After cooling to room temperature, the reaction was quenched with saturated aqueous sodium bicarbonate, and extracted with 3 x 100 mL of dichloromethane. The organic layers were combined, washed with brine, dried over anhydrous sodium sulfate, and concentrated under vacuum to provide 1.8 g (crude) of 25.4 as a yellow solid.
[00344] Synthesis of compound 25.5. Into a 100-mL round-bottom flask, was placed a solution of 25.4 (2.0 g, 9.89 mmol, 1.00 equiv) in 1,4-dioxane (15 mL) and POCl3 (4.5 g, 29.35 mmol, 2.97 equiv) under nitrogen. The resulting solution was stirred for 4 h at 90 °C. The excess amount of POCl3 was removed under reduced pressure and the residue was diluted with DCM. The resulting mixture was poured into a cooled saturated aqueous sodium bicarbonate, extracted with DCM, dried over anhydrous sodium sulfate and concentrated under vacuum. The residue was purified by silica gel column chromatography with ethyl acetate/petrolum ether (1 : 10) to afford 25.5 (0.9 g, 41%) as a white solid.
[00345] Example 26: N-((lr,4r)-4-morpholinocyclohexyl)benzo[4,5]thieno[2,3- d]pyrimidin-4-amine (1-65).
Figure imgf000113_0001
1-65
[00346] A solution of 25.5 (1.25 g, 5.66 mmol, 1.00 equiv) in N,N-dimethylformamide (20 mL) was added trans-dimethylcyclohexane-l,4-diamine dihydrochloride (1.66 g, 7.72 mmol, 1.36 equiv) and triethylamine (1.14 g, 11.27 mmol, 1.99 equiv) at room temperature under nitrogen. The resulting solution was stirred overnight at 50 °C in an oil bath. The resulting mixture was partially evaporated under reduced pressure. The solids were collected by filtration, washed with 10 mL of DCM and 5 mL of water. After dried in an oven at 45°C for 2 h, the desired compound 1-65 was obtained (1.0403 g, 51%) as an off-white solid. MS (ES): m/z 327 [M-0.95HC1+H]+. 1H-NMR: (300 MHz, d6-DMSO) δ 1.50-1.80 (4H, m), 2.00-2.20 (4H, m), 2.74 (6H, s), 3.09-3.25 (1H, m), 4.25-4.45 (1H, m), 7.03 (1H, d, J = 8.1 Hz), 7.52-7.63 (2H, m), 8.09 (1H, d, J= 7.2 Hz), 8.44 (1H, d, J= 7.2 Hz), 8.51(1H, s), 10.45 (1H, brs).
[00347] Example 27: Synthesis of (lr,4r)-Nl-(benzo[4,5]thieno[2,3-d]pyrimidin-4-yl)- N4,N4-dimethylcyclohexane-l,4-diamine (1-66)
Figure imgf000113_0002
25.5 1-66
[00348] Compound 1-66 was prepared from intermediate 25.5 in a manner analogous to the synthesis of 1-65. Isolated 256.6 mg (61%) of a white solid. MS (ES): m/z 369 (M+H)+. 1H NMR: (300 MHz, d6-DMSO) δ 1.41(2H, m), 1.58-1.71(2H, m), 1.82-1.93 (2H, m), 1.99-2.07 (2H, m), 2.20-2.28 (1H, m), 2.49-2.51 (4H, m), 3.56-3.59 (4H, t, J = 4.5Hz), 4.22-4.33 (1H, m), 6.90 (1H, d, J = 7.8Hz), 7.50-7.61 (2H, m), 8.06-8.10 (1H, m), 8.34-8.48 (1H, d, J = 13.5 Hz), 8.50 (1H, s).
[00349] Example 28: Synthesis of 2-(((lr,4r)-4-(benzo[4,5]thieno[2,3-d]pyrimidin-4- lamino)cyclohexyl)(methyl)amino)-l-(pyrrolidin-l-yl)ethanone. (1-67)
Figure imgf000114_0001
[00350] To a solution of intermediate 25.5 (300 mg, 1.36 mmol, 1.00 equiv) in CH3CN (25 mL) was added 3-(4-aminocyclohexyl)-l-(pyrrolidin-l-yl)butan-l-one (937 mg, 3.93 mmol, 3.00 equiv), potassium carbonate (1.88 g, 13.60 mmol, 10.00 equiv) and triethylamine (411 mg, 4.06 mmol, 3.00 equiv) subsequently at room temperature under nitrogen. The resulting solution was stirred overnight at 50 °C in an oil bath and quenched with water. The resulting solution was extracted with 3 x 40 mL of ethyl acetate and combined organic layers were washed with brine, dried over anhydrous sodium sulfate and concentrated under vacuum. The residue was applied onto a silica gel column with dichloromethane/methanol/NH4OH (10: 1 :0.05) to afford 1-67 (232.3 mg, 40%) as a white solid. MS (ES): m/z 424 [M+H]+. 1H NMR (400 MHz, CDC13): δ 8.60 (s, 1H), 7.95-7.93 (d, 1H), 7.81-7.79 (d, 1H), 7.58-7.49 (m, 2H), 5.36-5.34 (d, 1H), 4.32- 4.28 (m, 1H), 3.57-3.50 (m, 4H), 3.32 (s, 2H), 2.70 (m, 1H), 2.43-2.37 (m, 5H), 2.05-2.02 (m, 4H), 2.00-1.91 (m, 2H), 1.89-1.86 (m, 2H), 1.63-1.54 (dd, 2H), 1.48-1.38 (m, 2H).
[00351] Example 29: Synthesis of (lr,4r)-4-(benzo[4,5]thieno[2,3-d]pyrimidin-4-yloxy)- Ν,Ν-dimethylcyclohexanamine. (1-68)
Figure imgf000114_0002
[00352] Compound 1-68 was prepared from 25.5 in a manner analogous to the synthesis of 10.1 from 1.8. Isolated 22.1 mg (10%) of a white solid. MS (ES): m/z 328 (M+H)+; 1H-NMR (400 MHz, CDCls) δ 1.50-1.61 (2H, m), 1.69-1.79 (2H, m), 2.05-2.09 (2H, m), 2.36 (6H, s), 2.38-2.44 (2H, m), 5.42 (1H, m), 7.51-7.57 (2H, m), 7.89-7.92 (1H, m), 8.41-8.45 (1H, m), 8.72 (1H, s).
[00353] Example 30: Synthesis of Intermediate 30.9.
Figure imgf000115_0001
"*"■' 30.8 30.9
[00354] Synthesis of compound 30.2. Into a 1 L pressure tank reactor (60 atm) containing a solution of 3-hydroxybenzoic acid (30 g, 217.20 mmol, 1.00 equiv), Raney Ni (5 g) and sodium hydroxide (6.4 g, 160.00 mmol, 0.74 equiv) in water (500 mL) was introduced H2 (gas, 60 atm) and the resulting solution was stirred overnight at 150°C. After completion of the reaction, the reaction temperature was cooled down to room temperature and the solids were filtered out by filtration. The resulting solution was neutralized with 12 M HC1, extracted with 6 x 100 mL of tetrahydrofuran. The combined organic layers were dried over sodium sulfate and concentrated under vacuum to provide 30.2 (11 g, crude) as a white solid.
[00355] Synthesis of compound 30.3. The Jones oxidation reagent was prepared from sulfuric acid (30 mL), Cr03 (8.1 g) and H20 (30 mL) in an ice/water bath. To a solution of 30.2 (11 g, 76.3 mmol, 1.00 equiv) in acetone (150mL) was added slowly the prepared Jones reagent at 0 °C in 30 min. After addition, the resulting solution was stirred for 2 h at room temperature and the solids were filtered out. The resulting solution was extracted with DCM (3 xlOO mL). The combined organic layers were washed with brine and dried over sodium sulfate and concentrated under vacuum to give the desired 30.3 (8.7 g, crude) as a yellow oil.
[00356] Synthesis of compound 30.4. A solution of 30.3 (8.7 g, 61.20 mmol, 1.00 equiv) and ethanol (21 mL) in toluene (150 mL) was added /?-toluenensulfonic acid (250 mg) at room temperature and the resulting solution was heated to reflux overnight in presence of a Dean- Stark. After completion, the resulting mixture was concentrated under vacuum and the residue was applied onto a silica gel column with ethyl acetate/petroleum ether (1 :4) to give the desired 30.4 (7.5 g, 72%) as white oil.
[00357] Synthesis of compound 30.5. Into a 500-mL round-bottom flask, was placed 30.4 (7 g, 41.13 mmol, 1.00 equiv), ethyl 2-cyanoacetate (6.5 g, 57.46 mmol, 1.40 equiv), AcOH (0.8 mL) and NH4OAc (300 mg) in toluene (150 mL). The resulting solution was heated to reflux overnight in presence of a Dean-Stark. After the starting material was consumed completely, the resulting mixture was concentrated under vacuum. The residue was applied onto a silica gel column with ethyl acetate/petroleum ether (1 :9) to give the corresponding 30.5 (8.0 g, 73%) as a yellow oil.
[00358] Synthesis of compound 30.6. A solution of 30.5 (8 g, 30.15 mmol, 1.00 equiv) in methanol (100 mL) was added S (970 mg, 30.31 mmol, 1.01 equiv) and Et2NH (3.2 mL) and the resulting solution was stirred for 2 h at 50 °C under nitrogen. The resulting mixture was concentrated under vacuum and the residue was applied onto a silica gel column with ethyl acetate/petroleum ether (1 :9) to give 30.6 (8.0 g, 88%) as a yellow solid.
[00359] Synthesis of compound 30.7. A solution of 30.6 (8 g, 26.90 mmol, 1.00 equiv) in formamide (150 mL) was added formamidine acetate (10 g, 96.05 mmol, 3.57 equiv) at room temperature under nitrogen. The resulting solution was stirred for 4 h at 180°C. After cooling, the resulting solution was diluted with 200 mL of EtOAc, washed with water and brine, dried over anhydrous sodium sulfate and concentrated under vacuum. The residue was applied onto a silica gel column with ethyl acetate/petroleum ether (1 :4) give the desired 30.7 (4.1 g, 55%) as a yellow solid. [00360] Synthesis of compound 30.8. To a 250-mL round-bottom flask was placed a solution of 30.7 (4.0 g, 14.39 mmol, 1.00 equiv) in 1.4-dioxane (50 mL) and then DDQ (8 g, 35.24 mmol, 2.45 equiv) was added at room temperature. The resulting solution was stirred for 3 h at 90 °C under nitrogen. After cooling, the reaction was quenched with saturated aqueous Na2C03. The solids were filtered out and the filtration was extracted with 3 x 100 mL of dichloromethane. The combined organic layers were washed with brine, dried over anhydrous sodium sulfate and concentrated under vacuum. The residue was applied onto a silica gel column with ethyl acetate/petroleum ether (1 : 1) to give 30.8 (2.2 g, 56%) as a brown solid.
[00361] Synthesis of intermediate 30.9. Into a 100-mL round-bottom flask containing a solution of 30.8 (2.2 g, 4.01 mmol, 1.00 equiv) in 1.4-dioxane (60 mL) was added POCl3 (30 mL) at room temperature under nitrogen. The resulting solution was stirred for 2 h at 90°C and concentrated under vacuum. The residue was dissolved in DCM and poured into a cooled saturated aqueous Na2C03 and extracted with 3 x 100 mL of dichloromethane. The combined organic layers were washed with brine, dried over anhydrous sodium sulfate and concentrated under reduced pressure. The residue was applied onto a silica gel column with ethyl acetate/petroleum ether (1 :9) to give 1.8 g (77%) of the corresponding 30.9 as a yellow solid.
[00362] Example 31: Synthesis of (4-(((lr,4r)-4-
(dimethylamino)cyclohexyl)amino)benzo [4,5] thieno [2,3-d] pyrimidin-6-yl)methanol (1-69)
Figure imgf000117_0001
1-69
[00363] Synthesis of compound 31.1. To a solution of 30.9 (500 mg, 1.71 mmol, 1.00 equiv) in butan-2-ol (15 mL) was added triethylamine (300 mg, 2.96 mmol, 1.74 equiv) and trans-1- N,l-N-dimethylcyclohexane-l,4-diamine hydrochloride (730 mg, 3.41 mmol, 2.00 equiv) and the resulting solution was stirred for 5 h at 60 °C under nitrogen. After cooling, the resulting mixture was concentrated under vacuum. The residue was applied onto a silica gel column with dichloromethane/methanol/NH40H (10:1 :0.05) to provide 600 mg (88%) of the corresponding 31.1 as a yellow solid. MS (ES): m/z 399 [M+H]+. 1H-NMR (400 MHz, CDC13): δ 1.27-1.33 (1H, m), 1.33-1.66 (6H, m), 2.04-2.07 (2H, m), 2.34-2.50 (9H, m), 4.28-4.31 (1H, m), 4.48-4.52 (2H, m), 5.45-5.47(lH, d), 7.97-7.99 (1H, d), 8.15-8.17 (1H, d), 8.52 (1H, s), 8.63(1H, s).
[00364] Synthesis of Compound 1-69. To a solution of 31.1 (80 mg, 0.20 mmol, 1.00 equiv) in 5 mL of distilled THF was added LiAlH4 (16 mg, 0.42 mmol, 2.10 equiv) at 0 °C under nitrogen. The resulting solution was stirred for 2 h at room temperature. After completion, the mixture was diluted with THF (50 mL) and quenched with Na2S04 H20. The solids were filtered out and the filtrate was concentrated under vacuum. The residue was applied onto a silica gel column with dichloromethane/methanol/NH4OH (5: 1 :0.05) to give 16.8 mg (23%) of the desired compound as a white solid. MS (ES): m/z 357 [M+H]+. 1H-NMR (300 MHz, CD3OD) δ 1.56- 1.85 (4H, m), 2.10-2.19 (2H, m), 2.29-2.33 (2H, m), 2.73 (6H, s), 2.73-2.88 (1H, m), 4.32-4.37 (1H, m), 4.84 (2H, s), 7.53-7.55 (1H, d), 7.95-7.97 (1H, d), 8.19 (1H, s), 8.46 (1H, s).
[00365] Example 32: Synthesis of 4-((lr,4r)-4-(benzo[4,5]thieno[2,3-d]pyrimidin-4- yloxy)cyclohexyl)morpholine (1-70)
Figure imgf000118_0001
1-70
[00366] Compound 1-70 was prepared from 25.5 in a manner analogous to the synthesis of 10.1 from 1.8. Isolated 29 mg (14%) of a white solid. MS (ES): m/z 370 (M+H)+. 1H-NMR (300 MHz, CD3OD) δ 1.45-1.60 (2H, m), 1.66-1.80 (2H, m), 2.10 (2H, m), 2.41 (2H, m), 2.68 (4H, m), 3.78 (4H, m), 5.40 (1H, m), 7.52-7.56 (2H, m), 7.96 (1H, m), 8.40 (1H, m), 8.65 (1H, s).
[00367] Example 33: Synthesis of Intermediate 33.1.
Figure imgf000119_0001
1.2 33.1
[00368] Synthesis of compound 33.1. To a 10-L 4-necked round-bottom flask, purged and maintained with an inert atmosphere of nitrogen, was added a solution of 1.2 (500 g, 1.68 mol, 1.00 equiv) in formamide (5 L) at room temperature. The resulting solution was stirred for 5 h at 180 °C in an oil bath. The reaction mixture was cooled to room temperature and then quenched by the addition of 10 L of water/ice. The resulting solution was extracted with 3 x 5 L of ethyl acetate and the organic layers were combined. The mixture was washed with 3 x 3000 mL of brine, dried over anhydrous sodium sulfate and concentrated under vacuum. The solids were collected by filtration to afford 200 g (43%) of 33.1 as a yellow solid.
[00369] Example 34. Synthesis of Intermediates 34.3 and 34.4.
Figure imgf000119_0002
34.4
[00370] Synthesis of compound 34.1. A solution of 33.1 (15 g, 53.89 mmol, 1.00 equiv) and POCI3 (100 mL) in 100 mL of dioxane was heated at reflux for 3 h under nitrogen. After concentration under reduced pressure, the resulting solution was poured dropwise into saturated aqueous NaHC03 and extracted with ethyl acetate (3 x 150 mL). The combined organic layers were washed with brine and dried over sodium sulfate. After evaporation in vacuo, the residue was purified by column chromatography on silica gel with ethyl acetate/petroleum ether (1 :7) to afford 34.1 (15 g, 94%) as a light yellow oil. MS: m/z 297, 299 (M+H)+.
[00371] Synthesis of compound 34.2. To a 500-mL round-bottom flask under an atmosphere of nitrogen was added 34.1 (6 g, 20.22 mmol, 1.00 equiv) in 100 mL of distilled THF at -50 °C. DIBAL-H (25% w/w in hexane, 50 mL) was added dropwise and the resulting solution was stirred for 2 h at under -30 °C under nitrogen. The reaction was quenched with saturated aqueous ammonium chloride and extracted with ethyl acetate (2 x 150 mL). The combined organic layers were washed with brine, dried over anhydrous sodium sulfate and concentrated under vacuum. The residue was purified by column chromatography on silica gel with EtO Ac/petroleum ether (1 :5 to 1 : 1) to afford 34.2 (5.0 g, 97%) as a yellow solid. MS: m/z 255, 257 (M+H)+.
[00372] Synthesis of compounds 34.3 and 34.4. The enantiomers of racemic 34.2 (5.0 g, 19.6 mmol) were separated by chiral-SFC under the following conditions: column: CHIRALPAK IA; 20% methanol with C02; flow rate: 250 mL/min; UV detection at 254 nm. The fractions corresponding to the peak with tR = 1.63 were collected and the methanol removed in vacuo to give enantiomerically pure 34.3 (2.0 g) in 100% ee. Similar treatment of the fractions corresponding to the peak with tR = 2.69 gave enantiomerically pure 34.4 (2.0 g) in 100% ee.
[00373] Example 35: Synthesis of Intermediate 35.1.
HC
Figure imgf000120_0001
[00374] Intermediate 34.3 (1.3 g, 5.10 mmol, 1.00 equiv) was treated with imidazole (500 mg, 7.44 mmol, 1.40 equiv) and TBDMSC1 (920 mg, 6.10 mmol, 1.20 equiv) in distilled DMF (10 mL) for 2 h at room temperature under nitrogen. The reaction was then quenched with water and extracted with ethyl acetate (3 x 50 mL). The combined organic layers were washed with brine and dried over anhydrous sodium sulfate. After filtration and concentration under reduced pressure, the residue was loaded onto a silica gel column with ethyl acetate/petroleum ether (1 :5) and purified to give 35.1 (1.8 g, 96%) as a yellow oil.
[00375] Example 36: Synthesis of (S)-2-hydroxy-3-((R)-4-(((lr,4R)-4-(pyrrolidin-l- yl)cyclohexyl)oxy)-6,7-dihydro-5H-cyclopenta [4,5] thieno [2,3-d] pyrimidin-5- yl)propanamide (1-41) and Example 37: Synthesis of (R)-2-hydroxy-3-((R)-4-(((lr,4R)-4- (pyr rolidin- l-yl)cyclohexyl)oxy)-6,7-dihydro-5H-cyclopenta [4,5] thieno [2,3-d] pyrimidin-5- yl)propanamide (1-24)
Figure imgf000121_0001
35.1 36.1
36.2 36.3
Figure imgf000121_0002
36.4 36.5
Figure imgf000121_0003
36.6 36.7
Figure imgf000121_0004
36.8 1-41 I-24
[00376] Synthesis of compound 36.1. Compound 36.1 was prepared from 35.1 and trans-4- (pyrrolidin-l-yl)cyclohexan-l-ol in a manner analogous to the synthesis of 10.1 from 1.8. Isolated 1.09 g (89%) of 36.1 as a yellow oil.
[00377] Synthesis of compound 36.2. To a solution of 36.1 (1.05 g, 2.09 mmol, 1.00 equiv) in dichloromethane (50 mL) was added hydrochloric acid (12 M, 1 mL) and stirred for 1 h at 0 °C. The pH value of the solution was adjusted to 8 with saturated aqueous sodium bicarbonate, extracted with 3 x 50 mL of dichloromethane. The combined organic layers were washed with brine, dried over sodium sulfate and concentrated under vacuum. The residue was applied onto a silica gel column with dichloromethane/methanol (10/1) to afford 0.6 g (74%) of 36.2 as a colorless oil.
[00378] Synthesis of compound 36.3. To a solution of 36.2 (600 mg, 1.55 mmol, 1.00 equiv) in dichloromethane (20 mL) was added Dess-Martin periodinane (0.98 g, 1.50 equiv) at 0 °C. The resulting solution was stirred for 3 h at room temperature and diluted with water, extracted with 3 x 50 mL of dichloromethane. The combined organic layers were washed with brine, dried over sodium sulfate and concentrated under vacuum. The residue was applied onto a silica gel column with dichloromethane/methanol (10/1) to afford the desired 36.3 (0.45 g, 75%) as a white solid.
[00379] Synthesis of compound 36.4. Into a 100 mL round-bottom flask containing a solution of 36.3 (450 mg, 1.17 mmol, 1.00 equiv) in dichloromethane (20 mL) was added trimethylsilanecarbonitrile (360 mg, 3.63 mmol, 3.00 equiv) and TEA (60 mg, 0.59 mmol, 0.50 equiv) and the resulting solution was stirred for 2 h at room temperature under nitrogen. The resulting solution was quenched with water and extracted with 3 x 50 mL of dichloromethane. The combined organic layers were concentrated under vacuum to give 0.52 g (crude) of 36.4 as a yellow oil.
[00380] Synthesis of compound 36.5. To a solution of 36.4 (520 mg, 1.07 mmol, 1.00 equiv) in 20 mL of MeOH was added acetic acid (1.0 mL) at 0 °C and the resulting solution was stirred for 1 h at room temperature. The pH value of the solution was adjusted to 8 with saturated aqueous sodium bicarbonate and extracted with 3 x 50 mL of dichloromethane. The combined organic layers were dried over sodium sulfate and concentrated under vacuum. The residue was applied onto a silica gel column with dichloromethane/methanol (8/1). This resulted in 0.37 g (84%) of 36.5 as a white solid.
[00381] Synthesis of compound 36.6. Into a 50-mL round-bottom flask containing a solution of 36.5 (370 mg, 0.90 mmol, 1.00 equiv) in dichloromethane (20 mL) was added TBSC1 (0.41 g, 3.00 equiv), imidazole (0.24 g, 4.00 equiv) and 4-dimethylaminopyridine (24 mg) sequentially at room temperature. The resulting solution was stirred overnight at ambient temperature and quenched with water and extracted with 3 x 50 mL of dichloromethane. The organic layers were combined, dried over sodium sulfate and concentrated under reduced pressure. The residue was purified by silica gel column with DCM/MeOH (30: 1 to 10: 1) to provide 0.33 g (70%) of 36.6 as a yellow oil. [00382] Synthesis of compound 36.7. Tris(triphenylphosphine)rhodium(I) chloride (18.0 mg, 0.019 mmol) was added to a stirred solution of 36.6 (330 mg, 0.63 mmol) and acetaldoxime (0.23 mL, 3.60 mmol) in toluene (5.0 mL) and the reaction mixture heated at reflux overnight. Then tris(triphenylphosphine)rhodium(I) chloride (4.6 mg, 0.005 mmol) and acetaldoxime (62 μί, 1.0 mmol) were again added and heating continued for 2 h. After completion, the mixture was concentrated, diluted with ethyl acetate, and the organic layer washed with water, brine, dried, and concentrated in vacuo. The residue was purified by chromatography on silica gel eluting with a gradient of 0-10% MeOH in DCM to give the desired product 36.7 (250 mg) as a light yellow foam.
[00383] Synthesis of compound 36.8. Into a 100-mL round-bottom flask, a solution of material 36.7 (250 mg, 0.46 mmol, 1.00 equiv) in methanol (10 mL) was added hydrochloric acid (2 M, 0.8 mL) and stirred for 2 h in a water/ice bath. After completion, the reaction was quenched with saturated aqueous sodium bicarbonate and extracted with 3 x 30 mL of DCM. The organic phase was dried over sodium sulfate and concentrated under vacuum. The residue was purified by preparative TLC (DCM/MeOH: 10/l) to afford the desired product 36.8 (150 mg, 76%) as a white solid.
[00384] Synthesis of Compound 1-41 and 1-24. The enantiomers of racemic alcohol 36.8 (150 mg, 96%o purity) were separated by chiral HPLC under the following conditions (Gilson G x 281): Column: Chiralpak IA, 2*25 cm, 5 μιη; mobile phase: phase A: hexanes (0.1% IPA) (HPLC grade), phase B: EtOH (HPLC grade), gradient: 20% B in 7.6 min; flow rate: 20 mL/min; UV detection at 220/254 nm. The fractions of the first enantiomer to elute were collected and evaporated under reduced pressure and lyophilized overnight to afford 1-41 (36.2 mg) with 100% ee as a white solid. The fractions of the second enantiomer to elute were concentrated to give 1-24 (40.0 mg) with 94.2% ee, which was resubjected to the chiral HPLC conditons to give 30 mg with 98.4% ee as a white solid. The ee values of the two isomers were determined by chiral HPLC under the following conditions (SHIMADZU-PDA): Column: Chiralpak IA-3, 0.46* 15 cm, 3 μιη; mobile phase: hexanes (0.2% IPA): EtOH = 80:20; flow rate: 1.0 mL/min; UV detection at 254 nm.
[00385] Analytical data for Compound 1-41: MS (ES): m/z 431 (M+H)+. 1H-NMR (300 MHz, CD3OD): δ 8.45 (s, 1H), 5.31-5.24 (m, 1H), 4.06 (dd, J = 10.8, 2.4 Hz, 1H), 3.72-3.61 (m, 1H), 3.22-3.08 (m, 1H), 3.05-2.89 (m, 1H), 2.80-2.60 (m, 5H), 2.50-2.09 (m, 7H), 1.95-1.62 (m, 7H), 1.56-1.40 (m, 2H).
[00386] Analytical data for Compound 1-24: MS (ES): m/z 431 (M+H)+. 1H-NMR (300 MHz, CD3OD): δ 8.46 (s, 1H), 5.38-5.20 (m, 1H), 4.14 (dd, J = 7.5, 5.7 Hz, 1H), 3.60-3.50 (m, 1H), 3.15-3.06 (m, 1H), 3.04-2.92 (m, 4H), 2.78-2.60 (m, 2H), 2.55-2.41 (m, 2H), 2.40-2.15 (m, 4H), 1.94 (brs, 4H), 1.85-1.45 (m, 5H).
[00387] Example 38: Synthesis of (2S)-2-hydroxy-3-[(3R)-12-[[4-
(methylamino)cyclohexyl]oxy]-7-thia-9,ll-diazatricyclo[6.4.0.0[2,6]]dodeca-l(8),2(6),9,ll- tetraen-3-yl]propanamide (1-25)
Figure imgf000124_0001
38.9
[00388] Synthesis of compound 38.5. Compound 38.5 was prepared from 35.1 in a manner analogous to 36.4, substituting tert-butyl ((lr,4r)-4-hydroxycyclohexyl)(methyl)carbamate for trans-4-(pyrrolidin-l-yl)cyclohexan-l-ol and using TBAF/THF rather than HCl/DCM to cleave the TBS group in the second step. Isolated 402 mg of a yellow oil in 89% yield from 35.1.
[00389] Synthesis of compound 38.6. A 38.1 (402 mg, 0.74 mmol, 1.00 equiv) and TBAF.3H20 (349 mg, 1.11 mmol, 1.50 equiv) in tetrahydrofuran (20 mL) was stirred for 1 h at room temperature. The resulting mixture was concentrated under vacuum and purified onto a silica gel column with ethyl acetate/petroleum ether (1 : 1). This resulted in 38.2 (301 mg, 86%) as a colorless oil.
[00390] Synthesis of compound 38.7. A solution of 38.6 (301 mg, 0.64 mmol, 1.00 equiv), imidazole (73 mg, 1.07 mmol, 1.69 equiv) and TBSC1 (150 mg, 1.00 mmol, 1.57 equiv) in N,N- dimethylformamide (7 mL) was stirred overnight at room temperature. The resulting solution was diluted with 30 mL of water, extracted with 3x40 mL of ethyl acetate, washed with 50 mL of brine, concentrated under vacuum and purified onto a silica gel column with ethyl
acetate/petroleum ether (1 :3). This resulted in 38.7 (350 mg, 94%) as a colorless oil.
[00391] Synthesis of compounds 38.8 and 38.9. A solution of 38.7 (420 mg, 0.72 mmol, 1.00 equiv), LiOH.H20 (60 mg, 1.43 mmol, 2.00 equiv) and H202 (30%) (0.5 mL) in methanol (20 mL) was stirred for 2 h at 0 °C in a water/ice bath. The reaction was then quenched by the addition of 30 mL of saturated Na2S03. The resulting solution was extracted with 3x40 mL of ethyl acetate, concentrated under vacuum and purified by preparative TLC (PE/EA = 1 : 1). This resulted in 38.8 (180 mg) as a colorless oil and 38.9 (160 mg) as a colorless oil.
[00392] Synthesis of Compound 1-25. 1-25 was prepared from 38.4 according to the method for the preparation of 1-7 from 4.5. Isolated 48.3 mg (47%) as a white solid. The product was confirmed by LCMS and H-NMR.
1H NMR (400 MHz, CD3OD, ppm) δ 8.46 (s, 1H), 5.30 (m, 1H), 4.16 (m, 1H), 3.59 (m, 1H), 3.16-2.94 (m, 2H), 2.77-2.68 (m, 1H), 2.58-2.52 (m, 6H), 2.48-2.10 (m,4H), 1.75-1.72 (m, 3H), 1.66-1.34 (m, 2H).
MS (ES): m/z = 391 [M+H]+.
[00393] Example 39: Synthesis of (2S)-3-[(3R)-12-[[4-(ethylamino)cyclohexyl]oxy]-7-thia-
9,ll-diazatricyclo[6.4.0.0[2,6]]dodeca-l(8),2(6),9,ll-tetraen-3-yl]-2-hydroxypropanamide
(1-26)
Figure imgf000126_0001
Figure imgf000126_0002
39.4 39.5 1-26
[00394] Synthesis of compound 39.3. Compound 39.3 was prepared from 35.1 in a manner analogous to the synthesis of 38.3, substituting 39.1 for 38.1 in the first step.
[00395] Compound 1-26 was prepared in a manner analogous to Compound 1-25, substituting tert-butyl ((lr,4r)-4-hydroxycyclohexyl)(ethyl)carbamate for tert-butyl ((lr,4r)-4- hydroxycyclohexyl)(methyl)carbamate in the first step. Isolated 47.3 mg (22% overall yield) as a white solid. MS (ES): m/z 405 [M+H]+ . 1H-NMR (300 MHz, CD3OD, ppm): δ 8.474(lH,s), 5.24-5.26 (1H, m), 4.08-4.12 (1H, d), 3.53(1H, m), 3.04 -3.28 (1H, m), 2.92-2.95(lH, m), 2.50- 2.71 (4H, m), 2.40-2.47(2H, m),2.15-2.23(2H, m), 1.98-2.07 (2H, m),1.61-1.96(3H, m), 1.26- 1.34(2H, m), 1.08-1.13(3H, d).
[00396] Example 40: Synthesis of (2S)-3-[(3R)-12-[[4-(dimethylamino)cyclohexyl]oxy]-7- thia-9,11-diazatricyclo [6.4.0.0 [2,6] ] dodeca-l(8),2(6),9,l l-tetraen-3-yl]-2- hydroxypropanamide formate (1-27)
Figure imgf000127_0001
Figure imgf000127_0002
40.2 1-27
[00397] Synthesis of compound 40.1. 650 mg of the racemic 38.6 was separated by Prep- Chiral-HPLC with the following conditions (Gilson Gx 281): column, Chiralpak IC(SFC), 2*25cm, 5um; mobile phase: hex : EtOH = 80:20; flow rate: 20 mL/min; UV detection at 254/220 nm. The product-containing eluents were collected and evaporated to remove the solvents under reduced pressure to give the desired enantiomerically pure compound (0.23 g, tR = 9.83) with 100% ee.
[00398] Synthesis of compound 40.2. Compound 40.2 was prepared in a manner analogous to the synthesis of 38.8. Isolated a white solid in 46% yield (110 mg). MS: (ES, m/z 491(M+H) +.
[00399] Synthesis of Compound 1-27. Into a 10-mL round-bottom flask was placed a solution of 40.2 (110 mg, 0.22 mmol, 1.00 equiv) in dichloromethane (5.5 mL) at 0 °C under nitrogen. Then hydrochloric acid (12 M, 0.5 mL) was added and the resulting solution was stirred for 2 h at 0 °C. After completion of the reaction, the solvents were evaporated under reduced pressure. The residue was neutralized with 2 M aqueous sodium bicarbonate and extracted with 3 x 20 mL of dichloromethane. The organic layers were combined and washed with brine, dried over anhydrous sodium sulfate and concentrated under vacuum to give 80 mg (crude) of (2S)-2-hydroxy-3-[(3R)-12-[[4-(methylamino)cyclohexyl]oxy]-7-thia-9,l 1- diazatricyclo[6.4.0.0[2,6]]dodeca-l(8),2(6),9,l l-tetraen-3-yl]propanamide as a yellow oil. The corresponding product (80 mg, crude) and HCHO (37%, 0.8 mL) in methanol (4 mL) was stirred at room temperature for 30 min. NaBH3CN (39 mg, 0.62 mmol) was added and the resulting solution was stirred for 12 h at room temperature. The crude product (65 mg) was purified by preparative HPLC with the following conditions (SHIMADZU): column: SunFire Prep CI 8, 19* 150mm 5um; mobile phase: water with 0.1% HCOOH and CH3CN (6.0% CH3CN up to 55.0% in 19 min); UV detection at 254/220 nm. The product containing fractions were collected and evaporated to remove the solvents under reduced pressure to afford the resulted 1-27 (39.9 mg) as a white solid. MS: (ES, m/z 405(M+H) +. 1H-NMR (300 MHz, CD3OD) δ 8.48 (1H, br s), 8.42 (2H, d), 5.29 (1H, m), 4.10 (1H, m), 3.62 (1H, m), 2.90-3.13 (9H, m), 2.68 (1H, m),2.14- 2.55 (6H, m), 1.76 (5H, m).
[00400] Example 41: Synthesis of 3-[(3R)-12-[[4-(dimethylamino)cyclohexyl]oxy]-7-thia- 9,ll-diazatricyclo[6.4.0.0[2,6]]dodeca-l(8),2(6),9,ll-tetraen-3-yl]-2-hydroxypropanamide
(1-28
Figure imgf000128_0001
1-28
Compound 1-28 was prepared from 38.6 in a manner analogous to the synthesis of 1-27, except that the chiral resolution step was omitted. Isolated 26.4 mg (19%) of a white solid. MS (ES): m/z 405 (M+H) +. 1H-NMR (400MHz, CDC13) δ 8.52 (1H, s),6.49 (1H, d), 5.72 (1H, d), 5.23 (1H, m), 4.16 (1H, m), 3.68 (2H, m), 3.14 (1H, m), 3.01 (1H, m), 2.74 (1H, m), 2.23-2.39 (10H, m), 2.01 (3H, m), 1.27-1.47 (4H, m).
[00401] Example 42: Synthesis of (2S)-3-[(3R)-12-[(4-aminocyclohexyl)oxy]-7-thia-9,ll- iazatricyclo [6.4.0.0 [2,6] ] dodeca-l(8),2(6),9,l l-tetraen-3-yl] -2-hydroxypropanamide (1-29)
Figure imgf000128_0002
1-29 [00402] Compound 1-29 was prepared in a manner analogous to Compound 1-25, substituting tert-butyl ((lr,4r)-4-hydroxycyclohexyl)carbamate for tert-butyl ((lr,4r)-4- hydroxycyclohexyl)(methyl)carbamate in the first step. Isolated 47.3 mg (22% overall yield) as a white solid. Isolated 30.9 mg (10% overall yield) as a white solid. MS (ES): m/z 377 [M+H]+. 1H NMR (400 MHz ,CD3OD): δ 8.47 (1H, br s), 5.33 (lH,m) 4.15(1H, m), 3.50 (1H, m), 3.12 (2H, m), 3.00 (1H, m), 2.98 (1H, m), 2.88-2.53(3H, m), 2.47-2.20(4H,m), 1.87-1.66(5H,m), 1.42(2H,m)
[00403] Example 43: Synthesis of 3-[[4-(morpholin-4-yl)cyclohexyl]amino]-8-thia-4,6- diazatricyclo[7.4.0.0[2,7]]trideca-l(9),2,4,6,10,12-hexaene-12-carboxamide (1-30)
Figure imgf000129_0001
1-30
[00404] Synthesis of compound 43.1. Compound 43.1 was prepared from 30.9 in a manner analogous to the synthesis of 1-67 from 25.4. Isolated 500 mg 66%>) as a yellow solid. MS: (ES, m/z) 441 [M+H]+. 1H-NMR (300 MHz, CD3OD): δ 1.44-1.53 (4H, m), 1.57-1.65 (3H, m), 2.24- 2.29 (2H, m), 2.25-2.33 (3H, m), 2.56-2.67 (4H, m), 3.45-3.75 (4H, m), 4.31-4.43 (1H, m), 4.45- 4.50 (2H, m), 8.07-8.16 (2H, m), 8.49 (1H, s), 8.78 (1H, s).
[00405] Synthesis of compound 43.2. A solution of 43.1 (100 mg, 0.23 mmol, 1.00 equiv) in 6 M aqueous HC1 (20 mL) was stirred for 2 h at 80 °C. The resulting mixture was concentrated under vacuum to give 85 mg (crude) of 43.2 as a yellow solid.
[00406] Synthesis of compound 43.3. A solution of 43.2 (85 mg, crude) in dichloromethane (5 mL) was added thionyl chloride (4 mL) at room temperature under nitrogen. The resulting solution was stirred for 2 h at 50 °C and concentrated under vacuum to give 43.3 (80 mg, crude) as a yellow solid which was used directly for next step without further purification.
[00407] Synthesis of Compound 1-30. Into a 25-mL round-bottom flask purged and maintained with an inert atmosphere of nitrogen, was placed a solution of 43.3 (40 mg, 0.09 mmol, 1.00 equiv) in dichloromethane (5 mL) at 0 °C under nitrogen. Then ammonia-THF (5 mL) was added via syringe and the resulting solution was stirred overnight at room temperature. The resulting mixture was concentrated under vacuum. The crude product (50 mg) was purified by prep-HPLC with the following conditions (Waters): column: Xbridge Prep C 18, 5um, 19*50 mm; mobile phase: Water with 0.05% NH4HCO3 and CH3CN (10% CH3CN up to 25% in 10 min, up to 95% in 1.5 min, down to 10% in 1.5 min); flow rate: 20 mL/min; UV detection at 254/220 nm. After collection and evaporation in vacuo, 1-30 (19 mg, 50%) was obtained as a white solid. MS: (ES, m/z) 412 [M+H]+. 1H-NMR (300 MHz, d6-DMSO): δ 1.36-1.44 (2H, m), 1.60-1.64 (2H, m), 1.91-1.95 (2H, m), 2.10-2.13 (2H, m), 2.27-2.50 (1H, m), 2.50-2.72 (4H, m), 3.57-3.59 (4H, m), 4.24-4.26 (1H, m), 7.01-7.04 (1H, d), 7.58 (1H, s), 7.96-7.99 (1H, d), 8.14- 8.21 (2H, m), 8.52-8.58 (2H, m).
[00408] Example 44: Synthesis of Intermediate 44.1.
Figure imgf000130_0001
44.1
[00409] Intermediate 44.1 was prepared from 2-oxocyclopentanane-l-carboxylate in a manner analogous to the synthesis of 34.1 from 1.1. Isolated 260 g (69%) of intermediate 44.1 as a white solid. Example 45: Synthesis of intermediate 45.1.
Figure imgf000131_0001
45.1
[00411] Synthesis of compound 45.1. Intermediate 45.1 was prepared from 44.1 in a manner analogous to the synthesis of 35.1 from 34.1. Isolated 2.0 g (35%) as a yellow oil in 100% ee. MS (ES): m/z 355 (M+H)+.
[00412] Example 46: Synthesis of 2-cyano-3-((R)-4-(((lr,4R)-4- morpholinocyclohexyl)oxy)-6,7-dihydro-5H-cyclopenta [4,5] thieno [2,3-d] pyrimidin-5- yl)propanamide (1-31)
Figure imgf000131_0002
Mol. Wt: 354.97 step 1 Mol. Wt: 503.77 step 2
Mol. Wt: 389.51 step 3 Mol. Wt: 499.41
46.4
Figure imgf000131_0003
[00413] Synthesis of compound 46.2. 46.2 was prepared from 45.1 in a manner analogous to the synthesis of 10.1 from 1.8. Isolated 350 mg (69%) of 46.2 as a light yellow oil. MS: (ES, m/z 504 [M+H]+.
[00414] Synthesis of compound 46.3. To a solution of 46.2 (350 mg, 0.69 mmol, 1.00 equiv) in methanol (30 mL) was added hydrochloric acid (3 M in water, 1 mL) at 0 °C and the resulting solution was stirred for 1 h at room temperature. The reaction was quenched with NaHCOs/brine and extracted with EtOAc ((3 x 80 mL). The organic layers were combined and dried over anhydrous sodium sulfate. After filtration and concentration in vacuo, the residue was purified by a silica gel column with DCM/MeOH (30: 1-10:1) to provide 260 mg (96%) of 46.3 as a light yellow semi-solid. MS (ES): m/z 390 [M+H]+.
[00415] Synthesis of compound 46.4. A 20-mL round-bottom flask purged and maintained with an inert atmosphere of nitrogen was charged with a solution of 46.3 (260 mg, 0.67 mmol, 1.00 equiv) in DCM (20 mL). Imidazole (91 mg, 1.34 mmol, 2.00 equiv) and PPh3 (264 mg, 1.01 mmol, 1.50 equiv) were added successively, followed by the addition of I2 (255 mg, 1.00 mmol, 1.50 equiv) at room temperature. The resulting solution was stirred overnight at ambient temperature and diluted with DCM and washed with Na2S03 (sat.) and brine. The organic layer was dried over anhydrous sodium sulfate. After filtration and concentration under reduced pressure, the residue was purified by a silica gel column with EtO Ac/petroleum ether (1 : 1-3: 1) to provide the desired 46.4 (287 mg, 86%) as a light yellow solid. MS (ES): m/z 500 [M+H]+.
[00416] Synthesis of compound 46.5. A mixture of 46.4 (287 mg, 0.57 mmol, 1.00 equiv), 18-crown-6 (190 mg, 0.72 mmol, 1.25 equiv), potassium carbonate (141 mg, 1.02 mmol, 1.78 equiv) and ethyl 2-cyanoacetate (678 mg, 5.99 mmol, 10.43 equiv) in benzene (20 mL) was stirred overnight at 80 °C under nitrogen. After cooling to room temperature, the solvent was removed under vacuum and the residue was purified by a silica gel column with DCM/MeOH (30: 1-10: 1) to provide 230 mg (83%) of 46.5 as a light yellow solid. MS (ES): m/z 485 [M+H]+.
[00417] Synthesis of compound 1-31. NH3 (gas) was introduced into ethanol (60 mL) at 0 °C with stirring for lh. 46.5 (230 mg, 0.47 mmol, 1.00 equiv) was added and the resulting solution was stirred overnight at room temperature. After concentration in vacuo, the residue was purified by preparative HPLC under the following conditions (Waters): column: Xbridge Prep CI 8, 19* 150mm 5um; mobile phase: water with 100 mM NH4HC03 and CH3CN (20.0% CH3CN up to 63.0% in 12 min); flow rate: 20 mL/min; UV detection at 254/220 nm. The product-containing fractions were collected and partially evaporated to remove water and CH3CN under reduced pressure. The residue was lyophilized overnight to give the desired 1-31 (154.3 mg, 71%) as a white solid. MS (ES): m/z 456 [M+H]+. 1H NMR (400 MHz, CD3OD): δ 8.48 (1H, s), 5.36-5.29 (1H, m), 3.73(4H, t), 3.62-3.59 (1H, m), 3.39-3.33 (1H, m), 3.25-3.02 (2H, m), 2.82-2.59 (6H, m), 2.48-2.23 (4H, m), 2.18-2.06 (2H, m), 1.91-1.72 (3H, m), 1.55- 1.43(2H, m). [00418] Example 47: Synthesis of (S)-2-hydroxy-3-((R)-4-(((lr,4R)-4-(piperidin-l- yl)cyclohexyl)oxy)-6,7-dihydro-5H-cyclopenta [4,5] thieno [2,3-d] pyrimidin-5- yl)propanamide (1-32)
Figure imgf000133_0001
47.4 47.5 47.6 1-32
[00419] Synthesis of compound 47.1. Compound 47.1 was prepared from 35.1 and trans-4- (piperidin-l-yl)cyclohexan-l-ol in a manner analogous to the synthesis of compound 36.4. Isolated 0.5 g of a yellow oil.
[00420] Synthesis of compound 47.5. Compound 47.5 was prepared from 47.1 in a manner analogous to the synthesis of 1-25, except that HC1/DCM was used in the first step rather than TBAF/THF. Isolated 0.1 g of a white solid in 14% overall yield.
[00421] Synthesis of Compounds 47.6 and 1-32. The enantiomers of 47.5 (100 mg, 0.22 mmol, 1.00 equiv) were separated by preparative chiral HPLC under the following conditions (Gilson): Column: CHIRALPAK OJ-H, 2*25 cm; mobile phase: hexanes (0.1% DEA) and IPA (0.2% DEA), gradient: (hold at 10% IPA in 23 min); flow rate: 20 mL/min; UV detection at 254/220 nm. Isolated 20 mg of 47.6 (tR = 12.05 min, 100% ee, 2% overall yield) and 22 mg of 1-32 (tR = 19.24 min, 100% ee, 2% overall yield), both as white solids. The ee values of the two isomers were determined by chiral HPLC under the following conditions (SHIMADZU): Column: Chiralpak AD-3, 0.46* 15 cm, 3 μιη (DAICEL); mobile phase: hexanes (0.1% TEA): IPA = 90: 10; UV detection at 254 nm. flow rate: 1.0 mL/min.
[00422] Analytical data for 1-32: MS (ES): m/z 445 (M+H)+. 1H-NMR (300 MHz, CD3OD): δ 8.45 (1H, s), 5.29-5.21 (1H, m), 4.14 (1H, dd), 3.65-3.50 (1H, m), 3.20-3.05 (1H, m), 3.02-2.89 (1H, m), 2.80-2.41 (8H, m), 2.40-2.20 (2H, m), 2.07 (2H, d), 1.80-1.40 (10H, m). [00423] Example 48: (2S)-2-hydroxy-3-[(3R)-12-[(4-[methyl[2-oxo-2-(pyrrolidin-l- yl)ethyl] amino] cyclohexyl)oxy]-7-thia-9,l 1-diazatricyclo [6.4.0.0A [2,6] ] dodeca- l(8),2 6),9,ll-tetraen-3-yl]propanamide (1-33)
Figure imgf000134_0001
1-25 1-33
[00424] In a 50-mL round-bottom flask a solution of 1-25 (110 mg, 0.26 mmol, 1.00 equiv), potassium carbonate (179 mg, 1.30 mmol, 5.03 equiv) and 2-chloro-l-(pyrrolidin-l-yl)ethan-l- one (152 mg, 1.03 mmol, 4.00 equiv) in N,N-dimethylformamide (6 mL) was stirred overnight at room temperature. The resulting solution was diluted with 20 mL of water. The resulting solution was extracted with 4x20 mL of ethyl acetate and the organic layers combined and concentrated under vacuum. The crude product was purified by flash preparative HPLC with the following conditions (IntelFlash-1): column: silica gel; mobile phase; detector, UV 254 nm. This resulted in 1-33 (63.8 mg, 49%) as a white solid. The product was confirmed by LCMS and 1H-NMR. 1H NMR (400 MHz, CD3OD, ppm): δ 8.47 (s, 1H), 5.28 (m, 1H), 4.17-4.14 (m, 1H), 3.59-3.32 (m, 7H), 3.15-2.98 (m, 2H), 2.76-2.70 (m, 2H), 2.52-2.33 (m, 7H), 2.04-1.89 (m, 6H), 1.74-1.53 (m, 5H). MS: m/z = 502 [M+H]+.
[00425] Example 49: Synthesis of Intermediate 49.1.
Figure imgf000134_0002
step 1 step 2
35.1 49.1 49.2
Figure imgf000135_0001
step 3 49 3
[00426] Intermediate 49.3 was prepared from 35.1 in a manner analogous to the synthesis of 36.3. Isolated 150 mg of a white solid in 57% overall yield. MS (ES): m/z 402 [M+H]+.
[00427] Example 50: Synthesis of Intermediate 50.4.
Figure imgf000135_0002
49.3 50.1 50.2
Figure imgf000135_0003
50.3 50.4
[00428] Intermediate 50.4 was prepared from 49.3 in a manner analogous to the synthesis of 1-25, except that HCl/MeOH rather than TBAF/THF was used in the second step. Isolated 124 mg of a white solid in 48% overall yield. MS (ES): m/z 447 [M+H]+. 1H NMR (400 MHz, CDCls): δ 8.46 (s, 1H), 5.28-5.25 (m, 1H), 4.17-4.06 (m, 51H), 3.74-3.72 (m, 5H), 3.37-2.98 (m, 2H), 2.72-2.28 (m, 10H), 2.11-2.08 (m, 2H), 1.79-1.46 (m, 5H).
[00429] Example 51: Synthesis of (S)-2-hydroxy-3-((R)-4-(((lr,4R)-4- morpholinocyclohexyl)oxy)-6,7-dihydro-5H-cyclopenta [4,5] thieno [2,3-d] pyrimidin-5- yl)propanamide (1-34) and Example 52: Synthesis of (R)-2-hydroxy-3-((R)-4-(((lr,4R)-4- morpholinocyclohexyl)oxy)-6,7-dihydro-5H-cyclopenta [4,5] thieno [2,3-d] pyrimidin-5- yl)propanamide (1-44)
Figure imgf000136_0001
[00430] The racemic 50.4 (1.6 g, 96.5% purity) was separated by Chiral-HPLC with the following conditions (Gilson G x 281): column: Chiralpak AD-H, 2*25 cm Chiral-P(AD-H); mobile phase: phase A: hex (O. P/oDEA) (HPLC grade), phase B: IPA (HPLC grade), gradient: 30% B in 9 min; flow rate: 20 mL/min; UV detection at 220/254 nm. The former fractions (tR = 4.75 min) were collected and evaporated under reduced pressure and lyophilized overnight to afford 1-44 (520 mg) with 100% ee as a white solid. And the latter fractions (tR = 5.82 min) were handled as the former fractions to give the desired 1-34 (510 mg) with 99.6%> ee as a white solid. The ee values of the two isomers were determined by the chiral-HPLC with the following conditions (SHIMADZU-SPD-20A): column: Chiralpak AD-H, 0.46*25 cm, 5um (DAICEL); mobile phase: hex (0.1% TEA): IPA = 85:15; UV detection at 254 nm. Flow rate: 1.0 mL/min. tR (1-44) = 7.939 min and tR (1-34) = 11.918 min.
[00431] Analytical data for 1-44: MS: (ES, m/z) 447 [M+H]+. 1H NMR (400 MHz, CD3OD+CDCI3): δ 8.47 (s, 1H), 5.32-5.22 (m, 1H), 4.08 (dd, 1H), 4.89-4.62 (m, 5H), 3.20-3.10 (m, 1H), 3.05-2.95 (m, 1H), 2.75-2.55 (m, 5H), 2.44-2.38 (m, 2H), 2.34-2.28 (m, 3H), 2.10 (d, 2H), 1.82-1.62 (m, 3H), 1.58-1.40 (m, 2H).
[00432] Analytical data for 1-34: MS: (ES, m/z) 447 [M+H]+. 1H NMR (400 MHz, CDC13): δ 8.46 (s, 1H), 5.32-5.22 (m, 1H), 4.15 (t, 1H), 3.73 (t, 4H), 3.59 (td, 1H), 3.19-3.08 (m, 1H), 3.02- 2.92 (m, 1H), 2.78-2.70 (m, 1H), 2.69-2.60 (m, 4H), 2.58-2.20 (m, 5H), 2.10 (d, 2H), 1.75-1.63 (m, 3H), 1.53-1.40 (m, 2H).
[00433] Example 53: Synthesis of 2,2-difluoro-3-[(3R)-12-[[4-(morpholin-4- yl)cyclohexyl]oxy]-7-thia-9,ll-diazatricyclo[6.4.0.0[2,6]]dodeca-l(8),2(6),9,ll-tetraen-3- yl]propanamide (1-35)
Figure imgf000137_0001
Figure imgf000137_0002
53.2 53.3 1-35
[00434] Synthesis of compound 53.1. A solution of 50.1 (400 mg, 0.93 mmol, 1.00 equiv) and hydrogen chloride (12 N) (0.6 mL) in methanol (10 mL). The resulting solution was stirred overnight at room temperature. The reaction was then quenched by the addition of 20 mL of sodium bicarbonate (sat.). The resulting solution was extracted with 3x50 mL of ethyl acetate and the organic layers combined and concentrated under vacuum. The residue was applied onto a silica gel column with dichloromethane/methanol (20: 1). This resulted in 230 mg (53%) of 53.1 as a colorless oil.
[00435] Synthesis of compound 53.2. A solution of 53.1 (230 mg, 0.50 mmol, 1.00 equiv), Dess-Martin (317 mg) in dichloromethane (10 mL). The resulting solution was stirred for 3 h at 25 °C. The reaction was then quenched by the addition of 20 mL of sodium bicarbonate (sat.). The resulting solution was extracted with 3x50 mL of dichloromethane and the organic layers combined and concentrated under vacuum. The residue was applied onto a silica gel column with dichloromethane/methanol (30: 1). This resulted in 180 mg (79%) of 53.2 as a colorless oil.
[00436] Synthesis of compound 53.3. A solution of 53.3 (150 mg, 0.33 mmol, 1.00 equiv), DAST (150 mg, 0.93 mmol, 2.85 equiv) in dichloromethane (10 mL). The resulting solution was stirred overnight at 25 °C. The reaction was then quenched by the addition of 20 mL of sodium bicarbonate(sat). The resulting solution was extracted with 3x50 mL of ethyl acetate and the organic layers combined and concentrated under vacuum. The residue was applied onto a silica gel column with dichloromethane/methanol (40: 1). This resulted in 120 mg (76%) of 53.3 as a colorless oil. [00437] Synthesis of Compound 1-35. Into a 50-mL seal-tube, NH3(g) (30 mL) was introduced to ethanol (20 mL) at 0 °C. This was followed by the addition of 53.3 (120 mg, 0.25 mmol, 1.00 equiv). The resulting solution was stirred overnight at 25 °C. The resulting mixture was concentrated under vacuum. The crude product (100 mg) was purified by preparative HPLC with the following conditions (l#-Pre-HPLC-016(Waters)): column: SunFire Prep C18, 19* 150mm 5um; mobile phase: water with 50mL NH4C03 and CH3CN (5.0% CH3CN up to 45.0%) in 11 min, up to 95.0% in 2 min,down to 5.0%> in 2 min); detector, uv 254/220nm. 31.9 mg product was obtained. This resulted in 31.9 mg (27%>) of 1-35 as a white solid.
MS (ES, m/z 467 [M+H]+. 1H-NMR (300 MHz, CD3OD, ppm): δ 8.474(lH,s) 5.226-5.333 (1H, m), 3.70-3.73(5H, m), 2.728-3.172 (4H, m), 2.62-2.65 (4H, m), 2.07-2.49 (7H, m), 1.63- 1.75(2H, m),1.41-1.53(2H, m).
[00438] Example 54: Synthesis of Intermediate 54.3.
Figure imgf000138_0001
54.3
[00439] Synthesis of compound 54.1. A 50-mL round-bottom flask containing a solution of intermediate 38.2 (1.0 g, 2.23 mmol, 1.00 equiv) and TEA (339 mg, 3.35 mmol, 1.50 equiv) in dichloromethane (25 mL) was added methanesulfonyl chloride (306 mg, 2.67 mmol, 1.20 equiv) at 0 °C under nitrogen. The resulting solution was stirred for 2 h at room temperature and quenched by the addition of water. The reaction mixture was extracted with dichloromethane (2 x 40 mL). The combined organic layers were washed with brine and dried over anhydrous sodium sulfate. After filtration and concentration under reduced pressure, the residue was loaded onto a silica gel column with ethyl acetate/petroleum ether (1 :2) and purified to afford compound 54.1 (1.1 g, 94%) as a yellow oil. MS: m/z 526 (M+H)+.
[00440] Synthesis of compound 54.2. 54.1 (1.1 g, 2.09 mmol, 1.00 equiv) was placed in a 50 mL round-bottom flask and treated with NaCN (308 mg, 6.29 mmol, 3.00 equiv) in DMSO (15 mL) for 4 h at 80 °C under nitrogen. The reaction was then quenched with water and extracted with ethyl acetate (3 x 40 mL). The organic layers were combined, washed with brine and dried over anhydrous sodium sulfate. After filtration and concentration under reduced pressure, the residue was purified by column chromatography on silica gel with ethyl acetate/petroleum ether (1 :3) to give the desired intermediate 54.2 (920 mg, 96%) as a yellow solid. MS: m/z 457 (M+H)+.
[00441] Synthesis of compound 54.3. Intermediate 54.3 was prepared from 54.2 in a manner analogous to the synthesis of 1-27 from 40.2. Isolated 86.4 mg (54%) as a white solid. 1H NMR (300 MHz, CD3OD): δ 8.49 (s, 1H), 5.32 (m, 1H), 3.33 (m, 1H), 3.07 (m, 2H), 2.74 (m, 1H), 2.35-2.54 (m, 12H), 2.11 (m, 2H), 1.74 (m, 1H), 1.44-1.66 (m, 4H). MS: m/z 371 (M+H)+.
[00442] Example 55: Synthesis of 3-[(35)-12-[[4-(dimethylamino)cyclohexyl]oxy]-7- thia-9,ll-diazatricyclo[6.4.0.0[2,6]]dodeca-l(8),2(6),9,ll-tetraen-3-yl]propanamide. -36)
Figure imgf000139_0001
1-36
[00443] Synthesis of compound 55.1. To a solution of intermediate 54.2 (200 mg, 0.44 mmol, 1.00 equiv) and potassium carbonate (166 mg, 1.20 mmol, 2.75 equiv) in DMSO (10 mL) was added H202 (30%>, 3 mL) at 0 °C. The resulting solution was stirred for 18 h at room temperature, quenched with saturated aqueous Na2S03 and extracted with ethyl acetate (3 x 60 mL). The combined organic layers were washed with brine, dried over anhydrous sodium sulfate and concentrated under vacuum. Compound 55.1 (110 mg, 53%) was obtained as a white solid. MS: m/z 475 (M+H)+. [00444] Synthesis of Compound 1-36. 1-36 was prepared from 54.2 in a manner analogous to the synthesis of 1-27 from 40.2. Isolated Compound 1-36 (67.5 mg, 76%) as a white solid. 1H NMR (300 MHz, CDC13) δ 8.46 (s, 1H), 5.78 (d, 2H), 5.22 (m, 1H), 3.44 (m, 1H), 3.02 (m, 2H), 2.67 (m, 1H), 2.33 (m, 13H), 1.90 (m, 3H), 1.44 (m, 4H). MS: m/z 389 (M+H)+.
[00445] Example 56: Synthesis of 3-[(3R)-12-[[4-(dimethylamino)cyclohexyl]oxy]-7-thia- 9,ll-diazatricyclo[6.4.0.0[2,6]]dodeca-l(8),2(6),9,ll-tetraen-3-yl]-2,2-difluoropropanamide -37)
Figure imgf000140_0001
[00446] Synthesis of compound 56.1. Compound 56.1 was prepared from 38.6 in a manner analogous to the synthesis of 53.1 from 50.1. Isolated 375 mg (crude) as a yellow oil.
[00447] Synthesis of compound 56.2. A solution of 56.1 (375 mg, 0.92 mmol, 1.00 equiv), TEA (0.5 mL) and di-tert-butyl dicarbonate (303 mg, 1.39 mmol, 1.50 equiv) in dichloromethane (20 mL) was stirred overnight at room temperature. The resulting solution was diluted with 30 mL of water. The resulting solution was extracted with 3x30 mL of ethyl acetate and the organic layers combined and concentrated under vacuum. The residue was purified onto a silica gel column with ethyl acetate/petroleum ether (1 :3). This resulted in 56.2 (275 mg, 59%) as a colorless oil.
[00448] Synthesis of compound 56.3. A solution of 56.2 (275 mg, 0.54 mmol, 1.00 equiv) and Dess-Martin oxidant (230 mg, 0.54 mmol, 1.00 equiv) in dichloromethane (15 mL) was stirred for 2 h at room temperature. The reaction was then quenched by the addition of 20 mL of saturated sodium bicarbonate, extracted with 3x30 mL of ethyl and concentrated under vacuum. The residue was applied onto a silica gel column with ethyl acetate/petroleum ether (1 :4). This resulted in 56.3 (205 mg, 75%) as a colorless oil.
[00449] Synthesis of compound 56.4. 56.4 was prepared from 56.5 in a manner analogous to the synthesis of 53.3 from 53.2. Isolated 180 mg (quant.) as a colorless oil.
[00450] Synthesis of compound 56.6. 56.6 was prepared from 56.4 in a manner analogous to the synthesis of 1-27 from 40.2. Isolated 105 mg (85%) as a yellow oil.
[00451] Synthesis of compound 56.7. A solution of 56.6 (130 mg, 0.31 mmol, 1.00 equiv) was dissolved in methanol (15 mL). Sulfuroyl dichloride (72 mg, 0.61 mmol, 1.98 equiv) was added dropwise. The resulting solution was stirred overnight at 30 °C. The reaction was then quenched by the addition of 20 mL of saturated sodium bicarbonate, extracted with chloroform, dried over anhydrous sodium sulfate and concentrated under vacuum. This resulted in 56.7 (103 mg, crude) as a yellow oil.
[00452] Synthesis of Compound 1-37. 1-37 was prepared from 56.7 in a manner analogous to the synthesis of 1-35 from 53.3. Isolated 41.3 mg (42%) as a white solid. 1H NMR (300 MHz, CD3OD): δ 8.51 (s, 1H), 5.30-5.23 (m, 1H), 3.66-3.51 (m, 1H), 3.32-2.70 (m, 4H), 2.51-2.01 (m, 13H), 1.74-1.44 (m, 4H). MS: m/z 425 (M+H)+.
[00453] Example 57. Synthesis of (S)-2-hydroxy-3-((R)-4-(((lr,4R)-4- morpholinocyclohexyl)amino)-6,7-dihydro-5H-cyclopenta [4,5] thieno [2,3-d] pyrimidin-5- yl)propanamide (1-38)
Figure imgf000141_0001
Chiral separation
Figure imgf000142_0001
1-38
[00454] Synthesis of compound 57.1. Compound 57.1 was prepared from 34.3 in a manner analogous to the synthesis of 50.4 from 49.2. Isolated 236 mg (99%) of 57.1 as a white solid. MS: m/z 395 (M+H)+.
[00455] Synthesis of compound 57.2. To a solution of 57.1 (150 mg, 0.38 mmol, 1.00 equiv) in NMP (1 mL) was added triethylamine (310 mg, 3.06 mmol, 8.00 equiv) and 4-(morpholin-4- yl)cyclohexan-l -amine (560 mg, 3.04 mmol, 8.00 equiv) at room temperature. The resulting solution was stirred overnight at 75 °C under nitrogen. After cooling to room temperature, the resulting solution was diluted with DCM (30 mL), washed with brine, dried over anhydrous sodium sulfate and concentrated under vacuum. The residue was applied onto a silica gel column with dichloromethane/methanol (20: 1) to give 185 mg (90%) of 57.2 as a white solid. MS : m/z 542 (M+H)+.
[00456] Synthesis of Compound 1-38. Compound 1-38 was prepared from 57.2 in a manner analogous to the synthesis of 1-34 from 50.2. Isolated 1 13 mg (31 % overall yield from 57.2) with 100% ee as a white solid. MS: m/z 446 (M+H)+. 1H NMR (400 MHz, CD3OD): δ 8.23 (s, 1H), 4.21-4.1 1 (M, 1H), 4.10 (t, 1H), 3.80-3.60 (m, 5H), 3.20-3.05 (m, 1H), 3.01-2.85 (m, 1H), 2.74- 2.65 (m, 5H), 2.45-2.23 (m, 2H), 2.25-1.95 (m, 6H), 1.78-1.43 (m, 4H).
[00457] Example 58: Synthesis of (R)-3-((R)-4-(((lr,4R)-4-
(dimethylamino)cyclohexyl)oxy)-6,7-dihydro-5H-cyclopenta [4,5] thieno [2,3-d] pyrimidin-5- yl)-l,l,l-trifluoropropan-2-ol (1-39)
Figure imgf000142_0002
57.1 58.1
Figure imgf000143_0001
[00458] Synthesis of compound 58.1. To a solution of 57.1 (267 mg, 1.06 mmol, 1.00 equiv) in 10 mL of distilled THF was added trimethyl(trifluoromethyl)silane (280 mg, 1.97 mmol, 1.86 equiv) and TEA (0.1 mL). The reaction was stirred at 0 °C for 30 min under nitrogen. Then TBAF (0.01 mL) was added via syringe at 0 °C and the resulting solution was stirred for 5 min. The resulting solution was diluted with 15 mL of water and extracted with ethyl acetate (100 mL). The organic layer was washed with brine, dried over sodium sulfate and concentrated under vacuum. The residue was applied onto a silica gel column with ethyl acetate/petroleum ether (1 : 1) to afford 58.1 (320 mg, 77%) as a red oil.
[00459] Synthesis of compound 58.2. Compound 58.2 was prepared from 58.1 in a manner analogous to the synthesis of 10.1 from 1.8. The residue was applied onto a silica gel column with dichloromethane/methanol/NH40H (10: 1 :0.1) and purified to give the product as a mixture of diastereomers (180 mg).
[00460] Synthesis of Compound 1-39. The diastereomers of 58.2 (180 mg) were separated by preparative HPLC (SHIMADZU) under the following conditions: column: Xbridge Prep CI 8 5um, 19* 150mm; mobile phase: water (0.1% HCOOH) and MeOH (6.0% MeOH up to 53.0% in 19 min); UV detection at 254/220 nm. The product-containing fractions were collected and evaporated (to remove the water and CH3OH) to give 1-39 (74.1 mg) as a white solid. 1H NMR (300 MHz, CD3OD) δ 8.44 (s, 2H), 5.33-5.31 (m, 1H), 4.10-4.05 (m, 1H), 3.47-3.32 (m, 1H), 3.29-2.90 (m, 3H), 2.84-2.68 (m, 7H), 2.55-2.17 (m, 7H), 1.77-1.62 (m, 5H). MS: m/z 430 (M+H)+.
[00461] Example 59: Synthesis of 2-(((lR,4r)-4-(((R)-5-(2-hydroxyethyl)-6,7-dihydro-5H- cyclopenta[4,5]thieno[2,3-d]pyrimidin-4-yl)oxy)cyclohexyl)(methyl)amino)-l-(pyrrolidin-l- yl)ethanone. (1-48)
Figure imgf000144_0001
[00462] Synthesis of compound 59.2. 59.2 was prepared from 38.3 in a manner analogous to the synthesis of 1-7 from 4.5. Isolated 118 mg (77%) as a yellow solid. MS (ES): m/z 348 [M+H]+.
[00463] Synthesis of Compound 1-48. 1-48 was prepared from 59.2 in a manner analogous to the synthesis of 1-33 from 1-25. Isolated 130 mg (82%) as a white solid. MS (ES): m/z 459 [M+H]+. 1H NMR (400 MHz, CD3OD): δ 8.47 (IH, s), 5.32-5.20 (IH, m), 3.66 (2H, t), 3.58 (2H, t), 3.50-3.41 (3H, m), 3.37 (2H, s), 3.15-3.05 (IH, m), 3.00-2.92 (IH, m), 2.75-2.60 (2H, m), 2.37 (3H, s), 2.33-2.15 (4H, m), 2.03-1.88 (6H, m), 1.71-1.54 (4H, m).
[00464] Example 60: Synthesis of 3-((S)-4-(((lr,4S)-4-(methyl(2-oxo-2-(pyrrolidin-l- yl)ethyl)amino)cyclohexyl)oxy)-6,7-dihydr o-5H-cyclopenta [4,5] thieno [2,3-d] pyrimidin-5- yl)propanamide (1-40)
Figure imgf000144_0002
steP 1 step 2
I-48 60.1 60.2
Figure imgf000145_0001
Prep. HPLC
1-40
[00465] Synthesis of compound 60.1. A 25-mL round-bottom flask, purged and maintained with an inert atmosphere of nitrogen, was charged with a solution of 1-48 (100 mg, 0.22 mmol, 1.00 equiv) in 5 mL of distilled DMF. MsCl (38 mg, 0.33 mmol, 1.50 equiv) and triethylamine (66.7 mg, 0.66 mmol, 3.00 equiv) were added at 0 °C. The resulting solution was stirred overnight at room temperature. The reaction was then quenched with water, extracted with DCM. The organic phase was washed with brine, dried over anhydrous sodium sulfate and concentrated under vacuum. The residue was purified by column chromatography on silica gel with DCM/MeOH (20: 1 to 10: 1) to give 60.1 (107 mg) as a white solid. MS (ES): m/z 537 [M+H]+.
[00466] Synthesis of compound 60.2. To a solution of 60.1 (107 mg, 0.20 mmol, 1.00 equiv) in DMSO (5 mL) were added NaCN (58.8 mg, 1.20 mmol, 6.00 equiv) and 4- dimethylaminopyridine (2.4 mg, 0.02 mmol, 0.10 equiv) at room temperature. The resulting solution was stirred for 2 h at 80 °C. After cooling to room temperature, the reaction was then quenched by the addition of aqueous FeS04 solution and extracted with DCM. The organic phase was washed with brine, dried over anhydrous sodium sulfate and concentrated under vacuum. The residue was applied onto a silica gel column with DCM/MeOH (10: 1) to give the desired 60.2 (80 mg, 86%) as a yellow solid. MS (ES): m/z 468 [M+H]+.
[00467] Synthesis of Compound 1-40. Compound 1-40 was prepared from 60.2 in a manner analogous to the preparation of 38.8 and 38.9 from 38.3. Isolated 82.5 mg (quant.) as a white solid. MS (ES): m/z 486 [M+H]+; 1H NMR (400 MHz, CD3OD): δ 8.47 (1H, s), 5.32-5.22 (1H, m), 3.58 (2H, t), 3.45 (2H, t), 3.35 (2H, s), 3.20-3.09 (1H, m), 3.03-2.95 (1H, m), 2.73-2.65 (2H, m), 2.37 (3H, s), 2.33-2.19 (5H, m), 2.03-1.88 (6H, m), 1.66 (2H, m), 1.69-1.52 (4H, m). [00468] Example 61: Synthesis of (2R)-l-[(3R)-12-[[4-
(dimethylamino)cyclohexyl] oxy] -7-thia-9, 11-diazatricyclo [6.4.0.0 [2,6] ] dodeca- -tetraen-3-yl]propan-2-ol (I-42)
Figure imgf000146_0001
61.3 I-42
[00469] Synthesis of compound 61.1. To a solution of 38.4 (450 mg, 1.01 mmol, 1.00 equiv) in distilled tetrahydrofuran (20 mL) was added dropwise bromo(methyl)magnesium (0.5 mL, 1.50 equiv) at 0°C under nitrogen. The resulting solution was stirred for 2 h at room temperature and quenched with water and extracted with ethyl acetate. After drying over Na2S04 and evaporating under reduced pressure, the residue was applied onto a silica gel column with ethyl acetate / petroleum ether (1 : 10-1 :5) to give 61.2 (0.11 g) as a light yellow oil and 61.1 (0.16 g) as a light yellow oil, respectively.
[00470] Synthesis of Compound 1-42. 1-42 was prepared from 61.1 in a manner analogous to the synthesis of 1-27 from 40.2. The product-containing fractions were collected and evaporated to remove the solvents to afford Compound 1-42 (46.3 mg, 58%) as a white solid. 1H NMR (300 MHz, CD3OD) : δ 8.48 (s, 1H), 5.20-5.30 (m, 1H), 3.85- 3.95 (m, 1H), 3.50-3.60 (m, 1H), 3.05-3.15 (m, 1H), 2.90-3.00 (m, 1H), 2.60-2.70 (m, 1H), 2.25-2.40 (m, 10H), 2.15-2.25 (m, 1H), 2.00-2.14 (m, 2H), 1.63-1.68 (m, 2H), 1.40-1.52 (m, 3H), 1.21 (d, 3H). MS: m/z 376 (M+H)+.
[00471] Example 62: Synthesis of 2-[(12R)-3-[[4-(dimethylamino)cyclohexyl]oxy]-8-thia- -diazatricyclo[7.4.0.0[2,7]]trideca-l(9),2(7),3,5-tetraen-12-yl]ethan-l-ol formate. (1-43)
Figure imgf000147_0001
[00472] Synthesis of compound 62.2. Into a 1 L round-bottom flask was placed a solution of cyclohex-2-en-l-one (15.00 g, 156.04 mmol, 1.00 equiv) in anhydrous THF (300 mL). 1,3- diethyl 2-bromopropanedioate (56.00 g, 234.25 mmol, 1.50 equiv), In powder (18.00 g, 1.00 equiv) and TMSC1 (87.00 g, 800.81 mmol, 5.00 equiv) were added under nitrogen. The resulting solution was stirred for 30 min at room temperature and quenched by the addition of 200 mL of saturated aqueous sodium carbonate, extracted with 3 x 300 mL of ethyl acetate. The combined organic layers were washed with brine, dried over anhydrous sodium sulfate and concentrated under vacuum. The residue was applied onto a silica gel column with ethyl acetate/petroleum ether (1 :5) to give 1,3-diethyl 2-(3-oxocyclohexyl)propanedioate (28.50 g, 71%) as a yellow oil.
[00473] Synthesis of compound 62.3. A solution of 1,3-diethyl 2-(3- oxocyclohexyl)propanedioate (28.50 g, 111.20 mmol, 1.00 equiv) and sodium chloride (7.02 g, 1.10 equiv) in a mixture of water (4 mL) and DMSO (80 mL) was heated for 24 h at 180 °C in an oil bath. After cooling to r.t, the reaction was diluted with water and extracted with 3 x 300 mL of ethyl acetate. The organic layers were combined, washed with water and brine, dried over anhydrous sodium sulfate and concentrated under vacuum to give the desired ethyl 2-(3- oxocyclohexyl)acetate (22.7 g, crude) as a yellow oil.
[00474] Purification of compound 62.8. Compound 62.8 was prepared from 62.3 in a manner analogous to the synthesis of 34.4 from 1.1. Isolated 790 mg (10% overall yield) as an off-white solid with 98.5% ee.
[00475] Synthesis of compound 62.9. To a solution of 62.8 (300 mg, 1.12 mmol, 1.00 equiv) in 5 mL of distilled DMF was added TBSC1 (252 mg, 1.50 equiv) and imidazole (137 mg, 2.01 mmol, 1.80 equiv) at room temperature under nitrogen. The resulting solution was stirred for 1 h at ambient temperature and then quenched water, extracted with 3 x 50 mL of ethyl acetate. The combined organic layers were washed with brine, dried over anhydrous sodium sulfate and concentrated under reduced pressure. The residue was applied onto a silica gel column with ethyl acetate/petroleum ether (1 :5) to provide 62.9 (420 mg, 98%) as a light yellow solid. MS (ES): m/z 384, 386 [M+H]+.
[00476] Synthesis of compound 62.10. Compound 62.10 was prepared from 62.9 and dimethylaminocyclohexanol in a manner analogous to the synthesis of 10.1, except that the reaction temperature was kept at room temperature rather than 60 °C. Isolated 105 g (41%) as a light yellow solid. MS (ES): m/z 490 [M+H]+.
[00477] Synthesis of Compound 1-43. To a solution of 62.10 (110 mg, 0.22 mmol, 1.00 equiv) in methanol (5 mL) was added hydrochloric acid (12 M, 0.5 mL) at 0 °C. The resulting solution was stirred for 1 h at room temperature. The solvent was removed in vacuo and the crude product (80 mg) was purified by preparative HPLC with the following conditions (SHIMADZU): column: SunFire Prep CI 8, 19* 150mm 5um; mobile phase: water with 0.1% HCOOH and CH3CN (6.0% CH3CN up to 53.0% in 16 min); flow rate: 20 mL/min; UV detection at 254/220 nm. The product-containing fractions were collected and partially evaporated to remove water and CH3CN under reduced pressure. The residue was lyophilized overnight to give 1-43 (46 mg, 57%) as a white semi-solid. MS (ES): m/z 376 [M-HCOOH+H] +. 1H-NMR: (300 MHz, CDC13): δ 8.49 (d, 2H,) 5.22 (s, 1H), 3.83 (t, 2H), 3.15 (dd, 2H), 2.88 (s, 2H), 2.65 (s, 6H), 2.48 (t, 3H), 2.07 (t, 4H), 1.70 (m, 7H).
[00478] Example 63: Synthesis of (S)-l-((R)-4-(((lr,4R)-4-morpholinocyclohexyl)oxy)- 6,7-dihydro-5H-cyclopenta [4,5] thieno [2,3-d] pyrimidin-5-yl)propan-2-ol (1-45)
Figure imgf000149_0001
step 4
49.3 1-45
[00479] A solution of 49.3 (150 mg, 0.37 mmol, 1.00 equiv) in 20 mL of distilled THF was cooled down to 0 °C under nitrogen. A solution of bromo(methyl)magnesium (1 M in hexane, 0.74 mL, 2.00 equiv) was added slowly via syringe and stirred for 3 h in a water/ice bath. After completion, the reaction was quenched with NH4CI (sat.), extracted with 3 x 80 mL of ethyl acetate. The combined organic layers were concentrated under vacuum, the residue (200 mg) was purified by preparative HPLC under the following conditions (SHIMADZU): column: SunFire Prep CI 8, 19* 150mm 5um; mobile phase: water with 100 mM NH4HC03 and CH3CN (6.0% CH3CN up to 60% in 20 min); flow rate: 20 mL/min; UV detection at 254/220 nm. The product-containing fractions were collected and partially evaporated to remove water and CH3CN under reduced pressure. The residue was lyophilized overnight to give (R)-l-((R)-4- (((lr,4R)-4-morpholinocyclohexyl)oxy)-6,7-dihydro-5H-cyclopenta[4,5]thieno[2,3-d]pyrimidin- 5-yl)propan-2-ol (32 mg) as a white solid and 1-45 (55.5 mg) as a white solid. MS (ES): m/z 418 [M-1.56HCOOH+H]+. 1H NMR (300 MHz, CD3OD): δ 8.42 (s, 2H), 5.29-5.26 (m, 1H), 3.90- 3.83 (m, 5H), 3.56-3.51 (m, 1H), 3.32-3.27 (m, 1H), 3.04-2.88 (m, 7H), 2.68-2.61 (m, 1H), 2.40- 2.90 (m, 6H), 1.75-1.15 (m, 8H). [00480] Example 64: Synthesis of 2-[(4-[[(3R)-3-[(2R)-2-hydroxypropyl]- diazatricyclo [6.4.0.0 [2,6] ] dodeca-l(8),2(6),9,l l-tetraen-12- yl]oxy]cyclohexyl)(methyl)amino]-l-(pyrrolidin-l-yl)ethan-l-one. (1-46)
Figure imgf000150_0001
[00481] 1-46 was prepared from 61.1 in a manner analogous to the synthesis of 1-33 from 38.8. Isolated 52.7 mg (29%, two steps) as a white solid. MS (ES): m/z 473 (M+H)+. 1H NMR (400MHz, CD3OD): δ 8.46 (1H, s), 5.32-5.18 (1H, m), 3.94-3.92 (1H, m), 3.57 (2H, t), 3.45 (2H, t), 3.36 (3H, m), 3.08-3.10 (1H, m), 2.96-2.98 (1H, m), 2.67-2.62 (2H, m), 2.41-2.29 (6H, m), 2.070-1.97 (5H, m), 1.95-1.87 (2H, m), 1.64-1.52 (5H, m), 1.30 (3H, d).
[00482] Example 65: Synthesis of 3-[[4-(dimethylamino)cyclohexyl]oxy]-8-thia-4,6, 12- triazatricyclo[7.4.0.0[2,7]]trideca-l(9),2(7),3,5-tetraene-12-sulfonamide. (1-47)
Figure imgf000150_0002
step 3
65.3 65.4 Formamidine acetate
Formamide, 100°C
Figure imgf000151_0001
step 4 step 5
65.5
Figure imgf000151_0002
step 6 step 7
65.6 65.7
Figure imgf000151_0003
step 8 step 9
65.8 65.9
Figure imgf000151_0004
step 10 step 1 1
6 .10 65.11
Figure imgf000151_0005
step 1
65.12 I-47
[00483] Synthesis of compound 65.2. A solution of 4-aminobutan-l-ol (18 g, 201.94 mmol, 1.00 equiv), 65.1 (44.5 g, 203.02 mmol, 1.01 equiv) and TEA (28 g, 276.71 mmol, 1.37 equiv) in 500 mL of THF was heated to reflux overnight. The reaction mixture was cooled to room temperature, quenched with water and extracted with ethyl acetate. The combined organic layers were washed with brine and dried over sodium sulfate. The solids were filtered out and the filtrate was concentrated under vacuum. The residue was applied onto a silica gel column with PE/EtOAc (10: 1 to 1 : 1) to give 30 g (68%) of 65.2 as a white solid. MS (ES): m/z 220 (M+H)+.
[00484] Synthesis of compound 65.3. To a 250-mL round-bottom flask, purged and maintained with an inert atmosphere of nitrogen, was added a solution of oxalic dichloride (2.52 g, 19.85 mmol, 1.98 equiv) in 40 mL of anhydrous dichloromethane. This solution was cooled to -78 °C under nitrogen. A solution of DMSO (1.56 g, 19.97 mmol, 1.99 equiv) in dichloromethane (10 mL) was added dropwise with stirring at -78 °C and the resulting solution was stirred for 20 min at -78 °C. A solution of 65.2 (2.2 g, 10.03 mmol, 1.00 equiv) in 20 mL of dichloromethane was added dropwise with stirring. Stirring was continued for 20 min at -78 °C, and then TEA (8.1 g, 80.20 mmol, 7.99 equiv) was added via syringe at the same temperature. The resulting solution was allowed to react, with stirring, for an additional 60 min at 0 °C. After completion, the resulting mixture was diluted with 1 M aqueous hydrochloric acid, extracted with DCM, washed with 3 x 100 mL of brine, dried over sodium sulfate and concentrated under vacuum. The residue was purified by a silica gel column with ethyl acetate/petroleum ether (1 : 1) to provide 65.3 (1.9 g, 87%) as a white solid. MS (ES): m/z 218 (M+H)+.
[00485] Synthesis of compound 65.4. A mixture of 65.3 (2.2 g, 10.13 mmol, 1.00 equiv), TEA (2 g, 19.76 mmol, 1.95 equiv), S (340 mg, 10.61 mmol, 1.05 equiv) and ethyl 2- cyanoacetate (1.2 g, 10.61 mmol, 1.05 equiv) in 50 mL of ethanol was stirred for 20 hours at 65 °C under nitrogen. The reaction mixture was cooled to room temperature and concentrated in vacuo. The residue was diluted with EtOAc, washed with brine, dried over anhydrous sodium sulfate and concentrated under vacuum. The residue was applied onto a silica gel column with ethyl acetate/petroleum ether (1 :1) to afford 2.4 g (69%) of 65.4 as an off-white solid. MS (ES): m/z 345 (M+H)+.
[00486] Synthesis of compound 65.5. To a 250-mL round-bottom flask, purged and maintained with an inert atmosphere of nitrogen, was added a solution of 65.4 (3.4 g, 9.87 mmol, 1.00 equiv) and formamidine acetate (1.5 g, 14.41 mmol, 1.46 equiv) dissolved in 50 mL of N,N- dimethylformamide. This solution was heated at 100 °C for 12 hrs. The reaction was cooled to room temperature and quenched with water. The resulting mixture was extracted with 3 x 100 mL of ethyl acetate and the combined organic layers were washed with brine, dried over sodium sulfate and concentrated under vacuum. The residue was applied onto a silica gel column with ethyl acetate/petroleum ether (1 : 1) to yield 1.76 g (55%) of 65.5 as an off-white solid. MS (ES): m/z 326 (M+H)+.
[00487] Synthesis of compound 65.6. A solution of 65.5 (3.2 g, 9.84 mmol, 1.00 equiv) in 50 mL of ethanol was added NH2NH2 (80% in water, 10 mL) at room temperature and heated to reflux for 2 hours. The reaction mixture was cooled to room temperature and the solids were filtered out. The filtrate was concentrated under vacuum and the crude product (3.0 g) was purified by flash preparative HPLC with the following conditions (CombiFlash-1): column: CI 8 silica gel; mobile phase: CH3CN and water (0%> H20 increasing to 30%> within 35 min); flow rate: 20 mL/min; UV detection at 254 nm. After concentration in vacuo, this resulted in 65.6 (1.76 g, 58%>) as a yellow solid.
[00488] Synthesis of compound 65.7. To a solution of 65.6 (2.0 g, 10.24 mmol, 1.00 equiv) and TEA (5 g, 49.41 mmol, 4.82 equiv) in 50 mL of THF was added Boc20 (7.0 g, 32.07 mmol, 3.13 equiv), in portions, at 0 °C under nitrogen. The resulting solution was warmed up to room temperature and stirred for 12 h at 40 °C. TLC analysis indicated that the amine was consumed completely and the resulting mixture was concentrated under vacuum. The residue was diluted with 50 mL of ethyl acetate, washed with brine, dried over Na2S04 and concentrated under reduced pressure. The residue was applied onto a silica gel column with ethyl acetate/petroleum ether (1 :4) to give 3.6 g (89%) of 65.7 as a white solid. MS (ES): m/z 396 (M+H)+.
[00489] Synthesis of compound 65.8. To a 25-mL round-bottom flask containing a solution of 65.7 (370 mg, 0.94 mmol, 1.00 equiv) in 10 mL of methanol was added NH4OH (2.1 mL) at room temperature and the resulting solution was stirred for 2 hours under N2. After completion, the resulting mixture was concentrated under vacuum and the residue was applied onto a silica gel column with ethyl acetate/petroleum ether (1 :1) to afford 252 mg (91%) of 65.8 as a white solid. MS (ES): m/z 296 (M+H)+.
[00490] Synthesis of compound 65.9. To a solution of 65.8 (2.3 g, 7.79 mmol, 1.00 equiv) in DCE (300 mL) was added PPh3 (4.1 g, 15.63 mmol, 2.00 equiv). The solution was stirred for 1 h at room temperature under nitrogen. Then CC14 (3.6 g, 23.38 mmol, 3.00 equiv) was added and the resulting solution was stirred at 72 °C overnight. After cooling, the resulting mixture was concentrated under vacuum. The residue was applied onto a silica gel column with ethyl acetate/petroleum ether (1 :5 to 1 :3) to give 65.9 (1.0 g) as a light yellow solid. MS (ES): m/z 314 and 316 (M+H)+. [00491] Synthesis of compound 65.10. Sodium hydride (80 mg, 2.00 mmol, 4.83 equiv, 60% dispersion in mineral oil) was treated with trans-4-(dimethylamino)cyclohexan-l-ol (80 mg, 0.56 mmol, 1.35 equiv) in 10 mL of distilled THF for 30 min at 0 °C under nitrogen. Then a solution of 65.9 (130 mg, 0.41 mmol, 1.00 equiv) in 3 mL of THF was added and stirred at room temperature for 12 hours. After cooling to 0 °C, the reaction was quenched with water and extracted with 3 x 30 mL of ethyl acetate. The organic layers were combined, washed with brine, dried over anhydrous sodium sulfate and concentrated under vacuum. The residue was applied onto a silica gel column with dichloromethane/methanol (4: 1) to give 140 mg (80%) of 65.10 as a white solid. MS (ES): m/z 421 (M+H)+.
[00492] Synthesis of compound 65.11. To a solution of 65.10 (140 mg, 0.33 mmol, 1.00 equiv) in 5 mL of DCM was added CF3COOH (0.5 mL) at room temperature. The resulting solution was stirred for 5 h at ambient temperature and then concentrated under vacuum to give 65.11 (100 mg) as a yellow oil which was used directly without further purification. MS (ES): m/z 321 (M+H)+.
[00493] Synthesis of compound 65.12. To a 25 -mL round-bottom flask (1 atm), purged and maintained with an inert atmosphere of nitrogen, was placed 65.11 (100 mg, 0.31 mmol, 1.00 equiv) in 25 mL of 1,2-dichloroethane. Then paraformaldehyde (80 mg, 2.67 mmol, 8.55 equiv) and CF3COOH (0.5 mL) were added simultaneously at 0 °C and the resulting mixture was heated to 45 °C and stirred for 12 h. The reaction mixture was cooled to room temperature with a water bath and concentrated under reduced pressure. The crude product was purified by preparative HPLC under the following conditions (Waters): Column: Xbridge Prep C18, 5 μιη, 19*50mm; mobile phase: water with 0.01% TFA and CH3CN (10% CH3CN up to 35% in 11 min, up to 95% in 1.5 min, down to 10% in 1.5 min); flow rate: 20 mL/min; UV detection at 254/220 nm. This resulted in 40 mg (39%) of 65.12 as a light yellow solid. MS (ES): m/z 333 (M+H)+.
[00494] Synthesis of 1-47. Into a 50-mL round-bottom flask was placed a solution of 65.12 (HC1 salt) (50 mg, -75 purity) in anhydrous DCM (5 mL) cooled to 0 °C. Then sulfamoyl chloride (26 mg, 0.23 mmol, 1.50 equiv) was added, followed by addition of TEA (45 mg, 0.44 mmol, 2.96 equiv) and the resulting solution was stirred for 2 h at this temperature under nitrogen. The resulting mixture was concentrated under vacuum. The crude product (60 mg) was purified by preparative HPLC under the following conditions (Waters): Column: XBridge Shield RP18 OBD 5 μιη, 19* 150 mm; mobile phase: water with 0.01% NH4HC03 and acetonitrile (gradient: 20%-24% CH3CN in 10 min); flow rate: 15 ml/min; UV detection at 254 nm. This resulted in 10.7 mg (17%) of 1-47 as a white solid. MS (ES): m/z 412 (M+H)+. 1H-NMR (300MHz, CD3OD): δ 8.47 (1H, s), 5.26-5.15 (1H, m), 4.51 (2H, s), 3.52 (2H, t), 3.03 (2H, t), 2.43-2.25 (9H, m), 2.03 (2H, d), 1.67-1.45 (4H, m).
[00495] Example 66: Synthesis of (12S)-3-[[4-(morpholin-4-yl)cyclohexyl]oxy]-8-thia-4,6- diazatricyclo[7.4.0.0[2,7]]trideca-l(9),2(7),3,5-tetraene-12-carboxamide. (1-54)
Figure imgf000155_0001
Figure imgf000155_0002
66.3 1-54
[00496] Synthesis of compound 66.1. Compound 66.1 was prepared from ethyl 3- oxocyclohexane-l-carboxylate in a manner analogous to the synthesis of 34.3 from 1.1. Isolated 404 mg of 66.1 in 98.7% ee (as determined by analytical chiral HPLC with the following conditions, column: Lux Cellulose-4, 0.46* 15cm,5um, 4.6*250mm, 5um; mobile phase: hex:IPA=80:20; flow rate: 1 mL/min; UV detection at 254 um) and in 17% overall yield.
[00497] Synthesis of compound 66.2. To a solution of 66.1 (253 mg, 0.99 mmol, 1.00 equiv) in acetone (10 mL) was added dropwise Jones reagent (2 mL) at 0 °C. The resulting mixture was stirred for 20 min at this temperature, quenched with aqueous saturated NaHS03, extracted with EtOAc (2 x 100 mL). The combined organic layers were washed with brine, dried over anhydrous sodium sulfate and concentrated under vacuum to yield 247 mg (93%>) of 66.2 as a white solid.
[00498] Synthesis of compound 66.3. 66.3 was prepared from 66.2 in a manner analogous to the synthesis of 2.1 from 1.9. Isolated 200 mg (quant., crude) as a white solid.
[00499] Synthesis of Compound 1-54. 1-54 was prepared from 66.3 in a manner analogous to the synthesis of 1-1 from 2.3. Isolated 79.6 mg (38%) of 1-54 as a white solid. MS (ES): m/z 417 [M+H . 1H-NMR (300 MHz, CD3OD) δ 8.45 (1H, s), 5.30-5.22 (1H, m), 3.74-3.71 (4H, t), 3.33-3.26 (1H, m), 3.09-2.97 (3H, m), 2.87-2.70 (1H, m), 2.65-2.62 (4H, t), 2.45-2.30 (3H, m), 2.23-2.18 (1H, m), 2.11-2.07 (2H, m), 2.07-1.91 (1H, m), 1.64-1.44 (4H, m).
[00500] Example 67: Synthesis of intermediate 67.6.
Figure imgf000156_0001
67.1 67.2 67.3
Figure imgf000156_0002
[00501] Synthesis of compound 67.2. To a solution of 4,4-difluoropiperidine hydrochloride (1.58 g, 10.0 mmol, 1.00 equiv) in dichloromethane (75 mL) was added 1,4- dioxaspiro[4.5]decan-8-one (1.56 g, 10.0 mmol, 1.0 equiv), acetic acid (0.5 mL, 2.00 equiv) and NaBH(OAc)3 (4.24 g, 20 mmol, 2.0 equiv) at room temperature under nitrogen. The resulting solution was stirred for 24 h at ambient temperature. The pH value of the solution was adjusted to 12 with 1 M aqueous NaOH solution and extracted with 3 x 50 mL of dichloromethane. The combined organic layers were washed with brine, dried over sodium sulfate and concentrated in vacuo. The residue was applied onto a silica gel column with DCM/MeOH (80: 1 to 50:1) to give the desired 67.2 (2.0 g) as a white solid.
[00502] Synthesis of compound 67.3. A mixture of 67.2 (200 mg, 0.77 mmol, 1.00 equiv) and hydrochloric acid (4 M, 6 mL) in 30 mL of THF was stirred for 24 h at 60 °C. The resulting mixture was concentrated under vacuum. The pH value of the solution was adjusted to 8 with saturated aqueous sodium bicarbonate and extracted with 5 x 50 mL of dichloromethane. The organic layers were combined, dried over sodium sulfate and concentrated under vacuum to yield 150 mg (90%) of 67.3 as a light yellow oil.
[00503] Synthesis of compound 67.4. To a solution of 67.3 (100 mg, 0.46 mmol, 1.00 equiv) in methanol (10 mL) was added NaBH4 (53 mg, 1.40 mmol, 3.00 equiv) and the resulting solution was stirred for 3 h at room temperature. The reaction was quenched with saturated aqueous NH4C1 and extracted with 4 x 50 mL of dichloromethane. The combined organic layers were washed with brine, dried over anhydrous sodium sulfate and concentrated under vacuum to give 80 mg (79%) of 67.4 as a white solid.
[00504] Synthesis of compound 67.5. Into a 100-mL round-bottom flask containing a solution of 67.4 (100 mg, 0.46 mmol, 1.00 equiv) in dry dichloromethane (20 mL) was added 4- nitrobenzoyl chloride (170 mg, 0.92 mmol, 2.00 equiv), followed by addition of sodium bicarbonate (115 mg, 1.37 mmol, 3.00 equiv) and 4-dimethylaminopyridine (5 mg, 0.04 mmol, 0.10 equiv) at room temperature. The resulting mixture was stirred 4h at ambient temperature. After completion, the resulting mixture was diluted with water and extracted with DCM, dried and concentrated under reduced pressure. The residue was purified by chromatography on silica gel with DCM/MeOH (50: 1 to 30: 1) to afford the corresponding 67.5 (120 mg) as a light yellow solid.
[00505] Synthesis of compound 67.6. To the pure compound 67.5 (368 mg, 1.0 mmol, 1.0 equiv) in a mixture of MeOH/THF/water (4mL/4mL/2mL) was added LiOH H20 (126 mg, 3.0 mmol, 3.0 equiv) at room temperature and followed by stirring for 1 h. After evaporation in vacuo, the residue was diluted with water and acidified with 1 M hydrochloric acid to pH 6 and extracted with DCM, washed with brine and dried, concentrated under reduced pressure to give 67.6 (200 mg) as a light yellow solid.
[00506] Example 68: Synthesis of (12S)-3-[[4-(4,4-difluoropiperidin-l-yl)cyclohexyl]oxy]- 8-thia-4,6-diazatricyclo[7.4.0.0[2,7]]trideca-l(9),2(7),3,5-tetraene-12-carboxamide (I-51)
Figure imgf000157_0001
66.2 68.1
Figure imgf000158_0001
[00507] Synthesis of compound 68.1. Compound 68.1 was prepared in a manner analogous to the synthesis of 66.10. Isolated 100 mg (quant.) of a yellow solid.
[00508] Synthesis of compound 68.2. A solution of 68.1 (100 mg, crude) in 5 mL of DCM was added oxalic dichloride (250 mg, 5.00 equiv) dropwise at 0 °C, followed by addition of DMF (1 drop) under nitrogen. The resulting solution was stirred for 1 h at room temperature and concentrated under reduced pressure to give 110 mg (crude) of 68.2 as a light yellow solid.
[00509] Synthesis of 1-51. A solution of 68.2 (110 mg, crude) in 5 mL of DCM was added dropwise to 30 mL of ammonia solution and extracted with 4 x 50 mL of dichloromethane and the organic layers combined and dried over anhydrous sodium sulfate and concentrated under vacuum. The crude product (60 mg) was purified by preparative HPLC with the following conditions (Waters): column: XBridge Shield RP18 OBD column: 5 um, 19* 150 mm; mobile phase: water with 0.01% NH4HCO3 and acetonitrile (Gradient B% 10%~33% in 12 min); flow rate: 15 ml/min; UV detection at 254 nm. This resulted in 9.2 mg of 1-51 as a white solid. MS (ES): m/z 451 [M+H]+. 1H-NMR (300 MHz, CD3OD) δ 8.47 (1H, s), 5.35-5.15 (1H, m), 3.27- 3.25 (1H, d, J = 6.0 Hz), 3.05-2.97 (3H, m), 2.89-2.60 (6H, m), 2.33-2.22 (2H, m), 2.20 (1H, d), 2.19-1.85 (7H, m), 1.75-1.57 (4H, m).
[00510] Example 69: Synthesis of (12S)-3-[[4-(dimethylamino)cyclohexyl]oxy]-8-thia-4,6- diazatricyclo[7.4.0.0[2,7]]trideca-l(9),2(7),3,5-tetraene-12-carboxamide. (1-49)
Figure imgf000158_0002
66.9 1-49
69.1 [00511] Synthesis of compound 69.1. 69.1 was prepared in a manner analogous to 1-1. Isolated 120 mg of a light yellow solid in 90% yield.
[00512] Synthesis of 1-49. 1-49 was prepared in a manner analogous to the synthesis of 2.1. Isolated 8.5 mg (11%) of a white solid.MS (ES): m/z 375 (M+H)+. 1H-NMR (300 MHz, CDC13): δ 8.49 (1H, s), 5.66-5.58 (1H, m), 5.48-5.41 (1H, m), 5.25-5.21 (1H, m), 3.33-3.25 (1H, m), 3.15-2.91 (3H, m), 2.89-2.63 (2H, m), 2.55 (6H, s), 2.46-2.36 (2H, m), 2.26-2.10 (3H, m), 2.10- 1.98 (lH, m), 1.61-1.60 (4H, m).
[00513] Example 70: Synthesis of 2-[(3R)-12-[[4-(4,4-difluoropiperidin-l- yl)cyclohexyl] oxy] -7-thia-9, 11-diazatricyclo [6.4.0.0 [2,6] ] dodeca-l(8),2(6),9, 1 l-tetraen-3- yl]acetamide. (1-50)
Figure imgf000159_0001
70.2 1-50
[00514] Synthesis of compound 70.1. Compound 70.1 was prepared according to the method for the preparation of compound 66.2. Isolated 18.0 g (90%>, crude) of 70.1 as a yellow solid. MS (ES): m/z 269 and 271 (M+H)+.
[00515] Synthesis of compound 70.2. 70.2 was prepared from 70.1 and intermediate 67.6 in a manner analogous to the synthesis of 2.1. Isolated 150 mg (quant.) of a red solid.
[00516] Synthesis of Compound 1-50. 1-50 was prepared in a manner analogous to 1-54. Isolated 56 mg of a white solid 56% yield. MS (ES): m/z 451 (M+H)+. 1H-NMR (300 MHz, CD3OD): δ 8.37 (s, 1H), 5.22-5.08 (m, 1H), 3.77-3.65 (m, 1H), 3.09-2.84 (m, 3H), 2.72-2.62 (m, 5H), 2.52-2.45 (m, 1H), 2.32-2.13 (m, 4H), 1.90-1.81 (m, 6H) , 1.60-1.45 (m, 4H). [00517] Example 71: Synthesis of 2-[(3R)-12-[[4-(piperidin-l-yl)cyclohexyl]oxy]-7-thia-9, -diazatricyclo [6.4.0.0 [2,6] ] dodeca-l(8),2(6),9, 1 l-tetraen-3-yl] acetamide. (1-52)
Figure imgf000160_0001
71.3 71.4 I-52
[00518] Synthesis of compound 71.2. Compound 71.2 was prepared from 45.1 in a manner analogous to the synthesis of 46.3. Isolated 250 mg of a yellow oil in 85% overall yield.
[00519] Synthesis of compound 71.4. Compound 71.4 was prepared from 71.2 in a manner analogous to the synthesis of 54.2. Isolated 120 mg (quant., two steps) as a yellow oil.
[00520] Synthesis of Compound 1-52. Compound 1-52 was prepared from 71.4 in a manner analogous to the synthesis of 38.8. Isolated 22 mg (18%) of a white solid. MS (ES): m/z 415 (M+H)+. 1H NMR (400 MHz, CD3OD): δ 8.49 (s, 1H), 5.29-5.24 (m, 1H), 3.81 (m, 1H), 3.33- 3.25 (m, 1H), 3.03-2.98 (m, 2H), 2.85-2.68 (m, 5H), 2.53 (m, 1H), 2.33-2.25 (m, 4H), 2.09-2.06 (m, 2H), 1.68-1.31 (m, 10H).
21] Example 72: Synthesis of Intermediate 72.1
Figure imgf000160_0002
72.1 [00522] Intermediate 72.1 was prepared from 45.1 in a manner analogous to the synthesis of 77.5, except that TBAF/THF was used in the second step rather than HCl/MeOH. Isolated 132 mg of a white solid in 18% overall yield.
[00523] Example 73: Synthesis of 2-[(3R)-12-[(4-aminocyclohexyl)oxy]-7-thia-9,ll- diazatricyclo[6.4.0.0[2,6]]dodeca-l(12),2(6),8,10-tetraen-3-yl]acetamide (1-53)
Figure imgf000161_0001
72.1 I-53
[00524] Compound 1-53 was prepared from 72.1 in a manner analogous to the synthesis of 1-25. Isolated 33.2 mg (36%) of a white solid. MS (ES): m/z 347 [M+H]+. 1H-NMR (300 MHz, CD3OD): δ 1.32-1.46 (m, 2H), 1.61-1.70 (m, 2H), 1.98-2.03(m, 2H), 2.17-2.30(m, 4H), 2.62- 2.74 (m, 1H), 2.82-3.20 (m, 4H), 2.72-3.80 (m, 1H), 5.23-5.3 l(m, 1H), 8.42 (s, 1H).
[00525] Example 74: Synthesis of 2-[(3R)-12-[[4-(piperazin-l-yl)cyclohexyl]oxy]-7-thia- -diazatricyclo[6.4.0.0[2,6]]dodeca-l(12),2(6),8,10-tetra -3-yl]acetamide (I-55)
Figure imgf000161_0002
step 1 step 2
74.3
Figure imgf000161_0003
74.4 '"55
[00526] Synthesis of compound 74.2. To a solution of bis(2-chloroethyl)amine (20 g, 112.05 mmol, 1.00 equiv) and triethylamine (17 mL) in dichloromethane (250 mL) was added di-tert- butyl dicarbonate (26.9 g, 123.25 mmol, 1.10 equiv). The resulting solution was stirred for 30 min at 0 °C in a water/ice bath. The resulting mixture was washed with 2x200 mL of H20. The mixture was dried over anhydrous magnesium sulfate. The solids were filtered out. The resulting mixture was concentrated under vacuum. This resulted in 34.6 g (crude) of tert-butyl N,N-bis(2- chloroethyl)carbamate as an off-white oil.
[00527] Synthesis of compound 74.3. Into a 500-mL round-bottom flask was placed tert- butyl 2-chloroethyl N-(2-chloroethyl)carbamate (24 g, 98.70 mmol, 1.00 equiv), 4- aminocyclohexan-l-ol (12 g, 104.19 mmol, 1.06 equiv), KI (20 g), potassium carbonate (41 g, 296.65 mmol, 3.01 equiv) and CH3CN (200 mL). The resulting solution was heated to reflux for 3 h. The solids were filtered out. The resulting mixture was concentrated under vacuum. The residue was applied onto a silica gel column with dichloromethane/methanol (7: 1). This resulted in 5.5 g (20%) of l-tert-butyl-6-(4-hydroxycyclohexyl)-l [3],3,6-oxadiazocan-2-one as a yellow oil.
[00528] Synthesis of compound 74.4. Compound 74.4 was prepared from 45.1 and 74.3 in a manner analogous to the synthesis of 1-52. Isolated 260 mg of a yellow oil in 39% yield.
[00529] Synthesis of Compound 1-55. To a solution of 74.4 (120 mg, 0.23 mmol, 1.00 equiv) in dichloromethane (4 mL) was added trifluoroacetic acid (2 mL). The resulting solution was stirred for 2 h at room temperature. The resulting mixture was concentrated under vacuum. The crude product (110 mg) was purified by preparative HPLC under the following conditions: column: SunFire Prep CI 8, 19* 150mm 5um; mobile phase: water with 50mL NFL;C03 and CH3CN (15.0% CH3CN up to 36.0% in 13 min, up to 95.0% in 2 min, down to 15.0% in 2 min); detector: 254/220nm. This resulted in 64.7 mg (67%) of 1-55 as a white solid. MS: m/z 416 (M+H)+. 1H NMR (300MHz, CD3OD): δ 1.46-1.86 (4H, m), 2.03-2.24 (2H, m) 2.29-2.44 (5H, m), 2.69-2.77 (5H, m), 2.96-3.01 (6H, m), 3.01-3.10 (1H, m), 3.59-3.79 (1H, m), 5.00-5.26 (1H, m), 8.47 (1H, s).
[00530] Example 75: Synthesis of 5-[[(3R)-12-[[4-(dimethylamino)cyclohexyl]oxy]-7-thia- 9,ll-diazatricyclo[6.4.0.0[2,6]]dodeca-l(8),2(6),9,ll-tetraen-3-yl]methyl]-l,3-oxazolidine- 2,4-dione (1-56)
Figure imgf000163_0001
1-27 1-56
[00531] A solution of 1-27 (25 mg, 0.06 mmol, 1.00 equiv), diethyl carbonate (73 mg, 0.62 mmol, 10.00 equiv), sodium methylate (33.5 mg) in methanol (5 mL) was stirred for 5 h at 80 °C under nitrogen. The resulting mixture was concentrated under vacuum. The crude product (25 mg) was purified by preparative HPLC with the following conditions (Waters): column: Xbridge Prep CI 8, 5um, 19*50mm; mobile phase: water with 0.05% NH4HCO3 and CH3CN (10% CH3CN up to 35%) in 10 min, up to 95% in 2.5 min, down to 10%> in 2.5 min); flow rate: 20mL/min; UV detection at 254/220 nm. After concentration of the fractions under reduced pressure, the desired 1-56 (19.8 mg, 74%) was obtained as a white solid. LCMS: (ES, m/z) 431 [M+H]+. 1H NMR (300 MHz, CD3OD) δ 8.42 (s, 1H), 5.26-5.30 (m, 1H), 4.70-4.74 (m, 1H), 3.60-3.68 (m, 1H), 2.96-3.07 (m, 3H), 2.65-2.73 (d, 7H), 2.08-2.43 (m, 5H), 1.59-1.93 (m, 5H).
[00532] Example 76: Synthesis of 2-((R)-4-(((lr,4R)-4-morpholinocyclohexyl)oxy)-6,7- dihydro-5H-cyclopenta [4,5] thieno [2,3-d] pyrimidin-5-yl)acetamide (1-57)
Figure imgf000163_0002
[00533] Synthesis of compound 76.1. Alcohol 49.2 (185 mg, 0.46 mmol, 1.00 equiv) was oxidized with dipyridinium dichromate (752 mg, 2.00 mmol, 4.36 equiv) in 50 mL of DMF for 24 h at room temperature. The resulting solution was diluted with water and extracted with 3 x 50 mL of mixed solutions of CHCl3/iso-PrOH. The organic layers were combined, dried (Na2S04) and concentrated under vacuum. The residue was applied onto a silica gel column with dichloromethane/methanol (5: 1 to 1 : 1) and purified to afford 105 mg (55%) of acid 76.1 as a yellow oil.
[00534] Synthesis of Compound 1-57. A 50 mL round-bottom flask containing a solution of acid 76.1 (105 mg, 0.25 mmol, 1.00 equiv), NH4C1 (80 mg, 1.50 mmol, 6.00 equiv), EDCI (57 mg, 0.3 mmol, 1.2 equiv), 4-dimethylaminopyridine (37 mg, 0.3 mmol, 1.2 equiv) and HOBt (40 mg, 0.3 mmol, 1.2 equiv) in 5 mL of anhydrous DMF was stirred for 24 h at room temperature. The resulting solution was diluted with water and extracted with 4 x 50 mL of mixed solution of CHCl3:iso-PrOH. The combined organic layers were concentrated under vacuum. The crude product was purified by preparative HPLC (SHIMADZU) under the following conditions: column: SunFire Prep CI 8, 19* 150mm 5um; mobile phase: water (0.05% NH4C03) and CH3CN (6.0% CH3CN up to 50.0% in 25 min); UV detection at 254/220 nm. The product-containing fractions were collected and concentrated to give Compound 1-57 (22.5 mg) as a white solid. 1H NMR (300 MHz, CD3OD) δ 8.43 (s, 1H), 5.27-5.20 (m, 1H), 3.80-3.70 (m, 5H), 3.29-3.27 (m, 1H), 3.12-2.90 (m, 2H), 2.73-2.67 (m, 5H), 2.49-2.42 (m, 1H), 2.32-2.19 (m, 4H), 2.10-2.06 (d, 2H), 1.67-1.46 (m, 4H). MS: m/z 417 (M+H)+.
[00535] Example 77: Synthesis of Intermediate 77.12.
Figure imgf000164_0001
77.3 77.4 OH
Figure imgf000165_0001
Figure imgf000165_0002
77.7 77.8
Figure imgf000165_0003
77.9 77.10 77.11 77.12
[00536] Synthesis of compound 77.2. A 250-mL round-bottom flask, purged and maintained with an inert atmosphere of nitrogen, was charged with 4-hydroxycyclohexane-l-carboxylic acid (15.7 g, 108.90 mmol, 1.00 equiv) in methanol (90 mL). Sulfuric acid (0.8 mL) was added to the mixture slowly. The resulting solution was stirred for 10 h at 60 °C. The reaction was then quenched by the addition of 200 mL of sodium bicarbonate (sat.). The resulting solution was extracted with 3 x 200 mL of ethyl acetate and the organic layers combined and dried over anhydrous sodium sulfate and concentrated under reduced pressure. The resulting 77.2(16.6 g, 96%) was obtained as a light yellow oil.
[00537] Synthesis of compound 76.7. A 500-mL round-bottom flask, purged and maintained with an inert atmosphere of nitrogen, was charged with a solution of methyl trans-4- hydroxycyclohexane-l-carboxylate (16.6 g, 104.93 mmol, 1.00 equiv) and imidazole (14.28 g, 180.53 mmol, 1.72 equiv) in distilled DMF (25 mL). Tert-butyl(chloro)dimethylsilane (28.3 g, 187.76 mmol, 1.79 equiv) was added slowly and the resulting solution was stirred for 14 hrs at room temperature. After completion, the reaction was then quenched with water and extracted with 3 x 200 mL of ethyl acetate. The combined organic layers were washed with water, brine and dried over sodium sulfate. After concentration under reduced pressure, the residue was applied onto a silica gel column with ethyl acetate/petroleum ether (10: 1) to give the desired 77.3 (27.4 g, 96%) as a colorless oil.
[00538] Synthesis of compound 77.4. A 500-mL round-bottom flask containing a solution of diisopropylamine (10 g, 99.01 mmol, 3.00 equiv) in freshly distilled THF (100 mL) was cooled down to -78 °C under nitrogen. Then n-BuLi (2.5 M in hexane, 39.6 mL) was added dropwise and the resulting solution was stirred at -78°C for 1 h. A solution of methyl trans -4-[(tert- butyldimethylsilyl)oxy]cyclohexane-l-carboxylate (9 g, 33.03 mmol, 1.00 equiv) in THF (20 mL) was added via syringe and the reaction mixture was held at -78°C for another 1 h. Iodoethane (25.74 g, 165.04 mmol, 5.00 equiv) was added to the mixture and stirred for additional 2 h at -78°C. Then the reaction temperature was raised to room temperature in 1 h with stirring. The reaction was then quenched with saturated aqueous NH4CI and extracted with 3 x 100 mL of ethyl acetate. The combined organic layers were washed with brine, dried over sodium sulfate and concentrated under vacuum. The residue was applied onto a silica gel column with ethyl acetate/petroleum ether (1 : 15) to give the major 77.4 (8.2 g, 83%) as a yellow oil. The desired cz's-compound was confirmed by 1H NMR spectroscopy.
[00539] Synthesis of compound 77.5. To a 500-mL round-bottom flask containing a solution of 77.4 (8.2 g, 27.29 mmol, 1.00 equiv) in 150 mL of THF was added TBAF 3H20 (12.9 g, 40.95 mmol, 1.50 equiv) and the resulting solution was stirred for 4 h at 30 °C. The resulting mixture was concentrated under vacuum and the residue was applied onto a silica gel column with ethyl acetate/petroleum ether (1 :2) to give 77.5 (4.5 g, 89%>) as a yellow oil.
[00540] Synthesis of compound 77.6. A solution of 77.5 (3.5 g, 18.79 mmol, 1.00 equiv) in 100 mL of THF was added 4-nitrobenzoic acid (6.3 g, 37.70 mmol, 2.01 equiv), PPh3 (9.85 g, 37.55 mmol, 2.00 equiv) and DIAD (7.6 g, 37.58 mmol, 2.00 equiv) successively at room temperature under N2. The resulting solution was stirred for 48 hrs at ambient temperature. After completion, the reaction was quenched with water and extracted with EtOAc (3 x 100 mL). The combined organic layers were dried over sodium sulfate and concentrated under vacuum. The residue was applied onto a silica gel column with ethyl acetate/petroleum ether (1 : 15) to give the desired 77.6 (2.3 g, 36%) as a white solid. 1H NMR (300 MHz, CDC13): δ 8.32 (2H, d), 8.21 (2H, d), 5.28-5.20 (1H, M), 3.72 (3H, s), 2.10-2.04 (2H, m), 1.98-1.90 (2H, m), 1.84-1.73 (2H, m), 1.68-1.56 (5H, m), 0.88 (3H, t). [00541] Synthesis of compound 77.7. To a 50-mL round-bottom flask containing a solution of 77.6 (2.3 g, 6.86 mmol, 1.00 equiv) in a mixture of methanol (15 mL) and water (3 mL) was added potassium carbonate (2.84 g, 20.55 mmol, 3.00 equiv) and the resulting solution was stirred for 2 h at 40 °C. The resulting solution was quenched with water and extracted with 3 x 50 mL of ethyl acetate. The combined organic layers were washed with brine and dried over anhydrous sodium sulfate and concentrated under vacuum. The residue was purified by column chromatography on silica gel with ethyl acetate/petroleum ether (1 : 18) to afford the corresponding 77.7 (1.2 g, 94%) as a white solid. 1H NMR (300 MHz, CDC13): δ 3.89-3.85 (1H, m), 3.69 (3H, s), 1.95-1.79 (2H, m), 1.72-1.54 (8H, m), 0.80 (3H, t).
[00542] Synthesis of compound 77.8. To a solution of 77.7 (1.0 g, 5.37 mmol, 1.00 equiv) and DIEA (2.08 g, 16.09 mmol, 3.00 equiv) in dichloromethane (20 mL) was added SEMC1 (1.79 g) slowly at room temperature. The resulting solution was stirred for 14 h at ambient temperature. The reaction was then quenched with water and extracted with 3 x 40 mL of ethyl acetate. The combined organic layers were washed with brine, dried over sodium sulfate and concentrated under vacuum. The residue was applied onto a silica gel column with ethyl acetate/petroleum ether (1 : 10) to provide 77.8 (1.5 g, 88%) as a colorless oil. 1H NMR (300 MHz, CD3OD): δ 4.65 (2H, s), 3.71-3.58 (6,H, m), 1.86-1.82 (2H, m), 1.68-1.49 (8H, m), 0.90 (2H, t), 0.78 (3H, t).
[00543] Synthesis of compound 77.9. A solution of 77.8 (1.5 g, 4.74 mmol, 1.00 equiv) in a mixed methanol (20 mL)/water(5 mL) was added sodium hydroxide (948 mg, 23.70 mmol, 5.00 equiv) and the resulting solution was stirred for 14 h at 75 °C. After cooling down to r.t, the pH value of the mixture was adjusted to 4 with 2 M aqueous hydrochloric acid and extracted with 3 x 50 mL of ethyl acetate. The combined organic layers were washed with brine, dried over anhydrous sodium sulfate and concentrated under vacuum. The desired 77.9 (1.3 g, 91%>) was obtained as a yellow oil. 1H NMR (300 MHz, CD3OD): δ 4.69 (2H, s), 3.80-3.70 (1H, m), 3.67 (2H, t), 1.87-1.82 (2H, m), 1.75-1.54 (8H, m), 0.95 (2H, t), 0.92 (3H, t).
[00544] Synthesis of compound 77.10. A solution of 77.9 (1.24 g, 4.10 mmol, 1.00 equiv), DPPA (2.03 g, 7.38 mmol, 1.80 equiv) and TEA (1.24 g, 12.25 mmol, 2.99 equiv) in 2- methylpropan-2-ol (30 mL) was stirred for 14 h at reflux under nitrogen. The reaction mixture was quenched with water, extracted with 3 x 50 mL of EtOAc. The organic layers were combined, washed with brine, dried over sodium sulfate and concentrated under vacuum. The residue was applied onto a silica gel column with ethyl acetate/petroleum ether (1 :6) to give 77.10 (0.9 g, 73%) as a colorless oil. 1H NMR (400 MHz, CD3OD): δ 4.70 (2H, s), 3.89-3.80 (1H, m), 3.67 (2H, t), 1.80-1.73 (6H, m), 1.65-1.58 (4H, m), 1.00 (3H, t), 0.94 (2H, t).
[00545] Synthesis of compound 77.11. Into a 25-mL round-bottom flask contained a solution of 77.10, trimethylsilane (840 mg, 2.80 mmol, 1.00 equiv) in tetrahydrofuran (10 mL) was added hydrochloric acid (5 M, 2 mL) slowly and the resulting solution was stirred for 14 hrs at 30°C. The resulting mixture was concentrated under vacuum to provide 77.11 (410 mg, crude) as a white solid.
[00546] Synthesis of Intermediate 77.12. A solution of 77.11 (380 mg, 2.11 mmol, 1.00 equiv) and sodium hydroxide (127 mg, 1.13 mmol, 0.54 equiv) in a mixture of THF (30 mL)/water (5mL) was added (Boc)20 (462 mg, 2.12 mmol, 1.00 equiv) at 0 °C. The resulting solution was stirred for 3 h at room temperature and diluted with water, extracted with 3 x 50 mL of ethyl acetate. The organic layers were combined, washed with brine, dried over anhydrous sodium sulfate and concentrated under vacuum. The residue was purified by column chromatography on silica gel with ethyl acetate/petroleum ether (1 :6) to give the desired 77.12 (450 mg, 87%) as a colorless oil. 1H NMR (300 MHz, CD3OD): δ 3.80-3.72 (1H, m), 1.69-1.61 (8H, m), 1.50-1.32 (11H, m), 0.78 (3H, t).
[00547] Example 78. Synthesis of 2-((R)-4-(((lr,4R)-4-(dimethylamino)-4- ethylcyclohexyl)oxy)-6,7-dihydro-5H-cyclopenta [4,5] thieno [2,3-d] pyrimidin-5-yl)ethanol (1-58)
TBS
SO-"\
Figure imgf000169_0001
35.1
Figure imgf000169_0002
78.2 I-58
[00548] Synthesis of compound 78.1. 78.1 was prepared from 35.1 and 77.12 in a manner analogous to the synthesis of 10.1 from 1.8. Isolated 480 mg (43%) as a colorless oil. MS: 576 [M+H]+.
[00549] Synthesis of Compound 1-58. 1-58 was prepared from 78.1 in a manner analogous to the synthesis of 1-27 from 40.2. Isolated 41.4 mg (14%) as a white semi-solid. MS (ES): m/z 390 [M+H]+. 1H NMR (400 MHz, CD3OD): δ 8.47 (s, 1H), 5.44-5.46 (m, 1H), 3.64-3.69 (m, 2H), 3.32-3.33 (m, 1H), 3.10-3.12 (m, 1H), 2.99-3.02 (m, 1H), 2.68 (m, 1H), 2.09-2.39 (m, 10H), 1.61-1.86 (m, 9H), 0.93-0.96 (t, 3H).
Figure imgf000169_0003
38.3 79.1
[00551] Compound 79.1 was prepared from 38.3 in a manner analogous to the synthesis of 76.1 from 49.2. Isolated 0.9 g (76%) as a colourless oil. [00552] Example 80: 2-[(3R)-12-[[4-(methylamino)cyclohexyl]oxy]-7-thia-9,ll- diazatricyclo[6.4.0.0A[2,6]]dodeca-l(8),2(6),9,ll-tetraen-3-yl]acetamide (1-60)
Figure imgf000170_0001
[00553] Synthesis of compound 80.1. Into a 50-mL round-bottom flask containing a solution of 79.1 (220 mg, 0.48 mmol, 1.00 equiv) in 4 mL of distilled DMF was added HOBT (96.6 mg), 4-dimethylaminopyridine (86.6 mg), EDCI (136.7 mg) and NH4C1 (153.18 mg, 2.86 mmol, 6.01 equiv) successively at room temperature under nitrogen. The resulting solution was stirred for 14 h at 25°C and diluted with water, extracted with 3 x 50 mL of ethyl acetate. The combined organic layers was washed with brine, dried over sodium sulfate and concentrated under vacuum. The residue was applied onto a silica gel column with ethyl acetate/petroleum ether (2: 1) to give 80.1 (176 mg, 80%) as a colorless oil. MS (ES): m/z 461 [M+H]+.
[00554] Synthesis of Compound 1-60. 1-60 was prepared in a manner analogous to the synthesis of 1-7 from 4.5. Isolated 83 mg (62%) as a white solid. MS (ES): m/z 361 [M+H]+. 1H NMR (300 MHz, CD3OD): δ 8.43(lH,s), 5.28(1H, m), 3.76 (1H, m), 3.00-3.28 (1H, m), 2.97- 2.99 (2H, m), 2.66-2.73 (1H, m), 2.42-2.49 (1H, m), 2.36 (3H, s), 2.29-2.18 (4H, m), 2.00-2.03 (2H, m), 1.65-1.57 (2H-m), 1.26-1.35 (2H, m).
[00555] Example 81: 2-[(3R)-12-[(4-[methyl[2-oxo-2-(pyrrolidin-l- yl)ethyl] amino] cyclohexyl)oxy]-7-thia-9,l 1-diazatricyclo [6.4.0.0 [2,6] ] dodeca-l(8),2(6),9,l 1- tetraen-3-yl]acetamide (1-59)
Figure imgf000170_0002
[00556] Synthesis of Compound 1-59: A solution of 1-60 (90 mg, 0.25 mmol, 1.00 equiv) in 8 mL of DMF was added 2-chloro-l-(pyrrolidin-l-yl)ethan-l-one (55 mg, 0.37 mmol, 1.50 equiv) and potassium carbonate (69 mg, 0.50 mmol, 2.00 equiv) and the resulting solution was stirred overnight at room temperature. The reaction was quenched with water and extracted with DCM. The organic phase was washed with brine, dried over anhydrous sodium sulfate and concentrated under vacuum. The crude product (80 mg) was purified by preparative HPLC under the following conditions (SHIMADZU): column: SunFire Prep C18, 19* 150mm 5 um; mobile phase: water with 0.05% NH4HCO3 and CH3CN (6.0% CH3CN up to 50.0% in 16 min); flow rate: 20 mL/min; UV detection at 254/220 nm. The product-containing fractions were collected and partially evaporated to remove water and CH3CN under reduced pressure. The residue was lyophilized overnight to give the desired 1-59 (50 mg) as a white solid. LCMS (ES): m/z 472
[M+H]+. 1H NMR (300 MHz, CD3OD): δ 8.43 (1H, s), 5.25-5.20 (1H, m), 3.85-3.69 (1H, m), 3.52 (2H, t), 3.42 (2H, t), 3.37 (2H, s), 3.15-2.85 (2H, m), 2.73-2.58 (2H, m), 2.29-2.18 (7H, m), 1.96-1.79 (6H, m), 1.67-1.47 (4H, m).
[00557] Example 82: Synthesis of 2-((R)-4-(((lr,4R)-4-(ethylamino)cyclohexyl)oxy)-6,7- dihydro-5H-cyclopenta [4,5] thieno [2,3-d] pyrimidin-5-yl)acetamide. (1-61)
Figure imgf000171_0001
1-61
[00558] Synthesis of Compound 1-61. 1-61 was prepared from intermediate 39.3 in a manner analogous to the synthesis of 1-60 from 38.3. Isolated 114.4 mg (31% yield, three steps) as a white solid. MS (ES): m/z 375 [M+H]+. 1H-NMR (300 MHz, CD3OD): δ 8.47 (1H, s), 5.29 (1H, m), 4.82 (1H, m), 3.07-3.10 (1H, m), 2.93-2.99 (2H, m), 2.58-2.73 (4H, m), 2.23-2.23 (4H, m), 2.03-2.18 (2H, m), 1.66 (2H, m), 1.40-1.30 (2H, m), 1.03-1.18 (3H, t).
[00559] Example 83. 2-((R)-4-(((lr,4R)-4-(methyl(2-(pyridin-2- yl)ethyl)amino)cyclohexyl)oxy)-6,7-dihydr o-5H-cyclopenta [4,5] thieno [2,3-d] pyrimidin-5- yl)acetamide (1-62)
Figure imgf000172_0001
80.1 I-60
Figure imgf000172_0002
I-62
[00560] Synthesis of Compound 1-60 (HC1 salt). To a solution of 80.1 (524 mg, 1.14 mmol, 1.00 equiv) in DCM (6 mL) was added 6 M aqueous hydrochloric acid (1 mL) at 0 °C and the resulting solution was stirred for 2 h at room temperature. After concentration under reduced pressure, the desired 1-60 HC1 salt (451 mg, crude) was obtained as a light yellow solid.
[00561] Synthesis of Compound 1-62. To the hydrochloride (111 mg, 0.29 mmol, 1.00 equiv) in 4 mL of distilled DMF was added DIEA (119 mg, 0.92 mmol, 3.13 equiv) and 2-(2- bromoethyl)pyridine (57 mg, 0.31 mmol, 1.04 equiv) at room temperature. The resulting solution was stirred overnight at 100 °C in an oil bath. The resulting mixture was diluted with water and extracted with DCM and concentrated under vacuum. The crude product (110 mg) was purified by preparative HPLC under the following conditions (Waters): column: Xbridge Prep C18, 5um, 19*50mm; mobile phase: water with 0.05% NH4HCO3 and CH3CN (10% CH3CN up to 25% in 10 min, up to 95% in 1.5 min, down to 10% in 1.5 min); UV detection at 254/220 nm. The product-containing fractions were collected and partially evaporated under reduced pressure to remove CH3CN and water. The residue was lyophilized overnight to give the desired 1-62 (16.3 mg) as a light yellow solid. MS: m/z 466 (M+H)+. 1H NMR (400 MHz, CD3OD): δ 8.48 (d, 2H), 7.80 (t, 1H), 7.79 (d, 1H), 7.77-7.27 (m, 1H), 5.29-5.23 (m, 1H), 3.81-3.79 (m, 1H), 3.16-3.10 (m, 1H), 3.03-2.93 (m, 6H), 2.78-2.73 (m, 2H), 2.44 (s, 3H), 2.33-2.26 (m, 4H), 2.02 (d, 3H), 1.68-1.59 (m, 4H), 1.57-1.54 (m, 1H). [00562] Example 84: Synthesis of 2-[(3R)-12-[[4-(pyrrolidin-l-yl)cyclohexyl]oxy]-7-thia- ll-diazatricyclo[6.4.0.0[2,6]]dodeca-l(8),2(6),9, ll-tetraen-3-yl]acetamide. (1-63)
Figure imgf000173_0001
84.2
Figure imgf000173_0002
1-63
[00563] Synthesis of compound 84.2. Into a 50-mL round-bottom flask purged and maintained with an inert atmosphere of nitrogen was placed a mixture of trans-4- aminocyclohexan-l-ol (1.0 g, 8.7 mmol, 1.00 equiv), KI (1.87 g), potassium carbonate (0.3 g, 2.2 mmol, 0.25 equiv) and 1 ,4-dibromobutane (1.93 g, 8.9 mmol, 1.03 equiv) in CH3CN (30 mL). The resulting solution was heated to reflux for 2 h and filtered to remove the solids. The filtrate was concentrated in vacuo and the residue was diluted with water and extracted with 3 x 50 mL of ethyl acetate. The combined organic layers were dried over sodium sulfate and concentrated under vacuum. The residue was applied onto a silica gel column with dichloromethane/methanol (10/1) to give in 1.4 g (95%) of 84.2 as a yellow solid.
[00564] Synthesis of compound 1-63. 1-63 was prepared from 84.2 and 45.1 in a manner analogous to the synthesis of 1-52 from 45.1. Isolated 116 mg (38%, 5 steps) was obtained as a white solid. MS (ES): m/z 401 (M+H)+. 1H-NMR (400 MHz, CD3OD): δ 8.47 (1H, s), 5.31 (1H, m), 4.80 (1H, m), 3.14 (1H, m), 3.02 (2H, m), 2.70 (5H, m), 2.15-2.33 (7H, m), 1.84 (4H, s), 1.68 (2H, m), 1.63 (2H, m).
[00565] Example 85: Synthesis of 2-[(3R)-12-[[4-(dimethylamino)-4- ethylcyclohexyl] oxy] -7-thia-9,l 1 -diazatricyclo [6.4.0.0 [2,6] ] dodeca- 1 (8),2(6),9, 1 l-tetraen-3- yljacetamide (1-64)
Figure imgf000174_0001
Figure imgf000174_0002
85.2
[00566] Synthesis of compound 85.1. A solution of 78.1 (1.4 g, 2.43 mmol, 1.00 equiv) and TBAF (1.15 g, 4.40 mmol, 1.81 equiv) in tetrahydrofuran (15 mL) was stirred for 2 h at 25 °C. The reaction was then quenched by the addition of water, extracted with 3x100 mL of ethyl acetate and the organic layers combined and concentrated under vacuum. The residue was applied onto a silica gel column with ethyl acetate/petroleum ether (1 :3). This resulted in 1.1 g (98%) of 85.1 as a colorless oil.
[00567] Synthesis of compound 85.2. 85.2 was prepared in a manner analogous to the synthesis of 76.1 from 49.2. Isolated 100 mg (32%) as a colorless oil. MS (ES): m/z 476 [M+H]+.
[00568] Synthesis of Compound 1-64. To a 50-mL round-bottom flask containing a solution of 85.2 (60 mg, 0.13 mmol, 1.00 equiv) in dichloromethane (4 mL) was added hydrochloric acid (8 M, 0.5 mL) at 0 °C. The resulting solution was stirred for 1 h at the same temperature. The reaction mixture was concentrated in vacuo to give the 2-[(3R)-12-[(4-amino-4- ethylcyclohexyl)oxy]-7-thia-9,l l-diazatricyclo[6.4.0.0[2,6]]dodeca-l(8),2(6),9,l l-tetraen-3- yljacetamide hydrochloride (50 mg, crude) as a white solid. The solid (50 mg, crude) was dissolved in 5 mL of methanol, HCHO (37%, 0.5 mL) was added and the solution was stirred at room temperature for 30 min. NaBH3CN (22 mg) was added to the mixture and stirred overnight. The reaction was then quenched by the addition of 30 mL of water, extracted with 3 x 40 mL of chloroform/isopropanol. The combined organic layers were concentrated under vacuum. The crude product (50 mg) was purified by preparative HPLC under the following conditions (SHIMADZU): column: SunFire Prep C18, 19* 150mm 5um; mobile phase: water with 50 mM NH4HCO3 and CH3CN (6.0% CH3CN up to 52.0% in 14 min); flow rate: 20 mL/min; UV detection at 254/220 nm. The product-containing fractions were collected and partially evaporated to remove water and CH3CN under reduced pressure. The residue was lyophilized overnight to give 1-64 (25.1 mg) as a white solid. MS (ES): m/z 403 [M+H]+. 1H NMR (300 MHz, CD3OD): δ 8.47 (1H, s), 5.29 (1H, m), 3.85-3.89 (1H, m), 2.9-3.20 (3H, m), 2.70-2.88 (1H, m),2.22-2.46 (8H, m), 2.06-2.17 (2H, m), 1.65-1.93 (8H, m), 0.98-1.07 (3H, m).
[00569] Example 86: Synthesis of 12-[[4-(morpholin-4-yl)cyclohexyl]oxy]-7-thia-9,ll- diazatricyclo[6.4.0.0[2,6]]dodeca-l(8),2(6),9,ll-tetraene (1-75)
Figure imgf000175_0001
1-75
[00570] Synthesis of compound 86.4. 86.4 was prepared from cyclopentanone in a manner analogous to the synthesis of 34.1. Isolated 6.6 g (29%, 3 steps) was obtained as a yellow solid. LC-MS (ES): m/z 211 (M+H)+.
[00571] Synthesis of Compound 1-75. 1-75 was prepared from 86.4 in a manner analogous to the synthesis of 10.1 from 1.8. Isolated 55.4 mg (23%) as a white solid. LC-MS (ES): m/z 360 [M+H]+ and 401 [M+H+CH3CN]+. 1H-NMR (300 MHz, CD3OD): δ 8.44 (1H, s), 5.25 (1H, m), 3.72 (4H, t), 3.02 (4H, t), 2.63 (4H, t), 2.51 (2H, m), 2.40-2.29 (3H, m), 2.10-2.00 (2H, m), 1.65- 1.54 (4H, m).
[00572] Example 87: Synthesis of (lr,4r)-Nl-(6,7-dihydro-5H-cyclopenta[4,5]thieno[2,3- d] pyrimidin-4-yl)-N4,N4-dimethylcyclohexane- 1 ,4-diamine (1-72)
Figure imgf000176_0001
[00573] Synthesis of compound 87.1. The mixture of 86.4 (1.0 eq) and trans-4- aminocyclohexylamine (3.0 eq) in iPrOH ([86.4] = 0.24 M) was heated at reflux for 12 h. The solvent was removed under vacuum and water (20 mL) was added. The aqueous phase was extracted by CH2C12 (3 x 40 mL). The combined organic phases was washed (brine), dried (Na2S04), filtered and concentrated. The residue was purified by column chromatography on silica gel. Isolated a white solid in 87% yield. 1H NMR (400 MHz, DMSO-d6) δ 1.16-1.23 (m, 2H), 1.44-1.53 (m, 2H), 1.80-1.92 (m, 4H), 2.40-2.43 (m, 2H), 2.58-2.63 (m, 1H), 2.92 (t, J= 7.2 Hz, 2H), 3.06 (t, J= 7.2 Hz, 2H), 4.00-4.04 (m, 1H), 5.97 (d, J= 8.0 Hz, 1H), 8.26 (s, 1 H). MS: m/z 289.1 (M+H) +.
[00574] Synthesis of Compound 1-72. A mixture of compound 86.4 (1.97 g, 6.83 mmol, 1.00 equiv) and formaldehyde (2.05 g, 68.24 mmol, 9.99 equiv) in formic acid (20 mL) was heated to reflux for 16 h. The reaction mixture was cooled with a water/ice bath. The pH value of the solution was adjusted to 12 with sodium hydroxide (2N). The solids were collected by filtration. The crude product was purified by re-crystallization from ethanol to give 984.4 mg (53%) of Compound 1-72 as a yellow solid, m. p. = 97-98 °C. 1H NMR (400 MHz, CDC13) δ 8.36 (1H, s), 7.26-7.07 (4H, m), 4.85-4.83 (1H, d), 4.12-4.02 (1H, m), 3.00-2.97 (4H, m), 2.76-2.70 (4H, m), 2.58-2.49 (3H, m), 2.37 (3H, s), 2.27-2.23 (2H, d), 1.94-1.89 (2H, d), 1.71-1.47 (2H, q), 1.29- 1.25 (2H, q). MS: m/z 317 (M+H)+.
[00575] Example 88. Synthesis of l-N-methyl-l-N-[2-(pyridin-2-yl)ethyl]-4-N-[7-thia- 9,ll-diazatricyclo[6.4.0.0A[2,6]]dodeca-l(8),2(6),9,ll-tetraen-12-yl]cyclohexane-l,4- diamine (1-73)
Figure imgf000177_0001
Figure imgf000177_0002
[00576] Synthesis of compound 88.1. A mixture of 87.1 (4 g, 13.87 mmol, 1.00 equiv), triethylamine (2.1 g, 20.75 mmol, 1.50 equiv) and di-tert-butyl dicarbonate (3.3 g, 15.12 mmol, 1.09 equiv) in dichloromethane (60 mL) was stirred for 2 h at room temperature. The reaction was then quenched by the addition of 50 mL of water. The organic phase was dried and concentrated under vacuum to give 4.8 g (crude) of 88.1 as a yellow solid.
[00577] Synthesis of compound 88.2. Lithium aluminium tetrahydride (590 mg, 15.53 mmol, 3.00 equiv) was added into a solution of 88.1 (2 g, 5.15 mmol, 1.00 equiv) in tetrahydrofuran (50 mL) in several batches at 0 °C in 10 min under an inert atmosphere of nitrogen.. The resulting solution was stirred for 2 h at 80 °C in an oil bath. The reaction was then quenched by the addition of 10 mL of 15% aqueous sodium hydroxide. The solids were collected by filtration and dried under reduced pressure to get 1.5 g (crude) of 88.2 as a yellow solid.
[00578] Synthesis of Compound 1-73. 1-73 was prepared from 87.2 in a manner analogous to the synthesis of 1-33 from 1-25, substituting 2-(2-bromoethyl)pyridine for 2-chloro-l- (pyrrolidin-l-yl)ethanone. Isolated 27.1 mg (9%) of 1-73 as a light yellow oil. 1H NMR (300 MHz, CD3OD) δ 8.47-8.45 (1H, m), 8.21 (1H, s), 7.80-7.74 (1H, m), 7.38-7.35 (1H, m), 7.29- 7.25 (1H, m), 4.07-4.04 (1H, m), 3.07-2.85 (8H, m), 2.61-2.44 (3H, m), 2.39 (3H, s), 2.22-2.14 (2H, m), 1.96-1.94 (2H, m), 1.55-1.45 (4H, m). MS: m/z 408 (M+H)+.
[00579] Example 89: 6-((lr,4r)-4-((6,7-dihydro-5H-cyclopenta[4,5]thieno[2,3- d]pyrimidin-4-yl)oxy)cyclohexyl)-2-oxa-6-azaspiro[3.3]heptane (1-74)
Figure imgf000178_0001
[00580] Synthesis of compound 89.2. To a solution of 4-aminocyclohexan-l-ol (345 mg, 3.00 mmol, 1.00 equiv) in CH3CN (10 mL) was added KI (124 mg, 0.75 mmol, 0.25 equiv), K2C03 (1.7 g, 12.32 mmol, 4.00 equiv), KOH (168 mg, 2.99 mmol, 1.00 equiv) and 3,3- bis(chloromethyl)oxetane (558 mg, 3.60 mmol, 1.20 equiv). The resulting solution was stirred overnight at 80 °C. The mixture was diluted with DCM (30 mL), washed with brine (30 mL), dried over anhydrous sodium sulfate and concentrated under vacuum. The residue was applied onto a silica gel column with dichloromethane/methanol (5: 1) to result 70 mg (12%) of 89.2 as a colorless oil.
[00581] Synthesis of compound 1-74. 1-74 was prepared from 89.2 in a manner analogous to the synthesis of 10.1 from 1.8. Isolated 32 mg (18%) as a white solid. MS: (ES, m/z) 372 [M+H]+. H-NMR(400 MHz, CDC13) δ 8.50 (s, 1H), 5.26-5.21 (m, 1H), 4.77 (s, 4H), 3.50 (s, 4H), 3.04 (t, 2H, J = 7.2 Hz), 3.02 (t, 2H, J = 7.2 Hz), 2.53 (quint, 2H, J = 7.2 Hz), 2.21-2.19 (m, 2H), 2.08-2.05 (m, 1H), 1.89-1.85 (m, 2H), 1.49-1.62 (m, 2H), 1.21-1.32(m, 2H).
[00582] Example 90: Synthesis of 2-[(3R)-10-[(l-methyl-lH-pyrazol-4-yl)amino]-12- [ [trans-4-(morpholin-4-yl)cyclohexyl] oxy] -7-thia-9,l 1-diazatricyclo [6.4.0.0 [2,6] ] dodeca- l(8),2(6),9,ll-tetraen-3-yl]ethan-l-ol (I-5)
E
Figure imgf000179_0001
step 1 step 2
Figure imgf000179_0002
step 3
90.2 90.3
Figure imgf000179_0003
[00583] Synthesis of compound 90.1. A solution of 1.9 (330 mg, 1.0 mmol, 1.0 equiv) in 8 mL of freshly distilled THF cooled down to -78 °C was added DIBAL-H (1 M in hexane, 2.5 mL, 2.5 equiv) via syringe under nitrogen. The resulting solution was warmed slowly up to -30 °C in 1 h. After completion of the reaction, the mixture was quenched with saturated aqueous NH4CI, extracted with EtO Ac (3 x 40 mL). The combined organic layers were washed with brine, dried over sodium sulfate and concentrated in vacuo to afford 260 mg (90%) of 90.1 as a light yellow oil. MS (ES): m/z 289 and 291 (M+H)+.
[00584] Synthesis of compound 90.2. A solution of 90.1 (260 mg, 0.9 mmol, 1.0 equiv) in dry DCM (8 mL) was added imidazole (122.4 mg, 1.8 mmol, 2.0 equiv), followed by addition of TBSCI (163 mg, 1.08 mmol, 1.2 equiv) at 0 °C under nitrogen. After stirred for 3 h at room temperature, the reaction mixture was diluted with saturated aqueous NH4CI, extracted with EtOAc (3 x 40 mL). The combined organic layers were washed with 0.5 M HC1, brine, dried over sodium sulfate and concentrated in vacuo to afford 90.2 (327 mg, 90%) as a light yellow oil. MS (ES): m/z 404 and 406 (M+H)+.
[00585] Synthesis of compound 90.4. Compound 90.4 was prepared from 90.2 in a manner consistent with the synthesis of 2.2 from 1.9. Isolated 120 mg (30%, two steps) of a light yellow oil. MS (ES): m/z 552 and 554 (M+H)+.
[00586] Synthesis of Compound 1-5. A solution of 90.4 (120 mg, 0.20 mmol, 1.00 equiv) in methanol/water (4 mL/1 mL) was added AcOH (3 mL) and the resulting solution was stirred for 4 h at room temperature. The resulting mixture was concentrated under vacuum. The crude product (120 mg) was purified by preparative HPLC with the following conditions (SHIMADZU): column: SunFire Prep C18, 19* 150mm 5um; mobile phase: water with 1% HCOOH and CH3CN (3.0% CH3CN up to 10.0% in 15 min); flow rate: 15 mL/min; UV detection at 254/220 nm. The product-containing fractions were collected and partially evaporated to remove solvents under reduced pressure. The residue was lyophilized overnight to give 50.4 mg of 1-5 as a white solid. MS (ES): m/z 499 (M+H)+. 1H-NMR (300 MHz, CD3OD) δ 7.90 (1H, s), 7.59 (1H, s), 5.28-5.08 (1H, m), 3.90 (3H, s), 3.85-3.75 (4H, m), 3.65 (2H, t), 3.30 (1H, m), 3.05-2.92 (1H, m), 2.90-2.75 (5H, m), 2.70-2.50 (2H, m), 2.50-2.30 (2H, m), 2.29-2.10 (4H, m), 1.80-1.45 (5H, m).
[00587] Example 91: Synthesis of Intermediates 91.3 and 91.4.
Figure imgf000180_0001
91.1A
[00588] Synthesis of compound 91.1. Compound 91.1 was prepared from ethyl 3- oxocyclohexanecarboxylate in a manner analogous to the synthesis of 1.4. Isolated 18 g of a light yellow oil as an approximately 13:5 mixture of 91.1 and 91.1A in 46% overall yield. MS (ES): m/z 445 (M+H)+.
[00589] Synthesis of compound 91.2. A solution of mixture 91.1 and 91.1A (1 g, 2.25 mmol, 1.00 equiv) in POCI3 (10 mL) was heated at 110 °C for 24 h under nitrogen. The excess POCI3 was removed under reduced pressure and the residue was diluted with EtOAc, poured into cooled saturated aqueous NaHC03, extracted with EtOAc, washed with brine, dried over sodium sulfate and concentrated in vacuo. The residue was applied onto a silica gel column with ethyl acetate/petroleum ether (1 : 100 to 1 :50) to afford the desired 200 mg of pure 91.2 as a white solid. MS (ES): m/z 331 and 333 (M+H)+.
[00590] Synthesis of compound 91.3 and 91.4. Into a 100-mL round-bottom flask containing a solution of 91.2 (150 mg, 0.45 mmol, 1.00 equiv) in 10 mL of distilled THF was cooled down to -70 °C under nitrogen. Then a solution of DIBAL-H (1 M in hexane, 0.9 mL) was added via syrine at -70 °C and the resulting solution was stirred for 1 h at -30 °C. After the starting material was consumed completely, the reaction was then quenched with saturated aqueous NH4C1, extracted with 3 x 50 mL of EtOAc. The organic phase was washed with brine, dried over anhydrous sodium sulfate and concentrated under vacuum. The residue was applied onto a silica gel column with EtOAc / PE (1 :30 to 1 : 10) to give 110 mg (85%) of [3,5-dichloro-8-thia-4,6- diazatricyclo[7.4.0.0A[2,7]]trideca-l(9),2(7),3,5-tetraen-12-yl]methanol as a white solid. MS (ES): m/z 289 and 291 (M+H)+.
[00591] The enantiomers of the latter compound (550 mg) were resolved by chiral-preparative HPLC with the following conditions: column: Chiralpak IC, 2*25cm, 5um; mobile phase: ethanol and hex (v:v = 15:85); flow rate: 20 mL/min; UV detection at 220/254 nm. The first peak fractions were collected and evaporated under reduced pressure to give 240 mg of 91.3 as a white solid; the second peak fractions give 220 mg of 91.4. MS (ES): m/z 290 (M+H)+.
[00592] Example 92: Synthesis of (12R)-5-[(l-methyl-lH-pyrazol-4-yl)amino]-3-[[(lr,4r)- 4-(morpholin-4-yl)cyclohexyl] oxy] -8-thia-4,6-diazatricyclo [7.4.0.0 [2,7] ] trideca- 1 (9),2,4,6- tetraene-12-carboxamide (1-11)
Figure imgf000182_0001
Figure imgf000182_0002
step 3 step 4
92.3 1-11
[00593] Synthesis of compound 92.1. Compound 92.1 was prepared from 91.4 in a manner analogous to the synthesis of 66.2 from 66.1. Isolated 160 g (76%, crude) as a white solid. MS (ES): m/z 304 and 306 (M+H)+.
[00594] Synthesis of compound 92.2. Compound 92.2 was prepared from 92.1 in a manner analogous to the synthesis of 1-1 from 2.3. Isolated 120 mg (75%) as a white solid. MS (ES): m/z 303 and 305 (M+H)+.
[00595] Synthesis of compound 92.3. Compound 92.3 was prepared from 92.2 in a manner analogous to the synthesis of 2.1 from 1.9. Isolated 160 g (76%, crude) as a white solid. MS (ES): m/z 451 and 453 (M+H)+.
[00596] Synthesis of Compound 1-11. 1-11 was prepared from 92.3 in a manner analogous to the synthesis of 2.2 from 2.1. Isolated 15.1 mg (10%) of a white solid. MS (ES): m/z 512 (M+H)+. 1H NMR (300 MHz, CD3OD) δ 7.86 (s, 1H), 7.52 (s, 1H), 5.17-5.15 (m, 1H), 3.85 (s, 3H), 3.72-3.67 (m, 4H), 3.20-3.11 (m, 1H), 2.89-2.79 (m, 3H), 2.70-2.57 (m, 5H) , 2.31-2.77 (m, 3H) , 2.13-2.041 (m, 3H) , 1.99-1.81 (m, 1H) , 1.67-1.31 (m, 4H).
[00597] Example 93: Synthesis of (12S)-5-[(l-methyl-lH-pyrazol-4-yl)amino]-3-[[trans-4- (morpholin-4-yl)cyclohexyl] oxy] -8-thia-4,6-diazatricyclo [7.4.0.0 [2,7] ] trideca- 1 (9),2,4,6- tetraene-12-carboxamide (1-17)
Figure imgf000183_0001
1-17
[00598] 1-17 was prepared from 91.3 in a manner analogous to the synthesis of 1-11 from 91.4. Isolated 5.3 mg (1% yield, 4 steps) of a white solid. MS (ES): m/z 512 (M+H)+. 1H NMR (300 MHz, CD3OD) δ 7.89 (s, 1H), 7.56 (s, 1H), 5.22-5.13 (m, 1H), 3.88 (s, 3H), 3.74-3.71 (m, 4H), 3.22-3.15 (m, 1H), 2.99-2.83 (m, 3H), 2.75-2.62 (m, 5H) , 2.35-2.32 (m, 3H) , 2.21-2.12 (m, 3H) , 2.08-1.94 (m, 3H) , 1.70-1.42 (m, 4H).
[00599] Example 94. Synthesis of 2-(4-((4-(((lr,4r)-4-morpholinocyclohexyl)amino)-6,7- dihydro-5H-cyclopenta[4,5]thieno[2,3-d]pyrimidin-2-yl)amino)-lH-pyrazol-l-yl)acetate, Example 1-76
Figure imgf000183_0002
[00600] To a solution of intermediate 3.4 (50 mg, 0.13 mmol, 1.00 equiv) and 2-(4-amino- lH-pyrazol-l-yl)acetic acid (36 mg, 0.26 mmol, 2.00 equiv) in isopropanol (5 mL) was added hydrochloric acid (4 M in dioxane, 0.1 mL) at room temperature. The reaction was stirred for 2 h at 140 °C and irradiated in microwave. The resulting mixture was concentrated under vacuum and resulting crude was purified to yield compound 1-76 as a yellow solid. LCMS (ES, m/z) 540 [M+H]+. 1H NMR (300 MHz, d6-OMSO, ppm): δ 8.99 (s, 1H), 7.90 (s, 1H), 7.51 (s, 1H), 7.26-7.06 (m, 1H), 5.79-5.71 (m, 1H), 4.99-4.91 (m, 3H), 4.01-3.99 (m, 1H), 3.77-51 (m, 4H), 3.05-2.98 (m, 2H), 2.83-2.73 (m, 2H), 2.61-2.53 (m, 5H), 2.40-2.35 (m, 2H), 2.01-1.94 (m, 2H), 1.49-1.35 (m, 4H), 1.20-1.15 (m, 6H). [00601] Example 95. Synthesis of 4-(4-((4-(((lr,4r)-4-morpholinocyclohexyl)amino)-6,7- dihydro-5H-cyclopenta [4,5] thieno [2,3-d] pyrimidin-2-yl)amino)- lH-pyrazol- 1- yl)tetrahydro-2H-thiopyran 1,1-dioxide, 1-77
Figure imgf000184_0001
3.4 I-77
[00602] Compound 1-77 was prepared from compound 3.4 and compound 117.6 using procedure described in Example 94. as a white solid. LCMS (ES, m/z): 572 [M+H]+; 1H NMR (300 MHz, d6-OMSO)d 8.92 (IH, s), 7.86 (IH, s), 7.53 (IH, s), 5.69 (IH, s), 4.58-4.51 (IH, m), 4.01-3.97 (IH, m), 3.57-3.56 (4H, m), 3.40-3.36 (2H, d), 3.23-3.19 (2H, d), 2.97-2.95 (2H, d), 2.82-2.80 (2H, d), 2.50-2.49 (4H, m), 2.39-2.17 (7H, m), 2.05-1.89 (4H, m), 1.52-1.44 (4H, m).
[00603] Example 96. Synthesis of N4-((lr,4r)-4-morpholinocyclohexyl)-N2-(lH-pyrazol- 4-yl)-6,7-dihydr o-5H-cyclopenta [4,5] thieno [2,3-d] pyrimi
Figure imgf000184_0002
3.4 1-78
[00604] Synthesis of compound 96.2 ^ Pd/C, H2 ^MH
6 N-— (' J. H.N— (' I
^N
MeOH / R.T
96.1 96.2
[00605] A solution of 4-nitro-lH-pyrazole (200 mg, 1.77 mmol, 1.00 equiv) and 10% Palladium on carbon (40 mg) in methanol (5 mL) was degassed with H2 three times and the reaction mixture was stirred for 1 h at room temperature. The catalyst was filtered out and the filtrate was concentrated under vacuum to give 120 mg (crude) of compound 96.2 as a light yellow solid.
[00606] Synthesis of compound 1-78. Compound 1-78 was prepared from compounds 3.4 and 96.2 using protocol described in Example 94. . LCMS (ES, m/z): 440 [M+H]+; 1H-NMR (300 MHz, DMSO) δ 8.87 (1H, s), 7.82 (1H, brs), 7.52 (1H, brs), 5.71 (1H, d), 4.05-3.89 (1H, m), 3.58 (4H, brs), 3.01-2.95 (2H, m), 2.84-2.79 (2H, m), 2.50 (4H, m), 2.42-2.34 (2H, m), 2.26- 2.18 (1H, m), 2.05 (2H, d), 1.92 (2H, d), 1.47-1.30 (4H, m).
[00607] Example 97. Synthesis of compound N4-((lr,4r)-4-morpholinocyclohexyl)-N2-(l- (tetrahydro-2H-pyran-4-yl)-lH-pyrazol-4-yl)-6,7-dihydro-5H-cyclopenta[4,5]thieno[2,3- d]pyrimidine-2,4-diamine, 1-79
Figure imgf000185_0001
3.1 I-79
[00608] Synthesis of compound 97.2.
Figure imgf000185_0002
97.1 97.2
[00609] A solution of oxan-4-ol, compound 97.1 (7.0 g, 68.54 mmol, 1.00 equiv) in dichloromethane (100 mL) was added triphenylphosphine (27.0 g, 102.94 mmol, 1.50 equiv) and imidazole (7.0 g, 102.82 mmol, 1.50 equiv) at room temperature. This was followed by addition of iodine (18.3 g, 72.05 mmol, 1.05 equiv) in several batches at 0°C. The resulting solution was stirred for 2 h at room temperature under nitrogen. After completion, the reaction was quenched with 5% HC1 solution, extracted with dichloromethane (100 mL x 3). The combined organic layers were washed with brine, dried over anhydrous sodium sulfate and concentrated under vacuum. Crude was purified via flash column chromatography to afford 9.6 g of 4-iodooxane, compound 97.2 as colorless oil.
[00610] Synthesis of compound 97.3
Figure imgf000186_0001
[00611] To a solution of 4-nitro-lH-pyrazole (2.45 g, 21.67 mmol, 1.00 equiv) in freshly distilled DMF (70 mL) was added 4-iodooxane, compound 97.2 (9.2 g, 43.39 mmol, 2.00 equiv) and cesium carbonate (22.2 g, 68.14 mmol, 3.00 equiv) and the resulting solution was stirred for 3 h at 80 °C under nitrogen. The resulting mixture was diluted with 200 mL of water, extracted with CH2CI2 (100 mL x 3) and the organic layers were combined, washed with brine, dried over anhydrous sodium sulfate and concentrated under vacuum. Crude was purified via flash column chromatography to give 2.0 g of 4-nitro-l-(oxan-4-yl)-lH-pyrazole, compound 97.3 as a light yellow solid.
[00612] Synthesis of compound 97.4
Figure imgf000186_0002
[00613] To a solution of 4-nitro-l-(oxan-4-yl)-lH-pyrazole, compound 97.3 (2.0 g, 10.14 mmol, 1.00 equiv) in methanol (40 mL) was added 10% palladium on activated carbon (200 mg) at room temperature under nitrogen. Solution was flushed with H2 (~2 atm) three times and stirred for 2 h at ambient temperature. The catalyst was filtered out and the filtrate was added to HCI (2 M in MeOH) and stirred for another 1 h. The resulting mixture was concentrated under vacuum and crystalized from MeOH/Et20 (1 :5) to give 1.8 g of l-(oxan-4-yl)-lH-pyrazol-4- amine hydrochloride, compound 97.4 as a gray solid.
[00614] Synthesis of compound 1-79. Compound 1-79 was prepared from compound 3.4 and compound 97.4 using proyocol described in Example 94. LCMS (ES, m/z): 524 [M+H]+; 1H NMR (300 MHz, CD3OD): δ 7.94 (s, 1H), 7.60 (s, 1H), 4.42-4.30 (m, 1H), 4.18-4.08 (m, 3H), 3.80-3.70 (m, 4H), 3.65-3.52 (m, 2H), 2.99 (t, 2H), 2.87 (t, 2H), 2.68-2.60 (m, 4H), 2.48 (quintet, 2H), 2.43-2.30 (m, 1H), 2.20-2.15 (m, 2H), 2.12-1.98 (m, 6H), 1.55-1.35 (m, 4H).
[00615] Example 98. Synthesis of compound N4-((lr,4r)-4-morpholinocyclohexyl)-N2-(l- (oxetan-3-yl)- lH-pyrazol-4-yl)-6,7-dihydro-5H-cyclopenta [4,5] thieno [2,3-d] pyrimidine-2,4- diamine, 1-80
Figure imgf000187_0001
3.4 I-80
-(oxetan-3-yl)-lH-pyrazol-4-amine. P κdα//Cυ,, H M22 ( (11 aattmm))
Figure imgf000187_0002
MeOH / R.T H2 '
98.1 98.2 98.3
[00617] Synthesis of compound 98.2. To a solution of 4-nitro-lH-pyrazole (1.13 g, 9.99 mmol, 1.00 equiv) in distilled DMF (15 mL) were added 3-iodooxetane (1.84 g, 10.00 mmol, 1.00 equiv) and CS2CO3 (6.52 g, 20.00 mmol, 2.00 equiv) at room temperature. The resulting mixture was stirred for 1 h at 100 °C in an oil bath. After cooling, the reaction was quenched with water, extracted with 3 x 100 mL of ethyl acetate. Combined organic layers were washed with brine, dried over anhydrous sodium sulfate and concentrated under vacuum. Crude was purified via flash column chromatography to give 1.6 g (95%) of compound 98.2 as a yellow solid.
[00618] Synthesis of compound 98.3. To solution of compound 98.2 (1.60 g, 9.46 mmol, 1.00 equiv) in methanol (50 mL) was added 10% palladium on activated carbon (200 mg) and the mixture was flushed with hydrogen three times and stirred for 5 hours at room temperature. After the starting material was consumed completely, the catalyst was removed by filtration and the filtrate was concentrated in vacuo to afford 1.2 g (91%) of l-(oxetan-3-yl)-lH-pyrazol-4- amine, compound 98.3 as a pink solid.
[00619] Synthesis of compound 1-80. To a 50-mL round-bottom flask containing a solution of compound 3.4 (180 mg, 0.46 mmol, 1.00 equiv) and compound 98.3 (76 mg, 0.55 mmol, 1.20 equiv) in 1,4-dioxane (10 mL) were added CS2CO3 (448 mg, 1.37 mmol, 3.00 equiv), XantPhos (53 mg, 0.20 equiv) and Pd2(dba)3 (47 mg, 0.05 mmol, 0.10 equiv) subsequently under nitrogen. Reaction was heated to reflux for 3 hr. The resulting mixture was concentrated under vacuum and the residue was purified via flash column chromatography to give 121.5 mg (54%) of compound 1-80 as a white solid. LCMS (ES, m/z): 496 [M+H]+; 1H NMR (400 MHz, <¾-DMSO): δ 8.99 (s, 1H), 8.00 (s, 1H), 7.63 (s, 1H), 5.73 (brs, 1H), 5.56 (quintet, 1H), 4.90 (quintet, 4H), 4.10-3.92 (m, 1H), 3.59 (t, 4H), 2.99 (t, 2H), 2.84 (t, 2H), 2.51 (s, 4H), 2.45-2.35 (m, 2H), 2.30- 2.15 (m, 1H), 2.08 (d, 2H), 1.92 (d, 2H), 1.58-1.28 (m, 4H).
[00620] Example 99. Synthesis of compound N2-(l-methyl-lH-pyrazol-4-yl)-N4-((lr,4r)- 4-morpholino-cyclohexyl)-5,6,7,8-tetrahydr obenzo [4,5] thieno [2,3-d] pyrimidine-2,4- diamine, 1-81
Figure imgf000188_0001
Figure imgf000189_0001
[00621] Synthesis of compound 99.3. Compound 99.3 was prepared in a manner analogous to the synthesis of compound 3.3, substituting cyclohexanone for cyclopentanone in the first step. Isolated 4.8 g of a brown solid in 71% yield from cyclohexanone.
[00622] Synthesis of compound 99.4. A mixture of 99.3 (3 g, 13.50 mmol, 1.00 equiv) in 30 mL of phosphoroyl trichloride was heated at 110 °C for 4 h in an oil bath under nitrogen. The resulting mixture was concentrated under vacuum. The residue was diluted with 100 mL of EtOAc and poured into a solution of cooled saturated aqueous sodium bicarbonate. The resulting solution was extracted with 2 x 300 mL of ethyl acetate. The combined organic layers were washed with brine, dried over anhydrous sodium sulfate. The crude product was purified by re- crystallization from EtOAc to yield 99.4 (2.4 g, 69%) as a light yellow solid.
[00623] Synthesis of compound 99.5. Compound 99.5 was prepared in a manner analogous to the synthesis of compound 3.1. Isolated 621 mg a white solid in 19% yield.
Synthesis of comound 1-81. Into a 10-mL vial containing a suspension of compound 99.5 (100 mg, 0.25 mmol, 1.00 equiv) in dry isopropanol (4 mL) was added 1 -methyl- lH-pyrazol-4-amine hydrochloride (50 mg, 0.37 mmol, 1.50 equiv), followed by hydrochloric acid (4M in dioxane, 0.2 mL). Sealed vial was irradiated for 1.5 h at 140°C in microwave. After reaction completion, the resulting mixture was concentrated under vacuum and crude was purified byflash column chromatography give 50 mg of desired compound 1-81. Pure compound 1-81 (38 mg) was obtained by precipitation from Ct^C Vhexane (1 : 10) as a off-white solid. LCMS (ES, m/z): 468 [M+H]+. 1H NMR (300 MHz, DMSO) δ 8.84 (s, 1H), 7.78 (s, 1H), 7.43 (s, 1H), 5.63 (s, 1H), 4.01-3.98 (m, 1H), 3.78 (s, 3H), 3.52-3.61 (m, 4H), 2.90-2.81 (m, 2H) , 2.66-2.59 (m, 2H), 2.29- 2.20 (m, 1H), 3.57 (m, 3H) , 2.14-2.05 (m, 2H), 1.9-1.75 (m, 6H), 1.52-1.30 (m, 4H).
[00624] Example 100. Synthesis of compound N2-(5-methyl-lH-pyrazol-3-yl)-N4- ((1 r,4r)-4-morpholino-cyclohexyl)-6,7-dihydro-5H-cyclopenta [4,5] thieno [2,3-d] pyrimidine- 2,4-diamine, 1-82
Figure imgf000190_0001
3.4 I -82
[00625] To a solution of intermediate 3.4 (150 mg, 0.38 mmol, 1.00 equiv) in 6 mL of anhydrous dioxane were added Cs2C03 (352 mg, 1.08 mmol, 3.00 equiv), 5 -methyl- lH-pyrazol- 3-amine (74 mg, 0.76 mmol, 2.00 equiv), Pd2(dba)3 (16 mg, 0.02 mmol, 0.05 equiv) and BINAP (32 mg, 0.05 mmol, 0.10 equiv) at room temperature. The resulting mixture was degassed three times with nitrogen and stirred for 2 h at 100 °C in an oil bath. The resulting mixture was concentrated under vacuum. The residue was purified via flash column chromatography to afford 67.3 mg (39%) of compound 1-80, as a light yellow solid. LCMS (ES, m/z): 454 [M+H]+, 1H NMR (300 MHz, DMSO) ^ 11.70 (s, 1H), 5.77 (s, 1H), 4.00-3.85 (m, 1H), 3.53 (s, 4H), 3.01- 2.71 (m, 4H), 2.50-2.28 (m, 6H), 2.25-2.01 (m, 4H), 2.08-1.81 (m, 4H) ,1.50-1.27 (m, 4H).
[00626] Example 101. Synthesis of compound N4-((lr,4r)-4-morpholinocyclohexyl)-N2- (5-(trifluoromethyl)- IH-pyr azol-3-yl)-6,7-dihydr o-5H-cyclopenta [4,5] thieno [2,3-d] pyri- midine-2,4-diamine, 1-83
Figure imgf000190_0002
3.4 I-83
[00627] Compound 1-83 was prepared from compound 3.4 and compound 101.1 using procedure described in Example 100. LCMS (ES, m/z): 508 [M+H]+ , 1H NMR (300 MHz, d6- DMSO, ppm) : δ 12.86 (s, 1H), 9.81 (s, 1H), 6.36 (1, 1H), 5.94 (d, 1H), 4.03-3.91 (m, 1H), 3.59- 3.50 (m, 4H), 3.08-2.92 (m, 2H), 2.91-2.78 (m, 2H), 2.58-32 (m, 6H), 2.29-2.17 (m, 1H), 2.05- 1.98 (m, 2H), 1.90-1.84 (m, 2H), 1.53-1.25 (m, 4H). [00628] Example 102. Synthesis of compound N2-(l-(difluoromethyl)-lH-pyrazol-4-yl)- N4-((lr,4r)-4-morpholinocyclohexyl)-6,7-dihydro-5H-cyclopenta[4,5]thieno[2,3- d]pyrimi ine-2 4-diami -84
Figure imgf000191_0001
3.4 I-84
[00629] Compound 1-84 was prepared from compound 3.4 and compound 102.1 using procedure described in Example 94. . LCMS (ES, m/z): 490 [M+H]+. 1H NMR (300 MHz, d6- DMSO, ppm): δ 9.25 (s, 1H), 8.27 (s, 1H), 7.76 (s, 1H), 7.73 (t, 1H), 5.85 (d, 1H), 4.08-3.89 (m, 1H), 3.70-3.57 (m, 4H), 3.14-2.91 (m, 2H), 2.89-2.78 (m, 2H), 2.50-2.27 (m, 7H), 2.07-2.04 (m, 2H), 1.99-1.91 (m, 2H), 1.54-1.34 (m, 4H).
[00630] Example 103. Synthesis of compound N2-(l-(tert-butyl)-lH-pyrazol-4-yl)-N4- ((1 r,4r)-4-morpholino-cyclohexyl)-6,7-dihydro-5H-cyclopenta [4,5] thieno [2,3-d] pyrimidine- 2,4-diamine, 1-
Figure imgf000191_0002
[00631] Compound 1-85 was prepared from compound 3.4 and compound 103.1 using procedure described in Example 94. LCMS (ES, m/z): 496 [M+H]+; 1H NMR (400 MHz, CDCls): δ 7.94 (1H, s), 7.58 (1H, s), 6.54 (1H, s), 4.75 (1H, d), 4.12-3.98 (1H, m), 3.90-3.65 (4H, m), 2.95-2.88 (4H, m), 2.75-2.55 (4H, m), 2.53-2.42 (2H, m), 2.36-2.25 (3H, m), 2.15-1.92 (2H, m), 1.65-1.48 (HH, m), 1.35-1.18 (2H, m). [00632] Example 104. Synthesis of compound N4-((lr,4r)-4-morpholinocyclohexyl)-N2- (l-(piperidin-4-yl)-lH-pyrazol-4-yl)-6,7-dihydro-5H-cyclopenta[4,5]thieno[2,3- d]pyrimidine-2,4-diamine, 1-86
Figure imgf000192_0001
I-86
[00633] Synthesis of compound 104.1 To a 50-mL round-bottom flask under nitrogen, were added intermediate 3.4 (111 mg, 0.28 mmol, 1.00 equiv), 6-(4-amino-lH-pyrazol-l-yl)-l-tert- butyl-l [3],3-oxazocan-2-one (90 mg, 0.34 mmol, 1.20 equiv), CS2CO3 (183 mg, 0.56 mmol, 2.00 equiv), Pd2(dba)3-CHC13 (28 mg, 0.03 mmol, 0.10 equiv) and XantPhos (34 mg, 0.06 mmol, 0.20 equiv) in dioxane (15 mL). The resulting mixture was stirred for 2 h at 100 °C in an oil bath. After cooling, the resulting mixture was diluted with water, extracted with EtOAc, organic layers were dried over sodium sulfate and solvents under vacuum. The residue was purified via flash column chromatography to provide 138 mg (78%) of compound 104.1.
[00634] Synthesis of compound 1-86. To a solution of compound 104.1 (173 mg, 0.28 mmol, 1.00 equiv) in 10 mL of CH2C12 was added 1.0 mL of CF3COOH at 0 °C. The resulting solution was stirred for 1 h at room temperature and concentrated under vacuum. The residue was diluted with CH2C12, washed with aqueous NaHC03 and brine and concentrated under reduced pressure. The residue was purified by flash column chromatography) to give 64.8 mg (44.7%) of compound 1-83 as a white solid. LCMS (ES, m/z): 523 [M+H]+; 1H-NMR (400 MHz, <¾-DMSO, ppm) δ 8.92 (brs, 1H), 7.88 (s, 1H), 7.44 (s, 1H), 5.75 (brs, 1H), 4.20-4.10 (m, 1H), 4.06-3.94 (m, 1H), 3.62-3.54 (m, 4H), 3.15-3.05 (d, 2H), 2.98 (t, 2H), 2.84 (t, 2H), 2.65-2.55 (m, 2H), 2.51 (brs, 4H), 2.45-2.30 (m, 2H), 2.35-2.20 (m, 1H), 2.06 (d, 2H), 1.98-1.85 (m, 4H), 1.85-1.75 (m, 2H), 1.55-1.25 (m ,4H).
[00635] Example 105. Synthesis of compound N2-(l-isopropyl-lH-pyrazol-4-yl)-N4-
((1 r,4r)-4-morpholino-cyclohexyl)-5,6,7,8-tetrahydrobenzo [4,5] thieno [2,3-d] pyrimidine-2,4- diamine, 1-87
Figure imgf000193_0001
99.5 1-87
[00636] Compound 1-87 was prepared from compopund 99.5 and compound 105.1 using protocol described in Example 94. LCMS (ES, m/z): 496 [M+H]+. 1H NMR003065 (300 MHz, CD3OD): δ 7.85 (s, 1H), 7.60 (s, 1H), 4.48 (quintet, 1H), 4.15-4.00 (m, 1H), 3.73 (t, 4H), 2.86 (brs, 4H), 2.70 (m, 2H), 2.65 (t, 4H), 2.45-2.30 (m, 1H), 2.30-2.20 (m, 2H), 2.15-2.06 (m, 2H), 1.89 (brs, 4H), 1.52-1.35 (m, 10H).
[00637] Example 106. Synthesis of compound 2-(2-(4-((4-(((lr,4r)-4- morpholinocyclohexyl)amino)-6,7-dihydro-5H-cyclopenta [4,5] thieno [2,3-d] pyrimidin-2- yl)amino)-lH-pyrazol-l-yl)ethoxy)ethanol, 1-88
Figure imgf000193_0002
[00638] Compound 1-88 was prepared from compound 3.4 and compound 106.1 using protocol described in Example 94. LCMS (ES, m/z): 528 [M+H]+; 1H NMR (300 MHz, CD3OD) δ 7.94 (s, 1H), 7.61 (s, 1H), 4.28 (t, 2H), 4.15-4.02 (m, 1H), 3.84 (t, 2H), 3.73 (t, 4H), 3.64 (t, 2H), 3.51 (t, 2H), 2.98 (t, 2H), 2.89 (t, 2H), 2.65 (t, 4H), 2.49 (quintet, 2H), 2.40-2.28 (m, 1H), 2.25 (d, 2H), 2.09 (d, 2H), 1.58-1.35 (m, 4H).
[00639] Example 107. Synthesis of N2-(l-methyl-lH-pyrazol-4-yl)-N4-((lr,4r)-4- morpholino-cyclohexyl)-6,7,8,9-tetrahydro-5H-cyclohepta [4,5] thieno [2,3-d] pyrimidine-2,4- diamine compound 1-89
Figure imgf000194_0001
107.5 I-89
[00640] Synthesis of compound 107.2. To a solution of cycloheptanone (10.8 g, 96.28 mmol, 1.00 equiv) in ethanol (50 mL) was added ethyl 2-cyanoacetate (12.54 g, 1 10.86 mmol, 1.15 equiv), diethylamine (13.04 g, 178.30 mmol, 1.80 equiv) and sufur (3.42 g, 106.67 mmol, 1.10 equiv) at room temperature. The resulting mixture was stirred for 14 h at 50°C in an oil bath under nitrogen. The reaction mixture was cooled to room temperature, diluted with 100 mL of ice-cooled water, extracted with 2 x 150 mL of ethyl acetate. The organic phase was washed with brine, dried over anhydrous sodium sulfate. Solids were filtered off and the filtrate was concentrated under vacuum. Crude was purified via flash column chromatography to give 10.6 g (44%) of compound 107.2 as a yellow solid. LCMS (ES, m/z): 240 [M+H]+.
[00641] Synthesis of compound 107.3. To a solution of compound 107.2 (1.0 g, 4.18 mmol, 1.00 equiv) in dry CH2C12 (50 mL) was cooled down to -60°C under nitrogen. Solution of [(chlorosulfonyl)imino]methanone (700 mg, 4.95 mmol, 1.20 equiv) in 5 mL of CH2C12 was added dropwise at this temperature. After addition, the reaction mixture was stirred for another 30 minutes until starting material was consumed. Solvent was removed under reduced pressure and the residue was diluted with 10 M aqueous sodium hydroxide (100 mL) and stirred for 1.5 h at room temperature. Resulting mixture was allowed to react, with stirring, for an additional 30 minutes while the temperature was maintained at 75°C in an oil bath. The reaction mixture was cooled to room temperature and the pH value of the mixture was adjusted to ~1 with Cone, hydrochloric acid. Resulting solids were collected by filtration and the cake was washed with water, dried in an oven at 50°C under reduced pressure to give 900 mg (crude) of compound 107.3 as a pink solid. LCMS (ES, m/z): 237 [M+H]+.
[00642] Synthesis of compound 107.4. A solution of compound 107.3 (840 mg, 3.55 mmol, 1.00 equiv) in 20 mL of phosphoroyl trichloride was stirred for 24 h at 110°C in an oil bath under nitrogen. After completion, the reaction mixture was cooled to room temperature and concentrated under reduced pressure. Resulting residue was diluted with EtOAc and poured into saturated aqueous NaHC03, extracted with EtOAc, washed with brine, dried over sodium sulfate. Solvents were removed under vacuum. Crude was purified using flash column chromatography to give 190 mg of desired compound 107.4 as a white solid. LCMS (ES, m/z) 273 and 275 [M+H]+.
[00643] Synthesis of compound 107.5. To a solution of compound 107.4 (190 mg, 0.70 mmol, 1.00 equiv) in anhydrous MeCN (7 mL) were added trans-4-(morpholin-4-yl)cyclohexan- 1-amine (154 mg, 0.84 mmol, 1.00 equiv) and potassium carbonate (480 mg, 5.00 equiv). Reaction was stirred for 3 days at 85°C in an oil bath. The resulting mixture was concentrated under vacuum and the residue was purified via flash colyumn chromatography to give 304 mg of compound 107.5 as a yellow solid. LCMS (ES, m/z): 421 and 423 [M+H]+.
[00644] Synthesis of compound 1-89. To a solution of compound 107.5 (121 mg, 0.29 mmol, 1.00 equiv) in iso-propanol (5 mL) was added 1 -methyl- lH-pyrazol-4-amine (40 mg, 0.41 mmol, 1.40 equiv) . Reaction was irradiated in microwave for 2 h at 140°C. Upon completion of the reaction, solvent was removed under vacuum. Resulting crude residue was purified via flash column chromatography to afford 29.1 mg of N2-(l -methyl- 1 H-pyrazol-4-yl)- N4-((lr,4r)-4-morpho-lino-cyclohexyl)-6,7,8,9-tetrahydro-5H-cyclohepta[4,5]thieno[2,3- d]pyrimidine-2,4-diamine, 1-89 as a white solid. LCMS (ES, m/z): 482 [M+H]+; 1H NMR (300 MHz, CD3OD) δ 7.82 (s, 1H), 7.55 (s, 1H), 4.15-4.02 (m, 1H), 3.87 (s, 3H), 3.73 (t, 4H), 3.12- 2.95 (m, 2H), 2.88-1.78 (m, 2H), 2.70-2.58 (m, 4H), 2.40-2.21 (m, 3H), 2.18-2.08 (m, 2H), 1.95- 1.75 (m, 6H), 1.55-1.35 (m, 4H). [00645] Example 108. Synthesis of N2-(l-ethyl-lH-pyrazol-4-yl)-N4-((lr,4r)-4- morpholino-cyclohexyl)-6,8-dihydro-5H-pyrano [4',3 ' :4,5] thieno [2,3-d] pyrimidine-2,4- diamine, 1-90
Figure imgf000196_0001
17.5 I-90
[00646] To a solution of compound 17.5 (100 mg, 0.24 mmol, 1.00 equiv) in 1,4-dioxane (10 mL) was added 1 -ethyl- lH-pyrazol-4-amine (33 mg, 0.30 mmol, 1.20 equiv), Cs2C03 (239 mg, 0.73 mmol, 3.00 equiv), Pd2(dba)3-CHC13 (25 mg, 0.02 mmol, 0.10 equiv) and XantPhos (28 mg, 0.05 mmol, 0.20 equiv). The resulting mixture was heated to reflux for 3 h under nitrogen. Upon completion of the reaction solvents were removed under vacuuo . Crude was purified via flash column chromatography to give 32.2 mg of compound 1-90 of a yellow solid. LCMS (ES, m/z): 484 [M+H]+; 1H NMR (300 MHz, d6-OMSO, ppm): δ 1.32-1.47(m, 7H), 1.88-1.92 (d, 2H), 2.06- 2.10 (m, 2H), 2.23-2.27 (m, 1H), 2.95 (m, 2H), 3.55-3.59 (m, 4H), 3.89-3.93 (m, 3H), 4.03-4.11 (q, 2H), 4.66 (s, 2H), 5.68-5.71 (m, 1H), 7.45 (s, 1H), 7.82 (s, 1H), 8.93 (s, 1H).
[00647] Example 109. Synthesis of N4-((lr,4r)-4-morpholinocyclohexyl)-N2-(l- (tetrahydro-2H-pyr an-4-yl)- IH-pyr azol-4-yl)-6,8-dihydro-5H-pyrano [4 ',3 ' : 4,5] thieno [2,3- d]pyrimidine-2,4-diamine, compound 1-91
Figure imgf000196_0002
17.5 1-91
[00648] Compound 1-91 was prepared from compound 17.5 and compound 97.4 using procedure described in Example 94. . LCMS (ES, m/z): 540 [M+H]+; 1H NMR (300 MHz, d6- DMSO): δ 8.95 (s, IH), 7.88 (s, IH), 7.46 (s, IH), 5.78-5.62 (m, IH), 4.75 (s, 2H), 4.40-4.20 (m, IH), 4.10-3.85 (m, 5H), 3.57 (brs, 4H), 3.43 (td, 2H), 2.95 (s, 2H), 2.50 (s, 4H), 2.28-2.18 (m, IH), 2.15-2.02 (m, 2H), 2.01-1.79 (m, 6H), 1.65-1.25 (m, 4H).
[00649] Example 110. Synthesis of N4-((lr,4r)-4-morpholinocyclohexyl)-N2-(l-(oxetan-3- yl)-lH-pyrazol-4-yl)-6,8-dihydro-5H-pyrano [4',3 ' :4,5] thieno [2,3-d] pyrimidine-2,4-diamine, compound 1-92
Figure imgf000197_0001
[00650] Compound 1-92 was prepared from compound 17.5 and compound 98.3 using protocol described in Example 108. LCMS (ES, m/z): 512 [M+H]+; 1H-NMR (300 MHz, d6- OMSO, ppm): δ 9.01(1H, s), 7.97 (IH, s), 7.62 (IH, s), 5.70 (IH, d), 5.53 (IH, quintet), 4.92- 4.87 (4H, m), 4.66 (2H, s), 4.10-3.93 (IH, m), 3.91 (2H, t), 3.62-3.56 (4H, m), 2.96 (2H, s), 2.50 (4H, brs), 2.30-2.18 (IH, m), 2.15-2.06 (2H, m), 1.88 (2H, d), 1.55-1.28 (4H, m).
[00651] Example 111. Synthesis 2-[(3R)-12-[[trans-4-[morpholin-4-yl]cyclohexyl]oxy]-7- thia-9,ll-diazatricyclo[6.4.0.0A[2,6]]dodeca-l(8),2(6),9,ll-tetraen-3-yl]acetamide-d8, compound 1-93
Figure imgf000197_0002
THF / 0 °C to R.T o THF / overnight D ,X D MgS04 / dioxane / 100°C
111.4 111.5
Figure imgf000198_0001
[00652] Synthesis of compound 112.2. To a solution of compound 111.1 (5.0 g, 37.29 mmol, 1.00 equiv) in methanol (100 mL) was added sulfuric acid (0.5 mL) and the resulting solution was stirred overnight at 80°C. Reaction was concentrated under vacuo and the residue was diluted with 100 mL of CH2CI2, washed with 2 x 50 mL of 2 M sodium bicarbonate followed brine. Organic layers were combined, dried over sodium sulfate and solvents were removed under reduced pressure to give 3.1 g (51%) of compound 111.2 as a light yellow solid. 1H NMR (300 MHz, CDC13) δ4.22 (s, 4H), 3.74 (s, 6H).
[00653] Synthesis of compound 111.3. Sodium metal (461 mg, 20.05 mmol, 0.50 equiv) was dissolved in dry MeOD (20 mL) and compound 111.2 (6.5 g, 40.09 mmol, 1.00 equiv) was added. Reaction was stirred for 24 hrs under nitrogen at room temperature. The resulting solution was evaporated and a fresh portion of MeOD was added and the mixture was stirred for another 24 hrs. Replacement of the 'used' MeOD by 'fresh' one was repeated until the proton peak in 1H NMR completely disappeared, usually requiring 3 cycles. After concentrated in vacuo, the residue was diluted with 20 mL of CH2CI2, washed with 2 x 10 mL of D20 and dried over anhydrous sodium sulfate and concentrated under vacuum to give 2.8 g (42%) of compound 111.3 as a yellow oil. 1H NMR (400 MHz, CDC13) £3.80 (s, 6H).
[00654] Synthesis of compound 111.4. A solution of compound 111.3 (2.0 g, 12.04 mmol, 1.00 equiv) in dry THF (50 mL) was added L1AID4 (964 mg, 22.96 mmol, 1.91 equiv) at 0 °C under nitrogen. Reaction was stirred for 4 hours at room temperature. The resulting solution was diluted with 100 mL of EtOAc, quenched with NaSO4 10H2O. The solids were filtered out and washed with 2 x 10 mL of EtOAc. The filtrate was concentrated under vacuum and crude was purified using flash column chromatography to give 530 mg of compound 111.4 as a light yellow oil.
[00655] Synthesis of compound 111.5. To a aqueous solution of NaOH (650 mg) was added compound 111.4 (530 mg, 4.64 mmol, 1.00 equiv). Reaction flask was cooled to 0 °C and tosylchloride (2.23 g, 11.7 mmol) in THF (25 ml) was added dropwise over a period of 1 h. Resulting mixture was stirred overnight at room temperature. The pH value of the solution was adjusted to 5 with hydrogen chloride (1 mol/L) and solution was extracted with 3 x 100 mL of ethyl acetate. The combined organic layers were dried over sodium sulfate and concentrated under vacuum. Crude was purified using flash column chromatography to furnish compound
42.5 as an off-white solid. 1H NMR (300 MHz, CDC13) ^7.78 (d, 4H), 7.34 (d, 4H), 2.45 (s, 6H).
[00656] Synthesis of compound 111.6. A solution of compound 111.5 (1 g, 2.37 mmol, 1.00 equiv) and trans-4-aminocyclohexan-l-ol (1.36 g, 11.81 mmol, 4.99 equiv) in 1,4-dioxane (30 mL) was added magnesium sulfate (1 g) and the resulting mixture was stirred overnight at 100°C under nitrogen. Reaction was cooled to ambient temperature, the solids were filtered out and filter cake was washed with EtOAc (3 x 50 mL). Filtrate was concentrated under vacuum and the residue was purified using flash column chromatography to afford 430 mg (94%) of compound
42.6 as a yellow solid. LCMS (ES, m/z): 194 [M+H]+.
[00657] Synthesis of compound 111.8. Compound 111.6 (400 mg, 2.07 mmol, 1.00 equiv) was treated with NaHMDS (3.1 mL, 2 M in THF) in 20 mL of distilled THF at 0 °C for 10 min under nitrogen. A solution of compound 111.7 (555 mg, 2.07 mmol, 1.00 equiv) in 10 mL of THF was added dropwise via syringe at this temperature and the reaction was stirred for 1 h. The resulting solution was diluted with 30 mL of H20 and the pH value of the solution was adjusted to 5.0 using 1 M HC1 The resulting solution was extracted with CH2C12 , organic layers were combined and dried over anhydrous sodium sulfate. Solvents were removed in vacuo to give 1.5 g of compound 111.8 as a yellow solid. LCMS (ES, m/z): 426 [M+H]+.
[00658] Synthesis of compound 1-93. To a solution of compound 111.8 (1.5 g, 3.52 mmol, 1.00 equiv) in anhydrous CH2C12 (40 mL) was added oxalic dichloride (1.79 g, 14.10 mmol, 4.00 equiv) at 0°C, followed by addition of DMF (0.1 mL) under nitrogen. Reaction was stirred for 30 minuntes at room temperature and concentrated under vacuum to give 1.8 g crude acyl chloride. Into a 250-mL round-bottom flask charged with ammonia (100 mL) and 100 mL of CH2CI2 was added dropwise a solution of acyl chloride (1.8 g, crude) in dry CH2CI2 (20 mL). After addition, the resulting solution was diluted with 50 mL of H2O, extracted with 3 x 100 mL of CH2CI2. The combined organic layers were dried over sodium sulfate and concentrated under vacuum.Crude was purified using flash column chromatography to furnish 480 mg of compound 1-93 as a white solid. LCMS (ES, m/z): 425 [M+H]+; 1H NMR (400 MHz, CD3OD) ^8.49 (1H, s), 5.32- 5.22 (1H, m), 3.89-3.79 (1H, m), 3.17-3.10 (1H, m), 3.04-2.97 (2H, m), 2.78-2.71 (1H, m), 2.40- 2.22 (5H, m), 2.10 (2H, d), 1.72-1.58 (2H, m), 1.56-1.45 (2H, m).
[00659] Example 112. Synthesis of 6,6-dimethyl-N2-(l-methyl-lH-pyrazol-4-yl)-N4- ((1 r ,4r)-4-morpholinocyclohexyl)-6,8-dihydro-5H-pyrano [4 ',3 ' : 4,5] thieno [2,3-d] pyrimidine- 2,4-diamine, compound 1-94
Figure imgf000200_0001
1-94 [00660] Synthesis of compound 112.2. Into a 100-mL 3-necked round-bottom flask under nitrogen were added compound 112.1 (3 g, 23.41 mmol, 1.00 equiv), ethyl 2-cyanoacetate (2.9 g, 25.64 mmol, 1.10 equiv), S (3.0 g) and morpholine (0.75 g) in 50 mL of dry ethanol. The resulting mixture was stirred overnight at 50 °C. Upon completion of the reaction solvent was removed under vacuum and crude was purified via flash column chromatography to afford 5.6 g (94%) of compound 112.2 as a yellow solid.
[00661] Synthesis of compound 112.3. Toa solution of compound 112.2 (5.6 g, 21.93 mmol, 1.00 equiv) in anhydrous CH2CI2 (100 mL) cooled to -60 °C was added dropwise sulfurisocyanatidic chloride (4.6 g) via syringe under nitrogen. The resulting mixture was stirred for 30 min at this temperature. After reaction completion, solvent was removed in vacuo and the residue was dissolved in water (150 mL). Sodium hydroxide (5.0 g, 125.00 mmol, 3.49 equiv) was added and the mixture was stirred for 30 min at 85 °C. pH value of the solution was adjusted to 1.0 using concentrated HCl and resulting solids were collected by filtration. Filter cake was washed with water and dried in an oven at 50 °C under reduced pressure to give 5.0 g (55%) of compound 112.3 as a red solid.
[00662] Synthesis of compound 112.4. Compound 112.3 (5 g, 19.82 mmol, 1.00 equiv) in POCI3 (30 mL) was stirred for 5 h at 110 °C under nitrogen. After the consumption of starting materials, excess POCI3 was removed under vacuum. Crude was dissolved in 500 mL of EtOAc, washed with saturated aqueous sodium bicarbonate, brine and dried over sodium sulfate. Solvents were removed under vacuum. Crude was purified via flash column chromatography to give 1.6 g of compound 112.4 as a white solid.
[00663] Synthesis of compound 112. 5. A mixture of compound 112.4 (500 mg, 1.73 mmol, 1.00 equiv), trans-4-(morpholin-4-yl)cyclohexan-l -amine dihydrochloride (667.4 mg, 2.60 mmol, 1.5 equiv) and potassium carbonate (958 mg, 6.93 mmol, 4.00 equiv) in 30 mL of CH3CN (15 mL) was heated for two days at 85 °C. The resulting mixture was concentrated under vacuum and the crude was purified via flash column chromatography to furnish 700 mg (93%) of compound 112.5 as a white solid.
[00664] Synthesis of compound 1-94. To a 20-mL sealed microwave tube containing compound 112.5 (120 mg, 0.27 mmol, 1.00 equiv), 1 -methyl- lH-pyrazol-4-amine hydrochloride (77.7 mg, 0.58 mmol, 2.10 equiv) in 5 mL of isopropanol was added 0.02 mL of hydrochloric acid (4 M in dioxane) at room temperature. The reaction mixture was heated on microwave for 2.5 h at 140 °C and resulting solids were collected by filtration. Crude collected was purified via flash column chromatography to give 72.2 mg (53%) of compound 1-94 as a off- white solid. LCMS (ES, m/z): 498 [M+H]+. 1H NMR(400MHz, <¾-DMSO): δ 8.94 (s, IH), 7.80 (s, IH), 7.46 (s, IH), 5.75-5.60 (s, IH), 4.65 (s, 2H), 34.08-3.95 (m, IH), 3.80 (s, 3H), 3.59 (t, 4H), 2.85 (s, 2H), 2.52 (s, 4H), 2.30-2.22 (m, IH), 2.10 (d, 2H), 1.92 (d, 2H), 1.55-1.32 (m, 4H), 1.28 (s, 6H).
[00665] Example 113. Synthesis of 4-(4-((6,6-dimethyl-4-(((lr,4r)-4-morpholino- cyclohexyl)amino)-6,8-dihydro-5H-pyrano [4 ',3 ' : 4,5] thieno [2,3-d] pyrimidin-2-yl)amino)- lH- razol-l-yl)tetrahydro-2H-thiopyran 1,1-dioxide, compound 1-95
Figure imgf000202_0001
112.5 I-95
[00666] Compound 1-95 was prepared from compound 112.5 and compound 117.6 using protocol described in Example 94 . LCMS (ES, m/z) : 616 [M+H]+. 1H NMR (300 MHz, d6- DMSO): δ 8.95 (IH, s), 7.84 (IH, s), 7.55 (IH, s), 5.67 (IH, s), 4.63 (3H, s), 4.04-3.98 (IH, m), 3.56 (4H, s), 3.31 (IH, s), 3.23 (2H, s), 2.83 (2H, s), 2.51 - 2.49 (2H, m), 2.48 - 2.45 (2H, m), 2.41 - 2.30 (4H, m), 2.27-2.17 (IH, m), 2.08-2.04 (2H, d), 1.93-1.90 (2H, d), 1.48-1.45 (4H, m), 1.26 (7H, s).
[00667] Example 114. Synthesis of 2-(4-((4-(((lr,4r)-4-morpholinocyclohexyl)amino)-6,7- dihydro-5H-cyclopenta[4,5]thieno[2,3-d]pyrimidin-2-yl)amino)-lH-pyrazol-l-yl)ethanol, compound 1-96
Figure imgf000203_0001
3.4 1-96
[00668] Compound 1-96 was prepared from compound 3.4 and compound 114.1 using protocol described in Example 94. LCMS (ES, m/z): 484 [M+H]+; 1H NMR (300 MHz, CD3OD) δ 7.96 (s, 1H), 7.60 (s, 1H), 4.21 (t, 2H), 4.18-4.02 (m, 1H), 3.89 (t, 2H), 3.74 (t, 4H), 2.99 (t, 2H), 2.90 (t, 2H), 2.68 (brs, 4H), 2.50 (quintet, 2H), 2.42-2.32 (m, 1H), 2.26 (d, 2H), 2.06 (d, 2H), 1.62-1.35 (m, 4H).
[00669] Example 115. Synthesis of compound N4-((lr,4r)-4-(4,4-difluoropiperidin-l- yl)cyclohexyl)-N2-(l-ethyl-lH-pyrazol-4-yl)-6,7-dihydro-5H-cyclopenta[4,5]thieno[2,3- d]pyrimidine-2,4-diamine, 1-97
Figure imgf000203_0002
[00670] Synthesis of compound 115.2. A mixture of 4,4-difluoropiperidine hydrochloride (1.0 g, 6.35 mmol, 1.00 equiv) in methanol (100 mL) was added tert-butyl N-(4- oxocyclohexyl)carbamate (1.63 g, 7.64 mmol, 1.20 equiv), compound 115.1 and NaBH(OAc)3 (4.72 g, 22.26 mmol, 3.50 equiv) at room temperature and stirred overnight at 40 °C under nitrogen. The resulting mixture was concentrated under vacuum and the residue was diluted with 50 mL of water, extracted with 3 x 50 mL of ethyl acetate. The combined organic layers were dried over sodium sulfate and concentrated under vacuum. Crude was purified via flash column chromatography to give 1.15 g (57%) of compound 115.2 as a yellow solid.
[00671] Synthesis of compound 115.3. To a solution of compound 115.2 (1.15 g, 3.61 mmol, 1.00 equiv) in dichloromethane (6 mL) was added trifluoroacetic acid (6 mL) at 0 °C. Reaction was stirred overnight at room temperature. The resulting mixture was concentrated under vacuum to give 1.2 g (crude) of compound 115.3 as a yellow oil.
[00672] Synthesis of compound 115.4. To a solution of compound 3.3 (150 mg, 0.61 mmol, 1.00 equiv) in acetonitrile (10 mL) was added compound 115.3 (350 mg, 1.11 mmol, 1.82 equiv) and potassium carbonate (253 mg, 1.83 mmol, 3.00 equiv) at room temperature. Reaction was stirred overnight at 80 °C under nitrogen. After cooling, solvents were removed under vacuum and crude was purified via flash column chromatography to give 280 mg of compound 115.4 as a yellow solid.
[00673] Separation of compounds 115.5 and 115.6. Compounds 115.5 and 115.6 were obtained by separation on preparative HPLC of compound 115.4 (280 mg, 0.66 mmol, 1.00 equiv). Compound 115.5 was obtained in 21% yield (60 mg) as a off-white solid. Compound 70.6 was obtained in 14% yield (40 mg) a off-white solid.
[00674] Analytical data for compound 115.5: 1H NMR (300 MHz, DMSO d6): δ 6.39 (d, 1H), 4.05-3.85 (m, 1H), 3.05 (t, 2H), 2.91 (t, 2H), 2.65-2.58 (m, 4H), 2.48-2.32 (m, 3H), 1.98-1.70 (m, 8H), 1.59-1.30 (m, 4H).
[00675] Analytical data for compound 115.6: 1H NMR (300 MHz, DMSO d6): δ 6.19 (d, 1H), 4.20-4.08 (m, 1H), 3.11 (t, 2H), 2.94 (t, 2H), 2.60 (brs, 4H), 2.46-2.30 (m, 3H), 2.02-1.82 (m, 6H), 1.71-1.50 (m, 6H). [00676] Synthesis of compound 1-97. To a solution of compound 115.5 (55 mg, 0.13 mmol, 1.00 equiv) in isopropanol (2 mL) was added 0.05 mL of hydrochloric acid (4 M in dioxane) followed by 1 -ethyl- lH-pyrazol-4-amine hydrochloride (23 mg, 0.16 mmol, 1.21 equiv) at room temperature. Reaction mixture was irradiated in microwave for 2 h at 140 °C. Upon completion solvent was removed under vacuum and crude was purified via flash column chromatography. Additional purification was done via trituration with Cf^CVhexanes to give 19.6 mg (30%) of N4-(( lr,4r)-4-(4,4-difluoropiperidin- 1 -yl)cyclohexyl)-N2-( 1 -ethyl- 1 H-pyrazol-4-yl)-6,7-dihydro- 5H-cyclopenta[4,5]thieno[2,3-d]pyrimidine-2,4-diamine, 1-97 as a off-white solid. LCMS (ES, m/z): 502 [M+H]+, 1H NMR (400 MHz, CDC13): δ 7.86 (1H, s), 7.52 (1H, s), 6.56 (1H, s), 4.76 (1H, d), 4.17 (2H, q), 4.08-3.95 (1H, m), 2.93 (4H, brs), 2.76 (4H, brs), 2.62-2.45 (3H, m), 2.31 (2H, d), 2.20-1.92 (6H, m), 1.60-1.45 (5H, m), 1.35-1.18 (4H, m).
[00677] Example 116. Synthesis of (R)-3-((R)-4-(((lr,4R)-4-morpholinocyclohexyl)oxy)- 6,7-dihydro-5H-cyclopenta [4,5] thieno [2,3-d] pyrimidin-5-yl)propane-l ,2-diol, 1-98
Figure imgf000205_0001
49.3 116.1 116.2
Chiral separation
Figure imgf000205_0002
[00678] Synthesis of compound 116.1. To a mixture of Ph3PCH3Br (650 mg) in 15 mL of anhydrous THF was added dropwise 1.3 mL of NaHMDS (1 M in THF) at 0 °C under nitrogen. After stirring for 1 h, a solution of compound 49.3 (500 mg, 1.25 mmol, 1.00 equiv) in 5 mL of THF was added and the reaction was stirred for 3 h at 0 °C. The reaction was then quenched with water, extracted with 3 x 100 mL of ethyl acetate. The combined organic layers were washed with brine, dried over sodium sulfate and concentrated under vacuum. The crude was purified using flash column chromatography to give 300 mg (60%) of compound 116.1 as a white solid.
[00679] Synthesis of compound 116.2. To a solution of compound 116.1 (240 mg, 0.60 mmol, 1.00 equiv) in tert-butanol / water (8/0.8 mL) was added NMO (210 mg, 1.79 mmol, 2.98 equiv) and Os04 (20 mg) at room temperature. Reaction was stirred for 3 h at room temperature and quenched with Na2S03 (aq.), extracted with 3 x 100 mL of ethyl acetate. The combined organic layers were washed with brine and dried over sodium sulfate and concentrated under vacuum. Crude was purified using flash column chromatography to give 100 mg (38%) of compound 116.2 as a white solid.
[00680] Synthesis of compound 1-98. Racemic compound 116.2 (100 mg, 0.23 mmol, 1.00 equiv) was resolved using preparative chiral HPLC to furnish 21.3 mg (21%) of compound 116.3 and 19.3 mg (19%) of compound 1-98 as a white solids.
[00681] Analysis for compound 1-98: LCMS (ES, m/z): 434 [M+H]+; 1H NMR (300 MHz, CD3OD, ppm): δ 8.46 (1H, s), 5.38-5.18 (1H, m), 3.89-3.68 (5H, m), 3.62 (1H, dd), 3.55-3.33 (2H, m), 3.20-3.06 (1H, m), 3.05-2.88 (1H, m), 2.78-2.58 (5H, m), 2.55-2.22 (4H, m), 2.21-2.02 (3H, m), 1.77-1.58 (2H, m), 1.56-1.40 (3H, m).
[00682] Example 117. Synthesis of 4-(4-((4-(((lr,4r)-4-morpholinocyclohexyl)amino)-6,8- dihydro-5H-pyrano[4',3':4,5]thieno[2,3-d]pyrimidin-2-yl)amino)-lH-pyrazol-l- yl)tetrahydro-2H-thiopyran 1,1-dioxide, 1-99
Figure imgf000206_0001
117.1 117.2 117.3
Figure imgf000206_0002
Figure imgf000207_0001
[00683] Synthesis of compound 117.2. To a solution of thian-4-one (1.0 g, 8.61 mmol, 1.00 equiv) in methanol (10 mL) was added NaBH4 (655 mg, 17.31 mmol, 2.00 equiv) portionwise with stirring at 0 °C and the reaction was stirred for 1 hour at room temperature. The resulting mixture was concentrated under vacuum and the residue was diluted with H20, extracted with 3 x 30 mL of dichloromethane. The combined organic layers were washed with brine, dried over sodium sulfate and concentrated under vacuum to give 0.95 g (crude) of compound 117.2 as a white solid.
[00684] Synthesis of compound 117.3. To solution of compound 117.2 (1 g, 8.46 mmol, 1.00 equiv), PPh3 (3.33 g, 12.70 mmol, 1.50 equiv) and lH-imidazole (863 mg, 12.68 mmol, 1.50 equiv) in dichloromethane (10 mL) was added I2 (2.25 g, 1.05 equiv) at 0 °C under nitrogen. Reacttion was stirred for 1 hour at room temperature and diluted with water, extracted with 3 x 30 mL of dichloromethane. The combined organic layers were washed with water and concentrated under vacuum. The crude product was re-crystallized give 1.0 g (crude) of compound 117.3 as light yellow oil.
[00685] Synthesis of compound 117.4. To a solution of compound 117.3 (1.0 g, crude) and 4-nitro-lH-pyrazole (582 mg, 5.15 mmol, 1.00 equiv) in anhydrous DMF (20 mL) was added Cs2C03 (5.07 g, 3.00 equiv). Reaction was stirred for 3 hours at 85 °C under nitrogen. The resulting solution was diluted with H20, extracted with 3 x 80 mL of ethyl acetate. The combined organic layers were washed with brine, dried over sodium sulfate and concentrated under vacuum. Crude was purified using flash column chromatography to give 200 mg (18%) of compound 117.4 as a white solid.
[00686] Synthesis of compound 117.5. To a solution of compound 117.4 (200 mg, 0.94 mmol, 1.00 equiv) in chloroform (10 mL) was added m-CPBA (578 mg, 3.35 mmol, 2.50 equiv) in portions with stirring at 0 °C. Reaction was stirred for 1 h at room temperature and quenched with saturated aqueous Na2S03 and extracted with 3 x 30 mL of ethyl acetate. The combined organic layers were washed with sodium bicarbonate (aq.) and brine. Organic layer was dried over sodium sulfate and concentrated under vacuum. Crude was purified using flash column chromatography to afford 150 mg (65%) of compound 117.5 as a white solid.
[00687] Synthesis of compound 117.6. To a solution of compound 117.5 (130 mg, 0.53 mmol, 1.00 equiv) in methanol (10 mL) was added 10% Palladium on activated carbon (13 mg) under nitrogen. Suspension was degassed with H2 gas three times and stirred for 1 hr at room temperature. The solids were filtered out and the filtrate was concentrated under vacuum to afford 90 mg (79%) of compound 117.6 as a pink solid.
[00688] Synthesis of compound 1-99. Compound 1-99 was prepared from compound 17.5 and compound 117.6 using protocol described in Example 94. LCMS (ES, m/z): 588 [M+H]+; 1H NMR (300 MHz, d6-OMSO): δ 8.96 (1H, s), 7.85 (1H, s), 7.55 (1H, s), 5.72-5.58 (1H, m), 4.66 (2H, s), 4.59-4.49 (1H, m), 3.93-3.89 (3H, m), 3.58-3.56 (4H, m), 3.36-3.32 (2H, m), 3.23- 3.15 (2H, m), 2.95 (2H, s), 2.60-2.58 (2H, m), 2.50-2.49 (2H, m), 2.41-2.33 (5H, m), 2.09- 2.04 (2H, m), 1.99-1.89 (2H, m), 1.42-1.33 (4H, m).
[00689] Example 118. Synthesis of N2-(l-methyl-lH-pyrazol-4-yl)-N4-((lr,4r)-4- morpholinocyclohexyl)benzo[4,5]thieno[2,3-d]pyrimidine-2,4-diamine, compound I- 100, |
Figure imgf000208_0001
1-81 1-100
[00690] To a solution of compound 1-81 (240 mg, 0.51 mmol, 1.00 equiv) in 1 ,4-dioxane (10 mL) was added DDQ (292 mg, 1.29 mmol, 2.51 equiv) at room temperature. Reaction was stirred for 36 h at reflux under nitrogen. Upon completion of the reaction solvents were removed under vacuum and the residue was diluted with 100 mL of EtOAc, washed with 3 x 50 mL of saturated sodium bicarbonate. Combined organic layers were dried over anhydrous sodium sulfate and concentrated under vacuum. Crude product was purified using preparative HPLC to furnish 20 mg of compound 1-100 as a grey solid. LCMS (ES, m/z): 464 [M+H]+; 1H NMR (400 MHz, CD3OD) δ 8.00 (d, 1H), 7.96 (s, 1H), 7.81 (d, 1H), 7.63 (s, 1H), 7.47 (t, 1H), 7.34 (t, 1H), 4.35-4.18 (m, 1H), 3.92 (s, 3H), 3.76 (brs, 4H), 2.68 (brs, 4H), 2.48-2.28 (m, 3H), 2.14 (d, 2H), 1.65-1.41 (m, 4H).
[00691] Example 119. Synthesis of N4-((lr,4r)-4-(dimethylamino)cyclohexyl)-N2-(l- (tetrahydro-2H-pyran-4-yl)-lH-pyrazol-4-yl)-6,7-dihydro-5H-cyclopenta[4,5]thieno[2,3- d]pyrimidine-2,4-diamine, compound 1-101
Figure imgf000209_0001
5.1 1-101
[00692] Compound 1-101 was prepared from compound 5.1 and compound 97.4 using procedure described in Example 94. LCMS (ES, m/z): 482 [M+H]+; 1H NMR (300 MHz. d6- DMSO): δ 8.90 (brs, 1H), 7.89 (s, 1H), 7.45 (s, 1H), 5.69 (d, 1H), 4.45-4.22 (m, 1H), 4.02-3.95 (m, 3H), 3.48-3.35 (m, 2H), 2.95 (t, 2H), 2.82 (t, 2H), 2.45-2.35 (m, 2H), 2.22-2.15 (m, 7H), 2.05 (d, 2H), 1.98-1.68 (m, 6H), 1.60-1.25 (m, 4H).
[00693] Example 120. Synthesis of N4-((lr,4r)-4-(6-azaspiro[2.5]octan-6-yl)cyclohexyl)- N2-(l-methyl-lH-pyrazol-4-yl)-6,7-dihydro-5H-cyclopenta[4,5]thieno[2,3-d]pyrimidine-2,4- diamine, Compound 1-102
CbzCI / THF Jones reagent
Figure imgf000209_0002
NaOH / H20 acetone / 0°C
120.1 120.2 120.3
Figure imgf000209_0003
120.4 120.5
Figure imgf000210_0001
1-102
[00694] Synthesis of compound 120.2. To a 250-mL round-bottom flask charged with a solution of compound 120.1 (5.0 g, 43.41 mmol, 1.00 equiv) in 100 mL of THF/H20 (v:v = 1 : 1) was added benzyl chloroformate (11.08 g, 64.95 mmol, 1.50 equiv) and sodium hydroxide (8.7 g, 217.52 mmol, 5.01 equiv) at room temperature. Reaction was stirred overnight at ambient temperature and concentrated under vacuum to remove THF. Resulting solids were collected by filtration and dried in an oven at 40°C overnight to give 8.4 g (78%) of compound 120.2 as a white solid.
[00695] Synthesis of compound 120.3. To a solution of compound 120.2 (7.0 g, 28.08 mmol, 1.00 equiv) in acetone (100 mL) was added Jones reagent (-10 mL) dropwise at 0 °C. The reaction was monitored by TLC. After stirring for 30 minutes, reaction was quenched with saturated aqueous NaHSOs and extracted with 3 x 100 mL of EtOAc. Combined organic layers were washed with brine, dried over anhydrous sodium sulfate and concentrated under vacuum to give 5.0 g (72%) of compound 120.3 as a white solid.
[00696] Synthesis of compound 120.4. To a solution of 6-azaspiro[2.5]octane (1.9 g, 17.09 mmol, 1.00 equiv) in dichloromethane (60 mL) was added compound 120.3 (6.34 g, 25.64 mmol, 1.50 equiv) and NaBH(OAc)3 (10.89 g, 51.38 mmol, 3.01 equiv) at room temperature. Reaction was stirred overnight at ambient temperature under nitrogen. Upon completion reaction was diluted with 100 mL of H20 and extracted with 3 x 100 mL of dichloromethane. Combined organic layers were dried over anhydrous sodium sulfate and concentrated under vacuum, resulting crude was purified using flash column chromatography to give 2.0 g (34%) of compound 120.4 as a yellow solid.
[00697] Synthesis of compound 120.5. Compound 120.4 (3.1 g, 9.05 mmol, 1.00 equiv) was resolved using preparative chiral HPLC to give 1.4 g of compound 120.5 as a white solid. LCMS (ES, m/z): 343 [M+H]+; 1H NMR (300 MHz, CDC13, ppm) 7.36-7.28 (m, 5H), 5.32 (s, 2H), 4.57 (d, 1H), 3.50-3.35 (m, 1H), 2.62 (brs, 4H), 2.50-2.32 (m, 1H), 2.10 (d, 2H), 1.98-1.88 (m, 2H), 1.55-1.30 (m, 6H), 1.25-1.05 (m, 2H), 0.25 (s, 4H).
[00698] Synthesis of compound 120.6. To a solution of compound 120.5 (300 mg, 0.88 mmol, 1.00 equiv) in methanol (10 mL) was added 10% palladium on carbon (60 mg) under nitrogen at room temperature. Then ¾ (g) was introduced and exchanged three times and the resulting mixture was stirred for 3 hours at ambient temperature. After completion, the solids were filtered out and the filtrate was concentrated under vacuum to give 190 mg (crude) of compound 120.6. as yellow oil.
[00699] Synthesis of compound 120.7. To a solution of compound 3.3 (267 mg, 1.09 mmol, 1.19 equiv) and compound 120.6 (190 mg, 0.91 mmol, 1.00 equiv) in MeCN (10 mL) was added K3PC"4 (581 mg, 2.74 mmol, 3.00 equiv). Reaction was stirred overnight at 80 °C in an oil bath under nitrogen. The resulting mixture was concentrated under vacuum and crude was purified using flash column chromatography to give 275 mg (72%) of compound 120.7 as a yellow solid.
[00700] Synthesis of compound 1-102. Into a 25-mL round-bottom flask charged with a solution of compound 120.7 (120 mg, 0.29 mmol, 1.00 equiv) in 2-butanol (5 mL) was added 1- methyl-lH-pyrazol-4-amine hydrochloride (80.4 mg, 0.58 mmol, 2.00 equiv) at room temperature. Reaction was stirred overnight at 110 °C under nitrogen. After cooling, the solids were collected by filtration, dissolved into water and neutralized with 1 M aqueous sodium hydroxide. Resulting solids were collected by filtration and dried in an oven at 40 °C under reduced pressure to give 60 mg (44%) of compound 1-102 as a grey solid. LCMS (ES, m/z) 478 [M+H]+; 1H NMR (400 MHz, d6-OMSO): δ 8.89 (1H, s), 7.80 (1H, s), 7.45 (1H, s), 5.71 (1H, brs), 4.02-3.95 (1H, s), 3.79 (3H, s), 2.97 (2H, t), 2.84 (2H, t), 2.54-5.50 (4H, m), 2.42-2.29 (3H, m), 2.12-1.99 (2H, m), 1.86-1.79 (2H, m), 1.55-1.35 (4H, m), 1.30 (4h, brs), 0.22 (4H,s). [00701] Example 121. Synthesis of N4-((lr,4r)-4-(6-azaspiro[2.5]octan-6-yl)cyclohexyl)- N2-(l-(tetrahydro-2H-pyran-4-yl)-lH-pyrazol-4-yl)-6,7-dihydro-5H-cyclopenta[4,5]thieno- -d]pyrimidine-2,4-diamine, compound 1-103
Figure imgf000212_0001
[00702] Compound 1-103 was prepared from compound 120.7 and compound 97.4 using procedure described in Example 94. LCMS (ES, m/z): 548 [M+H]+; 1H NMR (300 MHz, DMSO) δ 8.92 (s, 1H), 7.91 (s, 1H), 7.43 (s, 1H), 5.71 (d, 1H), 4.40-4.20 (m, 1H), 4.05-3.88 (m, 3H), 3.51-3.34 (m, 2H), 2.97 (t, 2H), 2.81 (t, 2H), 2.60-2.52 (m, 4H), 2.46-2.40 (m, 3H), 2.11- 1.78 (m, 9H), 1.54-1.38 (m, 4H), 1.35-1.25 (brs, 4H), 0.22 (s, 4H).
[00703] Example 122: FLT3 Inhibition Assay
[00704] The aqueous Base Reaction Buffer was freshly prepared using 20 mM Hepes (pH 7.5), 10 mM MgCl2, 1 mM EGTA, 0.02% Brij35, 0.02 mg/mL BSA, 0.1 mM Na3V04, 2 mM DTT, and 1% DMSO. The kinase substrate (Abltide, [EAIYAAPFAKK ]), and required co- factors (1.5 mM CaCl2, 16 ug/mL Calmodulin, and 2 mM MnCl2) were dissolved in Base Reaction Buffer for each kinase reaction performed. The appropriate FLT3 kinase (wild type: GenBank # NP 004110; FLT3-D835Y; or FLT3-ITD: residues 591-601 duplicated) was added to the substrate solution and gently mixed. Compounds to be tested were dissolved in DMSO and added to the kinase reaction mixture. 33P-ATP (10 uCi/uL) was added to the kinase reaction mixture to initiate the reaction. The kinase reaction was incubated for 120 minutes at room temperature. Each reaction was spotted onto P81 ion exchange paper (Whatman #3698-915), and the paper was thoroughly washed with 0.75%> phosphoric acid. The radioactivity was then quantified and used to calculate the percent of radiolabel incorporated onto the substrate.
[00705] Tables 2, 3, and 4 show the activity of selected compounds of this invention in the FLT3 activity inhibition assay using wild type FLT3 (Table 2), FLT-ITD (Table 3), and FLT- D835Y (Table 4). The compound numbers correspond to the compound numbers in Table 1. Compounds having an activity designated as "A" provided an IC50 < 1 μΜ; compounds having an activity designated as "B" provided an IC50 of 1-2.5 μΜ; compounds having an activity designated as "C" provided an IC50 of 2.5-5 μΜ; and compounds having an activity designated as "D" provided an IC50 > 5 μΜ. "NA" stands for "not assayed."
Table 2. FLT3 (wild type) Activity Inhibition Data
Figure imgf000213_0001
Table 3. FLT3-ITD Activity Inhibition Data
Figure imgf000214_0001
Table 4. FLT3-D835Y Activity Inhibition Data
Figure imgf000214_0002
[00706] While we have described a number of embodiments of this invention, it is apparent that our basic examples may be altered to provide other embodiments that utilize the compounds and methods of this invention. Therefore, it will be appreciated that the scope of this invention is to be defined by the appended claims rather than by the specific embodiments that have been represented by way of example.

Claims

CLAIMS We claim:
1. A method of inhibiting FLT3 kinase comprising contacting said kinase with a compound of formula I:
Figure imgf000215_0001
I
or a pharmaceutically acceptable salt thereof, wherein:
Ring A is a 3-7 membered saturated or partially unsaturated carbocyclic ring or a 4-7 membered saturated or partially unsaturated heterocyclic ring having 1-3 heteroatoms independently selected from nitrogen, oxygen, or sulfur;
n is 0-4;
each R1 is independently -R, halogen, -CN, -N02, -OR, -CH2OR, - SR, -N(R)2, -S02R, -S02N(R)2, -SOR, -C(0)R, -C02R, -C(0)N(R)2, -C(0)N(R)-OR, - NRC(0)OR, -NRC(0)N(R)2, Cy, or -NRS02R; or R1 is selected from one of the following formulas:
Figure imgf000215_0002
two R1 groups are taken together with their intervening atoms to form an optionally substituted 4-7 membered fused, spiro-fused, or bridged bicyclic ring having 0-2 heteroatoms independently selected from nitrogen, oxygen, or sulfur;
each Cy is an optionally substituted monocyclic or bicyclic ring selected from a 3-7 membered saturated or partially unsaturated carbocyclic ring or a 4-7 membered saturated or partially unsaturated heterocyclic monocyclic or bicyclic ring having 1-3 heteroatoms independently selected from nitrogen, oxygen, or sulfur;
each R is independently hydrogen, or an optionally substituted group selected from Ci_6 aliphatic, phenyl, 4-7 membered saturated or partially unsaturated heterocyclic having 1-2 heteroatoms independently selected from nitrogen, oxygen, or sulfur, or 5-6 membered heteroaryl ring having 1-4 heteroatoms independently selected from nitrogen, oxygen, or sulfur, or:
two R groups on the same nitrogen are taken together with their intervening atoms to form a 4-7 membered saturated, partially unsaturated, or heteroaryl ring having 0-3 heteroatoms, in addition to the nitrogen, independently selected from nitrogen, oxygen, or sulfur;
Ring B is a 4-8 membered partially unsaturated carbocyclic fused ring; a 4-7 membered partially unsaturated heterocyclic fused ring having 1-2 heteroatoms selected from nitrogen, oxygen, or sulfur; a benzo fused ring; or a 5-6 membered heteroaromatic fused ring having 1-3 heteroatoms independently selected from nitrogen, oxygen, and sulfur wherein said Ring B may be optionally substituted by one or more oxo, thiono, or imino groups;
m is 0-4;
p is 0-2;
W is N or -C(R3)-;
Rz is R, CN, N02, halogen, -C(0)N(R)2, -C(0)OR, -C(0)R, -N(R)2, -NH-[Ar], -N(R)C(0)OR,
-NRC(0)N(R)2, -OR, or -S02N(R)2;
R3 is hydrogen, halogen, -CN, Ci_4 aliphatic, Ci_4 haloaliphatic, -OR, -C(0)R, or -C(0)N(R)2; [Ar] is an optionally substituted phenyl or heteroaromatic ring;
L1 is a covalent bond or a Ci_6 bivalent hydrocarbon chain wherein one or two methylene units of the chain are optionally and independently replaced by -NR-, -N(R)C(0)-, -C(0)N(R)- , -N(R)S02-, -S02N(R)-, -0-, -C(O)-, -OC(O)-, -C(0)0-, -S-, -SO- or -S02-;
each L2 is independently a covalent bond or a Ci_6 bivalent hydrocarbon chain wherein one or two methylene units of the chain are optionally and independently replaced by -NR-, - N(R)C(0)-, -C(0)N(R)-, -N(R)S02-, -S02N(R)-, -0-, -C(O)-, -OC(O)-, -C(0)0-, -S-, -SO- or -S02-;
each R4 is independently halogen, -CN, -N02, -OR,
SR, -N(R)2, -S02R, -S02N(R)2, -SOR, -C(0)R, -C02R, -C(0)N(R)2, -NRC(0)R, - NRC(0)N(R)2, -C(0)N(R)OR, -N(R)C(0)OR, -N(R)S(0)2N(R)2, -NRS02R, or an optionally substituted group selected from Ci_6 aliphatic, phenyl, 4-7 membered saturated or partially unsaturated heterocyclic having 1-2 heteroatoms independently selected from nitrogen, oxygen, or sulfur, or 5-6 membered heteroaryl ring having 1-4 heteroatoms independently selected from nitrogen, oxygen, or sulfur, or:
two -L2(R4)P-R4 groups are taken together with their intervening atoms to form an optionally substituted 4-7 membered fused, spiro-fused, or bridged bicyclic ring having 0-2 heteroatoms independently selected from nitrogen, oxygen, or sulfur.
2. The method of claim 1 wherein the compound is of formula II:
Figure imgf000217_0001
II
or a pharmaceutically acceptable salt thereof.
3. The method of claim 2 wherein the compound is of formula III:
Figure imgf000217_0002
III
or a pharmaceutically acceptable salt thereof.
4. The method of claim claim 3 wherein the compound is of formula IV:
Figure imgf000217_0003
IV
or a pharmaceutically acceptable salt thereof.
5. The method of claim 4 wherein the compound is of formula V:
Figure imgf000218_0001
V
or a pharmaceutically acceptable salt thereof.
6. The method of claim 1 wherein the compound is of formula XXIV:
Figure imgf000218_0002
XXIV
or a pharmaceutically acceptable salt thereof.
7. The method of claim 1 wherein the FLT3 is in a biological sample.
8. The method of claim 1 wherein the compound is selected from those depicted in Table 1, or pharmaceutically acceptable salts thereof.
9. A method of treating an FLT3 -mediated disorder, disease, or condition in a patient comprising administering to said patient a pharmaceutical composition comprising a compound of formula I:
Figure imgf000218_0003
I
or a pharmaceutically acceptable salt thereof, wherein: Ring A is a 3-7 membered saturated or partially unsaturated carbocyclic ring or a 4-7 membered saturated or partially unsaturated heterocyclic ring having 1-3 heteroatoms independently selected from nitrogen, oxygen, or sulfur;
n is 0-4;
each R1 is independently -R, halogen, -CN, -N02, -OR, -CH2OR, - SR, -N(R)2, -S02R, -S02N(R)2, -SOR, -C(0)R, -C02R, -C(0)N(R)2, -C(0)N(R)-OR, - NRC(0)OR, -NRC(0)N(R)2, Cy, or -NRS02R; or R1 is selected from one of the following formulas:
Figure imgf000219_0001
two R1 groups are taken together with their intervening atoms to form an optionally substituted 4-7 membered fused, spiro-fused, or bridged bicyclic ring having 0-2 heteroatoms independently selected from nitrogen, oxygen, or sulfur;
each Cy is an optionally substituted monocyclic or bicyclic ring selected from a 3-7 membered saturated or partially unsaturated carbocyclic ring or a 4-7 membered saturated or partially unsaturated heterocyclic monocyclic or bicyclic ring having 1-3 heteroatoms independently selected from nitrogen, oxygen, or sulfur;
each R is independently hydrogen, or an optionally substituted group selected from Ci_6 aliphatic, phenyl, 4-7 membered saturated or partially unsaturated heterocyclic having 1-2 heteroatoms independently selected from nitrogen, oxygen, or sulfur, or 5-6 membered heteroaryl ring having 1-4 heteroatoms independently selected from nitrogen, oxygen, or sulfur, or:
two R groups on the same nitrogen are taken together with their intervening atoms to form a 4-7 membered saturated, partially unsaturated, or heteroaryl ring having 0-3 heteroatoms, in addition to the nitrogen, independently selected from nitrogen, oxygen, or sulfur;
Ring B is a 4-8 membered partially unsaturated carbocyclic fused ring; a 4-7 membered partially unsaturated heterocyclic fused ring having 1-2 heteroatoms selected from nitrogen, oxygen, or sulfur; a benzo fused ring; or a 5-6 membered heteroaromatic fused ring having 1-3 heteroatoms independently selected from nitrogen, oxygen, and sulfur wherein said Ring B may be optionally substituted by one or more oxo, thiono, or imino groups; m is 0-4;
p is 0-2;
W is N or -C(R3)-;
Rz is R, CN, N02, halogen, -C(0)N(R)2, -C(0)OR, -C(0)R, -N(R)2, -NH-[Ar], - N(R)C(0)OR, -NRC(0)N(R)2, -OR, or -S02N(R)2;
R3 is hydrogen, halogen, -CN, Ci_4 aliphatic, Ci_4 haloaliphatic, -OR, -C(0)R, or -C(0)N(R)2; [Ar] is an optionally substituted phenyl or heteroaromatic ring;
L1 is a covalent bond or a Ci_6 bivalent hydrocarbon chain wherein one or two methylene units of the chain are optionally and independently replaced by -NR-, -N(R)C(0)-, -C(0)N(R)- , -N(R)S02-, -S02N(R)-, -0-, -C(O)-, -OC(O)-, -C(0)0-, -S-, -SO- or -S02-;
each L2 is independently a covalent bond or a Ci_6 bivalent hydrocarbon chain wherein one or two methylene units of the chain are optionally and independently replaced by -NR-, - N(R)C(0)-, -C(0)N(R)-, -N(R)S02-, -S02N(R)-, -0-, -C(O)-, -OC(O)-, -C(0)0-, -S-, -SO- or -S02-;
each R4 is independently halogen, -CN, -N02, -OR,
SR, -N(R)2, -S02R, -S02N(R)2, -SOR, -C(0)R, -C02R, -C(0)N(R)2, -NRC(0)R, - NRC(0)N(R)2, -C(0)N(R)OR, -N(R)C(0)OR, -N(R)S(0)2N(R)2, -NRS02R, or an optionally substituted group selected from Ci_6 aliphatic, phenyl, 4-7 membered saturated or partially unsaturated heterocyclic having 1-2 heteroatoms independently selected from nitrogen, oxygen, or sulfur, or 5-6 membered heteroaryl ring having 1-4 heteroatoms independently selected from nitrogen, oxygen, or sulfur, or:
two -L2(R4)P-R4 groups are taken together with their intervening atoms to form an optionally substituted 4-7 membered fused, spiro-fused, or bridged bicyclic ring having 0-2 heteroatoms independently selected from nitrogen, oxygen, or sulfur.
10. The method of claim 9 wherein the disorder is selected from acute myeloid leukemia, solid tumors, gliomas, myelodysplasia syndrome, renal cell carcinoma, glioblastoma, prostate cancer, melanoma, acute lymphoblastic leukemia, myeloproliferative disorder, nasopharyngeal carcinoma, breast tumors, thrombocytopenia, polycythemia vera, myelofibrosis, chronic myelocytic leukemia, lung tumors, colorectal tumors, and autoimmune diseases.
11. The method of claim 9 wherein the disorder is is selected from gliomas, myelodysplasia syndrome, acute lymphoblastic leukemia, myeloproliferative disorder, nasopharyngeal carcinoma, polycythemia vera, and myelofibrosis.
12. The method of claim 9 wherein the disorder is acute myeloid leukemia.
13. The method of claim 12 wherein the acute myeloid leukemia is associated with one or more mutations of FLT3.
14. The method of claim 13 wherein the mutations are selected from internal tandem duplications and D835Y point mutations.
15. The method of claim 13 wherein the acute myeloid leukemia is resistant to one or more other FLT3 inhibitors.
16. The method of claim 15 wherein the acute myeloid leukemia is resitant to one or more FLT3 inhibitors selected from quizartinib, midostaurin, lestaurtinib, sorafenib, and sunitinib.
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US11053195B2 (en) 2013-03-15 2021-07-06 Global Blood Therapeutics, Inc. Compounds and uses thereof for the modulation of hemoglobin
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