BR112018011178B1 - CONJUGATE SALTS, PHARMACEUTICAL COMPOSITION, AND THEIR USE IN CANCER THERAPY - Google Patents

Abstract

SALTS OF CONJUGATES FOR CANCER THERAPY. The present invention relates to pharmaceutically acceptable salts of conjugates comprising a chemotherapeutic drug and an amino acid or a derivative thereof, which are readily taken up by a target cell and reduce the side effects induced by the chemotherapeutic drug. In particular, the present invention relates to pharmaceutically acceptable salts of conjugates comprising cytidine and aspartic or glutamic acid analog drugs and analogs thereof, pharmaceutical compositions comprising these conjugates and use thereof for the treatment of cancer or a precancerous condition or disorder.

Description

FIELD OF INVENTION

[001] The present invention relates to pharmaceutically acceptable salts of conjugates comprising a chemotherapeutic drug and an amino acid or a derivative thereof, which are readily taken up by a cancer cell. In particular, the present invention relates to pharmaceutically acceptable salts of conjugates of cytidine and aspartic or glutamic acid analogue drugs and analogues thereof, pharmaceutical compositions comprising these conjugates and use thereof for the treatment of cancer or a precancerous condition or disorder.

FUNDAMENTALS OF THE INVENTION Antiproliferative Drugs

[002] Antiproliferative drugs, also known as antimetabolites, antineoplastic agents, and covalent DNA-binding drugs, act by inhibiting essential metabolic pathways and are commonly used in the treatment of malignant diseases. However, their high toxicity to normal cells and severe side effects limit their use as therapeutic agents. Undesirable side effects include anemia, emesis, and baldness due to cytotoxic effects on rapidly dividing normal cells such as stem cells in the bone marrow, epithelial cells of the intestinal tract, hair follicle cells, etc.

[003] Another major problem associated with antiproliferative drugs is the inherent or acquired resistance of tumors to the drugs. For example, although the initial remission rate after treatment with L-asparaginase is quite high in patients with acute lymphoblastic leukemia (ALL), relapsed and associated drug resistance represents a significant clinical problem. Studies have demonstrated increased expression of asparagine synthetase (AS) in asparaginase-resistant cells, which has led to the hypothesis that elevated AS activity allows the survival of drug-resistant malignant cells.

Nucleotide/nucleoside analogues

[004] Nucleoside analogues compete with their physiological counterparts for incorporation into nucleic acids and have gained an important place in the treatment of acute leukaemia. The most important are the arabinose nucleosides; a unique class of antimetabolites originally isolated from the sponge Cryptothethya crypta but now produced synthetically. They differ from the physiological deoxyribonucleosides by the presence of a 2'-OH group in the cis configuration relative to the N-glycosyl bond between cytosine and the arabinoside sugar. Several arabinose nucleosides have useful antitumor and antiviral effects. The most active cytotoxic agent of this class is the cytosine arabinoside (Cytarabine). A related nucleoside, adenine arabinoside, also exerts antitumor activity, and its analogue, fludarabine phosphate (2-fluoro-ara-adenosine monophosphate), has potent antitumor activity in lymphomas and chronic lymphocytic leukemia (Warrell & Berman, 1986). Another member of the group is arabinosyl-5-azacytidine, a synthetic analogue that has failed in the clinic (Dalal et al, 1986).

[005] One goal of analogue development in the area of cytidine antimetabolites has been to find compounds that preserve the inhibitory activity of ara-C but are resistant to deamination. A number of deaminase-resistant analogues have been developed, including cyclocytidine (Ho DHW, 1974) and N4-behenoya ara-C (Kodama et al., 1989) which showed antileukemic activity in some clinical trials but had undesirable side effects (Woodcock et al., 1980). Other representative compounds are N4-Palmitoyl-ara, 2´-Azido-2´-deoxy ara-C, 5´-(Cortisone 21-phosphoryl)ester of ara-C, 5´-acyl esters of ara-C (e.g., 5´-palmitate ester), N4-Beenoyl-ara-C, Ara-C conjugate with poly-H5(2-hydroxyethyl)-L-glutamine, Dihydro-5-azacytidine, 5-Aza-arabinosylcytosine, 5-Aza 2´-deoxycytidine, and 2´-2´-difluorodeoxycytidine (Hartel et al., 1990 and Heineman et al., 1988).

[006] Gemcitabine (2,2-difluorodeoxycytidine, dFdC) is the most important cytidine analogue to enter clinical trials since ara-C. It has been incorporated into standard first-line therapy for patients with pancreatic cancer, lung cancer, and transitional cell cancer of the bladder.

[007] Nucleotide analogues have also been used in non-oncological applications. For example, flucytosine, a fluorinated analogue of cytosine, is used as an antifungal agent.

Amino Acids and Proliferative Disease

[008] Asparagine is a non-essential amino acid that is required for rapidly proliferating cells. Mammalian cells can synthesize asparagine from aspartate using the ATP-dependent enzyme asparagine synthetase (EC 6.3.5.4), which transfers the amino group from the amide of glutamine to the β-carboxyl of aspartate in a reaction that can be represented as: Glutamine + Aspartate + ATP + H2O = Glutamate + Asparagine + AMP + PPi.

[009] Asparagine synthetase deficiency occurs in certain tumors, causing them to depend on an external supply of asparagine from other sources, such as serum. This observation led to the development of the enzyme L-asparaginase (type CE-2, CE 3.5.1.1) as a chemotherapeutic agent. L-asparaginase hydrolyzes L-asparagine to aspartate and ammonia, depleting serum L-asparagine and inhibiting tumor growth. L-asparaginase is primarily used in the treatment of acute lymphoblastic leukemia (ALL) and shows some activity against other hematologic cancers, including acute nonlymphocytic leukemia.

[0010] L-asparaginase used in the clinic is available in two unmodified (native) forms purified from bacterial sources, and one as a PEGylated compound. U.S. Patent No. 4,179,337 describes PEGylated L-asparaginase, in which the enzyme is coupled to PEG having a molecular weight of about 500 to 20,000 Daltons.

[0011] In vivo downregulation of asparagine synthetase may provide an efficient mechanism to inhibit tumor growth. However, cells respond to amino acid deprivation by a concerted increase in asparagine synthetase mRNA, protein, and enzyme activity that involves transcriptional control of the asparagine synthetase gene.

[0012] Initially, a metabolic approach was used to inhibit asparagine synthetase activity by generating analogs of L-asparagine and L-aspartic acid. Analogs including 5-carboxamido-4-amino-3-isoxazolidone (Stammer et al., 1978) and N-substituted sulfonamides and N-substituted sulfonylhydrazides were prepared as sulfur analogs of L-asparagine (Brynes S et al., 1978a Brynes S et al., 1978b). U.S. Patent No. 4,348,522 teaches the N-phosphonacetyl-L-aspartic acid salt, PALA, which has been shown to exhibit antitumor activity and is currently in clinical trials as combination chemotherapy for colorectal and pancreatic cancers.

[0013] The aspartic acid analogue of Ara-C was used as a starting material for the further synthesis of Ara-C T-peptide conjugates to target CD4-positive cells (Manfredini et al., 2000).

[0014] The use of prodrugs to impart desired characteristics, such as increased bioavailability or increased site specificity, is a recognized concept in the art of pharmaceutical development. For example, direct or indirect conjugation of a drug to an antibody creates a stable conjugate that can reach the target site with minimal drug dissociation. Drug targeting can be combined with a selective drug release mechanism for maximum potency.

[0015] US Patent No. 4,296,105 describes doxorubicin derivatives linked to an amino acid optionally substituted at the hydroxy group of the amino acid residue, which have in vitro greater antitumor activity and lower toxicity than doxorubicin.

[0016] U.S. Patent No. 5,962,216 teaches tumor-activated prodrugs that are unable to enter the cell until cleaved by a factor or factors secreted by a target cell.

[0017] US Patent No. 5,650,386 describes compositions comprising at least one active agent, and at least one modified non-alpha amino acid or polyamino acid, which acts as a carrier for the active agent. Modification of amino acids includes acylation or sulfonation of at least one free amino group.

[0018] U.S. Patent Nos. 6,623,731, 6,428,780, and 6,344,213 teach non-covalent mixtures comprising modified amino acids as carriers for biologically active agents.

[0019] US Patent No. 5,106,951 describes a conjugate comprising an aromatic drug non-covalently intercalated between two aromatic side chains in an oligopeptide, and an antibody or antibody fragment covalently linked to the oligopeptide for targeting to cancer cells.

[0020] U.S. Patent No. 6,617,306 teaches a carrier for in vivo delivery of a therapeutic agent, the carrier and the therapeutic agent linked by a disulfide bond. In that patent, the carrier comprises a polymer, and at least one thiol compound conjugated to the polymer such that the thiol group of the thiol compound and the thiol group of the therapeutic agent form a disulfide bond.

[0021] International patent application publication No. WO 00/33888 teaches cleavable anti-tumor and anti-inflammatory compounds comprising a therapeutic agent capable of entering a target cell, an oligopeptide, a stabilizing group and an optional linker.

[0022] International patent application publication No. WO 2005/072061 and U.S. Patent No. 7,989,188 to some of the inventors of the present invention describe compounds comprising a drug covalently linked to an amino acid through a side chain with a functional group useful for targeting drugs to neoplastic cells.

Pharmaceutical Salts

[0023] Selection of optimal pharmaceutical salts of a drug can increase the efficacy and improve drug delivery of the drug. Establishing such active drug salts is nontrivial and requires unique methods in the development and analysis of such salts.

[0024] There remains an unmet medical need for compounds and compositions capable of targeting tumors while avoiding cytotoxic damage to normal tissues.

SUMMARY OF THE INVENTION

[0025] It is an object of the present invention to target drugs to malignant and pre-malignant cells while reducing the side effects associated with current drug therapy.

[0026] The present invention provides pharmaceutically acceptable salts of conjugates comprising a first chemical moiety covalently linked to a second chemical moiety, wherein the first chemical moiety is an amino acid comprising in its side chain at least one functional group selected from the group consisting of an amino group, a carboxyl group, a sulfhydryl group and a hydroxyl group, and the second chemical moiety is an anticancer drug selected from the group consisting of a cytotoxic agent, a cytostatic agent and a chemotherapeutic agent. The anticancer drug is linked to the amino acid through the functional group of the side chain of the amino acid. The amino acid is preferably aspartic acid or glutamic acid or derivatives or analogues thereof covalently linked to the anticancer drug, preferably a nucleotide/nucleoside analogue such as cytarabine or gemcitabine. The pharmaceutically acceptable salt is preferably selected from an organic or inorganic acid or residue of an acid (i.e., its conjugate base). The present invention further provides pharmaceutical compositions comprising the conjugate salts, and use thereof for the treatment of cancer and pre-cancerous conditions or disorders. The present invention further provides methods for preparing the salt forms of the conjugates of the present invention, in particular, by reacting a protected precursor compound with an acid that forms the relevant salt.

[0027] The salt forms of the present invention can serve as a delivery vehicle for a drug or prodrug wherein the drug undergoes rapid uptake by cancer cells. The salt forms can also serve to increase the stability and solubility of the conjugate.

[0028] The conjugates of the present invention are typically prepared from precursors that contain one or more protecting groups on susceptible moieties, such as amines or carboxylic acids. For example, the amino group of the amino acid is typically protected by a t-butoxycarbonyl (BOC) group. It is well known that BOC groups are typically removed from amino groups in acidic media, with trifluoroacetic acid (TFA) being the most commonly used deprotecting agent. It is further known that TFA is difficult to remove and therefore deprotected amines typically contain residual amounts of TFA. Due to its toxic effects, compounds that are devoid of TFA are required for pharmaceutical use. In some embodiments, the present invention avoids the need to use TFA as a deprotecting agent. It has now been unexpectedly discovered that salts of the conjugates of the present invention can be prepared by deprotecting the BOC groups using the same reagent that ultimately forms the salt. It is now described that certain pharmaceutically acceptable acids can simultaneously remove the protecting group(s) and generate the salt of the conjugate of the present invention in a single step, thereby obviating an intermediate TFA deprotection step prior to salt formation. Thus, in certain embodiments, the conjugates of the present invention are not exposed to TFA for deprotection, and thus the conjugate salts of the present invention are free of TFA residues. The resulting salts are therefore highly advantageous in that they are devoid of toxic TFA and are therefore suitable for pharmaceutical use.

[0029] According to a first aspect, the present invention provides a pharmaceutically acceptable salt of a compound, the salt represented by the structure of formula (I) wherein, A designates an amino acid comprising in its side chain at least one functional group selected from the group consisting of a carboxyl group, an amino group, a sulfhydryl group and a hydroxyl group; D denotes a residue of a drug selected from the group consisting of a cytotoxic agent, a cytostatic agent and a chemotherapeutic agent, wherein D is linked to A through the side chain functional group of A; and Y is a pharmaceutically acceptable acid or organic or inorganic residue of an acid (i.e., the conjugate base of said acid), selected from the group consisting of hydrochloric acid, acetic acid, methanesulfonic acid, phosphoric acid, citric acid, lactic acid, succinic acid, tartaric acid, boric acid, benzoic acid, toluenesulfonic acid, benzenesulfonic acid, ascorbic acid, sulfuric acid, maleic acid, formic acid, malonic acid, nicotinic acid, and oxalic acid, wherein the pharmaceutically acceptable salt contains 0.1% or less of trifluoroacetic acid (TFA) as an impurity. Each possibility represents a separate embodiment of the present invention.

[0030] The amino acid (A) may be an α-amino acid (D or L), b-amino acid, g-amino acid, d-amino acid, or e-amino acid. In a currently preferred embodiment, the amino acid (A) is an α-amino acid (D or L). In some embodiments, the amino acid (A) is selected from the group consisting of aspartic acid (Asp), glutamic acid (Glu), asparagine (Asn), glutamine (Gln), α-aminoadipic acid (Aad), α-aminopimelic acid, g-carboxy-glutamic acid, g-hydroxy-glutamic acid, aminoglycine, amino-isobutyric acid (Aib), arginine (Arg), citrulline (Cit), cysteine (Cys), cystine, diaminobutanoic acid, diaminobutyric acid (Dab), diaminopropionic acid (Dpr), dihydroxyphenylalanine, dimethylarginine, pyroglutamic acid (p-Glu), histidine (His), 1-methyl-histidine, 3-methyl-histidine, homoserine (Hse), homocitrulline, hydroxyproline (Hyp), lysine (Lys), methyl-lysine, dimethyl lysine, trimethyl lysine, azidolysine, methionine (Met), methionine sulfoxide, methionine sulfone, ornithine (Orn), sarcosine (Sar), selenocysteine (Sec), serine (Ser), phospho-serine, methyl-serine, aminoserine (Ams), thienylalanine (Thi), threonine (Thr), phospho-threonine, tryptophan (T rp), tyrosine (Tyr), methyl-tyrosine, phosphor-tyrosine, sulfo-tyrosine, α-aminosuberic acid, 3,5-diiodotyrosine, penicillamine (Pen), 4-ethylamine phenylglycine, 4-aminophenylglycine, 4-sulfophenylalanine, 4-aminophenylalanine and 2-amino-4[4-(2- [amino]-pyrimidinyl]butanoic acid, 3-aminopropionic acid, 6-aminohexanoic acid (e-Ahx), p-aminobenzoic acid, isonipecotic acid, statin (Sta), 2-aminobutyric acid (Abu) and 4-aminobutyric acid, and derivatives and analogues thereof. Each possibility represents a separate embodiment of the present invention.

[0031] In some embodiments, the amino acid (A) is selected from the group consisting of aspartic acid, glutamic acid, asparagine, glutamine, and derivatives and analogues thereof. In a currently preferred embodiment, the amino acid (A) is aspartic acid. In another currently preferred embodiment, the amino acid (A) is glutamic acid.

[0032] Group (D) may be derived from any drug selected from the group consisting of a cytotoxic agent, a cytostatic agent, and a chemotherapeutic agent. In some embodiments, the cytotoxic, cytostatic, or chemotherapeutic agent D is the residue of a drug selected from the group consisting of a pyrimidine analog, a purine analog, an antifolate, hydroxyurea, an antimicrotubule agent, an alkylating agent, an antitumor antibiotic, a topoisomerase targeting agent, an antimetabolite, a DNA binding agent, and a DNA antagonist.

[0033] In some embodiments, the cytotoxic, cytostatic, or chemotherapeutic agent D is the residue of a pyrimidine analog selected from cytosine arabinoside (ara-C, cytarabine), gemcitabine, fluorouracil, 5-fluorodeoxyuridine (5-FUDR), ftorafur, capecitabine, carmofure, BOF-A2, 5-chloro-2,4-dihydroxypyridine, decitabine, leflunomide, trifluridine, iodoxuridine, zidovudine, telbivudine, trimethoprim, fluorocytosine, deoxythymidine, and 5-amino-6-nitrouracil. Each possibility represents a separate embodiment of the present invention. In a currently preferred embodiment, the cytotoxic, cytostatic, or chemotherapeutic agent D is the residue of cytarabine. In another currently preferred embodiment, the cytotoxic, cytostatic or chemotherapeutic agent D is the gemcitabine residue.

[0034] In other embodiments, the cytotoxic, cytostatic, or chemotherapeutic agent D is the residue of a purine analog selected from clofarabine, decarbazine, cladribine, mercaptopurine, nelarabine, pentostatin, thioguanine, gefitinib, azathiopurine, 8-azaguanine, immucillin-G, 2-fluoro-2'-deoxyadenosine, 9-beta-D-xylofuranosyl-adenine, 3-dexoxyguanine, 6-methyl-formycin A, 2-fluoroadenosine, 1-deaza-adenosine, N-ethyl-5-carboxamido-adenosine, 3'-oxo-adenosine, 2-amino-adenosine, 6-O-cyclomethyl-guanine fludarabine, and 8-iodo-guanine. Each possibility represents a separate embodiment of the present invention.

[0035] In other embodiments, the cytotoxic, cytostatic, or chemotherapeutic agent D is the residue of a drug selected from the group consisting of 2,4-dioxo-5-fluoropyrimidine (5-FU), azacitidine (5-AZC), 6-mercaptopurine (6-MP), 6-thioguanine (6-TG), chlorodeoxyadenosine (2-CDA), and pentostatin (dCF). Each possibility represents a separate embodiment of the present invention.

[0036] In some embodiments, the salts of formula (I) are prepared in a substantially pure form. The term “substantially pure” refers to at least about 95% purity as determined by HPLC, preferably at least about 97% purity, more preferably at least about 98% purity, and most preferably at least about 99% purity.

[0037] As noted above, in the conjugates of the present invention, the cytotoxic, cytostatic or chemotherapeutic agent D is linked to the amino acid (A) through the side chain functional group of (A). Some non-limiting embodiments of such conjugates are described below. According to some embodiments, the amino acid (A) is selected from the group consisting of aspartic acid and glutamic acid, and the compound is represented by the structure of formula (II) or (III): where D and Y are as defined above.

[0038] In a specific embodiment of formula (II), A is aspartic acid and D is the cytarabine residue. In this embodiment, the compound is a salt represented by the structure of formula (1):

[0039] In one embodiment of formula (1), the compound is a salt with acetic acid (i.e., an acetate salt), which is represented by the structure of formula (1A):

[0040] In another embodiment of formula (1), the compound is a salt with hydrochloric acid (i.e., a hydrochloride salt), which is represented by the structure of formula (1B):

[0041] In another specific embodiment of formula (II), A is aspartic acid and D is the residue of gemcitabine and the compound is represented by the structure of formula (2):

[0042] In a specific embodiment of formula (III), A is glutamic acid and D is the cytarabine residue and the compound is represented by the structure of formula (3):

[0043] In another specific embodiment of formula (III), A is glutamic acid and D is the residue of gemcitabine and the compound is represented by the structure of formula (4):

[0044] In any of the above compounds of formula (I), (II), (III), (1), (2), (3) and (4), Y is a pharmaceutically acceptable acid selected from the group consisting of acetic acid, hydrochloric acid, methanesulfonic acid, phosphoric acid, citric acid, lactic acid, succinic acid, tartaric acid, boric acid, benzoic acid, toluenesulfonic acid, benzenesulfonic acid, ascorbic acid, sulfuric acid, maleic acid, formic acid, malonic acid, nicotinic acid and oxalic acid. Each possibility represents a separate embodiment of the present invention.

[0045] In a currently preferred embodiment, the pharmaceutically acceptable acid Y is acetic acid, i.e., an organic acid. In another currently preferred embodiment, the pharmaceutically acceptable acid Y is hydrochloric acid (HCl), i.e., an inorganic acid.

[0046] It is understood by one skilled in the art that any of the aforementioned pharmaceutically acceptable acids can form a pharmaceutically acceptable salt with the compounds of the invention. In some embodiments, the pharmaceutically acceptable salt is selected from the group consisting of: acetate, hydrochloride, methanesulfonate, phosphate, citrate, lactate, succinate, tartrate, borate, benzoate, toluenesulfonate, benzenesulfonate, ascorbate, sulfate, maleate, formate, malonate, nicotinate, and oxalate. Each possibility represents a separate embodiment of the present invention.

[0047] In a currently preferred embodiment, the pharmaceutically acceptable acid is acetic acid (CH3COOH). In this embodiment, the salt is an acetate salt. In another preferred embodiment, the pharmaceutically acceptable salt is a hydrochloric acid (HCl) salt. In this embodiment, the salt is a hydrochloride salt.

[0048] In another aspect, the present invention provides a pharmaceutical composition comprising as active ingredient a salt of formula (I), or any compound covered by that formula, for example, compounds (II), (III), (1), (1A), (1B), (2), (3) or (4), as described herein and a pharmaceutically acceptable carrier or diluent.

[0049] In another aspect, the present invention provides a method of treating a cancer including metastases thereof, comprising the step of administering to a subject in need thereof a therapeutically effective amount of a salt of formula (I), or any compound covered by such formula, e.g., compounds (II), (III), (1), (1A), (1B), (2), (3) or (4), as described herein. In another aspect, the present invention provides a salt of formula (I), or any compound covered by such formula, e.g., compounds (II), (III), (1), (1A), (1B), (2), (3) or (4), as described herein, for use in treating a cancer including metastases thereof. In some embodiments, the cancer is characterized by a non-solid tumor or a solid tumor or a combination thereof.

[0050] In other embodiments, the cancer is a hematologic cancer. In other specific embodiments, the hematologic cancer is selected from the group consisting of leukemias, lymphomas, and multiple myeloma. In some embodiments, the leukemia is selected from the group consisting of acute myeloid leukemia (AML) and acute lymphoblastic leukemia (ALL). In some embodiments, the leukemia is a relapsed/refractory leukemia. In other embodiments, the leukemia is newly diagnosed leukemia.

[0051] In some specific embodiments, the cancer characterized by a solid tumor selected from the group consisting of central nervous system (CNS) tumors, liver cancer, colorectal carcinoma, breast cancer, gastric cancer, pancreatic cancer, bladder carcinoma, cervical carcinoma, head and neck tumors, vulvar cancer, and dermatologic neoplasms including melanoma, squamous cell carcinoma, and basal cell carcinomas.

[0052] In another aspect, the present invention provides a method of treating a pre-cancer condition or disorder, comprising the step of administering to a subject in need thereof a therapeutically effective amount of a salt of formula (I), or any compound covered by such formula, for example, compounds (II), (III), (1), (1A), (1B), (2), (3) or (4), as described herein.

[0053] In another aspect, the present invention provides a salt of formula (I), or any compound covered by that formula, for example, compounds (II), (III), (1), (1A), (1B), (2), (3) or (4), as described herein, for use in the treatment of a pre-cancerous condition or disorder.

[0054] In some embodiments, the pre-cancer condition or disorder is Myelodysplastic Syndrome (MDS).

[0055] In other embodiments, the present invention relates to a method of treating an individual who is a candidate for a bone marrow transplant, comprising the step of pre-treating said individual, prior to the transplant, with a salt of formula (I) or any compound covered by such formula, for example, compounds (II), (III), (1), (1A), (1B), (2), (3) or (4), as described herein.

[0056] In other embodiments, the present invention provides a salt of formula (I), or any compound covered by that formula, e.g., compounds (II), (III), (1), (1A), (1B), (2), (3) or (4), as described herein, for use in pretreatment prior to a bone marrow transplant.

[0057] The salt forms of the present invention can be used to treat any mammal, preferably humans.

[0058] Further described is a method for preparing a pharmaceutically acceptable salt of a compound represented by formula (I) of the present invention, comprising the step of

[0059] (a) reacting a compound of formula (IV): wherein A and D are as defined above; n is an integer from 1 to 6; and each P is independently at each occurrence a protecting group; wherein each P is attached to any functional group available for protection on the amino acid (A), the drug residue (D), or on both the amino acid (A) and the drug residue (D); with a reagent capable of removing the protecting group(s) P; and (b) adding a reagent capable of introducing the group Y, so as to generate a salt represented by the formula: where Y is as defined above.

[0060] In one embodiment, it has now been unexpectedly discovered that the salts of the present invention can be prepared by deprotecting a precursor compound using the same pharmaceutically acceptable acid that ultimately forms the salt. Thus, in accordance with the principles of the present invention, the pharmaceutically acceptable acid Y capable of introducing the group Y can simultaneously remove the protecting group(s) P and generate the salt of formula (I) in one step, thereby obviating an intermediate deprotection step prior to salt formation. In this way, the purity level of the salt product is improved as residual amounts of deprotecting agent (i.e. impurities) are avoided. For example, protecting groups such as t-butoxycarbonyl (BOC) are typically removed in acidic media, for example with trifluoroacetic acid (TFA). However, TFA is difficult to remove and therefore the deprotected product includes residual amounts of this acid. This inevitably leads to increased levels of TFA impurities in the final salt forms, which is unacceptable from a pharmaceutical development perspective. The present invention provides a solution to this problem by performing the final step of the synthesis using the same reagent (e.g. acid) that will ultimately form the salt.

[0061] Thus, in one embodiment, the process of the invention involves the use of a reagent capable of removing the protecting group(s) P and simultaneously introducing the group Y, for example, an acid. According to this embodiment, the protecting group(s) P is removed and the group Y is introduced in a single step.

[0062] According to some embodiments, the pharmaceutically acceptable salt of the conjugate of the present invention contains about 5% or less residual TFA. According to additional embodiments, the pharmaceutically acceptable salt of the conjugate of the present invention contains about 1% or less residual TFA. According to additional embodiments, the pharmaceutically acceptable salt of the conjugate of the present invention contains about 0.1% or less residual TFA. According to a certain embodiment, the pharmaceutically acceptable salt of the conjugate of the present invention is substantially devoid of TFA as an impurity. According to another embodiment, the pharmaceutically acceptable salt of the conjugate of the present invention is completely devoid of TFA as an impurity.

[0063] Compounds of formula (I) can generally be prepared by coupling a drug moiety with an amino acid, preferably in the presence of a coupling agent, and conversion to the salt form according to the process described above. This process is exemplified herein for compounds of formula (II) and (III).

[0064] In some embodiments, the present invention provides a process for preparing a compound of formula (II), comprising the step of: (a) coupling a compound of formula (i) with a drug of formula D or a protected derivative thereof of formula D-(P3)n in the presence of a coupling reagent to provide an intermediate of formula (ii); and (b) removing the protecting groups P1, P2 and P3 (if present), with an acid (Y) or with a deprotecting agent followed by an acid (Y) so as to generate a salt represented by formula (II): where P1 is a carboxy protecting group, P2 is an amino protecting group, P3 is a protecting group located on one or more functional groups in the drug D and n is 0, 1, 2 or 3.

[0065] In other embodiments, the present invention provides a process for preparing a compound of formula (III), comprising the step of: (a) coupling a compound of formula (iii) with a drug of formula D or a protected derivative thereof of formula D-(P3)n in the presence of a coupling reagent to provide an intermediate of formula (iv); and (b) removing the protecting groups P1, P2 and P3 (if present), with an acid (Y) or with a deprotecting agent followed by an acid (Y) so as to generate a salt represented by formula (II). where P1 is a carboxy protecting group, P2 is an amino protecting group, P3 is a protecting group located on one or more functional groups in the drug D and n is 0, 1, 2 or 3.

[0066] In one embodiment, the drug is cytarabine. In other embodiments, the drug is protected cytarabine comprising protecting groups at its 2’, 3’, and/or 5’ hydroxyl moieties, such as, triphenylmethyl-cytarabine (CAS 7075-13-0), benzoyl-cytarabine (CAS 34270-10-5), admantoyl-cytarabine (CAS 23113-01-1), and trimethylsilyl-cytarabine. In other embodiments, the drug is gemcitabine. In other embodiments, the drug is a protected gemcitabine comprising protecting groups at its 3’ and/or 5’ hydroxyl moieties, such as, triphenylmethyl-gemcitabine (CAS 1642862-24-5).

[0067] In some embodiments, the protecting groups are removed simultaneously with the introduction of the Y group, using an acid that is capable of removing the protecting groups and introducing the Y group.

[0068] These and other embodiments of the present invention will become apparent in conjunction with the following figures, description and claims.

BRIEF DESCRIPTION OF THE FIGURES

[0069] FIG. 1: Mass spectrum of Asp(Cytarabine) •acetate salt (Y = CH3COOH).

[0070] FIG. 2: Mass spectrum of Asp(Cytarabine) methanesulfonate salt (Y = CH3SO3H).

[0071] FIG. 3: Mass spectrum of Asp(Cytarabine) benzenesulfonate salt (Y = C6H5SO3H).

[0072] FIG. 4: 1H NMR of purified BOC-Asp-OtBu-(Cytarabine).

[0073] FIG. 5: 1H NMR of Asp(Cytarabine-HCl.

DETAILED DESCRIPTION OF THE INVENTION

[0074] The present invention provides pharmaceutically acceptable salt forms of amino acid conjugates covalently linked to therapeutic agents. In particular, the present invention relates to pharmaceutically acceptable salts of conjugates comprising cytidine and aspartic or glutamic acid analog drugs and analogs thereof, pharmaceutical compositions comprising these conjugates, and use thereof for the treatment of cancer or a pre-cancer condition or disorder.

[0075] The art does not teach or suggest methods for creating acid addition salt forms of a drug covalently linked to an amino acid through a side chain functional group using direct transformation modes. The present invention describes that pharmaceutical salts can be created from a drug conjugate precursor having protected groups removable by the same compound (e.g., acid) that is used to generate the pharmaceutical salt form.

Conjugated

[0076] The conjugates of the present invention are provided as pharmaceutically acceptable salt forms of conjugates, which are biologically active as inhibitors of cancer cell proliferation. The salt forms are represented by the structure of formula (I): wherein, A designates an amino acid comprising in its side chain at least one functional group selected from the group consisting of a carboxyl group, an amino group, a sulfhydryl group and a hydroxyl group; D denotes a residue of a drug selected from the group consisting of a cytotoxic agent, a cytostatic agent and a chemotherapeutic agent, wherein D is linked to A through the side chain functional group of A; and Y is selected from the group consisting of: a pharmaceutically acceptable organic or inorganic acid or residue of an acid (i.e., the conjugate base of said acid), a pharmaceutically acceptable inorganic cation, a pharmaceutically acceptable organic amine; and an amino acid.

Pharmaceutical Salt

[0077] The term “pharmaceutical salt” as used herein refers to “pharmaceutically acceptable salts” of drug substances according to the IUPAC conventions. A pharmaceutical salt is an inactive ingredient in salt form combined with a drug. The term “pharmaceutically acceptable salt” as used herein refers to salts of compounds of general formula (I), formula (II) and formula (III), for example, compounds (1), (1A), (1B), (2), (3) or (4), or any other salt form encompassed by the generic formula, which are substantially non-toxic to living organisms. Typical pharmaceutically acceptable salts include those salts prepared by reacting the compounds of the present invention with a mineral, a base, an acid or a pharmaceutically acceptable salt as described herein. Acid salts are also known as acid addition salts.

[0078] Pharmaceutical salts, as they are known in the art (Stahl and Wermuth, 2011, Handbook of pharmaceutical salts, Second Edition), the contents of which are incorporated herein by reference in their entirety, are exemplified herein below in some non-limiting embodiments.

[0079] In the compounds of the present invention, the group Y represents the salt form. Y may be (or may be derived from) any pharmaceutically acceptable organic or inorganic acid or base, non-limiting examples of which are provided below:

(i) Acid Addition Salts:

[0080] In one embodiment, Y is an acid or a pharmaceutically acceptable organic or inorganic residue of an acid, selected from the group consisting of acetic acid, hydrochloric acid, methanesulfuric acid, phosphoric acid, citric acid, lactic acid, succinic acid, tartaric acid, boric acid, benzoic acid, 2-(4-hydroxybenzoyl)-benzoic acid, toluenesulfonic acid, benzenesulfonic acid, ascorbic acid, sulfuric acid, maleic acid, formic acid, malonic acid, nicotinic acid, oxalic acid, camphorsulfonic acid, cyclamic acid, 2,2-dichloroacetic acid, di(t-butyl)-naphthalenesulfonic acid, di(t-butyl)-naphthalenedisulfonic acid, dodecylsulfonic acid, ethane-1,2-disulfonic acid, ethanesulfonic acid, fumaric acid, galactaric (mucic) acid, gentisic acid, glucaric acid, gluconic acid, glycerophosphoric acid, hydrobromic acid, hydroiodic acid, 2-hydroxyethanesulfonic (isethionic) acid, 1-hydroxy-2-naphthoic acid, medonic (bisphosphonic) acid, metaphosphoric acid, methylboronic acid, naphthalene-1,5-disulfonic acid, naphthalene-2-sulfonic acid, nitric acid, orotic acid, 2-oxo-glutaric (ketoglutaric) acid, pamoic (embonic) acid, pyruvic acid, saccharinic acid, salicylic acid, 4-aminosalicylic acid, and thiocyanic acid. Each possibility represents a separate embodiment of the present invention.

[0081] In some currently preferred embodiments, Y is a pharmaceutically acceptable acid selected from the group consisting of acetic acid, hydrochloric acid, methanesulfonic acid, phosphoric acid, citric acid, lactic acid, succinic acid, tartaric acid, boric acid, benzoic acid, toluenesulfonic acid, benzenesulfonic acid, ascorbic acid, sulfuric acid, maleic acid, formic acid, malonic acid, nicotinic acid, and oxalic acid. Each possibility represents a separate embodiment of the present invention.

[0082] Other embodiments of pharmaceutical acid salt forms may be created from acids including aceturic, 4-acetamidobenzoic, adipic, aminohippuric, 4-aminosalicylic, alginic, aspartic, boric, butyric, capric (decanoic), caproic (hexanoic), carbonic, camphoric, camphorsulfonic, caprylic (octanoic), cyclamic, cinnamic, 2,2-dichloroacetic, di(t-butyl)-naphthalenesulfonic, di(t-butyl)-naphthalenedisulfonic, dehydroacetic, diatrizoic, dodecylsulfuric, ethane-1,2-disulfonic, edetic, ethanesulfonic, 2-ethylhexanoic, erythorbic, formic, fumaric, galactaric (mucic), gentisic, glycoheptanoic, gluconic, glucuronic, glutamic, glutaric, glycerophosphoric, glycolic, hippuric, hydrochloric, hydrobromic, iodiodic, 2-(4-hydroxybenzoyl)-benzoic, 2-hydroxy-ethanesulfonic (isethionic), 1-hydroxy-2-naphthoic, isobutyric, lactic, lactobionic, lauric, iodoxamic, isostearic, maleic, malic, malonic, mandelic, medronic, methanesulfonic, metaphosphoric, methylboronic, myristic, naphthalene-1,5-disulfonic, naphthalene-2-sulfonic, nicotinic, oleic, oxalic, palmitic, pentetic, propionic, propanoic, pyroglutamic, pyruvic, phosphoric, sebacic, sorbic, stearic (octadecanoic), suberic, succinic, sulfuric, tartaric, thiazoximic, thiocyanic, toluenesulfonic, trifluoroacetic and undecylenic (undec-10-enoic). Each possibility represents a separate embodiment of the present invention.

(ii) Basic Addition Salts:

[0083] In one embodiment, Y is a pharmaceutically acceptable organic or inorganic base or residue of a base, selected from the group consisting of alkali metals, alkaline earth metals, aluminum, zinc, and ammonium.

[0084] In some currently preferred embodiments, Y is a pharmaceutically acceptable inorganic cation selected from the group consisting of lithium, sodium, potassium, calcium, magnesium, aluminum, zinc, and ammonium. Each possibility represents a separate embodiment of the present invention.

[0085] In another currently preferred embodiment, Y is a pharmaceutically acceptable organic amine selected from the group consisting of ammonium, a primary amine, a secondary amine, a tertiary amine, a quaternary ammonium compound, an amino alcohol, and an amino sugar. In some currently preferred embodiments, Y is an organic amine base selected from the group consisting of benethamine, benzathine, betaine, t-butylamine (erbumine), deanol, dicyclohexylamine, diethylamine, 2-diethylaminoethanol, diethanolamine, ethanolamine, ethylenediamine, hydrabamine, morpholine, 4-(2-hydroxyethyl)morpholine, 1-(2-hydroxyethyl)pyrrolidine (epolamine), imidazole, N-methylglucamine (meglumine), 4-phenylcyclohexylamine, piperazine, and tromethamine. Each possibility represents a separate embodiment of the present invention.

[0086] Other basic forms of pharmaceutical salts can be created from bases including aluminum hydroxide, ammonia, arginine, benetamine, benzathine, betaine, t-butylamine (erbumine), calcium hydroxide, choline hydroxide, deanol, diethylamine, 2-diethylaminoethanol, diethanolamine, ethanolamine, ethylenediamine, hydrabamine, 4-(2-hydroxyethyl)morpholine, 1-(2-hydroxyethyl)pyrrolidine (epolamine), imidazole, lithium hydroxide, lysine, N-methylglucamine (meglumine), magnesium hydroxide, 4-phenylcyclohexylamine, piperazine, potassium hydroxide, sodium hydroxide, tromethamine, and zinc hydroxide. Each possibility represents a separate embodiment of the present invention.

[0087] As understood by one skilled in the art, the aforementioned acids may react with a basic moiety (e.g., an amino group) in the amino acid-drug conjugate to form a salt. For example, when the acid is a carboxylic acid and the conjugate comprises a basic amine moiety, the salt may be represented by the structure R-COO-. R'-NH3+. Similar acid-base reaction products may form with any of the other acids described herein.

(iii) Salts with Amino Acids

[0088] In another current preferred embodiment, Y is an amino acid capable of forming salt forms, such as arginine or lysine. Each possibility represents a separate embodiment of the present invention.

(iv) Other salts

[0089] Other salt dosage forms may be created from salts including ammonium acetate, ammonium sulfate, calcium acetate, calcium carbonate, calcium chloride, calcium glutate, calcium lactate, calcium phosphate, calcium sulfate, calcium disodium edatate, disodium edatate, sodium edatate, iodine, magnesium carbonate, magnesium chloride, magnesium nitrate, magnesium sulfate, potassium acetate, potassium carbonate, potassium citrate, potassium metabisulfite, potassium phosphate, potassium sorbate, sodium acetate, sodium acid pyrophosphate, sodium alginate, sodium alkyl sulfate, sodium aminobenzoate, sodium ascorbate, sodium benzoate, sodium bicarbonate, sodium bisulfate, sodium bisulfate, sodium borate, sodium carbonate, sodium carrageenan, sodium chlorate, sodium chloride, sodium citrate, sodium dithionate, hypochlorite sodium iodide, sodium lactate, sodium metabisulfite, sodium phosphate, sodium pyrophosphate, sodium succinate, sodium sulfate, sodium sulfite, sodium tartrate, sodium thioglycolate, sodium thiosulfate, stannous chloride, zinc acetate, zinc chloride, and zinc sulfate. Each possibility represents a separate embodiment of the present invention.

Amino acids (A)

[0090] According to one embodiment, the amino acid (A) is an L configuration, D configuration, or mixture thereof. Each possibility represents a separate embodiment of the present invention.

[0091] Amino acid A may be an a-amino acid, b-amino acid, g-amino acid, d-amino acid or e-amino acid, with each possibility representing a separate embodiment of the present invention. The currently preferred amino acids are a-amino acids.

[0092] In some embodiments, the amino acid (A) is selected from the group consisting of aminoisobutyric acid (Aib), arginine (Arg), asparagine (Asn), aspartic acid (Asp), citrulline (Cit), cysteine (Cys), cystine, diaminobutanoic acid, diaminobutyric acid (Dab), diaminopropionic acid (Dpr), dihydroxyphenylalanine, dimethylarginine, glutamic acid (Glu), pyroglutamic acid (p-Glu), glutamine (Gln), histidine (His), 1-methyl-histidine, 3-methyl-histidine, homoserine (Hse), homocitrulline, hydroxyproline (Hyp), lysine (Lys), methyl-lysine, dimethyl-lysine, trimethyl-lysine, azidolysine, methionine (Met), methionine sulfoxide, methionine sulfone, ornithine (Orn), sarcosine (Sar), selenocysteine (Sec), serine (Ser), phosphorus-serine, methyl-serine, aminoserine (Ams), thienylalanine (Thi), threonine (Thr), phospho-threonine, tryptophan (Trp), tyrosine (Tyr), methyl-tyrosine, phosphotyrosine, sulfo-tyrosine, a-aminosuberic acid, 3,5-diiodotyrosine, penicillamine (Pen), 4-ethylamine phenylglycine, 4-aminophenylglycine, 4-sulfophenylalanine, 4-aminophenylalanine, and 2-amino-4[4-(2-amino)-pyrimidinyl]butanoic acid, 3-aminopropionic acid, 6-aminohexanoic acid (e-Ahx), p-aminobenzoic acid, isonipecotic, statin (Sta), 2-aminobutyric acid (Abu) and 4-aminobutyric acid, α-aminoadipic acid (Aad), α-aminopimelic acid, g-carboxyglutamic acid, g-hydroxyglutamic acid, aminoglycine and derivatives and analogues thereof. Each possibility represents a separate embodiment of the present invention.

[0093] In some embodiments, the amino acid (A) is selected from the group consisting of asparagine, aspartic acid, glutamine, glutamic acid, and derivatives and analogues thereof. In a currently preferred embodiment, the amino acid (A) is glutamic acid. In another currently preferred embodiment, the amino acid (A) is aspartic acid.

Drug (D)

[0094] Group D can be derived from any drug selected from the group consisting of a cytotoxic agent, a cytostatic agent, and a chemotherapeutic agent. In some embodiments, the cytotoxic, cytostatic, or chemotherapeutic agent D is the residue of a drug selected from the group consisting of a pyrimidine analog, a purine analog, an antifolate, hydroxyurea, an antimicrotubule agent, an alkylating agent, an antitumor antibiotic, a topoisomerase targeting agent, an antimetabolite, a DNA binding agent, and a DNA antagonist.

[0095] In some embodiments, the cytotoxic, cytostatic, or chemotherapeutic agent D is the residue of a pyrimidine analog selected from the group consisting of cytosine arabinoside (ara-C, cytarabine), gemcitabine, fluorouracil, 5-fluorodeoxyuridine, ftorafur, capecitabine, carmofur, BOF-A2, 5-chloro-2,4-dihydroxypyridine, decitabine, leflunomide, trifluridine, iodoxuridine, zidovudine, telbivudine, trimethoprim, fluorocytosine, deoxythymidine, and 5-amino-6-nitrouracil. Each possibility represents a separate embodiment of the present invention.

[0096] In other embodiments, the cytotoxic, cytostatic, or chemotherapeutic agent D is the residue of a purine analog selected from clofarabine, decarbazine, cladribine, mercaptopurine, nelarabine, pentostatin, thioguanine, gefitinib, azathiopurine, 8-azaguanine, immucillin-G, 2-fluoro-2'-deoxyadenosine, 9-beta-D-xylofuranosyl-adenine, 3-dexoxyguanine, 6-methyl-formycin A, 2-fluoroadenosine, 1-deaza-adenosine, N-ethyl-5-carboxamido-adenosine, 3'-oxo-adenosine, 2-amino-adenosine, 6-O-cyclomethyl-guanine, fludarabine, and 8-iodo-guanine. Each possibility represents a separate embodiment of the present invention.

[0097] In other embodiments, the cytotoxic, cytostatic, or chemotherapeutic agent D is the residue of an anticancer drug selected from the group consisting of 2,4-dioxo-5-fluoropyrimidine (5-FU), fluorodeoxyuridine (5-FUDR), azacitidine (5-AZC), 6-mercaptopurine (6-MP), 6-thioguanine (6-TG), chlorodeoxyadenosine (2-CDA), and pentostatin (dCF). Each possibility represents a separate embodiment of the present invention.

[0098] In a currently preferred embodiment, the cytotoxic, cytostatic or chemotherapeutic agent D is the cytarabine residue. In another currently preferred embodiment, the cytotoxic, cytostatic or chemotherapeutic agent D is the gemcitabine residue.

[0099] The present invention also relates to an asparagine drug conjugate wherein the cytotoxic, cytostatic or chemotherapeutic agent drug D is an antimetabolite, which can inhibit enzymes involved in the metabolism of asparagine. An important enzyme that can be inhibited by the asparagine drug conjugate is asparagine synthetase, which is essential for the synthesis of asparagine in mammalian cells. Other enzymes involved in the metabolism of glycoproteins, especially those that have N-linked sugars attached to the asparagine amino acid in the protein including, but not limited to, glucosidase I, glucosidase II, calnexin and alpha-glycosyltransferase can potentially be inhibited by an asparagine drug conjugate. N-glycosylation on protein N-glycans occurs on asparagine in the consensus sequence Asn-X-Ser/Thr, and interference with glycosylation metabolism disrupts glycoprotein folding and secretion. Inhibition of glycosylation of essential glycoproteins will cause cell arrest and cell death.

[00100] An asparagine drug conjugate may also affect the metabolism of other amino acids, such as ornithine, since asparagine has been shown to be involved in the membrane Na+/H+ antiport in the induction of ornithine decarboxylase (Fong and Law, 1988).

[00101] An asparagine drug conjugate can undergo rapid absorption by aspartate and glutamate transporters, which mediate the delivery of glutamic/glutamine and aspartic/asparagine amino acids across the blood-brain barrier (BBB), and can be applied as an efficient drug delivery system for transporting chemotherapeutic drugs to treat CNS tumors. It has been demonstrated that cerebrospinal fluid (CSF) and plasma levels of asparagine are significantly lower in patients with primary and secondary Central Nervous System (CNS) tumors (Piek et al., 1987).

[00102] Hepatoma cancer cells have been shown to express a glutamine transporter, which shows a much higher rate of glutamine uptake in human hepatoma cells than in normal hepatocytes, indicating that the drug-glutamine conjugate can be used for the treatment of liver cancers.

[00103] As noted above, in the conjugates of the present invention, the drug residue (D) is linked to the amino acid (A) through the side chain functional group of (A). Some non-limiting embodiments of such conjugates are described below.

[00104] In specific embodiments, the compound comprises the amino acid residue of aspartic acid or glutamic acid conjugated to a drug selected from the group consisting of purine analogs and pyrimidine analogs. In specific embodiments, the drug is selected from cytarabine and gemcitabine. In specific embodiments, the salt form of the amino acid-drug conjugate is an acid salt form selected from acetic acid and hydrochloric acid.

[00105] Without wishing to be bound by theory, the conjugated amino acid salts of the present invention are transported into the cell via amino acid transporters thereby bypassing multidrug resistance (MDR) mechanisms, and either prevent cell growth or kill the cell from within.

[00106] According to some embodiments, the amino acid (A) is selected from the group consisting of aspartic acid and glutamic acid, and the compound is represented by the structure of formula (II) or (III) and Y are as defined above.

[00107] In a specific embodiment of formula (II), A is aspartic acid and D is the cytarabine residue and the compound is represented by the structure of formula (1):

[00108] In one embodiment of formula (1), the compound is a salt with acetic acid (i.e., an acetate salt) that is represented by the structure of formula (1A):

[00109] In another embodiment of formula (1), the compound is a salt with hydrochloric acid (i.e., a hydrochloride salt), which is represented by the structure of formula (1B):

[00110] In another specific embodiment of formula (II), A is aspartic acid and D is the residue of gemcitabine and the compound is represented by the structure of formula (2):

[00111] In a specific embodiment of formula (III), A is glutamic acid and D is the cytarabine residue and the compound is represented by the structure of formula (3):

[00112] In another specific embodiment of formula (III), A is glutamic acid and D is the gemcitabine residue and the compound is represented by the structure of formula (4):

[00113] In any of the above compounds of formula (1), (2), (3) and (4), Y may be any of the options described above, it may preferably be a pharmaceutically acceptable acid selected from the group consisting of acetic acid, hydrochloric acid, methanesulfonic acid, phosphoric acid, citric acid, lactic acid, succinic acid, tartaric acid, boric acid, benzoic acid, toluenesulfonic acid, benzenesulfonic acid, ascorbic acid, sulfuric acid, maleic acid, formic acid, malonic acid, nicotinic acid and oxalic acid.

Definitions

[00114] For convenience and clarity, certain terms employed in the specification, examples, and claims are described herein.

[00115] The term “drug residue” refers to a drug excluding the functional group that was used to attach the amino acid in the formation of the amino acid-drug conjugate A-D.

[00116] The term “residue of an acid” refers to the conjugate base of that acid. For example, the conjugate base of hydrochloric acid (HCl) is chloride (Cl-). The conjugate base of acetic acid (CH3COOH) is acetate (CH3COO-).

[00117] The term “drug” denotes any pharmacologically active agent capable of arresting cell growth, or inducing death of the hyperproliferative cell in which it is present and includes known cytotoxic, cytostatic, antiproliferative drugs, as are known in the art, as exemplified above. The classification of drugs contained herein is for convenience only and is not intended to limit any component to a particular application or applications listed.

[00118] The term “cytotoxic agent,” as used herein, refers to an agent that is toxic to, damages, and/or destroys, cells, e.g., cancer cells. Chemotherapy by chemotherapy drugs is a form of cytotoxic therapy.

[00119] The term “cytostatic drug,” as used herein, refers to any substance that inhibits cell growth and division.

[00120] The term “chemotherapeutic agent,” as used herein, refers to a drug used in chemotherapy.

[00121] As used herein, “stopping cell growth” and “inducing death” of the hyperproliferative cell, e.g., neoplastic cell, refers to slowing, interrupting, halting, or stopping its growth and metastasis and does not necessarily indicate a total elimination of neoplastic growth.

[00122] The amino acids used in this invention are those that are commercially available or are available by routine synthetic methods. Coded natural amino acids and their derivatives are represented by three-letter codes according to IUPAC conventions. When there is no indication, the L or D isomer may be used. D isomers are indicated by “D” before the residue abbreviation.

[00123] As used herein, the term “stereoisomer” refers to a compound consisting of the same atoms bonded by the same bonds but having different three-dimensional structures, which are not interchangeable. Three-dimensional structures are called configurations. As used herein, the term “enantiomer” refers to two stereoisomers whose molecules are non-superimposable mirror images of each other. The term “chiral center” refers to a carbon atom to which four different groups are attached. As used herein, the term “diastereoisomers” refers to stereoisomers, which are not enantiomers. In addition, two diastereoisomers, which have a different configuration at only one chiral center, are referred to herein as “epimers”. The terms “racemate”, racemic mixture or “racemic modification” refer to a mixture of equal parts of enantiomers. The present invention includes enantiomers of the compounds of general formula (I), (II) and (III), and the specific compounds described herein.

Synthetic Methods

[00124] In the conjugate of the present invention, the drug is covalently linked to the amino acid or amino acid analogue. One skilled in the art will be able to optimize the appropriate binding and position of the drug moiety within the compound. Several concerns should be taken into consideration to guide the skilled in the art in this decision, such as selection of the specific drug, selection of derivatives, selection of the binding position to the drug species, and requirements for the host intracellular enzymes for activation of the drug.

[00125] Thus, according to another aspect, the present invention provides a method for preparing a pharmaceutically acceptable salt represented by the structure of formula I, comprising the step of

[00126] (a) reacting a compound of formula (IV): wherein A and D are as defined above, n is an integer from 1 to 6; and each P is independently at each occurrence a protecting group; wherein each P is attached to any functional group available for protection on the amino acid (A), the drug residue (D), or on both the amino acid (A) and the drug residue (D); with a reagent capable of removing the protecting group(s) P; and (b) adding a reagent capable of introducing the group Y, so as to generate a salt represented by the formula: where Y is as defined above.

[00127] As mentioned above, it has now unexpectedly been discovered that the salts of the present invention can be prepared by deprotecting a precursor compound containing one or more protecting groups (P) using the same reagent that ultimately forms the salt. Thus, in accordance with the principles of the present invention, the reagent capable of introducing the group Y can simultaneously remove the protecting group(s) P and generate the salt of formula (I) in one step, thereby obviating an intermediate deprotection step prior to salt formation, which ultimately leads to a pharmaceutical product of higher purity. Specifically, the use of TFA for BOC deprotection is avoided, leading to the generation of a salt of the conjugate of the present invention represented by formula I, which salt is devoid or free of toxic TFA. Specific embodiments of this process are described below. The compounds described herein comprise a pharmaceutical salt form of an anticancer drug conjugated to an amino acid or amino acid derivatives or analogues. The compounds of the present invention can be readily prepared from amino acids by methods familiar to one of skill in the art. For example, the conjugates or their precursors can be prepared according to the methods described in International Patent Application Publication No. WO 2005/072061 and in US Patent No. 7,989,188, the contents of which are incorporated herein by reference. The conjugates (or their protected precursors) can then be reacted with a reagent capable of introducing the Y group (e.g., the reagent is an acid or base) to form the compounds of formula (I). Additional methods useful for preparing the compounds of the invention are, for example, the methods described in International Patent Application Publication Nos. WO96/30036 and WO97/36480, and in US Patent Nos. 5,643,957 and 5,650,386, among others. For example, the compounds can be prepared by reacting the single amino acid with the appropriate catalytic agent, which reacts with a free moiety present on the amino acid and conjugating it to a free moiety on the drug to form an ester or an amide. Protecting groups can be used to prevent undesirable side reactions, as would be known to those skilled in the art. The contents of each of the above references are incorporated herein by reference.

[00128] For example, for purposes of illustration and not limitation, conjugates of formula (II) (aspartic acid-drug conjugates) or formula (III) (glutamic acid-drug conjugates) can be prepared by reacting aspartic acid or glutamic acid that are protected at the amine and alpha carboxyl (compound (i) and (iii) in Schemes 1A and 1B), with the activated and/or protected drug or a derivative thereof in the presence of a coupling reagent, followed by deprotection. Drug (D) can be deprotected or protected at any available functional group (e.g., OH, NH2, SH, carboxyl, etc.), with any of the protecting groups described herein. Exemplary processes are depicted in Schemes 1A and 1B: where P1 is a carboxyl protecting group, P2 is an amino protecting group, P3 is a protecting group located on one or more functional groups on the drug D and n is 0, 1, 2, or 3. Depending on the number of functional groups on the drug, n could also be 4, 5, 6, etc.

[00129] In one embodiment, the drug is cytarabine. In other embodiments, the drug is protected cytarabine comprising protecting groups on its 2’, 3’, and/or 5’ hydroxyl units, such as triphenylmethyl-cytarabine (CAS 7075-13-0), benzoyl-cytarabine (CAS 34270-10-5), admantoyl-cytarabine (CAS 23113-01-1), and trimethylsilyl-cytarabine. In other embodiments, the drug is gemcitabine. In other embodiments, the drug is a protected gemcitabine comprising protecting groups on its 3’ and/or 5’ hydroxyl units, such as triphenylmethyl-gemcitabine (CAS 1642862-24-5).

[00130] A non-limiting example of a protected drug is a protected cytarabine, wherein at least one of the 2', 3' or 5' hydroxyls is protected with a P3 group as defined above. In a non-limiting embodiment described below for the purpose of illustration, the compound is represented by formula E, wherein each of the 2', 3' or 5' hydroxyls is protected with a P3 group.

[00131] In other embodiments, only the 5’-OH group is protected, while the 2’ and 3’ hydroxyls are unprotected.

[00132] Another non-limiting example of a protected drug is a protected gemcitabine, wherein at least one of the 3' or 5' hydroxyls is protected with a P3 group as defined above. In a non-limiting embodiment described below for the purpose of illustration, the compound is represented by formula F, wherein each of the 3' or 5' hydroxyls is protected with a P3 group.

[00133] In other embodiments, only the 5’-OH group is protected, while the 3’-hydroxyl is not unprotected.

[00134] In one embodiment, referred to herein as “Route A”, the deprotection step is conducted in the presence of trifluoroacetic acid (TFA), followed by reaction of an acid to introduce the Y group. According to this embodiment, the present invention provides a pharmaceutically acceptable salt preferably comprising less than about 5% TFA as a residual impurity, preferably less than about 1% TFA, more preferably less than about 0.1% TFA. Most preferably, the salt of the present invention is substantially devoid of TFA as an impurity.

[00135] In another embodiment, referred to herein as “Route B”, the deprotection step is conducted directly with the reagent capable of introducing the Y group (i.e., an acid) to directly provide the salt of the present invention. According to this embodiment, the protecting groups are removed simultaneously with the introduction of the Y group, and the resulting pharmaceutically acceptable salt is completely devoid of TFA as an impurity.

Protection Groups (P)

[00136] The precursors of compound (I) may include one or more protecting groups (P), exemplified above as P1, P2 and P3, which may be any protecting group known to one skilled in the art. The term “protecting group” refers to chemical residues used to block reactive sites during chemical synthesis, which allow the chemical reaction to be carried out selectively at one reaction site in a multifunctional compound, other reactive sites must be temporarily blocked. The residues used to block these reactive sites called protecting groups.

[00137] The protecting group may be a hydroxyl protecting group, an amino protecting group, a carboxy protecting group, etc. As used herein, the term “OH protecting group” or “hydroxy protecting group” refers to a readily cleavable group attached to hydroxyl groups. As used herein, the term “NH protecting group” or “amino protecting group” refers to a readily cleavable group attached to amino groups. As used herein, the term “carboxy protecting group” refers to a readily cleavable group attached to carboxy groups.

[00138] According to one embodiment, the protecting group P is selected from the group acetamidomethyl (Acm), acetyl (Ac), acetonide, adamantyloxy (AdaO), alpha-allyl (OAll), Alloc, benzoyl (Bz), benzyl (Bzl), benzyloxy (BzlO), benzyloxycarbonyl (Z), benzyloxymethyl (Bom), bis-dimethylamino (NMe2), 2-bromobenzyloxycarbonyl (2-Br-Z), t-butoxy (tBuO), t-butoxycarbonyl (Boc), t-butoxymethyl (Bum), t-butyl (tBu), t-butylthio (tButio), 2-chlorobenzyloxycarbonyl (2-Cl-Z), 2-chlorotrityl (2-Cl-Trt), cyclohexyloxy (cHxO), 1-cyclopropyl-1-methyl-ethyl (Dmcp) initrophenyl (Dnp), fluorenylmethoxycarbonyl (Fmoc), formyl (For), Mesitylene-2-sulfonyl (Mts), 4-methoxybenzyl, 4-methoxy-2,3,6-trimethylbenzenesulfonyl (Mtr), 4-methoxytrityl (Mmt), 4-methylbenzyl (MeBzl), 3-methylpent-3-yl (Mpe), 1- methyl-1-phenyl-ethyl (PhiPr), Methyl, 4-methyltrityl (Mtt), 3-nitro-2-pyridinesulfenyl (Npys), 2,2,4,6,7-pentamethyl-dihydrobenzofuran-5-sulfonyl (Pbf), 2,2,5,6,8-pentamethyl-chroman-6-sulfonyl (Pmc), Tosyl (Tos), Trifluoroacetyl (Tfa), Trimethylacetamidomethyl (Tacm), Triphenylmethyl (Trityl, Trt) and Xanthyl (Xan). Each possibility represents a separate embodiment of the present invention.

[00139] A non-limiting example of a hydroxyl protecting group is an acyl group (COR where R = alkyl, aryl, etc.). A presently preferred acyl group is an acetyl group (i.e., OR' = acetate, OAc). Another example of a hydroxy protecting group is a silyl group, which may be substituted with alkyl (trialkylsilyl), with an aryl (triarylsilyl), or a combination thereof (e.g., dialkylphenylsilyl). A preferred example of a silyl protecting group is trimethylsilyl (TMS) or di-t-butyldimethylsilyl (TBDMS), Triisopropylsilyl (TIPS), triethylsilyl (TES). Other examples of hydroxy protecting groups include, for example, C1-C4 alkyl, -CO-(C1-C6 alkyl), -SO2-(C1-C6 alkyl), -SO2-aryl, -CO-Ar, where Ar is an aryl group as defined above, and -CO-(C1-C6 alkyl)Ar (for example, a carboxybenzyl (Bz) group).

[00140] Examples of amino protecting groups include t-butoxycarbonyl (BOC), benzyloxycarbonyl, acetyl, phenylcarbonyl or a silyl group, which may be substituted with alkyl (trialkylsilyl), with an aryl (triarylsilyl) or a combination thereof (e.g. dialkylphenylsilyl), e.g. trimethylsilyl (TMS) or t-butyldimethylsilyl (TBDMS). Other examples of hydroxy protecting groups include, for example, C1-C4 alkyl (e.g., methyl, ethyl, propyl, butyl, and the like), -CH2Ph (benzyl or bzl), allyl (All), (allyl)-CO-(C1-C6 alkyl), -SO2-(C1-C6 alkyl), -SO2-aryl, -CO-Ar, wherein Ar is an aryl group as defined above, and -CO-(C1-C6 alkyl)Ar (e.g., a carboxybenzyl group (Bz)). Other examples of hydroxy protecting groups include acid-sensitive protecting groups such as tetrahydropyranyl (THP), methoxymethyl (MOM), triphenylmethyl (Trityl), and dimethoxytrityl (DMT). Each possibility represents a separate embodiment of the present invention.

[00141] Representative carboxy protecting groups include, but are not limited to, methyl, ethyl, t-butyl, benzyl (Bn), p-methoxybenzyl (PMB), 9-fluorenylmethyl (Fm), trimethylsilyl (TMS), t-butyldimethylsilyl (TBDMS), diphenylmethyl (benzhydryl, DPM), and the like. Protecting groups may be removed by deprotecting agents which, in accordance with the principles of the present invention, are preferably the same as the pharmaceutical salt moiety. In certain embodiments, pharmaceutical acids suitable for removing acid-labile protecting groups are: acetic, aceturic, 4-acetamidobenzoic, adipic, aminohippuric, 4-aminosalicylic, ascorbic, alginic, aspartic, benzenesulfonic, benzoic, boric, butyric, capric (decanoic), caproic (hexanoic), carbonic, citric, camphoric, camphorsulfonic, caprylic (octanoic), cyclamic, cinnamic, 2,2-dichloroacetic, di(t-butyl)-naphthalenesulfonic, di(t-butyl)-naphthalenedisulfonic, dehydroacetic, diatrizoic, dodecylsulfuric, ethane-1,2-disulfonic, edetic, ethanesulfonic, 2-ethylhexanoic, erythorbic, formic, fumaric, galataric (mucic), gentisic, glucaric, glucoheptanoic, gluconic, glucuronic, glutamic, glutaric, glycerophosphoric, glycolic, hippuric, hydrochloric, hydrobromic, hydroiodic, 2-(4-hydroxybenzoyl)-benzoic, 2-hydroxyethanesulfonic (isethionic), 1-hydroxy-2-naphthoic, isobutyric, lactic, lactobionic, lauric, iodoxamic, isostearic, maleic, malic, malonic, mandelic, medronic, methanesulfonic, metaphosphoric, methylboronic, myristic, naphthalene-1,5-disulfonic, naphthalene-2-sulfonic, nicotinic, nitric, oleic, orotic, oxalic, 2-oxo-glutaric (ketoglutaric), palmitic, pamoic (embonic), pentetic, propionic, propanoic, pyroglutamic, pyruvic, phosphoric, saccharin, salicylic, sebacic, sorbic, stearic (octadecanoic), suberic, succinic, sulfuric, tartaric, thiazoximic, thiocyanic, toluenesulfonic, trifluoroacetic and undecylenic (undec-10-enco). Each possibility represents a separate embodiment of the present invention.

[00142] In specific embodiments, the pharmaceutically acceptable acid that reacts with the precursor compound of formula (IV) and removes the protecting group is selected from the group consisting of acetic acid, methanesulfonic acid, hydrochloric acid, phosphoric acid, citric acid, lactic acid, succinic acid, tartaric acid, boric acid, benzoic acid, toluenesulfonic acid, benzenesulfonic acid, ascorbic acid, sulfuric acid, maleic acid, formic acid, malonic acid, nicotinic acid, and oxalic acid. According to such embodiments, the resulting salt is selected from the group consisting of acetate, hydrochloride, methanesulfonate, phosphate, citrate, lactate, succinate, tartrate, borate, benzoate, toluenesulfonate, benzensulfonate, ascorbate, sulfate, maleate, formate, malonate, nicotinate, and oxalate. Each possibility represents a separate embodiment of the present invention.

[00143] In certain embodiments, suitable pharmaceutical bases for removing base-labile protecting groups are: aluminum hydroxide, ammonia, arginine, benzamine, benzathine, betaine, t-butylamine (erbumine), calcium hydroxide, choline hydroxide, deanol, diethylamine, 2-diethylaminoethanol, diethanolamine, ethanolamine, ethylenediamine, hydrabamine, 4-(2-hydroxyethyl)morpholine, 1-(2-hydroxyethyl)pyrrolidine (epolamine), imidazole, lithium hydroxide, lysine, N-methylglucamine (meglumine), magnesium hydroxide, 4-phenylcyclohexylamine, piperazine, potassium hydroxide, sodium hydroxide, tromethamine, and zinc hydroxide. Each possibility represents a separate embodiment of the present invention.

[00144] In other embodiments, H2 Pd/C may be used as the deprotecting agent when the protecting group is, for example, benzyl.

[00145] Precursor compounds containing protecting groups can generally be described in Schemes 1A and 1B above.

[00146] In specific embodiments, the precursor is Boc-Asp(Cytarabine)-OtBu (formula A), while the protecting groups are removable by an acid to yield Asp(Cytarabine)•Y (formula 1), while Y is an acid salt form of a pharmaceutical acid.

[00147] In specific embodiments, the precursor is Boc-Asp(Gemcitabine)-OtBu (formula B), wherein the protecting groups are removable by an acid to yield Asp(Gemcitabine)•Y (formula 2), wherein Y is an acid salt form of a pharmaceutical acid.

[00148] In other specific embodiments, the precursor is Boc-Glu(Cytarabine)-OtBu (formula C), while the protecting groups are removable by an acid to yield Glu(Cytarabine) •Y (formula 3), while Y is an acid salt form of a pharmaceutical acid.

[00149] In other specific embodiments, the precursor is Boc-Glu(Gemcitabine)-OtBu (formula D), wherein the protecting groups are removable by an acid to yield Glu(Gemcitabine)•Y (formula 4), wherein Y is an acid salt form of a pharmaceutical acid. Other suitable protecting groups are described, for example, in Wuts and Greene, 2007, “Greene’s protecting groups in organic synthesis”, Fourth Edition, the contents of which are incorporated herein by reference.

[00150] While it is preferred that the protecting group be removed by a pharmaceutical compound that creates a salt, the present invention is not limited to such embodiments. It is apparent to one skilled in the art that the protecting group can be removed by other deprotection agents, and the salt form of the drug can be generated in a separate subsequent step.

[00151] The drug may be used as such or as an activated or protected derivative thereof. For example, one embodiment of the present invention includes the use of cytarabine or gemcitabine that are protected at the hydroxyl groups of the sugar moieties (C2, C3 and/or C5) so as to prevent or minimize the formation of a diamino acid-Cytarabine or diamino acid-Gemcitabine byproduct (i.e., a second amino acid attached to the 5'-OH of the arabinose moiety). The protecting group on the sugar moiety may be any of the protecting groups described above and may be removed with the appropriate deprotection agent as described herein.

[00152] The principles that apply to the selection of the organic/amino acid binding site, protecting groups and pharmaceutical salt form, etc., will be detailed here for exemplary compounds. The principles may be generalized as follows: a) Selection of the organic acid/amino acid or amino acid derivative or analogue: aspartate/asparagine and glutamate/glutamine and any of their derivatives or analogues that can undergo uptake by malignant or cancerous cells are suitable, as are other organic acids that have improved drug solubility, stability, bioavailability and patient compliance; b) Selection of the organic acid/amino acid protecting groups: the groups may be selected from protecting groups suitable for the chemically functional groups on the organic acid/amino acid that can be removed by a compound that creates a pharmaceutical salt, such as other removable protecting groups; c) Selection of the drug: the drug may be selected from antiproliferative agents, cytotoxic agents and cytostatic agents; d) Selection of the drug protecting groups: The groups may be selected from suitable protecting groups for the chemically functional groups on the drug that can be removed by a compound that creates a pharmaceutical salt, such as other removable protecting groups; e) Selection of the reaction conditions for the conjugation of the drug to the organic acid: The optimum solvents, catalysts, additives, and the time and temperature of the various reaction steps. f) Selection of the conditions for removing the protecting groups: By a pharmaceutical salt or other suitable compound for removing protecting groups and scavengers, as is known to those skilled in the art (Wuts and Greene, 2007). g) Selection of the pharmaceutical salt: The salt may be selected from a mineral, an additional acid salt, a base salt, and a combination thereof. h) Selection of the conditions for creating the pharmaceutical salt: The optimum solvents/antisolvents, the concentrations, and the time and temperature of the reaction.

[00153] Compounds and conditions for the conjugation reaction are exemplified, but not limited to, by the following components: Solvents: dimethylformamide (DMF), dimethylacetamide (DMA), ethanol, diethyl ether, dimethyl ethyl acetate sulfoxide (DMSO), dichloromethane (DCM), dichloroethane (DCE), heptane, hexane, pyridine, 2-propanol, methanol, tert-butylmethyl ether, N-methylpyrrolidone (NMP), tetrahydrofuran (THF), trifluoroethanol (TFE), water, and combinations thereof; Catalysts/coupling reagents: BOP (benzotriazol-1-yl-oxy-tris-(dimethylamino)-phosphonium hexafluorophosphate), CDI (N,N'-carbonyldiimidazole), COMU (1-[(1-(Cyano-2-ethoxy-2-oxoethylideneaminooxy)dimethylaminomorpholino)]uranium), DCC (N,N'-dicyclohexylcarbodiimi da), DIC (N,N'- diisopropylcarbodiimide), EDC (1-ethyl-3-(3-dimethylaminopropyl)carbodiimide), DEPBT (3-(diethoxyphosphoryloxy)-1,2,3-benzo[d]triazin-4(3H)-one), HATU, HBTU hexetluorophosphate de (2-(1H-benzotriazol-1-yl)- - phosphonium); Additives: DIPEA (diisopropylethylamine), DMAP (dimethylaminopyridine), pyridine, triethylamine, HOAt (N-hydroxy-9-azabenzotriazole), HOBt (N-hydroxybenzotriazole), pure oxime, NaclO4, LiCl, KSCN and Triton.

[00154] Reaction temperatures of about 0°C to about 80°C, preferably about 4°C to about 25°C; Reaction times of about 3 minutes to about 72 hours, preferably about 3 to about 24 hours; Compounds and conditions for removing protecting groups are exemplified, but not limited to, the following components: Pharmaceutical salts: including pharmaceutically acceptable mineral, base, acid or salt; the preferred acid and salt for removing protecting groups have a pKa ≥ 3 and the preferred base and salt have a pKa > 9.

[00155] The preferred acid for removing protecting groups having a pKa £ 3 is selected from the group consisting of: benzenesulfonic, camphorsulfonic, cyclamic, 2,2-dichloroacetic, di(t-butyl)-naphthalenesulfonic, di(t-butyl)-naphthalenedisulfonic, dodecylsulfuric, ethane-1,2-disulfonic, ethanesulfonic, fumaric, galactaric (mucic), gentisic, glucaric, gluconic, glycerophosphoric, hydrochloric, hydrobromic, hydroiodic, 2-hydroxy-ethanesulfonic (isethiononic), 1-hydroxy-2-naphthoic, maleic, malonic, medronic (bisphosphonic), methanesulfonic, methophosphoric, methylboronic, naphthalene-1,5-disulfonic, naphthalene-2-sulfonic, nitric, orotic, oxalic, 2-oxo-glutaric (ketoglutaric), pamoic (embonic), pyruvic, phosphoric, saccharin, salicylic, sulfuric, tartaric, thiocyanic, toluenesulfonic and trifluoroacetic acids.

[00156] Other protecting group removing compounds including, but not limited to: TMSBr, HBF4, HF, piperidine, Pd(Ph3P)4, NH2NH2, I2 and Tl, Hg, Ag and Pd compounds; Scavengers: H2O, TIS, TES, EDT, m-cresol, thioanisole, anisole, phenol; Temperatures from -20°C to 80°C, preferably from 0°C to 25°C; Times from 1 minute to 24 hours, preferably 1-3 hours; Compounds and conditions for creating pharmaceutical salt are selected from: Solvents for dissolving the compound with the pharmaceutical acid: H2O, dioxane, ethyl acetate, methanol, ethanol, isopropanol, DMSO, tetrahydrophorane (THF) and trifluoroethanol (TFE) or any combination thereof. In a currently preferred embodiment, the solvent is H2O. In another currently preferred embodiment, the solvent is dioxane; Anti-solvent to precipitate the product indicated in formula (I): diethyl ether, t-butyl methyl ether (tBME), hexane, heptane, DCM.

[00157] Molar ratio of 1:1 to 1:30 of compound to pharmaceutical salt, preferably 1:1.1 to 1:3, at room temperature (25°C), incubated for 1 hour.

[00158] This invention further describes hydrates of the pharmaceutically acceptable salts of general formula (I).

[00159] In a particular embodiment, the pharmaceutically acceptable acid is HCl and the solvent used is H2O or dioxane. It has unexpectedly been found that when dioxane/HCl is used, the product precipitates during the deprotection reaction, and there is no need for a separate precipitation step.

Pharmaceutical Compositions

[00160] The present invention further provides pharmaceutical compositions comprising at least one of the compounds of the present invention and one or more pharmaceutically acceptable excipients or diluents.

[00161] Compositions comprising the compounds of the present invention have utility in delivering active agents to selected biological systems and in an increased or improved bioavailability of the active agent compared to administration of the active agent without the delivery agent. Delivery may be improved by delivering more active agent over a period of time, or by delivering active agent in a specific period of time (e.g., faster or delayed delivery) or over a period of time (such as extended delivery).

[00162] The pharmaceutical compositions of the present invention can be manufactured by processes well known in the art, for example, by means of conventional mixing, dissolving, granulating, grinding, pulverizing, dragee production, levigation, emulsification, encapsulation, capture or lyophilization processes.

[00163] As used herein, a “pharmaceutical composition” refers to a preparation of one or more of the compounds described herein, or their physiologically acceptable salts or solvents, with other chemical components, such as physiologically suitable carriers and excipients. The purpose of a pharmaceutical composition is to facilitate the administration of a compound to a subject.

[00164] The pharmaceutical compositions of the invention may be administered by any suitable route of administration selected from the group consisting of parenteral and oral routes of administration. According to some embodiments, the route of administration is parenteral route of administration. According to some embodiments, the route of administration is intravenous, intraarterial, intramuscular, subcutaneous, intraperitoneal, intracerebral, intracerebroventricular, intrathecal, or intradermal route of administration. For example, the pharmaceutical compositions may be administered systemically, for example, by intravenous (i.v.) or intraperitoneal (i.p.) injection or infusion. According to a certain embodiment, the pharmaceutical composition is administered by intravenous infusion over 30 minutes to 2 hours, such as over 1 hour. The compositions of the invention may be administered locally and may further comprise an additional active agent and/or excipient.

[00165] The pharmaceutical compositions for use according to the present invention may thus be formulated in conventional manner using one or more physiologically acceptable carriers comprising excipients and auxiliaries, which facilitate the processing of the active compounds into preparations which can be used pharmaceutically. The appropriate formulation depends on the route of administration chosen.

[00166] For injection, the compounds of the invention may be formulated in aqueous solutions, preferably in physiologically compatible buffers such as Hank's or Ringer's solution or physiological saline buffer. For transmucosal and transdermal administration, penetrants appropriate to the barrier to be permeated may be used in the formulation. Such penetrants, including, for example, DMSO or polyethylene glycol, are known in the art.

[00167] For oral administration, the compounds can be readily formulated by combining the active compounds with pharmaceutically acceptable carriers well known in the art. Such carriers enable the compounds of the invention to be formulated as tablets, pills, dragees, capsules, liquids, gels, syrups, slurries, suspensions and the like, for oral ingestion by a patient. Pharmacological preparations for oral use can be made using a solid excipient, optionally grinding the resulting mixture, and processing the granule mixture, after the addition of suitable auxiliaries, if desired, to obtain tablets or dragee cores. Suitable excipients are, in particular, fillers, such as, sugars, including lactose, sucrose, mannitol or sorbitol; cellulose preparations, such as, for example, corn starch, wheat starch, rice starch, potato starch, gelatin, gum tragacanth, methyl cellulose, hydroxypropylmethyl cellulose, sodium carbomethyl cellulose; and/or physiologically acceptable polymers such as polyvinylpyrrolidone (PVP). If desired, disintegrating agents such as cross-linked polyvinylpyrrolidone, agar or alginic acid or a salt thereof such as sodium alginate may be added.

[00168] Furthermore, enteric coating is useful because it is desirable to avoid exposure of the compounds of the invention to the gastric environment.

[00169] Pharmaceutical compositions, which can be used orally, include softgel capsules made of gelatin, as well as soft, sealed capsules made of gelatin and a plasticizer such as glycerol or sorbitol. The softgel capsules may contain the active ingredients in admixture with fillers such as lactose, binders such as starches, lubricants such as talc or magnesium stearate and, optionally, stabilizers.

[00170] In soft capsules, the active compounds may be dissolved or suspended in suitable liquids, such as fatty oils, liquid paraffin or liquid polyethylene glycols. In addition, stabilizers may be added. All formulations for oral administration must be in dosages appropriate for the chosen route of administration.

[00171] For buccal administration, the compositions may take the form of tablets or lozenges formulated in a conventional manner.

[00172] For administration by inhalation, the active compound for use according to the present invention is conveniently dispensed in the form of an aerosol spray presentation from a pressurised pack or nebuliser using a suitable propellant, for example dichlorodifluoromethane, trichlorofluoromethane, dichlorotetrafluoroethane or carbon dioxide. In the case of a pressurised aerosol, the dosage unit may be determined by providing a valve to deliver a metered amount. Capsules and cartridges of, for example, gelatin for use in an inhaler or insufflator may be formulated containing a powder mix of the peptide and a suitable powder base, such as lactose or starch.

[00173] Pharmaceutical compositions for parenteral administration include aqueous solutions of the active ingredients in water-soluble form. Additionally, suspensions of the active compounds may be prepared as suitable oily injection suspensions. Suitable lipophilic solvents or vehicles include fatty oils, such as sesame oil, or synthetic fatty acid esters, such as ethyl oleate, triglycerides or liposomes. Aqueous injection suspensions may contain substances which increase the viscosity of the suspension, such as sodium carboxymethyl cellulose, sorbitol or dextran. Optionally, the suspension may also contain suitable stabilizers or agents which increase the solubility of the compounds, to enable the preparation of highly concentrated solutions.

[00174] The active compounds of the present invention may also be formulated into rectal compositions, such as suppositories or retention enemas, using, for example, conventional suppository bases, such as cocoa butter or other glycerides.

[00175] Depending on the severity and responsiveness of the condition being treated, the dosage may also be a single administration of a slow release composition, with a course of treatment lasting from several days to several weeks or until a cure is effected or abatement of the disease state is achieved. The amount of a composition to be administered will, of course, be dependent on many factors including the individual being treated, the severity of the condition, the mode of administration, and the judgment of the attending physician.

[00176] According to a certain embodiment, the formulations of the pharmaceutically acceptable salts of the invention are provided as an aqueous isotonic solution having an osmolarity of about 300 mOsm and a pH of 4-8. Thus, the pharmaceutically acceptable carrier of the compound of the invention can be a buffered saline solution, a buffered dextrose solution, or a buffered glycerol solution having an osmolarity of about 300 mOsm and a pH of 4-8.

[00177] In another embodiment, the buffer solution may be a pharmaceutically acceptable mono-ionic buffer system or a poly-ionic buffer system having an ionization pK in the range of 4-8.

[00178] Various buffers having a pK of 4-8 may be employed to adjust the pH of the solution containing the pharmaceutically acceptable salt of the present invention, such as, for example, ACES (N-(acetamido)-2-aminoethanesulfonic acid); Acetate; N-(2-acetamido)-2-iminodiacetic acid; BES (N,N-bis[2-hydroxyethyl]-2-aminoethanesulfonic acid); Bicine (2-(bis(2-hydroxyethyl)amino)acetic acid); Bis-Trismethane (2-[bis(2-hydroxyethyl)amino]-2-(hydroxymethyl)propane-1,3-diol); BisTris propane (1,3-bis(tris(hydroxymethylmethylamino)propane); Carbonate; Citrate; 3,3-dimethylglutarate; DIPSO (3-[N,N-bis(2-hydroxyethyl)amino]-2-hydroxypropansulfonic acid); N-ethylmorpholine; Glycerol-2-phosphate; Glycine; Glycine-amide; HEPBS (N-(2-hydroxyethyl)piperazin-N’-4-butanesulfonic acid); HEPES (N-(2-hydroxyethyl)piperazin-N’-2-ethanesulfonic acid); HEPPS (N-(2-hydroxyethyl)piperazin-N’-(3-propanesulfic acid)); HEPPSO (N-(2-hydroxyethyl)piperazin-N’-(2-hydroxypropanesulfonic acid); Histidine; Hydrazine; Imidazole; Maleate; 2-methylimidazole; MES (2-(N-morpholino)ethanesulfonic acid); MOBS (4-(N-morpholino)-butanesulfonic acid); MOPS (3-(N-morpholino)-propanesulfonic acid; MOPSO (3-(N-morpholino)-2-hydroxypropanesulfonic acid); Oxalate; Phosphate; Piperazine; PIPES (1,4-Piperazine-diethanesulfonic); POPSO (piperazine-N,N’-bis(2-hydroxypropanesulfonic acid)); Succinate; Sulfite; TAPS (3-[[1,3-dihydroxy-2-(hydroxymethyl)propan-2-yl]amino]propane-1-sulfonic acid); TAPSO (3-[[1,3-dihydroxy-2-(hydroxymethyl)propan-2-yl]amino]-2-hydroxypropane-1-sulfonic acid); TES (2-[[1,3-dihydroxy-2-(hydroxymethyl)propan-2-yl]amino]ethanesulfonic acid); THAM (Tris) (2-Amino-2-hydroxymethyl-propane-1,3-diol); and Tricine (N-(2-hydroxy-1,1-bis(hydroxymethyl)ethyl)glycine).

[00179] According to some embodiments, the buffer is a sulfonic acid derivative buffer including, but not limited to, ACES, BES, DIPSO, HEPBS, HEPES, HEPPS, HEPPSO, MES, MOBS, MOPS, MOPSO, PIPES, POPSO, Sulfite, TAPS, TAPSO, and TES buffer.

[00180] According to additional embodiments, the buffer is a carboxylic acid derived buffer including, but not limited to, Acetate, N-(2-acetamido)-2-iminodiacetic acid, 2-(bis(2-hydroxyethyl)amino)acetic acid, Carbonate, Citrate, 3,3-dimethyl-glutarate, Lactate, Maleate, Oxalate, Succinate, and Tartaric acid buffer.

[00181] According to other embodiments, the buffer is an amino acid-derived buffer including, but not limited to, Bicine, Glycine, Glycine-amide, Histidine, and Tricine buffer.

[00182] According to still other embodiments, the buffer is a phosphoric acid-derived buffer including, but not limited to, Glycerol-2-phosphate and phosphate buffer.

[00183] Alternatively, the buffered saline formulation may be, for example, Hank's balanced salt solution, Earle's balanced salt solution, Gey's balanced salt solution, HEPES-buffered saline, phosphate-buffered saline, Plasma-lyte, Ringer's solution, Ringer's acetate, saline citrate, or Tris-buffered saline.

[00184] The buffered dextrose solution formulation may be, for example, acid-citrate-dextrose solution or Elliott's solution B.

[00185] According to exemplary embodiments, the injection solutions are Plasma-Lyte A or Compound Sodium Lactate purchased from Baxter.

Therapeutic Use

[00186] The compounds of the present invention are useful for preventing cell growth, or inducing death of the hyperproliferative cell in which it is present and include known cytotoxic, cytostatic and antiproliferative drugs, as are known in the art. As such, compounds of the present invention are useful in the treatment and prevention of a variety of cancers, as well as pre-treatment prior to a bone marrow transplant.

[00187] Thus, in some embodiments, the present invention provides a method of treating a cancer including metastases, or a pre-cancer condition or disorder, comprising the step of administering to a subject in need thereof a therapeutically effective amount of a salt of formula (I), or any compound covered by such formula, for example, compounds (II), (III), (1), (1A), (1B), (2), (3) or (4), as described herein.

[00188] In other embodiments, the present invention provides a salt of formula (I), or any compound covered by that formula, for example, compounds (II), (III), (1), (1A), (1B), (2), (3) or (4), as described herein, for use in the treatment of a cancer including metastases thereof, or a pre-cancerous condition or disorder.

[00189] In some embodiments, the cancer is characterized by a non-solid or solid tumor or a combination thereof. In other embodiments, the cancer is a hematologic cancer.

[00190] In some specific embodiments, the cancer is characterized by a non-solid tumor selected from lymphoproliferative disorders including leukemias and lymphomas.

[00191] In other specific embodiments, the cancer characterized by a solid tumor is selected from the group consisting of central nervous system (CNS) tumors, liver cancer, colorectal carcinoma, breast cancer, gastric cancer, pancreatic cancer, bladder carcinoma, cervical carcinoma, head and neck tumors, vulvar cancer, and dermatologic neoplasms including melanoma, squamous cell carcinoma, and basal cell carcinomas.

[00192] The salt forms of the present invention can be used to treat any mammal, preferably humans. In other embodiments, the present invention relates to a method of treating a subject who is a candidate for a bone marrow transplant, comprising the step of pre-treating said subject, prior to the transplant, with a salt of formula (I) or any compound covered by such formula, for example, compounds (II), (III), (1), (1A), (1B), (2), (3) or (4), as described herein. In other embodiments, the present invention relates to a salt of formula (I), or any compound covered by such formula, for example, compounds (II), (III), (1), (1A), (1B), (2), (3) or (4), as described herein, for use in pre-treatment prior to a bone marrow transplant.

[00193] In some embodiments, the cancer may be selected from hematologic cancers or non-hematologic cancers.

[00194] Hematologic cancers include leukemias, lymphomas, and myelomas, including but not limited to myeloid leukemia, lymphocytic leukemia, e.g., acute myeloid leukemia (AML), chronic myeloid leukemia (CML), acute lymphocytic leukemia (ALL), chronic lymphocytic leukemia (CLL), Hodgkin's lymphoma, non-Hodgkin's lymphoma, multiple myeloma, and Waldenstrom's macroglobulinemia. Hematologic cancer may be a relapsed/refractory cancer or a newly diagnosed cancer.

[00195] Non-hematologic cancers also known as solid tumors include, but are not limited to, sarcoma, carcinoma, fibrosarcoma, myxosarcoma, liposarcoma, chondrosarcoma, osteogenic sarcoma, chordoma, angiosarcoma, endotheliosarcoma, mesothelioma, leiomyosarcoma, Ewing's tumor, rhabdomyosarcoma, squamous cell carcinoma, basal cell carcinoma, adenocarcinoma, sweat gland carcinoma, sebaceous gland carcinoma, papillary carcinoma, papillary adenocarcinoma, cystadenocarcinoma, medullary carcinoma, bronchogenic carcinoma, renal cell carcinoma, hepatoma bile duct carcinoma, choriocarcinoma, seminoma, embryonal carcinoma, Wilms tumor cervical cancer, testicular tumor, lung carcinoma, small cell lung carcinoma, bladder carcinoma, epithelial carcinoma, astrocytoma, sarcoma Kaposi's disease and melanoma. Each possibility represents a separate embodiment of the invention.

[00196] Non-hematological cancers include organ cancers, where cancer of an organ includes, but is not limited to, breast cancer, bladder cancer, colon cancer, rectal cancer, endometrial cancer, kidney cancer, lung cancer, cervical cancer, pancreatic cancer, prostate cancer, testicular cancer, thyroid cancer, ovarian cancer, brain cancer, including ependymoma, glioma, glioblastoma, medulloblastoma, craniopharyngioma, pinealoma, acoustic neuroma, hemangioblastoma, oligodendroglioma, menangioma, neuroblastoma, retinoblastoma, and their metastases. Each possibility represents a separate embodiment of the invention.

[00197] In another aspect, the present invention provides a method of treating a pre-cancer condition or disorder, comprising the step of administering to a subject in need thereof a therapeutically effective amount of a salt of formula (I), or any compound covered by such formula, for example, compounds (II), (III), (1), (1A), (1B), (2), (3) or (4), as described herein.

[00198] In another aspect, the present invention provides a salt of formula (I), or any compound covered by that formula, for example, compounds (II), (III), (1), (1A), (1B), (2), (3) or (4), as described herein, for use in the treatment of a pre-cancerous condition or disorder.

[00199] The term precancerous disorder or condition includes Myelodysplastic Syndromes (MDS). The term “myelodysplastic syndromes” (MDS) refers to a heterogeneous group of hematopoietic premalignancies characterized by blood cytopenias, ineffective hematopoiesis, and hypercellular bone marrow. MDSs are preleukemic conditions, in which transformation to acute myeloid leukemia (AML) occurs in approximately 30 to 40% of cases. Unless allogeneic stem cell transplantation can be offered, MDS is generally considered an incurable condition.

[00200] The present invention further provides a method for treating an infection caused by a viral agent that is a cancer-causing virus comprising the step of administering a salt of formula (I), or any compound covered by such formula, for example, compounds (II), (III), (1), (1A), (1B), (2), (3) or (4), as described herein. Thus, the invention provides a method for treating a viral infection caused by a viral oncogene. Non-limiting examples of such viruses include human papillomavirus, hepatitis B, hepatitis C, Epstein-Barr virus, human T-lymphotropic virus, Kaposi's sarcoma-associated herpesvirus and Merkel cell polyomavirus. Each possibility represents a separate embodiment of the invention.

[00201] A viral disease may be caused by other viruses including, but not limited to, human immunodeficiency virus (HIV), herpes simplex virus (HSV), cytomegalovirus (CMV), and varicella zoster virus (VZV). Each possibility represents a separate embodiment of the invention. According to one embodiment, the viral disease is Herpes viral encephalitis.

[00202] The method of the present invention may be useful for treating a neoplastic disease in a subject having an immunological disease or disorder. Immunological diseases or disorders include, but are not limited to, rheumatoid arthritis (RA), psoriatic arthritis, systemic lupus erythematosus (SLE), lupus nephritis, inflammatory bowel disease (IBD), irritable bowel syndrome, type I diabetes, immune thrombocytopenic purpura, multiple sclerosis, Sjorgren's syndrome, Hashimoto's thyroiditis, Grave's disease, primary biliary cirrhosis, Wegener's granulomatosis, tuberculosis, and Waldenstrom's macroglobulemia.

[00203] The method of the present invention may be useful for treating a neoplastic disease in individuals with organ dysfunction, such as liver dysfunction, kidney dysfunction, pancreatic dysfunction, bone marrow dysfunction, and cerebellar dysfunction.

[00204] The term “liver dysfunction” refers to a state in which liver function is decreased compared to a normal state. In general, liver dysfunction is a state characterized by any one or more measurement values of inspection items for liver function (e.g., levels of AST, ALT, TTT, ZTT, total bilirubin, total protein, albumin, lactate dehydrogenase, choline esterase and the like in the blood) are deviated from the range of normal values (reference values). Liver dysfunction is characteristic of diseases such as, for example, fulminant hepatitis, chronic hepatitis, viral hepatitis, alcoholic hepatitis, liver fibrosis, liver cirrhosis, liver cancer, autoimmune hepatitis, drug-allergic liver disease and primary biliary cirrhosis.

[00205] Renal dysfunction is characteristic of diseases such as, for example, acute renal failure, glomerulonephritis, chronic renal failure, azotemia, uremia, immune renal disease, acute nephrotic syndrome, rapidly progressive nephritic syndrome, nephrotic syndrome, Berger's disease, chronic nephritic/proteinuria syndrome, tubulointerstitial disease, nephrotoxic disorders, renal infarction, atheroembolic renal disease, renal cortical necrosis, malignant nephroangiosclerosis, renal vein thrombosis, renal tubular acidosis, renal glycosuria, nephrogenic diabetes insipidus, Bartter's syndrome, Liddle's syndrome, polycystic kidney disease, interstitial nephritis, acute hemolytic uremic syndrome, medullary cystic disease, medullary sponge kidney, hereditary nephritis, and nail-patella syndrome.

[00206] Pancreatic dysfunction is characteristic of diseases such as diabetes, hyperglycemia, impaired glucose tolerance and insulin resistance.

[00207] Bone marrow dysfunction is characteristic of diseases such as, for example, osteomyelitis, dyshematopoiesis, ion deficiency anemia, pernicious anemia, megaloblastosis, hemolytic anemia and aplastic anemia.

[00208] Cerebellar dysfunction is characteristic of motor and neurobehavioral disorders such as hypotonia, dysarthria, dysmetria, dysdiadochokinesia, impaired reflex and intention tremor.

[00209] In one embodiment, the individual is a medically compromised individual who is unsuitable for treatment with an anticancer agent, e.g., cytarabine. The medically compromised individual may be selected from the group consisting of elderly individuals, individuals with hepatic dysfunction, individuals with renal dysfunction, individuals having pancreatic dysfunction, individuals with bone marrow dysfunction, individuals having cerebellar dysfunction, individuals having an immunological disorder, individuals having relapsed or refractory hematologic cancer, and any combination thereof.

[00210] The toxicity and therapeutic efficacy of the compounds described herein can be determined by standard pharmaceutical procedures in cell cultures or experimental animals, for example, by determining the IC50 (the concentration that provides 50% inhibition of cell growth) and the MTD (maximum tolerated dose in test animals) for a compound in question. Data obtained from cell culture assays and animal studies can be used in formulating a dosage range for use in human subjects. The dosage may vary depending on the dosage form employed and the route of administration used. The exact formulation, route of administration and dosage can be chosen by the individual physician in view of the patient's condition. (See, for example, Fingl et al., 1975, in “The Pharmacological Basis of Therapeutics”, chap. 1 p. 1).

[00211] The amount of active agent used in a composition of the present invention is an amount effective to achieve the purpose of the particular active agent for the target indication. The amount of an active agent in the compositions is typically a therapeutically, pharmacologically, biologically or chemically effective amount. However, the amount may be less than this amount when the composition is used in a unit dosage form because the unit dosage form may contain a plurality of active compounds or agents in a single composition or may contain a divided pharmacologically, biologically, therapeutically or chemically effective amount. The total effective amount may then be administered in cumulative units containing, in total, an effective amount of the active agent.

[00212] The salt of the invention can be administered in a daily dose ranging from about 0.3 g/m2 to about 10 g/m2 of the subject's surface area. According to some embodiments, the salt can be administered in a daily dose ranging from about 0.5 g/m2 to about 5 g/m2 of the subject's surface area. According to some embodiments, the salt can be administered in a daily dose ranging from about 0.5 g/m2 to about 4.5 g/m2 of the subject's surface area. According to other embodiments, the salt is administered in a daily dose of about 0.3; 0.5; 0.8; 1; 1.5; 2; 2.3; 2.5; 3; 3.5; 4; 4.5; 5; 6; 7; 8; 9; 10 g/m2 of the subject's surface area. Each possibility represents a separate embodiment of the invention.

[00213] According to some embodiments, the salt of the invention is administered by intravenous infusion at a daily dose ranging from 0.3 g/m2 to 4.5 g/m2 of the subject's surface area.

[00214] According to some embodiments, the pharmaceutical composition is administered at least once per month. According to additional embodiments, the pharmaceutical composition is administered at least twice per month. According to other embodiments, the pharmaceutical composition is administered at least once per week. According to still other embodiments, the pharmaceutical composition is administered at least twice per week. According to still other embodiments, the pharmaceutical composition is administered once per day for at least one week. According to other embodiments, the pharmaceutical composition is administered at least once per day for at least one week or until the subject is cured.

[00215] According to some embodiments, the pharmaceutical composition is administered once daily for at least 2, 3, 4, 5, 6, 8, 10, 12 or at least 14 consecutive days once a month. Alternatively, the pharmaceutical composition is administered once daily for at least 2, 3, 4, 5, 6 or 12 days twice a month, or alternatively the pharmaceutical composition is administered every day or twice a week until the patient is cured.

[00216] In some embodiments, where the pharmaceutical composition is used to prevent cancer recurrence, the pharmaceutical composition may be administered regularly for extended periods of time in accordance with the clinician's instructions.

[00217] In some cases it may be advantageous to administer a large loading dose followed by periodic (e.g., weekly) maintenance doses during the treatment period. Compounds may also be dispensed by slow-release dispensing systems, pumps, and other known dispensing systems for continuous infusion. Dosing regimens may be varied to provide desired circulating levels of a particular compound based on its pharmacokinetics. Thus, doses are calculated so that the desired circulating level of a therapeutic agent is maintained.

[00218] Typically, the effective dose is determined by the activity and efficacy of the compound and the condition of the individual, as well as the body weight or surface area of the individual being treated. The dose and dosage regimen are also determined by the existence, nature and extent of any adverse side effects accompanying the administration of the compounds in a particular individual.

[00219] According to additional embodiments, the individual to be treated is an individual who is 50 or older, such as 60, 70, 75 or older. Each possibility represents a separate embodiment of the invention.

[00220] According to other embodiments, the compound of the invention is administered in a daily dose of at least 2, 3, 5, 10, 15, 20, or at least 30 times greater than the maximum standard of care dose of cytarabine alone. Each possibility represents a separate embodiment of the invention. The following examples are to be considered only as illustrative and not limiting in nature. It will be apparent to one skilled in the art to which the present invention relates that many modifications, permutations and variations can be made without departing from the scope of the invention. EXAMPLES The following abbreviations are used in the examples: BOC: t-butyloxycarbonyl DCC: N,N'-dicyclohexylcarbodiimide DMF: dimethylformamide DCM: dichloromethane DIC: diisopropyl carbodiimide DIEA: diisopropylethylamine DMAP: dimethyl aminopyridine Fmoc: 9-fluorenylmethyloxycarbonyl HBTU: 2-(1H-benzotriazol-1-yl)-1,1,3,3-tetramethyluronium hexafluorophosphate HOBt: N-hydroxybenzotriazole HPLC: High Performance Liquid Chromatography MS: Mass Spectrometry NMP: N-methyl 2-pyrrolidone OtBu: t-butyl ester TFA: trifluoroacetic acid Example 1: Synthesis of BOC-Asp(Cytarabine)-OtBu(A)- a protected precursor of the salt form Asp(Cytarabine) •Y(1)

[00221] BOC-Asp-OtBu (23.7 g), EDC (15.7 g), HOBT (11.1 g) and Ara-C (20 g) were dissolved in 400 ml of DMF and mixed for 24 h at room temperature. The mixture was evaporated at 80°C to give oily residue. The residue was dissolved in 300 ml of ethyl acetate and extracted successively with 5% NaHCO3, 0.1 M HCl, 5 M NaCl. The mixture was evaporated completely to give the solid product of the protected precursor BOC-Asp(Cytarabine)-OtBu. The precursor was analyzed by mixed mode chromatography HPLC on primesep 100 at a gradient of acetonitrile/H2O to 0.1% TFA to give a peak at retention time of 29.6 min. The yield of the precursor product was 80-90%, off-white crystals, HPLC purity 95%.

[00222] The protected precursor BOC-Asp(Cytarabine)-OtBu (A) is further used for the creation of variety salt forms of the final product Asp(Cytarabine) •Y (Y = acid addition salt): Example 2: Synthesis of Asp(Cytarabine)acetate (Y=CH3COOH), (formula IV) - Route A using TFA for deprotection

[00223] The protected precursor BOC-Asp(Cytarabine)-OtBu (from example 1) was subjected to TFA cleavage. Cleavage of the protecting groups (BOC, OtBu) was carried out in 400 ml of TFA + 2.5% H2O for 2.5 hours. The solution was transferred to 1.6 L of tBME for precipitation of the product. The precipitate was dried to give off-white crystals of Asp(Cytarabine)•TFA, TFA salt form (M.W. = 472 g/mol). Asp(Cytarabine)•TFA was further dissolved in concentrated acetic acid until complete dissolution. The product was precipitated with tBME and dried to give Asp(Cytarabine)•Acetate. The product yield was 75-85%, white crystals. The crude product was analyzed by mixed mode chromatography HPLC on primesep 100 at a gradient of acetonitrile/H2O to 0.1% TFA, at 240 nm, to give a peak at retention time 9.2 min, purity 85.3%. MS results show MW = 359 g/mol (MS + 1H). The MS results are presented in FIG. 1. Optical density at 213, 245, and 298 nm. NMR results: 1H NMR (600 MHz, DMSO-6d) d ppm (600MHz, DMSO-6d) δ ppm 2.98 (dd, 1H), 3.08 (dd, 1H), 3.61 (d, 1H), 3.84 (ddd, 1H), 3.93 (td, 1H), 4.06 (td, 1H), 3 (dtd, 1H),5.50 (s,1H), 6.05 (d,1H), 7.12 (d, 1H), 8.09 (d, 1H), 11.10 (s, 1H). 13C NMR (151MHz, DMSO-6d) δ ppm 36.61 (s, 1C), 48.23 (s, 1C), 61.02 (s, 1C), 74.57 (s, 1C), 76.15 (s, 1C), 85.82 (s, 1C), 87.06 (s, 1C), 94.30 (s, 1C), 147.01 (s, 1C), 154.33 (s, 1C), 161.84 (s, 1C), 170.00 (s, 1C), 170.04 (s, 1C). NMR results are in agreement with the anticipated structure.

[00224] Purification of Asp(Cytarabine). Acetate 95-100% was performed as follows: 7 g of crude product (85.3%) was dissolved in 15 ml of H2O and separated by preparative HPLC as described in Table 1. Solution A - 0.1% TFA or 1% acetic acid dissolved in water. Solution B - 0.1% TFA or 1% acetic acid dissolved in acetonitrile/water = 9/1. Injection volume: 15 ml, Wavelength: 220nm, Flow rate: 40 ml/min, Column: primesep100, Dimension: 60x300mm, Particle size: 10 μm 100A. Fraction collection volume: 30 ml. The fractions collected from minutes 18 to 36 showed 95% purity of Asp(Cytarabine) •Acetate. The fractions were freeze-dried to obtain purified Asp(Cytarabine) •Acetate powder in a yield of 3.18 g. Table 1 Example 3: Synthesis of Asp(Cytarabine) •methanesulfonate by acid substitution - Route A using TFA for deprotection

[00225] Two grams of BOC-protected precursor Asp(Cytarabine)-OtBu were subjected to TFA cleavage as described in Example 1. 1 g of Asp(Cytarabine)•TFA was dissolved in 70% methanesulfonic acid in H2O to complete dissolution. The product Asp(Cytarabine)•methanesulfonate was precipitated with isopropanol/tBME and the pellet was dried. The product yield was 51%, white crystals. The product was analyzed by HPLC and MS as described in Example 2. The product yielded a peak at retention time 10.6 minutes, purity 80.6%.

[00226] Purification of 95-100% Asp(Cytarabine) methanesulfonate was carried out accordingly: 20 g of G-10 resin was swelled in 100 ml of H2O for 1 hour. After swelling, G-10 was transferred to sintered glass and washed with 20% ethanol in H2O (x3 column volumes), H2O and 0.1% methanesulfonic acid in H2O. A crude powder of Asp(Cytarabine) methanesulfonate (80.6%) was dissolved with 5 ml of H2O. The solution was loaded onto G-10 resin and extracted with 0.1% methanesulfonic acid in H2O. 5 ml fractions were collected. The purified Asp(Cytarabine) methanesulfonate fractions were greater than 95% pure. Example 4: Synthesis of salt forms of Asp(Cytarabine) •Y variety by acid substitution

[00227] Route A can be used for various salt forms by replacing TFA with other acids by the same method described in Examples 2 and 3. Acids such as: benzenesulfonic, benzoic, boric, citric, ethanesulfonic, formic, hydrochloric, lactic, maleic, malonic, methanesulfonic, methophosphoric, nicotinic, oxalic, phosphoric, salicylic, succinic, sulfuric, tartaric and toluenesulfonic. A variety of acids with selected concentrations and ratios from 1:1 to 1:20 were analyzed for the substitution reactions. After precipitation, the products were analyzed by HPLC and MS. The conditions and results are presented in Table 2. The products Asp(Cytarabine) •Y are further purified using purification methods described in Examples 2 and 3. Table 2: Example 5: Synthesis of the methanesulfonate salt form of Asp(Cytarabine) by cleavage of methanesulfonic acid - Route B

[00228] 0.25 g of BOC-Asp(Cytarabine)-OtBu precursor (from example 1) was subjected to cleavage by methanesulfonic acid. 0.25 g of precursor was added with 1 mL of 70% MSA in H2O. After 2 h of cleavage the solution was transferred to 10 mL of tBME, a sediment of the product Asp(Cytarabine) methanesulfonate was formed. The product was washed with tBME and dried. The yield of the product was 65%, white crystals. The product was analyzed by mixed mode chromatography by HPLC on primesep 100 at a gradient of Acetonitrile/H2O to 0.1% TFA, at 220 nm, to give a peak at retention time of 9.6 minutes, purity of 56%. The MS results of Asp(Cytarabine) methanesulfonate show M.W. = 453 and 454 g/mol (MS±1H). The MS results are presented in FIG. 2. Purify Asp(Cytarabine) methanesulfonate as described in Examples 2 and 3. If the purification method of Example 2 is used, the acid used in Solutions A and B is replaced by the acid addition salt. Solution A - 0.1% methanesulfonic acid dissolved in water. Solution B - 0.1% methanesulfonic acid dissolved in acetonitrile/water = 9/1. Example 6: Synthesis of the salt form of Asp(Cytarabine) •benzenesulfonate by cleavage of benzenesulfonic acid - Route B

[00229] The salt form of Asp(Cytarabine)benzenesulfonate was synthesized from the precursor BOC-Asp(Cytarabine)-OtBu by cleavage with benzenesulfonic acid as described in Example 5. The yield of the product was 31%, white crystals. The product was analyzed as described in Example 5. HPLC results show two peaks at retention times of 9.3 and 14.7 minutes, purity of 20 and 44%, respectively. MS results of Asp(Cytarabine)benzenesulfonate show M.W. = 515 and 516 g/mol (MS±1H). The MS results are presented in FIG. 3. Asp(Cytarabine) benzenesulfonate is purified as described in Examples 2, 3 and 5. Example 7: Synthesis of the phosphate salt form of Asp(Cytarabine) by cleavage with phosphoric acid - Route B

[00230] The phosphate salt form of Asp(Cytarabine) was synthesized from the BOC-Asp(Cytarabine)-OtBu precursor by concentrated phosphoric acid cleavage as described in Example 5. The product yield was 34%, white crystals. The product was analyzed as described in Example 5. HPLC results show the peak at retention times of 9.0 minutes, purity of 84.3%. MS results of Asp(Cytarabine) phosphate show MW = 455 and 456 g/mol (MS±1H). Asp(Cytarabine) phosphate is purified as described in Examples 2, 3 and 5. Example 8: Synthesis of the sulfate salt form of Asp(Cytarabine) by sulfuric acid cleavage - Route B

[00231] The phosphate salt form of Asp(Cytarabine) was synthesized from the BOC-Asp(Cytarabine)-OtBu precursor by concentrated sulfuric acid cleavage as described in Example 5. The product yield was 77%, white crystals, M.W. = 456 g/mol. The product was analyzed as described in Example 5. HPLC results show the peak at retention times of 9.3 minutes, purity of 53%. MS results of Asp(Cytarabine) sulfate show M.W. = 455 and 456 g/mol (MS ± 1H). Asp(Cytarabine) sulfate is purified as described in Examples 2, 3 and 5. Example 9: Synthesis of the salt form of Asp(Cytarabine) •hydrochloride by hydrochloric acid cleavage - Route C

[00232] 1 g of BOC-Asp(Cytarabine)-OtBu precursor (from example 1) was subjected to HCl cleavage. 3 ml of 37% HCl/H2O was added to the precursor and incubated for 1 hour to remove the BOC and OtBu protecting groups and form the salt form. The solution was titrated with 1N NaOH to pH = 5-6. During the titration, white sediment was formed. The received sediment was dried, dissolved in 2 ml of H2O and lyophilized. The product was analyzed as described in example 5. HPLC results show the peak at retention times of 9.46 minutes, purity of 75.9%. Asp(Cytarabine) • hydrochloride is purified as described in Examples 2, 3 and 5. Example 10: Synthesis of variety of Asp(Cytarabine) •Y salt forms by acid cleavage

[00233] Routes B and C can be used for the formation of various salt forms by cleavage with the acid that creates the salt form by the same methods described in Examples 5 and 9. Preferred acids for removing protecting groups and forming the salt form have a pKa <3, with a ratio of 1:1 to 30:1 (acid:precursor). For example, acids such as: benzenesulfonic, camphorsulfonic, cyclamic, 2,2-dichloroacetic, di(t-butyl)-naphthalenesulfonic, di(t-butyl)-naphthalenedisulfonic, dodecylsulfuric, ethane-1,2-disulfonic, ethanesulfonic, fumaric, galactaric, gentisic, glucaric, gluconic, glycerophosphoric, hydrochloric, hydrobromic, hydroiodic, 2-hydroxy-ethanesulfonic (isethiononic), 1-hydroxy-2-naphthoic, maleic, malonic, medronic (bisphosphonic), methanesulfonic, methophosphoric, methylboronic, naphthalene-1,5-disulfonic, naphthalene-2-sulfonic, nitric, orotic, oxalic, 2-oxo-glutaric (ketoglutaric), pamoic (embonic), pyruvic, phosphoric, saccharin, salicylic, sulfuric, tartaric, thiocyanic, toluenesulfonic, and trifluoroacetic acid and the like. A variety of acids with selected concentrations from 10% to 100% (concentrated acid) were used for the cleavage of the BOC-Asp(Cytarabine)-OtBu precursor to remove the protecting group and form the salt form. The products were further analyzed by HPLC and MS. The conditions and results are presented in Table 3. The products Asp(Cytarabine) •Y are purified as described in Examples 2, 3, and 5. Table 3 Example 11: Preparation of addition salt of Asp(Cytarabine) •Y with salts (such as NaH2PO4)

[00234] The protected BOC-Asp(Cytarabine)-OtBu precursor was synthesized as described in Example 1. Asp(Cytarabine).TFA was synthesized as described in Example 2. 0.25 g of Asp(Cytarabine).TFA was further dissolved in 1 ml of 1M NaH2PO4 until complete dissolution. The product was precipitated with tBME and dried to afford Asp(Cytarabine) •phosphate product.

[00235] This reaction can be used with other salts such as: KH2PO4 and the like. Example 12: Synthesis of BOC-Asp(Gemcitabine)-OtBu (B)- a protected precursor of the salt form Asp(Gemcitabine) •Y (2)

[00236] The BOC-protected precursor Asp(Gemcitabine)-OtBu was synthesized as described in Example 1.

[00237] The protected precursor is further used for the creation of a variety of salt forms of the final product Asp(Gemcitabine) •Y (Y = acid addition salt) (2). Example 13:Synthesis of salt form Asp(Gemcitabine) •acetate

[00238] Salt form of Asp(Gemcitabine) •acetate (Y = CH3COOH) was synthesized from precursor B by acid substitution as described in Example 2. Example 14: Synthesis of the methanesulfonate salt form of Asp(Gemcitabine)

[00239] The methanesulfonate salt form of Asp(Gemcitabine) (Y = CH3SO3H) was synthesized from precursor B by cleavage with methanesulfonic acid as described in Example 5. Example 15: Synthesis of BOC-Glu(Cytarabine)-OtBu(C)- a protected precursor of the Glu(Cytarabine)•Y salt form (3)

[00240] The protected precursor of BOC-Glu(Cytarabine)-OtBu was synthesized as described in Example 1.

[00241] The protected precursor is further used to create variety salt forms of the final product Glu(Cytarabine)•Y (Y = acid addition salt) (3). Example 16:Synthesis of salt form of Glu(Cytarabine) •acetate

[00242] Salt form Glu(Cytarabine)acetate (Y = CH3COOH) was synthesized from precursor C via acid substitution as described in Example 2. Example 17: Synthesis of the methanesulfonate salt form of Glu(Cytarabine)

[00243] Methanesulfonate salt form of Glu(Cytarabine) (Y = CH3SO3H) was synthesized from precursor C by cleavage with methanesulfonic acid as described in Example 5. Example 18: Synthesis of BOC-Glu(Gemcitabine)-OtBu(D)- a protected precursor of the salt form Glu(Gemcitabine)•Y(4)

[00244] The BOC-protected precursor-Glu(Gemcitabine)-OtBu was synthesized as described in Example 1.

[00245] The protected precursor of the salt form Glu(Gemcitabine) •OtBu is further used to create a variety of salt forms of the final product Glu(Gemcitabine) •Y (4) ((Y = acid addition salt). Example 19: Synthesis of salt form of Glu (Gemcitabine) •acetate

[00246] Salt form Glu(Gemcitabine) •acetate (Y = CH3COOH) was synthesized from precursor D by acid substitution as described in Example 2. Example 20: Synthesis of salt form Glu(Gemcitabine) •methanesulfonate

[00247] Salt form of Glu(Gemcitabine)•methanesulfonate (Y = CH3SO3H) was synthesized from precursor D by cleavage with methanesulfonic acid as described in Example 5. Example 21: Synthesis of the salt form of Asp(Cytarabine)•HCl (Y = HCl) Stage 1: Synthesis of Boc-Asp(Cytarabine)-OtBu

[00248] Cytarabine (0.50 g, 2.1 mmol) was suspended in 5 mL of dry DMF. The mixture was heated to 70 °C until a clear solution was obtained. Boc-Asp-OtBu (0.40 g, 1.4 mmol) was dissolved at room temperature in 5 mL of dry DMF. HOBt (0.22 g, 1.5 mmol) was added and dissolved, followed by EDC (0.28 g, 1.5 mmol). The solution was stirred at room temperature for 30 min.

[00249] At 45°C, the activated amino acid cytarabine was added over 2 h and then stirred at 45°C overnight.

[00250] The solvent was evaporated under reduced pressure at 60°C. The oily residue was dissolved in ethyl acetate (55 mL) and washed with 3 x 5% NaHCO3, 3 x 0.1 M HCl, and 3 x brine (15 mL each wash). After drying over sodium sulfate and filtration, the solvent was evaporated and the crude product was obtained as a foamy solid. The crude product contains an impurity of ~5-15% (Boc-Asp-OtBu)2-(Cytarabine). Stage 2: Purification of Boc-Asp(Cytarabine)-OtBu

[00251] The crude product is purified using a Biotage Isolera® purification system. 3.97 g of the crude product (containing less than 1% (Boc-Asp-OtBu) of the 2-cytarabine impurity) was purified using a SNAP cartridge, KP-SIL 100 g Silica column with the following gradient at a flow rate of 40 mL/min (Monitoring at 254 nm).

[00252] The product fractions were combined and evaporated to dryness, yielding 3.78 g of purified product. The 1H NMR spectrum (MeOD) of purified Boc-Asp-OtBu-(Cytarabine) is shown in FIG. 4 Stage 3: Synthesis of Asp(Cytarabine)•HCl

[00253] Boc-Asp-OtBu-(Cytarabine) (3.6 g, 7.0 mmol) was dissolved in 8.8 mL of dry dioxane. After obtaining a clear solution, HCl in dioxane (4 M, 17.5 mL, 70 mmol) was added, and the mixture was stirred overnight (after 1 min, a white solid formed) at room temperature. H2O (0.26 mL, 7.0 mmol) was added dropwise, and stirring was continued for 3 h. The precipitate was filtered (4 sintered glass filter) and washed with dry dioxane (2 x 25 mL). The obtained white solid was dried under vacuum at room temperature to give a white powder (3.34 g). The 1H NMR spectrum (MeOD) of Asp(Cytarabine)•HCl is shown in FIG. 5 Example 22: Synthesis of the Asp(Cytarabine)•HCl (Y=HCl) salt form of protected cytarabine. Stage 1: Protection of cytarabine

[00254] 2.44 g of cytarabine and 1.5 g of t-butyldimethylsilyl chloride (TBDMS-Cl) were dissolved in 20 ml of pyridine, the reaction was stirred for 10 hours. The solution was evaporated and the pellet was washed with water and ethyl acetate to give a white solid of TBDMS-cytarabine (5'-OH protected). Stage 2: Synthesis of Boc-Asp(TBDMS-cytarabine)-OtBu

[00255] 1 g of TBDMS-cytarabine was dissolved in 10 mL of DMF. 0.8 g of Boc-Asp-OtBu was dissolved in 5 mL of DMF. 0.44 g of HOBt was added and dissolved, followed by 0.56 g of EDC. The solution is stirred at room temperature for 30 min. The activated amino acid was added dropwise into TBDMS-cytarabine solution and then stirred overnight.

[00256] The solvent was evaporated under reduced pressure at 60°C. The oily residue was dissolved in ethyl acetate and washed with 5% NaHCO3, 0.1 M HCl and brine. 2/3 of the solvent was evaporated and the crude product was precipitated by hexane to give a white solid. The crude product does not contain (Boc-Asp-OtBu)2-(TBDMS-Cytarabine) impurity, no column purification was required. Stage 3: Synthesis of Asp(Cytarabine)•HCl

[00257] Boc-Asp(TBDMS-cytarabine)-OtBu was used for the synthesis of Asp(Cytarabine) •HCl, using the same process described in step 3 of example 22. Example 23: Biological activity of Asp(Cytarabine) •Y salt forms, effect on the proliferation of leukemia cancer cells

[00258] L1210 leukemia cancer cells were cultured in DMEM medium with 10% donor horse serum, 1% glutamine, 1% non-essential amino acids, 0.1% amphotericin, and 0.1% gentamicin. Cells were diluted and seeded in 96-well plates of 2000 cells/ml, 400 cells/well, in a volume of 0.2 ml per well. Plates were incubated at 37°C for 180 min. After incubation, test substances were added at increasing concentrations from 1 μM to 100 nM formulated in saline in 6 replicates each. The substances tested were Asp(Cytarabine) acetate salt form, Asp(Cytarabine) hydrochloride salt form, Asp(Cytarabine) methanesulfonate salt form, Asp(Cytarabine) benzenesulfonate salt form and Asp(Cytarabine) phosphate salt form. The test plates also included 6 replicate wells of saline as a control. The plates were incubated at 37°C for 72 h prior to analysis. At the end of the exposure period, an MTT [3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide] assay was performed. MTT at a concentration of 5 mg/ml in a volume of 0.02 ml was added to each test. The plates were incubated at 37°C for 3 h, then centrifuged at 3500 rpm for 6 min and the supernatants were aspirated. The pellets containing MTT crystals were dissolved in 0.2 ml of DMSO each. The absorbance was determined using ELISA Reader at a wavelength of 570 nm. The IC50 values obtained are summarized in Table 4. Table 4: Example 24: Biological activity of the acetate salt form of Asp(Cytarabine), effect on proliferation of various cell lines

[00259] Human leukemia cancer cell line was cultured in RPMI medium with 10% fetal bovine serum, 1% glutamine, 0.1% amphotericin and 0.1% gentamicin. Cells were diluted and seeded as described in Example 23. The acetate salt form of Asp(Cytarabine) was added in increasing concentrations from 1μM to 100 nM formulated in saline in 3 replicates each. The MTT assay was performed as described in Example 23. The IC50 values obtained are summarized in Table 5. Table 5:

[00260] It will be appreciated by those skilled in the art that the present invention is not limited by what has been particularly shown and described above. On the contrary, the scope of the present invention includes combinations and subcombinations of various features described above, as well as variations and modifications. Accordingly, the invention should not be construed as restricted to the particularly described embodiments, and the scope and concept of the invention will be more readily understood by reference to the claims, which follow.

Claims (23)

1. Pharmaceutically acceptable salt characterized by the fact that it is represented by the structure of formula (1): wherein Y is a pharmaceutically acceptable organic or inorganic acid or a residue thereof, selected from HCl, HBr, methanesulfonic acid, TFA, H2SO4, H3PO4 or NaHSO4. 2. Pharmaceutically acceptable salt according to claim 1, characterized in that Y is HCl, represented by the structure of: 3. Pharmaceutically acceptable salt according to claim 1, characterized in that Y is methanesulfonic acid, represented by the structure of: 4. Pharmaceutically acceptable salt according to characterized by the fact that Y is TFA, represented by the structure of: 5. Pharmaceutically acceptable salt according to claim 1, characterized in that Y is H2SO4, represented by the structure of: 6. Pharmaceutically acceptable salt according to claim 1, characterized in that Y is H3PO4, represented by the structure of: 7. Pharmaceutically acceptable salt according to claim 1, characterized in that Y is NaHSO4, represented by the structure of: 8. Pharmaceutical composition characterized by the fact that it comprises a pharmaceutically acceptable salt as defined in any one of claims 1 to 7, and a pharmaceutically acceptable excipient. 9. Use of a pharmaceutically acceptable salt represented by the structure of formula (1) as defined in any one of claims 1 to 7 characterized in that it is to prepare a pharmaceutical composition for treating a cancer or a pre-cancerous condition or disorder. 10. Use according to claim 9, characterized in that the cancer is metastatic cancer. 11. Use according to claim 9, characterized in that the cancer is a non-solid tumor or a solid tumor or a combination thereof. 12. Use according to claim 11, characterized in that the non-solid tumor comprises leukemias, lymphomas and multiple myeloma. 13. Use according to claim 12, characterized in that the leukemia comprises acute myeloid leukemia (AML) and acute lymphoblastic leukemia (ALL). 14. Use according to claim 11, characterized in that the solid tumor comprises tumors in the central nervous system (CNS), liver cancer, colorectal carcinoma, breast cancer, gastric cancer, pancreatic cancer, bladder carcinoma, cervical carcinoma, head and neck tumors, vulvar cancer, and dermatological neoplasms. 15. Use according to claim 14, characterized in that the dermatological neoplasms comprise melanoma, squamous cell carcinoma and basal cell carcinomas. 16. Use according to claim 9, characterized in that the cancer is a hematologic cancer. 17. Use according to claim 16, characterized in that the hematologic cancer is leukemia, lymphoma and myeloma. 18. Use according to claim 17, characterized in that the cancer is myeloid leukemia, lymphocytic leukemia, acute myeloid leukemia (AML), chronic myeloid leukemia (CML), acute lymphocytic leukemia (ALL), chronic lymphocytic leukemia (CLL), Hodgkin's lymphoma, non-Hodgkin's lymphoma, multiple myeloma, or Waldenstrom's macroglobulinemia. 19. Use according to claim 9, characterized in that the pre-cancer condition or disorder is myelodysplastic syndrome (MDS). 20. Use according to claim 9, characterized in that the treatment is of a human individual. 21. Use of a pharmaceutically acceptable salt represented by the structure of formula (1) as defined in any one of claims 1 to 7 characterized in that it is to prepare a pharmaceutical composition for treating an individual who is a candidate for a bone marrow transplant. 22. Use of a pharmaceutically acceptable salt represented by the structure of formula (1) as defined in any one of claims 1 to 7 characterized in that it is to prepare a pharmaceutical composition for treating a neoplastic disease in an individual who has an immunological disease or disorder. 23. Use of a pharmaceutically acceptable salt represented by the structure of formula (1) as defined in any one of claims 1 to 7 characterized in that it is to prepare a pharmaceutical composition for treating a neoplastic disease in individuals who have organ dysfunction.