WO2024109660A1

WO2024109660A1 - Cdk inhibitors

Info

Publication number: WO2024109660A1
Application number: PCT/CN2023/132400
Authority: WO
Inventors: 朱伟波; 付家胜; 覃华; 毕方超; 鲍劲霄; 石谷沁; 孙大庆; 张雯; 徐开; 孙维梅; 陶维康
Original assignee: 上海齐鲁制药研究中心有限公司
Priority date: 2022-11-21
Filing date: 2023-11-17
Publication date: 2024-05-30

Abstract

The present disclosure discloses a novel class of compounds which act as selective CDK8/CDK19 inhibitors, pharmaceutical compositions comprising the compounds, useful intermediates for preparing the compounds, and methods for treating cell proliferative diseases, such as cancer, by using the compounds of the present disclosure. The compounds have structures as shown in formula (I).

Description

CDK inhibitors

This application claims priority to Chinese patent application CN202211453864.X with an application date of November 21, 2022, Chinese patent application CN202310553978.X with an application date of May 16, 2023, and Chinese patent application CN202310802087.3 with an application date of June 30, 2023, all of which are incorporated by reference into this application.

Technical Field

The present disclosure belongs to the field of medicinal chemistry, and specifically relates to novel compounds with CDK8/CDK19 inhibitory activity, pharmaceutical compositions containing the compounds, useful intermediates for preparing the compounds, and methods for treating cell proliferative diseases such as cancer using the compounds of the present disclosure.

Background technique

The cell cycle is a basic process of cell life activities, controlling cell growth, proliferation and differentiation. Cyclin-dependent kinases (CDKs) are an important class of cellular enzymes that work in conjunction with cyclins and play an important role in the regulation of the cell cycle.

CDK8 and CDK19 are tumor-regulated transcription kinases with high homology that are closely related in structure and function. Unlike other kinases in the CDK family, such as CDK1, CDK2 and CDK4/6, CDK8 does not play a role in cell cycle regulation, so blocking CDK8 does not inhibit the growth of normal cells. However, since CDK8 plays a vital role in the formation of the pluripotent stem cell phenotype, knocking out CDK8 in embryonic stem cells will lead to embryonic development arrest. CDK8 plays an important role in regulating transcription by binding to the mediator complex or by phosphorylating transcription factors. Numerous genetic and biochemical studies have identified CDK8 as a key oncogenic driver for many cancers. Specifically, CDK8-mediated oncogenic Wnt-β-catenin signaling activation, transcription of estrogen-induced genes, and repression of super-enhancer-associated genes contribute to the tumorigenesis of colorectal, breast, and hematological malignancies, respectively. Currently, CDK8 overexpression has been observed in about 50% of colon cancer, melanoma, and breast cancer, and is associated with poor prognosis. In addition, studies have shown that CDK8 inhibitors can provide an important method of anti-cancer treatment as a single drug or in combination with various anti-tumor therapies or agents that activate the immune system, such as the treatment of blood diseases such as AML, MM, myelodysplastic syndrome (MDS) and chronic lymphocytic leukemia (CLL).

Although there are a large number of existing technologies that disclose compounds that can serve as CDK8 or CDK8/CDK19 inhibitors, there are relatively few molecules in the clinical stage, and all of them are in the early clinical stage. Therefore, the development of a new class of selective CDK8/CDK19 inhibitors is of great research significance.

Public Content

The purpose of the present disclosure is to provide a new class of compounds with CDK8/CDK19 inhibitory activity, pharmaceutical compositions containing the compounds, useful intermediates for preparing the compounds, and uses of the compounds in preparing drugs for treating cancer.

The present disclosure provides a compound represented by formula (I), a pharmaceutically acceptable salt and a stereoisomer thereof,

in,

Structural units Selected from

Ring B is selected from phenyl, 5-6 membered heteroaryl, 5-6 membered heterocycloalkyl, C _5-6 cycloalkyl;

Ring C is selected from phenyl, 5-6 membered heterocycloalkyl, 5-6 membered heteroaryl;

^Ra is independently selected from hydrogen, halogen, _C1-4 alkyl, _C1-4 alkoxy, CN, 5-6 membered heteroaryl; wherein the 5-6 membered heteroaryl may be optionally substituted by 1-3 ^Rab , wherein ^Rab is selected from halogen, _C1-4 alkyl;

R ^b is each independently selected from hydrogen, halogen, C _1-4 alkyl, C _1-4 alkoxy;

R ^c is independently selected from hydrogen, C _1-4 alkyl, -C _0-4 alkyl CONR ^ca R ^cb ; wherein R ^ca , R ^cb are independently selected from hydrogen, C _1-4 alkyl, or R ^ca The N atom to which R ^cb is commonly connected is cyclized to form a 5-6 membered heterocycloalkyl, a 5-11 membered para-heterocyclyl, or a 5-11 membered spiro heterocycloalkyl; wherein the 5-6 membered heterocycloalkyl, the 5-11 membered para-heterocyclyl, or the _5-11 membered spiro heterocycloalkyl may be optionally substituted by 1-3 C _1-4 alkyl, halogen, or C _1-4 haloalkyl, wherein the C _1-4 alkyl may be optionally substituted by one or more of the following groups: hydroxyl, amino, or cyano;

R ^d is selected from H, NH ₂ , OH, D, halogen, C _1-6 alkyl;

n is selected from 0, 1, 2, 3;

m is selected from 0, 1, 2, 3;

p is selected from 0, 1, 2, 3;

Wherein, formula (I) is not the following compound:

in,

Structural units Selected from

R ^c is independently selected from hydrogen, C _1-4 alkyl, -C _0-4 alkyl CONR ^ca R ^cb ; wherein R ^ca , R ^cb are independently selected from hydrogen, C _1-4 alkyl, or R ^ca and R ^cb and the N atom to which they are commonly connected are cyclized to form a 5-6 membered heterocycloalkyl, a 5-11 membered heterocyclyl, a 5-11 membered spiro heterocycloalkyl; wherein the 5-6 membered heterocycloalkyl, the 5-11 membered heterocyclyl, the 5-11 membered spiro heterocycloalkyl can be optionally substituted with 1-3 C _1-4 alkyl, halogen, C _1-4 haloalkyl, wherein _{the C 1-4} _alkyl can be optionally substituted with one or more of the following groups: hydroxyl, amino, cyano;

R ^d is selected from H, NH ₂ , OH, D, halogen;

n is selected from 0, 1, 2, 3;

m is selected from 0, 1, 2, 3;

p is selected from 0, 1, 2, 3;

Wherein, formula (I) is not the following compound:

in,

Structural units Selected from Ring B is selected from phenyl, 5-6 membered heteroaryl, 5-6 membered heterocycloalkyl, C _5-6 cycloalkyl;

R ^b is each independently selected from halogen, C _1-4 alkyl, C _1-4 alkoxy;

R ^c is independently selected from hydrogen, C _1-4 alkyl, -C _0-4 alkyl CONR ^ca R ^cb ; wherein R ^ca and R ^cb are independently selected from hydrogen, C _1-4 alkyl, or R ^ca and R ^cb and the N atom to which they are commonly connected are cyclized to form 5-6 membered heterocycloalkyl, 5-11 membered para-heterocycloalkyl, 5-11 membered spiroheterocycloalkyl; wherein 5-6 membered heterocycloalkyl, 5-11 membered para-heterocycloalkyl, 5-11 membered spiroheterocycloalkyl can be optionally substituted with 1-3 C _1-4 alkyl, halogen, C _1-4 haloalkyl;

R ^d is selected from H, NH ₂ ;

n is selected from 0, 1, 2, 3;

m is selected from 0, 1, 2, 3;

p is selected from 0, 1, 2, 3;

Wherein, formula (I) is not the following compound:

In some embodiments of the present disclosure, the compound of formula (I), pharmaceutically acceptable salts and stereoisomers thereof,

in,

Structural units Selected from

Ring B is selected from phenyl;

Ring C is selected from 5-6 membered heteroaryl;

R ^b is selected from hydrogen;

R ^c is selected from -C _1-4 alkyl CONR ^ca R ^cb ; wherein R ^ca and R ^cb are each independently selected from hydrogen, C _1-4 alkyl, or R ^ca and R ^cb and the N atom to which they are commonly connected are cyclized to form a 5-6 membered heterocycloalkyl, a 5-11 membered heterocyclyl, or a 5-11 membered spiro heterocycloalkyl; wherein the 5-6 membered heterocycloalkyl, the 5-11 membered heterocyclyl, or the 5-11 membered spiro heterocycloalkyl may be optionally substituted with 1-3 C _1-4 alkyl, halogen, or C _1-4 haloalkyl, wherein the C _1-4 alkyl may be optionally substituted with one or more of the following groups: hydroxyl, amino, or cyano;

R ^d is selected from H, NH ₂ , OH, D, halogen, C _1-4 alkyl;

n is selected from 1, 2, and 3;

m is selected from 1, 2, and 3.

in,

Structural units Selected from

Ring B is selected from phenyl;

Ring C is selected from 5-6 membered heteroaryl;

R ^b is selected from hydrogen;

R ^d is selected from H, NH ₂ , OH, D, halogen;

n is selected from 1, 2, and 3;

m is selected from 1, 2, and 3.

In some embodiments of the present disclosure, Ra ^is independently selected from H, F, Cl, _-CH3 , -CN, _CH3O- ,

In some embodiments of the present disclosure, Ra ^is independently selected from

In some embodiments of the present disclosure, Ra ^is independently selected from H, F,

In some embodiments of the present disclosure, R ^b is selected from H.

In some embodiments of the present disclosure, R ^c is independently selected from

In some embodiments of the present disclosure, the structural unit Selected from

In some embodiments of the present disclosure, the structural unit Selected from wherein ^Ra , ^Rd , and n are as defined above.

In some embodiments of the present disclosure, the structural unit Selected from wherein ^Ra and ^Rd are as defined above.

In some embodiments of the present disclosure, the structural unit Selected from Preferably, the structural unit Selected from

In some embodiments of the present disclosure, ring B is selected from phenyl, pyridyl, pyrimidinyl, pyrazinyl, piperazinyl, piperidinyl.

In some embodiments of the present disclosure, ring B is selected from

In some embodiments of the present disclosure, the structural unit Selected from wherein R ^c and m are as described above.

In some embodiments of the present disclosure, the structural unit Selected from wherein R ^c is as described above.

In some embodiments of the present disclosure, the compound of formula (I), pharmaceutically acceptable salts and stereoisomers thereof are selected from the compounds represented by formula (II), (II-a), (II-b) and (II-c), pharmaceutically acceptable salts and stereoisomers thereof.

wherein Ring B, Ring C, ^Ra , ^Rb , ^Rc , ^Rd , n, m and p are as defined above.

In some embodiments of the present disclosure, the compound of formula (I), pharmaceutically acceptable salts and stereoisomers thereof are selected from the compound of formula (III-A), pharmaceutically acceptable salts and stereoisomers thereof.

wherein ^Ra , ^Rc and ^Rd are as defined above.

in,

^Ra is selected from a 5-6 membered heteroaryl group containing 1-3 atoms selected from nitrogen atoms and/or oxygen atoms and/or sulfur atoms, wherein the 5-6 membered heteroaryl group may be optionally substituted by 1-3 ^{Rab groups} , wherein ^Rab is selected from halogen, _C1-4 alkyl;

R ^c is selected from -CH ₂ CONR ^ca R ^cb , wherein R ^ca and R ^cb are each independently selected from C _1-4 alkyl, or R ^ca and R ^cb are cyclized with the N atom to which they are commonly connected to form a 5-6 membered heterocycloalkyl; wherein the 5-6 membered heterocycloalkyl may be optionally substituted with 1-3 C _1-4 alkyl, halogen, C _1-4 haloalkyl, wherein _the C _1-4 alkyl may be optionally substituted with one or more of the following groups: hydroxyl, amino, cyano;

R ^d is selected from H, NH ₂ , OH, D, halogen, and C _1-4 alkyl.

In some embodiments of the present disclosure, the compound of formula (I), pharmaceutically acceptable salts and stereoisomers thereof are selected from the compound of formula (III-B), pharmaceutically acceptable salts and stereoisomers thereof,

wherein ^Ra , ^Rc and ^Rd are as defined above.

in,

R ^c is selected from -CH ₂ CONR ^ca R ^cb , wherein R ^ca and R ^cb are each independently selected from C _1-4 alkyl, or R ^ca and R ^cb are cyclized with the N atom to which they are commonly attached to form a 5-6 membered heterocycloalkyl, wherein the 5-6 membered heterocycloalkyl may be optionally substituted with 1-3 C _1-4 alkyl, halogen, C _1-4 haloalkyl, wherein the C _1-4 alkyl may be optionally substituted with one or more of the following groups: hydroxyl, amino, cyano;

R ^d is selected from H, NH ₂ , OH, D, halogen, and C _1-4 alkyl.

In some embodiments of the present disclosure, the compound of formula (I), pharmaceutically acceptable salts and stereoisomers thereof are selected from the compound of formula (III-C), pharmaceutically acceptable salts and stereoisomers thereof,

in,

R ^c is selected from -CH ₂ CONR ^ca R ^cb , wherein R ^ca and R ^cb are each independently selected from C _1-4 alkyl, or R ^ca and R ^cb and the N atom to which they are commonly connected are cyclized to form a 5-6-membered heterocycloalkyl, a 5-11-membered para-heterocyclyl, or a 5-11-membered spiro heterocycloalkyl; wherein the 5-6-membered heterocycloalkyl, the 5-11-membered para-heterocyclyl, or the 5-11-membered spiro heterocycloalkyl may be optionally substituted with 1-3 C _1-4 alkyl, halogen, or C _1-4 haloalkyl, wherein the C _1-4 alkyl may be optionally substituted with one or more of the following groups: hydroxyl, amino, or cyano;

R ^d is selected from H, NH ₂ , OH, D, halogen, and C _1-4 alkyl.

In some embodiments of the present disclosure, the compound of formula (I), pharmaceutically acceptable salts and stereoisomers thereof are selected from the group consisting of formula (III-D), (III-E) The compounds, pharmaceutically acceptable salts and stereoisomers thereof,

Wherein, R ^c is selected from -CH ₂ CONR ^ca R ^cb , wherein R ^ca and R ^cb are each independently selected from C _1-4 alkyl, or R ^ca and R ^cb are cyclized with the N atom to which they are commonly connected to form a 5-6 membered heterocycloalkyl; wherein the 5-6 membered heterocycloalkyl may be optionally substituted with 1-3 C _1-4 alkyl, halogen, C _1-4 haloalkyl, wherein the C _1-4 alkyl may be optionally substituted with one or more of the following groups: hydroxyl, amino, cyano;

R ^d is selected from H, NH ₂ , OH, D, halogen, and C _1-4 alkyl.

The present disclosure also provides the following compounds, pharmaceutically acceptable salts and stereoisomers thereof, wherein the compound can be selected from any of the following structures:

The present disclosure also provides a pharmaceutical composition, which contains (preferably a therapeutically effective amount) the above compound or a pharmaceutically acceptable salt thereof and a pharmaceutically acceptable carrier.

The present disclosure also provides use of the above-mentioned compound or a pharmaceutically acceptable salt thereof or the above-mentioned pharmaceutical composition in the preparation of a drug for treating CDK8/CDK19-mediated cancer.

The present disclosure also provides the above-mentioned compound or a pharmaceutically acceptable salt thereof or the above-mentioned pharmaceutical composition for use in treating CDK8/CDK19-mediated cancer.

The present disclosure also provides a method for treating CDK8/CDK19-mediated cancer, comprising administering a therapeutically effective amount of the above-mentioned compound or a pharmaceutically acceptable salt thereof or the above-mentioned pharmaceutical composition to a patient.

In some embodiments of the present disclosure, the above uses and methods, wherein the cancer is selected from hematological tumors and solid tumors;

Preferably, blood tumors include acute myeloid leukemias (AMLs), myelodysplastic syndromes (MDSs) and myeloproliferative diseases (MPDs);

Solid tumors include breast cancer, gastric cancer, colorectal cancer and pancreatic cancer.

Technical Effects

The compounds disclosed herein have good CDK8/CDK19 kinase inhibitory activity; some compounds disclosed herein have weak inhibitory activity against CDK2/7/9 kinases and can selectively inhibit CDK8/CDK19 kinases.

The disclosed compound has good tumor cell proliferation inhibition activity.

Description and Definition

Unless otherwise specified, the following terms and phrases used herein are intended to have the following meanings. A particular term or phrase should not be considered ambiguous or unclear without special definition, but should be understood according to the ordinary meaning.

The term "pharmaceutically acceptable" refers to those compounds, materials, compositions and/or dosage forms which are, within the scope of sound medical judgment, suitable for use in contact with the tissues of humans and animals without excessive toxicity, irritation, allergic response, or other problems or complications, commensurate with a reasonable benefit/risk ratio.

The term "pharmaceutically acceptable salt" refers to a derivative of the disclosed compound prepared with a relatively non-toxic acid or base. These salts can be prepared during the synthesis, separation, and purification of the compound, or the purified free form of the compound can be used alone to react with a suitable acid or base. When the compound contains a relatively acidic functional group, it reacts with an alkali metal, alkaline earth metal hydroxide or an organic amine to obtain a base addition salt, including cations based on alkali metals and alkaline earth metals and non-toxic ammonium, quaternary ammonium and amine cations, and also covers salts of amino acids. When the compound contains a relatively basic functional group, it reacts with an organic acid or an inorganic acid to obtain an acid addition salt.

The compounds disclosed herein exist in geometric isomers and stereoisomers, such as cis-trans isomers, enantiomers, diastereomers, racemic mixtures and other mixtures, all of which are within the scope of the present disclosure.

The compounds of the present disclosure exist as "tautomers". The term "tautomer" refers to a type of functional group isomer that has different points of attachment due to the displacement of one or more double bonds, for example, a ketone and its enol form are keto-enol tautomers.

The term "enantiomer" refers to stereoisomers that are mirror images of one another.

The term "diastereomer" refers to stereoisomers that have two or more chiral centers and are not mirror images of each other.

The term "cis-trans isomers" refers to configurations in which double bonds or single bonds of ring carbon atoms in a molecule cannot rotate freely.

Unless otherwise specified, the key is a solid wedge. and dotted wedge key To indicate the absolute configuration of a stereocenter, use a straight solid bond. and straight dashed key Indicates the relative configuration of a stereocenter.

Stereoisomers of the disclosed compounds can be prepared by chiral synthesis or chiral reagents or other conventional techniques. For example, one enantiomer of a compound of the disclosed compounds can be prepared by asymmetric catalysis technology or chiral auxiliary derivatization technology. Alternatively, a compound of a single stereo configuration can be obtained from a mixture by chiral resolution technology. Alternatively, it can be prepared directly using chiral starting materials. The separation of optically pure compounds in the disclosed compounds is usually accomplished by preparative chromatography, using a chiral chromatographic column to achieve the purpose of separating chiral compounds.

The absolute stereo configuration of a compound can be confirmed by conventional techniques in the art. For example, single crystal X-ray diffraction can also be used to confirm the absolute configuration of a compound by the chiral structure of the raw material and the reaction mechanism of asymmetric synthesis. Compounds marked as "absolute configuration not determined" herein are usually separated into single isomers by chiral preparative SFC from racemic compounds, and then characterized and tested.

The term "pharmaceutically acceptable carrier" refers to a medium generally acceptable in the art for delivering biologically active agents to animals, particularly mammals, including, for example, adjuvants, excipients or excipients, such as diluents, preservatives, fillers, flow regulators, disintegrants, wetting agents, emulsifiers, suspending agents, sweeteners, flavoring agents, aromatics, antibacterial agents, antifungal agents, lubricants and dispersants, depending on the nature of the mode of administration and dosage form. Pharmaceutically acceptable carriers are formulated within the scope of ordinary technicians in the field according to a large number of factors. They include, but are not limited to: the type and nature of the active agent to be formulated, the subject to whom the composition containing the agent is to be administered, the intended route of administration of the composition, and the target therapeutic indication. Pharmaceutically acceptable carriers include both aqueous and non-aqueous media and a variety of solid and semi-solid dosage forms. In addition to the active agent, such carriers include many different ingredients and additives, and such additional ingredients included in the prescription for various reasons (e.g., stabilizing the active agent, adhesives, etc.) are well known to ordinary technicians in the field.

The term "effective preventive or therapeutic amount" refers to a sufficient amount of the compound of the present disclosure, its pharmaceutically acceptable salt or its stereoisomer to treat the disorder at a reasonable effect/risk ratio applicable to any medical treatment and/or prevention. However, it should be recognized that the total daily dosage of the compound of Formula I or its pharmaceutically acceptable salt and composition of the present disclosure must be determined by the attending physician within the scope of sound medical judgment. For any specific patient, the specific therapeutically effective dosage level must be determined based on a variety of factors, including the disorder being treated and the severity of the disorder; the activity of the specific compound used; the specific composition used; the patient's age, weight, general health, sex and diet; the administration time, route of administration and excretion rate of the specific compound used; the duration of treatment; drugs used in combination or simultaneously with the specific compound used; and similar factors known in the medical field.

The term "optionally" means that it may be substituted or not substituted. Unless otherwise specified, the type and number of substituents may be arbitrary based on chemical feasibility. For example, the term "optionally substituted with one or more R _d " means that it may be substituted with one or more R _d or not substituted with R _d .

When any variable (e.g., R _d ) occurs more than once in a compound's composition or structure, its definition at each occurrence is independent. It means that the cyclopentyl group is substituted by 3 R _d , and each R _d has independent options.

When a substituent has a bond that crosses two atoms in a ring, the substituent may be bonded to any atom in the ring. It means that the substituent _R1 can be substituted at any position on the benzene ring.

When a substituent is listed without specifying the atom through which it is bonded to the compound included in the general chemical formula but not specifically mentioned, the substituent may be bonded through any atom thereof. For example, pyrazole as a substituent means that any carbon atom or nitrogen atom on the pyrazole ring is bonded to the substituent. On the group of generation; when it appears in the structure , it indicates that the atom is a bonding atom, for example It indicates that the N atom on the morpholine ring is a bonding atom.

Unless otherwise specified, "ring" refers to saturated, partially saturated or unsaturated monocyclic and polycyclic rings, and "polycyclic rings" include spirocyclic, fused or bridged rings. Representative "rings" include substituted or unsubstituted cycloalkyl, heterocycloalkyl, cycloalkenyl, heterocycloalkenyl, cycloalkynyl, heterocycloalkynyl, aryl or heteroaryl. The term "hetero" refers to substituted or unsubstituted heteroatoms and oxidized forms of heteroatoms, also known as heteroatomic groups, wherein the heteroatoms are generally selected from N, O, S, and the oxidized forms generally include NO, SO, S(O) _2. The nitrogen atom may be substituted, i.e., NR (R is H or other substituents defined herein); the number of atoms on the ring is generally defined as the number of ring members, for example, "3-6 membered heterocycloalkyl" refers to a ring formed by 3-6 atoms arranged around each ring, each ring optionally containing 1 to 3 heteroatoms, i.e., N, O, S, NO, SO, S(O) ₂ or NR, each ring optionally substituted by an R group, and R is a group defined herein.

Unless otherwise specified, "cycloalkyl" refers to a saturated monocyclic or polycyclic hydrocarbon group. The cycloalkyl group is preferably a C _3-8 monocyclic alkyl group, more preferably a C _3-6 monocyclic alkyl group, and examples of these monocyclic alkyl groups include, but are not limited to, cyclopropyl, cyclobutyl, cyclopentyl, cyclohexyl, cycloheptyl, and cyclooctyl.

Unless otherwise specified, "heterocycloalkyl" refers to mono-heterocycloalkyl and poly-heterocycloalkyl containing a certain number of heteroatoms or heteroatoms in the ring, and the heteroatoms are generally selected from N, O, S, NO, SO, S(O) ₂ and NR. Heterocycloalkyl is preferably a 3-8-membered mono-heterocycloalkyl, more preferably a 3-6-membered mono-heterocycloalkyl, and more preferably a 5-6-membered mono-heterocycloalkyl. Examples of these mono-heterocycloalkyls include, but are not limited to, oxirane, tetrahydropyrrolyl, piperidinyl, piperazinyl, morpholinyl, tetrahydrofuranyl, tetrahydrothienyl, tetrahydropyranyl, 1,3-dioxolane, 1,4-dioxane, and the like.

Unless otherwise specified, "spirocyclyl" refers to a polycyclic system in which a carbon atom (called spiro atom) is shared between substituted or unsubstituted monocyclic rings, and each monocyclic ring may contain a certain number of double bonds. Spirocyclyl is preferably a 5-13-membered spirocyclyl, a 6-12-membered spirocyclyl, or a 7-11-membered spirocyclyl. Examples of spirocyclyl include, but are not limited to, spiro[2.2]pentyl, spiro[2.3]hexyl, spiro[2.4]heptyl, spiro[2.5]octyl, spiro[2.6]nonyl, spiro[3.3]heptyl, spiro[3.4]octyl, spiro[3.5]nonyl, spiro[3.6]decyl, spiro[4.4]nonyl, spiro[4.5]decyl, spiro[4.6]undecyl, spiro[5.5]undecyl, spiro[5.6]dodecyl, spiro[6.6]tridecyl, spiro[6.7]tetradecyl.

"Spiro heterocyclyl" refers to a spirocyclic group in which one or more carbon atoms in the spirocyclic skeleton structure are replaced by a heteroatom or a heteroatom group, wherein the heteroatom is selected from N, O, S, NO, SO, S(O) ₂ , etc. The spiro heterocyclyl is preferably a 5-13-membered spiro heterocyclyl, a 6-12-membered spiro heterocyclyl, a 5-11-membered spiro heterocycloalkyl, or a 7-11-membered spiro heterocyclyl. Examples of spiroheterocyclyl groups include, but are not limited to, 2-oxa-7-azaspiro[5.3]nonan-7-yl, 2-oxa-7-azaspiro[4.4]nonan-7-yl, 2-oxa-6-azaspiro[3.3]heptan-6-yl, 2-oxa-8-azaspiro[4.5]decan-8-yl, 1,4,9-triazaspiro[5.5]undecan-9-yl, 3-oxa-9-azaspiro[5.5]undecan-9-yl, 2,6-diazaspiro[3.3]heptan-2-yl, 2,7-diazaspiro[5.3]nonan-7-yl, 2,7-dioxaspiro[5.3]nonan-7-yl, 3,9-diazaspiro[5.5]undecan-8-yl, 2,6-diazaspiro[3.3]heptan-2-yl, 2,7-diazaspiro[5.3]nonan-7-yl, 2,7-dioxaspiro[5.3]nonan-8-yl, 3,9-diazaspiro[5.5]undecan-9-yl, 3,9-diazaspiro[5.5]undecan-9-yl, 3,9-diazaspiro[5.5]undecan-8 ...8-yl, 3,9-diazaspiro[5.5]undecan-8-yl, 3,9 [5.5]undec-3-yl, 1-oxa-4,9-diazaspiro[5.5]undec-9-yl, 1-oxa-4,8-diazaspiro[5.4]dec-8-yl, 3-azaspiro[5.5]undec-3-yl, 7-azaspiro[3.5]dec-7-yl, 1-oxa-4,9-diazaspiro[5.5]undec-4-yl, 6-oxa-2,9-diazaspiro[4.5]dec-9-yl, 9-oxa-2,6-diazaspiro[4.5]dec-6-yl, 3-azaspiro[5.5]undec-3-yl, 4-oxa-1,9-diazaspiro[5.5]undec-9-yl.

Unless otherwise specified, "paracyclic" refers to a polycyclic group in which each ring in the system shares a pair of adjacent carbon atoms with other rings in the system, wherein each ring may contain a certain number of unsaturated bonds, such as double bonds. When all ring systems do not contain unsaturated bonds, it is a paracyclic heterocyclic alkyl group. Paracyclic is preferably a 5-14-membered paracyclic group, more preferably a 7-12-membered paracyclic group, more preferably an 8-10-membered paracyclic group, and more preferably an 8-membered paracyclic group. "Parally heterocyclic" refers to a paracyclic group in which one or more carbon atoms constituting the paracyclic skeleton are replaced by a heteroatom or heteroatom group, and the heteroatom is selected from N, O, S, NO, SO, S(O) ₂ , etc. Paracyclic heterocyclic groups are preferably 5-14-membered paracyclic heterocyclic groups, preferably 5-11-membered paracyclic heterocyclic groups, preferably 7-12-membered paracyclic heterocyclic groups, preferably 8-10-membered paracyclic heterocyclic groups, and more preferably 8-membered paracyclic heterocyclic groups. Examples of paracyclic heterocyclic groups include but are not limited to Unless otherwise specified, the term "aryl" refers to an unsaturated, usually aromatic hydrocarbon group, which may be a single ring or multiple rings fused together. Preferably, C _5-10 aryl, more preferably C _5-8 aryl, most preferably a monocyclic C _5-6 aryl; examples of aryl include, but are not limited to, phenyl and naphthyl.

Unless otherwise specified, the term "heteroaryl" means a stable monocyclic or polycyclic aromatic hydrocarbon containing at least one heteroatom or heteroatom group (N, O, S, NO, SO, S(O) ₂ or NR). Preferably, a 5-membered or 6-membered monocyclic heteroaryl group is used. More preferably, a 5-membered or 6-membered monocyclic heteroaryl group containing a nitrogen atom is used. Examples of heteroaryl groups include, but are not limited to

Unless otherwise specified, the term "alkyl" is used to represent a straight or branched saturated hydrocarbon group. Preferably, the alkyl group is C _1-6 , and more preferably, the alkyl group is C _1-4 . Examples of the alkyl group include, but are not limited to, methyl, ethyl, n-propyl, isopropyl, butyl, isobutyl, pentyl, isopentyl, neopentyl, n-hexyl, and the like.

Unless otherwise specified, the term "alkoxy" refers to an alkyl group connected by an oxygen bridge, that is, a group obtained by replacing the hydrogen atom in a hydroxyl group with an alkyl group. Preferably, C _1-6 alkoxy is used, and more preferably C _1-4 alkoxy is used. Examples of alkoxy include, but are not limited to, methoxy, ethoxy, n-propoxy, isopropoxy, n-butoxy, sec-butoxy, tert-butoxy, n-pentoxy, neopentoxy, and n-hexyloxy.

Unless otherwise specified, the term "halogen" means a fluorine, chlorine, bromine or iodine atom.

Unless otherwise specified, the term "haloalkyl" refers to an alkyl group in which one or more hydrogen atoms are replaced by halogen atoms. Preferably, C _1-6 haloalkyl, more preferably C _1-6 haloalkyl ₄ -haloalkyl. Examples of haloalkyl include, but are not limited to, monofluoromethyl, difluoromethyl, trifluoromethyl, trichloromethyl, tribromomethyl, 2,2,2-trifluoroethyl, 2,2,2-trichloroethyl, and the like.

Specifically, all combinations of substituents and/or variations thereof described herein are permissible only if such combinations result in stable compounds.

In the examples disclosed herein, the naming of the title compound is converted from the compound structure with the aid of Chemdraw. If there is any inconsistency between the compound name and the compound structure, it can be determined by combining relevant information and reaction routes; if it cannot be confirmed by other means, the given compound structure shall prevail.

The preparation methods of some compounds in the present disclosure refer to the preparation methods of the aforementioned similar compounds. Those skilled in the art should be aware that when using or referring to the preparation methods cited, the feed ratio of reactants, reaction solvent, reaction temperature, etc. can be appropriately adjusted according to the different reactants.

The compounds disclosed herein can be prepared by a variety of synthetic methods well known to those skilled in the art, including the specific embodiments listed below, embodiments formed by combining them with other chemical synthetic methods, and equivalent substitutions well known to those skilled in the art. Preferred embodiments include but are not limited to the examples disclosed herein.

The abbreviations and their corresponding chemical names used in the embodiments of the present disclosure are as follows:

Detailed ways

The structures of the compounds disclosed in the present invention are determined by nuclear magnetic resonance (NMR) mass spectroscopy. NMR chemical shifts (δ) are given in parts per million (ppm). NMR measurements are performed using a Bruker Ascend 400 nuclear magnetic resonance instrument, with deuterated dimethyl sulfoxide (DMSO-d ₆ ), deuterated methanol (CD ₃ OD) and deuterated chloroform (CDCl ₃ ) as the solvents, and tetramethylsilane (TMS) as the internal standard in heavy water (D ₂ O).

Ultra-high performance liquid chromatography-mass spectrometry (UPLC-MS) was performed using a Waters UPLC H-class SQD2 mass spectrometer.

HPLC determination was performed using Waters e2695-2998.

Preparative HPLC used Waters 2555-2489 (10 μm, ODS 250 cm × 5 cm).

Chiral HPLC determination used Agilent 1220, column Daicel chiralpak AD-H (5um, 4.6*250mm) and Agilent 1260, column Daicel-OJ-RH (5um, 4.6*150mm), Daicel-OD-RH (5um, 4.6*150mm), Daicel-IA (5um, 4.6*150mm).

The thin layer chromatography silica gel plate used was HSGF254 silica gel plate produced by Yantai Jiangyou Silica Gel Development Co., Ltd. The specification used for TLC was 0.20 mm ± 0.03 mm, and the preparative type was 20 x 20 cm. The column chromatography used 200-300 mesh silica gel produced by Qingdao Hailang Silica Gel Desiccant Co., Ltd. as the carrier.

The starting materials in the embodiments of the present disclosure are known and commercially available, or can be synthesized using or according to methods known in the art.

Unless otherwise specified, all reactions disclosed herein are carried out under continuous magnetic stirring in a dry nitrogen or argon atmosphere, the solvent is a dry solvent, and the reaction temperature is in degrees Celsius.

1. Preparation Example

Embodiment 1:

2-(4-(5-(isoquinolin-4-yl)pyridin-2-yl)-1H-pyrazol-1-yl)-N,N-dimethylacetamide

Steps:

Step A: 5-bromo-2-iodopyridine (500 mg, 1.76 mmol), isoquinoline-4-boronic acid (300 mg, 1.76 mmol), [1,1'-bis(diphenylphosphino)ferrocene]palladium dichloride (130 mg, 0.176 mmol) and sodium carbonate (466 mg, 4.4 mmol) were added to 1,4-dioxane/water (4/1, 15 ml), replaced with nitrogen three times, and reacted at 80 °C for 5 hours.

After TLC monitoring showed that the starting material disappeared, the mixture was cooled, ethyl acetate (20 ml) and water (10 ml) were added, stirred, and the mixture was separated. The residue was washed with water (10 ml), dried over anhydrous sodium sulfate, filtered, and the filtrate was concentrated under reduced pressure. The resulting residue was purified by silica gel column chromatography to give 4-(5-bromopyridin-2-yl)isoquinoline (220 mg).

Step B: 4-(5-bromopyridin-2-yl)isoquinoline (110 mg, 0.387 mmol), N,N-dimethyl-2-(4-(4,4,5,5-tetramethyl-1,3,2-dioxaborolan-2-yl)-1H-pyrazol-1-yl)acetamide (synthesized by reference US2007082900A1, 216 mg, 0.774 mmol), [1,1'-bis(diphenylphosphino)ferrocene]palladium dichloride (28 mg, 0.039 mmol) and sodium carbonate (82 mg, 0.774 mmol) were added to 1,4-dioxane/water (4/1, 5 ml), replaced with nitrogen three times, and reacted at 110°C for 3 hours.

After TLC monitoring showed that the starting material disappeared, the mixture was cooled, ethyl acetate (20 ml) and water (10 ml) were added, stirred, and the mixture was separated. The organic phase was washed with saturated brine (10 ml), dried over anhydrous sodium sulfate, filtered, and the filtrate was concentrated under reduced pressure. The resulting residue was purified by silica gel column chromatography to obtain 2-(4-(5-(isoquinolin-4-yl)pyridin-2-yl)-1H-pyrazol-1-yl)-N,N-dimethylacetamide (86.58 mg).

MS (ESI) M/Z: 358.2 [M+H] ⁺ .

¹ H NMR (400 MHz, DMSO-d ₆ ) δ 9.39 (s, 1H), 9.08 (d, J=2.3 Hz, 1H), 8.67 (s, 1H), 8.37–8.27 (m, 2H), 8.26–8.15 (m, 2H), 8.10 (s, 1H), 7.87–7.68 (m, 3H), 5.20 (s, 2H), 3.07 (s, 3H), 2.88 (s, 3H).

Embodiment 2:

2-(4-(6-(isoquinolin-4-yl)pyridin-3-yl)-1H-pyrazol-1-yl)-N,N-dimethylacetamide

Reaction route:

Steps:

Step A: 2-bromo-5-iodopyridine (500 mg, 1.755 mmol) and N,N-dimethyl-2-(4-(4,4,5,5-tetramethyl-1,3,2-dioxaborolan-2-yl)-1H-pyrazol-1-yl)acetamide (410 mg, 1.467 mmol) were dissolved in dioxane/water (4/1, 16 ml), sodium carbonate (388 mg, 3.658 mmol) and [1,1'-bis(diphenylphosphino)ferrocene]palladium dichloride (107 mg, 0.146 mmol) were added, the atmosphere was replaced with nitrogen three times, and the reaction was carried out at 80°C for 5 hours.

After TLC monitoring showed that the raw material disappeared, the mixture was cooled, filtered, and the filtrate was concentrated under reduced pressure. Water (20 ml) was added, and ethyl acetate (30 ml × 2 times) was used for extraction. The organic phases were combined, washed with saturated brine (20 ml), dried over anhydrous sodium sulfate, filtered, and finally concentrated under reduced pressure. The residue was slurried with petroleum ether/ethyl acetate = 1/1, filtered, and the solid was dried to obtain 2-(4-(6-bromopyridin-3-yl)-1H-pyrazol-1-yl)-N,N-dimethylacetamide (180 mg).

MS (ESI) M/Z: 309.1 [M+H] ⁺ .

Step B: Dissolve 2-(4-(6-bromopyridin-3-yl)-1H-pyrazol-1-yl)-N,N-dimethylacetamide (180 mg, 0.583 mmol) and 4-isoquinoline-boronic acid (121 mg, 0.70 mmol) in dioxane/water (4/1, 10 ml), add sodium carbonate (124 mg, 1.16 mmol) and [1,1'-bis(diphenylphosphino)ferrocene]palladium dichloride (43 mg, 0.058 mmol), replace with nitrogen three times, and react at 95 °C for 3 hours.

After LCMS monitoring showed the disappearance of the starting material, water (20 mL) was added and extracted with ethyl acetate (20 mL × 2 times). The organic phases were combined, washed with saturated brine (20 mL), then dried over anhydrous sodium sulfate, filtered, and finally concentrated under reduced pressure to give the crude product 2-(4-(6-(isoquinolin-4-yl)pyridin-3-yl)-1H-pyrazol-1-yl)-N,N-dimethylacetamide (43.89 mg).

MS (ESI) M/Z: 358.3 [M + H] ⁺ .

¹ H NMR (400 MHz, DMSO-d ₆ ) δ 9.39 (s, 1H), 9.09 (d, J=2.3 Hz, 1H), 8.67 (s, 1H), 8.35–8.28 (m, 2H), 8.25–8.17 (m, 2H), 8.11 (s, 1H), 7.87–7.70 (m, 3H), 5.20 (s, 2H), 3.07 (s, 3H), 2.88 (s, 3H).

Embodiment 3:

2-(4-(2-(isoquinolin-4-yl)pyrimidin-5-yl)-1H-pyrazol-1-yl)-N,N-dimethylacetamide

Steps:

Step A: 5-bromo-2-iodopyrimidine (500 mg, 1.75 mmol), isoquinoline-4-boronic acid (300 mg, 1.75 mmol), [1,1'-bis(diphenylphosphino)ferrocene]palladium dichloride (130 mg, 0.175 mmol) and sodium carbonate (470 mg, 4.38 mmol) were added to dioxane (12 ml) and water (3 ml), replaced with nitrogen three times, and reacted at 80 °C for 3 hours.

After TLC monitoring showed that the raw material disappeared, the mixture was cooled and filtered, and the filtrate was stirred with water (20 ml) and ethyl acetate (20 ml) to separate the liquids. The organic phase was washed with saturated brine (10 ml), dried over anhydrous sodium sulfate, and concentrated under reduced pressure. The resulting residue was purified by silica gel column chromatography to obtain 4-(5-bromopyrimidin-2-yl)isoquinoline (230 mg).

MS (ESI) M/Z: 286.1 [M+H] ⁺ .

Step B: 4-(5-bromopyrimidin-2-yl)isoquinoline (100 mg, 0.352 mmol), N,N-dimethyl-2-(4-(4,4,5,5-tetramethyl-1,3,2-dioxaborolan-2-yl)-1H-pyrazol-1-yl)acetamide (200 mg, 0.704 mmol), [1,1'-bis(diphenylphosphino)ferrocene]palladium dichloride (25 mg 0.035 mmol) and sodium carbonate (75 mg, 0.704 mmol) were added to dioxane (4 ml) and water (1 ml), the atmosphere was replaced with nitrogen three times, and the mixture was stirred at 90 °C for 3 hours.

After LCMS monitoring showed that the raw material disappeared, the mixture was cooled and filtered. Water (20 ml) and ethyl acetate (20 ml) were added to the filtrate and stirred to separate the liquids. The organic phase was washed with saturated brine (10 ml), dried over anhydrous sodium sulfate, and concentrated under reduced pressure. The resulting crude product was used to prepare 2-(4-(2-(isoquinolin-4-yl)pyrimidin-5-yl)-1H-pyrazol-1-yl)-N,N-dimethylacetamide (59.97 mg).

MS (ESI) M/Z: 359.2 [M+H] ⁺ .

¹ H NMR (400 MHz, DMSO-d ₆ ) δ 9.45 (s, 1H), 9.33 (s, 2H), 9.11 (s, 1H), 8.92–8.85 (m, 1H), 8.40 (s, 1H), 8.26 (d, J=8.2 Hz, 1H), 8.19 (s, 1H), 7.93–7.84 (m, 1H), 7.82–7.73 (m, 1H), 5.23 (s, 2H), 3.07 (s, 3H), 2.89 (s, 3H).

Embodiment 4:

2-(4-(5-(isoquinolin-4-yl)pyrimidin-2-yl)-1H-pyrazol-1-yl)-N,N-dimethylacetamide

Steps:

Step A: 5-bromo-2-iodopyrimidine (500 mg, 1.755 mmol) and N,N-dimethyl-2-(4-(4,4,5,5-tetramethyl-1,3,2-dioxaborolan-2-yl)-1H-pyrazol-1-yl)acetamide (408 mg, 1.463 mmol) were dissolved in dioxane/water (4/1, 16 ml), sodium carbonate (388 mg, 3.658 mmol) and [1,1'-bis(diphenylphosphino)ferrocene]palladium dichloride (107 mg, 0.146 mmol) were added, the atmosphere was replaced with nitrogen three times, and the reaction was carried out at 80°C for 5 hours.

After TLC monitoring showed that the raw material disappeared, the mixture was cooled, filtered, and the filtrate was concentrated under reduced pressure. Water (20 ml) was added, and ethyl acetate (30 ml × 2 times) was used for extraction. The organic phases were combined, washed with saturated brine (20 ml), dried over anhydrous sodium sulfate, filtered, and finally concentrated under reduced pressure. The residue was slurried with petroleum ether/ethyl acetate = 1/1, filtered, and the solid was dried to obtain 2-(4-(5-bromopyrimidin-2-yl)-1H-pyrazol-1-yl)-N,N-dimethylacetamide (190 mg).

MS (ESI) M/Z: 310.1 [M+H] ⁺ .

Step B: Dissolve 2-(4-(5-bromopyrimidin-2-yl)-1H-pyrazol-1-yl)-N,N-dimethylacetamide (190 mg, 0.613 mmol) and 4-isoquinolin-ylboronic acid (127 mg, 0.736 mmol) in dioxane/water (4/1, 10 ml), add sodium carbonate (130 mg, 1.226 mmol) and [1,1'-bis(diphenylphosphino)ferrocene]palladium dichloride (45 mg, 0.061 mmol), replace with nitrogen three times, and react at 95°C for 3 hours.

After LCMS monitoring showed that the starting material disappeared, water (20 mL) was added and extracted with ethyl acetate (20 mL × 2 times). The organic phases were combined and the organic phases were first The reaction mixture was washed with saturated brine (20 ml), dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure to give 2-(4-(5-(isoquinolin-4-yl)pyrimidin-2-yl)-1H-pyrazol-1-yl)-N,N-dimethylacetamide (43.89 mg).

MS (ESI) M/Z: 359.3 [M + H] ⁺ .

¹ H NMR (400 MHz, DMSO-d ₆ ) δ 9.44 (s, 1H), 8.99 (s, 2H), 8.59 (s, 1H), 8.41 (s, 1H), 8.28 (d, J=8.1 Hz, 1H), 8.14 (s, 1H), 7.96–7.90 (m, 1H), 7.89–7.83 (m, 1H), 7.83–7.76 (m, 1H), 5.25 (s, 2H), 3.07 (s, 3H), 2.86 (s, 3H).

Embodiment 5:

2-(4-(5-(isoquinolin-4-yl)pyrazol-2-yl)-1H-pyrazol-1-yl)-N,N-dimethylacetamide

Steps:

Step A: 5-bromo-2-iodopyridine (500 mg, 1.76 mmol), isoquinoline-4-boronic acid (300 mg, 1.73 mmol), [1,1'-bis(diphenylphosphino)ferrocene]palladium dichloride (107 mg, 0.146 mmol) and sodium carbonate (387.7 mg, 3.658 mmol) were added to 1,4-dioxane/water (4/1, 15 ml), replaced with nitrogen three times, and reacted at 80°C for 5 hours.

After TLC monitoring showed that the starting material disappeared, the mixture was cooled, ethyl acetate (20 ml) and water (10 ml) were added, stirred, and the liquids were separated. The organic phase was washed with saturated brine (10 ml), dried over anhydrous sodium sulfate, filtered, and the filtrate was concentrated under reduced pressure. The resulting residue was purified by silica gel column chromatography to obtain 4-(5-bromopyrazin-2-yl)isoquinoline (188 mg).

MS (ESI) M/Z: 286.3 [M + H] ⁺ .

Step B: 4-(5-bromopyrazin-2-yl)isoquinoline (188 mg, 0.66 mmol), N,N-dimethyl-2-(4-(4,4,5,5-tetramethyl-1,3,2-dioxaborolan-2-yl)-1H-pyrazol-1-yl)acetamide (368 mg, 1.32 mmol), [1,1'-bis(diphenylphosphino)ferrocene]palladium dichloride (48 mg, 0.066 mmol) and sodium carbonate (140 mg, 1.32 mmol) were added to 1,4-dioxane/water (4/1, 10 ml), replaced with nitrogen three times, and reacted at 110°C for 3 hours.

After TLC monitoring showed that the starting material disappeared, the mixture was cooled, ethyl acetate (20 ml) and water (10 ml) were added, stirred, and the mixture was separated. The organic phase was washed with saturated brine (10 ml), dried over anhydrous sodium sulfate, filtered, and the filtrate was concentrated under reduced pressure. The resulting residue was purified by column chromatography to give 2-(4-(5-(isoquinolin-4-yl)pyridin-2-yl)-1H-pyrazol-1-yl)-N,N-dimethylacetamide (61.69 mg).

MS (ESI) M/Z: 359.3 [M + H] ⁺ .

¹ H NMR (400 MHz, DMSO-d ₆ ) δ 9.45 (s, 1H), 9.22 (s, 1H), 8.97 (s, 1H), 8.76 (s, 1H), 8.46 (s, 1H), 8.33–8.16 (m, 3H), 7.90–7.72 (m, 2H), 5.25 (s, 2H), 3.07 (s, 3H), 2.89 (s, 3H).

Embodiment 6:

2-(4-(4-(isoquinolin-4-yl)phenyl)-1H-pyrazol-1-yl)-N,N,2-trimethylpropanamide

Steps:

Step A: Ethyl 2-(4-(4-bromophenyl)-1H-pyrazol-1-yl)-2-methylpropanoate (585 mg, 1.74 mmol) was dissolved in tetrahydrofuran (5 ml) and ethanol (5 ml). A solution of lithium hydroxide monohydrate (292.18 mg, 6.96 mmol) in water (5 ml) was added thereto, and the reaction solution was stirred at 60 degrees Celsius for 2 hours.

After TLC monitoring showed that the starting material disappeared, the mixture was cooled and the organic solvent was concentrated under reduced pressure. The residual aqueous phase was acidified with 1M hydrochloric acid and extracted with ethyl acetate (30 ml). The organic phase was washed with saturated brine (10 ml), dried over anhydrous sodium sulfate, and concentrated under reduced pressure to give 2-(4-(4-bromophenyl)-1H-pyrazol-1-yl)-2-methylpropanoic acid (494 mg).

Step B: 2-(4-(4-bromophenyl)-1H-pyrazol-1-yl)-2-methylpropanoic acid (494 mg, 1.60 mmol) and dimethylamine hydrochloride (394 mmol) were mixed. g, 4.81 mmol) was dissolved in N,N-dimethylformamide (6 ml), and HATU (915 mg, 2.41 mmol) and diisopropylethylamine (1.04 g, 8.02 mmol) were added under ice-water bath. After the addition, the temperature was raised to room temperature and the reaction was allowed to react for 2 hours.

After LCMS monitoring showed the disappearance of the starting material, aqueous sodium bicarbonate solution (20 ml) was added, and the mixture was extracted with ethyl acetate (10 ml × 2 times). The organic phases were combined, washed with saturated brine (10 ml), then dried over anhydrous sodium sulfate, filtered, and finally concentrated under reduced pressure. The crude product was purified by column chromatography to give 2-(4-(4-bromophenyl)-1H-pyrazol-1-yl)-N,N,2-trimethylpropanamide (510 mg).

MS (ESI) M/Z: 336.2 [M + H] ⁺ .

Step C: Add 2-(4-(4-bromophenyl)-1H-pyrazol-1-yl)-N,N,2-trimethylpropanamide (100 mg, 0.299 mmol), isoquinoline-4-boronic acid (77.5 mg, 0.448 mmol), [1,1'-bis(diphenylphosphino)ferrocene]palladium dichloride (21.9 mg, 0.03 mmol) and sodium carbonate (79.2 mg, 0.75 mmol) to 1,4-dioxane/water (4/1, 2.5 ml), replace with nitrogen three times, and react at 95 °C overnight.

After TLC monitoring showed that the starting material disappeared, the mixture was cooled, ethyl acetate (20 ml) and water (10 ml) were added, stirred, and the mixture was separated. The organic phase was washed with saturated brine (10 ml), dried over anhydrous sodium sulfate, filtered, and the filtrate was concentrated under reduced pressure. The resulting residue was purified by silica gel column chromatography to obtain 2-(4-(4-(isoquinolin-4-yl)phenyl)-1H-pyrazol-1-yl)-N,N,2-trimethylpropanamide (60.27 mg).

MS (ESI) M/Z: 385.4 [M + H] ⁺ .

¹ H NMR (400 MHz, DMSO-d ₆ ) δ 9.35 (s, 1H), 8.48 (d, J=8.3 Hz, 2H), 8.24 (d, J=7.9 Hz, 1H), 8.10 (s, 1H), 7.94–7.89 (m, 1H), 7.88–7.71 (m, 4H), 7.59–7.51 (m, 2H), 2.85 (s, 3H), 2.39 (s, 3H), 1.75 (s, 6H).

Embodiment 7:

2-(4-(4-(isoquinolin-4-yl)phenyl)-1H-1,2,3-triazol-1-yl)-N,N-dimethylacetamide

Steps:

Step A: 1-Bromo-4-ethynylbenzene (983 mg, 5.43 mmol) and ethyl 2-azidoacetate (500 mg, 3.88 mmol) were added to tert-butanol (10 ml), followed by the addition of cuprous iodide (36 mg, 0.19 mmol) and triethylamine (78 mg, 0.78 mmol), and the reaction mixture was stirred at 50 °C for 12 hours.

After TLC monitoring showed that the starting material disappeared, the reaction solution was concentrated under reduced pressure. The resulting residue was purified by silica gel column chromatography to give ethyl 2-(4-(4-bromophenyl)-1H-1,2,3-triazol-1-yl)acetate (1.1 g).

MS (ESI) M/Z: 310.36 [M+H] ⁺ .

Step B: Dissolve ethyl 2-(4-(4-bromophenyl)-1H-1,2,3-triazol-1-yl)acetate (200 mg, 0.654 mmol) and 4-isoquinolin-ylboronic acid (134 mg, 0.774 mmol) in dioxane/water (4/1, 10 ml), add sodium carbonate (137 mg, 1.29 mmol) and [1,1'-bis(diphenylphosphino)ferrocene]palladium dichloride (41 mg, 0.064 mmol), replace with nitrogen three times, and heat to 90 degrees Celsius for reaction overnight.

After TLC monitoring showed that the starting material disappeared, the reaction solution was concentrated to dryness under reduced pressure. The obtained crude 2-(4-(4-(isoquinolin-4-yl)phenyl)-1H-1,2,3-triazol-1-yl)acetic acid was directly used in the next step.

MS (ESI) M/Z: 331.0 [M+H] ⁺ .

Step C: Dissolve crude 2-(4-(4-(isoquinolin-4-yl)phenyl)-1H-1,2,3-triazol-1-yl)acetic acid (0.645 mmol) and dimethylamine hydrochloride (208 mg, 2.58 mmol) in DMF (6 ml), add 2-(7-azabenzotriazole)-N,N,N',N'-tetramethyluronium hexafluorophosphate (490 mg, 1.29 mmol) and N,N-diisopropylethylamine (416 mg, 3.225 mmol), and stir the reaction solution at room temperature overnight.

After LCMS monitoring showed the disappearance of the starting material, water (20 mL) was added, extracted with ethyl acetate (30 mL × 2 times), and the organic phases were combined, washed with saturated brine (20 mL), then dried over anhydrous sodium sulfate, filtered, and finally concentrated under reduced pressure. The resulting residue was purified by column chromatography to give 2-(4-(4-(isoquinolin-4-yl)phenyl)-1H-1,2,3-triazol-1-yl)-N,N-dimethylacetamide (46.07 mg).

MS (ESI) M/Z: 358.4 [M+H] ⁺ .

¹ H NMR (400 MHz, DMSO-d ₆ ) δ 9.37 (s, 1H), 8.57 (s, 1H), 8.51 (s, 1H), 8.25 (dd, J=7.9, 1.4 Hz, 1H), 8.10–8.03 (m, 2H), 7.96–7.90 (m, 1H), 7.86–7.80 (m, 1H), 7.79–7.73 (m, 1H), 7.68–7.63 (m, 2H), 5.54 (s, 2H), 3.11 (s, 3H), 2.91 (s, 3H).

Embodiment 8:

N,N-Dimethyl-2-(4-(4-(pyrido[3,4-b]pyrazin-8-yl)phenyl)-1H-pyrazol-1-yl)acetamide

Step A: 8-bromopyrido[3,4-b]pyrazine (100 mg, 0.476 mmol), N,N-dimethyl-2-(4-(4-(4,4,5,5-tetramethyl-1,3,2-dioxaborolan-2-yl)phenyl)-1H-pyrazol-1-yl)acetamide (prepared by the synthesis method of reference document EP3831829A1, 254 mg, 0.714 mmol), [1,1'-bis(diphenylphosphino)ferrocene]palladium dichloride (35 mg, 0.048 mmol) and sodium carbonate (126 mg, 1.19 mmol) were added to ethylene glycol dimethyl ether/water (5/1, 6 ml), replaced with nitrogen three times, and reacted at 90°C overnight.

After TLC monitoring showed that the starting material disappeared, the mixture was cooled, ethyl acetate (20 ml) and water (10 ml) were added, stirred, and the mixture was separated. The organic phase was washed with saturated brine (10 ml), dried over anhydrous sodium sulfate, filtered, and the filtrate was concentrated under reduced pressure. The resulting residue was subjected to column chromatography to obtain N,N-dimethyl-2-(4-(4-(pyrido[3,4-b]pyrazin-8-yl)phenyl)-1H-pyrazol-1-yl)acetamide (47.87 mg).

MS (ESI) M/Z: 359.3 [M + H] ⁺ .

¹ H NMR (400 MHz, DMSO-d ₆ ) δ 9.51 (s, 1H), 9.26–9.19 (m, 1H), 9.18–9.11 (m, 1H), 8.95 (s, 1H), 8.18 (s, 1H), 7.98 (s, 1H), 7.80–7.71 (m, 4H), 5.16 (s, 2H), 3.07 (s, 3H), 2.88 (s, 3H).

Embodiment 9:

2-(4-(4-(2,3-dimethylpyrido[3,4-b]pyrazin-8-yl)phenyl)-1H-pyrazol-1-yl)-N,N-dimethylacetamide

Steps:

Step A: 5-bromopyridine-3,4-diamine (1 g, 0.005 mol) and 2,3-butanedione (0.43 g, 0.005 mol) were added to ethanol (50 ml) and reacted at room temperature for 2 hours.

After TLC monitoring showed the disappearance of the starting material, the reaction solution was concentrated under reduced pressure to dryness to give 8-bromo-2,3-dimethylpyrido[3,4-b]pyrazine (1.2 g).

MS (ESI) M/Z: 238.1 [M+H] ⁺ .

Step B: 8-Bromo-2,3-dimethylpyrido[3,4-b]pyrazine (150 mg, 0.63 mmol) and N,N-dimethyl-2-(4-(4-(4,4,5,5-tetramethyl-1,3,2-dioxaborolan-2-yl)phenyl)-1H-pyrazol-1-yl)acetamide (335 mg, 0.95 mmol) were dissolved in dioxane/water (4/1, 10 ml), sodium carbonate (167 mg, 1.57 mmol) and [1,1'-bis(diphenylphosphino)ferrocene]palladium dichloride (46 mg, 0.06 mmol) were added, the atmosphere was replaced with nitrogen three times, and the mixture was stirred at 110°C for 2 hours.

After LCMS monitoring showed the disappearance of the starting material, water (20 mL) was added, extracted with ethyl acetate (30 mL × 2 times), and the organic phases were combined. The organic phase was first washed with saturated brine (20 mL), dried over anhydrous sodium sulfate, filtered, and finally concentrated under reduced pressure. The resulting residue was purified by column chromatography to give 2-(4-(4-(2,3-dimethylpyrido[3,4-b]pyrazin-8-yl)phenyl)-1H-pyrazol-1-yl)-N,N-dimethylacetamide (84.34 mg).

MS (ESI) M/Z: 387.4 [M+H] ⁺ .

¹ H NMR (400 MHz, DMSO-d ₆ ) δ 9.32 (s, 1H), 8.81 (s, 1H), 8.17 (s, 1H), 7.98 (s, 1H), 7.80–7.70 (m, 4H), 5.16 (s, 2H), 3.07 (s, 3H), 2.88 (s, 3H), 2.75 (s, 3H), 2.71 (s, 3H).

Embodiment 10:

2-(4-(4-(8-Fluoroisoquinolin-4-yl)phenyl)-1H-pyrazol-1-yl)-N,N-dimethylacetamide

By referring to the synthesis method of Example 8, 2-(4-(4-(8-fluoroisoquinolin-4-yl)phenyl)-1H-pyrazol-1-yl)-N,N-dimethylacetamide (27.77 mg) was obtained.

MS (ESI) M/Z: 375.6 [M + H] ⁺ .

¹ H NMR (400 MHz, DMSO-d ₆ ) δ 9.51 (s, 1H), 8.60 (s, 1H), 8.19 (s, 1H), 8.00 (s, 1H), 7.86–7.72 (m, 4H), 7.60–7.53 (m, 3H), 5.17 (s, 2H), 3.07 (s, 3H), 2.88 (s, 3H).

Embodiment 11:

2-(4-(4-(7-fluoroisoquinolin-4-yl)phenyl)-1H-pyrazol-1-yl)-N,N-dimethylacetamide

By referring to the synthesis method of Example 8, 2-(4-(4-(7-fluoroisoquinolin-4-yl)phenyl)-1H-pyrazol-1-yl)-N,N-dimethylacetamide (19.87 mg) was obtained.

MS (ESI) M/Z: 375.6 [M + H] ⁺ .

¹ H NMR (400 MHz, DMSO-d ₆ ) δ 9.34 (s, 1H), 8.48 (s, 1H), 8.19 (s, 1H), 8.05 (dd, J=9.3, 2.7 Hz, 1H), 8.04–7.95 (m, 2H), 7.82–7.68 (m, 3H), 7.55 (d, J=8.1 Hz, 2H), 5.17 (s, 2H), 3.07 (s, 3H), 2.88 (s, 3H).

Embodiment 12:

4-(4-(1-(2,2,2-trifluoroethyl)-1H-pyrazol-4-yl)phenyl)isoquinoline

Steps:

Step A: 4-isoquinolineboronic acid (1 g, 5.78 mmol), p-bromoiodobenzene (2.45 g, 8.67 mmol), potassium phosphate (2.45 g, 11.56 mmol) and [1,1'-bis(diphenylphosphino)ferrocene]palladium dichloride (211.3 mg, 0.289 mmol) were added to N,N-dimethylformamide (20 ml) and water (4 ml), replaced with nitrogen three times, and heated to 90°C for reflux overnight.

After TLC monitoring showed that the raw material disappeared, the mixture was cooled. Water (50 ml) and ethyl acetate (30 ml) were added and stirred for separation. The organic phase was washed with saturated brine (20 ml), dried over anhydrous sodium sulfate, and concentrated under reduced pressure. The obtained residue was purified by silica gel column chromatography to obtain 4-(4-bromophenyl)isoquinoline (800 mg).

MS (ESI) M/Z: 284.3 [M + H] ⁺ .

Step B: 4-(4-Bromophenyl)isoquinoline (800 mg, 2.83 mmol), 4-(4,4,5,5-tetramethyl-1,3,2-dioxaborolan-2-yl)-1H-pyrazole (658.1 mg, 3.39 mmol), potassium carbonate (780.2 mg, 5.65 mmol) and [1,1'-bis(diphenylphosphino)ferrocene]palladium dichloride (206.64 mg, 0.28 mmol) were added to dioxane (16 ml) and water (4 ml), replaced with nitrogen three times, and heated to 100°C for reflux for 3 hours.

After TLC monitoring showed that the raw material disappeared, the mixture was cooled. Water (50 ml) and ethyl acetate (30 ml) were added and stirred to separate the liquids. The organic phase was washed with saturated brine (30 ml), dried over anhydrous sodium sulfate, and concentrated under reduced pressure. The resulting residue was purified by silica gel column chromatography to obtain 4-(4-(1H-pyrazol-4-yl)phenyl)isoquinoline (430 mg).

Step C: 4-(4-(1H-pyrazol-4-yl)phenyl)isoquinoline (100 mg, 0.37 mmol), 2,2,2-trifluoroethyltrifluoromethanesulfonic acid (1.11 g, 4.80 mmol) and potassium carbonate (101.84 mg, 0.74 mmol) were added to DMF (5 ml) and reacted at room temperature overnight.

After TLC monitoring showed that the raw material disappeared, the mixture was cooled. Water (20 ml) and ethyl acetate (30 ml) were added and stirred to separate the liquids. The organic phase was washed with saturated brine (30 ml), dried over anhydrous sodium sulfate, and concentrated under reduced pressure. The obtained residue was prepared and purified to obtain 4-(4-(1-(2,2,2-trifluoroethyl)-1H-pyrazol-4-yl)phenyl)isoquinoline (2.29 mg).

MS (ESI) M/Z: 354.4 [M + H] ⁺ .

¹ H NMR (400 MHz, DMSO-d ₆ ) δ 9.35 (s, 1H), 8.48 (s, 1H), 8.40 (s, 1H), 8.24 (d, J=7.9 Hz, 1H), 8.16 (s, 1H), 7.92 (d, J=8.4 Hz, 1H), 7.85–7.78 (m, 3H), 7.78–7.71 (m, 1H), 7.62–7.52 (m, 2H), 5.27–5.13 (m, 2H).

Embodiment 13:

2-(4-(4-(isoquinolin-4-yl)phenyl)-1H-pyrazol-1-yl)acetonitrile

Steps:

Step A: Dissolve 4-(4-bromophenyl)-1H-pyrazole (300 mg, 1.35 mmol) in DMF (8 ml) and cool to 0°C in an ice bath. Add sodium hydride (60%, 81.1 mg, 2.03 mmol), stir for 30 minutes, then add bromoacetonitrile (243.24 mg, 2.03 mmol). Stir the reaction mixture at room temperature overnight.

After TLC monitoring showed that the raw material disappeared, an aqueous solution of ammonium chloride (30 ml) was added to quench the reaction. Ethyl acetate (30 ml) was added for extraction and separation. The organic phase was washed with saturated brine, dried over sodium sulfate, and concentrated to dryness under reduced pressure. The resulting residue was purified by silica gel column chromatography to obtain 2-(4-(4-bromophenyl)-1H-pyrazol-1-yl)acetonitrile (250 mg).

Step B: 2-(4-(4-bromophenyl)-1H-pyrazol-1-yl)acetonitrile (180 mg, 0.69 mmol), 4-isoquinolineboronic acid (178.97 mg, 1.04 mmol) were dissolved in dioxane (8 ml) and water (2 ml), and potassium carbonate (190.35 mg, 1.38 mmol) and 1,1'-bis(diphenylphosphinoferrocene)palladium dichloride (50.55 mg, 0.069 mmol) were added. The reaction liquid was replaced with nitrogen three times and reacted at 105 degrees Celsius for 3 hours.

After TLC monitoring of the disappearance of the starting material, water (10 ml) was added, and the mixture was extracted with ethyl acetate (20 ml × 2 times). The organic phases were combined, washed with saturated brine (50 ml), then dried over anhydrous sodium sulfate, filtered, and finally concentrated under reduced pressure. The crude product was purified by silica gel column chromatography to give 2-(4-(4-(isoquinolin-4-yl)phenyl)-1H-pyrazol-1-yl)acetonitrile (17.57 mg).

MS (ESI) M/Z: 311.2 [M+H] ⁺ .

¹ H NMR (400 MHz, DMSO-d ₆ ) δ 9.36 (s, 1H), 8.48 (s, 1H), 8.40 (s, 1H), 8.28–8.22 (m, 1H), 8.17 (s, 1H), 7.95–7.89 (m, 1H), 7.85–7.71 (m, 4H), 7.62–7.54 (m, 2H), 5.57 (s, 2H).

Embodiment 14:

N,N-Dimethyl-2-(4-(4-(3-(5-methyl-1,3,4-oxadiazol-2-yl)-1,7-naphthyridin-5-yl)phenyl)-1H-pyrazol-1-yl)acetamide

Steps:

Step A: Add 1,7-naphthyridine-3-carboxylic acid ethyl ester (1.0 g, 4.9 mmol) and NBS (1.005 mg, 5.9 mmol) to acetic acid (100 ml), replace with nitrogen three times, and react at 80 °C for 1 hour. After TLC monitoring shows that the starting material disappears, cool and filter, extract the filtrate with dichloromethane twice, wash with saturated brine, combine the filtrate, and concentrate under reduced pressure. The residue is purified by silica gel column chromatography to give 5-bromo-1,7-naphthyridine-3-carboxylic acid ethyl ester (760 mg).

MS (ESI) M/Z: 281.2 [M+H] ⁺ .

Step B: 5-bromo-1,7-naphthyridine-3-carboxylic acid ethyl ester (630 mg, 2.24 mmol) and hydrazine hydrate (630 mg) were added to 20 ml of methanol and stirred at 80 degrees Celsius overnight. After LCMS monitoring showed that the starting material disappeared, sodium bicarbonate aqueous solution (10 ml) was added, and the mixture was extracted with dichloromethane (10 ml × 2 times), and the organic phases were combined, washed with saturated brine (50 ml), then dried over anhydrous sodium sulfate, filtered, and finally concentrated under reduced pressure to obtain 5-bromo-1,7-naphthyridine-3-carbohydrazide crude product (480 mg), which was directly used in the next step.

MS (ESI) M/Z: 267.2 [M+H] ⁺ .

Step C: Dissolve the crude 5-bromo-1,7-naphthyridine-3-carbohydrazide (480 mg, 1.80 mmol) in 5 ml of acetic acid and react overnight at 95 degrees. The acetic acid was dried by rotary evaporation, phosphorus oxychloride (5 ml) was added, and refluxed at 120 degrees for 1.5 hours. After TLC monitoring showed that the raw material disappeared, it was cooled, potassium carbonate was added to adjust the reaction solution to alkalinity, filtered, and the filtrate was extracted with dichloromethane. The organic phase was dried over anhydrous sodium sulfate and concentrated under reduced pressure. The residue was purified by silica gel column chromatography to obtain 2-(5-bromo-1,7-naphthyridine-3-yl)-5-methyl-1,3,4-oxadiazole (330 mg).

MS (ESI) M/Z: 291.0 [M+H] ⁺

Step D: 2-(5-bromo-1,7-naphthyridin-3-yl)-5-methyl-1,3,4-oxadiazole (100 mg, 0.34 mmol), N,N-dimethyl-2-(4-(4-(4,4,5,5-tetramethyl-1,3,2-dioxaborolan-2-yl)phenyl)-1H-pyrazol-1-yl)acetamide (183 mg, 0.52 mmol), potassium carbonate (95 mg, 0.69 mmol), [1,1'-bis(diphenylphosphino)ferrocene]palladium dichloride (20 mg, 0.02 mmol) were added to 1,4-dioxane (2 ml) and water (1 ml), replaced with nitrogen three times, and reacted at 100 °C for 2 hours.

After TLC monitoring showed that the raw material disappeared, the mixture was cooled and water (20 ml) was added. The mixed solution was extracted with dichloromethane (20 ml × 2 times), and the organic phases were combined. The organic phases were first washed with saturated brine (20 ml), then dried over anhydrous sodium sulfate, filtered, and finally concentrated under reduced pressure. The resulting residue was purified by silica gel column chromatography to obtain N, N-dimethyl-2-(4-(4-(3-(5-methyl-1,3,4-oxadiazol-2-yl)-1,7-naphthyridin-5-yl)phenyl)-1H-pyrazol-1-yl)acetamide (62.23 mg).

MS (ESI) M/Z: 440.4 [M+H] ⁺ .

¹ H NMR (400 MHz, DMSO-d ₆ ) δ 9.62 (d, J = 2.1 Hz, 1H), 9.55 (s, 1H), 8.77 (s, 1H), 8.74-8.70 (m, 1H), 8.24 (s, 1H), 8.04 (s, 1H), 7.86 (d, J = 8.1 Hz, 2H), 7.65 (d, J = 8.2 Hz, 2H), 5.18 (s, 2H), 3.07 (s, 3H), 2.89 (s, 3H), 2.62 (s, 3H).

Embodiment 15:

4'-(Isoquinolin-4-yl)-N,N-dimethyl-[1,1'-biphenyl]-3-carboxamide

Steps:

Step A: Dissolve (3-(dimethylcarbamoyl)phenyl)boric acid (500 mg, 2.59 mmol) and p-bromoiodobenzene (879.3 mg, 3.11 mmol) in dioxane/water (4/1, 16 ml), add sodium carbonate (686.4 mg, 6.48 mmol) and [1,1'-bis(diphenylphosphino)ferrocene]palladium dichloride (189.9 mg, 0.26 mmol), replace with nitrogen three times, and react at 85 °C for 5 hours.

After TLC monitoring showed the disappearance of the starting material, the mixture was cooled, water (20 mL) was added, and extraction was performed with ethyl acetate (30 mL × 2 times). The organic phases were combined, washed with saturated brine (20 mL), then dried over anhydrous sodium sulfate, filtered, and finally concentrated under reduced pressure. The residue was subjected to column chromatography to give 4'-bromo-N,N-dimethyl-[1,1'-biphenyl]-3-carboxamide (655 mg).

MS (ESI) M/Z: 304.2 [M + H] ⁺ .

Step B: Dissolve 4'-bromo-N,N-dimethyl-[1,1'-biphenyl]-3-carboxamide (200 mg, 0.657 mmol) and 4-isoquinoline-boronic acid (136 mg, 0.788 mmol) in dioxane/water (4/1, 10 ml), add sodium carbonate (139 mg, 1.31 mmol) and [1,1'-bis(diphenylphosphino)ferrocene]dichloropalladium (48 mg, 0.066 mmol), replace with nitrogen three times, and react at 95 °C overnight.

After LCMS monitoring showed the disappearance of the starting material, water (20 mL) was added and extracted with ethyl acetate (20 mL × 2 times). The organic phases were combined, washed with saturated brine (20 mL), then dried over anhydrous sodium sulfate, filtered, and finally concentrated under reduced pressure. The crude product was purified by column chromatography to give 4'-(isoquinolin-4-yl)-N,N-dimethyl-[1,1'-biphenyl]-3-carboxamide (65.68 mg).

MS (ESI) M/Z: 353.3 [M + H] ⁺ .

¹ H NMR (400 MHz, DMSO-d ₆ ) δ 9.38 (s, 1H), 8.50 (s, 1H), 8.30–8.19 (m, 1H), 7.96–7.88 (m, 3H), 7.88–7.81 (m, 2H), 7.79–7.72 (m, 2H), 7.70–7.62 (m, 2H), 7.61–7.54 (m, 1H), 7.48–7.39 (m, 1H), 3.02 (s, 3H), 2.98 (s, 3H).

Embodiment 16:

4-(4-(isoquinolin-4-yl)phenyl)morpholine

Steps:

Step A: Dissolve 4-(4-bromophenyl)morpholine (100 mg, 0.41 mmol) and 4-isoquinolineboronic acid (85 mg, 0.50 mmol) in dioxane (8 ml) and water (2 ml), then add potassium carbonate (114 mg, 0.83 mmol) and 1,1-bis(diphenylphosphinoferrocene)palladium dichloride (30 mg, 0.041 mmol). Replace the reaction liquid with nitrogen three times and react at 105 degrees Celsius for 3 hours.

The raw material disappeared after TLC monitoring. Water (10 ml) was added and the mixture was extracted with ethyl acetate (20 ml x 2 times). The organic phases were combined. The organic phase was first washed with saturated brine (20 ml), then dried over anhydrous sodium sulfate, filtered, and finally concentrated under reduced pressure. The crude product was purified by column chromatography to obtain 4-(4-(isoquinolin-4-yl)phenyl)morpholine (84.2 mg).

MS (ESI) M/Z: 290.9 [M + H] ⁺ .

¹ H NMR (400 MHz, DMSO-d ₆ ) δ 9.29 (s, 1H), 8.41 (s, 1H), 8.20 (d, J = 8.0 Hz, 1H), 7.92 (d, J = 8.4 Hz, 1H), 7.83-7.67 (m, 2H), 7.43 (d, J = 8.2 Hz, 2H), 7.13 (d, J = 8.3 Hz, 2H), 3.88-3.71 (m, 4H), 3.26-3.18 (m, 4H).

Embodiment 17:

2-(4-(4-(1,7-naphthyridin-5-yl)phenyl)-1H-pyrazol-1-yl)-N,N-dimethylacetamide

By referring to the synthesis method of Example 8, 2-(4-(4-(1,7-naphthyridin-5-yl)phenyl)-1H-pyrazol-1-yl)-N,N-dimethylacetamide (84.2 mg) was obtained.

MS (ESI) M/Z: 358.3 [M + H] ⁺ .

¹ H NMR (400 MHz, DMSO-d ₆ ) δ 9.44 (s, 1H), 9.12 (dd, J=4.1, 1.6 Hz, 1H), 8.63 (s, 1H), 8.40–8.32 (m, 1H), 8.21 (s, 1H), 8.01 (s, 1H), 7.85–7.76 (m, 3H), 7.61–7.54 (m, 2H), 5.17 (s, 2H), 3.07 (s, 3H), 2.88 (s, 3H).

Embodiment 18:

4-(4-(1-methyl-1H-indazol-6-yl)phenyl)isoquinoline

Steps:

Step A: Dissolve (1-methyl-1H-indazol-6-yl)boronic acid (500 mg, 2.84 mmol) and p-bromoiodobenzene (1.21 g, 4.26 mmol) in DMF (10 ml) and water (2.5 ml), then add potassium phosphate (1.21 g, 5.68 mmol) and 1,1'-bis(diphenylphosphinoferrocene)palladium dichloride (103 mg, 0.14 mmol). Replace the reaction liquid with nitrogen three times and react at 90 degrees Celsius overnight.

After the disappearance of the starting material by TLC monitoring, water (30 ml) was added, and the mixture was extracted with ethyl acetate (30 ml × 2 times). The organic phases were combined, washed with saturated brine (50 ml), then dried over anhydrous sodium sulfate, filtered, and finally concentrated under reduced pressure. The crude product was purified by silica gel column chromatography to give 6-(4-bromophenyl)-1-methyl-1H-indazole (600 mg).

MS (ESI) M/Z: 287.0 [M+H] ⁺ .

Step B: 6-(4-bromophenyl)-1-methyl-1H-indazole (200 mg, 0.70 mmol) and 4-isoquinolineboronic acid (181.5 mg, 1.05 mmol) were dissolved in dioxane (10 ml) and water (2.5 ml), and potassium carbonate (193 mg, 1.4 mmol) and 1,1'-bis(diphenylphosphinoferrocene)palladium dichloride (51.26 mg, 0.07 mmol) were added. The reaction liquid was replaced with nitrogen three times and reacted at 105 degrees Celsius for 3 hours.

After TLC monitoring of the disappearance of the starting material, water (20 ml) was added, and the mixture was extracted with ethyl acetate (30 ml × 2 times). The organic phases were combined, washed with saturated brine (50 ml), then dried over anhydrous sodium sulfate, filtered, and finally concentrated under reduced pressure. The crude product was purified by silica gel column chromatography to give 4-(4-(1-methyl-1H-indazol-6-yl)phenyl)isoquinoline (120.68 mg).

MS (ESI) M/Z: 336.2 [M + H] ⁺ .

¹ H NMR (400 MHz, DMSO-d ₆ ) δ 9.39 (s, 1H), 8.53 (s, 1H), 8.26 (d, J=8.1 Hz, 1H), 8.13–8.04 (m, 2H), 8.04–7.98 (m, 2H), 7.98–7.93 (m, 1H), 7.91–7.81 (m, 2H), 7.81–7.74 (m, 1H), 7.74–7.67 (m, 2H), 7.61–7.51 (m, 1H), 4.14 (s, 3H).

Embodiment 19:

2-(4-(4-(4,5-dimethylpyridin-3-yl)phenyl)-1H-pyrazol-1-yl)-N,N-dimethylacetamide

Referring to the synthesis method of Example 8, 2-(4-(4-(4,5-dimethylpyridin-3-yl)phenyl)-1H-pyrazol-1-yl)-N,N-dimethylacetamide (8.91 mg) was obtained.

MS (ESI) M/Z: 335.1 [M+H] ⁺ .

¹ H NMR (400 MHz, DMSO-d ₆ ) δ 8.32 (s, 1H), 8.24 (s, 1H), 8.14 (s, 1H), 7.95 (s, 1H), 7.71–7.65 (m, 2H), 7.38–7.31 (m, 2H), 5.15 (s, 2H), 3.06 (s, 3H), 2.87 (s, 3H), 2.29 (s, 3H), 2.19 (s, 3H).

Embodiment 20:

(R)-1-(4-(Isoquinolin-4-yl)phenyl)-N,N-dimethylpyrrolidine-3-carboxamide

Step A: (R)-1-(tert-Butyloxycarbonyl)pyrrolidine-3-carboxylic acid (2 g, 9.3 mmol) and dimethylamine hydrochloride (1.52 g, 18.6 mmol) were dissolved in DCM/DMF (4:1, 20 ml) at room temperature, HATU (7.07 g, 18.6 mmol) and DIEA (4.8 g, 37.2 mmol) were added, and the mixture was stirred at room temperature for 6 hours.

After TLC/LCMS monitoring showed that the starting material disappeared, citric acid aqueous solution (40 ml) was added, and dichloromethane (50 ml) was used for extraction twice, and the organic phases were combined, washed with saturated brine, dried and concentrated. The crude product was mixed and passed through a column to obtain (R)-3-(dimethylcarbamoyl)pyrrolidine-1-carboxylic acid tert-butyl ester (2.14 g).

MS (ESI) M/Z: 243.2 [M+H] ⁺ .

Step B: Dissolve (R)-tert-butyl 3-(dimethylcarbamoyl)pyrrolidine-1-carboxylate (2.14 g, 8.84 mmol) in ethyl acetate (10 mL), add 6N HCl/EtOAc (10 mL) dropwise in an ice-water bath, and stir at room temperature for 1 hour.

After LCMS monitoring showed the disappearance of the starting material, the reaction mixture was concentrated to give (R)-3-(dimethylcarbamoyl)pyrrolidine hydrochloride (94 mg).

Step C: (R)-3-(Dimethylcarbamoyl)pyrrolidine hydrochloride (815 mg, 4.71 mmol), 1-bromo-4-iodobenzene (2 g, 7.07 mmol), sodium carbonate (1 g, 9.42 mmol), Pd(dppf)Cl ₂ (345 mg, 0.471 mmol) were added to 1,4-dioxane (20 ml) and water (5 ml), the atmosphere was replaced with nitrogen three times, and the reaction was carried out at 90°C overnight.

After TLC monitoring showed the disappearance of the starting material, the mixture was cooled, water (20 ml) was added, and extraction was performed with dichloromethane (20 ml x 2 times). The organic phases were combined, washed with saturated brine (20 ml), then dried over anhydrous sodium sulfate, filtered, and finally concentrated under reduced pressure. The residue was purified by silica gel column chromatography to give (R)-1-(4-bromophenyl)-N,N-dimethylpyrrolidine-3-carboxamide (530 mg).

MS (ESI) M/Z: 297.3 [M + H] ⁺ .

Step D: Isoquinoline-4-boronic acid (100 mg, 0.35 mmol), (S)-1-(4-bromophenyl)-N,N-dimethylpyrrolidine-3-carboxamide (94 mg, 0.525 mmol), cesium carbonate (514 mg, 1.575 mmol), [1,1'-bis(diphenylphosphino)ferrocene]palladium dichloride (32 mg, 0.035 mmol), 4,5-bis(diphenylphosphino)ferrocene-9,9-dimethylxanthene (40 mg, 0.07 mmol) were added to 1,4-dioxane (2 ml) and water (0.5 ml), the atmosphere was replaced with nitrogen three times, and the reaction was carried out at 80 °C for 3 hours.

After TLC monitoring showed that the raw material disappeared, the mixture was cooled and water (20 ml) was added. The mixed solution was extracted with ethyl acetate (20 ml × 2 times), and the organic phases were combined, washed with saturated brine (20 ml), then dried over anhydrous sodium sulfate, filtered, and finally concentrated under reduced pressure. The obtained residue was prepared to obtain (R)-1-(4-(isoquinolin-4-yl)phenyl)-N,N-dimethylpyrrolidine-3-carboxamide (6.63 mg).

MS (ESI) M/Z: 346.3 [M + H] ⁺ .

¹ H NMR (400 MHz, DMSO-d ₆ ) δ 9.26 (s, 1H), 8.38 (s, 1H), 8.19 (d, J=8.0 Hz, 1H), 7.96 (d, J=8.4 Hz, 1H), 7.81–7.75 (m, 1H), 7.74–7.68 (m, 1H), 7.42–7.32 (m, 2H), 6.78–6.68 (m, 2H), 3.60–3.52 (m, 2H), 3.44–3.39 (m, 3H), 3.10 (s, 3H), 2.87 (s, 3H), 2.28–2.18 (m, 1H), 2.17–2.07 (m, 1H).

Embodiment 21:

(S)-1-(4-(Isoquinolin-4-yl)phenyl)-N,N-dimethylpyrrolidine-3-carboxamide

Referring to the synthesis method of Example 20, (S)-1-(4-(isoquinolin-4-yl)phenyl)-N,N-dimethylpyrrolidine-3-carboxamide (15.08 mg) was obtained.

MS (ESI) M/Z: 346.0 [M+H] ⁺ .

¹ H NMR (400 MHz, DMSO-d ₆ ) δ 9.27 (s, 1H), 8.39 (s, 1H), 8.19 (d, J=8.0 Hz, 1H), 7.96 (d, J=8.4 Hz, 1H), 7.82–7.75 (m, 1H), 7.75–7.67 (m, 1H), 7.40–7.33 (m, 2H), 6.77–6.68 (m, 2H), 3.59–3.54 (m, 2H), 3.44–3.40 (m, 3H), 3.10 (s, 3H), 2.87 (s, 3H), 2.25–2.18 (m, 1H), 2.16–2.08 (m, 1H).

Embodiment 22:

2-(4-(4-(7-fluoroisoquinolin-4-yl)phenyl)-1H-pyrazol-1-yl)-1-morpholinoethan-1-one

Steps:

Step A: 4-(4,4,5,5-tetramethyl-1,3,2-dioxaborolan-2-yl)-1H-pyrazole (5 g, 0.026 mol), tert-butyl 2-bromoacetate (5.82 g, 0.03 mol) and cesium carbonate (21.18 g, 0.065 mol) were added to acetonitrile (50 ml) and reacted at 80 °C for 3 hours.

After TLC monitoring showed that the starting material disappeared, the mixture was cooled and filtered, and the filtrate was concentrated under reduced pressure to dryness to obtain crude tert-butyl 2-(4-(4,4,5,5-tetramethyl-1,3,2-dioxaborolan-2-yl)-1H-pyrazol-1-yl)acetate (4.5 g).

MS (ESI) M/Z: 308.9 [M + H] ⁺ .

Step B: Add tert-butyl 2-(4-(4,4,5,5-tetramethyl-1,3,2-dioxaborolan-2-yl)-1H-pyrazol-1-yl)acetate (4.5 g, 25.77 mmol), 1-bromo-4-iodobenzene (170 mg, 38.65 mmol), [1,1'-bis(diphenylphosphino)ferrocene]palladium dichloride (1.8 g, 2.58 mmol) and sodium carbonate (5.5 g, 51.54 mmol) to dioxane (140 ml) and water (35 ml), replace with nitrogen three times and stir at 90 °C overnight.

After LCMS monitoring showed the disappearance of the starting material, water (200 mL) was added, extracted with ethyl acetate (200 mL×2), and the organic phases were combined. The organic phases were first washed with saturated brine, then dried over anhydrous sodium sulfate, filtered, and finally concentrated under reduced pressure. The resulting residue was purified by silica gel column chromatography to give tert-butyl 2-(4-(4-bromophenyl)-1H-pyrazol-1-yl)acetate (5.5 g).

MS (ESI) M/Z: 337.1 [M+H] ⁺ .

Step C: Add tert-butyl 2-(4-(4-bromophenyl)-1H-pyrazol-1-yl)acetate (5.5 g, 16.2 mmol), bis(boron) (4.93 g, 19.4 mmol), potassium acetate (4.77 g, 48.6 mmol) and [1,1'-bis(diphenylphosphino)ferrocene]palladium dichloride (1.17 g, 1.6 mmol) to dioxane (100 ml), replace the atmosphere with nitrogen three times and stir at 100 °C overnight.

After LCMS monitoring showed the disappearance of the starting material, the mixture was filtered through celite, water (200 ml) was added to the filtrate, and the mixture was extracted with ethyl acetate (200 ml×2). The organic phases were combined, washed with saturated brine, dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure. The residue was purified by silica gel column chromatography to give tert-butyl 2-(4-(4-(4,4,5,5-tetramethyl-1,3,2-dioxaborolan-2-yl)phenyl)-1H-pyrazol-1-yl)acetate (5 g).

MS (ESI) M/Z: 385.2 [M + H] ⁺ .

Step D: 7-Fluoroisoquinoline (1 g, 6.8 mmol) was dissolved in acetic acid (30 ml), and N-bromosuccinimide (1.45 g, 8.2 mmol) was added in portions and stirred at 110 °C for 4 hours.

After LCMS monitoring showed that the starting material disappeared, the mixture was concentrated under reduced pressure and the resulting residue was purified by silica gel column chromatography to give 4-bromo-7-fluoroisoquinoline (450 mg).

MS (ESI) M/Z: 226.1 [M+H] ⁺ .

Step E: 4-bromo-7-fluoroisoquinoline (450 mg, 1.938 mmol), tert-butyl 2-(4-(4-(4,4,5,5-tetramethyl-1,3,2-dioxaborolan-2-yl)phenyl)-1H-pyrazol-1-yl)acetate (967.85 mg, 2.519 mmol), potassium carbonate (534.89 mg, 3.876 mmol) and [1,1'-bis(diphenylphosphino)ferrocene]palladium dichloride (141.81 mg, 0.194 mmol) were added to dioxane (20 ml) and water (5 ml), and the atmosphere was replaced with nitrogen three times, and the reaction was stirred at 110 °C for 3 hours.

After LCMS monitoring showed the disappearance of the starting material, the mixture was concentrated under reduced pressure and the resulting residue was purified by silica gel column chromatography to give tert-butyl 2-(4-(4-(7-fluoroisoquinolin-4-yl)phenyl)-1H-pyrazol-1-yl)acetate (600 mg).

MS (ESI) M/Z: 404.0 [M+H] ⁺ .

Step F: tert-Butyl 2-(4-(4-(7-fluoroisoquinolin-4-yl)phenyl)-1H-pyrazol-1-yl)acetate (600 mg, 1.488 mmol) was added to trifluoroacetic acid (5 ml) and dichloromethane (5 ml) and stirred at room temperature for 2 hours.

After LCMS monitoring showed the disappearance of the starting material, the reaction solution was concentrated under reduced pressure to dryness to give a crude product of 2-(4-(4-(7-fluoroisoquinolin-4-yl)phenyl)-1H-pyrazol-1-yl)acetic acid trifluoroacetate (800 mg).

MS (ESI) M/Z: 348.2 [M+H] ⁺ .

Step G: 2-(4-(4-(7-Fluoroisoquinolin-4-yl)phenyl)-1H-pyrazol-1-yl)acetic acid (100 mg, 0.288 mmol), N,N-diisopropylethylamine (185.76 mg, 1.44 mmol), morpholine (100.3 mg, 1.152 mmol) and 2-(7-azabenzotriazole)-N,N,N',N'-tetramethyluronium hexafluorophosphate (218.3 mg, 0.576 mmol) were added to N,N-dimethylformamide (2 ml) and stirred at room temperature for 1 hour.

After LCMS monitoring showed the disappearance of the starting material, water (10 ml) was added, extracted with ethyl acetate (20 ml×2), and the organic phases were combined, washed with saturated brine, then dried over anhydrous sodium sulfate, filtered, and finally concentrated under reduced pressure to obtain the residue to give 2-(4-(4-(7-fluoroisoquinolin-4-yl)phenyl)-1H-pyrazol-1-yl)-1-morpholinoethan-1-one (5.47 mg).

MS (ESI) M/Z: 416.5 [M+H] ⁺ .

¹ H NMR (400 MHz, DMSO-d ₆ ) δ 9.34 (s, 1H), 8.48 (s, 1H), 8.21 (s, 1H), 8.10–7.95 (m, 3H), 7.82–7.71 (m, 3H), 7.59–7.51 (m, 2H), 5.21 (s, 2H), 3.66–3.57 (m, 4H), 3.56–3.46 (m, 4H).

Embodiment 23:

2-(4-(4-(7-fluoroisoquinolin-4-yl)phenyl)-1H-pyrazol-1-yl)-1-(4-methylpiperazin-1-)ethan-1-one

Referring to the synthesis method of Example 22, 2-(4-(4-(7-fluoroisoquinolin-4-yl)phenyl)-1H-pyrazol-1-yl)-1-(4-methylpiperazin-1-)ethan-1-one (29.8 mg) was obtained.

MS (ESI) M/Z: 430.2 [M + H] ⁺ .

¹ H NMR (400 MHz, DMSO-d ₆ ) δ 9.34 (s, 1H), 8.48 (s, 1H), 8.21 (s, 1H), 8.10–7.93 (m, 3H), 7.82–7.68 (m, 3H), 7.55 (d, J=7.9 Hz, 2H), 5.20 (s, 2H), 3.58–3.43 (m, 4H), 2.44–2.29 (m, 4H), 2.23 (s, 3H).

Embodiment 24:

2-(4-(4-(7-fluoroisoquinolin-4-yl)phenyl)-1H-pyrazol-1-yl)-1-(piperidin-1-yl)ethan-1-one

Referring to the synthesis method of Example 22, 2-(4-(4-(7-fluoroisoquinolin-4-yl)phenyl)-1H-pyrazol-1-yl)-(1-piperidin-1-yl)ethan-1-one (19.89 mg) was obtained.

MS (ESI) M/Z: 415.2 [M + H] ⁺ .

¹ H NMR (400 MHz, DMSO-d ₆ ) δ 9.34 (s, 1H), 8.48 (s, 1H), 8.21 (s, 1H), 8.09–8.03 (m, 1H), 8.02–7.95 (m, 2H), 7.82–7.69 (m, 3H), 7.55 (d, J=7.9 Hz, 2H), 5.17 (s, 2H), 3.52–3.42 (m, 4H), 1.65–1.52 (m, 4H), 1.50–1.44 (m, 2H).

Embodiment 25:

2-(4-(4-(7-fluoroisoquinolin-4-yl)phenyl)-1H-pyrazol-1-yl)-1-(piperazin-1-yl)ethan-1-one

Steps:

Step A: 2-(4-(4-(7-fluoroisoquinolin-4-yl)phenyl)-1H-pyrazol-1-yl)acetic acid (100 mg, 0.288 mmol), tert-butyl piperazine-1-carboxylate (214 mg, 1.152 mmol), 2-(7-azabenzotriazole)-N,N,N',N'-tetramethyluronium hexafluorophosphate (218 mg, 0.576 mmol) and N,N-diisopropylethylamine (185 mg, 1.44 mmol) were added to N,N-dimethylformamide (2 ml) and reacted at room temperature for 1 hour.

After TLC monitoring showed that the starting material disappeared, sodium bicarbonate solution (20 ml) and ethyl acetate (30 ml x 2 times) were added for extraction. The organic phase was saturated with brine (20 ml), dried over anhydrous sodium sulfate and concentrated under reduced pressure. The resulting residue was purified by silica gel column chromatography to give tert-butyl 4-(2-(4-(7-fluoroisoquinolin-4-yl)phenyl)-1H-pyrazol-1-yl)acetyl)piperazine-1-carboxylate (70 mg).

MS (ESI) M/Z: 516.2 [M + H] ⁺ .

Step B: tert-Butyl 4-(2-(4-(7-fluoroisoquinolin-4-yl)phenyl)-1H-pyrazol-1-yl)acetyl)piperazine-1-carboxylate (70 mg, 0.136 mmol) was added to trifluoroacetic acid (1 ml) and dichloromethane (1 ml) and reacted at room temperature for 1 hour.

After LCMS monitoring showed the disappearance of the starting material, the mixture was concentrated under reduced pressure, and sodium bicarbonate solution (20 mL) was added, followed by extraction with ethyl acetate (30 mL × 2 times). The organic phases were combined, washed with saturated brine (20 mL), then dried over anhydrous sodium sulfate, filtered, and finally concentrated under reduced pressure to give 2-(4-(4-(7-fluoroisoquinolin-4-yl)phenyl)-1H-pyrazol-1-yl)-1-piperazin-1-ylethane-1-one (11.64 mg) as a crude product.

MS (ESI) M/Z: 416.2 [M+H] ⁺ .

¹ H NMR (400 MHz, DMSO-d ₆ ) δ 9.34 (s, 1H), 8.48 (s, 1H), 8.21 (s, 1H), 8.08–8.04 (m, 1H), 8.02–7.97 (m, 2H), 7.82–7.77 (m, 2H), 7.76–7.70 (m, 1H), 7.58–7.51 (m, 2H), 5.18 (s, 2H), 3.48–3.39 (m, 4H), 2.80–2.64 (m, 4H).

Embodiment 26:

2-(4-(4-(7-fluoroisoquinolin-4-yl)phenyl)-1H-pyrazol-1-yl)-1-(tetrahydro-1H-furan[3,4-c]pyrrol-5(3H)-yl)ethan-1-one

Referring to the synthesis method of Example 22, 2-(4-(4-(7-fluoroisoquinolin-4-yl)phenyl)-1H-pyrazol-1-yl)-1-(tetrahydro-1H-furan[3,4-c]pyrrol-5(3H)-yl)ethan-1-one (47.42 mg) was obtained.

MS (ESI) M/Z: 443.2 [M+H] ⁺ .

¹ H NMR (400 MHz, DMSO-d ₆ ) δ 9.34 (s, 1H), 8.48 (s, 1H), 8.21 (s, 1H), 8.09–8.04 (m, 1H), 8.02–7.95 (m, 2H), 7.84–7.77 (m, 2H), 7.76–7.70 (m, 1H), 7.63–7.47 (m, 2H), 5.19–5.01 (m, 2H), 3.84–3.72 (m, 3H), 3.67–3.44 (m, 5H), 3.07–2.98 (m, 1H), 2.96–2.88 (m, 1H).

Embodiment 27:

2-(4-(4-(7-fluoroisoquinolin-4-yl)phenyl)-1H-pyrazol-1-yl)-1-pyrrolidin-1-ylethan-1-one

Referring to the synthesis method of Example 22, 2-(4-(4-(7-fluoroisoquinolin-4-yl)phenyl)-1H-pyrazol-1-yl)-1-pyrrolidin-1-ylethan-1-one (16.39 mg) was obtained.

MS (ESI) M/Z: 401.1 [M+H] ⁺ .

¹ H NMR (400 MHz, DMSO-d ₆ ) δ 9.34 (s, 1H), 8.48 (s, 1H), 8.22 (s, 1H), 8.10–8.03 (m, 1H), 8.03–7.96 (m, 2H), 7.83–7.77 (m, 2H), 7.77–7.70 (m, 1H), 7.60–7.50 (m, 2H), 5.09 (s, 2H), 3.58–3.48 (m, 2H), 3.37–3.35 (m, 2H), 1.98–1.88 (m, 2H), 1.85–1.75 (m, 2H).

Embodiment 28:

2-(4-(4-(7-chloroisoquinolin-4-yl)phenyl)-1H-pyrazol-1-yl)-N,N-dimethylacetamide

Referring to the synthesis method of Example 8, 2-(4-(4-(7-chloroisoquinolin-4-yl)phenyl)-1H-pyrazol-1-yl)-N,N-dimethylacetamide (130 mg) was obtained.

MS (ESI) M/Z: 391.2 [M+H] ⁺ .

¹ H NMR (400 MHz, DMSO-d ₆ ) δ 9.34 (s, 1H), 8.52 (s, 1H), 8.39 (d, J=2.2 Hz, 1H), 8.19 (s, 1H), 8.00 (s, 1H), 7.97–7.91 (m, 1H), 7.84–7.80 (m, 1H), 7.80–7.77 (m, 2H), 7.58–7.52 (m, 2H), 5.17 (s, 2H), 3.05 (s, 3H), 2.88 (s, 3H).

Embodiment 29:

N,N-Dimethyl-2-(4-(4-(7-methylisoquinolin-4-yl)phenyl)-1H-pyrazol-1-yl)acetamide

Steps:

Step A: 2-(4-(4-(7-chloroisoquinolin-4-yl)phenyl)-1H-pyrazol-1-yl)-N,N-dimethylacetamide (120 mg, 0.30 mmol), methylboric acid (37 mg, 0.61 mmol), potassium phosphate (163 mg, 0.77 mmol), palladium acetate (14 mg, 0.06 mmol) and 2-dicyclohexylphosphino-2',6'-dimethoxybiphenyl (25 mg, 0.06 mmol) were added to tetrahydrofuran (2 ml) and toluene (2 ml), replaced with nitrogen three times, and reacted at 90 °C overnight.

After TLC monitoring showed that the raw material disappeared, the mixture was cooled and water (20 ml) was added. The mixed solution was extracted with ethyl acetate (20 ml × 2 times), and the organic phases were combined, washed with saturated brine (20 ml), then dried over anhydrous sodium sulfate, filtered, and finally concentrated under reduced pressure. The resulting residue was purified by silica gel column chromatography to obtain N,N-dimethyl-2-(4-(4-(7-methylisoquinolin-4-yl)phenyl)-1H-pyrazol-1-yl)acetamide (60.85 mg).

MS (ESI) M/Z: 371.5 [M + H] ⁺ .

¹ H NMR (400 MHz, DMSO-d ₆ ) δ 9.24 (s, 1H), 8.41 (s, 1H), 8.21–8.15 (m, 1H), 8.02–7.96 (m, 2H), 7.84 (d, J=8.6 Hz, 1H), 7.81–7.74 (m, 2H), 7.70–7.61 (m, 1H), 7.57–7.49 (m, 2H), 5.17 (s, 2H), 3.07 (s, 3H), 2.88 (s, 3H), 2.54 (s, 3H).

Embodiment 30:

2-(4-(4-(6-fluoroisoquinolin-4-yl)phenyl)-1H-pyrazol-1-yl)-N,N-dimethylacetamide

Referring to the synthesis method of Example 8, 2-(4-(4-(6-fluoroisoquinolin-4-yl)phenyl)-1H-pyrazol-1-yl)-N,N-dimethylacetamide (102.05 mg).

MS (ESI) M/Z: 375.2 [M + H] ⁺ .

¹ H NMR (400 MHz, DMSO-d ₆ ) δ 9.39 (s, 1H), 8.52 (s, 1H), 8.39 (dd, J=9.1, 5.8 Hz, 1H), 8.20 (s, 1H), 8.00 (s, 1H), 7.83–7.76 (m, 2H), 7.73–7.64 (m, 1H), 7.60–7.50 (m, 3H), 5.18 (s, 2H), 3.07 (s, 3H), 2.89 (s, 3H).

Embodiment 31:

2-(4-(4-(5-fluoroisoquinolin-4-yl)phenyl)-1H-pyrazol-1-yl)-N,N-dimethylacetamide

By referring to the synthesis method of Example 8, 2-(4-(4-(5-fluoroisoquinolin-4-yl)phenyl)-1H-pyrazol-1-yl)-N,N-dimethylacetamide (139.59 mg) was obtained.

MS (ESI) M/Z: 375.3 [M + H] ⁺ .

¹ H NMR (400 MHz, DMSO-d ₆ ) δ 9.42 (d, J=2.4 Hz, 1H), 8.40 (s, 1H), 8.16 (s, 1H), 8.13–8.08 (m, 1H), 7.97 (s, 1H), 7.79–7.71 (m, 1H), 7.70–7.64 (m, 2H), 7.63–7.55 (m, 1H), 7.50–7.43 (m, 2H), 5.16 (s, 2H), 3.07 (s, 3H), 2.88 (s, 3H).

Embodiment 32:

2-(4-(4-(7-fluoro-6-(5-methyl-1,3,4-oxadiazol-2-yl)isoquinolin-4-yl)phenyl)-1H-pyrazol-1-yl)-N,N-dimethylacetamide

Steps:

Step A: 6-bromo-7-fluoroisoquinoline (2.0 g, 8.85 mmol), bis(triphenylphosphine)palladium dichloride (310 mg, 0.44 mmol) and triethylamine (895 mg, 8.85 mmol) were added to methanol (300 ml), the mixture was added to an autoclave, carbon monoxide (2 MPa) was introduced, and the temperature was raised to 80 °C for reaction overnight.

After TLC monitoring showed that the starting material disappeared, the mixture was cooled and concentrated under reduced pressure. The resulting crude product was subjected to column chromatography to give methyl 7-fluoroisoquinoline-6-carboxylate (1.5 g).

MS (ESI) M/Z: 206.0 [M+H] ⁺ .

Step B: Methyl 7-fluoroisoquinoline-6-carboxylate (500 mg, 2.44 mmol) and 85% hydrazine hydrate (1.5 ml) were added to ethanol (20 ml), and the reaction solution was stirred at 80 °C overnight.

After LCMS monitoring showed that the starting material disappeared, the mixture was concentrated under reduced pressure and the resulting residue was purified by silica gel column chromatography to give 7-fluoroisoquinoline-6-hydrazide (370 mg).

MS (ESI) M/Z: 206.2 [M+H] ⁺ .

Step C: 7-Fluoroisoquinoline-6-hydrazide (370 mg, 1.8 mmol) and acetic acid (818 mg, 13.6 mmol) were added to phosphorus oxychloride (7 ml), and the reaction solution was stirred at 120 °C for 2 hours.

After LCMS monitoring showed that the raw material disappeared, the mixture was concentrated under reduced pressure. Saturated sodium bicarbonate aqueous solution (20 ml) and ethyl acetate (20 ml) were added to the residue, stirred, and separated. The organic phase was washed with saturated brine (10 ml), dried, and concentrated. The resulting residue was purified by silica gel column chromatography to obtain 2-(7-fluoroisoquinolin-6-yl)-5-methyl-1,3,4-oxadiazole (68 mg).

MS (ESI) M/Z: 230.2 [M + H] ⁺ .

Step D: 2-(7-Fluoroisoquinolin-6-yl)-5-methyl-1,3,4-oxadiazole (68 mg, 0.3 mmol) and N-bromosuccinimide (64 mg, 0.36 mmol) were added to acetic acid (1 ml), and the reaction solution was stirred at 110 °C for 3 hours.

After LCMS monitoring showed that the raw material disappeared, the mixture was cooled, saturated sodium bicarbonate aqueous solution (20 ml) and ethyl acetate (20 ml) were added, stirred, and separated. The organic phase was washed with saturated brine (10 ml), dried, and concentrated. The resulting residue was purified by silica gel column chromatography to obtain 2-(4-bromo-7-fluoroisoquinolin-6-yl)-5-methyl-1,3,4-oxadiazole (37 mg).

MS (ESI) M/Z: 308.08 [M+H] ⁺ .

Step E: 2-(4-bromo-7-fluoroisoquinolin-6-yl)-5-methyl-1,3,4-oxadiazole (37 mg, 0.12 mmol), N,N-dimethyl-2-(4-(4-(4,4,5,5-tetramethyl-1,3,2-dioxaborolan-2-yl)phenyl)-1H-pyrazol-1-yl)acetamide (47 mg, 0.13 mmol), potassium carbonate (41.4 mg, 0.3 mmol) and [1,1'-bis(diphenylphosphino)ferrocene]palladium dichloride (8.8 mg, 0.012 mmol) were added to 1,4-dioxane (4 ml) and water (1 ml), the atmosphere was replaced with nitrogen three times, and the reaction was heated under reflux for 3 hours.

After TLC monitoring showed that the raw material disappeared, the mixture was cooled. Water (10 ml) and ethyl acetate (20 ml) were added and stirred to separate the liquids. The organic phase was washed with saturated brine (10 ml), dried over anhydrous sodium sulfate, and concentrated under reduced pressure. The obtained residue was prepared and purified to obtain 2-(4-(4-(7-fluoro-6-(5-methyl-1,3,4-oxadiazol-2-yl)isoquinolin-4-yl)phenyl)-1H-pyrazol-1-yl)-N,N-dimethylacetamide (30.43 mg).

MS (ESI) M/Z: 457.3 [M + H] ⁺ .

¹ H NMR (400 MHz, DMSO-d ₆ ) δ 9.44 (s, 1H), 8.59 (s, 1H), 8.55 (d, J=6.7 Hz, 1H), 8.35 (d, J=11.0 Hz, 1H), 8.22 (s, 1H), 8.03 (s, 1H), 7.86–7.81 (m, 2H), 7.64–7.58 (m, 2H), 5.17 (s, 2H), 3.07 (s, 3H), 2.88 (s, 3H), 2.60 (s, 3H).

Embodiment 33:

2-(4-(4-(isoquinolin-4-yl)piperidin-1-yl)-1H-pyrazol-1-yl)-N,N-dimethylacetamide

Steps:

Step A: 4-bromoisoquinoline (1.0 g, 4.8 mmol), 4-(4,4,5,5-tetramethyl-1,3,2-dioxaboroborolan-2-yl)-3,6-dihydropyridine-1(2H)-carboxylic acid benzyl ester (1.98 g, 5.8 mmol) were added to dioxane/water (4/1, 10 ml), sodium carbonate (1.02 g, 9.6 mmol) and [1,1'-bis(diphenylphosphino)ferrocene] dichloropalladium dichloromethane complex (0.35 g, 0.5 mmol), the atmosphere was replaced with nitrogen three times, and the temperature was raised to 110 °C for reaction for 3 hours.

After TLC monitoring showed that the raw material disappeared, the mixture was cooled. Water (20 ml) and ethyl acetate (30 ml x 2) were added for extraction, and the organic phases were combined. The organic phase was washed with saturated brine (20 ml), dried over anhydrous sodium sulfate, and concentrated under reduced pressure. The obtained residue was purified by silica gel column chromatography to obtain 4-(isoquinolin-4-yl)-3,6-dihydropyridine-1(2H)-carboxylic acid benzyl ester (1.5 g).

MS (ESI) M/Z: 344.7 [M + H] ⁺ .

Step B: Benzyl 4-(isoquinolin-4-yl)-3,6-dihydropyridine-1(2H)-carboxylate (1.5 g, 4.35 mmol) was added to methanol (10 ml) and tetrahydrofuran (20 ml), palladium on carbon (400 mg) was added, and hydrogenation was carried out at 40 °C overnight.

After LCMS monitoring showed the disappearance of the starting material, the mixture was filtered through silica gel and the filtrate was concentrated under reduced pressure to give crude 4-(piperidin-4-yl)isoquinoline (1.18 g).

MS (ESI) M/Z: 213.3 [M + H] ⁺ .

Step C: 4-(Piperidin-4-yl)isoquinoline (300 mg, 1.413 mmol) and 2-(4-iodo-1H-pyrazol-1-yl)-N,N-dimethylacetamide (473 mg, 1.696 mmol) were added to dimethyl sulfoxide (10 ml), and L-proline (65 mg, 0.565 mmol), cuprous iodide (53 mg, 0.283 mmol) and potassium carbonate (584.98 mg, 4.239 mmol) were added. The atmosphere was replaced with nitrogen three times and the reaction was carried out at 90°C overnight.

After LCMS monitoring showed that the raw material disappeared, the mixture was cooled. Ammonia water (5 ml), water (20 ml) and ethyl acetate (30 ml x 2) were added for extraction, and the organic phases were combined. The organic phase was washed with saturated brine (20 ml), dried over anhydrous sodium sulfate, and concentrated under reduced pressure. The resulting residue was used to prepare 2-(4-(4-(isoquinolin-4-yl)piperidin-1-yl)-1H-pyrazol-1-yl)-N,N-dimethylacetamide (1.49 mg).

MS (ESI) M/Z: 364.1 [M+H] ⁺ .

¹ H NMR (400 MHz, DMSO-d ₆ ) δ 9.21 (s, 1H), 8.47 (s, 1H), 8.28–8.20 (m, 1H), 8.18–8.11 (m, 1H), 7.89-7.81 (m, 1H), 7.75–7.66 (m, 1H), 7.23 (s, 2H), 4.97 (s, 2H), 3.52–3.44 (m, 4H), 3.01 (s, 3H), 2.85 (s, 3H), 2.76-2.72 (m, 1H), 2.08–1.92 (m, 4H). C ₂₁ H ₂₅ N ₅ O.

Embodiment 34:

2-(4-(4-(8-fluoro-6-(5-methyl-1,3,4-oxadiazol-2-yl)isoquinolin-4-yl)phenyl)-1H-pyrazol-1-yl)-N,N-dimethylacetamide

Steps:

Step A: In a 250 ml autoclave, 6-bromo-8-fluoroisoquinoline (1 g, 4.4 mmol) was dissolved in methanol (180 ml), and triethylamine (445 mg, 4.4 mmol) and bistriphenylphosphine palladium dichloride (155 mg, 0.22 mmol) were added. Carbon monoxide was introduced into the autoclave and the reaction was carried out overnight at 80 degrees Celsius and 2 MPa pressure.

After the disappearance of the starting material as monitored by TLC, the residue was concentrated under reduced pressure and the crude product was purified by silica gel column chromatography to give methyl 8-fluoroisoquinoline-6-carboxylate (840 mg).

MS (ESI) M/Z: 206.3 [M + H] ⁺ .

Step B: Methyl 8-fluoroisoquinoline-6-carboxylate (840 mg, 4.10 mmol) was dissolved in anhydrous ethanol (30 ml), hydrazine hydrate (85%, 2.5 ml) was added, and the mixture was refluxed at 80°C overnight.

After the disappearance of the starting material as monitored by TLC, the mixture was concentrated under reduced pressure and the crude product was purified by silica gel column chromatography to give 8-fluoroisoquinoline-6-carbohydrazide (830 mg).

MS (ESI) M/Z: 206.0 [M+H] ⁺ .

Step C: 8-Fluoroisoquinoline-6-carbohydrazide (480 mg, 2.34 mmol) was dissolved in DMF (10 ml), acetic acid (210.5 mg, 3.51 mmol) and N,N-diisopropylethylamine (907.4 mg, 7.02 mmol) were added, and HATU (1.3 g, 3.51 mmol) was added under ice bath, and the reaction was carried out at room temperature for 2 hours.

After the disappearance of the raw material by TLC monitoring, sodium bicarbonate aqueous solution (20 ml) was added, extracted with ethyl acetate and a small amount of methanol (20 ml x 5), the organic phases were combined, washed with saturated brine, then dried over anhydrous sodium sulfate, filtered, and finally concentrated under reduced pressure. The crude product was purified by silica gel column chromatography to obtain N'-acetyl-8-fluoroisoquinoline-6-carbohydrazide (280 mg).

MS (ESI) M/Z: 247.9 [M + H] ⁺ .

Step D: N'-acetyl-8-fluoroisoquinoline-6-carbohydrazide (280 mg, 1.13 mmol) was dissolved in phosphorus oxychloride (5 ml) and reacted in a microwave oven at 100 °C overnight.

After the disappearance of the starting material monitored by TLC, the mixture was concentrated under reduced pressure, and an aqueous sodium bicarbonate solution was added for neutralization. The pH was adjusted to weak alkaline, and ethyl acetate (20 ml × 2) was added for extraction. The organic phase was first washed with saturated brine, then dried over anhydrous sodium sulfate, filtered, and finally concentrated under reduced pressure to obtain a crude product of 2-(8-fluoroisoquinolin-6-yl)-5-methyl-1,3,4-oxadiazole (140 mg).

MS (ESI) M/Z: 230.0 [M+H] ⁺ .

Step E: 2-(8-Fluoroisoquinolin-6-yl)-5-methyl-1,3,4-oxadiazole (140 mg, 0.61 mmol) was dissolved in acetic acid (3 ml), heated to 110 °C, N-bromosuccinimide (130.5 mg, 0.73 mmol) was added in portions, and the mixture was reacted at 110 °C for 3 hours.

After the starting material disappeared as monitored by TLC, the mixture was concentrated under reduced pressure and the crude product was purified by silica gel column chromatography to give 2-(4-bromo-8-fluoroisoquinolin-6-yl)-5-methyl-1,3,4-oxadiazole (170 mg).

MS (ESI) M/Z: 308.0 [M+H] ⁺ .

Step F: 2-(4-Bromo-8-fluoroisoquinolin-6-yl)-5-methyl-1,3,4-oxadiazole (170 mg, 0.55 mmol), N,N-dimethyl-2-(4-(4-(4,4,5,5-tetramethyl-1,3,2-dioxaborolan-2-yl)phenyl)-1H-pyrazol-1-yl)acetamide (215.5 mg, 0.61 mmol), anhydrous potassium carbonate (190 mg, 1.38 mmol) were dissolved in dioxane (4 ml) and water (1 ml), 1,1'-bis(di-phenylphosphino)ferrocenepalladium chloride (40.3 mg, 0.055 mmol) was added, the reaction liquid was replaced with nitrogen three times, and the reaction was carried out at 90°C overnight.

After the disappearance of the starting material as monitored by TLC, the mixture was concentrated under reduced pressure and the crude product was purified by silica gel column chromatography and then purified by preparative purification to give 2-(4-(4-(8-fluoro-6-(5-methyl-1,3,4-oxadiazol-2-yl)isoquinolin-4-yl)phenyl)-1H-pyrazol-1-yl)-N,N-dimethylacetamide (32.23 mg).

MS (ESI) M/Z: 456.8 [M+H] ⁺ .

¹ H NMR (400 MHz, DMSO-d ₆ ) δ9.59 (s, 1H), 8.74 (s, 1H), 8.28–8.22 (m, 2H), 8.08–8.02 (m, 2H), 7.85 (d, J=8.4 Hz, 2H), 7.62 (d, J=8.0 Hz, 2H), 5.18 (s, 2H), 3.07 (s, 3H), 2.89 (s, 3H), 2.59 (s, 3H). C ₂₅ H ₂₁ FN ₆ O ₂ .

Embodiment 35:

2-(4-(4-(7-fluoro-6-(1H-1,2,4-triazol-3-yl)isoquinolin-4-yl)phenyl)-1H-pyrazol-1-yl)-N,N-dimethylacetamide

Steps:

Step A: Methyl 7-fluoroisoquinoline-6-carboxylate (900 mg, 4.39 mmol) was dissolved in acetic acid (20 ml), heated to 110 °C, and N-bromosuccinimide (936.8 mg, 5.26 mmol) was added in portions, and the mixture was reacted at 110 °C for 3 hours.

After the disappearance of the starting material as monitored by TLC, the mixture was concentrated under reduced pressure and the crude product was purified by silica gel column chromatography to give methyl 4-bromo-7-fluoroisoquinoline-6-carboxylate (800 mg).

MS (ESI) M/Z: 284.0 [M+H] ⁺ .

Step B: Methyl 4-bromo-7-fluoroisoquinoline-6-carboxylate (400 mg, 1.41 mmol) was dissolved in ammonia methanol solution (20 ml) and refluxed at 50 °C for 64 hours.

After the starting material disappeared as monitored by TLC, the mixture was concentrated under reduced pressure to give crude 4-bromo-7-fluoroisoquinoline-6-carboxamide (380 mg).

MS (ESI) M/Z: 269.1 [M+H] ⁺ .

Step C: 4-bromo-7-fluoroisoquinoline-6-carboxamide (380 mg, 1.41 mmol) and N,N-dimethylformamide dimethyl acetal (3 ml) were added into a microwave tube and reacted at 100 °C for 3 hours.

After the disappearance of the starting material monitored by LCMS, the mixture was cooled, petroleum ether (12 ml) was added, the temperature was lowered to below 5°C for crystallization, and filtered to give (E)-4-bromo-N-((dimethylamino)methylene)-7-fluoroisoquinoline-6-carboxamide (346 mg).

MS (ESI) M/Z: 324.2 [M+H] ⁺ .

Step D: (E)-4-Bromo-N-((dimethylamino)methylene)-7-fluoroisoquinoline-6-carboxamide (346 mg, 1.07 mmol) was dissolved in acetic acid (5 ml), hydrazine hydrate (60 mg, 85%, 1.02 mmol) was added and the reaction was carried out at 98 °C overnight.

After the disappearance of the starting material monitored by TLC, petroleum ether was added and concentrated under reduced pressure to remove most of the acetic acid. The mixture was slurried with (petroleum ether/ethyl acetate=20/1) and filtered to give 4-bromo-7-fluoro-6-(1H-1,2,4-triazol-3-yl)isoquinoline (260 mg).

MS (ESI) M/Z: 292.8 [M+H] ⁺ .

Step E: 4-Bromo-7-fluoro-6-(1H-1,2,4-triazol-3-yl)isoquinoline (100 mg, 0.34 mmol), N,N-dimethyl-2-(4-(4-(4,4,5,5-tetramethyl-1,3,2-dioxaborolan-2-yl)phenyl)-1H-pyrazol-1-yl)acetamide (133.3 mg, 0.38 mmol), anhydrous potassium carbonate (117.3 mg, 0.85 mmol) were dissolved in dioxane (4 ml) and water (1 ml), 1,1'-bis(di-phenylphosphino)ferrocenepalladium chloride (25 mg, 0.034 mmol) was added, the reaction liquid was replaced with nitrogen three times, and the reaction was carried out at 90°C overnight.

After the disappearance of the starting material as monitored by TLC, the mixture was concentrated under reduced pressure and the crude product was purified by silica gel column chromatography to give 2-(4-(4-(7-fluoro-6-(1H-1,2,4-triazol-3-yl)isoquinolin-4-yl)phenyl)-1H-pyrazol-1-yl)-N,N-dimethylacetamide (46.84 mg).

MS (ESI) M/Z: 442.2 [M + H] ⁺ .

¹ H NMR (400 MHz, DMSO-d ₆ ) δ 14.39 (s, 1H), 9.36 (s, 1H), 8.73–8.61 (m, 2H), 8.51 (s, 1H), 8.23 (s, 1H), 8.22–8.11 (m, 1H), 8.03 (s, 1H), 7.82 (d, J=8.1 Hz, 2H), 7.63–7.55 (m, 2H), 5.18 (s, 2H), 3.07 (s, 3H), 2.89 (s, 3H). C ₂₄ H ₂₀ FN ₇ O.

Embodiment 36:

2-(4-(4-(7-fluoro-6-(1-methyl-1H-1,2,4-triazol-3-yl)isoquinolin-4-yl)phenyl)-1H-pyrazol-1-yl)-N,N-dimethylacetamide

Steps:

Step A: 4-bromo-7-fluoro-6-(1H-1,2,4-triazol-3-yl)isoquinoline (160 mg, 0.55 mmol) and anhydrous potassium carbonate (188.4 mg, 1.37 mmol) were dissolved in DMF (4 ml), and iodomethane (155.1 mg, 1.09 mmol) was added dropwise, and the mixture was reacted at room temperature for 60 hours.

After the disappearance of the starting material by TLC monitoring, water (10 ml) was added, and the mixture was extracted with ethyl acetate (20 ml × 2 times). The organic phases were combined, washed with saturated brine (20 ml), then dried over anhydrous sodium sulfate, filtered, and finally concentrated under reduced pressure. The crude product was purified by silica gel column chromatography to give 4-bromo-7-fluoro-6-(1-methyl-1H-1,2,4-triazol-3-yl)isoquinoline (168 mg).

MS (ESI) M/Z: 307.0 [M+H] ⁺ .

Step B: 4-Bromo-7-fluoro-6-(1-methyl-1H-1,2,4-triazol-3-yl)isoquinoline (168 mg, 0.55 mmol), N,N-dimethyl-2-(4-(4-(4,4,5,5-tetramethyl-1,3,2-dioxaborolan-2-yl)phenyl)-1H-pyrazol-1-yl)acetamide (214 mg, 0.60 mmol), anhydrous potassium carbonate (190 mg, 1.38 mmol) were dissolved in dioxane (4 ml) and water (1 ml), 1,1'-bis(di-phenylphosphino)ferrocenepalladium chloride dichloromethane complex (45 mg, 0.055 mmol) was added, the reaction liquid was replaced with nitrogen three times, and the reaction was carried out at 90°C overnight.

After the disappearance of the starting material as monitored by TLC, the mixture was concentrated under reduced pressure and the crude product was purified by silica gel column chromatography and then purified by preparative purification to give 2-(4-(4-(7-fluoro-6-(1-methyl-1H-1,2,4-triazol-3-yl)isoquinolin-4-yl)phenyl)-1H-pyrazol-1-yl)-N,N-dimethylacetamide (21.12 mg).

MS (ESI) M/Z: 456.1 [M + H] ⁺ .

¹ H NMR (400 MHz, DMSO-d ₆ ) δ 9.36 (s, 1H), 8.66–8.57 (m, 2H), 8.50 (s, 1H), 8.25–8.15 (m, 2H), 8.03 (s, 1H), 7.82 (d, J=7.8 Hz, 2H), 7.58 (d, J=8.1 Hz, 2H), 5.18 (s, 2H), 3.95 (s, 3H), 3.07 (s, 3H), 2.89 (s, 3H).

Embodiment 37:

2-(4-(4-(7-fluoro-6-(1H-pyrazol-3-yl)isoquinolin-4-yl)phenyl)-1H-pyrazol-1-yl)-N,N-dimethylacetamide

Steps:

Step A: 6-bromo-7-fluoroisoquinoline (500 mg, 2.212 mmol) was added to acetic acid (10 ml), the temperature was raised to 110 °C, N-iodosuccinimide (746 mg, 3.318 mmol) was added in batches and the mixture was reacted at 110 °C for 5 hours.

After TLC monitoring shows that the raw material disappears, cool it. Add sodium bicarbonate aqueous solution to adjust the pH to neutral, then add ethyl acetate (30 ml x 2) for extraction, and combine the organic phases. The organic phase is washed with saturated brine (20 ml), dried over anhydrous sodium sulfate, and concentrated under reduced pressure. The resulting residue is purified by silica gel column chromatography to obtain 6-bromo-7-fluoro-4-iodoisoquinoline (400 mg).

MS (ESI) M/Z: 351.9 [M + H] ⁺ .

Step B: 6-bromo-7-fluoro-4-iodoisoquinoline (400 mg, 1.137 mmol), N,N-dimethyl-2-(4-(4-(4,4,5,5-tetramethyl-1,3,2-dioxaboroborolan-2-yl)phenyl)-1H-pyrazol-1-yl)acetamide (403 mg, 1.137 mmol) were added to dioxane/water (4/1, 10 ml), potassium carbonate (313 mg, 2.274 mmol) and [1,1'-bis(diphenylphosphino)ferrocene]dichloropalladium dichloromethane complex (83 mg, 0.114 mmol), the atmosphere was replaced with nitrogen three times, and the temperature was raised to 100 °C for reaction for 3 hours.

After LCMS monitoring showed that the raw material disappeared, the mixture was cooled. Water (20 ml) and dichloromethane/methanol (30 ml x 2) were added for extraction, and the organic phases were combined. The organic phase was washed with saturated brine (20 ml), dried over anhydrous sodium sulfate, and concentrated under reduced pressure. The obtained residue was purified by silica gel column chromatography to obtain 2-(4-(4-(6-bromo-7-fluoroisoquinolin-4-yl)phenyl)-1H-pyrazol-1-yl)-N,N-dimethylacetamide (380 mg).

MS (ESI) M/Z: 453.1 [M+H] ⁺ .

Step C: Add 2-(4-(4-(6-bromo-7-fluoroisoquinolin-4-yl)phenyl)-1H-pyrazol-1-yl)-N,N-dimethylacetamide (180 mg, 0.397 mmol) and (1H-pyrazol-3-yl)boric acid (57 mg, 0.516 mmol) to dioxane/water (4/1, 10 ml), add potassium carbonate (109 mg, 0.794 mmol) and [1,1'-bis(diphenylphosphino)ferrocene] dichloropalladium dichloromethane complex (29 mg, 0.040 mmol), replace with nitrogen three times, and heat to 110 °C for 3 hours.

After LCMS monitoring showed that the raw material disappeared, the mixture was cooled. Water (20 ml) and ethyl acetate (30 ml x 2) were added for extraction, and the organic phases were combined. The organic phase was washed with saturated brine (20 ml), dried over anhydrous sodium sulfate, and concentrated under reduced pressure. The obtained residue was purified by silica gel column chromatography to obtain 2-(4-(4-(7-fluoro-6-(1H-pyrazol-3-yl)isoquinolin-4-yl)phenyl)-1H-pyrazol-1-yl)-N,N-dimethylacetamide (72.2 mg).

MS (ESI) M/Z: 441.1 [M+H] ⁺ .

¹ H NMR (400 MHz, DMSO-d ₆ ) δ 13.23 (s, 1H), 9.32 (s, 1H), 8.59 (d, J = 7.3 Hz, 1H), 8.46 (s, 1H), 8.22 (s, 1H), 8.14 (d, J = 11.6 Hz, 1H), 8.02 (s, 1H), 7.94–7.88 (m, 1H), 7.81 (d, J = 7.8 Hz, 2H), 7.58 (d, J = 7.9 Hz, 2H), 6.80–6.76 (m, 1H), 5.18 (s, 2H), 3.07 (s, 3H), 2.89 (s, 3H).

Embodiment 38:

2-(4-(4-(7-fluoro-6-(1H-imidazol-2-yl)isoquinolin-4-yl)phenyl)-1H-pyrazol-1-yl)-N,N-dimethylacetamide

Steps:

Step A: 4-Bromo-7-fluoroisoquinoline-6-carbonitrile (200 mg, 0.8 mmol) was dissolved in methanol (2 ml), sodium methoxide (4.32 mg, 0.08 mmol) was added, and the mixture was reacted at 40°C for 1 hour.

The reaction solution was cooled to room temperature, 2,2-dimethoxyethylamine (84.11 mg, 0.8 mmol) and acetic acid (0.2 ml) were added, and the mixture was reacted at 70 degrees Celsius for 1 hour. The reaction solution was cooled to room temperature, methanol (1.2 ml) and hydrochloric acid (6M, 1.2 ml) were added, and the mixture was reacted at 70 degrees Celsius for 4 hours.

TLC monitoring showed that the reaction of the raw material was incomplete. After post-treatment, saturated aqueous sodium bicarbonate solution (15 ml) was added, and the mixed solution was extracted with dichloromethane (30 ml × 2 times). The organic phases were combined, washed with saturated brine (20 ml), then dried over anhydrous sodium sulfate, filtered, and finally concentrated under reduced pressure. The crude product was purified by silica gel column chromatography to obtain 4-bromo-7-fluoro-6-(1H-imidazol-2-yl)isoquinoline (80 mg).

MS (ESI) M/Z: 291.8 [M+H] ⁺ .

Step B: 4-Bromo-7-fluoro-6-(1H-imidazol-2-yl)isoquinoline (80 mg, 0.275 mmol) and N,N-dimethyl-2-(4-(4,4,5,5-tetramethyl-1,3,2-dioxaborolan-2-yl)-1H-pyrazol-1-yl)acetamide (126.87 mg, 0.36 mmol) were dissolved in dioxane (8 ml) and water (2 ml). Potassium carbonate (75.88 mg, 0.55 mmol) and 1,1'-bis(diphenylphosphinoferrocene)palladium dichloride (40.19 mg, 0.055 mmol) were added. The reaction liquid was replaced with nitrogen three times and reacted at 105 degrees Celsius for 3 hours.

After the disappearance of the starting material by TLC monitoring, water (20 ml) was added, and the mixture was extracted with ethyl acetate (30 ml × 2 times). The organic phases were combined, washed with saturated brine (20 ml), then dried over anhydrous sodium sulfate, filtered, and finally concentrated under reduced pressure. The crude product was prepared and purified to give 2-(4-(4-(7-fluoro-6-(1H-imidazol-2-yl)isoquinolin-4-yl)phenyl)-1H-pyrazol-1-yl)-N,N-dimethylacetamide (7.11 mg).

MS (ESI) M/Z: 440.0 [M+H] ⁺ .

¹ H NMR (400 MHz, DMSO-d ₆ ) δ 12.51 (s, 1H), 9.33 (s, 1H), 8.61 (d, J = 7.2 Hz, 1H), 8.48 (s, 1H), 8.25-8.16 (m, 2H), 8.02 (s, 1H), 7.82 (d, J = 8.0 Hz, 2H), 7.58 (d, J = 8.0 Hz, 2H), 7.23 (s, 2H), 5.18 (s, 2H), 3.07 (s, 3H), 2.89 (s, 3H).

Embodiment 39:

2-(4-(4-(7-fluoro-6-(5-methyl-4H-1,2,4-triazol-3-yl)isoquinolin-4-yl)phenyl)-1H-pyrazol-1-yl)-N,N-dimethylacetamide

Steps:

Step A: Add 7-fluoroisoquinoline-6-carbohydrazide (250 mg, 1.22 mmol), acetamidoamine hydrochloride (230.4 mg, 2.44 mmol), sodium ethoxide (131.8 mg, 2.44 mmol) and anhydrous ethanol (5 ml) into a microwave tube and react at 98 °C overnight.

After the disappearance of the starting material by TLC monitoring, the residue was concentrated under reduced pressure. The crude product was purified by silica gel column chromatography to give 7-fluoro-6-(5-methyl-4H-1,2,4-triazol-3-yl)isoquinoline (230 mg).

MS (ESI) M/Z: 228.9 [M + H] ⁺ .

Step B: 7-Fluoro-6-(5-methyl-4H-1,2,4-triazol-3-yl)isoquinoline (230 mg, 1.01 mmol) was dissolved in acetic acid (5 ml), heated to 110 °C, N-bromosuccinimide (215.2 mg, 1.21 mmol) was added in portions, and the mixture was reacted at 110 °C for 3 hours.

After the starting material disappeared as monitored by TLC, the mixture was concentrated under reduced pressure and the crude product was purified by silica gel column chromatography to give 4-bromo-7-fluoro-6-(5-methyl-4H-1,2,4-triazol-3-yl)isoquinoline (150 mg).

MS (ESI) M/Z: 307.1 [M+H] ⁺ .

Step C: 4-bromo-7-fluoro-6-(5-methyl-4H-1,2,4-triazol-3-yl)isoquinoline (75 mg, 0.24 mmol), N,N-dimethyl-2-(4-(4-(4,4,5,5-tetramethyl-1,3,2-dioxaborolan-2-yl)phenyl)-1H-pyrazol-1-yl)acetamide (95.4 mg, 0.27 mmol), anhydrous potassium carbonate (84.2 mg, 0.61 mmol) were dissolved in dioxane (4 ml) and water (1 ml), 1,1'-bis(di-phenylphosphino)ferrocenepalladium chloride (17.9 mg, 0.024 mmol) was added, the reaction liquid was replaced with nitrogen three times, and the reaction was carried out at 90°C overnight.

After the disappearance of the starting material as monitored by TLC, the mixture was concentrated under reduced pressure and the crude product was purified by silica gel column chromatography and then purified by preparative purification to give 2-(4-(4-(7-fluoro-6-(5-methyl-4H-1,2,4-triazol-3-yl)isoquinolin-4-yl)phenyl)-1H-pyrazol-1-yl)-N,N-dimethylacetamide (25.35 mg).

MS (ESI) M/Z: 456.3 [M + H] ⁺ .

¹ H NMR (400 MHz, DMSO-d ₆ ) δ 13.96 (s, 1H), 9.35 (s, 1H), 8.60 (d, J = 7.0 Hz, 1H), 8.48 (s, 1H), 8.25–8.12 (m, 2H), 8.03 (s, 1H), 7.82 (d, J = 8.4 Hz, 2H), 7.58 (d, J = 8.4 Hz, 2H), 5.18 (s, 2H), 3.07 (s, 3H), 2.89 (s, 3H), 2.40 (s, 3H).

Embodiment 40:

N,N-Dimethyl-2-(4-(4-(6-(5-methyl-1,3,4-oxadiazol-2-yl)isoquinolin-4-yl)phenyl)-1H-pyrazol-1-yl)acetamide

Steps:

Step A: 2-(4-bromoisoquinolin-6-yl)-5-methyl-1,3,4-oxadiazole (60 mg, 0.21 mmol) and N,N-dimethyl-2-(4-(4-(4,4,5,5-tetramethyl-1,3,2-dioxaborolan-2-yl)phenyl)-1H-pyrazol-1-yl)acetamide (95.5 mg, 0.27 mmol) were dissolved in dioxane (8 ml) and water (2 ml). Potassium carbonate (57 mg, 0.41 mmol) and [1,1'-bis(diphenylphosphino)ferrocene]dichloropalladium dichloromethane complex (15 mg, 0.02 mmol) were added. The reaction liquid was replaced with nitrogen three times and reacted at 110 degrees Celsius for 3 hours.

After the disappearance of the starting material by TLC monitoring, water (20 ml) was added, and the mixture was extracted with ethyl acetate (30 ml × 2 times). The organic phases were combined, washed with saturated brine (20 ml), then dried over anhydrous sodium sulfate, filtered, and finally concentrated under reduced pressure. The crude product was prepared and purified to give N,N-dimethyl-2-(4-(4-(6-(5-methyl-1,3,4-oxadiazol-2-yl)isoquinolin-4-yl)phenyl)-1H-pyrazol-1-yl)acetamide (18.92 mg).

MS (ESI) M/Z: 439.3 [M + H] ⁺ .

¹ H NMR (400 MHz, DMSO-d ₆ ) δ 9.47 (d, J = 0.8 Hz, 1H), 8.60 (s, 1H), 8.48–8.41 (m, 2H), 8.33–8.20 (m, 2H), 8.03 (s, 1H), 7.84 (d, J = 8.4 Hz, 2H), 7.61 (d, J = 8.4 Hz, 2H), 5.18 (s, 2H), 3.07 (s, 3H), 2.89 (s, 3H), 2.59 (s, 3H).

Embodiment 41:

2-(4-(4-(6-cyano-7-fluoroisoquinolin-4-yl)phenyl)-1H-pyrazol-1-yl)-N,N-dimethylacetamide

Steps:

Step A: 4-Bromo-7-fluoroisoquinoline-6-carbonitrile (60 mg, 0.24 mmol) and N,N-dimethyl-2-(4-(4-(4,4,5,5-tetramethyl-1,3,2-dioxaborolan-2-yl)phenyl)-1H-pyrazol-1-yl)acetamide (110 mg, 0.31 mmol) were dissolved in dioxane (8 ml) and water (2 ml). Potassium carbonate (66 mg, 0.47 mmol) and [1,1'-bis(diphenylphosphino)ferrocene]dichloropalladium dichloromethane complex (17 mg, 0.02 mmol) were added. The reaction liquid was replaced with nitrogen three times and reacted at 110 degrees Celsius for 3 hours.

After TLC monitoring of the disappearance of the starting material, water (20 ml) was added, and the mixture was extracted with ethyl acetate (30 ml × 2 times). The organic phases were combined, washed with saturated brine (20 ml), then dried over anhydrous sodium sulfate, filtered, and finally concentrated under reduced pressure. The crude product was purified by silica gel column chromatography (eluent: ethyl acetate) to give 2-(4-(4-(6-cyano-7-fluoroisoquinolin-4-yl)phenyl)-1H-pyrazol-1-yl)-N,N-dimethylacetamide (45.07 mg).

MS (ESI) M/Z: 399.8 [M+H] ⁺ .

¹ H NMR (400 MHz, DMSO-d ₆ ) δ 9.46 (s, 1H), 8.65 (s, 1H), 8.51 (d, J=6.1 Hz, 1H), 8.38 (d, J=9.6 Hz, 1H), 8.21 (s, 1H), 8.01 (s, 1H), 7.81 (d, J=8.0 Hz, 2H), 7.60 (d, J=8.2 Hz, 2H), 5.17 (s, 2H), 3.07 (s, 3H), 2.88 (s, 3H).

Embodiment 42:

2-(4-(4-(7-fluoro-6-(5-methyl-1,3,4-oxadiazol-2-yl)isoquinolin-4-yl)phenyl)-1H-pyrazol-1-yl)-1-(pyrrolidin-1-yl)ethan-1-one

Referring to the synthesis method of Example 22, 2-(4-(4-(7-fluoro-6-(5-methyl-1,3,4-oxadiazol-2-yl)isoquinolin-4-yl)phenyl)-1H-pyrazol-1-yl)-1-(pyrrolidin-1-yl)ethan-1-one (88.72 mg) was obtained.

MS (ESI) M/Z: 483.2 [M + H] ⁺ .

¹ H NMR (400 MHz, DMSO-d ₆ ) δ 9.43 (s, 1H), 8.59 (s, 1H), 8.54 (d, J = 6.4 Hz, 1H), 8.34 (d, J = 11.0 Hz, 1H), 8.25 (s, 1H), 8.04 (s, 1H), 7.83 (d, J = 8.0 Hz, 2H), 7.61 (d, J = 8.1 Hz, 2H), 5.09 (s, 2H), 3.53 (t, J = 6.8 Hz, 2H), 3.35 (t, J = 6.9 Hz, 2H), 2.60 (s, 3H), 1.99–1.89 (m, 2H), 1.87–1.74 (m, 2H).

Embodiment 43:

2-(4-(4-(7-fluoro-6-(1H-pyrazol-3-yl)isoquinolin-4-yl)phenyl)-1H-pyrazol-1-yl)-1-(pyrrolidin-1-yl)ethan-1-one

Referring to the synthesis method of Example 37, 2-(4-(4-(7-fluoro-6-(1H-pyrazol-3-yl)isoquinolin-4-yl)phenyl)-1H-pyrazol-1-yl)-1-(pyrrolidin-1-yl)ethan-1-one (25.83 mg) was obtained.

MS (ESI) M/Z: 467.0 [M+H] ⁺ .

¹ H NMR (400 MHz, DMSO-d ₆ ) δ 13.23 (s, 1H), 9.32 (s, 1H), 8.59 (d, J = 7.2 Hz, 1H), 8.46 (s, 1H), 8.24 (s, 1H), 8.15 (d, J = 11.6 Hz, 1H), 8.03 (s, 1H), 7.91 (s, 1H), 7.82 (d, J = 7.9 Hz, 2H), 7.62–7.46 (m, 2H), 6.78 (dd, J = 4.7, 2.3 Hz, 1H), 5.09 (s, 2H), 3.53 (t, J = 6.8 Hz, 2H), 3.37–2.32 (m, 2H), 1.99–1.89 (m, 2H), 1.87–1.75 (m, 2H).

Embodiment 44:

2-(4-(4-(7-fluoro-6-(5-methyl-4H-1,2,4-triazol-3-yl)isoquinolin-4-yl)phenyl)-1H-pyrazol-1-yl)-1-(pyrrolidin-1-yl)ethan-1-one

Referring to the synthesis method of Example 39, 2-(4-(4-(7-fluoro-6-(5-methyl-4H-1,2,4-triazol-3-yl)isoquinolin-4-yl)phenyl)-1H-pyrazol-1-yl)-1-(pyrrolidin-1-yl)ethan-1-one (50.05 mg) was obtained.

MS (ESI) M/Z: 482.1 [M+H] ⁺ .

¹ H NMR (400 MHz, DMSO-d ₆ ) δ 13.95 (s, 1H), 9.35 (s, 1H), 8.60 (d, J = 7.0 Hz, 1H), 8.48 (s, 1H), 8.25 (s, 1H), 8.16 (d, J = 11.0 Hz, 1H), 8.03 (s, 1H), 7.82 (d, J = 8.4 Hz, 2H), 7.58 (d, J = 7.9 Hz, 2H), 5.09 (s, 2H), 3.53 (t, J = 6.8 Hz, 2H), 3.37-3.33 (m, 2H), 2.40 (s, 3H), 1.99-1.89 (m, 2H), 1.87-1.75 (m, 2H).

Embodiment 45:

1-(4,4-difluoropiperidin-1-yl)-2-(4-(4-(7-fluoroisoquinolin-4-yl)phenyl)-1H-pyrazol-1-yl)ethan-1-one

Referring to the synthesis method of Example 22, 1-(4,4-difluoropiperidin-1-yl)-2-(4-(4-(7-fluoroisoquinolin-4-yl)phenyl)-1H-pyrazol-1-yl)ethan-1-one (27.31 mg) was obtained.

MS (ESI) M/Z: 451.2 [M+H] ⁺ .

¹ H NMR (400 MHz, DMSO-d ₆ ) δ 9.34 (s, 1H), 8.48 (s, 1H), 8.20 (s, 1H), 8.10–7.95 (m, 3H), 7.82–7.68 (m, 3H), 7.59–7.52 (m, 2H), 5.27 (s, 2H), 3.69–3.50 (m, 4H), 2.19–1.88 (m, 4H).

Embodiment 46:

2-(4-(4-(7-fluoroisoquinolin-4-yl)phenyl)-1H-pyrazol-1-yl)-1-(2-methyl-2,6-dihydropyrrolo[3,4-c]pyrazol-5(4H)-yl)ethan-1-one

Referring to the synthesis method of Example 22, 2-(4-(4-(7-fluoroisoquinolin-4-yl)phenyl)-1H-pyrazol-1-yl)-1-(2-methyl-2,6-dihydropyrrolo[3,4-c]pyrazol-5(4H)-yl)ethan-1-one (101.75 mg) was obtained.

MS (ESI) M/Z: 453.2 [M + H] ⁺ .

¹ H NMR (400 MHz, DMSO-d ₆ ) δ 9.35 (s, 1H), 8.49 (s, 1H), 8.25 (s, 1H), 8.10–7.96 (m, 3H), 7.84–7.77 (m, 2H), 7.76–7.70 (m, 1H), 7.63–7.53 (m, 3H), 5.23 (s, 2H), 4.73 (s, 2H), 4.43 (s, 2H), 3.86 (s, 3H).

Embodiment 47:

2-(4-(4-(7-fluoroisoquinolin-4-yl)phenyl)-1H-pyrazol-1-yl)-1-(2-oxa-7-azaspiro[3.5]nonan-7-yl)ethan-1-one

Referring to the synthesis method of Example 22, 2-(4-(4-(7-fluoroisoquinolin-4-yl)phenyl)-1H-pyrazol-1-yl)-1-(2-oxa-7-azaspiro[3.5]nonan-7-yl)ethan-1-one (49.62 mg) was obtained.

MS (ESI) M/Z: 457.3 [M + H] ⁺ .

¹ H NMR (400 MHz, DMSO-d ₆ ) δ 9.34 (s, 1H), 8.48 (s, 1H), 8.20 (s, 1H), 8.06 (dd, J=9.3, 2.7 Hz, 1H), 8.04–7.95 (m, 2H), 7.82–7.69 (m, 3H), 7.59–7.52 (m, 2H), 5.19 (s, 2H), 4.39–4.29 (m, 4H), 3.50–3.35 (m, 4H), 1.85 (t, J=5.7 Hz, 2H), 1.75 (t, J=5.7 Hz, 2H).

Embodiment 48:

2-(4-(2-(4-(4-(7-fluoroisoquinolin-4-yl)phenyl)-1H-pyrazol-1-yl)acetyl)piperazin-1-yl)acetonitrile

Referring to the synthesis method of Example 22, 2-(4-(2-(4-(4-(7-fluoroisoquinolin-4-yl)phenyl)-1H-pyrazol-1-yl)acetyl)piperazin-1-yl)acetonitrile (38.11 mg) was obtained.

MS (ESI) M/Z: 455.3 [M + H] ⁺ .

¹ H NMR (400 MHz, DMSO-d ₆ ) δ 9.34 (s, 1H), 8.48 (s, 1H), 8.22 (s, 1H), 8.10–7.95 (m, 3H), 7.82–7.68 (m, 3H), 7.59–7.52 (m, 2H), 5.23 (s, 2H), 3.82 (s, 2H), 3.62–3.50 (m, 4H), 2.61–2.45 (m, 4H).

Embodiment 49:

2-(4-(4-(7-fluoro-6-(1H-imidazol-2-yl)isoquinolin-4-yl)phenyl)-1H-pyrazol-1-yl)-1-(pyrrolidin-1-yl)ethan-1-one

Referring to the synthesis method of Example 38, 2-(4-(4-(7-fluoro-6-(1H-imidazol-2-yl)isoquinolin-4-yl)phenyl)-1H-pyrazol-1-yl)-1-(pyrrolidin-1-yl)ethan-1-one (23.09 mg) was obtained.

MS (ESI) M/Z: 467.2 [M+H] ⁺ .

¹ H NMR (400 MHz, DMSO-d ₆ ) δ 12.50 (s, 1H), 9.33 (s, 1H), 8.61 (d, J = 7.2 Hz, 1H), 8.48 (s, 1H), 8.27–8.16 (m, 2H), 8.03 (s, 1H), 7.82 (d, J = 8.2 Hz, 2H), 7.62–7.55 (m, 2H), 7.34 (dd, J = 2.2, 1.1 Hz, 1H), 7.12 (d, J = 1.2 Hz, 1H), 5.09 (s, 2H), 3.53 (t, J = 6.8 Hz, 2H), 3.37–3.32 (m, 2H), 1.99–1.89 (m, 2H), 1.87–1.75 (m, 2H).

Embodiment 50:

1-(3,3-Difluoropyrrolidin-1-yl)-2-(4-(4-(7-fluoroisoquinolin-4-yl)phenyl)-1H-pyrazol-1-yl)ethan-1-one

Referring to the synthesis method of Example 22, 1-(3,3-difluoropyrrolidin-1-yl)-2-(4-(4-(7-fluoroisoquinolin-4-yl)phenyl)-1H-pyrazol-1-yl)ethan-1-one (78.87 mg) was obtained.

MS (ESI) M/Z: 437.0 [M+H] ⁺ .

¹ H NMR (400 MHz, DMSO-d ₆ ) δ 9.34 (s, 1H), 8.48 (s, 1H), 8.22 (d, J=3.5 Hz, 1H), 8.09–7.95 (m, 3H), 7.83–7.69 (m, 3H), 7.60–7.50 (m, 2H), 5.16 (d, J=22.9 Hz, 2H), 4.15–4.01 (m, 1H), 3.88–3.72 (m, 2H), 3.62–3.53 (m, 1H), 2.60–2.53 (m, 1H), 2.47–2.35 (m, 1H).

Embodiment 51:

2-(4-(4-(8-amino-3-(5-methyl-1,3,4-oxadiazol-2-yl)-1,7-naphthyridin-5-yl)phenyl)-1H-pyrazol-1-yl)-N,N-dimethylacetamide

Steps:

Step A: Ethyl 8-((tert-butoxycarbonyl)amino)-1,7-naphthyridine-3-carboxylate (1 g, 3.15 mmol), sodium hydroxide (378 mg, 9.45 mmol), THF (10 ml), and water (5 ml) were reacted at room temperature overnight.

After TLC monitoring showed that the raw material disappeared, citric acid (40 ml) and dichloromethane (60 ml) were added, stirred, and separated. The organic phase was washed with saturated brine (10 ml), dried over anhydrous sodium sulfate, and concentrated under reduced pressure to obtain 8-((tert-butoxycarbonyl)amino)-1,7-naphthyridine-3-carboxylic acid (720 mg).

MS (ESI) M/Z: 290.0 [M+H] ⁺ .

Step B: 8-((tert-Butyloxycarbonyl)amino)-1,7-naphthyridine-3-carboxylic acid (720 mg, 2.49 mmol), acetohydrazide (371 mg, 5 mmol), HATU (1.87 g, 4.92 mmol), DIPEA (972 mg, 7.53 mmol) were dissolved in DMF (10 mL) and dichloromethane (2 mL) and stirred at room temperature overnight.

After TLC monitoring showed that the raw material disappeared, water (30 ml) and dichloromethane (40 ml) were added, stirred, and separated. The organic phase was washed with saturated brine (10 ml), dried over anhydrous sodium sulfate, and concentrated to dryness under reduced pressure. The obtained residue was purified by silica gel column chromatography to obtain tert-butyl (3-(2-acetylhydrazine-1-carbonyl)-1,7-naphthyridin-8-yl)carbamate (400 mg).

MS (ESI) M/Z: 346.3 [M + H] ⁺ .

Step C: tert-Butyl (3-(2-acetylhydrazine-1-carbonyl)-1,7-naphthyridin-8-yl)carbamate (400 mg, 1.16 mmol), TsCl (332 mg, 1.74 mmol) and triethylamine (351 mg, 3.48 mmol) were dissolved in acetonitrile (10 mL) and stirred at room temperature overnight.

After TLC monitoring shows that the raw material disappears, cool it. Add sodium bicarbonate aqueous solution (50 ml) and ethyl acetate (50 ml), stir and separate. The organic phase is washed with saturated brine (10 ml), dried over anhydrous sodium sulfate, and concentrated to dryness under reduced pressure. The resulting residue is purified by silica gel column chromatography to obtain 3-(5-methyl-1,3,4-oxadiazol-2-yl)-1,7-naphthyridine-8-amine (370 mg).

MS (ESI) M/Z: 227.8 [M+H] ⁺ .

Step D: 3-(5-methyl-1,3,4-oxadiazol-2-yl)-1,7-naphthyridine-8-amine (200 mg, 0.61 mmol) and NBS (120 mg, 0.67 mmol) were added to acetic acid (5 ml), and the mixture was stirred at 100 °C for 5 hours.

After TLC monitoring shows that the raw material disappears, cool. Add sodium bicarbonate aqueous solution (30 ml) and ethyl acetate (30 ml), stir and separate. Wash the organic phase with saturated brine (10 ml), dry over anhydrous sodium sulfate, and concentrate to dryness under reduced pressure. The resulting residue is purified by silica gel column chromatography to obtain 5-bromo-3-(5-methyl-1,3,4-oxadiazol-2-yl)-1,7-naphthyridine-8-amine (100 mg).

MS (ESI) M/Z: 305.9 [M + H] ⁺ .

Step E: 5-Bromo-3-(5-methyl-1,3,4-oxadiazol-2-yl)-1,7-naphthyridin-8-amine (100 mg, 0.33 mmol), N,N-dimethyl-2-(4-(4-(4,4,5,5-tetramethyl-1,3,2-dioxaborolan-2-yl)phenyl)-1H-pyrazol-1-yl)acetamide (141 mg, 0.396 mmol), potassium carbonate (91 mg, 0.66 mmol), Pd(dppf) _Cl2 (24 mg, 0.033 mmol) were added to 1.4-dioxane (5 ml) and water (1 ml), and the mixture was stirred at 100 °C for 2 hours.

After TLC monitoring shows that the raw material disappears, cool. Add water (30 ml) and ethyl acetate (30 ml), stir, and separate. The organic phase is washed with saturated brine (10 ml), dried over anhydrous sodium sulfate, and concentrated to dryness under reduced pressure. The obtained residue is prepared and purified to obtain 2-(4-(4-(8-amino-3-(5-methyl-1,3,4-oxadiazol-2-yl)-1,7-naphthyridin-5-yl)phenyl)-1H-pyrazol-1-yl)-N,N-dimethylacetamide (0.82 mg).

MS (ESI) M/Z: 454.9 [M + H] ⁺ .

¹ H NMR (400 MHz, DMSO-d ₆ ) δ 9.34 (s, 1H), 8.62–8.52 (m, 1H), 8.21–8.16 (m, 1H), 8.04–7.95 (m, 2H), 7.81–7.73 (m, 2H), 7.54–7.48 (m, 2H), 7.31 (s, 2H), 5.16 (s, 2H), 3.07 (s, 3H), 2.88 (s, 3H), 2.61 (s, 3H).

Embodiment 52:

2-(4-(4-(7-fluoro-6-(2-methyl-2H-tetrazol-5-yl)isoquinolin-4-yl)phenyl)-1H-pyrazol-1-yl)-N,N-dimethylacetamide

Steps:

Step A: 6-Bromo-7-fluoroisoquinoline (500.0 mg, 2.22 mmol) and zinc cyanide (520.0 mg, 4.44 mmol) were dissolved in DMF (12 ml). Tetrakis(triphenylphosphine)palladium (256.8 mg, 0.22 mmol) was added. The reaction liquid was replaced with nitrogen three times and reacted at 98 degrees Celsius overnight.

After TLC monitoring of the disappearance of the raw material, water (20 ml) was added, filtered, and the filtrate was extracted with ethyl acetate (30 ml × 2 times). The organic phases were combined, washed with saturated brine (20 ml), then dried over anhydrous sodium sulfate, filtered, and finally concentrated under reduced pressure. The crude product was purified by silica gel column chromatography to obtain 7-fluoroisoquinoline-6-carbonitrile (380.0 mg).

MS (ESI) M/Z: 173.0 [M+H] ⁺ .

Step B: 7-Fluoroisoquinoline-6-carbonitrile (380 mg, 2.21 mmol) was dissolved in acetic acid (10 ml), heated to 110 °C, and N-bromosuccinimide (471.42 mg, 2.65 mmol) was added, and the mixture was reacted at 110 °C for 3 hours.

After the disappearance of the starting material by TLC monitoring, aqueous sodium bicarbonate solution (60 ml) was added for neutralization, and the mixed solution was extracted with ethyl acetate (30 ml × 2 times), and the organic phases were combined, washed with saturated brine (20 ml), then dried over anhydrous sodium sulfate, filtered, and finally concentrated under reduced pressure. The crude product was purified by silica gel column chromatography to obtain 4-bromo-7-fluoroisoquinoline-6-carbonitrile (340 mg).

MS (ESI) M/Z: 251.0 [M+H] ⁺ .

Step C: 4-Bromo-7-fluoroisoquinoline-6-carbonitrile (340 mg, 1.36 mmol) and dibutyltin oxide (507.96 mg, 2.04 mmol) were dissolved in toluene (12 ml), and trimethylsilyl azide (783.43 mg, 6.8 mmol) was added and reacted at 130 °C for 5 hours.

After the disappearance of the starting material as monitored by TLC, the mixture was concentrated under reduced pressure and the crude product was purified by silica gel column chromatography to give 4-bromo-7-fluoro-6-(2H-tetrazol-5-yl)isoquinoline (300.0 mg).

MS (ESI) M/Z: 293.8 [M+H] ⁺ .

Step D: 4-bromo-7-fluoro-6-(2H-tetrazolyl-5-yl)isoquinoline (100 mg, 0.34 mmol) and potassium carbonate (70.65 mg, 0.51 mmol) were dissolved in DMF (6 ml), and iodomethane (72.7 mg, 0.51 mmol) was added. The reaction mixture was reacted at room temperature overnight.

After the disappearance of the starting material by TLC monitoring, water (20 ml) was added, and the mixture was extracted with ethyl acetate (30 ml × 2 times). The organic phases were combined, washed with saturated brine (20 ml), then dried over anhydrous sodium sulfate, filtered, and finally concentrated under reduced pressure. The crude product was purified by silica gel column chromatography to give 4-bromo-7-fluoro-6-(2-methyl-2H-tetrazol-5-yl)isoquinoline (80 mg).

MS (ESI) M/Z: 307.91 [M+H] ⁺ .

¹ H NMR (400 MHz, DMSO-d ₆ ) δ 9.37 (s, 1H), 8.85–8.74 (m, 2H), 8.31 (d, J=10.7 Hz, 1H), 4.54 (s, 3H).

Step E: 4-bromo-7-fluoro-6-(2-methyl-2H-tetrazol-5-yl)isoquinoline (80 mg, 0.26 mmol) and N,N-dimethyl-2-(4-(4,4,5,5-tetramethyl-1,3,2-dioxaborolan-2-yl)-1H-pyrazol-1-yl)acetamide (120.26 mg, 0.34 mmol) were dissolved in dioxane (8 ml) and water (2 ml). Potassium carbonate (71.92 mg, 0.52 mmol) and 1,1'-bis(diphenylphosphinoferrocene)palladium dichloride (38.1 mg, 0.052 mmol) were added. The reaction liquid was replaced with nitrogen three times and reacted at 105 degrees Celsius for 3 hours.

After the disappearance of the starting material by TLC monitoring, water (20 ml) was added, and the mixture was extracted with ethyl acetate (30 ml × 2 times). The organic phases were combined, washed with saturated brine (20 ml), then dried over anhydrous sodium sulfate, filtered, and finally concentrated under reduced pressure. The crude product was prepared and purified to give 2-(4-(4-(7-fluoro-6-(2-methyl-2H-tetrazol-5-yl)isoquinolin-4-yl)phenyl)-1H-pyrazol-1-yl)-N,N-dimethylacetamide (32.37 mg).

MS (ESI) M/Z: 456.8 [M+H] ⁺ .

¹ H NMR (400 MHz, DMSO-d ₆ ) δ 9.42 (s, 1H), 8.65 (d, J = 6.8 Hz, 1H), 8.56 (s, 1H), 8.31 (d, J = 10.9 Hz, 1H), 8.23 (s, 1H), 8.03 (s, 1H), 7.83 (d, J = 8.0 Hz, 2H), 7.60 (d, J = 8.2 Hz, 2H), 5.18 (s, 2H), 4.47 (s, 3H), 3.07 (s, 3H), 2.89 (s, 3H).

Embodiment 53:

2-(4-(4-(7-fluoroisoquinolin-4-yl)phenyl)-1H-pyrazol-1-yl)-1-(4-isopropylpiperazin-1-yl)ethan-1-one

Referring to the synthesis method of Example 22, 2-(4-(4-(7-fluoroisoquinolin-4-yl)phenyl)-1H-pyrazol-1-yl)-1-(4-isopropylpiperazin-1-yl)ethan-1-one (69.04 mg) was obtained.

MS (ESI) M/Z: 458.2 [M + H] ⁺ .

¹ H NMR (400 MHz, DMSO-d ₆ ) δ 9.34 (s, 1H), 8.48 (s, 1H), 8.19 (s, 1H), 8.06–7.95 (m, 3H), 7.82–7.68 (m, 3H), 7.59–7.52 (m, 2H), 5.21 (s, 2H), 3.60-3.33 (m, 4H), 2.58-2.27 (m, 5H), 1.08–0.91 (m, 6H). C ₂₇ H ₂₈ FN ₅ O.

Embodiment 54:

2-(4-(4-(7-fluoroisoquinolin-4-yl)phenyl)-1H-pyrazol-1-yl)-1-(4-(2,2,2-trifluoroethyl)piperazin-1-yl)ethan-1-one

Referring to the synthesis method of Example 22, 2-(4-(4-(7-fluoroisoquinolin-4-yl)phenyl)-1H-pyrazol-1-yl)-1-(4-(2,2,2-trifluoroethyl)piperazin-1-yl)ethan-1-one (18.21 mg) was obtained.

MS (ESI) M/Z: 498.2 [M+H] ⁺ .

¹ H NMR (400 MHz, DMSO-d ₆ ) δ 9.34 (s, 1H), 8.48 (s, 1H), 8.21 (s, 1H), 8.06 (dd, J=9.3, 2.7 Hz, 1H), 8.04–7.95 (m, 2H), 7.82–7.68 (m, 3H), 7.59–7.52 (m, 2H), 5.21 (s, 2H), 3.56–3.46 (m, 4H), 3.27 (d, J=10.2 Hz, 2H), 2.80–2.56 (m, 4H).

Embodiment 55:

2-(4-(4-(7-chloro-6-(5-methyl-1,3,4-oxadiazol-2-yl)isoquinolin-4-yl)phenyl)-1H-pyrazol-1-yl)-1-(pyrrolidin-1-yl)ethan-1-one

Steps:

Step A: 2-(4-bromo-7-chloroisoquinolin-6-yl)-5-methyl-1,3,4-oxadiazole (50 mg, 0.155 mmol) and 1-(pyrrolidin-1-yl)-2-(4-(4,4,5,5-tetramethyl-1,3,2-dioxaborolan-2-yl)phenyl)-1H-pyrazol-1-yl)ethan-1-one (70.91 mg, 0.186 mmol) were dissolved in dioxane (4 ml)/water (1 ml), and potassium carbonate (42.78 mg, 0.31 mmol) and 1,1'-bis(diphenylphosphinoferrocene)palladium dichloride (11.7 mg, 0.016 mmol) were added. The reaction solution was replaced with nitrogen and refluxed at 105 degrees Celsius for 3 hours.

After the disappearance of the starting material by TLC monitoring, water (20 ml) was added, and the mixture was extracted with ethyl acetate (30 ml × 2 times). The organic phases were combined, washed with saturated brine (20 ml), dried over anhydrous sodium sulfate, filtered, and finally concentrated under reduced pressure to obtain a crude product to prepare 2-(4-(4-(7-chloro-6-(5-methyl-1,3,4-oxadiazol-2-yl)isoquinolin-4-yl)phenyl)-1H-pyrazol-1-yl)-1-(pyrrolidin-1-yl)ethan-1-one (18.88 mg).

MS (ESI) M/Z: 499.1 [M+H] ⁺ .

¹ H NMR (400 MHz, DMSO-d ₆ ) δ 9.46 (s, 1H), 8.65 (d, J = 13.6 Hz, 2H), 8.47 (s, 1H), 8.23 (s, 1H), 8.02 (s, 1H), 7.86–7.78 (m, 2H), 7.64–7.57 (m, 2H), 5.09 (s, 2H), 3.52 (t, J = 6.8 Hz, 2H), 3.34 (t, J = 6.9 Hz, 2H), 2.60 (s, 3H), 1.97–1.89 (m, 2H), 1.85–1.76 (m, 2H).

Embodiment 56:

2-(4-(4-(7-fluoro-6-(5-methyl-1,3,4-oxadiazol-2-yl)isoquinolin-4-yl-1-deuterio)phenyl)-1H-pyrazol-1-yl)-N,N-dimethylacetamide

Steps:

Step A: Dissolve 2-(7-fluoroisoquinolin-6-yl)-5-methyl-1,3,4-oxadiazole (2.38 g, 10.38 mmol) in dichloromethane (30 ml), add 85% m-chloroperbenzoic acid (5.38 g, 31.14 mmol), and stir the reaction solution at room temperature overnight.

After TLC monitoring showed that the starting material disappeared, 1M sodium hydroxide aqueous solution (50 ml) was added, stirred, and the liquids were separated. The organic phase was washed with saturated brine (10 ml), dried over anhydrous sodium sulfate, and concentrated to dryness under reduced pressure. The resulting residue, crude 7-fluoro-6-(5-methyl-1,3,4-oxadiazol-2-yl)isoquinoline 2-oxide, was directly used in the next step.

MS (ESI) M/Z: 246.0 [M+H] ⁺ .

Step B: Dissolve the crude product of (7-fluoro-6-(5-methyl-1,3,4-oxadiazol-2-yl)isoquinoline 2-oxide in chloroform (20 ml), add phosphorus oxychloride (3.75 g, 24.48 mmol), heat the reaction solution to 70 °C, and react for 2 hours.

After TLC monitoring shows that the raw material disappears, cool it. Add saturated sodium bicarbonate aqueous solution (100 ml) to quench, and extract with dichloromethane (100 ml). The organic phase is washed with saturated brine (40 ml), dried over anhydrous sodium sulfate, and concentrated to dryness under reduced pressure. The resulting residue is purified by silica gel column chromatography to obtain 2-(1-chloro-7-fluoroisoquinolin-6-yl)-5-methyl-1,3,4-oxadiazole (1.2 g).

MS (ESI) M/Z: 264.1 [M+H] ⁺ .

Step C: Dissolve 2-(1-chloro-7-fluoroisoquinolin-6-yl)-5-methyl-1,3,4-oxadiazole (1.2 g, 4.56 mmol) in anhydrous tetrahydrofuran (10 ml), add N,N,N',N'-tetramethylethylenediamine (795 mg, 6.84 mmol), [1,1'-bis(diphenylphosphino)ferrocene]dichloropalladium (33.38 mg, 0.046 mmol) and sodium deuterated borohydride (406.76 mg, 9.72 mmol). Replace the reaction liquid with nitrogen and stir at room temperature for 3 hours.

After TLC monitoring showed that most of the starting materials disappeared, saturated sodium bicarbonate aqueous solution (50 ml) and ethyl acetate (50 ml) were added, stirred, separated, and the organic phase was washed with saturated brine (30 ml), dried over anhydrous sodium sulfate, and concentrated to dryness under reduced pressure. The resulting residue was purified by silica gel column chromatography to obtain 2-(7-fluoroisoquinolin-6-yl-1-deuterio)-5-methyl-1,3,4-oxadiazole (700 mg).

MS (ESI) M/Z: 231.0 [M+H] ⁺ .

Step D: 2-(7-Fluoroisoquinolin-6-yl-1-deuterium)-5-methyl-1,3,4-oxadiazole (700 mg, 3.04 mmol) was dissolved in acetic acid (10 ml), heated to 110 °C, NBS (650 mg, 3.65 mmol) was added in portions, and the reaction solution was stirred at 110 °C for 3 hours.

After TLC monitoring shows that the raw material disappears, cool the mixture. Add saturated sodium bicarbonate aqueous solution (200 ml) and ethyl acetate (100 ml), stir and separate. Wash the organic phase with saturated brine (30 ml), dry over anhydrous sodium sulfate, and concentrate under reduced pressure. The resulting residue is purified by silica gel column chromatography to obtain 2-(4-bromo-7-fluoroisoquinolin-6-yl-1-deuterium)-5-methyl-1,3,4-oxadiazole (490 mg).

MS (ESI) M/Z: 309.1 [M+H] ⁺ .

Step E: 2-(4-bromo-7-fluoroisoquinolin-6-yl-1-deuterium)-5-methyl-1,3,4-oxadiazole (200 mg, 0.65 mmol), N,N-dimethyl-2-(4-(4-(4,4,5,5-tetramethyl-1,3,2-dioxaborolan-2-yl)phenyl)-1H-pyrazol-1-yl)acetamide (230.5 mg, 0.65 mmol), potassium carbonate (179 mg, 1.3 mmol) and [1,1'-bis(diphenylphosphino)ferrocene]palladium dichloride (47 mg, 0.065 mmol) were added to N,N-dimethylformamide (8 ml) and water (2 ml), replaced with nitrogen three times, and heated to 110 °C for 3 hours.

After TLC monitoring showed that the raw material disappeared, the mixture was cooled. Water (30 ml) and ethyl acetate (50 ml) were added and stirred to separate the liquids. The organic phase was washed with saturated brine (10 ml), dried over anhydrous sodium sulfate, and concentrated under reduced pressure. The resulting residue was purified by silica gel column chromatography to obtain 2-(4-(4-(7-fluoro-6-(5-methyl-1,3,4-oxadiazol-2-yl)isoquinolin-4-yl-1-deuterium)phenyl)-1H-pyrazol-1-yl)-N,N-dimethylacetamide (120.37 mg).

MS (ESI) M/Z: 458.3 [M + H] ⁺ .

¹ H NMR (400 MHz, DMSO-d ₆ ) δ 8.59 (s, 1H), 8.54 (d, J = 6.7 Hz, 1H), 8.34 (d, J = 11.0 Hz, 1H), 8.22 (s, 1H), 8.03 (s, 1H), 7.88–7.79 (m, 2H), 7.64–7.55 (m, 2H), 5.18 (s, 2H), 3.07 (s, 3H), 2.88 (s, 3H), 2.60 (s, 3H).

Embodiment 57:

2-(4-(4-(8-fluoro-6-(5-methyl-1,3,4-oxadiazol-2-yl)isoquinolin-4-yl-1-deuterio)phenyl)-1H-pyrazol-1-yl)-N,N-dimethylacetamide

Steps:

Step A: Dissolve 2-(8-fluoroisoquinolin-6-yl)-5-methyl-1,3,4-oxadiazole (650 mg, 2.84 mmol) in dichloromethane (7 ml), add 85% m-chloroperbenzoic acid (1.47 g, 8.51 mmol), and stir the reaction solution at room temperature overnight.

After TLC monitoring showed that the starting material disappeared, 1M sodium hydroxide aqueous solution (10 ml) was added, stirred, and the liquids were separated. The organic phase was washed with saturated brine (10 ml), dried over anhydrous sodium sulfate, and concentrated to dryness under reduced pressure. The resulting residue, crude 8-fluoro-6-(5-methyl-1,3,4-oxadiazol-2-yl)isoquinoline 2-oxide, was directly used in the next step.

MS (ESI) M/Z: 246.3 [M + H] ⁺ .

Step B: Dissolve the crude 8-fluoro-6-(5-methyl-1,3,4-oxadiazol-2-yl)isoquinoline 2-oxide obtained in the previous step in chloroform (15 ml), add phosphorus oxychloride (1.38 g, 9 mmol), and heat the reaction solution to 70°C for 2 hours.

After TLC monitoring shows that the raw material disappears, cool it. Add saturated sodium bicarbonate aqueous solution (50 ml) to quench, and extract with dichloromethane (50 ml). The organic phase is washed with saturated brine (20 ml), dried over anhydrous sodium sulfate, and concentrated to dryness under reduced pressure. The resulting residue is purified by silica gel column chromatography to obtain 2-(1-chloro-8-fluoroisoquinolin-6-yl)-5-methyl-1,3,4-oxadiazole (170 mg).

MS (ESI) M/Z: 264.1 [M+H] ⁺ .

Step C: Dissolve 2-(1-chloro-8-fluoroisoquinolin-6-yl)-5-methyl-1,3,4-oxadiazole (170 mg, 0.65 mmol) in anhydrous tetrahydrofuran (5 ml), add N,N,N',N'-tetramethylethylenediamine (112.7 mg, 0.97 mmol), [1,1'-bis(diphenylphosphino)ferrocene]dichloropalladium (47.6 mg, 0.0065 mmol) and sodium deuterated borohydride (57.6 mg, 1.38 mmol). Replace the reaction liquid with nitrogen and stir at room temperature for 3 hours.

After TLC monitoring showed that most of the starting materials disappeared, saturated sodium bicarbonate aqueous solution (10 ml) and ethyl acetate (20 ml) were added, stirred, separated, and the organic phase was washed with saturated brine (10 ml), dried over anhydrous sodium sulfate, and concentrated to dryness under reduced pressure. The resulting residue was purified by silica gel column chromatography to obtain 2-(8-fluoroisoquinolin-6-yl-1-deuterium)-5-methyl-1,3,4-oxadiazole (94 mg).

Step D: Dissolve 2-(8-fluoroisoquinolin-6-yl-1-d)-5-methyl-1,3,4-oxadiazole (94 mg, 0.4 mmol) in acetic acid (3 ml), heat to 110 °C, add NBS (73.7 mg, 0.41 mmol) in portions, and stir the reaction at 110 °C for 3 hours.

After TLC monitoring showed that most of the raw materials disappeared, the mixture was cooled. Saturated aqueous sodium bicarbonate solution (40 ml) and ethyl acetate (30 ml) were added, stirred, and separated. The organic phase was washed with saturated brine (10 ml), dried over anhydrous sodium sulfate, and concentrated to dryness under reduced pressure. The obtained residue was purified by silica gel column chromatography to obtain 2-(4-bromo-8-fluoroisoquinolin-6-yl-1-deuterium)-5-methyl-1,3,4-oxadiazole (58 mg).

MS (ESI) M/Z: 309.2 [M+H] ⁺ .

Step E: 2-(4-bromo-8-fluoroisoquinolin-6-yl-1-deuterio)-5-methyl-1,3,4-oxadiazole (58 mg, 0.19 mmol), N,N-dimethyl-2-(4-(4-(4,4,5,5-tetramethyl-1,3,2-dioxaborolan-2-yl)phenyl)-1H-pyrazol-1-yl)acetamide (67 mg, 0.19 mmol), potassium carbonate (52.44 mg, 0.19 mmol) were added to the mixture. mg, 0.38 mmol) and [1,1'-bis(diphenylphosphino)ferrocene]palladium dichloride (15.5 mg, 0.019 mmol) were added to N,N-dimethylformamide (4 ml) and water (1 ml), the atmosphere was replaced with nitrogen three times, and the mixture was heated to 110 °C for 3 hours.

After TLC monitoring showed that the raw material disappeared, the mixture was cooled. Water (20 ml) and ethyl acetate (30 ml) were added and stirred to separate the liquids. The organic phase was washed with saturated brine (10 ml), dried over anhydrous sodium sulfate, and concentrated under reduced pressure. The obtained residue was prepared and purified to give 2-(4-(4-(8-fluoro-6-(5-methyl-1,3,4-oxadiazol-2-yl)isoquinolin-4-yl-1-deuterium)phenyl)-1H-pyrazol-1-yl)-N,N-dimethylacetamide (67.7 mg).

MS (ESI) M/Z: 458.3 [M + H] ⁺ .

¹ H NMR (400 MHz, DMSO-d ₆ ) δ 8.74 (s, 1H), 8.25 (d, J = 18.0 Hz, 2H), 8.09-8.00 (m, 2H), 7.85 (d, J = 7.9 Hz, 2H), 7.62 (d, J = 8.0 Hz, 2H), 5.18 (s, 2H), 3.07 (s, 3H), 2.88 (s, 3H), 2.59 (s, 3H).

Embodiment 58:

2-(4-(4-(1-chloro-7-fluoro-6-(5-methyl-1,3,4-oxadiazol-2-yl)isoquinolin-4-yl)phenyl)-1H-pyrazol-1-yl)-N,N-dimethylacetamide

Steps:

Step A: Dissolve the compound 2-(4-(4-(7-fluoro-6-(5-methyl-1,3,4-oxadiazol-2-yl)isoquinolin-4-yl)phenyl)-1H-pyrazol-1-yl)-N,N-dimethylacetamide (1.08 g, 2.37 mmol) obtained in Example 32 in dichloromethane (20 ml), add m-chloroperbenzoic acid (1.23 g, 7.11 mmol) and react at room temperature overnight.

After TLC monitoring showed that the starting material disappeared, sodium hydroxide aqueous solution was added to adjust the pH to neutral, dichloromethane (20 ml) was added, stirred, and the liquids were separated. The organic phase was first washed with saturated brine (10 ml), then dried over anhydrous sodium sulfate, filtered, and finally concentrated under reduced pressure to obtain 4-(4-(1-(2-(N,N-dimethylamino)-2-oxoethyl)-1H-pyrazol-4-yl)phenyl)-7-fluoro-6-(5-methyl-1,3,4-oxadiazol-2-yl)isoquinoline 2-oxide (1.12 g).

MS (ESI) M/Z: 473.1 [M+H] ⁺ .

Step B: Dissolve 4-(4-(1-(2-(N,N-dimethylamino)-2-oxoethyl)-1H-pyrazol-4-yl)phenyl)-7-fluoro-6-(5-methyl-1,3,4-oxadiazol-2-yl)isoquinoline 2-oxide (1.12 g, 2.37 mmol) in chloroform (13 ml), add phosphorus oxychloride (1.09 g, 7.12 mmol) dropwise, and reflux at 70 °C for 2 hours.

After TLC monitoring showed that the raw material disappeared, the mixture was cooled to room temperature. After adding sodium bicarbonate aqueous solution to adjust the pH to neutral, the mixture was extracted with dichloromethane (20 ml × 2). The organic phases were combined, washed with saturated brine (20 ml), dried over anhydrous sodium sulfate, filtered, and finally concentrated under reduced pressure. The crude product was prepared and purified to obtain 2-(4-(4-(1-chloro-7-fluoro-6-(5-methyl-1,3,4-oxadiazol-2-yl)isoquinolin-4-yl)phenyl)-1H-pyrazol-1-yl)-N,N-dimethylacetamide (750 mg).

MS (ESI) M/Z: 491.2 [M+H] ⁺ .

¹ H NMR (400 MHz, DMSO-d ₆ ) δ 8.58 (d, J = 6.7 Hz, 1H), 8.41 (s, 1H), 8.36 (d, J = 11.6 Hz, 1H), 8.23 (s, 1H), 8.03 (s, 1H), 7.84 (d, J = 7.9 Hz, 2H), 7.61 (d, J = 7.9 Hz, 2H), 5.17 (s, 2H), 3.07 (s, 3H), 2.88 (s, 3H), 2.61 (s, 3H). C ₂₅ H ₂₀ ClFN ₆ O ₂ .

Embodiment 59:

2-(4-(4-(7-fluoro-1-methyl-6-(5-methyl-1,3,4-oxadiazol-2-yl)isoquinolin-4-yl)phenyl)-1H-pyrazol-1-yl)-N,N-dimethylacetamide

Steps:

Step A: The compound 2-(4-(4-(1-chloro-7-fluoro-6-(5-methyl-1,3,4-oxadiazol-2-yl)isoquinolin-4-yl)phenyl)-1H-pyrazol-1-yl)-N,N-dimethylacetamide (100 mg, 0.204 mmol) obtained in Example 58 was dissolved in dioxane/water (2 ml/0.2 ml), and methylboric acid (37 mg, 0.612 mmol), potassium carbonate (85 mg, 0.612 mmol) and [1,1'-bis(diphenylphosphino)ferrocene]dichloropalladium dichloromethane complex (8 mg, 0.0102 mmol) were added. After nitrogen replacement three times, the reaction was carried out at 100°C overnight.

After TLC monitoring showed that the starting material disappeared, water (20 ml) and ethyl acetate (15 ml) were added, stirred, and separated. The organic phase was first washed with saturated brine (10 ml), then dried over anhydrous sodium sulfate, filtered, and finally concentrated under reduced pressure. The crude product was prepared and purified to obtain 2-(4-(4-(7-fluoro-1-methyl-6-(5-methyl-1,3,4-oxadiazol-2-yl)isoquinolin-4-yl)phenyl)-1H-pyrazol-1-yl)-N,N-dimethylacetamide (33.28 mg).

MS (ESI) M/Z: 471.4 [M+H] ⁺ .

¹ H NMR (400 MHz, DMSO-d ₆ ) δ 8.53 (d, J = 7.2 Hz, 1H), 8.42 (s, 1H), 8.36 (d, J = 12 Hz, 1H), 8.21 (s, 1H), 8.01 (s, 1H), 7.81 (d, J = 7.9 Hz, 2H), 7.56 (d, J = 7.9 Hz, 2H), 5.17 (s, 2H), 3.07 (s, 3H), 2.97 (s, 3H), 2.88 (s, 3H), 2.60 (s, 3H).

Embodiment 60:

2-(4-(4-(1-amino-7-fluoro-6-(5-methyl-1,3,4-oxadiazol-2-yl)isoquinolin-4-yl)phenyl)-1H-pyrazol-1-yl)-N,N-dimethylacetamide

Steps:

Step A: Example 58 compound 2-(4-(4-(1-chloro-7-fluoro-6-(5-methyl-1,3,4-oxadiazol-2-yl)isoquinolin-4-yl)phenyl)-1H-pyrazol-1-yl)-N,N-dimethylacetamide (100 mg, 0.204 mmol) was dissolved in dioxane (2 ml), and diphenylmethylamine (74 mg, 0.408 mmol), 1,1'-binaphthyl-2,2'-bisdiphenylphosphine (12 mg, 0.0204 mmol), cesium carbonate (133 mg, 0.408 mmol), tris(dibenzylideneacetone)dipalladium (10 mg, 0.0102 mmol) were added, the atmosphere was replaced with nitrogen three times, and the mixture was refluxed at 90°C overnight.

After TLC monitoring showed that the raw material disappeared, the mixture was cooled to room temperature, and water (20 ml) was added and extracted with ethyl acetate (20 ml x 2). The organic phases were combined, washed with saturated brine (20 ml), dried over anhydrous sodium sulfate, filtered, and finally concentrated under reduced pressure to obtain 2-(4-(4-(1-(diphenylamine))-1-nitropropene The crude product of (4-(5-methyl)amino)-7-fluoro-6-(5-methyl-1,3,4-oxadiazol-2-yl)isoquinolin-4-yl)phenyl)-1H-pyrazol-1-yl)-N,N-dimethylacetamide was used directly in the next step (130 mg).

MS (ESI) M/Z: 635.2 [M + H] ⁺ .

Step B: Dissolve the crude product of 2-(4-(4-(1-(diphenylmethylene)amino)-7-fluoro-6-(5-methyl-1,3,4-oxadiazol-2-yl)isoquinolin-4-yl)phenyl)-1H-pyrazol-1-yl)-N,N-dimethylacetamide (130 mg, 0.204 mmol) in anhydrous methanol (2 ml), add sodium acetate (50 mg, 0.612 mmol) and hydroxylamine hydrochloride (28 mg, 0.408 mmol), and react at room temperature for 1.5 h.

After TLC monitoring showed that the starting material disappeared, sodium hydroxide aqueous solution was added to adjust the pH to neutral, and dichloromethane (20 ml x 2) was used for extraction. The organic phases were combined, washed with saturated brine (20 ml), dried over anhydrous sodium sulfate, filtered, and finally concentrated under reduced pressure. The crude product was prepared and purified to give 2-(4-(4-(1-amino-7-fluoro-6-(5-methyl-1,3,4-oxadiazol-2-yl)isoquinolin-4-yl)phenyl)-1H-pyrazol-1-yl)-N,N-dimethylacetamide (36.12 mg).

MS (ESI) M/Z: 472.4 [M+H] ⁺ .

¹ H NMR (400 MHz, DMSO-d ₆ ) δ 8.43–8.34 (m, 2H), 8.16 (s, 1H), 7.97 (s, 1H), 7.87 (s, 1H), 7.73 (d, J=8.0 Hz, 2H), 7.47 (d, J=7.9 Hz, 2H), 7.13 (s, 2H), 5.15 (s, 2H), 3.06 (s, 3H), 2.88 (s, 3H), 2.58 (s, 3H).

Example 61:

2-(4-(4-(7-fluoro-6-(5-methyl-1,3,4-thiadiazol-2-yl)isoquinolin-4-yl)phenyl)-1H-pyrazol-1-yl)-N,N-dimethylacetamide

Steps:

Step A: Dissolve 6-bromo-7-fluoroisoquinoline (500 mg, 2.22 mmol) in dioxane (15 ml), add bis-pinacol borate (620.74 mg, 2.45 mmol), potassium acetate (595.38 mg, 6.1 mmol), 1,1'-bis(diphenylphosphinoferrocene)palladium dichloride (295.65 mg, 0.41 mmol), replace nitrogen with vacuum, and react at 90 degrees Celsius for 2 hours.

TLC monitoring showed that the raw material was not completely reacted. The reaction mixture was cooled and used directly in the next step.

MS (ESI) M/Z: 192.3 [M + H] ⁺ .

Step B: Add 2-bromo-5-methyl-1,3,4-thiadiazole (360 mg, 2.022 mmol), potassium carbonate (837.2 mg, 6.1 mmol), 1,1'-bis(diphenylphosphino)ferrocenepalladium dichloride (295.65 mg, 0.41 mmol) to the reaction solution of the previous step, replace nitrogen with vacuum, and react at 90 degrees Celsius for 4 hours.

After TLC monitoring showed that the raw material disappeared, water (20 ml) was added to the mixture after cooling, and then extracted with ethyl acetate (20 ml x 2). The organic phases were combined, washed with saturated brine (20 ml), dried over anhydrous sodium sulfate, filtered, and finally concentrated under reduced pressure. The crude product was purified by silica gel column chromatography (eluent: petroleum ether: ethyl acetate = 1:1) to obtain 2-(7-fluoroisoquinolin-6-yl)-5-methyl-1,3,4-thiadiazole (186 mg).

MS (ESI) M/Z: 246.1 [M+H] ⁺ .

Step C: Dissolve 2-(7-fluoroisoquinolin-6-yl)-5-methyl-1,3,4-thiadiazole (186 mg, 0.76 mmol) in acetic acid (8 ml), add N-bromosuccinimide (148.63 mg, 0.84 mmol), and react at 110 °C for 3 hours.

After TLC monitoring showed that a small amount of raw materials remained, the mixture was cooled and neutralized with aqueous sodium bicarbonate solution, and extracted with ethyl acetate (20 ml x 2). The organic phases were combined, washed with saturated brine (20 ml), dried over anhydrous sodium sulfate, filtered, and finally concentrated under reduced pressure. The crude product was purified by silica gel column chromatography (eluent: petroleum ether: ethyl acetate = 4:1) to obtain 2-(4-bromo-7-fluoroisoquinolin-6-yl)-5-methyl-1,3,4-thiadiazole (40 mg).

MS (ESI) M/Z: 324.2 [M + H] ⁺ .

Step D: 2-(4-bromo-7-fluoroisoquinolin-6-yl)-5-methyl-1,3,4-thiadiazole (40 mg, 0.12 mmol), N,N-dimethyl-2-(4-(4-(4,4,5,5-tetramethyl-1,3,2-dioxaborolan-2-yl)phenyl)-1H-pyrazol-1-yl)acetamide (43.96 mg, 0.12 mmol), potassium carbonate (34.18 mg, 0.25 mmol), 1,1'-bis(diphenylphosphinoferrocene)palladium dichloride (9.05 mg, 0.012 mmol) were added to DMF (8 ml) and water (2 ml), the atmosphere was replaced with nitrogen under vacuum, and the reaction was carried out at 110 °C for 4 hours.

After TLC monitoring showed that the raw material disappeared, water (20 ml) was added after cooling, and ethyl acetate (20 ml x 2) was used for extraction. The organic phases were combined, washed with saturated brine (20 ml), dried over anhydrous sodium sulfate, filtered, and finally concentrated under reduced pressure. The crude product was prepared and purified to obtain 2-(4-(4-(7-fluoro-6-(5-methyl-1,3,4-thiadiazol-2-yl)isoquinolin-4-yl)phenyl)-1H-pyrazol-1-yl)-N,N-dimethylacetamide (9.47 mg, yellow solid, yield 16.20%).

MS (ESI) M/Z: 473.3 [M + H] ⁺ .

¹ H NMR (400 MHz, DMSO-d ₆ ) δ 9.43 (s, 1H), 8.81 (d, J = 6.7 Hz, 1H), 8.58 (s, 1H), 8.35 (d, J = 11.3 Hz, 1H), 8.22 (s, 1H), 8.02 (s, 1H), 7.83 (d, J = 7.8 Hz, 2H), 7.62 (d, J = 7.8 Hz, 2H), 5.17 (s, 2H), 3.07 (s, 3H), 2.89 (s, 3H), 2.82 (s, 3H).

Embodiment 62:

2-(4-(4-(7-fluoro-6-(pyridin-2-yl)isoquinolin-4-yl)phenyl)-1H-pyrazol-1-yl)-N,N-dimethylacetamide

By referring to the synthesis method of Example 61, 2-(4-(4-(7-fluoro-6-(pyridin-2-yl)isoquinolin-4-yl)phenyl)-1H-pyrazol-1-yl)-N,N-dimethylacetamide (43.56 mg) was obtained.

MS (ESI) M/Z: 452.4 [M + H] ⁺ .

¹ H NMR (400 MHz, DMSO-d ₆ ) δ 9.39 (s, 1H), 8.72 (d, J = 4.8 Hz, 1H), 8.54–8.44 (m, 2H), 8.26–8.17 (m, 2H), 8.02–7.88 (m, 3H), 7.79 (d, J = 7.9 Hz, 2H), 7.58 (d, J = 8.0 Hz, 2H), 7.49–7.42 (m, 1H), 5.16 (s, 2H), 3.07 (s, 3H), 2.88 (s, 3H).

Embodiment 63:

2-(4-(4-(7-fluoro-6-(pyrimidin-2-yl)isoquinolin-4-yl)phenyl)-1H-pyrazol-1-yl)-N,N-dimethylacetamide

By referring to the synthesis method of Example 61, 2-(4-(4-(7-fluoro-6-(pyrimidin-2-yl)isoquinolin-4-yl)phenyl)-1H-pyrazol-1-yl)-N,N-dimethylacetamide (66.36 mg) was obtained.

MS (ESI) M/Z: 453.2 [M + H] ⁺ .

¹ H NMR (400 MHz, DMSO-d ₆ ) δ 9.41 (s, 1H), 8.97 (d, J = 4.9 Hz, 2H), 8.62-8.51 (m, 2H), 8.21 (d, J = 11.2 Hz, 2H), 8.00 (s, 1H), 7.80 (d, J = 7.9 Hz, 2H), 7.62-7.52 (m, 3H), 5.16 (s, 2H), 3.07 (s, 3H), 2.88 (s, 3H).

Reference compound information:

Referring to the preparation method of WO2017202719A patent I-003, the following compounds were prepared:

2. Biological Activity Experiment

(I) CDK in vitro enzymatic assay

The inhibitory effect on the activities of CDK8/CycC/Med12 kinase and CDK19/CycC kinase was tested in vitro using the ADP-Glo Luminescent method, and the half-maximal inhibitory concentration IC ₅₀ of the compound on the activity of CDK8 kinase was obtained.

The inhibitory effect of the kinase activity of kinases CDK2/CycE1, CDK7/cyclinH/MAT1 and CDK9/cyclinT1 was tested in vitro using the time-difference fluorescence resonance energy transfer assay (LanceUltra), and the half-maximal inhibitory concentration IC50 of the compound on the kinase activity of CDK2/CDK7/CDK9 was obtained.

1. Experimental Materials

CDK8 was purified from Via Biotech, CDK19 was purchased from BPS Bioscience, substrate MBP was purchased from signalChem, ADP-Glo kit was purchased from Promega, DMSO was purchased from Sigma, and 384-well plates were purchased from Corning.

CDK2/CDK7/CDK9 were purchased from Carna, and substrate 18/substrate 8 were purchased from Gill Biochemicals.

2. Experimental methods

(1) Prepare 1× Kinase buffer; add FAM-labeled peptide and ATP to 1× Kinase buffer to make 2.5× Peptide solution.

(2) Preparation of compound concentration gradient: The test compound concentration starts at 10 μM, 3-fold dilution, 10 concentrations, and duplicate well detection. Dilute to 100 times the final concentration of 100% DMSO solution in the 384 source plate. Use the Echo 650 dispenser to transfer 50 nL of the 100 times final concentration of the compound to the destination plate 384-well plate. Add 200 nL DMSO to the positive and negative control wells.

(3) Prepare kinase solution by adding corresponding kinase into 1× Kinase buffer.

(4) Add 2.5 μL of kinase solution to the compound wells and positive control wells respectively; add 2.5 μL of 1× Kinase buffer to the negative control wells.

(5) Centrifuge at 1000 rpm for 30 seconds, shake the reaction plate to mix, and incubate at room temperature for 10 minutes.

(6) Add MBP-labeled peptide and ATP to 1× Kinase buffer to prepare a mixed solution of ATP and substrate at 2 times the final concentration.

(7) Add 2.5 μL or 10 μL of a mixed solution of ATP and substrate at twice the final concentration to start the reaction.

(8) The 384-well plate was centrifuged at 1000 rpm for 30 seconds, vortexed to mix, and then incubated at 37°C for 2 hours or 1 hour.

(9) Add 5 μL of ADP-Glo Reagent 1 to stop the kinase reaction, centrifuge at 1000 rpm for 30 seconds, shake to mix, and incubate at 37°C for 1 hour. Transfer 10 μL of ADP-Glo Reagent 2 to a 384-well plate; incubate the reaction at 37°C for 1 hour. Alternatively, add 20 μL of Stop Detection Solution to stop the kinase reaction, centrifuge at 1000 rpm for 30 seconds, shake to mix, and leave at room temperature for 1 hour.

(10) Read the RLU value using Envision 2104 Multilabel Reader.

Calculation formula:

Copy the RLU readings and convert the data to inhibition percentage using the formula: %inhibition = (max-Sample RLU)/(max-min)*100. Or copy the fluorescence readings (Lance signal ratio (665nm/615nm)); convert the data to inhibition percentage using the formula: %inhibition = (max-Sample Lance signal ratio)/(max-min)*100. Where: "min" is the reading of the control well without enzyme; "max" is the reading of the control well with DMSO added.

Fitting dose-effect curve

Import the data into MS Excel and use XLFit excel add-in version 5.4.0.8 for curve fitting; fitting formula: Y=Bottom+(Top-Bottom)/(1+(IC50/X)^HillSlope).

3. Experimental results

The IC ₅₀ data of the compounds disclosed herein on CDK kinase activity are shown in Table 1. It can be seen that the compounds disclosed herein have good CDK8/CDK19 kinase inhibitory activity; and some compounds disclosed herein have weak inhibitory activity on CDK2/7/9 kinases, and can selectively inhibit CDK8/CDK19 kinases.

Table 1

(II) Cell proliferation inhibition experiment

In this experiment, the method of measuring intracellular ATP content by fluorescence method (CellTiter-Glo) was used to detect the inhibitory effect of the compound on MV4-11 and KG-1 cell lines, and the half inhibitory concentration IC ₅₀ of the compound on the above cell lines was obtained.

1. Experimental Materials

RPMI-1640 medium, fetal bovine serum (FBS), 100X Pen/Strep, and GlutaMAX-I Supplement were purchased from GIBCO; MV4-11 and KG-1 cell lines were purchased from ATCC Cell Bank; Cell Titer-Glo luminescent cell viability detection reagent was purchased from Promega.

2. Experimental Methods

1) Inoculate cells into 384-well culture plates at a density of 300 MV4-11 or KG-1 cells per well, 100 μl per well, and incubate in an incubator (37° C., 5% CO ₂ ) overnight.

2) Day 0: D300e (TECAN) was used to add 50 nL of the compound to be tested in serial dilution (starting concentration was 30 μM, 10 concentrations, 1:3 dilution ratio) to the culture plate cells, the final DMSO concentration was 0.5%, and the culture plate was placed in a cell culture incubator for 168 hours (37° C., 5% CO ₂ ). For the blank control, 50 nL of DMSO was added to each well.

3) Day 7: Add 30 μL Cell Titer-Glo reagent to each well, shake at 500 rpm for 10 minutes, centrifuge at 1000 rpm for 1 minute, and incubate at room temperature in the dark for 20 minutes to stabilize the luminescent signal.

4) Envision microplate reader (PerkinElmer) was used to detect the luminescent signal.

5) GraphPad Prism 6 software was used for data analysis and calculation of IC ₅₀ of the compounds.

The inhibition results of some compounds disclosed in the present invention on MV4-11 and KG-1 cell lines are shown in Table 2. It can be seen that the compounds disclosed in the present invention have good tumor cell proliferation inhibition activity on MV4-11 cells and KG-1 cells.

Table 2

Biological test evaluation

Test Example 5: Pharmacokinetics of the compounds disclosed in this disclosure in mice

Mice were used as test animals to study the pharmacokinetic behavior of the disclosed compounds in mouse plasma after intravenous injection and oral injection. Plasma samples were collected at specific time points, and the compound concentration in plasma was detected by LC-MS/MS. PK parameters were calculated to reflect the pharmacokinetic behavior of the disclosed compounds in mouse plasma.

1. Experimental plan

1.1 Trial Drugs:

The present invention discloses compounds of Examples 11, 27, 34, 56, 57 and control compounds.

1.2 Experimental animals

Mouse, balb/c nude, female, supplier is Jiangsu Jicui Pharmaceutical Biotechnology Co., Ltd.

1.3 Administration

Information on mouse dosing of Examples 11, 27, 34, 56, 57 and control compounds: There were 3 mice in both IV (intravenous injection) and PO (oral) experimental groups. The IV dose for mice of Examples 11, 27, 34, 56, 57 and control compounds was 1 mg/kg, and the dosing volume was 5 mL/kg. The PO dose for mice of Examples 11, 27 and 34 was 10 mg/kg. The dose for mice of Examples 56 and 57 was 3 mg/kg, and the dosing volume was 10 mL/kg. The PO dose for mice of the control compound was 10 mg/kg, and the dosing volume was 10 mL/kg. The dosing solvent was 5 vol% DMSO/5 vol% Solutol/90 vol% Saline.

1.4 Experimental Equipment

The centrifuge and pipette were purchased from Eppendorf.

1.5 Sample collection

After administration to mice, 0.025 mL and 0.2 mL of venous blood were collected at 0.0833 (IV), 0.25, 0.5, 1, 2, 4, 8 and 24 hours, respectively, placed in EDTA-K2 tubes, centrifuged at 4°C, 2000 g for 10 min to separate plasma, and stored at -80°C.

1.6 Sample processing

Mouse plasma sample processing:

1) 15 μL of plasma sample was added to 200 μL of acetonitrile for precipitation, vortexed and centrifuged for 15 minutes.

2) The supernatant after treatment was diluted with water and the concentration of the test compound was analyzed by LC/MS/MS.

2. Experimental results

The pharmacokinetic parameters were calculated using WinNonlin 6.1. The pharmacokinetic parameters of intravenous, oral and intraperitoneal injections of drugs in mice are shown in Table 6 Wherein, C _max represents the maximum plasma concentration, CL represents the clearance rate, Vss represents the steady-state distribution volume, T _1/2 represents the terminal elimination half-life, MRT _Inf represents the mean residence time, AUC represents the area under the drug-time curve, and F represents the bioavailability.

Table 6 Pharmacokinetic parameters of some compounds of the present disclosure in mice after intravenous injection and oral administration

Note: “/” indicates not measured

Results: As can be seen from Table 6, the compounds of Examples 11, 27, 34, 56, and 57 of the present disclosure have higher exposure and better pharmacokinetic properties than the control compound.

Test Example 6: Pharmacokinetics of the compounds disclosed in the present invention in rats

Rats were used as test animals to study the pharmacokinetic behavior of the disclosed compounds in rat plasma after intravenous injection and oral injection. Plasma samples were collected at specific time points, and the concentration of the compound in plasma was detected by LC-MS/MS. PK parameters were calculated to reflect the pharmacokinetic behavior of the disclosed compounds in rat plasma.

1. Experimental plan

1.1 Trial Drugs:

The present invention discloses Examples 11, 27, 32, 34 and comparative compounds.

1.2 Experimental animals

Rats, Sprague-Dawley, male, were supplied by Zhejiang Weitong Lihua Experimental Animal Technology Co., Ltd.

1.3 Administration

The dosing information of Examples 11, 27, 32, 34 and control compounds in mice: There were 3 mice in both IV (intravenous injection) and PO (oral) experimental groups. The dosing information of Examples 11, 27, 32, 34 and control compounds in rats: The IV dose was 1 mg/kg, and the dosing volume was 5 mL/kg; the PO dose was 5 mg/kg, and the dosing volume was 10 mL/kg, and the dosing solvent was 5 vol% DMSO/30 vol% PEG400 (solubilizer)/65% Saline (physiological saline).

1.4 Experimental Equipment

The centrifuge and pipette were purchased from Eppendorf.

1.5 Sample collection

After administration to the rats, 0.025 mL and 0.2 mL of venous blood were collected at 0.0833 (IV), 0.25, 0.5, 1, 2, 4, 8 and 24 hours, respectively, placed in EDTA-K2 tubes, centrifuged at 4°C, 2000 g for 10 min to separate plasma, and stored at -80°C.

1.6 Sample processing

Rat plasma sample processing:

1) 50 μL of plasma sample was added with 200 μL of acetonitrile for precipitation, vortexed and centrifuged for 15 minutes.

2. Experimental results

The pharmacokinetic parameters were calculated using WinNonlin 6.1, and the pharmacokinetic parameters of drugs injected intravenously, orally and intraperitoneally in rats are shown in Table 7. Among them, _Cmax represents the maximum plasma concentration, CL represents the clearance rate, Vss represents the steady-state distribution volume, T1 _/2 represents the terminal elimination half-life, _MRTInf represents the mean residence time, AUC represents the area under the drug-time curve, and F represents the bioavailability.

Table 7 Pharmacokinetic parameters of some compounds of the present disclosure in rats after intravenous injection and oral administration

Note: “/” indicates not measured

Results: As can be seen from Table 7, the compounds of Examples 11, 27, 32, and 34 of the present disclosure have higher exposure, better bioavailability, and more excellent pharmacokinetic properties than the control compound.

Claims

The compound represented by formula (I), pharmaceutically acceptable salts and stereoisomers thereof,

in,

Structural units Selected from

Ring B is selected from phenyl, 5-6 membered heteroaryl, 5-6 membered heterocycloalkyl, C 5-6 cycloalkyl;

Ring C is selected from phenyl, 5-6 membered heterocycloalkyl, 5-6 membered heteroaryl;

Ra is independently selected from hydrogen, halogen, C1-4 alkyl, C1-4 alkoxy, CN, 5-6 membered heteroaryl; wherein the 5-6 membered heteroaryl may be optionally substituted by 1-3 Rab , wherein Rab is selected from halogen, C1-4 alkyl;

R b is each independently selected from hydrogen, halogen, C 1-4 alkyl, C 1-4 alkoxy;

R c is independently selected from hydrogen, C 1-4 alkyl, -C 0-4 alkyl CONR ca R cb ; wherein R ca , R cb are independently selected from hydrogen, C 1-4 alkyl, or R ca and R cb and the N atom to which they are commonly connected are cyclized to form a 5-6 membered heterocycloalkyl, a 5-11 membered heterocyclyl, a 5-11 membered spiro heterocycloalkyl; wherein the 5-6 membered heterocycloalkyl, the 5-11 membered heterocyclyl, the 5-11 membered spiro heterocycloalkyl can be optionally substituted with 1-3 C 1-4 alkyl, halogen, C 1-4 haloalkyl, wherein the C 1-4 alkyl can be optionally substituted with one or more of the following groups: hydroxyl, amino, cyano;

R d is selected from H, NH 2 , OH, D, halogen, C 1-6 alkyl;

n is selected from 0, 1, 2, 3;

m is selected from 0, 1, 2, 3;

p is selected from 0, 1, 2, 3;

Wherein, formula (I) is not the following compound:
The compound of formula (I), pharmaceutically acceptable salt and stereoisomer thereof as claimed in claim 1,

in,

Structural units Selected from

Ring B is selected from phenyl;

Ring C is selected from 5-6 membered heteroaryl;

Ra is independently selected from hydrogen, halogen, C1-4 alkyl, C1-4 alkoxy, CN, 5-6 membered heteroaryl; wherein the 5-6 membered heteroaryl may be optionally substituted by 1-3 Rab , wherein Rab is selected from halogen, C1-4 alkyl;

R b is selected from hydrogen;

R c is selected from -C 1-4 alkyl CONR ca R cb ; wherein R ca and R cb are each independently selected from hydrogen, C 1-4 alkyl, or R ca and R cb and the N atom to which they are commonly connected are cyclized to form a 5-6 membered heterocycloalkyl, a 5-11 membered heterocyclyl, or a 5-11 membered spiro heterocycloalkyl; wherein the 5-6 membered heterocycloalkyl, the 5-11 membered heterocyclyl, or the 5-11 membered spiro heterocycloalkyl may be optionally substituted with 1-3 C 1-4 alkyl, halogen, or C 1-4 haloalkyl, wherein the C 1-4 alkyl may be optionally substituted with one or more of the following groups: hydroxyl, amino, or cyano;

R d is selected from H, NH 2 , OH, D, halogen, C 1-4 alkyl;

n is selected from 1, 2, and 3;

m is selected from 1, 2, and 3.
The compound of formula (I), pharmaceutically acceptable salt and stereoisomer thereof according to any one of claims 1 to 2, characterized in that Ra is independently selected from H, F, Cl, -CH3 , -CN, CH3O- ,
The compound of formula (I), pharmaceutically acceptable salt and stereoisomer thereof according to any one of claims 1 to 3, characterized in that Ra is independently selected from H, F,
The compound of formula (I), pharmaceutically acceptable salt and stereoisomer thereof according to any one of claims 1 to 4, characterized in that R c is independently selected from
The compound of formula (I), pharmaceutically acceptable salt and stereoisomer thereof according to any one of claims 1 to 5, characterized in that the structural unit Selected from
The compound of formula (I), pharmaceutically acceptable salt and stereoisomer thereof according to any one of claims 1 to 6, characterized in that the structural unit Selected from Wherein, Ra , Rd , and n are as defined in claims 1 to 6; preferably, the structural unit Selected from wherein Ra and Rd are as defined in claims 1 to 6; more preferably, the structural unit Selected from
The compound of formula (I), pharmaceutically acceptable salt and stereoisomer thereof according to any one of claims 1 to 7, characterized in that ring B is selected from phenyl, pyridyl, pyrimidinyl, pyrazinyl, piperazinyl and piperidinyl.
The compound of formula (I), pharmaceutically acceptable salt and stereoisomer thereof according to claim 8, characterized in that ring B is selected from
The compound represented by formula (I) according to any one of claims 1 to 9, the pharmaceutically acceptable salt and the stereoisomer thereof, wherein The characteristic is that the structural unit Selected from wherein R c and m are as defined in any one of claims 1-9.
The compound represented by formula (I), pharmaceutically acceptable salt and stereoisomer thereof according to claim 10, characterized in that the structural unit Selected from wherein R c is as defined in claim 10.
The compound of formula (I), pharmaceutically acceptable salt and stereoisomer thereof according to claim 10, characterized in that the structural unit Selected from
The compound of formula (I), pharmaceutically acceptable salt and stereoisomer thereof according to any one of claims 1 to 12, characterized in that it is selected from the compounds represented by formula (II), (II-a), (II-b) and (II-c), pharmaceutically acceptable salt and stereoisomer thereof,

wherein ring B, ring C, Ra, Rb , Rc , Rd , n, m, and p are as defined in any one of claims 1 to 12.
The compound of formula (I), pharmaceutically acceptable salt and stereoisomer thereof according to any one of claims 1 to 12, characterized in that it is selected from the compounds represented by formula (III-A) and (III-B), pharmaceutically acceptable salt and stereoisomer thereof,

wherein Ra , Rc and Rd are as defined in any one of claims 1-12.
Compounds, pharmaceutically acceptable salts and stereoisomers thereof, characterized in that they are selected from,
A pharmaceutical composition comprising the compound according to any one of claims 1 to 15 or a pharmaceutically acceptable salt thereof and a pharmaceutically acceptable carrier.
Use of the compound according to any one of claims 1 to 15 or a pharmaceutically acceptable salt thereof or the pharmaceutical composition according to claim 16 in the preparation of a medicament for treating CDK8/CDK19-mediated cancer.
A method for treating CDK8/CDK19-mediated cancer, comprising administering to a patient a therapeutically effective amount of a compound according to any one of claims 1 to 15 or a pharmaceutically acceptable salt thereof, or a pharmaceutical composition according to claim 16.
The use according to claim 17 and the method according to claim 18, wherein the cancer is selected from blood tumors and solid tumors; preferably, blood tumors include acute myeloid leukemias (AMLs), myelodysplastic syndromes (MDSs) and myeloproliferative diseases (MPDs); solid tumors include breast cancer, gastric cancer, colorectal cancer and pancreatic cancer.