WO2025016439A1 - Compound used as cdk2/4 protein kinase inhibitor and use thereof - Google Patents

Abstract

Provided are a compound used as a CDK2/4 protein kinase inhibitor and the use thereof. Specifically, provided is a compound of formula (I), or a pharmaceutically acceptable salt, stereoisomer, tautomer, hydrate, solvate, isotope compound or prodrug thereof, wherein the definition of each group and substituent is as described in the description. The compound can also be used as an inhibitor of cyclin-dependent kinases (CDKs), and is used for preventing or treating cell proliferative diseases and conditions comprising various cancers, in particular for regulating and/or treating related diseases caused by overexpression and/or abnormal activity of cyclin-dependent kinase CDK2/4.

Description

Compounds used as CDK2/4 protein kinase inhibitors and their applications Technical Field

The present invention relates to the field of medical technology, and in particular to a class of novel compounds used as CDK2/4 protein kinase inhibitors, and their use in regulating CDK2/4 protein kinase activity or treating CDK2/4-related cell proliferation diseases and conditions, especially cancer.

Background Art

Protein kinases regulate a variety of biological functions, including DNA replication, transcription, translation, cell cycle progression, energy metabolism, migration, and cell growth, making them ideal targets for the treatment of proliferative diseases and conditions, including cancer. There is still a need for new compounds that can selectively inhibit protein kinase activity and are effective as therapeutic antiproliferative agents.

Cyclin-dependent kinases (CDKs) belong to the serine/threonine kinase family. They exert physiological functions by binding to corresponding cyclins (Cyclins) to form active dimer complexes, causing cell growth and proliferation. More than 20 CDKs have been discovered, which can be divided into two categories according to their main functions: CDKs that regulate the cell cycle and CDKs that regulate cell transcription. Among them, CDKs 1-6 and 14-18 and their cyclin partners (e.g., Cyclins A, B, D1, D2, D3, E, F, etc.) participate in the regulation of cell cycle progression and are considered to be cell cycle regulators; while CDKs 7-13 and 19-20 and their cyclin partners (e.g., Cyclins C, H, K, L1, L2, T1, T2, etc.) participate in the transcriptional regulation of cells and are considered to be transcriptional regulators. CDKs are therefore involved in the regulation of cell cycle control, apoptosis, differentiation and transcription. It has now been proven that CDKs inhibitors can be used to treat a variety of diseases including cancer.

CDK4/6 is involved in regulating the cell cycle from G1 to S phase after binding to Cyclin D. It is reported that abnormalities in the Cyclin D-CDK4/6-Rb pathway are associated with the progression of resistance to endocrine therapy. Currently, a variety of CDK4/6 inhibitors have been approved for clinical treatment, such as palbociclib, ribociclib, abemaciclib, etc., which have been approved for marketing and combined with endocrine therapy for the treatment of hormone receptor (HR)-positive, human epidermal growth factor 2 (HER2)-negative advanced or metastatic breast cancer. However, during the treatment with CDK4/6 inhibitors, hematological toxicity such as neutropenia and/or gastrointestinal toxic side effects often occur, leading to discontinuation of medication or intermittent administration, which seriously affects the efficacy and compliance of the drug. At present, research data show that the activity of the cell cycle protein Cyclin D3-CDK6 may be related to these side effects. At the same time, clinically, the CDK4/6 inhibitor palbociclib was found to produce drug resistance when combined with endocrine therapy, and the analysis of the transcriptional profile data of patient samples showed that its drug resistance mechanism was highly correlated with the activation of the MYC oncogene and Cyclin E-CDK2 activity. Currently, CDK2 has been proposed as a therapeutic target for a variety of cancers, such as KRAS-driven lung cancer, MYC-amplified neuroblastoma, melanoma, acute myeloid leukemia, glioblastoma, prostate cancer, and breast cancer. The cyclin binding partners of CDK2 (i.e., Cyclin E1 and Cyclin E2) are amplified in a considerable proportion of human cancers, and this phenotype indicates that the patient's overall survival is poor. By inhibiting the activity of Cyclin E-CDK2, it is expected to overcome the drug resistance problem of current clinical CDK4/6 inhibitors.

In summary, given the toxic side effects of current CDK4/6 dual-target inhibitors and the drug resistance they produce when used in combination with endocrine therapy, developing a selective CDK2/4 kinase inhibitor may have better safety and efficacy, as well as broader prospects for clinical application.

Summary of the invention

The purpose of the present invention is to provide a novel compound having CDK2/4 protein kinase inhibitory activity and better pharmacodynamics and pharmacokinetic properties.

In the first aspect of the present invention, a compound is provided, wherein the compound is a compound of formula I, or a pharmaceutically acceptable salt, stereoisomer, tautomer, hydrate, solvate, isotope compound or prodrug thereof,

in:

X is N or CR ₅ ;

_X1 is N or _CR5 ;

R ₁ is selected from the following group: H, halogen (such as F, Cl, Br), cyano, C _1-6 alkyl (such as methyl, ethyl), C _1-6 haloalkyl (such as CF ₃ ), C _1-6 alkoxy (such as methoxy), C _3-6 cycloalkyl (such as cyclopropyl);

_R2 is as shown in Formula Ia or Ib;

in,

W ₃ , W ₄ and W ₅ are each independently selected from: CH, N, NH;

One of _W2 and _W6 is C, and the other is C or N;

R ₆ is independently selected from the following group: halogen, C _1-6 alkyl, C _1-6 haloalkyl, C _1-6 hydroxyalkyl, C _3-6 cycloalkyl, -CO-C _1-6 alkyl (such as ), -COO-C _1-6 alkyl (such as ), 4 to 6 membered heterocyclyl, -C _1-4 alkylene-C _3-6 cycloalkyl, -C _1-4 alkylene-4 to 6 membered heterocyclyl; and wherein the alkyl, alkylene, cycloalkyl and heterocyclyl are optionally substituted by one or more substituents selected from the group consisting of halogen, hydroxyl, cyano, C _1-6 alkyl, C _1-6 haloalkyl, C _1-6 alkoxy;

The subscript p is 0, 1, or 2;

_W1 and _W7 are each independently CH or N;

Subscript o is 0 or 1;

R ₇ is each independently selected from the group consisting of halogen, C _1-6 alkyl, C _1-6 haloalkyl;

Subscript q is 0, 1, 2, or 3;

R ₃ is independently selected from the following group: H, C _1-6 alkyl (such as methyl), C _3-6 cycloalkyl (such as cyclopropyl), hydroxyl, amino, C _1-6 alkoxy (such as methoxy), C _1-6 haloalkyl (such as CF ₃ ), C _1-6 haloalkoxy, C _1-6 hydroxyalkyl, 5-8 membered heterocyclyl, 5-8 membered heteroaryl; or, two R ₃ located on the same atom together form an oxo group (=O);

_R4 is selected from the following groups: H, _C1-6 alkyl (such as methyl, ethyl, isopropyl), -CO- _C1-6 alkoxy (i.e. -CO- _OC1-6 alkyl, such as ), -CO-C _1-6 alkyl (such as ), C _3-8 cycloalkyl (such as: ), a 4-8 membered heterocycloalkyl containing 1, 2 or 3 heteroatoms selected from N, O or S (e.g.: ); wherein the alkyl, aryl, cycloalkyl and heterocycloalkyl are optionally substituted by one or more groups selected from the group consisting of halogen (such as F, Cl, or Br), cyano, hydroxyl, C _1-6 alkyl, -CO-C _1-6 alkyl, C _1-6 alkoxy (such as -O-methyl), amino (such as NH ₂ ), -COOH, -CONH ₂ , -CF ₂ CF ₃ , a 5-8 membered heterocycloalkyl group containing 1, 2 or 3 heteroatoms selected from N, O or S which is optionally substituted by C _1-6 alkyl, -CH(F) ₂ , -COO-C _1-6 alkyl (such as -COO-C(CH ₃ ) ₃ ) or -S(O) ₂ -C _6-10 aryl;

R ₅ is selected from the following group: H, halogen (such as F, Cl, Br), cyano, C _1-6 alkyl (such as methyl, ethyl), C _1-6 haloalkyl (such as CF ₃ ), -CO-C _1-6 alkoxy (i.e. -CO-OC _1-6 alkyl, such as )(Preferably, R ₅ is H);

The subscript m is selected from the following group: 0, 1, 2, 3, 4;

The subscript n is 0, 1, 2, 3 or 4 (preferably, 0, 1 or 2, more preferably, 0 or 1, and most preferably, 1).

In another preferred embodiment, the compound is a compound used as a CDK2/4 kinase inhibitor.

In another preferred embodiment,

X is N or CR ₅ ; and/or,

X ₁ is N or CR ₅ ; and/or,

R ₁ is selected from the following group: H, halogen (such as F, Cl, Br), C _1-6 alkyl (such as methyl), C _1-6 haloalkyl (such as CF ₃ ), C _3-6 cycloalkyl (such as cyclopropyl); and/or,

_R2 is as shown in Formula Ia or Ib; and/or,

in,

W ₃ , W ₄ and W ₅ are each independently selected from: CH, N, NH; and/or,

One of _W2 and _W6 is C, and the other is C or N; and/or,

R ₆ is each independently selected from the following group: halogen, C _1-6 alkyl, C _1-6 haloalkyl, C _1-6 hydroxyalkyl, C _3-6 cycloalkyl, 4 to 6 membered heterocyclyl, -C _1-4 alkylene-C _3-6 cycloalkyl, -C _1-4 alkylene-4 to 6 membered heterocyclyl; and wherein the alkyl, alkylene, cycloalkyl and heterocyclyl are optionally substituted by one or more substituents selected from the following group: halogen, hydroxyl, cyano, C _1-6 alkyl, C _1-6 haloalkyl, C _1-6 alkoxy; and/or,

subscript p is 0, 1 or 2; and/or,

_W1 and _W7 are each independently CH or N; and/or,

subscript o is 0 or 1; and/or,

R ₇ is each independently selected from the group consisting of halogen, C _1-6 alkyl, C _1-6 haloalkyl; and/or,

subscript q is 0, 1, 2 or 3; and/or,

R ₃ is independently selected from the following group: H, C _1-6 alkyl (such as methyl), C _3-6 cycloalkyl (such as cyclopropyl), hydroxyl, C _1-6 alkoxy (such as methoxy); and/or,

_R4 is selected from the following groups: H, _C1-6 alkyl (such as methyl, ethyl, isopropyl), -CO- _C1-6 alkoxy (i.e. -CO- _OC1-6 alkyl, such as ), -CO-C _1-6 alkyl (such as ); and/or,

R ₅ is selected from the following groups: H, C _1-6 alkyl (such as methyl, ethyl), -CO-C _1-6 alkoxy (i.e. -CO-OC _1-6 alkyl, such as )(Preferably, R ₅ is H); and/or,

The subscript m is selected from the group consisting of 0, 1, 2, 3, 4; and/or,

The subscript n is 0, 1, 2, 3 or 4 (preferably, 0, 1 or 2, more preferably, 0 or 1, and most preferably, 1).

In another preferred embodiment, R ₂ is as shown in Formula Ia.

In another preferred embodiment,

In formula (Ia),

_W1 is CH, _W2 is C, _W3 is NH, _W4 is CH, _W5 is N, _W6 is C and _W7 is CH; or,

_W1 is CH, _W2 is C, _W3 is CH, _W4 is CH, _W5 is N, _W6 is N and _W7 is CH; or,

_W1 is CH, _W2 is C, _W3 is N, _W4 is CH, _W5 is N, _W6 is N and _W7 is CH; or,

_W1 is CH, _W2 is N, _W3 is CH, _W4 is CH, _W5 is N, _W6 is C and _W7 is N; or,

_W1 is CH, _W2 is CH, _W3 is N, _W4 is CH, _W5 is CH, _W6 is N and _W7 is CH; or,

_W1 is CH, _W2 is CH, _W3 is N, _W4 is N, _W5 is CH, _W6 is N and _W7 is N; or,

_W1 is CH, _W2 is C, _W3 is NH, _W4 is CH, _W5 is N, _W6 is C and _W7 is N; or,

_W1 is CH, _W2 is C, _W3 is N, _W4 is CH, _W5 is N, _W6 is N and _W7 is N; or,

_W1 is CH, _W2 is C, _W3 is NH, _W4 is CH, _W5 is CH, _W6 is C and _W7 is CH; or,

_W1 is CH, _W2 is C, _W3 is NH, _W4 is N, _W5 is CH, _W6 is C and _W7 is CH; or,

_W1 is CH, _W2 is C, _W3 is NH, _W4 is CH, _W5 is N, _W6 is C and _W7 is CH; or,

_W1 is CH, _W2 is C, _W3 is CH, _W4 is N, _W5 is NH, _W6 is C and _W7 is CH; or,

_W1 is CH, _W2 is C, _W3 is N, _W4 is N, _W5 is NH, _W6 is C and _W7 is CH; or,

_W1 is CH, _W2 is C, _W3 is CH, _W4 is NH, _W5 is N, _W6 is C and _W7 is CH; or,

_W1 is N, _W2 is C, _W3 is CH, _W4 is NH, _W5 is N, _W6 is N and _W7 is CH; or,

_W1 is CH, _W2 is C, _W3 is CH, _W4 is CH, _W5 is N, _W6 is N and _W7 is N;

In formula (Ib),

_W2 is C, _W3 is CH, _W4 is CH, _W5 is N, _W6 is N and o is 0 or 1.

In another preferred embodiment, in formula (Ia), _W1 is CH, _W2 is C, _W3 is CH, _W4 is CH, _W5 is N, _W6 is N and _W7 is N.

In another preferred embodiment, in formula (Ia), _W1 is CH, _W2 is C, _W3 is NH, _W4 is CH, _W5 is N, _W6 is C and _W7 is CH.

In another preferred embodiment, in formula (Ia), _W1 is CH, _W2 is C, _W3 is CH, _W4 is NH, _W5 is N, _W6 is C and _W7 is CH.

In another preferred embodiment, R ₆ is independently selected from the following group: halogen, C _1-6 alkyl, C _1-6 haloalkyl, C _1-6 hydroxyalkyl, C _3-6 cycloalkyl, 4 to 6 membered heterocyclyl; and wherein the alkyl, alkylene, cycloalkyl and heterocyclyl are optionally substituted by one or more substituents selected from the following group: halogen, hydroxyl, cyano, C _1-6 alkyl, C _1-6 haloalkyl, C _1-6 alkoxy.

In another preferred embodiment, R ₆ is independently selected from the following group: halogen, C _1-4 alkyl, C _1-4 haloalkyl, C _1-4 hydroxyalkyl, C _3-6 cycloalkyl, 4 to 6 membered heterocycloalkyl, wherein the 4 to 6 membered heterocycloalkyl is selected from: pyrrolidinyl (such as ), piperidinyl (such as ).

In another preferred embodiment, in R ₆ , the alkyl, alkylene, cycloalkyl and heterocyclic groups are unsubstituted.

In another preferred embodiment, Select from the following group:

In another preferred embodiment, Select from the following group:

In another preferred embodiment, Select from the following group:

In another preferred embodiment, Select from the following group:

In another preferred embodiment, X is N.

In another preferred embodiment, _X1 is N.

In another preferred embodiment, the subscript n is 0 or 1.

In another preferred embodiment, R ₁ is selected from the following group: H, F, Cl, methyl, Br, CF ₃ .

In another preferred embodiment, R ₃ is independently selected from the following group: H, C _1-6 alkyl, C _3-6 cycloalkyl, hydroxyl, C _1-6 alkoxy.

In another preferred embodiment, R ₃ is independently selected from the following group: H, methyl, cyclopropyl, hydroxyl, methoxy, oxo.

In another preferred embodiment, R ₃ is selected from the following group: H, methyl, cyclopropyl, hydroxyl, methoxy.

In another preferred embodiment, R ₄ is selected from the following group: H, C _1-6 alkyl (such as methyl, ethyl, isopropyl), -CO-C _1-6 alkoxy (i.e. -CO-OC _1-6 alkyl, such as ), -CO-C _1-6 alkyl (such as ), C _3-8 cycloalkyl (such as: ), a 4-8 membered heterocycloalkyl containing 1, 2 or 3 heteroatoms selected from N, O or S (e.g.: ); wherein the alkyl, cycloalkyl and heterocycloalkyl are optionally substituted by one or more groups selected from the group consisting of halogen (such as F, Cl, or Br), hydroxyl.

In another preferred embodiment, _R4 is selected from the following group: H, methyl, ethyl,

In another preferred embodiment, _R4 is selected from the following group: H, methyl, ethyl,

In another preferred embodiment, the compound is as shown in Formula II;

wherein R ₃₁ is defined the same as R ₃ , m1 is 0, 1, 2 or 3; and R ₁ , R ₂ , R ₃ , R ₄ , X and X ₁ are defined the same as in formula (I).

In another preferred embodiment, the compound is as shown in formula II-1 or II-2;

wherein R ₃₁ is defined the same as R ₃ , m1 is 0, 1, 2 or 3; and R ₁ , R ₂ , R ₃ , R ₄ , X and X ₁ are defined the same as in formula (I).

In another preferred embodiment, in Formula II, Formula II-1 or Formula II-2, R ₃₁ is H or C _1-6 alkyl (such as methyl).

In another preferred embodiment, in Formula II, Formula II-1 or Formula II-2, R ₃ is independently selected from the following group: H, C _1-6 alkyl (such as methyl), C _3-6 cycloalkyl (such as cyclopropyl), hydroxyl, C _1-6 alkoxy (such as methoxy); or, two R ₃ located on the same atom together form an oxo group (＝O).

In another preferred embodiment, the isotope compound is a deuterated compound.

In another preferred embodiment, the deuterated compound refers to a compound in which one hydrogen atom (H) or multiple hydrogen atoms (H) in the compound (preferably, in R ₄ and/or R ₂ ; more preferably, in R ₄ and/or R ₆ ) is replaced by a deuterium atom (D).

In another preferred embodiment, in the deuterated compound, R ₄ is a deuterated C _1-6 alkyl group (such as -CD ₃ ).

In another preferred embodiment, in the deuterated compound, R ₆ is

In another preferred embodiment, X, X ₁ , R ₁ , R ₂ , R ₃ , R ₄ , R ₅ , subscript m and subscript n are each independently a corresponding group in the specific compounds shown in Table A or in the Preparation Examples.

In another preferred embodiment, the compound is selected from Table A:

Table A

In another preferred embodiment, the inorganic acid salt is selected from the following group: hydrochloride, hydrobromide, hydroiodide, sulfate, bisulfate, nitrate, phosphate, and acid phosphate.

In another preferred embodiment, the organic acid salt is selected from the following group: formate, acetate, trifluoroacetate, propionate, pyruvate, glycolate, oxalate, malonate, fumarate, maleate, lactate, malate, citrate, tartrate, methanesulfonate, ethanesulfonate, benzenesulfonate, p-toluenesulfonate, salicylate, picrate, glutamate, ascorbate, camphorate, camphorsulfonate.

In the second aspect of the present invention, a pharmaceutical composition is provided, which contains a preventive and/or therapeutically effective amount of the compound as described in the first aspect, and a pharmaceutically acceptable carrier.

In the third aspect of the present invention, there is provided a use of the compound according to the first aspect for preparing one or more drugs selected from the following group:

(a) A drug used as a CDK2/4 protein kinase inhibitor;

(b) drugs for modulating CDK2/4 protein kinase activity;

(c) Drugs for preventing or treating CDK2/4 related diseases.

In another preferred embodiment, the CDK2/4-related disease is selected from the following group: inflammation, cancer, cardiovascular disease, infection, immune disease, and metabolic disease.

In another preferred embodiment, the cancer is selected from the group consisting of lung cancer, breast cancer, prostate cancer, colorectal cancer, liver cancer, pancreatic cancer, ovarian cancer, leukemia, neuroblastoma, gastric cancer, kidney cancer, esophageal cancer, uterine cancer, and liposarcoma.

In the fourth aspect of the present invention, a method for preventing or treating a CDK2/4-related disease is provided, comprising administering to a subject in need thereof a compound as described in the first aspect, or a pharmaceutically acceptable salt, stereoisomer, tautomer, hydrate, solvate, isotope compound or prodrug thereof, or a pharmaceutical composition containing the compound, or a pharmaceutically acceptable salt, stereoisomer, tautomer, hydrate, solvate, isotope compound or prodrug thereof.

In another preferred embodiment, the CDK2/4 related disease is as defined above.

In the fifth aspect of the present invention, a method for inhibiting CDK2/4 protein kinase is provided, comprising: contacting CDK2/4 protein kinase with the compound as described in the first aspect, or a pharmaceutically acceptable salt, stereoisomer, tautomer, hydrate, solvate, isotope compound or prodrug thereof, thereby inhibiting CDK2/4 protein kinase.

In another preferred embodiment, the method is non-therapeutic in vitro.

In the sixth aspect of the present invention, a method for regulating the activity of CDK2/4 protein kinase is provided, comprising: contacting CDK2/4 protein kinase with the compound as described in the first aspect, or a pharmaceutically acceptable salt, stereoisomer, tautomer, hydrate, solvate, isotope compound or prodrug thereof, thereby regulating the activity of CDK2/4 protein kinase.

In another preferred embodiment, the method is non-therapeutic in vitro.

In a seventh aspect of the present invention, a method for inhibiting cell proliferation is provided, comprising: contacting cells with the compound as described in the first aspect, thereby inhibiting cell proliferation.

In another preferred embodiment, the cell is a cancer cell or a tumor cell.

In another preferred embodiment, the cells are selected from the group consisting of breast cancer cells (such as MCF-7), ovarian cancer cells (such as A2780), or a combination thereof.

In another preferred embodiment, the method is non-therapeutic in vitro.

It should be understood that within the scope of the present invention, the above-mentioned technical features of the present invention and the technical features specifically described below (such as embodiments) can be combined with each other to form a new or preferred technical solution. Due to space limitations, they will not be described one by one here.

DETAILED DESCRIPTION

After extensive and in-depth research, the inventor unexpectedly discovered a class of compounds with novel structures that have good CDK2/4 protein kinase inhibitory activity. These compounds have excellent inhibitory activity against CDK2/4 protein kinase and have better pharmacodynamics/pharmacokinetic properties. On this basis, the inventor completed the present invention.

the term

Unless otherwise stated, the following terms used in this application (including the specification and claims) have the definitions given below.

When substituents are described by conventional chemical formulas written from left to right, the substituents also include chemically equivalent substituents that would result if the formula were written from right to left. For example, _-CH2O- is equivalent to _-OCH2- .

As used herein, a group consisting of The bonds indicated represent the position at which the group is attached to the rest of the compound or molecule.

"Alkyl", alone or as part of another group, refers to a monovalent straight or branched saturated hydrocarbon group containing 1 to 12 carbon atoms (i.e., C _1-12 alkyl) consisting only of carbon and hydrogen atoms. Alkyl is preferably C _1-6 alkyl (i.e., an alkyl containing 1, 2, 3, 4, 5 or 6 carbon atoms), and more preferably C _1-4 alkyl (i.e., an alkyl containing 1, 2, 3, or 4 carbon atoms). Examples of alkyl groups include, but are not limited to, methyl, ethyl, propyl, isopropyl, isobutyl, sec-butyl, tert-butyl, pentyl, n-hexyl, octyl, dodecyl, and the like. Unless otherwise specified, in this application, alkyl is also intended to include substituted alkyl, i.e., one or more positions in the alkyl are substituted, especially 1-4 substituents, which can be substituted at any position. Unless otherwise specified, in this application, "substituted alkyl" includes, for example, haloalkyl, hydroxyalkyl, and the like. As used herein, "haloalkyl" refers to an alkyl as defined herein in which one or more hydrogens in the group are replaced by the same or different halogens. The haloalkyl group is preferably a C _1-6 haloalkyl group, more preferably a C _1-4 haloalkyl group. Examples of haloalkyl groups include -CH ₂ Cl, -CH ₂ CF ₃ , -CH ₂ CCl ₃ , perfluoroalkyl groups (e.g., -CF ₃ ) and the like. As used herein, "hydroxyalkyl" refers to an alkyl group as defined herein in which one or more hydrogens in the group are replaced by hydroxyl groups. The hydroxyalkyl group is preferably a C _1-6 hydroxyalkyl group, more preferably a C _1-4 hydroxyalkyl group. Examples of hydroxyalkyl groups include -CH ₂ OH, -C(CH ₃ ) ₂ OH and the like.

"Alkylene" refers to a divalent radical of an alkyl group as defined herein, preferably, the alkylene group has 1 to 4 carbon atoms, ie, _{C1-4 alkylene} . Examples of alkylene groups include, for example _{, -CH2- , -CH2CH2-} , and _-CH2CH2CH2- .

"Alkoxy", alone or as part of another group, refers to an alkyl group having an oxygen group attached thereto, which has an alkylO-structure, wherein alkyl has the definition as described above. Preferably, the alkoxy group is a C _1-6 alkoxy group (i.e., -OC _1-6 alkyl), more preferably, a C _1-4 alkoxy group (i.e., -OC _1-4 alkyl). Alkoxy groups include, but are not limited to, methoxy, ethoxy, propoxy, tert-butoxy, and the like. "Haloalkoxy" refers to a group of the formula -OR, wherein R is a haloalkyl group as defined herein. Examples of haloalkoxy groups include, but are not limited to, trifluoromethoxy, difluoromethoxy, 2,2,2-trifluoroethoxy, and the like.

"Thioalkyl" refers to an alkyl group as defined above in which a carbon is substituted with S, S(O) or S(O) ₂ , for example, -S- _1-6alkyl , -S(O)C _1-6alkyl or -S(O) ₂ C _1-6alkyl .

"Alkenyl", alone or as part of another group, refers to an aliphatic group containing at least one double bond, typically having 2 to 20 carbon atoms (i.e., _C2-20 alkenyl). Alkenyl is preferably _C2-6 alkenyl (i.e., alkenyl containing 2, 3, 4, 5 or 6 carbon atoms). Alkenyl includes, but is not limited to, for example, ethenyl, propenyl, butenyl, 1-methyl-2-buten-1-yl, etc. Unless otherwise defined, in the present invention, alkenyl also includes substituted alkenyl.

"Alkenylene" refers to an alkenyl group as defined above with two points of attachment, for example, "vinylene" refers to the group -CH=CH-. Alkenylene is preferably _C2-6 alkenylene (i.e., alkenylene containing 2, 3, 4, 5 or 6 carbon atoms). Unless otherwise defined, alkenylene may be in unsubstituted form or in substituted form with one or more substituents.

"Alkynyl", alone or as part of another group, refers to a straight or branched hydrocarbon chain containing more than 2 carbon atoms and characterized by having one or more triple bonds, typically having 2 to 20 carbon atoms (i.e., _C2-20 alkynyl). Alkynyl is preferably _C2-6 alkynyl (i.e., alkynyl having 2, 3, 4, 5 or 6 carbon atoms). Alkynyl includes, but is not limited to, ethynyl, propargyl and 3-hexynyl. One of the triple bond carbons may optionally be the point of attachment for an alkynyl substituent. In the present invention, unless otherwise defined, alkynyl also includes substituted alkynyl.

"Alkynylene" refers to an alkynyl group as defined above with two points of attachment. For example, "ethynylene" refers to the group: -C≡C-. Alkynylene is preferably a _C2-6 alkynylene (i.e., an alkynylene containing 2, 3, 4, 5 or 6 carbon atoms). Unless otherwise defined, an alkynylene may be in unsubstituted form or in substituted form with one or more substituents.

"Aliphatic group" refers to a straight chain, branched chain or cyclic hydrocarbon group, including saturated and unsaturated groups such as alkyl, alkenyl and alkynyl groups.

"Aromatic ring system" or "aromatic ring" refers to a monocyclic, bicyclic or polycyclic hydrocarbon ring system, wherein at least one ring is aromatic. Preferably, "aromatic ring system" or "aromatic ring" has 6-12 ring atoms, i.e., a C _6-12 aromatic ring, and examples of aromatic rings include benzene ring, naphthalene ring, anthracene ring, etc.

"Aryl", alone or as part of another group, refers to a monovalent group of an aromatic ring system (aromatic ring). Representative aryl groups include fully aromatic ring systems, such as phenyl, naphthyl and anthracenyl; and ring systems in which an aromatic carbocyclic ring is fused to one or more non-aromatic carbocyclic rings, such as indanyl, phthalimide, naphthylimide or tetrahydronaphthyl, etc. In the present invention, aryl is preferably a C _6-12 aryl group. In the present invention, unless otherwise defined, aryl also includes substituted aryl groups.

"Arylalkyl" or "aralkyl" refers to an alkyl moiety in which an alkyl hydrogen atom is replaced by an aryl group. Aralkyl includes groups in which one or more hydrogen atoms in an alkyl group are replaced by an aryl group, and aryl and alkyl are as defined above. Examples of "arylalkyl" or "aralkyl" include benzyl, 2-phenylethyl, 3-phenylpropyl, 9-fluorenyl, diphenylmethyl, and triphenylmethyl, etc.

"Aryloxy" means an -O-(aryl) group in which the aryl portion is as defined above.

"Heteroalkyl" refers to an alkyl group in which a carbon atom is replaced, having one or more backbone chain atoms selected from atoms other than carbon, for example, oxygen, nitrogen, sulfur, phosphorus, or a combination thereof. A numerical range may be given, for example, C _1-6 heteroalkyl refers to the number of carbons in the chain, which includes 1 to 6 carbon atoms. For example, a -CH ₂ OCH ₂ CH ₃ group is referred to as a "C3" heteroalkyl group. Connection to the rest of the molecule may be through a heteroatom or carbon in the heteroalkyl chain. "Heteroalkylene" refers to a divalent alkyl group in which a carbon atom is replaced, having one or more backbone chain atoms selected from atoms other than carbon, for example, oxygen, nitrogen, sulfur, phosphorus, or a combination thereof. Unless otherwise defined, "heteroalkyl" and "heteroalkylene" include substituted or unsubstituted forms.

"Carbocyclic ring system" or "carbocycle" refers to a monocyclic, bicyclic or polycyclic hydrocarbon ring system, wherein each ring is fully saturated or contains one or more unsaturated units, but wherein none of the rings is aromatic. Preferably, a "carbocyclic ring system" or "carbocycle" has 6-12 ring atoms, i.e., a C _6-12 carbocyclic ring. "Carbocyclyl" refers to a monovalent group of a carbocyclic ring system or carbocyclic ring as defined above. Preferably, a carbocyclyl has 6-12 ring atoms, i.e., a C _6-12 carbocyclyl. For example, examples of carbocyclyl include cycloalkyl (e.g., cyclopentyl, cyclobutyl, cyclopropyl, cyclohexyl, etc.) and cycloalkenyl (e.g., cyclopentenyl, cyclohexenyl, cyclopentadienyl, etc.), etc.

"Cycloalkyl" refers to a monovalent saturated carbocyclic group consisting of a mono- or bicyclic ring having 3-12 (i.e., _C3-12 cycloalkyl), preferably 3-10 (i.e., _C3-10 cycloalkyl), more preferably 3-8 ring atoms (i.e., _C3-8 cycloalkyl), and most preferably 3, 4, 5 or 6 ring atoms (i.e., _C3-6 cycloalkyl). Unless otherwise defined, a cycloalkyl may be optionally substituted with one or more substituents. Preferably, the substituents of a cycloalkyl may be independently hydroxy, alkyl, alkoxy, halogen, haloalkyl, amino, monoalkylamino or dialkylamino. Examples of cycloalkyl groups include, but are not limited to, cyclopropyl, cyclobutyl, cyclopentyl, cyclohexyl, cycloheptyl, etc.

"Cycloalkyloxy" refers to a radical of the formula -OR, where R is a cycloalkyl radical as defined herein. Exemplary cycloalkyloxy radicals include cyclopropyloxy, cyclobutyloxy, cyclopentyloxy, cyclohexyloxy, and the like. "Cycloalkylalkyl" or "cycloalkylalkylene" refers to -alkylene(cycloalkyl), where cycloalkyl and alkylene are as defined above. "Cycloalkylalkyl" or "cycloalkylalkylene" is bonded to the parent molecular structure through an alkyl(alkylene) radical.

"Heteroaromatic ring system" or "heteroaromatic ring" refers to a monocyclic (e.g., 5- or 6-membered), bicyclic (6-12-membered) or polycyclic ring system, wherein at least one ring is an aromatic ring containing at least one heteroatom (e.g., N, O or S) as a ring atom and the remaining ring atoms are carbon. In some cases, the aromatic ring containing at least one heteroatom contains 1, 2, 3 or 4 heteroatoms in the ring. In addition to the aromatic ring containing at least one heteroatom as a ring atom, the remaining rings in the "heteroaromatic ring system" or "heteroaromatic ring" can be saturated, partially unsaturated or fully unsaturated rings.

"Heteroaryl", alone or as part of another group, refers to a monovalent radical of a "heteroaromatic ring system" or "heteroaromatic ring" as defined above. The point of attachment of a heteroaryl group shall be on the aromatic ring. Examples of heteroaryl groups include, but are not limited to, imidazolyl, oxazolyl, isoxazolyl, thiazolyl, isothiazolyl, oxadiazolyl, thiadiazolyl, pyrazinyl, thienyl, furanyl, pyranyl, pyridinyl, pyrrolyl, pyrazolyl, pyrimidinyl, quinolyl, isoquinolyl, benzofuranyl, benzofuranyl, benzothiophenyl, benzothiopyranyl, benzimidazolyl, benzoxazolyl, benzoxadiazolyl, benzothiazolyl, benzothiadiazolyl, benzopyranyl, indolyl, isoindolyl, triazolyl, triazine, quinoxalinyl, purinyl, quinazolinyl, quinolizinyl, naphthyridinyl, pteridinyl, carbazolyl, aza Base, diazepine Heteroarylene refers to a heteroaryl group as defined above with two attachment sites. Unless otherwise defined, heteroaryl includes substituted or unsubstituted forms.

"Heterocyclic ring system" or "heterocycle" refers to monocyclic, bicyclic and polycyclic ring systems in which at least one ring is saturated or partially unsaturated (but nonaromatic) and the ring contains at least one heteroatom as a ring atom. The heterocyclic ring system or heterocycle may be attached to a pendant group at any heteroatom or carbon atom which creates a stable structure and any ring atom may be optionally substituted.

"Heterocyclyl" refers to a heterocyclic ring system or a monovalent radical of a heterocycle as defined above, generally a stable monocyclic ring (e.g., 3-8 members, i.e., 3, 4, 5, 6, 7 or 8 members) or bicyclic ring (e.g., 5-12 members, i.e., 5, 6, 7, 8, 9, 10, 11 or 12 members) or polycyclic ring (e.g., 7-14 members, i.e., 7, 8, 9, 10, 11, 12, 13 or 14 members), including fused, spiro and/or bridged ring structures, which are saturated, partially unsaturated, and which contain carbon atoms and 1, 2, 3 or 4 heteroatoms independently selected from N, O and S as ring atoms. The heterocyclyl is preferably a 3- to 14-membered heterocyclyl, more preferably a 3- to 8-membered heterocyclyl, and most preferably a 4- to 6-membered heterocyclyl. Representative heterocyclyl groups include those in which (1) each ring is non-aromatic and at least one ring contains a heteroatom, for example, tetrahydrofuranyl, tetrahydropyranyl, tetrahydrothiophenyl, pyrrolidinyl, pyrrolidonyl, piperidinyl, pyrrolinyl, decahydroquinolinyl, oxazolidinyl, piperazinyl, dioxanyl, dioxolanyl, diazepinyl, oxazepinyl, thiazepinyl, morpholinyl, and quinuclidinyl; and (2) at least one ring is non-aromatic. and contains heteroatoms as ring atoms and at least one other ring is an aromatic carbocyclic ring, for example, 1,2,3,4-tetrahydroquinolinyl, 1,2,3,4-tetrahydroisoquinolinyl; and (3) at least one ring is non-aromatic and contains heteroatoms and at least one other ring is aromatic and contains heteroatoms, for example, 3,4-dihydro-1H-pyrano[4,3-c]pyridine and 1,2,3,4-tetrahydro-2,6-naphthyridine. Heterocyclylene refers to a heterocyclyl as defined above having two attachment sites. In the present invention, preferably the heterocyclylene is a bicyclic ring, one of which is a heteroaryl group and is connected to the other parts in the general formula through the heteroaryl group. In the present invention, preferably the heterocyclylene is a 5-6-membered monocyclic heterocyclylene or an 8-10-membered bicyclic heterocyclylene. Unless otherwise defined, "heterocyclyl" and "heterocyclylene" include substituted or unsubstituted forms. When the heterocyclic group is saturated, the heterocyclic group may also be referred to as a heterocycloalkyl group.

"Heterocyclylalkyl" refers to an alkyl group substituted with a heterocyclyl group, wherein the heterocyclyl group and the alkyl group are as defined above.

The "alkylamino group" refers to a group having an alkyl-NR-structure, wherein R is H, or an alkyl group, a cycloalkyl group, an aryl group, a heteroaryl group, or the like as described above.

"Cycloalkylamino" refers to a radical of the formula -N( _Ra ) _Rb , wherein _Ra is H, alkyl as defined herein, or cycloalkyl as defined herein, and _Rb is cycloalkyl as defined herein, or _Ra and _Rb together with the N atom to which they are attached form a 3-10 membered N-containing monocyclic or bicyclic heterocyclic radical, such as tetrahydropyrrolyl. As used herein, _C3 - _C8 cycloalkylamino refers to an amine radical containing 3-8 carbon atoms.

In the present invention, "ester group" refers to a group having a -C(O)-O-R or R-C(O)-O- structure, wherein R independently represents hydrogen, alkyl, cycloalkyl, aryl, heteroaryl, or heterocyclic group, as defined above.

In the present invention, the term "amide group" refers to a group with the structure -CONRR', wherein R and R' can independently represent hydrogen, alkyl or substituted alkyl, cycloalkyl or substituted cycloalkyl, aryl or substituted aryl, heterocycle or substituted heterocycle, as defined above. R and R' can be the same or different in the dialkylamine fragment.

In the present invention, the term "sulfonamide" refers to a group with the structure _-SO2NRR ', wherein R and R' can independently represent hydrogen, alkyl or substituted alkyl, cycloalkyl or substituted cycloalkyl, aryl or substituted aryl, heterocycle or substituted heterocycle, as defined above. R and R' can be the same or different in the dialkylamine fragment.

"Ketocarbonyl" refers to R-C(=O)-, wherein R is alkyl, cycloalkyl, etc. as described above.

When a substituent is a non-terminal substituent, it is a substituent of the corresponding group, for example, alkyl for alkylene, cycloalkyl for cycloalkylene, heterocyclyl for heterocyclylene, alkoxy for alkyleneoxy, and the like.

In the present invention, each of the above-mentioned alkyl, alkoxy, cycloalkyl, heteroalkyl, aryl, heteroaryl, cycloheteroalkyl, alkenyl, alkyne, heterocycle, heterocyclic group and the like may be substituted or unsubstituted.

In the present invention, the term "substituted" refers to one or more hydrogen atoms on a specific group being replaced by a specific substituent. The specific substituent is a substituent described above, or a substituent appearing in the embodiments. Unless otherwise specified, a substituted group may have a substituent selected from a specific group at any substitutable site of the group, and the substituent may be the same or different at each position. It will be understood by those skilled in the art that the combinations of substituents contemplated by the present invention are those that are stable or chemically feasible. Typical substitutions include, but are not limited to, one or more of the following groups: such as hydrogen, deuterium, halogen (e.g., monohalogen substituents or polyhalogen substituents, the latter such as trifluoromethyl or alkyl containing Cl ₃ ), cyano, nitro, oxo (e.g., =O), trifluoromethyl, trifluoromethoxy, cycloalkyl, alkenyl, alkynyl, heterocyclic _, aromatic, _ORa , _SRa , S(=O) _Re , S(=O) _{2Re ,} P(=O) _2Re , S(= _O )2ORe, _P (=O) _2ORe , _{NRbRc , NRbS (} = _{O )} 2Re, _NRbP (=O _{) 2Re} , S(=O _{) 2NRbRc} , P(=O)2NRbRc, C(=O) _ORd , _{C (} =O) _{Ra ,} C(=O) _{NRbRc ,} OC(= _O ) _Ra , OC(＝O)NR _b R _c , NR _b C(＝O)OR _e , NR _d C(＝O)NR _b R _c , NR _d S(＝O) ₂ NR _b R _c , NR _d P(＝O) ₂ NR _b R _c , NR _b C(＝O)R _a , or NR _b P(＝O) ₂ R _e , wherein _Ra can independently represent hydrogen, deuterium, alkyl, cycloalkyl, alkenyl, alkynyl, heterocyclic ring or aromatic ring, R _b , R _c and R _d can independently represent hydrogen, deuterium, alkyl, cycloalkyl, heterocyclic ring or aromatic ring, or R _b and R _c together with the N atom can form a heterocyclic ring; _Re can independently represent hydrogen, alkyl, cycloalkyl, alkenyl, alkynyl, heterocyclic ring or aromatic ring. The above typical substituents, such as alkyl, cycloalkyl, alkenyl, cycloalkenyl, alkynyl, heterocyclic ring or aromatic ring can be optionally substituted. The substituents include, but are not limited to, halogen, hydroxyl, cyano, carboxyl (-COOH), _{C1 - C6} alkyl, C2- _C6 alkenyl, C2 _{- C6} alkynyl, C3- _{C8 cycloalkyl} , 3-12 membered heterocyclic group, aryl, heteroaryl, _C1 - _C8 aldehyde group, _C2 - _C10 acyl group, _C2 - _C10 ester group, amine group, _C1 - _C6 alkoxy group, _C1 - _C10 sulfonyl group, and _C1 - _C6 urea group.

"Cyano" refers to -CN.

"Nitro" refers to _-NO2 .

"Hydroxyl" refers to -OH.

"Amino" refers to _-NH2 or RNH-, wherein R is ketocarbonyl, sulfonyl, sulfonamide, Ra _{- C(=O)-, RaRbNC (} =O)-, etc., wherein _Ra and _Rb are alkyl, cycloalkyl, aryl or heteroaryl, etc.

"Halogen" refers to any halogen radical, for example, -F, -Cl, -Br or -I.

The term "deuterated compound" refers to a compound in which one or more hydrogen atoms (H) are replaced by deuterium atoms (D).

In the present invention, the term "plurality" independently refers to 2, 3, 4, or 5.

The structural formula of the carbamate group is -NH-C(=O)-O-R, wherein R is an alkyl group, an aryl group, a heteroaryl group, etc.

Active ingredients

As used herein, the term "compound of the present invention" or "active ingredient of the present invention" is used interchangeably to refer to a compound of formula I, or a pharmaceutically acceptable salt, hydrate, solvate, isotope compound (such as a deuterated compound or deuterated substance) or prodrug thereof. The term also includes racemates and optical isomers.

The compound of formula I has the following structure:

R ₁ , R ₂ , R ₃ , R ₄ , X, X ₁ , m and n are as defined above.

In another preferred embodiment, R ₁ , R ₂ , R ₃ , R ₄ , X, X ₁ , m and n are each independently a corresponding specific group in the specific compounds described in the present invention or the example compounds or the compounds shown in Table A.

The salts that may be formed by the compounds of the present invention also belong to the scope of the present invention. Unless otherwise specified, the compounds of the present invention are understood to include their salts. The term "salt" used herein refers to an acidic or basic salt formed with an inorganic or organic acid and a base. In addition, when the compound of the present invention contains a basic fragment, it includes but is not limited to pyridine or imidazole, and contains an acidic fragment, including but not limited to carboxylic acid, the zwitterions ("inner salts") that may be formed are included in the scope of the term "salt". Pharmaceutically acceptable (i.e., non-toxic, physiologically acceptable) salts are preferred, although other salts are also useful, for example, in separation or purification steps in the preparation process. The compounds of the present invention may form salts, for example, Compound I reacts with a certain amount of acid or base, such as an equivalent amount, and is salted out in a medium, or freeze-dried in an aqueous solution.

The compounds of the present invention contain basic fragments, including but not limited to amines or pyridine or imidazole rings, which may form salts with organic or inorganic acids. Typical acids that can form salts include acetates (such as acetic acid or trihaloacetic acid, such as trifluoroacetic acid), adipates, alginate, ascorbate, aspartate, benzoate, benzenesulfonate, bisulfate, borate, butyrate, citrate, camphor salt, camphorsulfonate, cyclopentanepropionate, diglycolate, dodecyl sulfate, ethanesulfonate, fumarate, glucoheptonate, glycerophosphate, hemisulfate, heptanoate, hexanoate, hydrochloride, hydrobromide, hydroiodide [0013] The invention also includes but is not limited to the following: esters, isothioates (e.g., 2-hydroxyethanesulfonate), lactates, maleates, methanesulfonates, naphthalenesulfonates (e.g., 2-naphthalenesulfonate), nicotinates, nitrates, oxalates, pectinates, persulfates, phenylpropionates (e.g., 3-phenylpropionate), phosphates, picrates, pivalates, propionates, salicylates, succinates, sulfates (e.g., formed with sulfuric acid), sulfonates, tartrates, thiocyanates, toluenesulfonates such as p-toluenesulfonates, dodecanoates, and the like.

Certain compounds of the present invention may contain acidic fragments, including but not limited to carboxylic acids, which may form salts with various organic or inorganic bases. Typical base-formed salts include ammonium salts, alkali metal salts such as sodium, lithium, potassium salts, alkaline earth metal salts such as calcium, magnesium salts and salts formed with organic bases (such as organic amines), such as benzathine, dicyclohexylamine, hepamine (salt formed with N,N-di(dehydroabietyl)ethylenediamine), N-methyl-D-glucamine, N-methyl-D-glucamide, tert-butylamine, and salts formed with amino acids such as arginine, lysine, etc. Basic nitrogen-containing groups can be combined with halide quaternary ammonium salts, such as small molecule alkyl halides (such as chlorides, bromides and iodides of methyl, ethyl, propyl and butyl), dialkyl sulfates (such as dimethyl sulfate, diethyl sulfate, dibutyl sulfate and dipentyl sulfate), long chain halides (such as chlorides, bromides and iodides of decyl, dodecyl, tetradecyl and tetradecyl), aralkyl halides (such as benzyl and phenyl bromides), etc.

Prodrugs and solvates (or solvates) of the compounds of the present invention are also within the scope of this invention.

The term "prodrug" herein refers to a compound that undergoes chemical transformation by metabolism or chemical processes to produce a compound, salt, or solvate of the present invention when treating a related disease. The compounds of the present invention include solvates, such as hydrates.

The compounds, salts or solvates of the present invention may exist in tautomeric forms (such as amides and imino ethers). These tautomers are all part of the present invention.

All stereoisomers of the compounds (e.g., those due to asymmetric carbon atoms that may exist for various substitutions), including enantiomeric and diastereomeric forms, are contemplated by the present invention. Individual stereoisomers of the compounds of the present invention may not exist simultaneously with other isomers (e.g., as a pure or substantially pure optical isomer having a particular activity), or may be mixtures, such as racemates, or mixtures with all other stereoisomers or portions thereof. The chiral centers of the present invention have two configurations, S or R, as defined by the 1974 recommendations of the International Union of Pure and Applied Chemistry (IUPAC). Racemic forms can be resolved by physical methods, such as fractional crystallization, or by crystallization of diastereoisomers derived therefrom, or by separation by chiral column chromatography. Individual optical isomers can be obtained from racemates by suitable methods, including but not limited to conventional methods, such as recrystallization after salt formation with an optically active acid.

The compounds of the present invention are prepared, separated and purified to obtain compounds with a weight content of equal to or greater than 90%, for example, equal to or greater than 95%, equal to or greater than 99% ("very pure" compounds), which are listed in the text description. Such "very pure" compounds of the present invention are also considered part of the present invention.

All configurational isomers of the compounds of the present invention are within the scope of the invention, whether in mixture, pure or very pure form. The definition of the compounds of the present invention includes both cis (Z) and trans (E) olefin isomers, as well as cis and trans isomers of carbocyclic and heterocyclic rings.

Throughout the specification, groups and substituents may be selected to provide stable fragments and compounds.

Specific functional groups and chemical term definitions are described in detail below. For purposes of the present invention, chemical elements are as defined in the Periodic Table of the Elements, CAS version, Handbook of Chemistry and Physics, ^75th Ed. Definitions of specific functional groups are also described therein. In addition, the basic principles of organic chemistry as well as specific functional groups and reactivity are also described in "Organic Chemistry", Thomas Sorrell, University Science Books, Sausalito: 1999, which is incorporated by reference in its entirety.

Certain compounds of the present invention may exist in specific geometric or stereoisomeric forms. The present invention encompasses all compounds, including their cis and trans isomers, R and S enantiomers, diastereomers, (D) isomers, (L) isomers, racemic mixtures and other mixtures. In addition, asymmetric carbon atoms may represent substituents, such as alkyl. All isomers and their mixtures are included in the present invention.

According to the present invention, the ratio of isomers contained in the mixture of isomers can be various. For example, in the mixture of only two isomers, there can be the following combinations: 50:50, 60:40, 70:30, 80:20, 90:10, 95:5, 96:4, 97:3, 98:2, 99:1, or 100:0, and all ratios of isomers are within the scope of the present invention. Similar ratios that are easily understood by ordinary technicians in this profession, and ratios for more complex mixtures of isomers are also within the scope of the present invention.

The present invention also includes isotopically labeled compounds that are equivalent to the original compounds disclosed herein. However, in practice, the substitution of one or more atoms by atoms having a different atomic mass or mass number will generally occur. Examples of isotopes of the compounds of the present invention include ^hydrogen , carbon, nitrogen, oxygen, phosphorus, sulfur, fluorine and chlorine isotopes, such as ^2H , ^3H , ^13C , ^{11C, 14C , 15N} , 18O, ^17O , ^31P , ^32P , ^35S , ^18F and ^36Cl , respectively. The compounds of the present invention, or enantiomers, diastereomers, isomers, or pharmaceutically acceptable salts or solvates containing isotopes or other isotopic atoms of the above compounds are within the scope of the present invention. Certain isotopically labeled compounds of the present invention, such as radioactive isotopes of ^3H and ^14C , are also included, which are useful in tissue distribution experiments of drugs and substrates. Tritium, i.e., ^3H and carbon-14, i.e., ^14C , are relatively easy to prepare and detect. Easy. It is the first choice among isotopes. In addition, heavier isotope substitutions such as deuterium, i.e. ² H, have advantages in certain therapies due to their good metabolic stability, such as increasing half-life in the body or reducing dosage, and therefore, may be preferred in some cases. Isotope-labeled compounds can be prepared by general methods by replacing readily available isotope-labeled reagents with non-isotopic reagents using the protocols disclosed in the examples.

If a synthesis of a specific enantiomer of the compound of the present invention is to be designed, it can be prepared by asymmetric synthesis, or derivatized with a chiral auxiliary, the resulting diastereomeric mixture can be separated, and the chiral auxiliary can be removed to obtain the pure enantiomer. In addition, if the molecule contains a basic functional group, such as an amino acid, or an acidic functional group, such as a carboxyl group, a diastereomeric salt can be formed with a suitable optically active acid or base, and then separated by conventional means such as fractional crystallization or chromatography, and then the pure enantiomer can be obtained.

As described herein, the compounds of the present invention can be replaced with any number of substituents or functional groups to expand their scope. Generally, the term "substituted" appears before or after the term "optional", and the general formula including substituents in the formulation of the present invention refers to replacing hydrogen free radicals with designated structural substituents. When multiple positions in a specific structure are substituted by multiple specific substituents, the substituents can be the same or different at each position. The term "substituted" used herein includes all allowed organic compound substitutions. In a broad sense, allowed substituents include non-cyclic, cyclic, branched non-branched, carbocyclic and heterocyclic, aromatic and non-aromatic organic compounds. In the present invention, heteroatom nitrogen can have hydrogen substituents or any allowed organic compounds described above to supplement its valence. In addition, the present invention is not intended to limit the allowed substitution of organic compounds in any way. The present invention believes that the combination of substituents and variable groups is very good in the treatment of diseases in the form of stable compounds. As used herein, the term "stable" refers to compounds that are stable and that maintain the structural integrity of the compound over a period of time sufficient to be detected, and preferably over a period of time sufficient to be effective, as used herein for the purposes described above.

The metabolites of the compounds and pharmaceutically acceptable salts thereof involved in the present application, as well as prodrugs that can be converted into the structures of the compounds and pharmaceutically acceptable salts involved in the present application and in vivo, are also included in the claims of the present application.

Preparation method

Methods for preparing compounds of Formula I are described in the following schemes and examples. Starting materials and intermediates were purchased from commercial sources, prepared by known procedures, or otherwise described. In some cases, the order in which the steps of the reaction schemes are carried out may be altered to facilitate the reaction or to avoid unwanted side reaction products.

The preparation method of the compound of formula I of the present invention is described in more detail below, but these specific methods do not constitute any limitation to the present invention. The compounds of the present invention can also be conveniently prepared by optionally combining various synthetic methods described in this specification or known in the art, and such a combination can be easily carried out by a technician in the field to which the present invention belongs.

Usually, in the preparation process, each reaction is usually carried out under the protection of inert gas in a suitable solvent at 0 to 150° C., and the reaction time is usually 2 to 24 hours.

Preferably the preparation method is as follows:

method:

Step 1: In a solvent (1,4-dioxane, tetrahydrofuran, 1,2-dichloroethane), SM1 and S1 react at 70-120 degrees to generate by palladium-catalyzed coupling or nucleophilic substitution reaction.

Step 2: In a solvent (1,4-dioxane, tetrahydrofuran, toluene), SM2 and S2 are reacted under alkaline conditions by palladium-catalyzed coupling or nucleophilic substitution to obtain product T

In the above formulae, R ₁ , R ₂ , R ₃ , R ₄ , X, X ₁ , m and n are as defined above.

Unless otherwise specified, the above starting materials can be purchased from commercial sources or synthesized according to reported literature.

Pharmaceutical compositions and methods of administration

Since the compounds of the present invention have excellent CDK2/4 protein kinase inhibitory activity, the compounds of the present invention and their various crystal forms, pharmaceutically acceptable inorganic or organic salts, hydrates or solvates, and pharmaceutical compositions containing the compounds of the present invention as the main active ingredient can be used to prevent and/or treat CDK2/4-related diseases. According to the prior art, the compounds of the present invention or pharmaceutical compositions containing the compounds of the present invention can be used to prevent and/or treat the following diseases: inflammation, cancer, cardiovascular disease, infection, immune disease, metabolic disease.

The compounds of the present invention may be used in combination with other drugs known to treat or improve similar conditions. When the compound of the present invention is administered in combination, the administration and dosage of the original drug may remain unchanged, while the compound of the present invention is taken simultaneously or subsequently. When the compound of the present invention is taken simultaneously with one or more other drugs, a pharmaceutical composition containing one or more known drugs and the compound of the present invention may be preferably used. Drug combination also includes taking the compound of the present invention and one or more other known drugs in overlapping time periods. When the compound of the present invention is used in combination with one or more other drugs, the dosage of the compound of the present invention or the known drug may be lower than the dosage of the drug alone.

The drugs or active ingredients that can be used in combination with the compounds of Formula I include but are not limited to: PD-1 inhibitors (such as nivolumab, pembrolizumab, JS-001, SHR-120, BGB-A317, IBI-308, GLS-010, GB-226, STW204, HX008, HLX10, BAT1306, AK105, LZM 009 or biosimilars of the above drugs, etc.), PD-L1 inhibitors (such as durvalumab, atezolizumab, CS1001, KN035, HLX20, SHR-1316, BGB-A333, JS003, CS1003, KL-A167, F 520, GR1405, MSB2311 or biosimilars of the above drugs, etc.), CD20 antibodies (such as rituximab, obinutuzumab, ofatumumab, tositumomab, ibritumomab tiuxetan, etc.), CD47 antibodies (such as Hu5F9-G4, CC-90002, TTI-621, TTI-622, OSE-172, SRF-231, ALX-148, NI-1701, SHR-1603, IBI188, IMM01), ALK inhibitors (such as ceritinib, alectinib, brigatinib, lorlatinib, ocatinib), PI3K inhibitors (such as idelalisib, dactolisib, taselisib, Bup arlisib, etc.), BTK inhibitors (such as ibrutinib, tirabrutinib, acalabrutinib, etc.), EGFR inhibitors (such as afatinib, gefitinib, erlotinib, lapatinib, dacomitinib, icotinib, canertinib, etc.), VEGFR inhibitors (such as sorafenib, pazopanib, rivatinib, cabozantinib, sunitinib, donafenib, etc.), HDAC inhibitors (such as givinostat, droxinostat, entinostat, dacilast, tekdinib, etc.), CDK inhibitors (such as palbociclib, ribociclib, abemaciclib, lerociclib, etc.), MEK inhibitors (such as selumetinib, etc.), (AZD6244), Trametinib (GSK1120212), PD0325901, U0126, AS-703026, PD184352 (CI-1040), etc.), Akt inhibitors (such as MK-2206, Ipatasertib, Capivasertib, Afuresertib, Uprosertib, etc.), mTOR inhibitors (such as Vistusertib, etc.), SHP2 inhibitors (such as RMC-4630, JAB-3068, TNO155, etc.), IGF-1R inhibitors (such as Ceritinib, Oclatinib, linsitinib, BMS-754807, etc.), , GSK1838705A, etc.), ER antagonists or degraders (such as tamoxifen, fulvestrant, etc.), aromatase inhibitors (such as letrozole, etc.), BCL2 or BCL-XL inhibitors (such as ABT-199, ABT-263, etc.), Hedgehog inhibitors (such as vismodegib, cyclopamine, etc.), chemotherapeutic drugs (such as cisplatin, etoposide, topotecan, etc.), PARP inhibitors (such as Olaparib, Veliparib, Rucaparib, etc.), ATR/ATM inhibitors (such as Ceralasertib, Berzosertib, etc.) or a combination thereof.

The dosage forms of the pharmaceutical composition of the present invention include (but are not limited to): injection, tablet, capsule, aerosol, suppository, film, pill, external ointment, controlled release or sustained release or nano preparation.

The pharmaceutical composition of the present invention comprises a safe and effective amount of the compound of the present invention or a pharmacologically acceptable salt thereof and a pharmacologically acceptable excipient or carrier. Wherein "safe and effective amount" means: the amount of the compound is sufficient to significantly improve the condition without causing serious side effects. Usually, the pharmaceutical composition contains 1-2000 mg of the compound of the present invention per dose, and more preferably, contains 10-1000 mg of the compound of the present invention per dose. Preferably, the "one dose" is a capsule or tablet.

"Pharmaceutically acceptable carrier" refers to: one or more compatible solid or liquid fillers or gel substances, which are suitable for human use and must have sufficient purity and sufficiently low toxicity. "Compatibility" here means that the components in the composition can be mixed with the compounds of the present invention and with each other without significantly reducing the efficacy of the compounds. Some examples of pharmaceutically acceptable carriers include cellulose and its derivatives (such as sodium carboxymethyl cellulose, sodium ethyl cellulose, cellulose acetate, etc.), gelatin, talc, solid lubricants (such as stearic acid, magnesium stearate), calcium sulfate, vegetable oils (such as soybean oil, sesame oil, peanut oil, olive oil, etc.), polyols (such as propylene glycol, glycerol, mannitol, sorbitol, etc.), emulsifiers (such as Tween ), wetting agents (such as sodium lauryl sulfate), colorants, flavoring agents, stabilizers, antioxidants, preservatives, pyrogen-free water, etc.

There is no particular limitation on the administration of the compound or pharmaceutical composition of the present invention. Representative administrations include, but are not limited to, oral, intratumoral, rectal, parenteral (intravenous, intramuscular or subcutaneous), and topical administration.

Solid dosage forms for oral administration include capsules, tablets, pills, powders and granules. In these solid dosage forms, the active compound is mixed with at least one conventional inert excipient (or carrier), such as sodium citrate or dicalcium phosphate, or with the following ingredients: (a) fillers or extenders, for example, starches, lactose, sucrose, glucose, mannitol, and silicic acid; (b) binders, for example, hydroxymethylcellulose, alginates, gelatin, polyvinyl pyrrolidone, sucrose, and acacia; (c) humectants, for example, glycerol; (d) disintegrants, for example, agar, calcium carbonate, potato starch or tapioca starch, alginic acid, certain complex silicates, and sodium carbonate; (e) solubilizers, for example, paraffin; (f) absorption accelerators, for example, quaternary ammonium compounds; (g) wetting agents, for example, cetyl alcohol and glyceryl monostearate; (h) adsorbents, for example, kaolin; and (i) lubricants, for example, talc, calcium stearate, magnesium stearate, solid polyethylene glycols, sodium lauryl sulfate, or mixtures thereof. In the case of capsules, tablets and pills, the dosage forms may also comprise buffering agents.

Solid dosage forms such as tablets, pills, capsules, pills and granules can be prepared using coatings and shell materials, such as enteric coatings and other materials known in the art. They may contain opacifiers, and the release of the active compound or compounds in such compositions can be delayed in a certain part of the digestive tract. Examples of embedding components that can be used are polymeric substances and waxes. If necessary, the active compound can also be formed into microencapsulated form with one or more of the above-mentioned excipients.

Liquid dosage forms for oral administration include pharmaceutically acceptable emulsions, solutions, suspensions, syrups or tinctures. In addition to the active compound, the liquid dosage form may contain an inert diluent conventionally used in the art, such as water or other solvents, solubilizers and emulsifiers, for example, ethanol, isopropanol, ethyl carbonate, ethyl acetate, propylene glycol, 1,3-butylene glycol, dimethylformamide and oils, in particular cottonseed oil, peanut oil, corn germ oil, olive oil, castor oil and sesame oil or mixtures of these substances.

Besides such inert diluents, the composition may also include adjuvants such as wetting agents, emulsifying and suspending agents, sweetening, flavoring, and perfuming agents.

Suspensions, in addition to the active compounds, may contain suspending agents such as, for example, ethoxylated isostearyl alcohols, polyoxyethylene sorbitol and sorbitan esters, microcrystalline cellulose, aluminum methanol and agar, or mixtures of these substances.

Compositions for parenteral injection may include physiologically acceptable sterile aqueous or anhydrous solutions, dispersions, suspensions or emulsions and sterile powders for reconstitution into sterile injectable solutions or dispersions. Suitable aqueous and non-aqueous carriers, diluents, solvents or excipients include water, ethanol, polyols and suitable mixtures thereof.

Dosage forms for topical administration of the compounds of the invention include ointments, powders, patches, sprays and inhalants. The active ingredient is mixed under sterile conditions with a physiologically acceptable carrier and any preservatives, buffers, or propellants that may be required.

The treatment method of the present invention can be used alone or in combination with other treatment methods or therapeutic drugs.

When using the pharmaceutical composition, a safe and effective amount of the compound of the present invention is applied to a mammal (such as a human) in need of treatment, wherein the dosage during administration is a pharmaceutically effective dosage, and for a person weighing 60 kg, the daily dosage is usually 1 to 2000 mg, preferably 50 to 1000 mg. Of course, the specific dosage should also take into account factors such as the route of administration and the health status of the patient, which are all within the skill of a skilled physician.

The present invention also provides a method for preparing a pharmaceutical composition, comprising the steps of: mixing a pharmaceutically acceptable carrier with the compound of the present invention or its crystal form, pharmaceutically acceptable salt, hydrate or solvate to form a pharmaceutical composition.

The present invention also provides a treatment method, which comprises the steps of administering the compound described in the present invention, or its crystal form, pharmaceutically acceptable salt, hydrate or solvate, or administering the pharmaceutical composition described in the present invention to a subject in need of treatment, for inhibiting CDK2/4 protein kinase.

The main advantages of the present invention include

(1) The compounds of the present invention have excellent inhibitory ability against CDK2/4 protein kinase;

(2) The compounds of the present invention have lower toxic side effects;

(3) The compounds of the present invention have better pharmacodynamics and pharmacokinetic properties.

The present invention is further described below in conjunction with specific examples. It should be understood that these examples are only used to illustrate the present invention and are not intended to limit the scope of the present invention. The experimental methods in the following examples where specific conditions are not specified are generally performed under conventional conditions, such as those described in Sambrook et al., Molecular Cloning: A Laboratory Manual (New York: Cold Spring Harbor Laboratory Press, 1989), or under conditions recommended by the manufacturer. Unless otherwise specified, percentages and parts are by weight. Ratio and weight parts.

Unless otherwise defined, all professional and scientific terms used herein have the same meanings as those familiar to those skilled in the art. In addition, any methods and materials similar or equivalent to those described herein can be applied to the methods of the present invention. The preferred implementation methods and materials described herein are for demonstration purposes only.

Preparation Example

Example 1: Synthesis of Compound T-1

The synthetic route of compound T-1 is as follows:

The specific steps are as follows:

Step 1:

Compound SM 1 (5.0 g, 1.0 eq) was dissolved in ultra-dry tetrahydrofuran solvent, placed in an ice-water bath, and solid cesium carbonate (17.1 g, 2.5 eq) was added. 3-chloropropylamine hydrochloride (2.7 g, 1.0 eq) was added under stirring. After the addition was completed, the temperature was naturally raised to room temperature for reaction. After the reaction was completed by TLC monitoring, the solid was removed by suction filtration, the filter cake was washed with an appropriate amount of EA, and water was added to extract the liquid. The organic phase was dried over anhydrous sodium sulfate, filtered and concentrated under reduced pressure, and purified by column chromatography to obtain a yellow solid SM 2 (4.7 g, 71.9%). ¹ H NMR (400 MHz, chloroform-d) δ7.35 (s, 1H), 6.81 (t, J = 2.0 Hz, 1H), 6.64 (dd, J = 12.0, 2.0 Hz, 1H), 3.67 (t, J = 4.0 Hz, 2H), 3.46 (q, J = 8.0 Hz, 2H), 2.16 (p, J = 8.0 Hz, H).

Step 2:

Compound SM 2 (4.3 g, 1.0 e.q.) was dissolved in a mixed solvent of methanol and water (v:v=4:1), iron powder (3.9 g, 5.0 e.q.) and solid ammonium chloride (1.5 g, 2.0 e.q.) were added, and the temperature was raised to reflux for reaction. TLC monitoring, after the reaction was completed, the mixture was cooled to room temperature, filtered, and the filter cake was washed with an appropriate amount of EA. The filtrate was concentrated under reduced pressure to remove most of the solvent, and extracted and separated with saturated sodium bicarbonate solution and EA. The organic phase was dried over anhydrous sodium sulfate, filtered and concentrated under reduced pressure, and purified by column chromatography to obtain compound SM 3 (2.0 g, 51.4%).

Step 3:

Compound SM 3 (1.3 g, 1.0 eq) and CDI (2.2 g, 2.5 eq) were dissolved in ultra-dry DMF solvent, heated to 50 °C for reaction, and after the reaction was completed, the mixture was cooled to room temperature and extracted with EA and water. The organic phase was precipitated with anhydrous sodium sulfate. After drying, suction filtration and concentration under reduced pressure, the product was separated and purified by column chromatography to obtain compound SM 4 (1.2 g, 83.3%).

Step 4:

Compound SM 4 (4.0 g, 1.0 eq) was dissolved in phosphorus oxychloride (40 mL), heated to 110 degrees Celsius and reacted for 24 hours. After the reaction was completed, the mixture was cooled to room temperature and slowly poured into ice water to quench. The mixture was extracted with EA and the organic phase was dried over anhydrous sodium sulfate. After filtration, the mixture was concentrated under reduced pressure and purified by column chromatography to obtain compound SM 5 (137 mg, 24.9%). ¹ H NMR (400 MHz, chloroform-d) δ 6.98 (dd, J = 10.0, 1.7 Hz, 1H), 6.93 (d, J = 1.7 Hz, 1H), 3.93 (t, J = 6.2 Hz, 2H), 3.48-3.31 (m, 2H), 3.22 (s, 3H), 2.26 (p, J = 6.0 Hz, 2H).

Step 5:

Compound SM 5 (8.0 g, 1.0 eq) was dissolved in 4.0 M methylamine ethanol solution (80 mL), heated to 80 degrees Celsius and reacted for 24 hours. After the reaction was completed, the mixture was cooled to room temperature, concentrated under reduced pressure to remove the solvent, extracted with EA and water, and the organic phase was dried over anhydrous sodium sulfate. After filtration, the mixture was concentrated under reduced pressure and purified by column chromatography to obtain compound SM 6 (3.5 g, 50.2%). ¹ H NMR (400 MHz, chloroform-d) δ 6.98 (dd, J = 10.0, 1.7 Hz, 1H), 6.93 (d, J = 1.7 Hz, 1H), 3.93 (t, J = 6.2 Hz, 2H), 3.48-3.31 (m, 2H), 3.22 (s, 3H), 2.26 (p, J = 6.0 Hz, 2H).

Step 6:

Under nitrogen protection, compound SM 6 (0.26 g, 1.0 eq), biboronic acid pinacol ester (0.46 g, 2.0 eq), Pd(dppf)Cl ₂ (34 mg, 0.05 eq) and potassium acetate (0.27 g, 3.0 eq) were added to a dry 50 mL three-necked flask in sequence, replaced with nitrogen three times, and ultra-dry 1,4-dioxane solvent was added, and the temperature was raised to reflux for reaction. TLC monitoring, after the reaction was completed, cooled to room temperature, Ea and water were extracted and separated, the organic phase was dried over anhydrous sodium sulfate, filtered and concentrated under reduced pressure, and purified by column chromatography to obtain compound SM7 (0.21 g, 69.3%). LCMS: [M+H] ⁺ = 332.6.

Step 7:

Under nitrogen protection, compound SM7 (0.21 g, 1.0 eq), 2,4,5-trichloropyrimidine (0.21 g, 2.0 eq), tetrakistriphenylphosphine palladium (71.5 mg, 0.1 eq) and sodium carbonate solid (0.2 g, 3.0 eq) were added to a 50 mL three-necked flask, and a mixture of 1,4-dioxane and water (v:v=10:3) was added. The nitrogen was replaced three times and the temperature was raised to reflux for reaction. TLC monitoring, after the reaction was completed, the mixture was cooled to room temperature, and Ea and water were extracted and separated. The organic phase was dried over anhydrous sodium sulfate, filtered and concentrated under reduced pressure, and purified by column chromatography to obtain compound SM8 (0.13 g, 59.2%). LCMS: [M+H] ⁺ = 352.5.

Step 8:

Compound SM8 (120 mg, 1.0 eq), pyrazolo[1,5-a]pyridine-5-amine (54.5 mg, 1.2 eq), palladium acetate (7.7 mg, 0.1 eq), BINAP (42.5 mg, 0.2 eq) and cesium carbonate (333.2 mg, 3.0 eq) were added to a 25 mL three-necked flask, 1,4-dioxane was added, nitrogen was replaced, and the temperature was raised to reflux for reaction. TLC monitoring, after the reaction was completed, cooled to room temperature, EA and water were used for extraction and separation, the organic phase was dried over anhydrous sodium sulfate, filtered, concentrated under reduced pressure and purified by column chromatography to obtain compound T-1 (70 mg, 45.8%). ¹ H NMR (400MHz, DMSO-d ₆ )δ10.24(s,1H),8.68(s,1H),8.58(d,J＝8.0Hz,1H),8.32(d,J＝2.3Hz,1H ),7.88(d,J＝2.0Hz,1H),7.58(d,J＝1.2Hz,1H),7.45(dd,J＝12.0,1.2Hz,1 H),7.09(dd,J＝7.5,2.4Hz,1H),6.41(d,J＝2.2Hz,1H),4.06(t,J＝6.0Hz,2H),3.41(t,J＝6.0Hz,2H),3.14(s,3H),2.19(t,J＝5.9Hz,2H).LC-MS[M+H] ⁺ =449.3.

Example 2-43: Synthesis of Compounds T-2 to T-43

Referring to the synthesis method of Example 1, the corresponding raw materials were used to synthesize the compounds shown in the following table:

Example 2: Synthesis of Compound T-81

The synthetic route is as follows:

The experimental process is as follows:

Step 1: Compound SM 1 (25.0 g, 1.0 eq) and DMF (2.16 mL, 0.1 eq) were dissolved in 500 mL of dry dichloromethane solvent, cooled to 0°C in an ice-water bath, and SOCl ₂ (30.5 mL, 1.5 eq) was slowly added dropwise. After the addition was completed, the temperature was raised to 40°C for reaction for 3 hours, and then the temperature was restored to room temperature for reaction, and a white solid precipitated. After TLC monitoring, the reaction was completed, and most of the solvent was removed by vacuum concentration. A white solid filter cake was obtained by suction filtration. The filter cake was washed with ethyl acetate and petroleum ether solvents in turn, and the product SM 2 (34 g, 84.2%) was obtained after drying. ¹ H NMR (400 MHz, deuterated methanol) δ 3.81-3.62 (m, 2H), 3.52 (m, 1H), 2.16 (m, 1H), 2.02 (m, 1H), 1.34 (d, J = 6.6 Hz, 3H).

Step 2: Dissolve compound SM 3 (56.0 g, 1.0 e.q.) in 500 mL of ultra-dry DMF solvent, add potassium carbonate solid (97.6 g, 3.0 e.q.), and add SM 2 (34.0 g, 1.0 e.q.) under stirring. After the addition is completed, the temperature is raised to 80°C for reaction. After the reaction is completed by TLC monitoring, the solid is removed by suction filtration, the filter cake is washed with an appropriate amount of EA, and water is added to extract the liquid. The organic phase is dried over anhydrous sodium sulfate, filtered and concentrated under reduced pressure, and purified by column chromatography to obtain a yellow solid SM 4 (58.5 g, 76.5%).

Step 3: Dissolve compound SM 4 (58.5 g, 1.0 eq) in a mixed solvent of ethanol and water (v:v = 3:1), add iron powder (60.3g, 6.0eq) and solid ammonium chloride (19.2g, 2.0eq), heated to reflux for 1 hour under nitrogen protection. TLC monitoring, after the reaction, cooled to room temperature, filtered, the filter cake was washed with an appropriate amount of EA, the filtrate was concentrated under reduced pressure to remove most of the solvent, extracted with saturated sodium bicarbonate solution and EA, the organic phase was dried over anhydrous sodium sulfate, filtered and concentrated under reduced pressure, and purified by column chromatography to obtain compound SM 5 (48.4g, 91.7%).

Step 4: Compound SM 5 (48.0 g, 1.0 e.q.) and CDI (66.3 g, 2.5 e.q.) were dissolved in 720 mL of ultra-dry DMF solvent, heated to 50 °C for reaction, and after the reaction was completed under TLC monitoring, cooled to room temperature, extracted with EA and water, and the organic phase was dried over anhydrous sodium sulfate, filtered and concentrated under reduced pressure, and purified by column chromatography to obtain compound SM 6 (48.0 g, 91.9%).

Step 5: Add compound SM 6 (48 g, 1.0 e.q.) to 167 mL of phosphorus oxychloride, heat to 100 °C and react for 18 hours. Monitor by TLC. After the reaction is completed, cool to room temperature, slowly pour into ice water to quench, extract with EA, dry the organic phase with anhydrous sodium sulfate, filter and concentrate under reduced pressure, separate and purify by column chromatography to obtain compound SM 7 (45.5 g, 89.7%).

Step 6: Add SM 7 (1.0 g, 1.0 e.q.), 2.0 M ethanol solution of methylamine (10 mL, 10.0 V), sodium iodide (2.2 g, 5.0 e.q.) and DMF (20 mL, 20 V) to a 100 mL jar, seal and heat to 70 ° C, and react for 12 hours. After the reaction is completed by TLC detection, cool to room temperature, extract with EA and water, and wash the organic phase three times with saturated brine, dry with anhydrous sodium sulfate, filter, concentrate the organic phase under reduced pressure, and separate and purify by silica gel column chromatography to obtain the target product SM 8 (400 mg, 47.8%), LCMS: [M+H] + = 284.1, 286.1.

Step 7: Under nitrogen protection, add 60% NaH solid powder (35.6 mg, 1.1 e.q.) to 15 mL of ultra-dry THF, place in an ice-water bath and add SM 8. After half an hour of reaction, add deuterated iodomethane THF solution (117 mg, 1.0 e.q.), and keep warm for 2 hours after the addition. After the reaction is completed by TLC monitoring, add appropriate amount of ice water to quench, extract with saturated ammonium chloride solution and EA, dry the organic phase with anhydrous sodium sulfate, filter and concentrate under reduced pressure, separate and purify by silica gel column chromatography to obtain the target compound SM 9 (200 mg, 82.3%), LCMS: [M+H]+＝301.2, 303.2.

Step 8: Under nitrogen protection, compound SM 9 (240 mg, 1.0 e.q.), bipyraclostrobin (406 mg, 2.0 e.q.), Pd(dppf)Cl2 (29 mg, 0.05 e.q.) and potassium acetate (235 mg, 3.0 e.q.) were added to a dry 100 mL three-necked flask in sequence, replaced with nitrogen three times, and 10 mL of ultra-dry 1,4-dioxane solvent was added, and the temperature was raised to 110 ° C for reaction. TLC monitoring, after the reaction was completed, cooled to room temperature, EA and water were used for extraction and separation, the organic phase was dried over anhydrous sodium sulfate, filtered and concentrated under reduced pressure, and purified by column chromatography to obtain compound SM 10 (200 mg, 72.7%), LCMS: [M+H]+＝349.4.

Step 9: Under nitrogen protection, compound SM 10 (220 mg, 1.0 e.q.), 2,4,5-trichloropyrimidine (0.11 mL, 1.5 e.q.), tetrakistriphenylphosphine palladium (36 mg, 0.05 e.q.) and sodium carbonate solid (201 mg, 3.0 e.q.) were added to a 50 mL three-necked flask in sequence, and a mixture of 1,4-dioxane and water (v:v=10:3) was added. The nitrogen was replaced three times and the temperature was raised to 110°C for reaction. TLC monitoring, after the reaction was completed, the mixture was cooled to room temperature, and the mixture was extracted and separated by Ea and water. The organic phase was dried over anhydrous sodium sulfate, filtered and concentrated under reduced pressure, and purified by column chromatography to obtain compound SM 11 (100 mg, 42.9%), LCMS: [M+H]+=369.3.

Step 10: Compound SM 11 (100 mg, 1.0 eq), pyrazolo[1,5-a]pyridine-5-amine (43.3 mg, 1.2 eq), palladium acetate (6.1 mg, 0.1 eq), BINAP (33.7 mg, 0.2 eq) and cesium carbonate (264.4 mg, 3.0 eq) were added to a 25 mL three-necked flask, 1,4-dioxane was added, nitrogen was replaced, and the temperature was raised to reflux for reaction. TLC monitoring, after the reaction was completed, cooled to room temperature, EA and water were used for extraction and separation, the organic phase was dried over anhydrous sodium sulfate, filtered, concentrated under reduced pressure and purified by column chromatography to obtain compound T-81, LCMS: [M+H]+=466.3. 1H NMR (400 MHz, chloroform-d) δ8.40 (s, 1H), 8.30 (d,J＝7.5Hz,1H),8.10(s,1H),7.83(s,1H),7.49(d,J＝11.9Hz,1H),7.45(s,1H),7.30(s,1H),6.75(s,1H),6.30(s,1H),4.57-4.3 9(m,1H),3.54(d,J＝3.2Hz,1H),3.24(d,J＝7.5Hz,1H),2.34(tt,J＝12.3,5.0Hz,1H),1.95(d,J＝16.3Hz,1H),1.46(d,J＝6.6Hz,3H).

Test Example 1 Enzyme Activity Test

The following is a biological activity test experiment on some compounds in the above examples and comparative examples.

The biological activity test experimental process is as follows:

1. Kinase activity test:

The IC ₅₀ values of the test compounds for CDK2 and CDK4 kinases were detected.

(I) Reagent information

(II) Equipment information

3. Research design

(1) Compound preparation:

① Prepare the test compound into a 0.5 mM DMSO solution, and prepare the positive control drug palbociclib into a 0.5 mM DMSO solution.

②Three-fold dilution to obtain 10 compound solutions of different concentrations.

(2) Enzyme assay:

① Prepare a 1.3x enzyme solution containing enzyme, substrate, and cofactors as shown below.

② Add 15 μL of 1.3x enzyme solution to each well and incubate at room temperature for 30 minutes.

③ Add 5 μL of 4x ATP solution to start the reaction. Each test well contains the ingredients in the list and the final volume is 20 μL.

④ Incubate for 60 minutes, then add 75 μL of buffer (containing 0.5 M EDTA) to stop the reaction.

⑤Use EZ to read the data of each test hole for analysis.

(3) Data analysis:

The percent inhibition was calculated using the concern ratio (CR) according to the following formula:

The wells treated with DMSO were used as positive controls, and the wells without enzyme were used as negative controls.

% (inhibition percentage) = 100-100*((CRPC-CRSample)/(CRPC-CRNC)).

Through the above detection, the inhibitory activity IC ₅₀ (nM) values of the tested samples on CDK2 and CDK4 kinases are shown in Table 1.

Table 1

As can be seen from the above table, through in vitro biological activity screening, with palbociclib as the reference substance, the compounds synthesized in the present application have a good inhibitory ability against CDK2/4 kinases, and are expected to be further developed into drugs for regulating CDK2/4 protein kinase activity or treating CDK2/4 related diseases.

Experimental Example 2 Cell Antiproliferation Experiment

1. Experimental materials and equipment:

Human breast cancer cells MCF-7, ovarian cancer cells A2780, human ovarian adenocarcinoma cells OVCAR3. DMEM medium (Bio-Channel), DMSO (dimethyl sulfoxide), MTT (thiazolyl blue), 0.25% EDTA-Tripsin (trypsin digestion solution), 1xPBS (phosphate buffer, pH 7.2), 96-well plate (Corning), fetal bovine serum (FBS), 10,000U/mL penicillin-G/streptomycin, high-speed refrigerated centrifuge (EPPENDORF 5810R), enzyme-linked immunosorbent assay (Tecan Spark).

2. Experimental preparation:

1. Cell plating

A) Tumor cells were cultured in DMEM (high glucose, containing 10% FBS and 100 U/mL penicillin-G/streptomycin) at 37°C, 5% _CO2 and saturated humidity to 80-90% confluence.

B) Remove the culture medium from the 10 cm culture dish;

C) Rinse the cells once with 10 ml 1xPBS;

D) Add 4 ml of 0.25% EDTA-Tripsin and place in a 37°C, 5% CO ₂ incubator for trypsin digestion for 5 minutes, transfer to a 15 ml centrifuge tube, centrifuge at 200 g for 5 minutes, and discard the supernatant to obtain the cell pellet;

E) Resuspend in 4 ml DMEM medium, count and adjust to 50,000 cells/ml.

F) The cell suspension was added to each well of a 96-well plate at a volume of 100 μL and cultured overnight in a 37°C, 5% CO ₂ incubator.

2. Compound treatment

Compound dilution

A) Prepare gradient dilution solutions of the test compound: Prepare the test compound into a 1mM stock solution. Then dissolve 1.5μl of the stock solution in 1.5ml of DMSO-free culture medium, and then perform 3-fold continuous gradient dilutions with 0.1% DMSO culture medium, for a total of 9 concentrations. The compound concentrations after dilution are as follows:

333.33nM, 111.11nM, 37.03nM, 12.35nM, 4.15nM, 1.37nM, 0.46nM, 0.15nM

B) After thorough mixing, 100 μL of the culture compound solution was taken to replace the culture medium in the cell culture plate, with 4 replicate wells for each concentration;

C) The cells were transferred to an incubator and incubated for 5 days.

3. MTT assay

A) Take out the cell culture plate and add 10μL of 5mg/ml MTT in the biosafety cabinet;

B) Place the cell culture plate back into the incubator and continue incubating for 3 hours;

C) Take out the cell culture plate, remove the culture medium, add 100 μL of isopropanol (containing 0.4 mM HCl, 0.1% NP-40), and shake at room temperature for 30 minutes;

D) Select 570 nm wavelength to measure the absorbance value on TECAN enzyme-linked immunosorbent assay instrument.

4. Data Analysis

The following formula is used to calculate the cell viability (% Cell Viability):

%Cell Viability＝100%×(Lum_Sample-Lum_LC)/(Lum_HC-Lum_LC)

Lum_HC: 0.1% DMSO control group cell reading

Lum_Sample: Cell readout with added compound

Lum_LC: Blank medium reading

The _IC50 value (in nM) was obtained by curve fitting using GraphPad Prism 8 software.

As shown in Table 2.

Table 2

It can be seen from Table 2 that the compounds of the present invention have excellent anti-proliferation activity against breast cancer cells, ovarian cancer cells and human ovarian adenocarcinoma cells.

Experimental Example 3 Preclinical rat pharmacokinetic study

1. Experimental materials and equipment:

Healthy adult SD rats, male, 6-8 weeks old, weighing 200-300 g, were purchased from XX Company. EDTA-Na2 anticoagulant. Analytical balance, animal weighing scale, magnetic stirrer, refrigerated centrifuge, single-channel manual pipette, etc.

2. Experimental process:

1. Drug preparation

Accurately weigh about 10 mg of the sample to be tested, add 5% DMSO to dissolve it, then add 10% solutol HS-15 and 85% saline, sonicate, and vortex to mix to obtain a solution with a concentration of 1 mg/mL; prepare freshly before use.

Pipette 0.2 mL of sample into a 1.5 mL centrifuge tube and store at -80°C for concentration analysis of the dosing solution.

2. Animal Preparation

The animals were kept in rat cages and fasted from the day before the experiment (for no less than 10 hours), but water was not allowed. They were weighed and marked on the tail on the day of the experiment. Blank blood was collected before administration. Blood was collected from the tail vein.

3. Drug administration

Route of administration: oral administration (p.o.)

Dosage: 10 mg/kg

Dosing volume: 10mL/kg

Operation procedure: Use your left hand wearing a bite-proof glove to hold the rat upright, insert a No. 16 gavage needle into the rat's mouth and throat, and test to see if there is any obvious resistance when inserting the needle, then inject the drug into the stomach.

4. Sample collection

Before administration and 0.5h, 1h, 2h, 4h, 6h, 8h, 12h, and 24h after administration, 0.1ml of whole blood was collected from the test animals in EDTA-Na2 anticoagulant tubes, inverted 3-4 times to mix, centrifuged at 4℃ 10000g for 5min to separate plasma, and stored at -80℃ for testing. Blood was collected from the tail vein.

The specific operation is to fix the rat on the fixator so that its tail can be completely exposed, wipe the rat's tail with alcohol, let the surface skin absorb the alcohol, achieve the effect of obvious venous dilation, select suitable veins from both sides, and insert the needle about one-third of the distance from the tail tip. The syringe uses an insulin syringe, and the needle is inserted with the bevel upward. After piercing the skin, it is immediately parallel. You can feel that the needle has little sliding resistance in the vein, and there is blood returning in the syringe. It has entered the vein and extracted about 0.1-0.2ml of whole blood. After pulling out the needle, press to stop bleeding.

3. Sample analysis:

Preparation of standard curve: Pipette 25 μL of blank rat plasma into a centrifuge tube, add 25 μL of the prepared standard series solution (prepared in methanol), then add 200 μL of internal standard solution (prepared in methanol), vortex mix for 2 min, and centrifuge at 4°C, 10000g for 10 min.

Treatment of unknown plasma samples: 25 μL of rat drug-containing plasma was taken, and 25 μL of methanol and 200 μL of internal standard solution were added in sequence, vortexed for 2 min, and centrifuged at 10000 g for 10 min at 4 °C. The supernatant was taken for LC/MS/MS detection.

4. Data Processing

The quantitative detection method of the test compound was established using Shimadzu liquid chromatography and Triple Quad TM 6500+AB mass spectrometry. The concentration of the original drug in plasma was measured. The blood drug concentration-time curve was drawn, and the non-compartmental model in Winnonlin Phoenix software was used to calculate the main pharmacokinetic parameters. The results are shown in Table 3.

Table 3

It can be seen from Table 3 that the compounds of the present invention all have excellent pharmacokinetic properties.

All documents mentioned in the present invention are cited as references in this application, just as each document is cited as reference individually. In addition, it should be understood that after reading the above teachings of the present invention, those skilled in the art can make various changes or modifications to the present invention, and these equivalent forms also fall within the scope defined by the claims attached to this application.

Claims (13)

A compound, characterized in that the compound is a compound of formula I, or a pharmaceutically acceptable salt, stereoisomer, tautomer, hydrate, solvate, isotope compound or prodrug thereof,
in: X is N or CR ₅ ; _X1 is N or _CR5 ; R ₁ is selected from the group consisting of H, halogen, cyano, C _1-6 alkyl, C _1-6 haloalkyl, C _1-6 alkoxy, C _3-6 cycloalkyl; _R2 is as shown in Formula Ia or Ib;
in, W ₃ , W ₄ and W ₅ are each independently selected from: CH, N, NH; One of _W2 and _W6 is C, and the other is C or N; R ₆ is each independently selected from the group consisting of halogen, C _1-6 alkyl, C _1-6 haloalkyl, C _1-6 hydroxyalkyl, C _3-6 cycloalkyl, -CO-C _1-6 alkyl, -COO-C _1-6 alkyl, 4 to 6-membered heterocyclyl, -C _1-4 alkylene-C _3-6 cycloalkyl, -C _1-4 alkylene-4 to 6-membered heterocyclyl; and wherein the alkyl, alkylene, cycloalkyl and heterocyclyl are optionally substituted by one or more substituents selected from the group consisting of halogen, hydroxyl, cyano, C _1-6 alkyl, C _1-6 haloalkyl, C _1-6 alkoxy; The subscript p is 0, 1, or 2; _W1 and _W7 are each independently CH or N; Subscript o is 0 or 1; R ₇ is each independently selected from the group consisting of halogen, C _1-6 alkyl, C _1-6 haloalkyl; Subscript q is 0, 1, 2, or 3; R ₃ is independently selected from the following group: H, C _1-6 alkyl, C _3-6 cycloalkyl, hydroxyl, amino, C _1-6 alkoxy (such as methoxy), C _1-6 haloalkyl (such as CF ₃ ), C _1-6 haloalkoxy, C _1-6 hydroxyalkyl, 5-8 membered heterocyclyl, 5-8 membered heteroaryl; or, two R ₃ located on the same atom together form an oxo group (=O); R ₄ is selected from the group consisting of H, C _1-6 alkyl, -CO-C _1-6 alkoxy, -COC _1-6 alkyl, C _3-8 cycloalkyl, and a 4-8 membered heterocycloalkyl containing 1, 2 or 3 heteroatoms selected from N, O or S; wherein the alkyl, aryl, cycloalkyl and heterocycloalkyl are optionally substituted by one or more groups selected from the group consisting of halogen, cyano, hydroxyl, C _1-6 alkyl, -CO-C _1-6 alkyl, C _1-6 alkoxy _, amino, -COOH, -CONH ₂ , -CF ₃ C(F) ₂ , a 1, ₂ or ₃ membered _{heterocycloalkyl} containing 1 _{, 2} or 3 heteroatoms selected from N, O or S; or a 5-8 membered heterocycloalkyl group having 3 heteroatoms selected from N, O or S; R ₅ is selected from the group consisting of H, hydroxy, halogen, cyano, C _1-6 alkyl, C _1-6 haloalkyl, -CO-C _1-6 alkoxy; The subscript m is selected from the following group: 0, 1, 2, 3, 4; Subscript n is 0, 1, 2, 3 or 4. The method according to claim 1, characterized in that X is N or CR ₅ ; _X1 is N or _CR5 ; R ₁ is selected from the group consisting of H, halogen, C _1-6 alkyl, C _1-6 haloalkyl, C _3-6 cycloalkyl; _R2 is as shown in Formula Ia or Ib;
in, W ₃ , W ₄ and W ₅ are each independently selected from: CH, N, NH; One of _W2 and _W6 is C, and the other is C or N; R ₆ is each independently selected from the group consisting of halogen, C _1-6 alkyl, C _1-6 haloalkyl, C _1-6 hydroxyalkyl, C _3-6 cycloalkyl, 4 to 6 membered heterocyclyl, -C _1-4 alkylene-C _3-6 cycloalkyl, -C _1-4 alkylene-4 to 6 membered heterocyclyl; and wherein the alkyl, alkylene, cycloalkyl and heterocyclyl are optionally substituted by one or more substituents selected from the group consisting of halogen, hydroxyl, cyano, C _1-6 alkyl, C _1-6 haloalkyl, C _1-6 alkoxy; The subscript p is 0, 1, or 2; _W1 and _W7 are each independently CH or N; Subscript o is 0 or 1; R ₇ is each independently selected from the group consisting of halogen, C _1-6 alkyl, C _1-6 haloalkyl; Subscript q is 0, 1, 2, or 3; R ₃ is each independently selected from the group consisting of H, C _1-6 alkyl, C _3-6 cycloalkyl, hydroxyl, C _1-6 alkoxy; R ₄ is selected from the group consisting of H, C _1-6 alkyl, -CO-C _1-6 alkoxy, -COC _1-6 alkyl; R ₅ is selected from the group consisting of H, C _1-6 alkyl, -CO-C _1-6 alkoxy; The subscript m is selected from the following group: 0, 1, 2, 3, 4; Subscript n is 0, 1, 2, 3 or 4. The compound according to claim 1 or 2, characterized in that _R2 is as shown in Formula Ia. The compound according to claim 1 or 2, characterized in that In formula (Ia), _W1 is CH, _W2 is C, _W3 is NH, _W4 is CH, _W5 is N, _W6 is C and _W7 is CH; or, _W1 is CH, _W2 is C, _W3 is CH, _W4 is CH, _W5 is N, _W6 is N and _W7 is CH; or, _W1 is CH, _W2 is C, _W3 is N, _W4 is CH, _W5 is N, _W6 is N and _W7 is CH; or, _W1 is CH, _W2 is N, _W3 is CH, _W4 is CH, _W5 is N, _W6 is C and _W7 is N; or, _W1 is CH, _W2 is CH, _W3 is N, _W4 is CH, _W5 is CH, _W6 is N and _W7 is CH; or, _W1 is CH, _W2 is CH, _W3 is N, _W4 is N, _W5 is CH, _W6 is N and _W7 is N; or, _W1 is CH, _W2 is C, _W3 is NH, _W4 is CH, _W5 is N, _W6 is C and _W7 is N; or, _W1 is CH, _W2 is C, _W3 is N, _W4 is CH, _W5 is N, _W6 is N and _W7 is N; or, _W1 is CH, _W2 is C, _W3 is NH, _W4 is CH, _W5 is CH, _W6 is C and _W7 is CH; or, _W1 is CH, _W2 is C, _W3 is NH, _W4 is N, _W5 is CH, _W6 is C and _W7 is CH; or, _W1 is CH, _W2 is C, _W3 is NH, _W4 is CH, _W5 is N, _W6 is C and _W7 is CH; or, _W1 is CH, _W2 is C, _W3 is CH, _W4 is N, _W5 is NH, _W6 is C and _W7 is CH; or, _W1 is CH, _W2 is C, _W3 is N, _W4 is N, _W5 is NH, _W6 is C and _W7 is CH; or, _W1 is CH, _W2 is C, _W3 is CH, _W4 is NH, _W5 is N, _W6 is C and _W7 is CH; or, _W1 is N, _W2 is C, _W3 is CH, _W4 is NH, _W5 is N, _W6 is N and _W7 is CH; or, _W1 is CH, _W2 is C, _W3 is CH, _W4 is CH, _W5 is N, _W6 is N and _W7 is N; In formula (Ib), _W2 is C, _W3 is CH, _W4 is CH, _W5 is N, _W6 is N and o is 0 or 1. The compound according to claim 1 or 2, characterized in that Select from the following group:

The compound according to claim 1 or 2, characterized in that Select from the following group:
The compound according to claim 1 or 2, characterized in that X is N, _X1 is N, and the subscript n is 0 or 1. The compound according to claim 1 or 2, characterized in that the compound has one or more of the following characteristics: R ₁ is selected from the group consisting of H, F, Cl, methyl, Br, CF ₃ , _R3 is selected from the group consisting of H, methyl, cyclopropyl, hydroxyl, methoxy; _R4 is selected from the group consisting of H, methyl, ethyl, The compound according to claim 1 or 2, characterized in that the isotope compound is a deuterated compound. The compound according to claim 1, characterized in that the compound is selected from Table A: Table A



A pharmaceutical composition, characterized in that it contains a preventive and/or therapeutically effective amount of the compound according to any one of claims 1 to 10, and a pharmaceutically acceptable carrier. A use of the compound according to claim 1 or the pharmaceutical composition according to claim 11, characterized in that it is used to prepare one or more drugs selected from the following group: (a) A drug used as a CDK2/4 protein kinase inhibitor; (b) drugs for modulating CDK2/4 protein kinase activity; (c) Drugs for preventing or treating CDK2/4 related diseases. The use according to claim 12, characterized in that the CDK2/4-related disease is selected from the group consisting of inflammation, cancer, cardiovascular disease, infection, immune disease, and metabolic disease.