Compound used as CDK4 kinase inhibitor and application thereof

Abstract

The invention relates to a compound used as a CDK4 kinase inhibitor and application thereof. Specifically, the compound disclosed by the invention has a structure as shown in a formula II, and the definitions of all groups and substituent groups are described in the specification. The compound provided by the invention can be used as an inhibitor of cyclin-dependent kinase (CDK), is used for treating or preventing proliferative diseases (such as cancers), and is particularly used for regulating and treating related diseases caused by abnormal activity of the cyclin-dependent kinase (CDK). # imgabs0 #

Description

Technical Field

[0001] The present invention relates to the field of medical technology, and in particular to compounds used as CDK4 kinase inhibitors, and applications thereof in regulating CDK4 kinase activity or treating CDK4-related diseases, especially cancer. Background Art

[0002] Cyclin-dependent kinases (CDKs) belong to the serine / threonine kinase family. They exert their physiological functions by binding to corresponding cyclins to form active dimer complexes, causing cell growth and proliferation. More than 20 CDKs have been discovered, which are divided into two categories according to their functions: CDKs that regulate the cell cycle and CDKs that regulate cell transcription. Among them, CDKs 1-6 and 14-18 are involved in the regulation of the cell cycle, and CDKs 7-13 and 19-20 are involved in the transcriptional regulation of cells. It has now been proven that CDK inhibitors can be used to treat cancer.

[0003] CDK4 and CDK6 participate in regulating the cell cycle from G1 to S phase after binding to Cyclin D. It is reported that abnormalities in the CyclinD-CDK4 / 6-Rb pathway are associated with the progression of resistance to endocrine therapy. At present, a variety of CDK4 / 6 inhibitors, such as palbociclib, ribociclib, and abemaciclib, have been approved for marketing and are used in combination with endocrine therapy to treat hormone receptor (HR)-positive, human epidermal growth factor 2 (HER2)-negative advanced or metastatic breast cancer. However, hematological toxicity and / or gastrointestinal toxic side effects such as neutropenia often occur during treatment with CDK4 / 6 inhibitors, 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 cyclin D3-CDK6 may be related to these side effects. In view of the toxic side effects of the current CDK4 / 6 dual-target inhibitors, the development of a selective CDK4 inhibitor may have better safety and efficacy. Summary of the invention

[0004] The present invention provides a novel compound having CDK4 kinase inhibitory activity and better pharmacodynamics and pharmacokinetic properties.

[0005] In a first aspect of the present invention, a compound used as a CDK4 kinase inhibitor is provided, wherein the compound is a compound of formula II, or a pharmaceutically acceptable salt, stereoisomer, tautomer, hydrate, solvate, isotope compound or prodrug thereof,

[0006]

[0007] in:

[0008] X1 is selected from the following group: N, CR3;

[0009] R1 is selected from the group consisting of H, CF3, F, Cl, Br, methyl, ethyl, isopropyl, cyclopropyl;

[0010] R2 is selected from the group consisting of H, F, CF3, CF2H, NH2, methyl;

[0011] Or R1 and R2 together with the C to which they are connected form a 5-membered heteroaryl group containing 2 N;

[0012] Each R3 is independently selected from the group consisting of H, CF3, F, Cl, Br, methyl, cyano;

[0013] Ring A is selected from the following group: Wherein, X2 is O or NR, X3 is N or CR3, and X4 is NR, SO or SO2;

[0014] Ring B is selected from the group consisting of:

[0015] wherein each R4 is independently selected from the group consisting of H, cyano, halogen, substituted or unsubstituted C1-C6 alkyl, substituted or unsubstituted C3-C6 cycloalkyl;

[0016] Each R5, R6, R9, R 10 and R 14 Each independently selected from the group consisting of H, halogen, hydroxyl, amino, Oxo, substituted or unsubstituted C1-C6 alkyl, substituted or unsubstituted C3-C6 cycloalkyl, substituted or unsubstituted phenyl, substituted or unsubstituted C 1-6 Alkoxy, substituted or unsubstituted -NR m R n ; or two R connected to the same C 14 Together with the C to which it is attached, it forms a C3-C6 cycloalkyl group;

[0017] or R9, R 10 Together with its connected N, it forms

[0018] Each R and R' is independently selected from the group consisting of H, C1-C6 alkyl, halogenated C1-C6 alkyl, C3-C6 cycloalkyl, halogenated C3-C6 cycloalkyl, phenyl;

[0019] Each R m Selected from the following group: H, C 1-6 Alkyl, C 3-6 Cycloalkyl, C 6-10 Aryl, 6-10 membered heteroaryl containing 1, 2 or 3 heteroatoms selected from N, O and S;

[0020] Each R n C 1-6 alkyl;

[0021] R4, R5, R6, R9, R 10 and R 14 The substitutions described in each of the above independently refer to substitutions by 1, 2 or 3 substituents selected from the group consisting of deuterium, halogen, hydroxy, amino, -N-(C1-C6 alkyl)2, C1-C6 alkoxy, phenyl, cyano;

[0022] Each m, n, and p are independently selected from the following group: 0, 1, 2, 3, 4;

[0023] q is selected from the following group: 1, 2, 3.

[0024] In another preferred embodiment,

[0025] R1 is selected from the group consisting of H, CF3, F, Cl, Br, methyl, ethyl, isopropyl, cyclopropyl;

[0026] R2 is selected from the group consisting of H, F, CF3, CF2H, NH2, methyl;

[0027] Ring A is selected from the following group: Wherein, X2 is O or NR, X3 is N or CR3, and X4 is NR, SO or SO2;

[0028] Ring B is selected from the group consisting of:

[0029] wherein each R4 is independently selected from the group consisting of H, cyano, halogen, substituted or unsubstituted C1-C6 alkyl, substituted or unsubstituted C3-C6 cycloalkyl;

[0030] Each R5, R6, R9, R 10 and R 14 Each independently selected from the group consisting of H, halogen, hydroxyl, amino, Substituted or unsubstituted C1-C6 alkyl, substituted or unsubstituted C3-C6 cycloalkyl, substituted or unsubstituted phenyl;

[0031] or R9, R 10 Together with its connected N, it forms

[0032] R and R' are each independently selected from the group consisting of H, C1-C6 alkyl, halogenated C1-C6 alkyl, C3-C6 cycloalkyl, halogenated C3-C6 cycloalkyl, phenyl;

[0033] R4, R5, R6, R9, R 10 and R 14 The substitutions described in each of the above independently refer to substitutions by 1, 2 or 3 substituents selected from the group consisting of deuterium, halogen, hydroxy, amino, -N-(C1-C6 alkyl)2, C1-C6 alkoxy, phenyl, cyano;

[0034] Each m, n, and p are independently selected from the following group: 0, 1, 2, 3, 4;

[0035] q is selected from the following group: 1, 2, 3.

[0036] In another preferred embodiment,

[0037] R1 is selected from the group consisting of Cl, Br;

[0038] R2 is H;

[0039] Ring A is selected from the following group:

[0040] Ring B is selected from the group consisting of:

[0041] in,

[0042] Each R5, R6, R9, R 10 and R 14 Each independently selected from the group consisting of H, halogen, hydroxyl, amino, Substituted or unsubstituted C1-C6 alkyl, substituted or unsubstituted C3-C6 cycloalkyl, substituted or unsubstituted phenyl;

[0043] or R9, R 10 Together with its connected N, it forms

[0044] R and R' are each independently selected from the group consisting of H, C1-C6 alkyl, halogenated C1-C6 alkyl, C3-C6 cycloalkyl, halogenated C3-C6 cycloalkyl, phenyl;

[0045] R5, R6, R9, R 10 and R 14 The substitutions described in each of the above independently refer to substitutions by 1, 2 or 3 substituents selected from the group consisting of deuterium, halogen, hydroxy, amino, -N-(C1-C6 alkyl)2, C1-C6 alkoxy, phenyl, cyano;

[0046] Each m, n, and p are independently selected from the following group: 0, 1, 2, 3, and 4.

[0047] In another preferred embodiment,

[0048] R1 and R2 together with the C to which they are connected form a 5-membered heteroaryl group containing 2 N;

[0049] Ring A is selected from the following group:

[0050] Ring B is selected from the group consisting of:

[0051] wherein each R4 is independently selected from the following group: H, substituted or unsubstituted C1-C6 alkyl, substituted or unsubstituted C3-C6 cycloalkyl;

[0052] Each R5, R6, R9, R 10 and R 14 Each independently selected from the group consisting of H, halogen, hydroxyl, amino, Substituted or unsubstituted C1-C6 alkyl, substituted or unsubstituted C3-C6 cycloalkyl, substituted or unsubstituted phenyl;

[0053] R and R' are each independently selected from the group consisting of H, C1-C6 alkyl, halogenated C1-C6 alkyl, C3-C6 cycloalkyl, halogenated C3-C6 cycloalkyl, phenyl;

[0054] R4, R5, R6, R9, R 10 and R 14 The substitutions described in each of the above independently refer to substitutions by 1, 2 or 3 substituents selected from the group consisting of deuterium, halogen, hydroxy, amino, -N-(C1-C6 alkyl)2, C1-C6 alkoxy, phenyl, cyano;

[0055] Each m, n, and p are independently selected from the following group: 0, 1, 2, 3, and 4.

[0056] In another preferred embodiment,

[0057] It has the following structure:

[0058] X is selected from the group consisting of N, CR3;

[0059] R3 is selected from the group consisting of H, CF3, F, Cl, Br, methyl, and cyano.

[0060] In a second aspect of the present invention, a compound used as a CDK4 kinase inhibitor 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,

[0061]

[0062] in:

[0063] X1 is selected from the following group: N, CR3;

[0064] R1 is selected from the group consisting of H, CF3, F, Cl, Br, methyl, ethyl, isopropyl, cyclopropyl;

[0065] R2 is selected from the group consisting of H, F, CF3, CF2H, NH2, methyl;

[0066] Or R1 and R2 together with the C to which they are connected form a 5-membered heteroaryl group containing 2 N;

[0067] R3 is selected from the group consisting of H, CF3, F, Cl, Br, methyl, cyano;

[0068] Ring A is selected from the following group:

[0069] Wherein, X2 is O or NR;

[0070] Ring B is selected from the group consisting of:

[0071] wherein each R4 is independently selected from the following group: H, cyano, halogen, substituted or unsubstituted C1-C6 alkyl, substituted or unsubstituted C3-C6 cycloalkyl,

[0072] Each R5, R6, R9, R 10 and R 14 Each independently selected from the group consisting of H, halogen, hydroxyl, amino, Substituted or unsubstituted C1-C6 alkyl, substituted or unsubstituted C3-C6 cycloalkyl, substituted or unsubstituted phenyl;

[0073] or R9, R 10 Together with its connected N, it forms

[0074] Each R and R' is independently selected from the group consisting of H, C1-C6 alkyl, halogenated C1-C6 alkyl, C3-C6 cycloalkyl, halogenated C3-C6 cycloalkyl, phenyl;

[0075] R4, R5, R6, R9, R 10 and R 14 The substitutions described in each of the above independently refer to substitutions by 1, 2 or 3 substituents selected from the group consisting of deuterium, halogen, hydroxy, amino, -N-(C1-C6 alkyl)2, C1-C6 alkoxy, phenyl, cyano;

[0076] Each m, n, and p are independently selected from the following group: 0, 1, 2, 3, and 4.

[0077] In another preferred embodiment,

[0078] R1 is selected from the group consisting of H, CF3, F, Cl, Br, methyl, ethyl, isopropyl, cyclopropyl;

[0079] R2 is selected from the group consisting of H, F, CF3, CF2H, NH2, methyl;

[0080] Ring A is selected from the following group:

[0081] Ring B is selected from the group consisting of:

[0082] wherein each R4 is independently selected from the following group: H, cyano, halogen, substituted or unsubstituted C1-C6 alkyl, substituted or unsubstituted C3-C6 cycloalkyl,

[0083] Each R5, R6, R9, R 10 and R 14 Each independently selected from the group consisting of H, halogen, hydroxyl, amino, Substituted or unsubstituted C1-C6 alkyl, substituted or unsubstituted C3-C6 cycloalkyl, substituted or unsubstituted phenyl;

[0084] or R9, R 10 Together with its connected N, it forms

[0085] R and R' are each independently selected from the group consisting of H, C1-C6 alkyl, halogenated C1-C6 alkyl, C3-C6 cycloalkyl, halogenated C3-C6 cycloalkyl, phenyl;

[0086] R4, R5, R6, R9, R 10 and R 14 The substitutions described in each of the above independently refer to substitutions by 1, 2 or 3 substituents selected from the group consisting of deuterium, halogen, hydroxy, amino, -N-(C1-C6 alkyl)2, C1-C6 alkoxy, phenyl, cyano;

[0087] Each m, n, and p are independently selected from the following group: 0, 1, 2, 3, and 4.

[0088] In another preferred embodiment,

[0089] R1 is selected from the group consisting of CF3, F, Cl, Br;

[0090] R2 is H;

[0091] Ring A is selected from the following group:

[0092] Ring B is selected from the group consisting of:

[0093] wherein each R4 is independently selected from the following group: H, substituted or unsubstituted C1-C6 alkyl, substituted or unsubstituted C3-C6 cycloalkyl,

[0094] Each R5, R6, R9, R 10 and R 14 Each independently selected from the group consisting of H, halogen, hydroxyl, amino, Substituted or unsubstituted C1-C6 alkyl, substituted or unsubstituted C3-C6 cycloalkyl, substituted or unsubstituted phenyl;

[0095] or R9, R 10 Together with its connected N, it forms

[0096] R and R' are each independently selected from the group consisting of H, C1-C6 alkyl, halogenated C1-C6 alkyl, C3-C6 cycloalkyl, halogenated C3-C6 cycloalkyl, phenyl;

[0097] R4, R5, R6, R9, R 10 and R 14The substitutions described in each of the above independently refer to substitutions by 1, 2 or 3 substituents selected from the group consisting of deuterium, halogen, hydroxy, amino, -N-(C1-C6 alkyl)2, C1-C6 alkoxy, phenyl, cyano;

[0098] Each m, n, and p are independently selected from the following group: 0, 1, 2, 3, and 4.

[0099] In another preferred embodiment,

[0100] R1 and R2 together with the C to which they are connected form a 5-membered heteroaryl group containing 2 N;

[0101] Ring A is selected from the following group:

[0102] Ring B is selected from the group consisting of:

[0103] wherein each R4 is independently selected from the following group: H, substituted or unsubstituted C1-C6 alkyl, substituted or unsubstituted C3-C6 cycloalkyl,

[0104] Each R5, R6, R9, R 10 and R 14 Each independently selected from the group consisting of H, halogen, hydroxyl, amino, Substituted or unsubstituted C1-C6 alkyl, substituted or unsubstituted C3-C6 cycloalkyl, substituted or unsubstituted phenyl;

[0105] R and R' are each independently selected from the group consisting of H, C1-C6 alkyl, halogenated C1-C6 alkyl, C3-C6 cycloalkyl, halogenated C3-C6 cycloalkyl, phenyl;

[0106] R4, R5, R6, R9, R 10 and R 14 The substitutions described in each of the above independently refer to substitutions by 1, 2 or 3 substituents selected from the group consisting of deuterium, halogen, hydroxy, amino, -N-(C1-C6 alkyl)2, C1-C6 alkoxy, phenyl, cyano;

[0107] Each m, n, and p are independently selected from the following group: 0, 1, 2, 3, and 4.

[0108] In another preferred embodiment,

[0109] It has the following structure:

[0110] X is selected from the group consisting of N, CR3;

[0111] R3 is selected from the group consisting of H, CF3, F, Cl, Br, methyl, and cyano.

[0112] In another preferred embodiment, the compound is selected from the following group:

[0113]

[0114]

[0115]

[0116]

[0117]

[0118]

[0119]

[0120]

[0121]

[0122]

[0123]

[0124] In another preferred embodiment, the pharmaceutically acceptable salt is an inorganic acid salt or an organic acid salt.

[0125] In another preferred embodiment, the inorganic acid salt is selected from the following group: hydrochloride, hydrobromide, hydroiodide, sulfate, bisulfate, nitrate, phosphate, and acid phosphate.

[0126] 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.

[0127] The third aspect of the present invention provides a pharmaceutical composition comprising a safe and effective amount of the compound described in the first aspect of the present invention and a pharmaceutically acceptable carrier.

[0128] The fourth aspect of the present invention provides a use of the compound described in the first aspect of the present invention for preparing a drug used as a CDK4 kinase inhibitor.

[0129] The fifth aspect of the present invention provides a use of the compound described in the first aspect of the present invention for preparing a drug for regulating CDK4 kinase activity or treating CDK4-related diseases.

[0130] In another preferred embodiment, the CDK4-related disease is selected from the following group: inflammation, cancer, cardiovascular disease, infection, immune disease, and metabolic disease.

[0131] 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, and uterine cancer.

[0132] In another preferred embodiment, the inflammation is selected from the following group: dermatitis, keratitis, conjunctivitis, prostatitis, hepatitis, and enteritis.

[0133] In another preferred embodiment, the infection is selected from the following group: bacterial infection and viral infection.

[0134] In another preferred embodiment, the bacterial infection is selected from the following group: Gram-positive bacterial infection, Gram-negative bacterial infection, mycoplasma infection, and fungal infection.

[0135] In another preferred embodiment, the viral infection is selected from the group consisting of coronavirus infection, influenza A, influenza B, and avian influenza.

[0136] In another preferred embodiment, the immune disease is selected from the group consisting of systemic lupus erythematosus, rheumatoid arthritis, scleroderma, and ulcerative colitis.

[0137] 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

[0138] After extensive and in-depth research, the inventors unexpectedly discovered a class of compounds with good CDK4 kinase inhibitory activity. In addition, the compounds have excellent inhibitory activity and selectivity against CDK4 kinase, and have better pharmacodynamics / pharmacokinetic properties. On this basis, the present invention was completed.

[0139] the term

[0140] Unless otherwise stated, the following terms used in this application (including the specification and claims) have the definitions given below.

[0141] 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-.

[0142] "Alkyl (alone or as part of another group)" refers to a monovalent straight or branched saturated hydrocarbon group containing 1 to 12 carbon atoms consisting only of carbon and hydrogen atoms. Alkyl is preferably C1-C6 alkyl (i.e., containing 1, 2, 3, 4, 5 or 6 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, etc. In the present 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. "Haloalkyl" refers to an alkyl as defined herein in which one or more hydrogens are replaced by the same or different halogens. Examples of haloalkyl include -CH2Cl, -CH2CF3, -CH2CCl3, perfluoroalkyl (e.g., -CF3), etc.

[0143] "Alkylene" refers to a divalent radical of an alkyl group, for example, -CH2-, -CH2CH2-, and -CH2CH2CH2-.

[0144] "Alkoxy (alone or as part of another group)" refers to an alkyl group to which an oxy group is attached, which has an alkylO-structure, wherein the alkyl group has the definition as described above. Preferably, the alkoxy group is a C1 to C6 alkoxy group. Alkoxy includes, but is 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.

[0145] "Thioalkyl" refers to an alkyl group in which a carbon is replaced by S, S(O) or S(O)2.

[0146] "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. In the present invention, "C2-C6 alkenyl" refers to an alkenyl containing 2, 3, 4, 5 or 6 carbon atoms. Alkenyl includes, but is not limited to, for example, vinyl, propenyl, butenyl, 1-methyl-2-butene-1-yl, etc. In the present invention, alkenyl includes substituted alkenyl.

[0147] "Alkenylene" refers to an alkenyl group having two points of attachment. For example, "vinylene" refers to the group -CH=CH-. Alkenylene can also be in unsubstituted form or in substituted form with one or more substituents.

[0148] "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. In the present invention, "C2-6 alkynyl" refers to an alkynyl group having 2, 3, 4, 5 or 6 carbon atoms. Alkynyl groups include, but are 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, alkynyl groups also include substituted alkynyl groups.

[0149] "Alkynylene" refers to an alkynyl group with two points of attachment. For example, "ethynylene" refers to the group: -C≡C-. Alkynylene can also be in unsubstituted form or in substituted form with one or more substituents.

[0150] "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.

[0151] "Aromatic ring system" means a monocyclic, bicyclic or polycyclic hydrocarbon ring system in which at least one ring is aromatic.

[0152] "Aryl (alone or as part of another group)" refers to a monovalent group of an aromatic ring system. 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 C6-C12 aryl group. In the present invention, aryl is also intended to include substituted aryl groups.

[0153] "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 are replaced by an aryl group, and aryl and alkyl have the definitions described above. Examples of "arylalkyl" or "aralkyl" include benzyl, 2-phenylethyl, 3-phenylpropyl, 9-fluorenyl, diphenylmethyl, and triphenylmethyl, etc.

[0154] "Aryloxy" means an -O-(aryl) group wherein the aryl portion is as defined herein.

[0155] "Heteroalkyl" refers to a substituted alkyl 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 can be given, for example, C1-C6 heteroalkyl refers to the number of carbons in the chain, which includes 1 to 6 carbon atoms. For example, a -CH2OCH2CH3 group is referred to as a "C3" heteroalkyl. Connection to the rest of the molecule can be through a heteroatom or carbon in the heteroalkyl chain. "Heteroalkylene" refers to an optionally substituted divalent alkyl having one or more backbone chain atoms selected from atoms other than carbon, for example, oxygen, nitrogen, sulfur, phosphorus, or a combination thereof.

[0156] "Carbocyclic ring system" refers to a monocyclic, bicyclic or polycyclic hydrocarbon ring system in which each ring is fully saturated or contains one or more units of unsaturation, but in which none of the rings are aromatic.

[0157] "Carbocyclyl" refers to a monovalent group of a carbon ring system, including, for example, cycloalkyl (cyclopentyl, cyclobutyl, cyclopropyl, cyclohexyl, etc.) and cycloalkenyl (cyclopentenyl, cyclohexenyl, cyclopentadienyl, etc.).

[0158] "Cycloalkyl" refers to a monovalent saturated carbocyclic group consisting of a mono- or bicyclic ring having 3-12, preferably 3-10, more preferably 3-8 ring atoms. The cycloalkyl group may be optionally substituted with one or more substituents, each of which is 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.

[0159] "Cycloalkyloxy" refers to a radical of the formula -OR, where R is a cycloalkyl as defined herein. Exemplary cycloalkyloxy groups include cyclopropyloxy, cyclobutyloxy, cyclopentyloxy, cyclohexyloxy, and the like. "Cycloalkylalkyl" refers to -(cycloalkyl)-alkyl, where the cycloalkyl and alkyl groups are as disclosed herein. "Cycloalkylalkyl" is bonded to the parent molecular structure through the cycloalkyl group.

[0160] "Heteroaromatic ring system" refers to a monocyclic (e.g., 5- or 6-membered), bicyclic (6-12-membered) or polycyclic ring system, wherein at least one ring is both aromatic and contains at least one heteroatom (e.g., N, O or S); and wherein none of the other rings is a heterocyclyl (as defined below). In some cases, the ring that is aromatic and contains heteroatoms contains 1, 2, 3 or 4 ring heteroatoms in the ring. At least one of the rings is heteroaromatic, and the remaining rings may be saturated, partially unsaturated or fully unsaturated rings.

[0161] "Heteroaryl" refers to a monocyclic (e.g., 5- or 6-membered), bicyclic (e.g., 8-10-membered) or tricyclic group of 5 to 12 ring atoms, which contains at least one aromatic ring containing 1, 2 or 3 ring heteroatoms selected from N, O or S, and the remaining ring atoms are C. It should be clear that the point of attachment of the heteroaryl group should be located on the aromatic ring. Examples of heteroaryl groups include, but are not limited to: imidazolyl, Azolyl, iso Azolyl, thiazolyl, isothiazolyl, oxadiazolyl, thiadiazolyl, pyrazinyl, thienyl, furanyl, pyranyl, pyridinyl, pyrrolyl, pyrazolyl, pyrimidinyl, quinolyl, isoquinolyl, benzofuranyl, benzofuranyl, benzothiophenyl, benzothiopyranyl, benzimidazolyl, benzo Azolyl, benzo oxadiazole, benzothiazolyl, benzothiadiazolyl, benzopyranyl, indolyl, isoindolyl, triazolyl, triazine, quinoxalinyl, purinyl, quinazolinyl, quinolizinyl, naphthyridinyl, pteridinyl, carbazolyl, aza Base, diazepine Heteroarylene refers to a heteroaryl group having two attachment sites.

[0162] "Heterocyclic ring system" refers to monocyclic, bicyclic and polycyclic ring systems in which at least one ring is saturated or partially unsaturated (but non-aromatic) and the ring contains at least one heteroatom. The heterocyclic ring system may be attached to a pendant group at any heteroatom or carbon atom that creates a stable structure and any ring atom may be optionally substituted.

[0163] "Heterocyclyl" refers to a monovalent group of a heterocyclic ring system, usually a stable monocyclic ring (such as 3-8 members, i.e. 3, 4, 5, 6, 7 or 8 members) or bicyclic ring (such as 5-12 members, i.e. 5, 6, 7, 8, 9, 10, 11 or 12 members) or polycyclic ring (such as 7-14 members, i.e. 7, 8, 9, 10, 11, 12, 13 or 14), including fused, spiro and / or bridged ring structures, which are saturated, partially unsaturated, and contain carbon atoms and 1, 2, 3 or 4 heteroatoms independently selected from N, O and S. Representative heterocyclyl groups include ring systems in which (1) each ring is non-aromatic and at least one ring contains heteroatoms, for example, tetrahydrofuranyl, tetrahydropyranyl, tetrahydrothiophenyl, pyrrolidinyl, pyrrolidonyl, piperidinyl, pyrrolinyl, decahydroquinolinyl, oxazolidinyl, piperazinyl, dioxanyl, dioxolanyl, diazepinyl, oxazepinyl, thiazepinyl, morpholinyl, and quinuclidinyl; (2) at least one ring is (3) at least one ring is non-aromatic and contains heteroatoms and at least one other ring is an aromatic carbocyclic ring, for example, 1,2,3,4-tetrahydroquinolyl, 1,2,3,4-tetrahydroisoquinolyl; and (4) 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. A heterocyclyl group refers to a heterocyclyl group having two attachment sites. In the present invention, preferably, the heterocyclyl group 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 heterocyclyl group is a 5-6-membered monocyclic heterocyclyl group or an 8-10-membered bicyclic heterocyclyl group.

[0164] "Heterocyclylalkyl" refers to an alkyl group substituted by a heterocyclyl group, wherein the heterocyclyl group and the alkyl group are as defined above.

[0165] 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.

[0166] "Cycloalkylamino" refers to a radical of the formula -NRaRb, wherein Ra is H, an alkyl radical as defined herein, or a cycloalkyl radical as defined herein, and Rb is a cycloalkyl radical 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.

[0167] In the present invention, "ester group" refers to a group having a -C(O)-OR or RC(O)-O- structure, wherein R independently represents hydrogen, alkyl, cycloalkyl, aryl, heteroaryl, heterocyclic group, as defined above.

[0168] 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.

[0169] 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.

[0170] "Ketocarbonyl" refers to RC(=O)-, where R is alkyl, cycloalkyl, etc. as described above.

[0171] 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.

[0172] 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.

[0173] 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 the substituent described above, or the 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 should be understood by those skilled in the art that the combination of substituents contemplated by the present invention is 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 substituent or polyhalogen substituent, the latter such as trifluoromethyl or alkyl containing Cl3), cyano, nitro, oxo (such as =O), trifluoromethyl, trifluoromethoxy, cycloalkyl, alkenyl, alkynyl, heterocycle, aromatic ring, OR a , SR a 、S(=O)R e 、S(=O)2R e 、P(=O)2R e 、S(=O)2OR e , P(=O)2OR e NR b R c NR b S(=O)2R e NR b P(=O)2R e 、S(=O)2NR b R c 、P(=O)2NR b R c 、C(=O)OR d 、C(=O)R a 、C(=O)NR b R c 、OC(=O)R a 、OC(=O)NR b R c NR b C(=O)OR e NR d C(=O)NR b R c NR d S(=O)2NR b R c NR d P(=O)2NR b R c NR b C(=O)R a , or NR b P(=O)2R e , where Ra may independently represent hydrogen, deuterium, alkyl, cycloalkyl, alkenyl, alkynyl, heterocyclic or aromatic rings, and R b , R c and R d may independently represent hydrogen, deuterium, an alkyl group, a cycloalkyl group, a heterocyclic ring or an aromatic ring, or R b and R c Together with the N atom, it can form a heterocyclic ring; R e It can independently represent hydrogen, alkyl, cycloalkyl, alkenyl, alkynyl, heterocycle or aromatic ring. The above typical substituents, such as alkyl, cycloalkyl, alkenyl, cycloalkenyl, alkynyl, heterocycle 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.

[0174] "Cyano" refers to a -CN group.

[0175] "Nitro" refers to -NO2.

[0176] "Hydroxyl" refers to -OH.

[0177] "Amino" refers to -NH2 or RNH-, where R is ketocarbonyl, sulfonyl, sulfonamide, R a -C(=O)-、R a R b NC(=O)- etc., where R a and R b It is an alkyl group, a cycloalkyl group, an aryl group or a heteroaryl group.

[0178] "Halogen" refers to any halogen radical, for example, -F, -Cl, -Br or -I.

[0179] The term "deuterated compound" refers to a compound in which one or more hydrogen atoms (H) are replaced by deuterium atoms (D).

[0180] In the present invention, the term "plurality" independently refers to 2, 3, 4, or 5.

[0181] Active ingredients

[0182] 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 II, or a pharmaceutically acceptable salt, hydrate, solvate, isotope compound (such as a deuterated compound) or prodrug thereof. The term also includes racemates and optical isomers.

[0183] The compound of formula II has the following structure:

[0184]

[0185] wherein each group is as defined above.

[0186] 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.

[0187] 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.

[0188] 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.

[0189] Prodrugs and solvates (or solvates) of the compounds of the present invention are also within the scope of this invention.

[0190] 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.

[0191] The compounds, salts or solvates of the present invention may exist in tautomeric forms (such as amides and imino ethers). All these tautomers are part of the present invention.

[0192] 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.

[0193] 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.

[0194] 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.

[0195] Throughout the specification, groups and substituents may be selected to provide stable fragments and compounds.

[0196] Specific functional groups and chemical term definitions are detailed below. For purposes of this invention, chemical elements are referred to in the Periodic Table of the Elements, CAS version, Handbook of Chemistry and Physics, 75 th 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.

[0197] 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.

[0198] 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.

[0199] The present invention also includes isotopically labeled compounds that are equivalent to the original compounds disclosed herein. However, in practice, it is common for one or more atoms to be replaced by atoms having a different atomic mass or mass number than the original atoms. Examples of isotopes of the compounds of the present invention include hydrogen, carbon, nitrogen, oxygen, phosphorus, sulfur, fluorine and chlorine isotopes, such as 2 H. 3 H. 13 C. 11 C. 14 C. 15 N. 18 O. 17 O. 31 P. 32 P. 35 S. 18 F and 36 The compounds of the present invention, or enantiomers, diastereomers, isomers, or pharmaceutically acceptable salts or solvates, wherein the isotopes or other isotopic atoms of the above compounds are all within the scope of the present invention. Certain isotope-labeled compounds of the present invention, such as 3 H and 14 Radioisotopes of C are also included and are useful in tissue distribution experiments of drugs and substrates. 3 H and carbon-14, i.e. 14 C, their preparation and detection are relatively easy. It is the first choice among isotopes. In addition, heavier isotope substitutions such as deuterium, i.e. 2 H, due to its good metabolic stability in certain therapies has advantages, such as increasing half-life or reducing dosage in vivo, therefore, in some cases, it can be preferred. Isotope-labeled compounds can be prepared by general methods by replacing readily available isotope-labeled reagents with non-isotopic reagents using the schemes disclosed in the examples.

[0200] 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.

[0201] 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.

[0202] 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.

[0203] In another preferred embodiment, in the compound, any one of ring A, ring B, R1, R2, and X1 is a corresponding group in the specific compound.

[0204] Preparation method

[0205] Methods for preparing compounds of Formula II are described in the following schemes and examples. Starting materials and intermediates are purchased from commercial sources, prepared by known procedures, or otherwise described. In some cases, the order of executing the steps of the reaction scheme may be changed to facilitate the reaction or avoid unwanted side reaction products.

[0206] The preparation method of the compound of formula II 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 combination can be easily carried out by a technician in the field to which the present invention belongs.

[0207] 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.

[0208] Preferably the preparation method is as follows:

[0209]

[0210] Step 1: In a solvent (such as 1,2-dichloroethane, dioxane, tetrahydrofuran), SM1 and M1 or M2 react at 70-120 degrees to generate SM2 through palladium-catalyzed coupling or nucleophilic substitution reaction;

[0211] Step 2: In an inert solvent (such as DMF, dioxane, ethylene glycol dimethyl ether, N-methylpyrrolidone, tetrahydrofuran, etc.), under alkaline conditions (such as diisopropylethylamine, potassium acetate, DBU, LiHDMS, etc.) or in the presence of a catalyst and a ligand (such as Pd(PPh3)4, Pd2(dba)3\t-BuXphos, etc.), M3 reacts with SM2 to generate T (i.e., compound of formula II).

[0212] In the above formulae, Ring A, Ring B, R1, R2 and X1 are as defined above.

[0213] Unless otherwise specified, the above starting materials can be purchased from commercial sources or synthesized according to reported literature.

[0214] Pharmaceutical compositions and methods of administration

[0215] The pharmaceutical composition of the present invention is used to prevent and / or treat the following diseases: inflammation, cancer, cardiovascular disease, infection, immune disease, and metabolic disease.

[0216] The compounds of formula II can be used in combination with other drugs known to treat or improve similar symptoms. When the combination is administered, the original drug administration method and dosage can remain unchanged, and the compound of formula II can be taken simultaneously or subsequently. When the compound of formula II is taken simultaneously with one or more other drugs, it is preferred to use a pharmaceutical composition containing one or more known drugs and the compound of formula II. The combination of drugs also includes taking the compound of formula II and one or more other known drugs in overlapping time periods. When the compound of formula II is used in combination with one or more other drugs, the dosage of the compound of formula II or the known drug may be lower than the dosage of the compound of formula II or the known drug when taken alone.

[0217] The drugs or active ingredients that can be used in combination with the compound of formula II 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, F520, 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, Tas Elisib, Buparlisib, 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, Tedecinaline, etc.), MEK inhibitors (such as selumetinib (AZD6244), trametinib (GSK1120 212), 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, Ocatinib, Linsitinib, BMS-754807, GSK1838705, etc.), A, 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.

[0218] 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.

[0219] 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.

[0220] "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 ), wetting agents (such as sodium lauryl sulfate), colorants, flavoring agents, stabilizers, antioxidants, preservatives, pyrogen-free water, etc.

[0221] 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.

[0222] 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.

[0223] 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.

[0224] 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.

[0225] Besides such inert diluents, the composition may also include adjuvants such as wetting agents, emulsifying and suspending agents, sweetening, flavoring, and perfuming agents.

[0226] 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, and the like.

[0227] 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.

[0228] 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.

[0229] The treatment method of the present invention can be used alone or in combination with other treatment methods or therapeutic drugs.

[0230] 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.

[0231] 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 formula II of the present invention or its crystal form, pharmaceutically acceptable salt, hydrate or solvate, thereby forming a pharmaceutical composition.

[0232] The present invention also provides a treatment method, which comprises the steps of administering the compound of formula II 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 CDK4.

[0233] Compared with the prior art, the present invention has the following main advantages:

[0234] (1) The compounds of the present invention have excellent inhibitory ability and selectivity against CDK4 kinase;

[0235] (2) The compounds of the present invention have lower toxic side effects;

[0236] (3) The compounds of the present invention have better pharmacodynamics and pharmacokinetic properties.

[0237] The present invention will be further described below in conjunction with specific examples. It should be understood that these examples are intended to illustrate the present invention only 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 stated, percentages and parts are calculated by weight.

[0238] 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.

[0239] Example B1

[0240] Synthesis of compound TB-1 of the present invention:

[0241]

[0242] The synthetic route is as follows:

[0243]

[0244] The experimental process is as follows:

[0245] Step 1:

[0246] Compound SM 1 (1.0 g, 1.0 eq) was dissolved in ultra-dry tetrahydrofuran solvent, placed in an ice-water bath to cool, cesium carbonate solid (1.37 g, 1.0 eq) was added, and isopropylamine (0.25 g, 1.0 eq) was added dropwise under stirring. After the addition was completed, the temperature was naturally raised to react. After the reaction was completed under 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, and after suction filtration, it was concentrated under reduced pressure, and column chromatography was separated and purified to obtain a yellow solid SM 2 (750 mg, 64.7%).

[0247] 1 H NMR (400MHz, Chloroform-d) δ6.76 (t, J = 1.9 Hz, 1H), 6.57 (dd, J = 10.9, 2.0 Hz, 1H), 3.73 (dq, J = 13.0, 6.5 Hz, 1H), 1.31 (d, J = 6.3 Hz, 6H).

[0248] Step 2:

[0249] Compound SM 2 (0.75 g, 1.0 eq) was dissolved in a mixed solvent of anhydrous ethanol and water (v:v = 4:1), iron powder (0.91 g, 6.0 eq) and solid ammonium chloride (0.29 g, 2.0 eq) were added in sequence, and the temperature was raised to 90°C for reaction. After the reaction was completed, most of the solvent was removed by concentration under reduced pressure, and the liquid was extracted with saturated sodium bicarbonate solution and EA, and the organic phase was dried over anhydrous sodium sulfate, filtered, concentrated under reduced pressure, and purified by column chromatography to obtain compound SM 3 (0.5 g, 74.7%)

[0250] 1 H NMR (400MHz, Chloroform-d) δ6.64(dd,J=9.4,2.1Hz,1H),6.53(t,J=1.8Hz,1H),3.56(p,J=6.3Hz,1H),3.50(s,1H),3.15(s,2H),1.23(d,J=6.2Hz,6H).

[0251] Step 3:

[0252] Compound SM 3 (1.3 g, 1.0 eq) and CDI (2.2 g, 2.5 eq) were dissolved in ultra-dry DMF solvent in turn, and the temperature was raised to 110°C for reaction. After the reaction was completed under TLC monitoring, the mixture was cooled to room temperature, extracted with EA and water, and the organic phase was dried over anhydrous sodium sulfate, filtered, concentrated under reduced pressure, and purified by column chromatography to obtain compound SM 4 (1.2 g, 83.3%).

[0253] 1 H NMR (400MHz, Chloroform-d) δ9.82 (s, 1H), 7.08 (s, 1H), 7.02 (dd, J = 8.0, 4.0Hz, 1H), 4.70 (m, 1H), 1.53 (d, J = 8.0Hz, 6H).

[0254] Step 4:

[0255] Compound SM 4 (1.2 g, 1.0 eq) was dissolved in dry toluene, phosphorus oxychloride (7.1 g, 1.0 eq) was added, and the temperature was raised to 90°C for two days. After the reaction was completed, the mixture was cooled to room temperature and poured into ice water for quenching. The organic phase was extracted with EA and dried over anhydrous sodium sulfate. After suction filtration, it was concentrated under reduced pressure and purified by column chromatography to obtain compound SM 5 (1.2 g, 94%).

[0256] 1H NMR (400MHz, Chloroform-d) δ7.45 (d, J = 2.0Hz, 1H), 7.14 (dd, J = 8.0, 2.0Hz, 1H), 4.88 (m, 1H), 1.64 (d, J = 8.0Hz, 6H).

[0257] Step 5:

[0258] Compound SM 5 (1.23 g, 1.0 eq) was dissolved in ultra-dry DMF, and dimethylamine hydrochloride (3.5 g, 10.0 eq) and triethylamine (8.8 g, 20.0 eq) were added in sequence, and the temperature was raised to 110°C for reaction. After the reaction was completed, the mixture was cooled to room temperature, extracted with EA and water, and the organic phase was dried over anhydrous sodium sulfate, filtered, concentrated under reduced pressure, and purified by column chromatography to obtain compound SM 6 (1.2 g, 94.5%), LCMS: [M+H] + =299.9,301.8.

[0259] Step 6:

[0260] Under nitrogen protection, compound SM 6 (0.27 g, 1.0 eq), diboronic acid pinacol ester (0.46 g, 2.0 eq), Pd(dppf)Cl2 (33 mg, 0.05 eq) and potassium acetate (0.26 g, 3.0 eq) were added to a dry 50 mL three-necked flask in sequence, and the nitrogen was replaced three times. Ultra-dry 1,4-dioxane solvent was added, and the temperature was raised to reflux for reaction. After the reaction was completed, it 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 SM 7 (0.3 g, 97%), LCMS: [M+H] + =348.3.

[0261] Step 7:

[0262] Under nitrogen protection, compound SM 7 (0.39 g, 1.0 eq), 2,4,5-trichloropyrimidine (0.27 g, 1.3 eq), tetrakistriphenylphosphine palladium (0.13 g, 0.1 eq) and sodium carbonate solid (0.36 g, 3.0 eq) 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 reflux. After the reaction was completed, the mixture was cooled to room temperature, and the mixture was extracted with 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 8 (0.2 g, 49%). LCMS: [M+H] + =368.2.

[0263] Step 8:

[0264] Under nitrogen, compound SM 8 (0.17 g, 1.0 eq), (3s, 4r)-4-aminooxane-3-ol hydrochloric acid (0.14 g, 2.0 eq) and DIPEA (0.24 g, 4.0 eq) were added to DMSO solvent in sequence, and the temperature was raised to 90°C for reaction. After the reaction, the mixture was cooled to room temperature, and extracted with saturated brine, water and EA in sequence. The organic phase was dried over anhydrous sodium sulfate, filtered and concentrated under reduced pressure, and purified by column chromatography to obtain compound TB-1 (90 mg, 43.3%) with an HPLC purity of 99.7%. LCMS: [M+H] + =449.4.

[0265] 1 H NMR(400MHz,Chloroform-d)δ8.30(s,1H),7.71(d,J=1.4Hz,1H),7.41(dd,J=11.4,1.4H z,1H),5.29(d,J=6.0Hz,1H),4.98(s,1H),4.68(h,J=7.0Hz,1H),4.02(ddd,J=27.1,11. 7,4.7Hz,2H),3.92–3.77(m,1H),3.63(td,J=9.5,4.9Hz,1H),3.46(td,J=11.9,2.2Hz,1 H),3.17(dd,J=11.4,9.8Hz,1H),2.98(s,6H),2.07(s,1H),1.62(dd,J=7.0,1.9Hz,6H).

[0266] With reference to the synthesis method of Example B1, the following compounds were synthesized:

[0267]

[0268]

[0269]

[0270]

[0271]

[0272] Example B2

[0273] Synthesis of compound TB-16 of the present invention:

[0274]

[0275] The synthetic route is as follows:

[0276]

[0277] The experimental process is as follows:

[0278] Synthesis of Compound TB-16

[0279] Under nitrogen protection, 5-[(4-ethylpiperazine-1-yl)methyl]pyridin-2-amine (180 mg, 1.5 eq) was dissolved in 4 mL of tetrahydrofuran ultra-dry solvent, placed in an ice-water bath, and a 1M concentration of LiHDMS in tetrahydrofuran solution (0.81 mL, 1.5 eq) was added dropwise. The mixture was reacted in an ice-water bath for half an hour, and then compound SM 1 (100 mg, 1.0 eq) was added and kept warm for reaction. After the reaction, Ea and water were extracted and separated, and the organic phase was dried over anhydrous sodium sulfate, filtered and concentrated under reduced pressure. After preparative separation and purification, 120 mg of compound TB-16 was obtained, with an HPLC purity of 98.14% and a yield of 82.0%. LCMS: [M+H] + =552.5.

[0280] 1 H NMR(400MHz,Chloroform-d)δ8.53(s,1H),8.47(s,1H),8.36–8.32(d,J=8.0Hz,1H),8.21(d,J=4.0Hz,1H),7.80(d,J=1.6Hz,1H),7.57(dd,J=8.0,4.0Hz ,1H),7.48(dd,J=12.0,1.6Hz,1H),4.65(p,J=8.0Hz,1H),3.48(s,2H),2.94 (s,6H),2.73(m,8H),1.58(d,J=8.0Hz,6H),1.25(t,J=6Hz,3H),1.19(m,2H).

[0281] Referring to the synthesis method of Example B2, the following compound was synthesized:

[0282]

[0283]

[0284]

[0285] Example B3

[0286] Synthesis of compound TB-23 of the present invention:

[0287] The synthetic route is as follows:

[0288]

[0289] The experimental process is as follows:

[0290] Step 1:

[0291] Compound SM 1 (1.5 g, 1.0 eq) was dissolved in a mixed solvent of ammonia water and methanol (45 mL, v:v = 2:1), and the temperature was raised to 110°C in a sealed autoclave for 24 hours. After the reaction was completed, the mixture was cooled to room temperature, the reaction solvent was concentrated under reduced pressure, and directly separated and purified by silica gel column chromatography to obtain 1.1 g of compound SM 2, with a yield of 78.6%, LCMS: [M+H] + =272.0,274.0.

[0292] Step 2:

[0293] Under ice-water bath, compound SM 2 (170 mg, 1.0 eq) was dissolved in 2 mL of ultra-dry THF solvent, and then triethylamine (316 mg, 5.0 eq) and 4-chlorobutyryl chloride (221 mg, 2.5 eq) were added in sequence, and the reaction was kept in ice-water bath. After the reaction was completed, EA and water were used to extract the liquid, and the organic phase was dried over anhydrous sodium sulfate and filtered. After the filtrate was concentrated under reduced pressure, it was separated and purified by silica gel column chromatography to obtain 180 mg of compound SM 3 with a yield of 76.6%. LCMS: [M+H] + =376.1,378.1.

[0294] Step 3:

[0295] Under ice-water bath conditions, compound SM 3 (1.1 g, 1.0 eq) was dissolved in 22 mL of ultra-dry DMF solvent, and then NaOH solid (0.21 g, 1.8 eq) was added, and the reaction was stirred in an ice bath. After the reaction was completed, the liquid was extracted with saturated ammonium chloride and EA, and the organic phase was dried over anhydrous sodium sulfate and filtered. After the filtrate was concentrated under reduced pressure, it was separated and purified by silica gel column chromatography to obtain 340 mg of compound SM 4, with a yield of 34%. LCMS: [M+H] + =340.1,342.1.

[0296] Step 4:

[0297] Under nitrogen protection, compound SM 4 (360 mg, 1.0 eq), diboronic acid pinacol ester (537 mg, 2.0 eq), Pd(dppf)Cl2 (38.7 mg, 0.05 eq) and potassium acetate (311 mg, 3.0 eq) were added to a dry 50 mL three-necked flask in sequence, and the nitrogen was replaced three times. Ultra-dry 1,4-dioxane solvent was added, and the temperature was raised to reflux for reaction. After the reaction was completed, it 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 SM 5 (310 mg, 75.6%), LCMS: [M+H] + =388.3.

[0298] Step 5:

[0299] Under nitrogen protection, compound SM 5 (310 mg, 1.0 eq), 2,4,5-trichloropyrimidine (200 mg, 1.5 eq), tetrakistriphenylphosphine palladium (92.5 mg, 0.1 eq) and sodium carbonate solid (254.5 mg, 3.0 eq) were added to a 50 mL three-necked flask in sequence, and 13 mL of a mixture of 1,4-dioxane and water (v:v=10:3) was added. The mixture was replaced with nitrogen three times, and the temperature was raised to reflux. After the reaction was completed, the mixture was cooled to room temperature, and the mixture was extracted with Ea and water. The organic phase was dried over anhydrous sodium sulfate, filtered, concentrated under reduced pressure, and purified by column chromatography to obtain compound SM 6 (240 mg, 73.6%). LCMS: [M+H] + =408.1.

[0300] Step 6:

[0301] Under nitrogen conditions, compound SM 6 (180 mg, 1.0 eq), (3s, 4r)-4-aminooxane-3-ol hydrochloric acid (102 mg, 1.5 eq) and DIPEA (228 mg, 4.0 eq) were added to 3 mL of DMSO solvent in sequence, and the temperature was raised to 90 ° C for reaction. After the reaction was completed, it was cooled to room temperature, and the liquid was extracted with saturated brine, water and EA in sequence. The organic phase was dried over anhydrous sodium sulfate, filtered and concentrated under reduced pressure, and purified by column chromatography to obtain compound TB-23 (120 mg, 56.0%) with an HPLC purity of 96.75%. LCMS: [M+H] + =489.4.

[0302] 1H NMR(400MHz,Chloroform-d)δ8.32(s,1H),7.88(d,J=1.2Hz,1H),7.45(dd,J=12.0,1.2Hz,1H),5.33(d ,J=8.0Hz,1H),4.82(s,1H),4.61(m,1H),4.11(t,J=8.0Hz,3H),4.05(dd,J=12.0,4.0Hz,1H),3.98(dd ,J=12.0,4.0Hz,1H),3.92–3.79(m,1H),3.63(td,J=8.0,4.0Hz,1H),3.46(td,J=12.0,2.0Hz,1H),3.1 7(t,J=8.0,1H),2.63(t,J=8.0Hz,2H),2.32(p,J=8.0Hz,2H),1.78–1.69(m,2H),1.68(d,J=8.0Hz,6H).

[0303] Example B4

[0304] Synthesis of compound TB-36 of the present invention:

[0305]

[0306] The synthetic route is as follows:

[0307]

[0308] The experimental process is as follows:

[0309] Step 1:

[0310] Under nitrogen protection, Int-1 (200 mg, 1.0 eq), N-methyl-3-aminobenzenesulfonamide (136.0 mg, 1.5 eq), cesium carbonate (476.5 mg, 3.0 eq) and Xantphos Pd G2 catalyst (21.6 mg, 0.05 eq) were added to a 25 mL three-necked flask, and 10 mL of ultra-dry 1,4-dioxane solvent was added. After nitrogen replacement three times, the temperature was raised to reflux reaction. After the reaction was completed, it was cooled to room temperature, and the system was extracted and separated with Ea and water. The organic phase was dried with anhydrous sodium sulfate, filtered and concentrated under reduced pressure. The isomers were separated by preparative chromatography to obtain compound TB-36 (20 mg, 7.3%), with an HPLC purity of 97.18%, LCMS: [M+H] + =561.0.

[0311] 1H NMR(400MHz,Chloroform-d)δ8.51(s,1H),8.25(t,J=2.0Hz,1H),8.03(d,J=1.3Hz,1H),7.84(d,J=8.3Hz,1H),7.62-7.43(m,4H),4. 79(p,J=6.9Hz,1H),4.66(t,J=7.7Hz,2H),4.51(q,J=5.4Hz,1H),4.40(t,J=7.7Hz,2H),2.69(d,J=5.4Hz,3H),1.72(d,J=6.9Hz,6H).

[0312] With reference to the synthetic method of Example B4, the following compounds were synthesized:

[0313]

[0314]

[0315]

[0316]

[0317] Example B5

[0318] Synthesis of compound TB-40 of the present invention:

[0319]

[0320] The synthetic route is as follows:

[0321]

[0322] The experimental process is as follows:

[0323] Step 1:

[0324] 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%).

[0325] 1H NMR (400MHz, Chloroform-d) δ7.35 (s, 1H), 6.81 (t, J = 2.0Hz, 1H), 6.64 (dd, J = 12. 0,2.0Hz,1H),3.67(t,J=4.0Hz,2H),3.46(q,J=8.0Hz,2H),2.16(p,J=8.0Hz,H).

[0326] Step 2:

[0327] Compound SM 2 (4.3 g, 1.0 eq) was dissolved in a mixed solvent of methanol and water (v:v=4:1), iron powder (3.9 g, 5.0 eq) and solid ammonium chloride (1.5 g, 2.0 eq) 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, 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 the filtrate was extracted and separated with a 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%).

[0328] Step 3:

[0329] Compound SM 3 (1.3 g, 1.0 eq) and CDI (2.2 g, 2.5 eq) were dissolved in super dry DMF solvent in turn, and the temperature was raised to 90°C for reaction. After the reaction was completed under TLC monitoring, the mixture was cooled to room temperature, extracted with EA and water, and the organic phase was dried over anhydrous sodium sulfate, filtered, concentrated under reduced pressure, and purified by column chromatography to obtain compound SM 4 (1.2 g, 83.3%).

[0330] Step 4:

[0331] Compound SM 4 (4.0 g, 1.0 eq) was dissolved in dry toluene, phosphorus oxychloride (36 mL, 30.0 eq) was added, and the temperature was raised to reflux for two days. TLC monitoring, after the reaction was completed, the mixture was cooled to room temperature, poured into ice water for quenching, extracted with EA, and the organic phase was dried over anhydrous sodium sulfate, filtered, concentrated under reduced pressure, and purified by column chromatography to obtain compound SM5 (137 mg, 24.9%), LCMS: [M+H] + =325.3,327.3.

[0332] Step 5:

[0333] Compound SM 5 (1.25 g, 1.0 eq) was added to 60 mL of 30% methylamine ethanol solution, and the temperature was raised to 60°C for reaction. After the reaction was completed, the mixture was cooled to room temperature, concentrated under reduced pressure to remove most of the solvent, and extracted with saturated brine and EA. The organic phase was dried over anhydrous sodium sulfate, filtered, concentrated, and purified by silica gel column chromatography to obtain compound SM6 (500 mg, 48.9%).

[0334] 1 H NMR(400MHz,Chloroform-d)δ6.98(dd,J=10.0,1.7Hz,1H),6.93(d,J=1.7Hz,1H ),3.93(t,J=6.2Hz,2H),3.48-3.31(m,2H),3.22(s,3H),2.26(p,J=6.0Hz,2H).

[0335] Step 6:

[0336] Under nitrogen protection, compound SM 6 (0.26 g, 1.0 eq), bipyraclostrobin (0.46 g, 2.0 eq), Pd(dppf)Cl2 (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 Int-1 (0.21 g, 69.3%), LCMS: [M+H] + =332.6.

[0337] Step 7:

[0338] Under nitrogen protection, compound Int-1 (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 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 Int-2 (0.13 g, 59.2%), LCMS: [M+H] + =352.5.

[0339] Step 8:

[0340] Under nitrogen, compound Int-2 (100 mg, 1.0 eq), (3s, 4r)-4-aminooxane-3-ol hydrochloric acid (87.2 mg, 2.0 eq) and DIPEA (0.2 mL, 4.0 eq) were added to DMSO solvent in sequence, and the temperature was raised to 90°C for reaction. After the reaction was completed, the mixture was cooled to room temperature, extracted with saturated saline, water and EA in sequence, and the organic phase was dried over anhydrous sodium sulfate, filtered and concentrated under reduced pressure, and purified by preparative liquid separation to obtain compound TB-40 (50 mg, 40.7%), with an HPLC purity of 91.7%, LCMS: [M+H] + =433.4.

[0341] 1 H NMR(400MHz,Chloroform-d)δ8.25(s,1H),7.42(dd,J=11.8,1.2Hz,1H),7.32(d,J=1.2 Hz,1H),5.27(d,J=6.0Hz,1H),5.12(s,1H),4.05(m,2H),4.00(t,J=6.0Hz,2H),3.89–3 .77(m,1H),3.62(td,J=9.6,4.8Hz,1H),3.50–3.43(m,1H),3.42(t,J=5.6Hz,2H),3.26 (s,3H),3.24–3.14(m,1H),2.28(p,J=5.8Hz,2H),2.09–1.98(m,1H),1.70–1.64(m,1H).

[0342] With reference to the synthetic method of Example B5, the following compounds were synthesized:

[0343]

[0344]

[0345]

[0346]

[0347]

[0348]

[0349]

[0350]

[0351]

[0352]

[0353]

[0354]

[0355]

[0356]

[0357] Example B6 Compound synthesized by the present invention:

[0358] The experimental process is as follows:

[0359] The synthesis route of compound TB-98 is as follows:

[0360]

[0361] The experimental process is as follows:

[0362] first step:

[0363] Compound SM 1 (7.7 g, 1.0 eq) was dissolved in ultra-dry tetrahydrofuran solvent, placed in an ice-water bath, and cesium carbonate solid (31.8 g, 3.0 eq) was added. (S)-3-aminobutyric acid methyl ester hydrochloride (5.0 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 (6.7 g, 61.5%), LCMS: [M+H] + =335.2,337.2.

[0364] Step 2:

[0365] Compound SM 2 (6.7 g, 1.0 eq) was dissolved in a mixed solvent of methanol and water (v:v=4:1), iron powder (6.7 g, 6.0 eq) and solid ammonium chloride (2.1 g, 2.0 eq) 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, 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 the filtrate was extracted and separated with a 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 (5.5 g, 90.0%), LCMS: [M+H] + =305.1,307.1.

[0366] Step 3:

[0367] Compound SM 3 (5.5 g, 1.0 eq) and CDI (4.4 g, 1.5 eq) were dissolved in ultra-dry DMF solvent in turn, and the temperature was raised to 50°C for reaction. After the reaction was completed under TLC monitoring, the mixture was cooled to room temperature, extracted with EA and water, and the organic phase was dried over anhydrous sodium sulfate, filtered, concentrated under reduced pressure, and purified by column chromatography to obtain compound SM 4 (5.4 g, 90.4%), LCMS: [M+H] + =331.2,333.1.

[0368] Step 4:

[0369] Compound SM 4 (5.4 g, 1.0 eq) was dissolved in phosphorus oxychloride (18.2 mL, 12.0 eq), and the temperature was raised to 100°C for reaction for 12 hours. After the reaction was completed, the mixture was cooled to room temperature and poured into ice water for quenching. The mixture was extracted with EA 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 5 (5.2 g, 91.2%). LCMS: [M+H] + =349.0,351.1.

[0370] Step 5:

[0371] Compound SM 5 (1.9 g, 1.0 eq) was added to 50 mL of 30% methylamine ethanol solution and kept in an ice-water bath for reaction. After the reaction was completed, most of the solvent was removed by concentration under reduced pressure, and the liquid was extracted with saturated brine and EA. The organic phase was dried over anhydrous sodium sulfate, filtered, concentrated, and purified by silica gel column chromatography to obtain compound SM 6 (1.7 g, 89.4%), LCMS: [M+H] + =348.1,350.1.

[0372] Step 6:

[0373] Compound SM 6 (1.5 g, 1.0 eq) was dissolved in 50 mL of ultra-dry tetrahydrofuran solvent, cooled in an ice-water bath, and 60% NaH powder was added in batches. After the addition was completed, the reaction was kept warm for 3 hours. After the reaction was completed, it was cooled to 0°C, and an appropriate amount of ice water was added to quench. The liquid was extracted with saturated brine and EA. The organic phase was dried over anhydrous sodium sulfate, filtered, and concentrated. The organic phase was separated and purified by silica gel column chromatography to obtain compound SM 7 (811 mg, 60.5%), LCMS: [M+H] + =312.1,314.0.

[0374] Step 7:

[0375] Under nitrogen protection, compound SM 7 (811 mg, 1.0 eq), bipyraclostrobin (1.32 g, 2.0 eq), Pd(dppf)Cl2 (95.1 mg, 0.05 eq) and potassium acetate (764.7 mg, 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 SM 8 (890 mg, 95.4%), LCMS: [M+H] + =360.3.

[0376] Step 7:

[0377] Under nitrogen protection, compound SM 8 (150 mg, 1.0 eq), 2,4,5-trichloropyrimidine (115 mg, 1.5 eq), tetrakistriphenylphosphine palladium (48.3 mg, 0.1 eq) and sodium carbonate solid (66.4 mg, 1.5 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, 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 9 (120 mg, 75.5%), LCMS: [M+H] + =380.1.

[0378] Step 8:

[0379] Under nitrogen, compound SM 9 (150 mg, 1.0 eq), (3s, 4r)-4-aminooxane-3-ol hydrochloric acid (121.2 mg, 2.0 eq) and DIPEA (0.28 mL, 4.0 eq) were added to NMP solvent in sequence, and the temperature was raised to 90°C for reaction. After the reaction was completed, the mixture was cooled to room temperature, extracted with saturated saline, water and EA in sequence, and the organic phase was dried over anhydrous sodium sulfate, filtered and concentrated under reduced pressure, and purified by preparative liquid separation to obtain compound TB-98 (76 mg, 40.7%), with an HPLC purity of 99.38%, LCMS: [M+H] + =461.3.

[0380] 1H NMR(400MHz,Chloroform-d)δ8.32(s,1H),7.53(s,1H),7.48(d,J=11.4Hz,1H),5.32(d,J=6.0H z,1H),4.75(p,J=6.9Hz,1H),4.05(dd,J=11.5,5.0Hz,1H),3.99(dd,J=11.7,4.1Hz,1H),3.90-3 .80(m,1H),3.67(dt,J=8.5,4.5Hz,1H),3.63(d,J=4.9Hz,1H),3.60(s,3H),3.46(ddd,J=14.1, 9.8,2.2Hz,2H),3.27-3.14(m,2H),2.11-1.99(m,1H),1.76-1.69(m,1H),1.47(d,J=6.7Hz,3H).

[0381] With reference to the synthesis method of Example B6, the following compounds were synthesized:

[0382]

[0383]

[0384] Example B7

[0385] The compound synthesized by the present invention:

[0386]

[0387] The experimental process is as follows:

[0388] Synthesis of compound TB-164

[0389] The synthetic route is as follows:

[0390]

[0391] The experimental process is as follows:

[0392] first step:

[0393] SM 1 (11.5 g, 1.0 eq) and potassium carbonate (13.4 g, 2.0 eq) were added to a 500 mL three-necked reaction bottle. Under nitrogen protection, 100 mL of DMF was added. 3-amino-3-methylbutan-1-ol (5 g, 1.0 eq) was added dropwise in an ice-water bath at 0°C. The mixture was replaced by N2 three times and the mixture was naturally heated to room temperature for 1-2 hours. TLC showed that new spots were generated. The raw materials could not react completely and the reaction was terminated. Filtered with diatomaceous earth, washed with an appropriate amount of ethyl acetate, the organic phase was directly concentrated, extracted with ethyl acetate and water, and the organic phase was washed with brine several times. The organic phase was dried over anhydrous sodium sulfate and concentrated. The yellow solid SM 2 was purified by silica gel chromatography with a yield of 60%.

[0394] Step 2:

[0395] Compound SM 2 (9.3 g, 1.0 eq) was added to a 100 mL three-necked flask, 40 mL of pyridine was added, acetic anhydride (8.2 mL, 3.0 eq) was added dropwise, and the mixture was reacted at room temperature under nitrogen protection. TLC showed that no raw material remained, and the reaction was completed. The reaction solution was extracted with ethyl acetate and water, the organic phase was dried over anhydrous sodium sulfate and concentrated, and separated and purified by silica gel column chromatography to obtain a yellow liquid SM 3, LCMS: [M+Na] + =385.0, with a yield of 96%.

[0396] Step 3:

[0397] Compound SM 3 (10.5 g, 1.0 eq), reduced iron powder (9.8 g, 6.0 eq) and NH4Cl (3.1 g, 2.0 eq) were added to a 500 mL three-necked flask in sequence, and a mixed solvent of methanol and water (v:v = 60 mL: 30 mL) was added. The mixture was heated to 70 °C under nitrogen protection and reacted for 1 hour. TLC showed that there was no residual raw material, and the reaction was completed. Cool to room temperature, filter with diatomaceous earth, wash with an appropriate amount of ethyl acetate, directly concentrate the organic phase, extract the organic phase with ethyl acetate and water, dry the organic phase with anhydrous sodium sulfate and concentrate it, separate and purify it by silica gel column chromatography to obtain a dark purple liquid SM 4, LCMS: [M+H] + =333.1,[M+Na] + =355.1, the yield is 92%.

[0398] Step 4:

[0399] SM 4 (8.8 g, 1.0 eq) was added to a 250 mL three-necked reaction bottle. Under nitrogen protection, 80 mL of DMF was added, and CDI (21.6 g, 5.0 eq) was added. The N2 was replaced three times, and the temperature was raised to 100 ° C for 1 hour. TLC showed that the raw material point disappeared and the reaction was completed. The organic phase was extracted with ethyl acetate and water, and the organic phase was washed with brine for many times. The organic phase was dried over anhydrous sodium sulfate and concentrated, and purified by silica gel chromatography to obtain a white solid SM 5. LCMS: [M+H] + =359.10,[M+Na] + =381.10, yield 72%.

[0400] Step 5:

[0401] Compound SM 5 (6.8 g, 1.0 eq) and K2CO3 (2.6 g, 1.0 eq) were added to a 250 mL three-necked flask, and 70 mL of methanol was added. The mixture was reacted at room temperature for 16 hours under nitrogen protection. TLC showed that no raw material remained, and the reaction was completed. The reaction solution was extracted with ethyl acetate and water, and the organic phase was dried over anhydrous sodium sulfate and concentrated. The white solid SM 6 was separated and purified by silica gel column chromatography, and the yield was 98%.

[0402] Step 6:

[0403] Compound SM 6 (5.9 g, 1.0 eq) was added to a 250 mL three-necked flask, followed by 100 mL of acetonitrile and DIPEA (13.0 mL, 4.0 eq), and MsCl (4.4 mL, 1.5 eq) and a mixed solvent of methanol and water (v:v = 60 mL:30 mL) were added dropwise in an ice-water bath at 0°C, and the mixture was replaced by N2 three times, and the mixture was naturally heated to room temperature for reaction. TLC showed that no raw material remained, and the reaction was completed. The organic phase was extracted with ethyl acetate and water, dried over anhydrous sodium sulfate, concentrated, and separated and purified by silica gel column chromatography to obtain a white solid SM 7, LCMS: [M+H] + =299.10, yield is 60%.

[0404] Step 7:

[0405] SM 7 (3.3 g, 1.0 eq), B2Pin2 (5.6 g, 2.0 eq), Pd(dppf)Cl2 (404 mg, 0.05 eq) and KOAc (3.3 g, 3.0 eq) were added to a 250 mL three-necked reaction bottle in sequence. Under nitrogen protection, 60 mL of 1,4-dioxane was added and the temperature was raised to 105 °C for 2 hours. TLC showed that the raw material point disappeared and the reaction was completed. The organic phase was extracted with ethyl acetate and water, dried over anhydrous sodium sulfate and concentrated, and purified by silica gel chromatography to obtain a light yellow solid SM 8, LCMS: [M+H]+ =347.40, with a yield of 95%.

[0406] Step 8:

[0407] Compound SM 8 (3.6 g, 1.0 eq), Pd(PPh3)4 (404 mg, 0.05 eq) and Na2CO3 (3.3 g, 3.0 eq) were added to a 250 mL three-necked reaction bottle in sequence. Under nitrogen protection, a mixed solvent of 1,4-dioxane: water (50 mL: 15 mL) was added, and 2,4,6-trichloropyrimidine (1.78 mL, 1.5 eq) was added. The temperature was raised to 105 ° C for 2 hours. TLC showed that there was no raw material remaining, and the reaction was completed. The reaction solution was extracted with ethyl acetate and water, the organic phase was dried over anhydrous sodium sulfate and concentrated, and the white solid SM 9 was obtained by separation and purification by silica gel column chromatography. LCMS: [M+H] + =367.20, with a yield of 97%.

[0408] Step 9:

[0409] Compound SM 9 (1.0 g, 1.0 eq) and (3S, 4R)-4-aminooxan-3-ol hydrochloride (840 mg, 2.0 eq) were added to a microwave tube, and 10 mL of NMP and DIPEA (2.0 mL, 4.0 eq) were added in sequence. The temperature was raised to 130 °C for reaction for 1 hour. TLC showed that no raw material remained, and the reaction was completed. The organic phase was extracted with ethyl acetate and water, and the organic phase was washed with brine several times. The organic phase was dried over anhydrous sodium sulfate and concentrated, and separated and purified by silica gel column chromatography to obtain a white solid TB-164, LCMS: [M+H] + =448.40, with a yield of 38%.

[0410] 1 H NMR(400MHz,Chloroform-d)δ8.31(s,1H),7.70(d,J=1.4Hz,1H),7.45(d,J=11.1Hz,1H) ,5.39(d,J=6.0Hz,1H),4.55-4.48(m,2H),4.05(dd,J=11.3,4.9Hz,1H),4.02-3.95(m,1 H),3.85(tt,J=10.8,5.4Hz,1H),3.64(td,J=9.5,5.0Hz,1H),3.46(td,J=11.9,2.2Hz,1 H),3.17(t,J=10.6Hz,1H),2.27-2.20(m,2H),1.77(s,6H),1.70(td,J=12.3,4.8Hz,3H).

[0411] With reference to the synthesis method of Example B7, the following compound was synthesized:

[0412]

[0413] Test Example 1 Enzyme Activity Test

[0414] The following is a biological activity test experiment on some compounds in the above examples and comparative examples.

[0415] The biological activity test experimental process is as follows:

[0416] 1. Kinase activity test:

[0417] The test compounds were tested for CDK4 and CDK6 kinase IC 50 Value detection.

[0418] (I) Reagent information

[0419] Reagents brand Part Number Palbociclib MCE HY-50767 FL-Peptide18 PerkinElmer 760362 FL-Peptide29 PerkinElmer 760429 FL-Peptide34 PerkinElmer 760643

[0420] (II) Equipment information

[0421] equipment brand Part Number Incubator Thermo Scientific - Shaker QILINBEIER - EZ Reader PerkinElmer 122919 Liquid Handler Labcyte Inc. Echo 550 Liquid Handler TECAN EVO200

[0422] 3. Research design

[0423] (1) Compound preparation:

[0424] 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.

[0425] ②Three-fold dilution to obtain 10 compound solutions of different concentrations.

[0426] (2) Enzyme assay:

[0427] ① Prepare a 1.3x enzyme solution containing enzyme, substrate, and cofactors as shown below.

[0428] ② Add 15 μL of 1.3x enzyme solution to each well and incubate at room temperature for 30 minutes.

[0429] ③ Add 5 μL of 4x ATP solution to start the reaction. Each test well contains the ingredients listed in the list, and the final volume is 20 μL.

[0430] ④ Incubate for 150 minutes, then add 75 μL of buffer (containing 0.5 M EDTA) to stop the reaction.

[0431] ⑤Use EZ to read the data of each test hole for analysis.

[0432] (3) Data analysis:

[0433] The percent inhibition was calculated using the concern ratio (CR) according to the following formula:

[0434] Wells treated with DMSO were used as positive controls, and wells without enzyme addition were used as negative controls.

[0435] % (inhibition percentage) = 100-100*((CRPC-CRSample) / (CRPC-CRNC)).

[0436]

[0437] Through the above detection, the inhibitory activity IC of the test sample on CDK4 and CDK6 kinases was obtained. 50 The (nM) values ​​are shown in Table 1.

[0438] Table 1

[0439]

[0440]

[0441]

[0442] As can be seen from Table 1, through in vitro biological activity screening, with Palbociclib as the reference substance, the compounds synthesized by the present invention have good inhibitory ability against CDK4 kinase, and the compounds synthesized by the present invention have very good selectivity for the kinase activity of CDK4 and CDK6, which will greatly reduce the side effects such as hematology caused by the inhibition of CDK6. Therefore, the compounds synthesized by the present invention are expected to be further developed into drugs for regulating CDK4 kinase activity or treating CDK4-related diseases.

[0443] Experimental Example 2 Cell Antiproliferation Experiment

[0444] 1. Experimental materials and equipment:

[0445] Human breast cancer cells MCF-7, ovarian cancer cells A2780, human mantle cell lymphoma cells JEKO-1. 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).

[0446] 2. Experimental preparation:

[0447] 1. Cell plating

[0448] 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.

[0449] B) Remove the culture medium from the 10 cm culture dish;

[0450] C) Rinse the cells once with 10 ml 1xPBS;

[0451] D) Add 4 ml of 0.25% EDTA-Tripsin and place in a 37°C, 5% CO2 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;

[0452] E) Resuspend in 4 ml DMEM medium, count and adjust to 50,000 cells / ml.

[0453] F) The cell suspension was added to a 96-well plate at a volume of 100 μL per well and cultured overnight in a 37°C, 5% CO2 incubator.

[0454] 2. Compound treatment

[0455] Compound dilution

[0456] A) Preparation of test compound gradient dilution solution: Prepare the test compound into 1 mM stock solution. Then dissolve 1.5 μl of the stock solution in 1.5 ml of DMSO-free culture medium, and then perform 3-fold continuous gradient dilution with 0.1% DMSO culture medium, for a total of 9 concentrations. The compound concentrations after dilution are as follows:

[0457] 333.33nM, 111.11nM, 37.03nM, 12.35nM, 4.15nM, 1.37nM, 0.46nM, 0.15nM

[0458] 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;

[0459] C) The cells were transferred to an incubator and incubated for 5 days.

[0460] 3. MTT assay

[0461] A) Take out the cell culture plate and add 10 μL of 5 mg / ml MTT in a biosafety cabinet;

[0462] B) Place the cell culture plate back into the incubator and continue incubating for 3 hours;

[0463] 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;

[0464] D) Select 570 nm wavelength to measure the absorbance value on TECAN enzyme-linked immunosorbent assay instrument.

[0465] 4. Data Analysis

[0466] The following formula was used to calculate the cell viability (% Cell Viability):

[0467] %Cell Viability=100%×(Lum_Sample-Lum_LC) / (Lum_HC-Lum_LC)

[0468] Lum_HC: 0.1% DMSO control group cell reading

[0469] Lum_Sample: Cell readout with added compound

[0470] Lum_LC: Blank medium reading

[0471] The IC was obtained by curve fitting using GraphPad Prism 8 software. 50 The numerical values ​​(unit: nM) are shown in Table 2, where A / B / C represent the following meanings: A≤500nM, 500nM<B≤2000nM, C>2000nM.

[0472] Table 2

[0473]

[0474]

[0475] As can be seen from Table 2, through in vitro biological activity screening, the compounds synthesized by the present invention have good inhibitory ability on human breast cancer cells MCF-7, ovarian cancer cells A2780, and human mantle cell lymphoma cells JEKO-1.

[0476] Test Example 3 Preclinical rat pharmacokinetic study

[0477] 1. Experimental materials and equipment:

[0478] Healthy adult SD rats, male, 6-8 weeks old, weighing 200-300 g. EDTA-Na2 anticoagulant. Analytical balance, animal weighing scale, magnetic stirrer, refrigerated centrifuge, single-channel manual pipette, etc.

[0479] 2. Experimental process:

[0480] 1. Drug preparation

[0481] 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 it freshly before use.

[0482] 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.

[0483] 2. Animal Preparation

[0484] 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.

[0485] 3. Drug administration

[0486] Route of administration: oral administration (po)

[0487] Dosage: 10 mg / kg

[0488] Dosing volume: 10mL / kg

[0489] 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.

[0490] 4. Sample collection

[0491] 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 10000g for 5min at 4℃ to separate plasma, and stored at -80℃ for testing. Blood was collected from the tail vein.

[0492] 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. That is, it has entered the vein and extracted about 0.1-0.2ml of whole blood. After pulling out the needle, press to stop bleeding.

[0493] 3. Sample analysis:

[0494] 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.

[0495] 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.

[0496] 4. Data Processing

[0497] The quantitative detection method of the test compound was established by 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 main pharmacokinetic parameters were calculated using the non-compartmental model in winnonlinPhoenix software. The detailed data are shown in Table 3.

[0498] Table 3

[0499]

[0500] As can be seen from Table 3, through PK test screening, the compounds synthesized by the present invention all have good pharmacokinetic properties.

[0501] 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

1. A compound used as a CDK4 kinase inhibitor, characterized in that The compound is a compound of formula II, or a pharmaceutically acceptable salt, stereoisomer, tautomer, hydrate, solvate, isotope compound or prodrug thereof, in: X1 is selected from the following group: N, CR3; R1 is selected from the group consisting of H, CF3, F, Cl, Br, methyl, ethyl, isopropyl, cyclopropyl; R2 is selected from the group consisting of H, F, CF3, CF2H, NH2, methyl; Or R1 and R2 together with the C to which they are connected form a 5-membered heteroaryl group containing 2 N; Each R3 is independently selected from the group consisting of H, CF3, F, Cl, Br, methyl, cyano; Ring A is selected from the following group: Wherein, X2 is O or NR, X3 is N or CR3, and X4 is NR, SO or SO2; Ring B is selected from the group consisting of: wherein each R4 is independently selected from the group consisting of H, cyano, halogen, substituted or unsubstituted C1-C6 alkyl, substituted or unsubstituted C3-C6 cycloalkyl; Each R5, R6, R9, R 10 and R 14 Each independently selected from the group consisting of: H, halogen, hydroxyl, amino, Oxo, substituted or unsubstituted C1-C6 alkyl, substituted or unsubstituted C3-C6 cycloalkyl, substituted or unsubstituted phenyl, substituted or unsubstituted C 1-6 Alkoxy, substituted or unsubstituted -NR m R n ; or two R connected to the same C 14 Together with the C to which it is attached, it forms a C3-C6 cycloalkyl group; or R9, R 10 Together with its connected N, it forms Each R and R' is independently selected from the group consisting of H, C1-C6 alkyl, halogenated C1-C6 alkyl, C3-C6 cycloalkyl, halogenated C3-C6 cycloalkyl, phenyl; Each R m Selected from the following group: H, C 1-6 Alkyl, C 3-6 Cycloalkyl, C 6-10 Aryl, 6-10 membered heteroaryl containing 1, 2 or 3 heteroatoms selected from N, O and S; Each R n C 1-6 alkyl; R4, R5, R6, R9, R 10 and R 14 The substitutions described in each of the above independently refer to substitutions by 1, 2 or 3 substituents selected from the group consisting of deuterium, halogen, hydroxy, amino, -N-(C1-C6 alkyl)2, C1-C6 alkoxy, phenyl, cyano; Each m, n, and p are independently selected from the following group: 0, 1, 2, 3, 4; q is selected from the following group: 1, 2, 3.

2. The compound according to claim 1, characterized in that R1 is selected from the group consisting of H, CF3, F, Cl, Br, methyl, ethyl, isopropyl, cyclopropyl; R2 is selected from the group consisting of H, F, CF3, CF2H, NH2, methyl; Ring A is selected from the following group: Wherein, X2 is O or NR, X3 is N or CR3, and X4 is NR, SO or SO2; Ring B is selected from the group consisting of: wherein each R4 is independently selected from the group consisting of H, cyano, halogen, substituted or unsubstituted C1-C6 alkyl, substituted or unsubstituted C3-C6 cycloalkyl; Each R5, R6, R9, R 10 and R 14 Each independently selected from the group consisting of: H, halogen, hydroxyl, amino, Substituted or unsubstituted C1-C6 alkyl, substituted or unsubstituted C3-C6 cycloalkyl, substituted or unsubstituted phenyl; or R9, R 10 Together with its connected N, it forms R and R' are each independently selected from the group consisting of H, C1-C6 alkyl, halogenated C1-C6 alkyl, C3-C6 cycloalkyl, halogenated C3-C6 cycloalkyl, phenyl; R4, R5, R6, R9, R 10 and R 14 The substitutions described in each of the above independently refer to substitutions by 1, 2 or 3 substituents selected from the group consisting of deuterium, halogen, hydroxy, amino, -N-(C1-C6 alkyl)2, C1-C6 alkoxy, phenyl, cyano; Each m, n, and p are independently selected from the following group: 0, 1, 2, 3, 4; q is selected from the following group: 1, 2, 3.

3. The compound according to claim 1, characterized in that R1 is selected from the group consisting of Cl, Br; R2 is H; Ring A is selected from the following group: Ring B is selected from the group consisting of: in, Each R5, R6, R9, R 10 and R 14 Each independently selected from the group consisting of: H, halogen, hydroxyl, amino, Substituted or unsubstituted C1-C6 alkyl, substituted or unsubstituted C3-C6 cycloalkyl, substituted or unsubstituted phenyl; or R9, R 10 Together with its connected N, it forms R and R' are each independently selected from the group consisting of H, C1-C6 alkyl, halogenated C1-C6 alkyl, C3-C6 cycloalkyl, halogenated C3-C6 cycloalkyl, phenyl; R5, R6, R9, R 10 and R 14 The substitutions described in each of the above independently refer to substitutions by 1, 2 or 3 substituents selected from the group consisting of deuterium, halogen, hydroxy, amino, -N-(C1-C6 alkyl)2, C1-C6 alkoxy, phenyl, cyano; Each m, n, and p are independently selected from the following group: 0, 1, 2, 3, and 4.

4. The compound according to claim 1, characterized in that R1 and R2 together with the C to which they are connected form a 5-membered heteroaryl group containing 2 N; Ring A is selected from the following group: Ring B is selected from the group consisting of: wherein each R4 is independently selected from the following group: H, substituted or unsubstituted C1-C6 alkyl, substituted or unsubstituted C3-C6 cycloalkyl; Each R5, R6, R9, R 10 and R 14 Each independently selected from the group consisting of: H, halogen, hydroxyl, amino, Substituted or unsubstituted C1-C6 alkyl, substituted or unsubstituted C3-C6 cycloalkyl, substituted or unsubstituted phenyl; R and R' are each independently selected from the group consisting of H, C1-C6 alkyl, halogenated C1-C6 alkyl, C3-C6 cycloalkyl, halogenated C3-C6 cycloalkyl, phenyl; R4, R5, R6, R9, R 10 and R 14 The substitutions described in each of the above independently refer to substitutions by 1, 2 or 3 substituents selected from the group consisting of deuterium, halogen, hydroxy, amino, -N-(C1-C6 alkyl)2, C1-C6 alkoxy, phenyl, cyano; Each m, n, and p are independently selected from the following group: 0, 1, 2, 3, and 4.

5. The compound according to claim 4, characterized in that It has the following structure: X is selected from the group consisting of N, CR3; R3 is selected from the group consisting of H, CF3, F, Cl, Br, methyl, and cyano.

6. A compound for use as a CDK4 kinase inhibitor, or a pharmaceutically acceptable salt, stereoisomer, tautomer, hydrate, solvate, isotope compound or prodrug thereof, characterized in that: The compound is selected from the group consisting of: 。 7. A pharmaceutical composition, characterized in that Contains a safe and effective amount of the compound according to any one of claims 1 to 8, and a pharmaceutically acceptable carrier.

8. Use of a compound according to any one of claims 1 to 6, characterized in that: Used for preparing drugs for regulating CDK4 kinase activity or treating CDK4 related diseases.

9. The use according to claim 8, characterized in that The disease is selected from the group consisting of inflammation, cancer, cardiovascular disease, infection, immune disease, and metabolic disease.

10. The use according to claim 9, characterized in that 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, and uterine cancer.

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