A pyrimidine benzo five-membered ring CDK inhibitor and its preparation method and application

Abstract

The present invention discloses a pyrimidine benzo five-membered ring CDK inhibitor and its preparation method and application, wherein the compound is shown in formula (S). The compound of the present invention can inhibit the continuous proliferation of cancer cells and can be used to prepare drugs for treating cancer or tumor-related diseases.

Description

Technical Field

[0001] The present invention relates to the field of pharmaceutical chemistry, and in particular to a pyrimidine benzo five-membered ring CDK inhibitor and a preparation method and application thereof. Background Art

[0002] Cyclin-dependent kinases (CDKs) play a vital role in the normal progression of the cell cycle. The upregulation of CDKs activity will lead to the acceleration of the cell cycle and cell proliferation, which will eventually lead to the occurrence and development of cancer. Therefore, inhibiting CDKs can prevent the malignant proliferation of cancer cells and can be used for the treatment of cancer. CDK6 plays an important regulatory role in the transition from the G1 phase to the S phase of the cell cycle, and CDK2 has a regulatory role in the transition from the G1 phase to the S phase of the cell cycle and in the S phase. Therefore, inhibiting CDK2 or CDK6 can significantly inhibit the proliferation of cancer cells and produce an anti-cancer effect. Summary of the invention

[0003] Purpose of the invention: The present invention provides a pyrimidine benzo five-membered ring CDK inhibitor, which inhibits CDK2 or CDK6. The present invention also provides a preparation method and application of the compound.

[0004] Technical solution: The compound of the present invention as shown in formula (S) or a pharmaceutically acceptable salt thereof:

[0005]

[0006] in,

[0007] X is selected from S or O;

[0008] R1 is selected from hydrogen or NR4R5, wherein R4 and R5 are selected from hydrogen, C1-C8 alkyl or C3-C8 cycloalkyl;

[0009] R2 is selected from halogen;

[0010] R3 is selected from -S(O)2C1-C8 alkyl or -C(O)C1-C8 alkyl.

[0011] Preferably:

[0012] Said X is selected from S or O;

[0013] R1 is selected from hydrogen or NR4R5, wherein R4 and R5 are selected from hydrogen, C1-C3 alkyl, cyclopentane or cyclohexane;

[0014] Said R2 is selected from F;

[0015] The R3 is selected from -S(O)2C1-C3 alkyl or -C(O)C1-C3 alkyl.

[0016] Preferably:

[0017] Said X is selected from S or O;

[0018] Said R1 is selected from hydrogen or NR4R5, wherein R4 and R5 are selected from hydrogen, methyl, ethyl, isopropyl, cyclopentane or cyclohexane;

[0019] Said R2 is selected from F;

[0020] The R3 is selected from -S(O)2CH3 or -C(O)CH3.

[0021] Preferably:

[0022] Said X is selected from S or O;

[0023] R1 is selected from hydrogen or NR4R5, wherein R4 is selected from hydrogen, methyl, ethyl, and R5 is selected from hydrogen, methyl, ethylisopropyl, and cyclopentane;

[0024] Said R2 is selected from F;

[0025] The R3 is selected from -S(O)2CH3 or -C(O)CH3.

[0026] In some specific embodiments of the present invention, the present invention also provides a compound selected from S-1 to S-10 or a pharmaceutically acceptable salt thereof, selected from the following compounds:

[0027]

[0028]

[0029] The above-mentioned pharmaceutically acceptable salts are acid addition salts of the compound of general formula (S), wherein the acid used for salt formation includes inorganic acids and organic acids, the inorganic acids include: hydrochloric acid, sulfuric acid, phosphoric acid and methanesulfonic acid, and the organic acids include acetic acid, trichloroacetic acid, propionic acid, butyric acid, maleic acid, p-toluenesulfonic acid, malic acid, malonic acid, cinnamic acid, citric acid, fumaric acid, camphoric acid, digluconic acid, aspartic acid and tartaric acid.

[0030] Preferably, the pharmaceutically acceptable salt described in the present invention is hydrochloride.

[0031] The preparation method of the compound of the general formula (S) of the present invention comprises preparing compound S by coupling reaction of compound A and compound B under the action of a palladium catalyst:

[0032]

[0033] in,

[0034] X is selected from S or O;

[0035] R1 is selected from hydrogen or NR4R5, wherein R4 and R5 are selected from hydrogen, C1-C8 alkyl or C3-C8 cycloalkyl;

[0036] R2 is selected from halogen;

[0037] R3 is selected from -S(O)2C1-C8 alkyl or -C(O)C1-C8 alkyl.

[0038] The present invention also provides a pharmaceutical composition of a compound as described in formula (S) or a pharmaceutically acceptable salt, ester, stereoisomer, solvent compound or prodrug thereof, and a pharmaceutically acceptable carrier.

[0039] A pharmaceutically acceptable carrier refers to an excipient or diluent that does not cause significant irritation to an organism and does not interfere with the biological activity and properties of the administered compound.

[0040] The present invention also provides a use of a compound of formula (S) or a pharmaceutically acceptable salt, ester, stereoisomer, solvent compound or prodrug thereof or the composition of the present invention in preparing a drug.

[0041] The present invention also provides a use of a compound of formula (S) or a pharmaceutically acceptable salt, ester, stereoisomer, solvent compound or prodrug thereof or the composition of the present invention in the preparation of a CDK2 or CDK6 inhibitor; the CDK2 or CDK6 kinase inhibitor is used to treat cancer or tumor-related diseases.

[0042] The use of the compound of the present invention or its pharmaceutically acceptable salt, ester, stereoisomer, solvate or prodrug or the composition of the present invention in the preparation of a drug for treating cancer or tumor-related diseases. The cancer or tumor is a cancer or tumor related to CDK2 or CDK6. Cancer or tumor-related diseases include but are not limited to leukemia, breast cancer, prostate cancer, lung cancer, multiple myeloma, liver cancer, gastric cancer, bone cancer, brain cancer, head and neck cancer, intestinal cancer, pancreatic cancer, bladder cancer, testicular cancer, ovarian cancer, esophageal cancer, liposarcoma, endometrial cancer, etc.

[0043] The compound of the general formula (S) or a pharmaceutically acceptable salt thereof described in the present invention has CDK2 or CDK6 target inhibitory activity and has a therapeutic effect on cell malignant proliferation tumors.

[0044] Beneficial effects: The compound represented by the general formula (S) of the present invention can inhibit the malignant proliferation of cancer cells, has good therapeutic effect, low toxicity, good drug metabolism characteristics, and is not prone to drug resistance. It can be used to prepare drugs for treating cancer or tumor-related diseases. DETAILED DESCRIPTION

[0045] The following examples are provided to facilitate a better understanding of the present invention, but are not intended to limit the present invention. The experimental methods in the following examples are conventional methods unless otherwise specified. The test materials used in the following examples are purchased from conventional biochemical reagent stores unless otherwise specified. The present application is described in detail below in conjunction with specific embodiments.

[0046] 1. Synthesis of intermediate reactants

[0047] The specific preparation method of the independently developed intermediate reactant (M) is as follows:

[0048] (1) Synthesis of 5-(2-chloro-5-fluoropyrimidin-4-yl)benzo[d]oxazole (M1)

[0049]

[0050] 5-Bromo-benzoxazole (3.96g, 20mmol), biboronic acid pinacol ester (5.58g, 22mmol), dissolved in 1,4-dioxane (100mL), then added Pd(dppf)Cl2 (731mg, 1mmol), potassium acetate (5.88g, 60mmol), replaced argon three times, heated to 100℃, and reacted for 12h. After the reaction was completed, water was added to quench, ethyl acetate was used, and the compound M1-1 was obtained by evaporation under reduced pressure and concentrated, without purification, and used directly in subsequent reactions. 2,4-Dichloro-5-fluoropyrimidine (4.01g, 24mmol), Pd(PPh3)2Cl2 (702mg, 1mmol), sodium carbonate (5.30g, 50mmol), water (20ml) and ethylene glycol dimethyl ether (20mL) were added to a three-necked flask, and argon was replaced three times. Compound M1-1 dissolved in ethylene glycol dimethyl ether (80 mL) was slowly added into a three-necked flask, heated to 80° C., reacted for 12 h, and purified by rapid silica gel column to obtain compound M1 (3.39 g, yield 68%). 1 H NMR (400MHz, Chloroform-d) δ8.61 (s, 1H), 8.56 (d, J = 3.1Hz, 1H), 8.28 (dd, J = 8.7, 1.8Hz, 1H), 8.20 (s, 1H), 7.74 (d, J = 8.7Hz, 1H).

[0051] (2) Synthesis of 5-bromobenzo[d]oxazol-2-amine (M2)

[0052]

[0053] 2-Amino-4-bromophenol (3.76 g, 20 mmol) and cyanogen bromide (2.54 g, 24 mmol) were dissolved in 100 mL of methanol, heated to 35°C, and reacted for 12 h. After the reaction was completed, a saturated aqueous solution of sodium bicarbonate was slowly added dropwise to quench, and the product was purified by rapid silica gel column to obtain compound M2 (3.23 g, yield 76%), MS (M+H) + :found,213.1.

[0054] (3) Synthesis of 2,5-dibromobenzo[d]oxazole (M3)

[0055]

[0056] Copper bromide (5.02 g, 22.2 mmol) and isoamyl nitrite (2.42 mL, 18 mmol) were added to 80 mL of methanol, stirred at 55 °C for 10 minutes, compound M2 (3.20 g, 15 mmol) was slowly added, the temperature was raised to 60 °C, and stirred for 1 h. Compound M3 (1.37 g, yield 33%) was obtained by rapid silica gel column purification, MS (M+H) + :found,277.9.

[0057] (4) Synthesis of 5-bromo-N-isopropylbenzo[d]oxazol-2-amine (M4)

[0058]

[0059] Compound M3 (1.38 g, 5 mmol) and isopropylamine (1.28 mL, 15 mmol) were dissolved in 1,4-dioxane (50 mL), heated to 80°C, and reacted for 12 h. Compound M4 (1.02 g, yield 80%) was obtained by rapid silica gel column purification. MS (M+H) + :found,255.1.

[0060] (5) Synthesis of 5-(2-chloro-5-fluoropyrimidin-4-yl)-N-isopropylbenzo[d]oxazol-2-amine (M5)

[0061]

[0062] Referring to the synthesis method of compound M1, the yield was 62%. 1 H NMR(400MHz,Chloroform-d)δ8.49(d,J=3.3Hz,1H),8.15(s,1H),7.93(dd,J=8.5,1.8Hz,1 H),7.36(d,J=8.5Hz,1H),5.02(d,J=7.9Hz,1H),4.16-4.07(m,1H),1.36(d,J=6.5Hz,6H).

[0063] (6) Synthesis of 5-(2-chloro-5-fluoropyrimidin-4-yl)benzo[d]thiazole (M6)

[0064]

[0065] Referring to the synthesis method of compound M1, the yield was 72%. 1 H NMR (400MHz, Chloroform-d) δ9.11 (s, 1H), 8.94 (s, 1H), 8.58 (d, J = 3.1Hz, 1H), 8.29 (dd, J = 8.6, 1.7Hz, 1H), 8.13 (dd, J = 8.6, 0.6Hz, 1H).

[0066] (7) Synthesis of 5-bromo-N,N-dimethylbenzo[d]thiazol-2-amine (M7)

[0067]

[0068] 5-Bromo-2-iodoaniline (5.96 g, 20 mmol), sodium thiamethoxam (5.72 g, 40 mmol), copper acetate (3.62 g, 20 mmol), potassium carbonate (5.52 g, 40 mmol) were added to N,N-dimethylformamide (150 mL), heated to 120°C, and reacted for 12 h. Compound M7 (4.37 g, yield 85%) was obtained by rapid silica gel column purification. MS (M+H) + :found,257.1.

[0069] (8) Synthesis of 5-(2-chloro-5-fluoropyrimidin-4-yl)-N,N-dimethylbenzo[d]thiazol-2-amine (M8)

[0070]

[0071] Referring to the synthesis method of compound M1, the yield was 65%. 1 H NMR (400MHz, Chloroform-d) δ8.49(d,J=3.3Hz,1H),8.35(s,1H),7.89(dd,J=8.3,1.0Hz,1H),7.72(d,J=8.3Hz,1H),3.24(s,6H).

[0072] (9) Synthesis of 6-(2-chloro-5-fluoropyrimidin-4-yl)benzo[d]thiazole (M9)

[0073]

[0074] Referring to the synthesis method of compound M1, the yield was 70%.1 H NMR (400MHz, Chloroform-d) δ9.18-9.17(m,1H),8.85-8.84(m,1H),8.59-8.57(m,1H),8.35-8.32(m,1H),8.29-8.26(m,1H).

[0075] (10) Synthesis of 6-bromo-N-cyclopentylbenzo[d]thiazol-2-amine (M10)

[0076]

[0077] 6-Bromo-2-chlorobenzothiazole (4.97 g, 20 mmol), cyclopentylamine (2.2 mL, 22 mmol), DIPEA (3.8 mL, 22 mmol) were dissolved in 100 mL of dimethyl sulfoxide, argon was replaced three times, heated to 80 ° C, and reacted for 12 h. Rapid silica gel column purification gave compound M10 (5.35 g, yield 90%), MS (M+H) + :found,297.1.

[0078] (11) Synthesis of 6-(2-chloro-5-fluoropyrimidin-4-yl)-N-cyclopentylbenzo[d]thiazol-2-amine (M11)

[0079]

[0080] Referring to the synthesis method of compound M1, the yield was 64%. 1 H NMR(400MHz,Chloroform-d)δ8.49(d,J=1.9Hz,1H),8.47(d,J=3.5Hz,1H),8.16(d,J=8.9Hz,1H ),7.59(d,J=8.9Hz,1H),6.03(s,1H),4.09-3.99(m,1H),2.18-2.11(m,2H),1.78-1.61(m,6H).

[0081] (12) Synthesis of 6-bromo-N,N-dimethylbenzo[d]thiazol-2-amine (M12)

[0082]

[0083] 4-Bromo-2-iodoaniline (5.96 g, 20 mmol), sodium thiamethoxam (5.72 g, 40 mmol), copper acetate (3.62 g, 20 mmol), potassium carbonate (5.52 g, 40 mmol) were added to N,N-dimethylformamide (150 mL), heated to 120°C, and reacted for 12 h. Compound M12 (4.37 g, yield 85%) was obtained by rapid silica gel column purification. MS (M+H) + :found,257.1.

[0084] (13) Synthesis of 6-(2-chloro-5-fluoropyrimidin-4-yl)-N,N-dimethylbenzo[d]thiazol-2-amine (M13)

[0085]

[0086] Referring to the synthesis method of compound M1, the yield was 63%. 1 H NMR (400MHz, Chloroform-d) δ8.49(d,J=1.9Hz,1H),8.45(d,J=3.6Hz,1H),8.16(dd,J=8.6,0.8Hz,1H),7.63(d,J=8.6Hz,1H),3.27(s,6H).

[0087] (14) Synthesis of 6-bromo-N,N-diethylbenzo[d]thiazol-2-amine (M14)

[0088]

[0089] 4-Bromo-2-iodoaniline (5.96 g, 20 mmol), sodium diethyldithiocarbamate trihydrate (9.09 g, 40 mmol), copper acetate (3.62 g, 20 mmol), potassium carbonate (5.52 g, 40 mmol) were added to N,N-dimethylformamide (150 mL), heated to 120°C, and reacted for 12 h. Compound M14 was obtained by rapid silica gel column purification (4.56 g, yield 80%), MS (M+H) + :found,285.1.

[0090] (15) Synthesis of 6-(2-chloro-5-fluoropyrimidin-4-yl)-N,N-diethylbenzo[d]thiazol-2-amine (M15)

[0091]

[0092] Referring to the synthesis method of compound M1, the yield was 66%. 1H NMR(400MHz,Chloroform-d)δ8.48(d,J=2.0Hz,1H),8.44(d,J=3.6Hz,1H),8.15(dd,J =8.6,0.9Hz,1H),7.59(d,J=8.6Hz,1H),3.63(q,J=7.2Hz,4H),1.32(t,J=7.2Hz,6H).

[0093] 2. Synthesis of Compounds S-1-S-10

[0094] Example 1: Synthesis of 4-(Benzo[d]oxazol-5-yl)-5-fluoro-N-(1-(methylsulfonyl)piperidin-4-yl)pyrimidin-2-amine (S-1)

[0095]

[0096] Compound M1 (498 mg, 2 mmol) and 1-methylsulfonyl-4-aminopiperidine (445 mg, 2.5 mmol) were dissolved in dioxane (6 mL), and then Pd2(dba)3 (92 mg, 0.1 mmol), Xantphos (116 mg, 0.2 mmol), cesium carbonate (1.3 g, 4 mmol) were added, argon was replaced three times, heated to 105 ° C, and reacted for 12 h. Cooling, filtration, concentration, column chromatography, and compound S-1 (375 mg, 48% yield) were obtained as a white solid. 1 H NMR(400MHz,Chloroform-d)δ8.54(s,1H),8.26(d,J=3.6Hz,1H),8.18(s,1H),8.15(d,J=9.4Hz,1H),7.69(dd,J=8.7,0.7Hz,1H), 5.07(d,J=7.8Hz,1H),4.05-3.97(m,1H),3.80-3.76(m,2H),3.00-2.93(m,2H),2.83(s,3H),2.25-2.20(m,2H),1.72-1.64(m,2H).

[0097] Example 2: Synthesis of 5-(5-fluoro-2-((1-(methylsulfonyl)piperidin-4-yl)amino)pyrimidin-4-yl)-N-isopropylbenzo[d]oxazol-2-amine (S-2)

[0098]

[0099] The synthetic method of reference example 1 has a yield of 40% and is a white solid. 1H NMR (300MHz, Chloroform-d) δ8.20(d,J=3.8Hz,1H),8.09(s,1H),7.80-7.77(m,1H),7.32(d,J=8.4Hz,1H),5.22(s,1H),5.19(s,1H),4.16-4. 05(m,1H),4.03-3.96(m,1H),3.80-3.73(m,2H),3.00-2.91(m,2H),2. 82(s,3H),2.25-2.18(m,2H),1.74-1.62(m,2H),1.36(d,J=6.5Hz,6H).

[0100] Example 3: Synthesis of 4-(Benzo[d]thiazol-5-yl)-5-fluoro-N-(1-(methylsulfonyl)piperidin-4-yl)pyrimidin-2-amine (S-3)

[0101]

[0102] The synthetic method of reference example 1 has a yield of 45% and is a white solid. 1 H NMR(400MHz,Chloroform-d)δ9.08(s,1H),8.87(s,1H),8.28(d,J=3.7Hz,1H),8.17(d,J=8.5Hz,1H),8.07(d,J=8.5Hz,1H),5.1 1(d,J=7.7Hz,1H),4.07-3.99(m,1H),3.81-3.76(m,2H),3.00-2.94(m,2H),2.83(s,3H),2.26-2.21(m,2H),1.74-1.67(m,2H).

[0103] Example 4: Synthesis of 5-(5-fluoro-2-((1-(methylsulfonyl)piperidin-4-yl)amino)pyrimidin-4-yl)-N,N-dimethylbenzo[d]thiazol-2-amine (S-4)

[0104]

[0105] The synthetic method of reference example 1 has a yield of 45% and is a white solid. 1H NMR(400MHz,Chloroform-d)δ8.27(s,1H),8.21(d,J=3.8Hz,1H),7.77(d,J=8.4Hz,1H),7.69(d,J=8.4Hz,1H),4.99(d,J=7.7Hz ,1H),4.06-3.99(m,1H),3.78-3.73(m,2H),3.24(s,6H),3.01-2.94(m,2H),2.82(s,3H),2.24-2.19(m,2H),1.72-1.62(m,2H).

[0106] Example 5: Synthesis of 4-(Benzo[d]thiazol-6-yl)-5-fluoro-N-(1-(methylsulfonyl)piperidin-4-yl)pyrimidin-2-amine (S-5)

[0107]

[0108] The synthetic method of reference example 1 has a yield of 47% and is a white solid. 1 H NMR (400MHz, DMSO-d6) δ9.54(s,1H),8.80(s,1H),8.49(d,J=3.7Hz,1H),8.24(d,J=8.6Hz,1H),8.16(d,J=8.6Hz,1H),7.39( d,J=7.8Hz,2H),3.95-3.89(m,1H),3.57-3.52(m,2H),2.95-2.91(m,2H),2.88(s,3H),2.03-1.99(m,2H),1.64-1.55(m,2H).

[0109] Example 6: Synthesis of N-cyclopentyl-6-(5-fluoro-2-((1-(methylsulfonyl)piperidin-4-yl)amino)pyrimidin-4-yl)benzo[d]thiazol-2-amine (S-6)

[0110]

[0111] The synthetic method of reference example 1 has a yield of 42% and is a white solid. 1H NMR (400MHz, Chloroform-d) δ8.35(d,J=1.8Hz,1H),8.19(d,J=4.0Hz,1H),8.06(d,J=9.4Hz,1H),7.58(d,J=8.6Hz,1H),5.65-5.45(m,1H),5.00(d,J=7.6 Hz,1H),4.10-4.02(m,1H),4.01-3.96(m,1H),3.79-3.74(m,2H),3.00-2.94( m,2H),2.82(s,3H),2.24-2.20(m,2H),2.17-2.00(m,2H),1.78-1.62(m,8H).

[0112] Example 7: Synthesis of 6-(5-fluoro-2-((1-(methylsulfonyl)piperidin-4-yl)amino)pyrimidin-4-yl)-N,N-dimethylbenzo[d]thiazol-2-amine (S-7)

[0113]

[0114] The synthetic method of reference example 1 has a yield of 48% and is a white solid. 1 H NMR (300MHz, DMSO-d6) δ8.42(s,1H),8.39(d,J=4.1Hz,1H),7.96(d,J=8.5Hz,1H),7.54(d,J=8.5Hz,1H),7.26(d,J=7.8Hz,1 H),3.93-3.82(m,1H),3.57-3.51(m,2H),3.19(s,6H),2.95-2.91(m,2H),2.88(s,3H),2.03-1.98(m,2H),1.63-1.52(m,2H).

[0115] Example 8: Synthesis of 1-(4-((4-(2-(dimethylamino)benzo[d]thiazol-6-yl)-5-fluoropyrimidin-2-yl)amino)piperidin-1-yl)ethan-1-one (S-8)

[0116]

[0117] The synthetic method of reference example 1 has a yield of 42% and is a white solid. 1H NMR(400MHz,Chloroform-d)δ8.36(d,J=1.8Hz,1H),8.19(d,J=4.0Hz,1H),8.07(dd,J=7.5,1.8Hz,1H),7.61(d,J=8.6Hz,1H),5.01(d,J=7.7 Hz,1H),4.53-4.48(m,1H),4.10-4.03(m,1H),3.85-3.80(m,1H),3.31 -3.24(m,7H),2.95-2.88(m,1H),2.21-2.09(m,5H),1.49-1.42(m,2H).

[0118] Example 9: Synthesis of N,N-diethyl-6-(5-fluoro-2-((1-(methylsulfonyl)piperidin-4-yl)amino)pyrimidin-4-yl)benzo[d]thiazol-2-amine (S-9)

[0119]

[0120] The synthetic method of reference example 1 has a yield of 47% and is a white solid. 1 H NMR (400MHz, Chloroform-d) δ8.35(d,J=1.8Hz,1H),8.18(d,J=4.0Hz,1H),8.05(d,J=9.4Hz,1H),7.58(d,J=8.6Hz,1H),5.01(d,J=7.6Hz,1H),4.04 -3.96(m,1H),3.79-3.74(m,2H),3.62(q,J=7.1Hz,4H),3.00-2.94(m,2H) ,2.83(s,3H),2.24-2.19(m,2H),1.72-1.65(m,2H),1.32(t,J=7.1Hz,6H).

[0121] Example 10: Synthesis of 1-(4-((4-(2-(diethylamino)benzo[d]thiazol-6-yl)-5-fluoropyrimidin-2-yl)amino)piperidin-1-yl)ethan-1-one (S-10)

[0122]

[0123] The synthetic method of reference example 1 has a yield of 43% and is a white solid. 1H NMR(400MHz,Chloroform-d)δ8.35(d,J=1.7Hz,3H),8.18(d,J=4.1Hz,3H),8.06 (d,J=8.6Hz,1H),7.58(d,J=8.6Hz,1H),5.00(d,J=7.7Hz,1H),4.53-4.48(m,1H) ,4.10-4.07(m,1H),3.84-3.80(m,1H),3.62(q,J=7.2Hz,4H),3.31-3.24(m,1H) ,2.96-2.89(m,1H),2.21-2.09(m,5H),1.49-1.42(m,2H),1.31(t,J=7.2Hz,6H).

[0124] 3. Biological Evaluation Experiment

[0125] (1) CDK2 kinase activity analysis method

[0126] a) Prepare ATP / substrate solution and CDK2 kinase solution using kinase reaction buffer.

[0127] b) Transfer compound dilutions to a 384-well plate; centrifuge and add CDK2 kinase solution to the 384-well plate, continue centrifugation for 1 minute, and then incubate at 25°C for 10 minutes.

[0128] c) Add ATP / substrate solution to the 384-well plate, centrifuge for 1 minute, and then incubate at 25°C for 60 minutes.

[0129] d) Add ADP-Glo ​​to the 384-well plate, centrifuge for 1 minute, and then incubate at 25°C for 40 minutes.

[0130] e) The reaction solution was transferred to another 384-well plate, centrifuged at 1000 rpm for 1 minute, and then incubated at 25°C for 40 minutes.

[0131] f) Read the luminescent signal with a multifunctional microplate reader and calculate the inhibition rate and IC of the compound on CDK2 kinase 50 value.

[0132] (2) CDK6 kinase activity analysis method

[0133] a) Prepare ATP / substrate solution and CDK6 kinase solution using kinase reaction buffer.

[0134] b) Transfer compound dilutions to a 384-well plate; centrifuge and add CDK6 kinase solution to the 384-well plate, continue centrifugation for 1 minute, and then incubate at 25°C for 10 minutes.

[0135] c) Add ATP / substrate solution to the 384-well plate, centrifuge for 1 minute, and then incubate at 25°C for 60 minutes.

[0136] d) Add ADP-Glo ​​to the 384-well plate, centrifuge for 1 minute, and then incubate at 25°C for 40 minutes.

[0137] e) The reaction solution was transferred to another 384-well plate, centrifuged at 1000 rpm for 1 minute, and then incubated at 25°C for 40 minutes.

[0138] f) Read the luminescent signal with a multifunctional microplate reader and calculate the inhibition rate and IC of the compound on CDK6 kinase 50 value.

[0139] Table 1 CDK kinase inhibitory activity of the compounds of the present invention

[0140]

[0141]

[0142] A:IC 50 ≤100nM; B:100nM <IC 50 ≤200nM; C:200nM <IC 50 ≤1000nM; “-”: not tested.

Claims

1. A compound represented by formula (S) or a pharmaceutically acceptable salt thereof, characterized in that: in, X is selected from S or O; R1 is selected from hydrogen or NR4R5, wherein R4 and R5 are selected from hydrogen, C1-C8 alkyl or cyclopentane; R2 is selected from halogen; R3 is selected from -S(O)2C1-C8 alkyl or -C(O)C1-C8 alkyl.

2. The compound according to claim 1 or a pharmaceutically acceptable salt thereof, characterized in that: Said X is selected from S or O; Said R1 is selected from hydrogen or NR4R5, wherein R4 and R5 are selected from hydrogen, C1-C3 alkyl or cyclopentane; Said R2 is selected from F; The R3 is selected from -S(O)2C1-C3 alkyl or -C(O)C1-C3 alkyl.

3. The compound according to claim 1 or a pharmaceutically acceptable salt thereof, characterized in that: Said X is selected from S or O; Said R1 is selected from hydrogen or NR4R5, wherein R4 and R5 are selected from hydrogen, methyl, ethyl, isopropyl or cyclopentane; Said R2 is selected from F; The R3 is selected from -S(O)2CH3 or -C(O)CH3.

4. The compound according to claim 1 or a pharmaceutically acceptable salt thereof, characterized in that: Said X is selected from S or O; R1 is selected from hydrogen or NR4R5, wherein R4 is selected from hydrogen, methyl, ethyl, and R5 is selected from hydrogen, methyl, ethyl, isopropyl or cyclopentane; Said R2 is selected from F; The R3 is selected from -S(O)2CH3 or -C(O)CH3.

5. The compound according to claim 1 or a pharmaceutically acceptable salt thereof, characterized in that: Select any of the following compounds:

6. The compound according to claim 1 or a pharmaceutically acceptable salt thereof, characterized in that: The pharmaceutically acceptable salt is an acid addition salt of the compound of formula (S), wherein the acid used for salt formation includes inorganic acids and organic acids, the inorganic acids include hydrochloric acid, sulfuric acid, phosphoric acid and methanesulfonic acid, and the organic acids include acetic acid, trichloroacetic acid, propionic acid, butyric acid, maleic acid, p-toluenesulfonic acid, malic acid, malonic acid, cinnamic acid, citric acid, fumaric acid, camphoric acid, digluconic acid, aspartic acid and tartaric acid.

7. A method for preparing the compound according to claim 1 or a pharmaceutically acceptable salt thereof, characterized in that: Compound S is prepared by coupling reaction of compound A and compound B under the action of palladium catalyst. X is selected from S or O; R1 is selected from hydrogen or NR4R5, wherein R4 and R5 are selected from hydrogen, C1-C8 alkyl or cyclopentane; R2 is selected from halogen; R3 is selected from -S(O)2C1-C8 alkyl or -C(O)C1-C8 alkyl.

8. A pharmaceutical composition, characterized in that: The invention comprises a compound according to any one of claims 1 to 6 or a pharmaceutically acceptable salt thereof, and a pharmaceutically acceptable carrier.

9. Use of the compound according to any one of claims 1 to 6 or a pharmaceutically acceptable salt thereof in the preparation of a CDK inhibitor.

10. Use of the compound according to any one of claims 1 to 6 or a pharmaceutically acceptable salt thereof in the preparation of a drug for preventing and / or treating tumors, wherein the tumor is a CDK-related tumor.

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