Coumarin KRAS-G12C inhibitor as well as preparation and application thereof

Abstract

The invention discloses a coumarin KRAS-G12C inhibitor as well as preparation and application of the coumarin KRAS-G12C inhibitor. The coumarin compound disclosed by the invention has a remarkable inhibition effect on non-small cell lung cancer cells (NCI-H23 and NCI-H358) with high expression of KRAS-G12C and pancreatic cancer cells (MIAPaCa-2), is particularly used for preparing medicines for treating and / or preventing non-small cell lung cancer and pancreatic cancer, and has a good prospect of development and application of antitumor medicines.

Description

Technical Field

[0001] The present invention relates to the fields of organic chemistry and medicinal chemistry, and specifically relates to a coumarin KRAS-G12C inhibitor and its preparation and application. Background Art

[0002] The rat sarcoma gene (Ras) was the first human proto-oncogene discovered. The RAS protein it encodes is a crucial molecular switch in cellular signaling pathways, playing a crucial role in regulating life processes such as cell proliferation, differentiation, and apoptosis. KRAS is the most commonly mutated subtype of the RAS family, accounting for approximately 85% of all RAS mutations, while NRAS and HRAS account for 12% and 3%, respectively. Clinical data indicate that the KRAS-G12C mutation accounts for 12% of all KRAS mutations in cancers, including 59% and 2% of all non-small cell lung cancer (NSCLC) and pancreatic cancer (PCA) cases, respectively. The KRAS-G12C mutation impairs its ability to bind to GAP proteins, thereby inhibiting GAP-induced GTP hydrolysis. With reduced GTPase hydrolysis capacity, GTP accumulates, leading to KRAS hyperactivation. This ultimately leads to the aberrant activation of multiple downstream signaling pathways, inducing the development and progression of malignancies.

[0003] To date, two drugs (AMG510 and MRTX849) have been approved for marketing internationally, and about 19 KRAS-G12C inhibitors have entered the clinical research stage in China. According to the reported KRAS-G12C inhibitors, they are mainly divided into quinazolines, piperidopyrimidines, fused tetracyclics, etc. according to the structure of the parent nucleus. However, the structural similarity of these inhibitors is high and homogeneous competition is severe. Therefore, the development of KRAS-G12C inhibitors with new structures is of great research significance. The present invention has discovered a coumarin compound that can lock KRAS-G12C in its inactive conformation and has potent inhibitory activity against KRAS-G12C protein and its mutated cells; in terms of preparation methods, the present invention can efficiently synthesize key intermediates and target compounds through simple methods. Summary of the Invention

[0004] The technical problem solved by the present invention is to provide a coumarin compound as a KRAS-G12C inhibitor.

[0005] The present invention provides a compound having the structural formula shown in Formula I or a pharmaceutically acceptable salt thereof:

[0006]

[0007] in:

[0008] X1 and X 2 are the same or different and are independently selected from hydrogen, halogen, allyl or alkoxy; X 1 Preferably, hydrogen, fluorine or chlorine, X 2 Preferably hydrogen;

[0009] Ar is selected from C 6-10 Aryl or C 5-10 Heteroaryl, wherein the heteroaryl contains 1-3 heteroatoms selected from N, O or S;

[0010] R 1 and R 2 the same or different, each independently selected from hydrogen or C 1-6 Alkyl or cyano;

[0011] R 3 Selected from C 1-4 Alkenyl, C 1-6 Halogenated alkyl, C 5-10 Heteroaryl, C 1-4 Halogenated alkenyl or C 3-6 Halogenated cycloalkyl;

[0012] In an embodiment of the present invention, Ar is selected from C 6-10 Aryl or C 5-10 heteroaryl;

[0013] In a further preferred embodiment of the present invention, Ar is further optimized to 2-fluorophenyl, 2-trifluoromethylphenyl, 2-fluoro-6-hydroxyphenyl, 3-methoxy-naphthyl-1, 3-hydroxy-naphthyl-1 or 5-methyl-1H-indazolyl-4.

[0014] In an embodiment of the present invention, R 1 and R 2 the same or different, each independently selected from hydrogen or C 1-6 alkyl;

[0015] In a further preferred embodiment of the present invention, R 1 Preferably, hydrogen, methyl, or S-configuration methyl, R 2 Preferred are hydrogen, methyl or R-configured methyl.

[0016] In an embodiment of the present invention, R 3 Selected from C 1-4 Alkenyl, C 1-6 Haloalkyl or C 5-10 heteroaryl;

[0017] In a further preferred embodiment of the present invention, R 3 is selected from vinyl, chloromethyl or 5-nitrofuryl-2-yl.

[0018] In the most preferred embodiment of the present invention, the following specific compounds are included:

[0019]

[0020] The present invention also provides a method for preparing the above-mentioned coumarin derivatives. The reaction flow of the preparation method is as follows:

[0021] Option 1:

[0022]

[0023] Option 2:

[0024]

[0025] The specific steps of Solution 1 are:

[0026] 1) Dissolve compound A in anhydrous ethanol solution and add R 1 or R 2 The substituted N-Boc piperazine was reacted at 100°C for 8 hours, and a yellow solid was precipitated after cooling to obtain compound B;

[0027] 2) Compound B is subjected to trifluoroacetic acid to remove Boc to obtain compound C, which is then directly acylated with acryloyl chloride to obtain compound D without purification;

[0028] 3) Compound D is dissolved in 1,4-dioxane solution, and subjected to Suzuki coupling reaction with arylboronic acid or arylboronic acid ester in the presence of a catalyst and an acidifying agent to obtain a reaction solution, which is evaporated to dryness and purified by column chromatography to obtain the target compound E.

[0029] The specific steps of Option 2 are:

[0030] Compound B is dissolved in 1,4-dioxane solution, and subjected to Suzuki coupling reaction with arylboronic acid or arylboronic acid ester catalyst and acidifying agent to obtain a reaction solution, which is concentrated and evaporated to dryness, and purified by column chromatography to obtain compound C-1;

[0031] Compound C-1 is subjected to trifluoroacetic acid-mediated removal of Boc to obtain compound D-1, which is then directly acylated with 5-nitro-2-furancarboxylic acid, acryloyl chloride or chloroacetyl chloride to obtain the target compound E without further purification.

[0032] In many cases, the compounds of the present invention are capable of forming acid and / or base salts by virtue of the presence of a piperazine group or groups similar thereto.

[0033] The beneficial effects of the present invention are as follows: the coumarin anti-tumor compounds and pharmaceutically acceptable salts thereof obtained by the present invention have excellent anti-tumor activity and safety, and can be used to treat cancers such as non-small cell lung cancer and pancreatic cancer containing KRAS-G12C mutations. DETAILED DESCRIPTION

[0034] The present invention provides the following examples. It should be particularly noted that the preparation methods in the examples of the present invention are merely used to illustrate the present invention, rather than to limit the present invention. Simple improvements to the preparation methods of the present invention based on the concept of the present invention fall within the scope of protection claimed in the present invention.

[0035] Example 1: Synthesis of tert-butyl (4-(7-bromo-6-chloro-2-oxo-2H-chromen-4-yl)piperazine-1-carboxylate

[0036] 7-Bromo-4,6-dichloro-2H-chromen-2-one (1.6 g, 5.44 mmol, 1.0 eq.) and N-Boc piperazine (1.52 g, 8.16 mmol, 1.5 eq.) were added to anhydrous ethanol. Triethylamine (0.08 mL, 0.54 mmol, 0.1 eq.) was added dropwise and heated under reflux for 10 h. The reaction mixture was cooled, the solid precipitated, filtered, and the filter cake collected. Drying afforded a white solid. The product assay data are as follows:

[0037] Yield: 76%; 1 H NMR (400MHz, Chloroform-d) δ7.63 (d, J=1.2Hz, 2H), 5.76 (s, 1H), 3.69-3.67 (m, 4H), 3.18-3.16 (m, 4H), 1.50 (s, 9H).

[0038] Example 2: Synthesis of 4-(4-acryloylpiperazin-1-yl)-7-bromo-6-fluoro-2H-chromen-2-one

[0039] To a solution of tert-butyl 4-(7-bromo-2-oxo-2H-chromen-4-yl)piperazine-1-carboxylate (880 mg, 1.98 mmol, 1.0 eq.) in DCM (12 mL) was added trifluoroacetic acid (3.7 mL, 49.5 mmol, 25.0 eq.) at room temperature and allowed to react for 30 min. The reaction solution was evaporated to dryness, and the crude product was used directly in the next step without purification.

[0040] The crude product obtained in the previous step was dissolved in DCM solution, and triethylamine (1.04 mL, 7.5 mmol, 2.93 eq.) was added. The temperature was lowered to -70°C, and acryloyl chloride (256 mg, 2.82 mmol, 1.1 eq.) was added dropwise. The reaction was incubated at -70°C for 20 min. Purified water was added, and the reaction solution was extracted with DCM. The organic phases were combined, washed once with saturated brine, evaporated to dryness, and purified by column chromatography to obtain the compound as a white solid. The product assay data are as follows:

[0041] Yield: 36%; 1 H NMR (400MHz, Chloroform-d) δ7.64 (d, J=6.4Hz, 1H), 6.61 (dd, J=16.8, 10.4Hz, 1 H), 6.52-6.30 (m, 1H), 5.82-5.80 (m, 2H), 3.89-3.87 (m, 4H), 3.24-3.21 (m, 2H).

[0042] Example 3: Synthesis of 4-(4-acryloylpiperazin-1-yl)-6-chloro-7-(3-hydroxynaphthalen-1-yl)-2-hydrogen-chromen-2-one

[0043] The compound 4-(4-acryloylpiperazin-1-yl)-7-bromo-6-fluoro-2H-chromen-2-one (150 mg, 0.37 mmol, 1.0 eq.), 3-hydroxynaphthalene borate (117 mg, 0.74 mmol, 2.0 eq.) and solid tripotassium phosphate (160 mg, 0.74 mmol, 2.0 eq.) were added sequentially to a mixed solution of 1,4-dioxane and water (5 mL, v / v, 4:1) under nitrogen protection. Tris(dibenzylideneacetone)dipalladium (Pd2(dba)3, 34.5 mg, 0.037 mmol, 0.1 eq.) and 2-dicyclohexylphosphine-2′,6′-dimethoxybiphenyl (sphos, 154.8 mg, 0.074 mmol, 0.2 eq.) were added, and the temperature was raised to 95°C for 3 hours. The mixture was cooled to room temperature, purified water was added and extracted (3 × 15 mL), the organic phases were combined, dried over anhydrous sodium sulfate, evaporated to dryness, and purified by column chromatography to obtain a white solid. The product test data are as follows:

[0044] Yield: 43% 1H NMR (400MHz, DMSO-d6) δ10.01 (s, 1H), 7.87 (s, 1H), 7.79 (d, J = 8.4Hz, 1H), 7.52 (s, 1H), 7.43-7.41 (m, 1H), 7.32-7.18 (m, 3H), 7.01 (d, J = 2.4Hz, 1 H), 6.87 (dd, J=16.4, 10.4Hz, 1H), 6.19 (dd, J=16.8, 2.4Hz, 1H), 5.87 (s, 1H), 5.76 (dd, J=10.4, 2.8Hz, 1H), 3.85-3.82 (m, 4H), 3.51-3.49 (m, 4H); 13 C NMR (150MHz, DMSO-d6) δ164.9, 161.1, 159.6, 154.9, 152.6, 142.1, 137.3, 135.3, 128.5, 128.4, 128.3, 127.1 , 126.9, 125.9, 125.8, 125.5, 123.9, 120.9, 119.9, 117.3, 110.2, 98.3, 55.4, 51.2, 44.9, 41.4.HRMS (ESI) m / z calculatedfor C 26 H 21 ClN2O4[M+H] + 461.1262, found 461.1240.

[0045] In the cell proliferation inhibition experiment described later, the sample number of this example is 1.

[0046] Example 4: Synthesis of the compound 4-(4-acryloyl-2-methylpiperazin-1-yl)-6-chloro-7-(3-hydroxynaphthalen-1-yl)-2H-methen-2-one

[0047] The operation was the same as in Example 7, except that 4-(4-acryloyl-2-methylpiperazin-1-yl)-7-bromo-6-chloro-2H-chromen-2-one was used instead of 4-(4-acryloylpiperazin-1-yl)-7-bromo-6-fluoro-2H-chromen-2-one to obtain a yellow solid. The product test data are as follows:

[0048] Yield: 47% 1H NMR (400MHz, DMSO-d6) δ10.03 (s, 1H), 7.85 (s, 1H), 7.79 (d, J=8.4Hz, 1H), 7.50 (d, J=5. 2Hz, 1H), 7.46-7.37 (m, 1H), 7.31-7.19 (m, 3H), 7.01 (dd, J=3.6, 2.4Hz, 1H), 6.96-6.77 (m, 1H), 6.23-6.16 (m, 1H), 5.88 (s, 1H), 5.76 (dd, J=10.4, 2.2Hz, 1H), 4.40 (d, J=12.8H z, 1H), 4.17-4.14 (m, 2H), 3.89-3.86 (m, 2H), 3.20 (t, J=12.6Hz, 2H), 1.09-1.06 (m, 3H); 13 C NMR (150MHz, DMSO-d6) δ165.4, 161.1, 159.0, 154.9, 152.6, 142.1, 137.3, 135.3, 128.2, 127.1, 126.9, 125.9, 1 25.8, 125.5, 123.9, 120.9, 120.1, 117.7, 110.1, 100.6, 79.7, 53.5, 53.3, 49.7, 46.0, 41.4, 13.9.HRMS (ESI) m / z calculatedfor C 27 H 23 ClN2O4[M+H] + 475.1418, found 475.1405.

[0049] In the cell proliferation inhibition experiment described later, the sample number of this example is 2.

[0050] Example 5: Synthesis of Compound (S)-4-(4-acryloyl-2-methylpiperazin-1-yl)-6-chloro-7-(3-hydroxynaphthalen-1-yl)-2H-methen-2-one

[0051] The operation was the same as in Example 7, except that (S)-4-(4-acryloyl-2-methylpiperazin-1-yl)-7-bromo-6-chloro-2H-chromen-2-one was used instead of 4-(4-acryloylpiperazin-1-yl)-7-bromo-6-fluoro-2H-chromen-2-one to obtain a white solid. The product test data are as follows:

[0052] Yield: 31% 1H NMR (600MHz, DMSO-d6) δ10.00 (d, J=6.0Hz, 1H), 7.86-7.83 (m, 1H), 7.79 (d, J=8.4Hz, 1H), 7.51 ( d, J=7.8Hz, 1H), 7.47-7.40 (m, 1H), 7.31-7.20 (m, 3H), 7.02 (dd, J=5.4, 2.4Hz, 1H), 6.93-6.78 ( m, 1H), 6.20 (t, J=15.0Hz, 1H), 5.89-5.86 (m, 1H), 5.77-5.74 (m, 1H), 4.40 (d, J=13.2Hz, 1H), 4. 25-4.10 (m, 2H), 4.00-3.77 (m, 1H), 3.59-3.41 (m, 2H), 3.23-3.07 (m, 1H), 1.08 (t, J=6.6Hz, 3H); 13 C NMR (150MHz, DMSO-d6) δ165.3, 161.1, 159.0, 154.9, 152.6, 142.1, 137.3, 135.3, 128.4, 128.2, 127.1, 126.9, 12 5.9, 125.7, 125.5, 123.9, 120.9, 119.9, 117.7, 110.1, 100.6, 79.7, 53.5, 49.7, 45.9, 41.5, 13.9.HRMS (ESI) m / z calculated for C 27 H 23 C1N2O4[M+H] + 475.1418, found 477.1169.

[0053] In the cell proliferation inhibition experiment described later, the sample number of this example is 3.

[0054] Example 6: Synthesis of 4-(4-acryloyl-3-methylpiperazin-1-yl)-6-chloro-7-(3-hydroxynaphthalen-1-yl)-2H-methen-2-one

[0055] The operation was the same as in Example 7, except that 4-(4-acryloyl-3-methylpiperazin-1-yl)-7-bromo-6-chloro-2H-chromen-2-one was used instead of 4-(4-acryloylpiperazin-1-yl)-7-bromo-6-fluoro-2H-chromen-2-one to obtain a yellow solid. The product test data are as follows:

[0056] Yield: 51% 1H NMR (600MHz, DMSO-d6) δ10.01 (s, 1H), 7.90 (d, J = 4.2Hz, 1H), 7.79 (d, J = 8.4Hz, 1H), 7 .52 (s, 1H), 7.43-7.40 (m, 1H), 7.30-7.20 (m, 3H), 7.01 (d, J=2.4Hz, 1H), 6.84 (dd, J=1 6.210.2Hz, 1H), 6.18 (d, J=15.6Hz, 1H), 5.86 (d, J=4.8Hz, 1H), 5.77-5.72 (m, 1H), 4.6 8-4.64(m, 1H), 4.11-4.08(m, 1H), 3.69-3.67(m, 3H), 3.11-3.08(m, 2H), 1.41(s, 3H); 13 C NMR (150MHz, DMSO-d6) δ164.9, 161.1, 160.1, 154.9, 152.6, 142.1, 137.3, 135.3, 128.7, 128.3, 127.1, 126.9, 125.9, 125.6, 125.5, 123.8, 120.9, 119.9, 117.3, 110.1, 98.4, 79.7, 55.0(2C), 50.9(2C), 29.5.HRMS(ESI)m / z calculated for C 27 H 23 ClN2O4[M+H] + 475.1418, found 475.1402.

[0057] In the cell proliferation inhibition experiment described later, the sample number of this example is 4.

[0058] Example 7: Synthesis of (R)-4-(4-acryloyl-3-methylpiperazin-1-yl)-6-chloro-7-(3-hydroxy-naphthalen-1-yl)-2H-methen-2-one

[0059] The operation was the same as in Example 7, except that (R)-4-(4-acryloyl-3-methylpiperazin-1-yl)-7-bromo-6-chloro-2H-chromen-2-one was used instead of 4-(4-acryloylpiperazin-1-yl)-7-bromo-6-fluoro-2H-chromen-2-one to obtain a yellow solid. The product test data are as follows:

[0060] Yield: 32% 1H NMR (400MHz, DMSO-d6) δ10.07 (s, 1H), 7.91 (s, 1H), 7.79 (d, J=12.0Hz, 1H), 7.60-7.36 (m, 2H), 7.24-7.21 (m, 3H), 7.03 (s, 1H), 6.88-6.85 (m, 1 H), 6.19 (d, J=24.6Hz, 1H), 5.95-5.69 (m, 2H), 4.67-4.64 (m, 1H), 4.24 -4.20(m, 1H), 3.69-3.65(m, 3H), 3.09-3.05(m, 2H), 1.45-1.40(m, 3H); 13 C NMR (150MHz, DMSO-d6) δ164.9, 161.1, 160.1, 154.9, 152.6, 142.1, 137.2, 135.3, 128.67, 128.3, 127.1, 126.9, 125.9, 125.6, 125.5, 125.5, 123.8, 120.9, 120.0, 117.3, 110.2, 98.4, 55.0(2C), 50.9(2C), 17.8. HRMS(ESI)m / zcalculated for C 27 H 23 ClN2O4[M+H] + 475.1418, found 475.1398.

[0061] In the cell proliferation inhibition experiment described later, the sample number of this example is 5.

[0062] Example 8: Synthesis of (R)-4-(4-acryloyl-3-methylpiperazin-1-yl)-6-chloro-7-(3-methoxy-naphthalen-1-yl)-2H-methen-2-one

[0063] The operation was the same as in Example 7, except that (R)-4-(4-acryloyl-3-methylpiperazin-1-yl)-7-bromo-6-chloro-2H-chromen-2-one and 3-methoxynaphthalene borate were used instead of 4-(4-acryloylpiperazin-1-yl)-7-bromo-6-fluoro-2H-chromen-2-one and 3-hydroxynaphthalene borate, respectively, to obtain a white solid. The product test data are as follows:

[0064] Yield: 42% 1H NMR (600MHz, DMSO-d6) δ8.04-7.81 (m, 2H), 7.59-7.43 (m, 3H), 7.31 (d, J=4.2H z, 2H), 7.16-7.06 (m, 1H), 6.84 (dd, J=16.2, 10.2Hz, 1H), 6.18 (d, J=16.6Hz, 1H ), 5.86 (d, J = 4.2Hz, 1H), 5.75 (d, J = 10.2Hz, 1H), 4.68-4.65 (m, 1H), 4.22-4.18 (m, 1H), 3.93 (s, 3H), 3.79-3.53 (m, 3H), 3.23-2.90 (m, 2H), 1.41-1.37 (m, 3H); 13 C NMR (150MHz, DMSO-d6) δ165.7, 161.1, 160.1, 156.8, 152.6, 141.8, 137.3, 135.0, 128.7, 128.3, 127.8, 127.2, 12 6.8, 125.6(2C), 125.5, 124.7, 121.1, 119.9, 117.4, 107.4, 98.4, 55.8, 55.0(2C), 50.9(2C), 16.9.HRMS(ESI)m / z calculated for C 28 H 25 ClN2O4[M+H] + 489.1575, found489.1556.

[0065] In the cell proliferation inhibition experiment described later, the sample number of this example is 6.

[0066] Example 9: Synthesis of 4-[(R)-4-acryloyl-3-methylpiperazin-1-yl)-6-chloro-7-(5-methyl-1H-indazol-4-yl)]-2H-methen-2-one

[0067] The operation was the same as in Example 7, except that (R)-4-(4-acryloyl-3-methylpiperazin-1-yl)-7-bromo-6-chloro-2H-chromen-2-one and 5-methyl-1-H-indazolenaphthalene borate were used instead of 4-(4-acryloylpiperazin-1-yl)-7-bromo-6-fluoro-2H-chromen-2-one and 3-hydroxynaphthalene borate, respectively, to obtain a white solid. The product test data are as follows:

[0068] Yield: 34%; 1H NMR (600MHz, DMSO-d6) δ13.18 (s, 1H), 7.93 (s, 1H), 7.71-7.30 (m, 4H), 6.84 (s, 1H), 6.18 (d, J=16.8Hz, 1H ), 5.86-5.75(m, 2H), 4.80-4.04(m, 2H), 3.69-3.64(m, 3H), 3.17-2.98(m, 2H), 2.18(s, 3H), 1.42(s, 3H); 13 C NMR (150MHz, DMSO-d6) δ164.97, 161.10, 160.18, 152.65, 141.16, 139.02, 132.53, 129.29, 128.46, 128.14, 1 27.32, 125.72, 123.31, 120.51, 117.20, 110.82, 98.35, 55.06(2C), 50.93(2C), 19.00, 16.15.HRMS(ESI)m / z calculated for C 25 H 23 ClN4O3[M+H] + 463.1531, found 463.1510.

[0069] In the cell proliferation inhibition experiment described later, the sample number of this example is 7.

[0070] Example 10: Synthesis of (R)-6-chloro-7-(3-methoxynaphthalen-1-yl)-4-[3-methyl-4-(5-nitrofuran-2-carbonyl)piperazin-1-yl]-2H-methen-2-one

[0071] To a DMF solution of compound (R)-6-chloro-7-(3-methoxynaphthalen-1-yl)-4-(3-methylpiperazin-1-yl)-2H-methen-2-one (100 mg, 0.24 mmol, 1.0 eq.) were added EDCI·HCl (91 mg, 0.48 mmol, 1.98 eq.), HOBt (12.8 mg, 0.096 mmol, 0.4 eq.), and 5-nitrofuran-2-carboxylic acid (42 mg, 0.27 mmol, 1.12 eq.). The temperature was lowered to 0°C, and DIPEA (0.13 mL, 0.72 mmol, 3.0 eq.) was added. The reaction mixture was allowed to warm to room temperature and allowed to react for 2 hours. 15% aqueous sodium bicarbonate solution was then added, and the reaction mixture was extracted with ethyl acetate (3 x 15 mL). The organic phases were combined, dried over anhydrous sodium sulfate, concentrated under reduced pressure, and purified by column chromatography to obtain a white solid. The product assay data are as follows:

[0072] Yield: 52% 1H NMR (600MHz, DMSO-d6) δ7.97-7.90 (m, 2H), 7.81 (d, J=4.2Hz, 1H), 7.54 (d, J=1 .8Hz, 1H), 7.53-7.49 (m, 1H), 7.47 (d, J=2-4Hz, 1H), 7.35 (d, J=4.2Hz, 1H), 7.3 2(t, J=3.0Hz, 2H), 7.14-7.10 (m, 1H), 5.88 (d, J=4.2Hz, 1H), 4.50-4.52 (m, 1H ), 3.74-3.64(m, 3H), 3.74-3.71(m, 3H), 3.28-3.03(m, 2H), 1.56-1.51(m, 3H). 13 C NMR (150MHz, DMSO-d6) δ161.0, 160.0, 157.3, 156.8, 152.6, 151.7, 148.0, 141.8, 137.3, 135.0, 128.4, 127.8, 127. 2, 126.8, 125.6, 125.5, 124.7, 121.0, 119.9, 117.7, 117.3, 113.4, 107.4, 98.4, 55.8, 55.4, 50.9, 50.8, 40.5, 16.6. HRMS(ESI)m / z calculated for C 30 H 24 ClN3O7[M+H] + 574.1375, found 574.1347.

[0073] In the cell proliferation inhibition experiment described later, the sample number of this example is 8.

[0074] Example 11: Synthesis of (R)-6-chloro-7-(3-hydroxynaphthalen-1-yl)-4-[3-methyl-4-(5-nitrofuran-2-carbonyl)piperazin-1-yl]-2H-methen-2-one

[0075] Under a nitrogen atmosphere at -78°C, compound (R)-6-chloro-7-(3-methoxynaphthalen-1-yl)-4-(3-methyl-4-(5-nitrofuran-2-carbonyl)piperazin-1-yl)-2H-methen-2-one (80 mg, 0.14 mmol, 1.0 eq.) was dissolved in dry DCM (0.7 mL). The temperature was cooled to -78°C, and 0.7 mL of boron tribromide (1 min DCM, 0.7 mmol) was slowly added. The mixture was kept under nitrogen and allowed to warm to room temperature for 2 hours. The reaction mixture was cooled to -30°C, and saturated sodium bicarbonate aqueous solution was added to alkalize the pH to approximately 8. The reaction mixture was extracted with DCM (3 × 10 mL). The organic phases were combined, washed once with water and once with salt, dried over anhydrous sodium sulfate, and evaporated to dryness to obtain an oily product, which was purified by column chromatography to obtain a white solid. The product assay data are as follows:

[0076] Yield: 28% 1 H NMR (400MHz, Chloroform-d) δ7.76 (s, 1H), 7.69 (d, J=8.4Hz, 1H), 7.39-7.36 (m , 1H), 7.35-7.31 (m, 2H), 7.27 (d, J=8.4Hz, 1H), 7.24-7.21 (m, 2H), 6.99 (t, J=3. 2Hz, 1H), 5.80 (d, J=1.6Hz, 1H), 5.67-5.61 (m, 1H), 4.89-4.85 (m, 1H), 4.53-4.5 1(m, 1H), 3.77-3.52(m, 2H), 3.18-3.05(m, 1H), 2.99-2.95(m, 1H), 1.65(s, 3H); 13 C NMR (150MHz, DMSO-d6) δ161.1, 160.0, 157.3, 154.9, 152.56, 151.7, 148.0, 142.2, 137.3, 135.3, 128.4, 127.1, 126.9, 1 25.9, 125.6, 125.5, 123.8, 120.9, 120.0, 117.7, 117.2, 113.4, 110.2, 99.2, 55.3, 50.9, 50.8, 40.5, 17.6.HRMS (ESI) m / z calculated for C 28 H 24 ClFN2O4[M+H] + 560.1218, found 560.1187.

[0077] In the cell proliferation inhibition experiment described later, the sample number of this example is 9.

[0078] Example 12: Synthesis of (R)-6-chloro-4-[4-(2-chloroacetyl)-3-methylpiperazin-1-yl]-7-(3-methoxy-naphthalen-1-yl)-2H-methen-2-one

[0079] Compound (R)-6-chloro-7-(3-methoxynaphthalen-1-yl)-4-[3-methyl-4-(5-nitrofuran-2-carbonyl)piperazin-1-yl]-2H-methen-2-one (120 mg, 0.28 mmol, 1.0 eq.) and triethylamine (0.08 mL, 0.26 mmol, 2.0 eq.) were dissolved in dry DCM solution (5 mL). The temperature was cooled to -70°C. Chloroacetyl chloride (35 mg, 0.3 mmol, 1.1 eq.) was added dropwise to the reaction solution, and the reaction was incubated at -70°C for 20 minutes. The reaction solution was extracted with DCM, and the organic phase was washed once with brine and dried over anhydrous sodium sulfate.

[0080] The solvent was evaporated and the product was purified by column chromatography to obtain a white solid. The product test data are as follows:

[0081] Yield: 47% 1 H NMR (600MHz, DMSO-d6) δ7.97-7.86 (m, 2H), 7.59-7.45 (m, 3H), 7.31 (d, J=4.2Hz, 1H), 7.11 (d, J=2.4Hz, 1H), 5.87 (s, 1H ), 4.68-4.53(m, 1H), 4.46(s, 2H), 4.31(s, 1H), 3.93(s, 3H), 3.86-3.55(m, 3H), 3.20-2.89(m, 2H), 1.65-1.22(m, 3H); 13 C NMR (150MHz, DMSO-d6) δ165.3, 161.1, 160.1, 156.8, 152.6, 141.8, 137.3, 135.0, 128.4, 127.8, 127.2, 126.8, 125.5(2C),124.7,121.0,119.9,117.4,107.4,98.4,55.9,50.7,49.2,45.5,42.6,40.8,15.9.HRMS(ESI)m / z calculated for C 27 H 24 Cl2N2O4[M+H] + 511.1185, found511.1163.

[0082] In the cell proliferation inhibition experiment described later, the sample number of this example is 10.

[0083] Example 13: Synthesis of Compound (R)-6-chloro-4-[4-(2-chloroacetyl)-3-methylpiperazin-1-yl)-7-(3-hydroxy-naphthalen-1-yl]-2H-methen-2-one

[0084] The operation process was the same as that of Example 15, except that (R)-6-chloro-4-[4-(2-chloroacetyl)-3-methylpiperazin-1-yl]-7-(3-methoxy-naphthalen-1-yl)-2H-methen-2-one was used instead of (R)-6-chloro-7-(3-methoxynaphthalen-1-yl)-4-(3-methyl-4-(5-nitrofuran-2-carbonyl)piperazin-1-yl)-2H-methen-2-one. Purification by column chromatography gave a white solid. The product test data are as follows:

[0085] Yield: 24% 1 H NMR (600MHz, DMSO-d6) δ10.01 (s, 1H), 7.90 (d, J = 4.2Hz, 1H), 7.79 (d, J = 8.4Hz, 1H), 7.52 (s, 1H), 7.43 (t, J = 7.2Hz, 1H), 7.33-7.15 (m, 3H), 7.02 (d, J=1.8Hz, 1H), 5.86 (s, 1H), 4.66-4.68 (m, 1H), 4.46 (s, 2H), 4.31 (s, 1H), 3.89-3.62 (m, 3H), 3.23-2.93 (m, 2H), 1.68-1.24 (m, 3H); 13 C NMR (150MHz, DMSO-d6) δ165.3, 161.1, 160.2, 154.9, 152.6, 152.5, 142.1, 137.3, 135.3, 128.4, 127.1, 126.9, 125 .9, 125.5, 123.8, 120.9, 119.9, 117.3, 110.2, 98.3, 54.6, 50.7, 49.2, 45.5, 42.6, 15.9.HRMS (ESI) m / zcalculated for C 26 H 22 Cl2N2O4[M+H] + 497.1029, found 497.1008.

[0086] In the cell proliferation inhibition experiment described later, the sample number of this example is 11.

[0087] Example 14: Proliferation Inhibitory Activity of Compounds 1-11

[0088] The in vitro cytotoxic activity of the compounds against KRAS-G12C high-expressing cell lines NCI-H23 or NCI-H358 (non-small cell lung cancer cells) was determined by the MTT assay. The in vitro cytotoxic activity of the compounds against KRAS-G12C mutant MIA PaCa-2 cells (pancreatic cancer cells) was determined by the CCK8 assay. The KRAS-G12C inhibitor ARS1620 was used as a positive control. The results are shown in Table 1.

[0089] MTT method: NCI-H23 or NCI-H358 cells (5×10 6 Cells (cells / well) were added to a 96-well plate containing 10% FBS and cultured in an incubator (37°C, 5.0% CO2) for 24 hours. Drugs at varying concentrations were then added and cultured for 72 hours. MTT solution (5 mg / mL) was then added and cultured for 4 hours. The culture medium was discarded, DMSO was added, and the cells were shaken to mix. The absorbance was measured at 490 nm using a microplate reader. IC values ​​were calculated using GraphPad Prism Software version 5.02. 50 value.

[0090] CCK8 method: The culture and drug addition methods are similar to those of the MTT method. CCK8 solution is used instead of the MTT solution in the above method to determine the in vitro anti-proliferative activity of pancreatic cancer cell MIA PaCa-2.

[0091] Example 15: Inhibitory activity of compound 1-11 on KRAS-G12C protein

[0092] The TR-FRET method was used to study the in vitro inhibitory activity of coumarin compounds 1-11 against KRAS-G12C using the KRAS-G12C inhibitor ARS1620 as the positive compound. The experimental results are shown in Table 1.

[0093] TR-FRET assay: Prepare compound solutions of varying concentration gradients and add them to a 384-well Source plate. Add equal amounts of positive control compound and DMSO. Prepare buffer and protein solution, centrifuge, and incubate. Add the prepared peptide solution and centrifuge. Add the test solution and continue centrifugation and incubation. After the compound binds to the enzyme protein, calculate the IC using GraphPad Prism 5. 50 value.

[0094] Table 1 Antiproliferative activity of compounds 1-11 against KRAS-G12C mutant cells

[0095]

[0096]

[0097] a The mean of two experiments

[0098] b The average of three experiments, time 72h

[0099] c MIA PaCa-2 cells were determined by CCK8 assay, and other cells were determined by MrT assay.

[0100] The results in Table 1 show that the coumarin compounds of the present invention have potent inhibitory activity against KARS-G12C protein and its mutant cell lines, and most compounds have better inhibitory and anti-proliferative activities than the positive control drug ARS1620; among them, compounds 8-10 have a half-maximal inhibitory concentration (IC50) against non-small cell lung cancer NCI-H23 and NCI-H358. 50 ) values ​​were all less than 2 μM, and the anti-proliferative activity was nearly 10-fold higher than that of ARS1620. Compounds 8-11 showed outstanding anti-proliferative activity against pancreatic cancer cells MIAPaCa-2, which was superior to ARS1620.

[0101] Comparative Example

[0102] Comparison of efficacy

[0103] The present invention presents a novel class of coumarin-based anti-tumor compounds. Compared to the active agent ARS1620, compounds 8-10 exhibited nearly 10-fold increased anti-proliferative activity against non-small cell lung cancer cells NCI-H23 and NCI-H358. Compound 9 exhibited potent anti-proliferative activity against pancreatic cancer cells MIA PaCa-2, with approximately 8-fold increased cytotoxicity compared to ARS1620, overcoming the problem of ARS1620's insensitivity to pancreatic cancer cells.

[0104] The preferred embodiments of the present invention have been described in detail above. However, the present invention is not limited to the specific details of the aforementioned embodiments. Within the scope of the technical concept of the present invention, various equivalent transformations can be made to the technical solutions of the present invention. To avoid unnecessary repetition, the present invention will not further describe various possible combinations. Any modifications, equivalent substitutions, or improvements made within the scope of the technical concept of the present invention are included in the scope of protection of the present invention.

Claims

1. A coumarin derivative or a pharmaceutically acceptable salt thereof, characterized in that: The compound is shown in Formula I. in: X 1 and X 2 are the same or different and are independently selected from hydrogen, halogen, allyl or alkoxy; Ar is selected from C 6-10 Aryl or C 5-10 heteroaryl; R 1 and R 2 the same or different, independently selected from hydrogen, C 1-6 Alkyl or cyano; R 3 Selected from C 1-4 Alkenyl, C 1-6 Halogenated alkyl, C 5-10 Heteroaryl, C 1-4 Halogenated alkenyl or C 3-6 Halogenated cycloalkyl.

2. The compound according to claim 1 and its pharmaceutically acceptable salt, characterized in that: The X 1 is hydrogen, fluorine or chlorine, X 2 is hydrogen; Ar is shown to be 2-fluorophenyl, 2-trifluoromethylphenyl, 2-fluoro-6-hydroxyphenyl, 3-methoxy-naphthyl-1, 3-hydroxy-naphthyl-1 or 5-methyl-1H-indazolyl-4; The R 1 is hydrogen, methyl, or S-configuration methyl, R 2 is hydrogen, methyl or R-configuration methyl; The R 3 is vinyl, chloromethyl or 5-nitrofuryl-2-yl.

3. Use of the coumarin derivatives or pharmaceutically acceptable salts thereof according to claims 1-2 for preparing KRAS-G12C inhibitor anticancer drugs.

4. Use of the coumarin derivatives or pharmaceutically acceptable salts thereof according to claims 1-2 in the preparation of drugs for treating non-small cell lung cancer.

5. Use of the coumarin derivatives or pharmaceutically acceptable salts thereof according to claims 1-2 in the preparation of anti-pancreatic cancer drugs.

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