Pol theta inhibitor and preparation method, pharmaceutical composition and application thereof

Abstract

The invention discloses a Pol theta inhibitor as well as a preparation method, a pharmaceutical composition and application thereof. The compound disclosed by the invention is shown as a general formula (I), further comprises stereoisomers, pharmaceutically acceptable salts or a mixture of the stereoisomers and the pharmaceutically acceptable salts, can effectively inhibit Pol theta enzyme activity, is used for preparing medicines for treating colon cancer and breast cancer, and is wide in application, the preparation method is high in applicability, and the structure is easy to expand. # imgabs0 #

Description

Technical Field

[0001] The present invention relates to a compound, a preparation method and an application thereof, and in particular to a preparation method, a pharmaceutical composition and an application of a Polθ inhibitor. Background Art

[0002] Targeting DNA repair defects has become an effective strategy for treating cancer. However, cancers with DNA repair defects often rely on alternative DNA repair pathways, which are also the basis for synthetic lethality. The successful example of synthetic lethality is the treatment of BRCA-deficient breast and ovarian cancers with poly (ADP-ribose) polymerase (PARP) inhibitors.

[0003] DNA damage repair processes are essential for genome maintenance and stability, among which double-strand breaks (DSBs) are primarily repaired by the nonhomologous end joining (NHEJ) pathway and homologous recombination (HR) during the S-G2 phase. A less readily resolvable alternative end joining (alt-EJ), also known as the microhomology-mediated end joining (MMEJ) pathway, is often considered a "backup" DSB repair pathway when NHEJ or HR is impaired. Many genetic studies have highlighted the role of Polθ in stimulating MMEJ in higher organisms.

[0004] Polθ includes a C-terminal DNA polymerase domain and an N-terminal ATPase / helicase domain belonging to the HELQ class of the SF2 helicase superfamily. In homologous recombination-deficient cells, Polθ can perform error-prone DNA synthesis at DNA damage sites via the alt-EJ pathway. Studies have shown that the helicase domain of Polθ inhibits the HR pathway by disrupting the formation of the Rad51 nucleoprotein complex, which is involved in the initiation of HR-dependent DNA repair reactions after ionizing radiation. This anti-recombinase activity of Polθ promotes the alt-EJ pathway. In addition, the helicase domain of Polθ facilitates microhomology-mediated strand annealing. When the ssDNA overhang contains microhomology greater than 2 bp, Polθ effectively promotes end joining in the alt-EJ pathway through this annealing activity. This reannealing activity is achieved through the coupling of Rad51 interactions, followed by ATPase-mediated displacement of Rad51 from the DSB damage site. After annealing, the primer strand of DNA can be extended by the polymerase domain of Polθ.

[0005] Polθ is not expressed in normal cells, but is upregulated in breast, lung, and ovarian cancers. In addition, increased Polθ expression is associated with poor prognosis in breast cancer. Studies have shown that cancer cells lacking HR, NHEJ, or ATM are highly dependent on Polθ expression. Summary of the invention

[0006] Objectives of the invention: The first objective of the present invention is to provide a compound as a Polθ inhibitor. The second objective of the present invention is to provide a method for preparing a Polθ inhibitor. The third objective of the present invention is to provide a pharmaceutical composition. The fourth objective of the present invention is to provide the use of the compound or a pharmaceutically acceptable salt thereof in the preparation of a drug for treating or preventing cancer with homologous recombination deficiency.

[0007] Technical solution: The present invention provides a Polθ inhibitor as shown in the general formula (I), its tautomer or pharmaceutically acceptable salt:

[0008]

[0009] in:

[0010] X is selected from C or N;

[0011] Y is selected from CH or N;

[0012] C is selected from -CH2-, O or no atom;

[0013] R1 is selected from C1-C4 alkyl, H or absent;

[0014] R2, R3 are selected from H or C1-C4 alkyl;

[0015] A is selected from:

[0016] or thienyl;

[0017] Z is selected from C or N;

[0018] R4 is selected from H, halogen, C1-C4 alkyl, CHO, CF3, NO2, CN;

[0019] R5 is selected from H, halogen, C1-C4 alkyl, CHO, CF3, NO2, CN;

[0020] R6 is selected from H, halogen;

[0021] B is selected from substituted phenyl, -(CH2) 0～3 -C≡CH, -(CH2) 0～3 -CH=CH2, C3-C6 cycloalkyl; the substituent is selected from at least one halogen, C1-C4 alkoxy, CN.

[0022] The Polθ inhibitor, its tautomer or pharmaceutically acceptable salt:

[0023] in:

[0024] X is selected from C or N;

[0025] Y is selected from CH or N;

[0026] C is selected from -CH2-, O or no atom;

[0027] R1 is selected from C1-C4 alkyl, H or absent;

[0028] R2, R3 are selected from H or CH3;

[0029] A is selected from:

[0030]

[0031] Z is selected from C or N;

[0032] R4 is selected from H, F, Cl, Br, I, CH3, CHO, CF3, NO2, CN;

[0033] R5 is selected from H, F, Cl, Br, I, CH3, CHO, CF3, NO2, CN;

[0034] R6 is selected from H, F, Cl, Br, I;

[0035] B is selected from substituted phenyl, ethynyl, vinyl, C3-C6 cycloalkyl; the substituent is selected from at least one of F, Cl, Br, I, OCH3, CN.

[0036] The Polθ inhibitor, its tautomer or pharmaceutically acceptable salt:

[0037] When X is C and Y is N, R1 is CH3; when X is N and Y is CH, R1 is absent;

[0038] C is selected from -CH2-, O or no atom;

[0039] R2, R3 are selected from at least one H or CH3;

[0040] A is selected from:

[0041]

[0042] Z is selected from C or N;

[0043] R4 is selected from H, F, Cl, CH3, CHO;

[0044] R5 is selected from H, F;

[0045] R6 is selected from H, F;

[0046] B is selected from substituted phenyl, alkynyl, alkenyl, C3-C6 cycloalkyl; the substituent is selected from at least one of F, Cl, OCH3, CN.

[0047] A Polθ inhibitor, a tautomer or a pharmaceutically acceptable salt thereof, selected from any one of the following compounds:

[0048]

[0049]

[0050] Use of the Polθ inhibitor, its tautomer or pharmaceutically acceptable salt in the preparation of Polθ inhibitor.

[0051] Use of the Polθ inhibitor, its tautomer or pharmaceutically acceptable salt according to claim 1 in the preparation of a drug for treating or preventing cancers defective in the homologous recombination repair pathway.

[0052] A pharmaceutical composition comprises the Polθ inhibitor, its tautomer or pharmaceutically acceptable salt and a pharmaceutically acceptable excipient.

[0053] Use of the Polθ inhibitor, its tautomer or pharmaceutically acceptable salt or the pharmaceutical composition in preparing drugs for treating or preventing colon cancer or breast cancer.

[0054] The method for preparing the Polθ inhibitor, its tautomer or pharmaceutically acceptable salt is characterized by comprising the following steps:

[0055] (a) Compound 1 and 2 are subjected to SUZUKI reaction to obtain compound 3;

[0056] (b) Compound 3 is hydrolyzed under alkaline conditions to obtain compound 4;

[0057] (c) Compound 5 reacts with selenium powder and sodium borohydride to produce compound 6;

[0058] (d) Compound 6 and compound 7 react with POCl3 to produce compound 8;

[0059] (e) Compound 4 and Compound 8 react with an amide condensation agent TCFH to generate compound SEP, i.e., a compound represented by general formula (I);

[0060]

[0061] When X is selected from C or N;

[0062] Y is selected from CH or N;

[0063] C is selected from -CH2-, O or no atom;

[0064] R1 is selected from C1-C4 alkyl, H or absent;

[0065] R2, R3 are selected from H or C1-C4 alkyl;

[0066] A is selected from:

[0067]

[0068] Z is selected from C or N;

[0069] R4 is selected from H, halogen, C1-C4 alkyl, CHO, CF3, NO2, CN;

[0070] R5 is selected from H, halogen, C1-C4 alkyl, CHO, CF3, NO2, CN;

[0071] R6 is selected from H, halogen;

[0072] B is selected from substituted phenyl, -(CH2) 0～3 -C≡CH, -(CH2) 0～3 -CH=CH2, C3-C6 cycloalkyl; the substituent is selected from at least one halogen, C1-C4 alkoxy, CN.

[0073] The method for preparing the Polθ inhibitor, its tautomer or pharmaceutically acceptable salt is characterized in that the specific reaction reagents and conditions are:

[0074] (a) Pd(dppf)Cl2, K2CO3, 1,4-Dioxane, H2O, 70-90℃; (b) NaOH, MeOH, 40-60℃; (c) CH3I, Se, N aBH4, EtOH, N2, -15-80°C; (d) POCl3, 1,4-Dioxane, 75-90°C; (e) TCFH, NMI, MeCN, N2, 40-70°C.

[0075] Beneficial effects: The present invention designs a novel class of Polθ inhibitors with selenium atom insertion. Polθ is involved in microhomology-mediated end joining (MMEJ), is almost not expressed in normal cells but is overexpressed in many tumors, and is a potential drug target for the treatment of HR-deficient tumors. DETAILED DESCRIPTION

[0076] The NMR spectra of the final products and intermediates in the examples were measured by Bruker's 300 MHz, 400 MHz or 600 MHz NMR spectrometer with DMSO-d6 or CDCl3-d3 as solvent and TMS as internal standard; the high-resolution mass spectrometry (HRMS) was measured by Agilent's Q-TOF 6520 mass spectrometer.

[0077] The reagents used in the synthesis, purification and separation of the compounds are as follows: (1) Column chromatography silica gel: 200 or 300 mesh silica gel were purchased from Qingdao Ocean Chemical Industry; (2) HSGF254 TLC thin layer chromatography plate: purchased from Yantai Chemical Research Institute; (3) Conventional solvents used in column chromatography elution systems such as petroleum ether, dichloromethane, ethyl acetate, methanol, etc. and chemical reagents required for the reaction, unless otherwise specified, were all commercially available chemically pure or analytically pure products.

[0078] Example 1: Synthesis of N-(5-(4-chlorophenethyl)-1,3,4-selenodiazol-2-yl)-3-(2-methoxyphenyl)isonicotinamide (SEP-1)

[0079] Synthesis route:

[0080]

[0081] Step 1:

[0082] At room temperature, compound 9 (1.0 g, 6.28 mmol) and compound 10 (1.2 g, 5.48 mmol) were dissolved in 8 ml 1,4-dioxane, potassium carbonate (1.5 g, 10.96 mmol), 1 ml water and Pd(dppf)Cl2 (144 mg, 0.03 mmol) were added in sequence, and the mixture was reacted at 80°C overnight under nitrogen protection. TLC monitored the completion of the reaction. After the reaction was cooled to room temperature, water was added, extracted with ethyl acetate, washed with water, washed with saturated brine, dried over anhydrous Na2SO4, distilled under reduced pressure, and column chromatography was performed to obtain 1.0 g of a white solid.

[0083] Step 2:

[0084] Compound 11 (1.0 g, 4.11 mmol) was dissolved in 10 mL methanol and water (1:1), and NaOH (493 mg, 12.33 mmol) was added. The mixture was reacted at 50°C for 3 h. The reaction was completed after TCL monitoring. The reaction solution was concentrated until no fraction flowed out, and the pH was adjusted to 4-5 with 10% dilute hydrochloric acid. A white solid was precipitated, filtered, and dried at 50°C to obtain 800 mg of a white solid. The product was directly added without purification.

[0085] Step 3:

[0086] Compound 13 (12.0 g, 131.86 mmol) and iodomethane (22.4 g, 157.84 mmol) were suspended in 120 ml of anhydrous ethanol, refluxed at 80 ° C for 2 h, crystallized at 0 ° C, filtered, and dried to obtain S-methylthiosemicarbazide, a white solid of 10.0 g. Sodium borohydride (8.6 g, 225 mmol) was added to a three-necked flask, replaced with nitrogen, and a suspension of selenium powder and anhydrous ethanol was slowly injected at -15 ° C. After the addition, it was stirred for 2 h, and the hot ethanol solution of S-methylthiosemicarbazide was injected into the system, and the reaction was allowed to proceed overnight at room temperature. After filtering, the filter cake was washed with anhydrous ethanol, and dried to obtain 13.0 g of off-white aminoselenourea.

[0087] Step 4:

[0088] Compound 14 (188.0 mg, 1.35 mmol) and compound 15 (250.0 mg, 1.35 mmol) were suspended in 2.5 ml 1,4-dioxane, phosphorus oxychloride (1.67 g, 12.15 mmol) was slowly added, and refluxed at 75°C for 1 h. After the reaction was completed by TLC monitoring, the reaction solution was poured into ice water, and the pH was adjusted to 8 with sodium bicarbonate, extracted, and concentrated under reduced pressure to obtain 150 mg of brown-yellow solid. It was directly cast without purification.

[0089] Step 5:

[0090] Compound 12 (143.8 mg, 0.63 mmol) and compound 16 (150 mg, 0.52 mmol) were suspended in 15 ml of acetonitrile, and 1.5 ml of N-methylimidazole and N,N,N′,N′-tetramethylchloroformamidine hexafluorophosphate (172 mg, 0.68 mmol) were added in sequence. The mixture was reacted at 50°C overnight under N2 protection, and the reaction was completed after monitoring by TCL. Water was added, the mixture was extracted with ethyl acetate, washed with water, washed with saturated brine, dried over anhydrous Na2SO4, distilled under reduced pressure, and column chromatography was performed to obtain 30 mg of a white solid. 1HNMR (300MHz, CDCl3) δ8.67 (d, J=5.1Hz, 1H), 8.59 (s, 1H), 7.57 (d, J=4.8Hz, 1H), 7.34-7.27 (m, 1H), 7.26 (s, 1H), 7.24-7.23 (m , 1H), 7.18-7.08 (m, 3H), 6.93 (t, J=6.6Hz, 1H), 6.80 (d, J=8.4Hz, 1H), 3.49 (s, 3H), 3.28 (t, J=7.5Hz, 2H), 3.03 (t, J=7.5Hz, 2H).

[0091] Example 2: Synthesis of N-(5-(4-fluorophenethyl)-1,3,4-selenodiazolyl-2-yl)-3-(2-methoxyphenyl)isonicotinamide (SEP-2)

[0092]

[0093] Referring to the synthesis method of Example 1, compound 15 was replaced with 3-(4-fluorophenyl)propionic acid, and other conditions remained unchanged. 1 H NMR (300MHz, CDCl3) δ11.04 (s, 1H), 8.73 (d, J=5.1Hz, 1H), 8.63 (s, 1H), 7.63 (d, J=5.1Hz, 1H), 7.40-7.34 (m, 1H), 7.22 ( dd, J=7.5, 1.8Hz, 1H), 7.20-7.14 (m, 2H), 7.05-6.95 (m, 3H), 6.88 (d, J=8.4Hz, 1H), 3.60 (s, 3H), 3.31 (t, J=6.9Hz, 2H),

[0094] 3.06 (t, J = 7.8 Hz, 2H).

[0095] Example 3: Synthesis of N-(5-(4-methoxyphenethyl)-1,3,4-selenodiazol-2-yl)-3-(2-methoxyphenyl)isonicotinamide (SEP-3)

[0096]

[0097] Referring to the synthesis method of Example 1, compound 15 was replaced with 3-(4-methoxyphenyl)propionic acid, and other conditions remained unchanged. 1 H NMR (400MHz, CDCl3) δ8.72 (d, J=5.2Hz, 1H), 8.63 (s, 1H), 7.61 (d, J=7.2Hz, 1H), 7.38-7.34 (m, 1H), 7.22 (dd, J=7.6, 1.6Hz, 1H), 7. 15-7.10 (m, 2H), 7.02 (t, J=7.2Hz, 1H), 6.88-6.83 (m, 3H), 3.79 (s, 3H), 3.59 (s, 3H), 3.31 (t, J=7.6Hz, 2H), 3.02 (t, J=7.6Hz, 2H).

[0098] Example 4: Synthesis of N-(5-(4-cyanophenyl)-1,3,4-selenodiazole-2-yl)-3-(2-methoxyphenyl)isonicotinamide (SEP-4)

[0099]

[0100] Referring to the synthesis method of Example 1, compound 15 was replaced with 3-(4-cyanophenyl)propionic acid, and other conditions remained unchanged. 1H NMR (300MHz, CDCl3) δ10.99 (s, 1H), 8.74 (d, J=5.1Hz, 1H), 8.64 (s, 1H), 7.65 (d, J=4.8Hz, 1H), 7.60-7.55 (m, 2H), 7.43-7.37 (m, 1H), 7. 34-7.29 (m, 2H), 7.26-7.21 (m, 1H), 7.08-7.02 (m, 1H), 6.90 (d, J=8.4Hz, 1H), 3.62 (s, 3H), 3.34 (t, J=6.9Hz, 2H), 3.16 (t, J=7.8Hz, 2H).

[0101] Example 5: Synthesis of N-(5-(2-cyclopropylethyl)-1,3,4-selenodiazol-2-yl)-3-(2-methoxyphenyl)isonicotinamide (SEP-5)

[0102]

[0103] Referring to the synthesis method of Example 1, compound 15 was replaced by 3-cyclopropylpropionic acid, and other conditions remained unchanged. 1HNMR (300MHz, CDCl3) δ8.68 (d, J=5.1Hz, 1H), 8.60 (s, 1H), 7.59 (d, J=5.1Hz, 1H), 7.34-7.28 (m, 1H), 7.19 (dd, J=7.5, 1.8Hz, 1H), 6.98-6.93 (m , 1H), 6.82 (d, J=8.1Hz, 1H), 3.55 (s, 3H), 3.08 (t, J=7.5Hz, 2H), 1.65 (q , J=7.3Hz, 2H), 0.86-0.69 (m, 1H), 0.53-0.42 (m, 2H), 0.12-0.07 (m, 2H).

[0104] Example 6: Synthesis of N-(5-(3-butynyl)-1,3,4-selenodiazol-2-yl)-3-(2-methoxyphenyl)isonicotinamide (SEP-6)

[0105]

[0106] Referring to the synthesis method of Example 1, compound 15 was replaced by 4-pentynoic acid, and other conditions remained unchanged. 1H NMR (300MHz, CDCl3) δ8.73 (d, J=5.1Hz, 1H), 8.63 (s, 1H), 7.63 (d, J=4.8Hz, 1H), 7.35 (td, J=7.8, 1.5Hz, 1H), 7.23 (dd, J=7.5, 1.7Hz, 1H ), 7.00 (t, J=7.5Hz, 1H), 6.86 (d, J=8.4Hz, 1H), 3.59 (s, 3H), 3.22 (t, J=6.6Hz, 2H), 2.63 (td, J=6.6, 2.4Hz, 2H), 2.11 (t, J=2.7Hz, 1H).

[0107] Example 7: Synthesis of N-(5-(3-butenyl)-1,3,4-selenodiazol-2-yl)-3-(2-methoxyphenyl)isonicotinamide (SEP-7)

[0108]

[0109] Referring to the synthesis method of Example 1, compound 15 was replaced by 4-pentenoic acid, and other conditions remained unchanged. 1 H NMR (300MHz, CDCl3) δ8.63 (d, J=4.8Hz, 1H), 8.56 (s, 1H), 7.55 (d, J=4.8Hz, 1H), 7.30-7.23 (m, 1H), 7.14 (dd, J=7.5, 1.8Hz, 1H), 6.90 (td .

[0110] Example 8: Synthesis of N-(5-(4-cyanophenyl)-1,3,4-selenodiazole-2-yl)-4-(5-chloro-2-methoxyphenyl)-6-methylnicotinamide (SEP-8)

[0111]

[0112] Referring to the synthesis method of Example 1, Compound 9 and Compound 10 were replaced by 5-chloro-2-methoxyphenylboronic acid and 4-bromo-6-methylnicotinate, respectively, and other conditions remained unchanged. 1H NMR (600MHz, CDCl3) δ8.91 (s, 1H), 7.59 (s, 1H), 7.57 (s, 1H), 7.38 (dd, J=9.0, 2.4Hz, 1H), 7.33 (s, 1H), 7.32 (s, 1H), 7.25 (d , J=2.4Hz, 1H), 7.15 (s, 1H), 6.85 (d, J=9.0Hz, 1H), 3.63 (s, 3H), 3.35 (t, J=7.8Hz, 2H), 3.18 (t, J=7.8Hz, 2H), 2.67 (s, 3H).

[0113] Example 9: Synthesis of N-(5-(4-cyanophenyl)-1,3,4-selenodiazole-2-yl)-4-(5-fluoro-2-methoxyphenyl)-6-methylnicotinamide (SEP-9)

[0114]

[0115] Referring to the synthesis method of Example 1, Compound 9 and Compound 10 were replaced by 5-fluoro-2-methoxyphenylboronic acid and 4-bromo-6-methylnicotinate, respectively, and other conditions remained unchanged. 1 H NMR (600MHz, CDCl3) δ8.92 (s, 1H), 7.58 (s, 1H), 7.57 (s, 1H), 7.33 (s, 1H), 7.31 (s, 1H), 7.14 (s, 1H), 7.09 (td, J=9.0, 3.0Hz, 1H), 6.99 (dd, J=8.4, 3.0Hz, 1H), 6.83 (dd, J=9.0, 4.2Hz, 1H), 3.59 (s, 3H), 3.33 (t, J=7.8Hz, 2H), 3.17 (t, J=7.8Hz, 2H), 2.66 (s, 3H).

[0116] Example 10: Synthesis of N-(5-(4-cyanophenyl)-1,3,4-selenodiazole-2-yl)-4-(2-fluoro-6-methoxy-3-methylphenyl)-6-methylnicotinamide (SEP-10)

[0117]

[0118] Referring to the synthesis method of Example 1, Compound 9 and Compound 10 were replaced by 2-fluoro-6-methoxy-3-methylphenylboronic acid and 4-bromo-6-methylnicotinate, respectively, and other conditions remained unchanged. 1H NMR (300MHz, CDCl3) δ9.04 (s, 1H), 7.57 (s, 1H), 7.54 (s, 1H), 7.32 (s, 1H), 7.29 (s, 1H), 7.23-7.15 (m, 2H), 6.6 3 (d, J=8.4Hz, 1H), 3.61 (s, 3H), 3.32 (t, J=6.9Hz, 2H), 3.15 (t, J=7.5Hz, 2H), 2.64 (s, 3H), 2.22-2.18 (m, 3H).

[0119] Example 11: Synthesis of N-(5-(4-cyanophenyl)-1,3,4-selenodiazole-2-yl)-6-methyl-4-(thiophen-3-yl)nicotinamide (SEP-11)

[0120]

[0121] Referring to the synthesis method of Example 1, Compound 9 and Compound 10 were replaced by thiophene-3-ylboronic acid and 4-bromo-6-methylnicotinate, respectively, and other conditions remained unchanged. 1 H NMR (300MHz, CDCl3) δ8.83 (s, 1H), 7.61 (s, 1H), 7.58 (s, 1H), 7.36-7.31 (m, 4H), 7.23 (s, 1 H), 7.03 (dd, J=4.8, 1.5Hz, 1H), 3.32 (t, J=6.9Hz, 2H), 3.15 (t, J=7.8Hz, 2H), 2.62 (s, 3H).

[0122] Example 12: Synthesis of 2′-chloro-N-(5-(4-cyanophenyl)-1,3,4-selenodiazole-2-yl)-5′-methoxy-6-methyl-[4,4′-bipyridine]-3-carboxamide (SEP-12)

[0123]

[0124] Referring to the synthesis method of Example 1, Compound 9 and Compound 10 were replaced by 2-chloro-5-methoxypyridine-4-boric acid and 4-bromo-6-methylnicotinate, respectively, and other conditions remained unchanged. 1 H NMR (300MHz, DMSO-d6) δ13.25 (s, 1H), 8.85 (s, 1H), 8.15 (s, 1H), 7.77 (s, 1H), 7.74 (s, 1H), 7.53 (s, 1H), 7.4 9 (s, 1H), 7.47 (s, 1H), 7.41 (s, 1H), 3.54 (s, 3H), 3.35 (d, J = 7.3Hz, 3H), 3.13 (t, J = 7.5Hz, 2H), 2.58 (s, 3H).

[0125] Example 13: Synthesis of N-(5-(4-cyanophenyl)-1,3,4-selenodiazole-2-yl)-4-(5-formyl-2-methoxyphenyl)-6-methylnicotinamide (SEP-13)

[0126]

[0127] Referring to the synthesis method of Example 1, Compound 9 and Compound 10 were replaced by 5-formyl-2-methoxyphenylboronic acid and 4-bromo-6-methylnicotinate, respectively, and other conditions remained unchanged. 1 H NMR (300MHz, CDCl3) δ9.89 (s, 1H), 9.03 (s, 1H), 7.91 (dd, J=8.7, 2.1Hz, 1H), 7.83 (d, J=2.1Hz, 1H), 7.58 (s, 1H), 7.56 (s, 1H), 7.3 3 (s, 1H), 7.31 (s, 1H), 7.22 (s, 1H), 6.98 (d, J = 8.5Hz, 1H), 3.63 (s, 3H), 3.31 (t, J = 8.1Hz, 2H), 3.16 (t, J = 7.8Hz, 2H), 2.68 (s, 3H).

[0128] Example 14: Synthesis of N-(5-(4-cyanophenyl)-1,3,4-selenodiazole-2-yl)-4-(2-fluoro-6-methoxyphenyl)-6-methylnicotinamide (SEP-14)

[0129]

[0130] Referring to the synthesis method of Example 1, Compound 9 and Compound 10 were replaced by 2-fluoro-6-methoxyphenylboronic acid and 4-bromo-6-methylnicotinate, respectively, and other conditions remained unchanged. 1 H NMR (600MHz, DMSO-d6) δ13.11 (s, 1H), 8.84 (s, 1H), 7.75 (s, 1H), 7.74 (s, 1H), 7.48 (s, 1H), 7.47 (s, 1H), 7.39 (td, J=8.4, 6.7Hz, 1 H), 7.31 (s, 1H), 6.90 (t, J=8.9Hz, 1H), 6.86 (d, J=8.4Hz, 1H), 3.51 (s, 3H), 3.31 (s, 2H), 3.15-3.10 (t, J=6.6Hz, 2H), 2.56 (s, 3H).

[0131] Example 15: Synthesis of N-(5-(4-cyanophenyl)-1,3,4-selenodiazole-2-yl)-4-(8-fluoroquinolin-5-yl)-6-methylnicotinamide (SEP-15)

[0132]

[0133] Referring to the synthesis method of Example 1, Compound 9 and Compound 10 were replaced by 8-fluoro-5-(4,4,5,5-tetramethyl-1,3,2-dioxaborin-2-yl)quinoline and 4-bromo-6-methylnicotinate, respectively, and other conditions remained unchanged. 1 H NMR (300MHz, CDCl3) δ9.41 (s, 1H), 8.99-8.97 (m, 1H), 7.87 (d, J = 8.6Hz, 1H), 7.58 (s, 1H), 7.55 (s, 1H), 7.49 (d, J = 8.0Hz, 1H), 7.44 (d, J = 3.2Hz, 1H), 7.43-7.39 (m, 1H), 7.36 (dd, J=8.6, 4.2Hz, 1H), 7.30-7.26 (m, 1H), 3.24 (t, J=7.5Hz, 2H), 3.10 (t, J=7.5Hz, 2H), 2.74 (s, 3H).

[0134] Example 16: Synthesis of 2′-chloro-N-(5-(4-cyanophenoxy)-1,3,4-selenadiazole-2-yl)-5′-methoxy-6-methyl-[4,4′-bipyridine]-3-carboxamide (SEP-16)

[0135]

[0136] Referring to the synthesis method of Example 1, Compound 9, Compound 10 and Compound 15 were replaced by 2-chloro-5-methoxypyridine-4-boric acid, 4-bromo-6-methylnicotinate and (4-cyanophenoxy)acetic acid, respectively, and other conditions remained unchanged. 1HNMR (300MHz, DMSO-d6) δ 13.50 (s, 1H), 8.87 (s, 1H), 8.14 (s, 1H), 7.84-7.82 (m, 1H), 7.81-7.79 (m, 1H), 7.55 (s, 1H), 7.43 (s, 1H), 7.27-7.25 (m, 1H), 7.24-7.22 (m, 1H), 5.56 (s, 2H), 3.55 (s, 3H), 2.59 (s, 3H).

[0137] Example 17: Synthesis of 2′-chloro-N-(5-(1-(4-cyanophenoxy)ethyl)-1,3,4-selenadiazol-2-yl)-5′-methoxy-6-methyl-[4,4′-bipyridine]-3-carboxamide (SEP-17)

[0138]

[0139] Referring to the synthesis method of Example 1, Compound 9, Compound 10 and Compound 15 were replaced by 2-chloro-5-methoxypyridine-4-boric acid, 4-bromo-6-methylnicotinate and 2-(4-cyanophenoxy)propionic acid, respectively, and other conditions remained unchanged. 1HNMR (300MHz, CD3OD) δ8.86 (s, 1H), 7.99 (s, 1H), 7.65 (s, 1H), 7.62 (s, 1H), 7.44 (s, 1H), 7.35 (s, 1H), 7.16 (s, 1H), 7.13 (s, 1H), 5.88-5.85 (m, 1H), 3.57 (s, 3H), 2.64 (s, 3H), 1.76 (d, J = 6.3 Hz, 3H).

[0140] Example 18: Synthesis of 2′-chloro-N-(5-(2-(4-cyanophenoxy)propan-2-yl)-1,3,4-selenadiazole-2-yl)-5′-methoxy-6-methyl-[4,4′-bipyridine]-3-carboxamide (SEP-18)

[0141]

[0142] Referring to the synthesis method of Example 1, Compound 9, Compound 10 and Compound 15 were replaced by 2-chloro-5-methoxypyridine-4-boric acid, 4-bromo-6-methylnicotinate and 2-(4-cyanophenoxy)-2-methylpropionic acid, respectively, and other conditions remained unchanged. 1 H NMR (300MHz, DMSO-d6) δ8.88(s, 1H), 8.12(s, 1H), 7.74-7.72(m, 1H), 7.71-7.68(m, 1H), 7.52(s , 1H), 7.41(s, 1H), 6.99-6.97(m, 1H), 6.96-6.94(m, 1H), 3.51(s, 3H), 2.58(s, 3H), 1.82(s, 6H).

[0143] Example 19: Synthesis of 2′-chloro-N-(5-(4-cyanobenzyl)-1,3,4-selenadiazole-2-yl)-5′-methoxy-6-methyl-[4,4′-bipyridine]-3-carboxamide (SEP-19)

[0144]

[0145] Referring to the synthesis method of Example 1, Compound 9, Compound 10 and Compound 15 were replaced by 2-chloro-5-methoxypyridine-4-boric acid, 4-bromo-6-methylnicotinate and 4-cyano-phenylacetic acid, respectively, and other conditions remained unchanged. 1H NMR (300MHz, DMSO-d6) δ13.35 (s, 1H), 8.85 (s, 1H), 8.13 (s, 1H), 7.82 (d, J = 1.7Hz, 1H), 7.80 (d, J = 1.9Hz, 1H), 7.56 (d, J=1.9Hz, 1H), 7.54 (d, J=1.8Hz, 1H), 7.51 (s, 1H), 7.40 (s, 1H), 4.45 (s, 2H), 3.54 (s, 3H), 2.57 (s, 3H).

[0146] Example 20: Synthesis of N-(5-((4-cyanophenoxy)methyl)-1,3,4-selenodiazol-2-yl)-4-(2-fluoro-6-methoxyphenyl)-6-methylnicotinamide (SEP-20)

[0147]

[0148] Referring to the synthesis method of Example 1, Compound 9, Compound 10 and Compound 15 were replaced by 2-fluoro-6-methoxyphenylboronic acid, 4-bromo-6-methylnicotinate and (4-cyanophenoxy)acetic acid, respectively, and other conditions remained unchanged. 1 H NMR (300MHz, DMSO-d6) δ13.43 (s, 1H), 8.86 (s, 1H), 7.84-7.78 (m, 2H), 7.43-7.32 (m , 2H), 7.27-7.21(m, 2H), 6.95-6.84(m, 2H), 5.56(s, 2H), 3.53(s, 3H), 2.57(s, 3H).

[0149] Example 21: Synthesis of N-(5-((4-chlorobenzyl)oxy)-1,3,4-thiadiazol-2-yl)-3-(2-methoxyphenyl)isonicotinamide (IDE95)

[0150]

[0151] The synthesis method of the positive control drug IDE95 refers to the synthesis method of compound 95 in patent WO2020243459A1. 1 H NMR (300MHz, DMSO-d6) δ12.89 (s, 1H), 8.71 (d, J=5.0Hz, 1H), 8.61 (s, 1H), 7.63 (d, J=5.0Hz, 1H), 7.50 (q, J=8. 5Hz, 4H), 7.38 (dd, J=8.0, 6.5Hz, 2H), 7.06 (t, J=7.4Hz, 1H), 6.98 (d, J=8.3Hz, 1H), 5.47 (s, 2H), 3.51 (s, 3H).

[0152] Example 22: Activity Test

[0153] (I) ADP-GloTM luminescence assay to determine Polθ inhibitory activity in vitro

[0154] Experimental Materials

[0155] ADP-Glo TM Kinase assay kit (Promega); ssDNA (Genscript); POLQ protein (Inhouse); neomycin (MCE); 96-well plate (Nunc); 384-well plate (PE); centrifuge (Cence); microplate reader (BMG).

[0156] Test Method

[0157] Add 50 μL of compound to a 384-well dilution plate, dilute the compound with DMSO at 1:3 per column (DMSO: compound = 1:3, v / v), and dilute 10 points in a row. Use the Echo 650 sonic pipetting system to transfer 0.1 μL of diluted compound solution per row to 384 assay plates, with 2 replicates for each compound. Add 5 μL of enzyme working solution (Aisip Technology Co., Ltd., S2201T-H02H) to the 384-well plate, centrifuge at 1000 rpm for 1 min, and incubate at 25°C for 10 minutes. Add 5 μL of working solution containing ssDNA (final concentration: 50 nM) and ATP (final concentration: 50 μM), centrifuge at 1000 rpm for 1 minute, and incubate at 25°C for 60 minutes. Add 5 μL of ADP-Glo TM Reagent, centrifuge at 1000 rpm for 1 minute, incubate at 25°C for 40 minutes. Add 10 μL ADP-Glo TM Kinase assay solution, centrifuge at 1000 rpm for 1 minute.

[0158] Experimental data processing: Enzyme inhibition activity (%) = 100*(ave High control-cpd well) / (ave High control-ave Low control). The IC50 values ​​of the compounds were fitted from the nonlinear regression equation using GraphPad Prism 8.4.3 software.

[0159] Experimental results:

[0160] The specific results are shown in Table 1. 50 <10nM(denoted as: A);IC 50 =10-100nM (denoted as: B); IC 50 =100-500nM (denoted as: C); IC 50>500nM (denoted as: D). Untested compounds are denoted as: -

[0161] Table 1 Inhibitory activity of compounds on Polθ

[0162]

[0163] The results in Table 1 show that the compounds of the present invention exhibit good inhibitory activity against Polθ.

[0164] (II) Antiproliferation experiments of compounds

[0165] 1. Experimental Materials

[0166] HCT116 cells, RPMI1640 incomplete medium (KEL biotech); MDA-MB-436 cells (Guangzhou Cellcook Biotech); L-15 medium (Guangzhou Cellcook Biotech); fetal bovine serum (Guangzhou Cellcook Biotech); ESCO CO2 incubator; microscope (Jiangnan Yongxin XD-202); centrifuge (DM0412); ELISA analyzer (Thermo); high pressure steam sterilizer (Zealway).

[0167] 2. Test Method

[0168] Take the cells in the logarithmic growth phase, pour out the old culture medium, wash twice with PBS, add 1mL 0.25% trypsin solution to digest for 1min, observe the cells become round under the microscope, add 4mL fresh culture medium containing 10% fetal bovine serum to stop digestion, transfer the solution to a centrifuge tube, centrifuge at 1000r / min for 5 minutes, and discard the supernatant. Add 1mL culture medium to resuspend the cells and count the cells. After counting, the cells are planted in a 96-well plate at a concentration of 600 cells per well, 100μL per well. The 96-well plate with cells is placed in a 37℃, 5% CO2 incubator and cultured for 12h. Use culture medium to dilute the drug gradient to 100μmol / L, 33.3μmol / L, 11.1μmol / L, 3.7μmol / L, 1.2μmol / L, 0.4μmol / L, and then add them to the 96-well plate, 90μL per well, and set three replicates for each concentration. The control group was added with the medium containing the corresponding concentration of solvent, and the zero-adjusted well was added with the same volume of blank medium, and incubated in a 5% CO2, 37°C incubator for 5 days. After adding 50 μL CTG to each well and incubating at room temperature for 10 minutes, the light signal value (Lum) was obtained using Envision microplate reader, and the inhibition rate and half inhibitory concentration (IC 50 ).

[0169] Inhibition rate (%) = 100*(Lum max -Lum compounds) / (Lum max -LUmmin)% where Lum max Lum refers to the light signal value of the well containing cells, and Lum compound refers to the light signal value of the well containing compounds and cells. min Refers to the light signal value containing culture medium.

[0170] The inhibitory effect of the disclosed compounds on the growth of tumor cells was determined by the above test, and the measured IC 50 See Table 2 for the values

[0171] Experimental results:

[0172] The specific results are shown in Table 2. 50 <1-20μM (denoted as: A); IC 50 >20μM (denoted as: B).

[0173] Table 2 Antiproliferative activity of compounds on different cells

[0174]

Claims

1. A Polθ inhibitor represented by the general formula (I), a tautomer or a pharmaceutically acceptable salt thereof: in: X is selected from C or N; Y is selected from CH or N; C is selected from -CH2-, O or no atom; R1 is selected from C1-C4 alkyl, H or absent; R2, R3 are selected from H or C1-C4 alkyl; A is selected from: or thienyl; Z is selected from C or N; R4 is selected from H, halogen, C1-C4 alkyl, CHO, CF3, NO2, CN; R5 is selected from H, halogen, C1-C4 alkyl, CHO, CF3, NO2, CN; R6 is selected from H, halogen; B is selected from substituted phenyl, -(CH2) 0～3 -C≡CH, -(CH2) 0～3 -CH=CH2, C3-C6 cycloalkyl; the substituent is selected from at least one halogen, C1-C4 alkoxy, CN.

2. The Polθ inhibitor, its tautomer or pharmaceutically acceptable salt according to claim 1, characterized in that: in: X is selected from C or N; Y is selected from CH or N; C is selected from -CH2-, O or no atom; R1 is selected from C1-C4 alkyl, H or absent; R2, R3 are selected from H or CH3; A is selected from: Z is selected from C or N; R4 is selected from H, F, Cl, Br, I, CH3, CHO, CF3, NO2, CN; R5 is selected from H, F, Cl, Br, I, CH3, CHO, CF3, NO2, CN; R6 is selected from H, F, Cl, Br, I; B is selected from substituted phenyl, ethynyl, vinyl, C3-C6 cycloalkyl; the substituent is selected from at least one of F, Cl, Br, I, OCH3, CN.

3. The Polθ inhibitor, its tautomer or pharmaceutically acceptable salt according to claim 1, characterized in that: When X is C and Y is N, R1 is CH3; when X is N and Y is CH, R1 is absent; C is selected from -CH2-, O or no atom; R2, R3 are selected from at least one H or CH3; A is selected from: Z is selected from C or N; R4 is selected from H, F, Cl, CH3, CHO; R5 is selected from H, F; R6 is selected from H, F; B is selected from substituted phenyl, alkynyl, alkenyl, C3-C6 cycloalkyl; the substituent is selected from at least one of F, Cl, OCH3, CN.

4. A Polθ inhibitor, a tautomer or a pharmaceutically acceptable salt thereof, characterized in that: Selected from any of the following compounds:

5. Use of the Polθ inhibitor, its tautomer or pharmaceutically acceptable salt according to claim 1 in the preparation of a Polθ inhibitor.

6. Use of the Polθ inhibitor, its tautomer or pharmaceutically acceptable salt according to claim 1 in the preparation of a drug for treating or preventing homologous recombination repair pathway-deficient cancer.

7. A pharmaceutical composition, characterized in that It comprises the Polθ inhibitor according to claim 1, its tautomer or pharmaceutically acceptable salt and pharmaceutically acceptable excipients.

8. Use of the Polθ inhibitor according to claim 1, its tautomer or pharmaceutically acceptable salt, or the pharmaceutical composition according to claim 6 in the preparation of drugs for treating or preventing colon cancer or breast cancer.

9. A method for preparing the Polθ inhibitor, its tautomer or pharmaceutically acceptable salt as claimed in claim 1, characterized in that: The steps include: (a) Compound 1 and 2 are subjected to SUZUKI reaction to obtain compound 3; (b) Compound 3 is hydrolyzed under alkaline conditions to obtain compound 4; (c) Compound 5 reacts with selenium powder and sodium borohydride to produce compound 6; (d) Compound 6 and compound 7 react with POCl3 to produce compound 8; (e) Compound 4 and Compound 8 react with an amide condensation agent TCFH to generate compound SEP, i.e., a compound represented by general formula (I); When X is selected from C or N; Y is selected from CH or N; C is selected from -CH2-, O or no atom; R1 is selected from C1-C4 alkyl, H or absent; R2, R3 are selected from H or C1-C4 alkyl; A is selected from: Z is selected from C or N; R4 is selected from H, halogen, C1-C4 alkyl, CHO, CF3, NO2, CN; R5 is selected from H, halogen, C1-C4 alkyl, CHO, CF3, NO2, CN; R6 is selected from H, halogen; B is selected from substituted phenyl, -(CH2) 0～3 -C≡CH, -(CH2) 0～3 -CH=CH2, C3-C6 cycloalkyl; the substituent is selected from at least one halogen, C1-C4 alkoxy, CN.

10. The method for preparing the Polθ inhibitor, its tautomer or pharmaceutically acceptable salt according to claim 9, characterized in that: The specific reaction reagents and conditions are: (a) Pd(dppf)Cl2, K2CO3, 1,4-Dioxane, H2O, 70-90℃; (b) NaOH, MeOH, 40-60℃; (c) CH3I, Se, N aBH4, EtOH, N2, -15-80°C; (d) POCl3, 1,4-Dioxane, 75-90°C; (e) TCFH, NMI, MeCN, N2, 40-70°C.

Images