Use of POLQ inhibitor in preparation of drug for treating or preventing tumor

Abstract

The present invention provides a use of a POLQ inhibitor in the preparation of a drug for treating or preventing a tumor. Specifically provided is a use of a POLQ inhibitor in the preparation of a drug for treating and / or preventing a tumor, the tumor including HR gene deficiency, TLS polymerase deficiency and / or POLQ overexpression. The POLQ inhibitor provided by the present invention exhibits good activity and has good inhibitory activity against tumors.

Description

Application of a POLQ inhibitor in the preparation of drugs for treating or preventing tumors

[0001] This application claims priority to Chinese patent application 2024105155981, filed on April 26, 2024. The entire contents of the aforementioned Chinese patent application are incorporated herein by reference. Technical Field

[0002] This invention relates to the use of a POLQ inhibitor in the preparation of drugs for treating or preventing tumors. Background Technology

[0003] DNA damage is a crucial mechanism by which many chemotherapeutic drugs exert their antitumor effects. Double-strand breaks (DSBs) are one of the most common types of cell damage, caused directly by ionizing radiation or induced by ultraviolet radiation, reactive oxygen species (ROS), or other mutagens, such as common chemotherapeutic drugs like cisplatin, 5-FU, and etoposide. Unrepaired DNA damage leads to the arrest of cellular functions such as transcription and replication, inducing apoptosis or necrosis, and is subsequently cleared by immune cells. Tumor cells possess highly efficient damage repair or alternative repair pathways. Targeting key proteins in these DNA damage repair pathways to allow unrepaired DNA damage to accumulate and ultimately lead to cell death is a key strategy in cancer treatment.

[0004] According to the "2022 National Cancer Report" released by the National Cancer Center in mid-2022, there were 4.064 million new cancer cases nationwide in 2016. Lung cancer, colorectal cancer, stomach cancer, liver cancer, and breast cancer were the top five cancers in terms of incidence in China. Among them, the incidence of colorectal cancer in Zhejiang Province was 26.56% higher than the national average! With the development of biomedical technology, the survival rate of cancer patients has improved significantly, but malignant tumors are still often incurable, seriously threatening people's lives and health.

[0005] This invention discloses a biomarker for the combined lethal treatment of cancer using POLQ small molecule inhibitors. This biomarker will have a better therapeutic effect than existing drugs on cancers such as lung cancer, breast cancer, ovarian cancer, gastric cancer, prostate cancer, colon cancer, and pancreatic cancer that have specific biomarkers, greatly prolonging the survival time of patients with these cancers and improving their quality of life.

[0006] DNA polymerase theta (Pol theta or Polθ) is a key protein in the microhomological end joining (Alt-NHEJ) pathway, also known as microhomological end joining (MMEJ) or TMEJ. It is a unique multifunctional polymerase composed of an N-terminal helicase domain, a central domain, and a C-terminal polymerase domain. Studies have shown that when homologous recombination (HR) and / or non-homologous end joining (NHEJ) are deficient, cells highly depend on MMEJ to repair broken DNA, and inhibition of MMEJ leads to apoptosis.

[0007] Studies have found that the Polθ polymerase domain is essential for DNA elongation at DSB damage repair sites, while the helicase domain and intermediate domain play crucial roles in the recognition and binding of Polθ to substrates. Polθ can deactivate the interaction between DNA and the damage repair complex (e.g., competitive binding of single-stranded DNA to RAD51), inhibiting the HR repair pathway. Furthermore, the Polθ helicase domain is also involved in DNA replication arrest; its loss of function leads to increased replication pressure in tumor cells, resulting in apoptosis.

[0008] Polθ is not expressed or is expressed at low levels in normal tissues and cells, but it is highly expressed in various tumors, including lung cancer, breast cancer, ovarian cancer, gastric cancer, prostate cancer, colon cancer, and pancreatic cancer, and is associated with poor prognosis. In particular, over 70% of breast cancers show Polθ overexpression. These phenomena suggest that Polθ may play an important role in these cancers and is a potential tumor-specific target.

[0009] Studies knocking down or knocking out Polθ in tumor cells have found that Polθ deficiency can sensitize these cells to radiation, induce DSB production, enhance replication fork instability, and sensitize tumor cells to mutagens, potentially enhancing the effects of radiotherapy and chemotherapy, making it a potential drug target. Research has also found a combined lethal effect between Polθ and HR deficiency; its small-molecule inhibitors can kill HR-deficient tumor cells in vitro and in vivo. Particularly in HR-deficient reversion mutation-resistant tumors resistant to PARP inhibitors (such as Olaparib), Polθ inhibitors can enable cells to overcome PARP resistance, providing a valuable therapeutic opportunity; however, further research on the applicable cancer diseases is lacking. Summary of the Invention

[0010] The technical problem this invention aims to solve is to overcome the deficiency that the specific types of tumors to which POLQ inhibitors are applicable are unclear. To this end, this invention provides an application of a POLQ inhibitor in the preparation of drugs for treating or preventing tumors. The POLQ inhibitor provided by this invention can effectively inhibit tumor cells with gene defects in certain HR pathways, and has good pharmaceutical prospects.

[0011] This invention provides the use of a POLQ inhibitor in the preparation of drugs for treating and / or preventing tumors, wherein the tumors include HR gene defects, TLS (translesion synthesis) polymerase defects, and / or POLQ overexpression.

[0012] Preferably, the tumor includes HR gene defects and / or TLS (translesion synthesis) polymerase defects.

[0013] Preferably, when the tumor includes an HR gene defect, it also includes a Shieldin complex defect. More preferably, the Shieldin complex defect includes one or more of the following target genes with decreased expression and / or abnormal protein function: RIF1, 53BP1, SHLD2, SHLD1, SHLD3, and REV7.

[0014] Preferably, the HR gene defect is a gene defect that can cause cells to produce HRD (homologous recombination deficiency), preferably including one or more of the following target genes with decreased expression and / or abnormal protein function: BRCA1, BRCA2, PALB2, RAD51, RAD51C, RAD51B, RAD51D, SLX4, FANCD2, FANCF, and XRCC3; more preferably, the HR gene defect includes one or more of the following target genes with decreased expression and / or abnormal protein function: BRCA1, BRCA2, RAD51C, and XRCC3.

[0015] Preferably, the TLS (translesion DNA synthesis) polymerase gene defect includes one or more of the following target genes with decreased expression and / or abnormal protein function: POLI, POLH, RAD18, REV1, REV3, POLK, POLL, PCNA, and REV7; more preferably, the TLS polymerase gene defect includes one or two of the following target genes with decreased expression and / or abnormal protein function: REV7 and REV3.

[0016] In one embodiment, the HR gene defect includes one or more of the following targets: decreased gene expression and / or abnormal protein function: BRCA2, BRCA1, XRCC3, RAD51B, RAD51C, and / or FANCF.

[0017] In one embodiment, the TLS polymerase deficiency includes one or more of the following targets: decreased gene expression and / or abnormal protein function: REV7, POLE2, and / or POLH.

[0018] In one embodiment, the Shieldin complex defect includes one or more of the following targets: decreased gene expression and / or abnormal protein function: 53BP1 and SHLD2.

[0019] In this invention, the decrease in expression includes the case of no expression.

[0020] In one embodiment, the tumor includes HR gene-deficient lung cancer, HR gene-deficient breast cancer, HR gene-deficient gastric cancer, HR gene-deficient ovarian cancer, HR gene-deficient prostate cancer, HR gene-deficient colon cancer, HR gene-deficient pancreatic cancer, HR gene-deficient uterine leiomyosarcoma, HR gene-deficient multiple myeloma, HR gene-deficient cervical cancer, HR gene-deficient urinary tract-related cancer, HR gene-deficient skin cancer, or HR gene-deficient kidney cancer. Preferably, the tumor further includes Shieldin complex deficiency (e.g., SHLD2 deficiency).

[0021] In one embodiment, the tumor includes TLS polymerase-deficient lung cancer, TLS polymerase-deficient breast cancer, TLS polymerase-deficient gastric cancer, TLS polymerase-deficient ovarian cancer, TLS polymerase-deficient prostate cancer, TLS polymerase-deficient colon cancer, TLS polymerase-deficient pancreatic cancer, TLS polymerase-deficient uterine leiomyosarcoma, TLS polymerase-deficient multiple myeloma, HR gene-deficient cervical cancer, HR gene-deficient urinary tract-associated cancer, HR gene-deficient skin cancer, or HR gene-deficient kidney cancer.

[0022] In one embodiment, the tumor is an ovarian cancer containing HR gene defects, including HR gene defective primary ovarian cancer and HR gene defective metastatic ovarian cancer. The HR gene defects may include BRCA2 defects, BRCA1 defects, XRCC3 defects, RAD51B defects and / or RAD51C defects. Preferably, the tumor further includes SHLD2 defects and / or 53BP1 defects.

[0023] In one embodiment, the HR gene-deficient tumor is an HR gene-deficient lung cancer, including HR gene-deficient primary lung cancer and HR gene-deficient metastatic lung cancer. The HR gene deficiency may include BRCA2 deficiency, BRCA1 deficiency, XRCC3 deficiency, RAD51B deficiency and / or RAD51C deficiency. Preferably, the tumor further includes SHLD2 deficiency, REV7 deficiency and / or 53BP1 deficiency.

[0024] In one embodiment, the HR gene-deficient tumor is an HR gene-deficient breast cancer, including HR gene-deficient primary breast cancer and HR gene-deficient metastatic breast cancer. The HR gene deficiency may include BRCA2 deficiency, BRCA1 deficiency, XRCC3 deficiency, RAD51B deficiency and / or RAD51C deficiency. Preferably, the tumor further includes SHLD2 deficiency, REV7 deficiency and / or 53BP1 deficiency.

[0025] In one embodiment, the HR gene-deficient tumor is an HR gene-deficient gastric cancer, including HR gene-deficient primary gastric cancer and HR gene-deficient metastatic gastric cancer. The HR gene deficiency may include BRCA2 deficiency, BRCA1 deficiency, XRCC3 deficiency, RAD51B deficiency and / or RAD51C deficiency. Preferably, the tumor further includes SHLD2 deficiency, REV7 deficiency and / or 53BP1 deficiency.

[0026] In one embodiment, the HR gene-deficient tumor is an HR gene-deficient prostate cancer, including HR gene-deficient primary prostate cancer and HR gene-deficient metastatic prostate cancer. The HR gene deficiency may include BRCA2 deficiency, BRCA1 deficiency, XRCC3 deficiency, RAD51B deficiency and / or RAD51C deficiency. Preferably, the tumor further includes SHLD2 deficiency, REV7 deficiency and / or 53BP1 deficiency.

[0027] In one embodiment, the HR gene-deficient tumor is an HR gene-deficient colorectal cancer, including HR gene-deficient primary colorectal cancer and HR gene-deficient metastatic colorectal cancer. The HR gene deficiency may include BRCA2 deficiency, BRCA1 deficiency, XRCC3 deficiency, RAD51B deficiency and / or RAD51C deficiency. Preferably, the tumor further includes SHLD2 deficiency, REV7 deficiency and / or 53BP1 deficiency.

[0028] In one embodiment, the HR gene-deficient tumor is an HR gene-deficient pancreatic cancer, including HR gene-deficient primary pancreatic cancer and HR gene-deficient metastatic pancreatic cancer. The HR gene deficiency may include BRCA2 deficiency, BRCA1 deficiency, XRCC3 deficiency, RAD51B deficiency and / or RAD51C deficiency. Preferably, the tumor further includes SHLD2 deficiency, REV7 deficiency and / or 53BP1 deficiency.

[0029] In one embodiment, the HR gene-deficient tumor is an HR gene-deficient uterine leiomyosarcoma, including HR gene-deficient primary uterine leiomyosarcoma and HR gene-deficient metastatic uterine leiomyosarcoma. The HR gene deficiency may further include BRCA2 deficiency, BRCA1 deficiency, XRCC3 deficiency, RAD51B deficiency and / or RAD51C deficiency. Preferably, the tumor further includes SHLD2 deficiency, REV7 deficiency and / or 53BP1 deficiency.

[0030] In one embodiment, the HR gene-deficient tumor is an HR gene-deficient multiple myeloma, including HR gene-deficient primary multiple myeloma and HR gene-deficient metastatic multiple myeloma. The HR gene deficiency may further include BRCA2 deficiency, BRCA1 deficiency, XRCC3 deficiency, RAD51B deficiency and / or RAD51C deficiency. Preferably, the tumor further includes SHLD2 deficiency, REV7 deficiency and / or 53BP1 deficiency.

[0031] In one embodiment, the TLS polymerase-deficient cancers include TLS polymerase-deficient primary cancers and TLS polymerase-deficient metastatic cancers, such as TLS polymerase-deficient primary lung cancer, TLS polymerase-deficient metastatic lung cancer, TLS polymerase-deficient primary breast cancer, TLS polymerase-deficient metastatic breast cancer, TLS polymerase-deficient primary gastric cancer, TLS polymerase-deficient metastatic gastric cancer, TLS polymerase-deficient primary ovarian cancer, TLS polymerase-deficient metastatic ovarian cancer, TLS polymerase-deficient metastatic prostate cancer, TLS polymerase-deficient primary prostate cancer, TLS polymerase-deficient primary colon cancer, TLS polymerase-deficient metastatic colon cancer, TLS polymerase-deficient primary pancreatic cancer, TLS polymerase-deficient metastatic pancreatic cancer, TLS polymerase-deficient primary uterine leiomyosarcoma, TLS polymerase-deficient metastatic uterine leiomyosarcoma, TLS polymerase-deficient multiple primary myeloma, or TLS polymerase-deficient multiple metastatic myeloma.

[0032] In one embodiment, the HR gene-deficient cancers include HR gene-deficient primary cancers and HR gene-deficient metastatic cancers, such as HR gene-deficient primary lung cancer, HR gene-deficient metastatic lung cancer, HR gene-deficient primary breast cancer, HR gene-deficient metastatic breast cancer, HR gene-deficient primary gastric cancer, HR gene-deficient metastatic gastric cancer, HR gene-deficient primary ovarian cancer, HR gene-deficient metastatic ovarian cancer, HR gene-deficient metastatic prostate cancer, HR gene-deficient primary prostate cancer, HR gene-deficient primary colon cancer, HR gene-deficient metastatic colon cancer, HR gene-deficient primary pancreatic cancer, HR gene-deficient metastatic pancreatic cancer, HR gene-deficient primary uterine leiomyosarcoma, HR gene-deficient metastatic uterine leiomyosarcoma, HR gene-deficient multiple primary myeloma, or HR gene-deficient multiple metastatic myeloma.

[0033] In one embodiment, the cancer (tumor) is BRCA2-deficient colon cancer, BRCA2-deficient primary colon cancer, BRCA2-deficient prostate cancer, RAD51B-deficient, POLE2-deficient, and POLH-deficient metastatic prostate cancer (preferably, the metastatic prostate cancer contains RAD51B, POLB, POLE2, POLH, MSH6, and RNASEH2B deficiencies), BRCA2-deficient ovarian cancer, BRCA2-deficient uterine leiomyosarcoma, BRCA-deficient breast cancer (e.g., BRCA1-deficient breast cancer), XRCC3-deficient colon cancer, RAD51C-deficient colon cancer, 53BP1-deficient breast cancer, BRCA-deficient and SHLD2-deficient breast cancer (e.g., BRCA1-deficient and SHLD2-deficient breast cancer or BRCA2-deficient and SHLD2-deficient breast cancer), 53BP1-deficient and SHLD2-deficient breast cancer, BRCA1-deficient and SHLD2-deficient breast cancer, FANCF-deficient ovarian cancer, POLQ-overexpressing gastric cancer, BRCA2-deficient and FANCD2-deficient multiple myeloma, or TLS polymerase-deficient colon cancer (e.g., REV7-deficient colon cancer).

[0034] In one embodiment, the POLQ inhibitor comprises a compound of Formula I or a pharmaceutically acceptable salt thereof;

[0035] in:

[0036] m can be 0, 1, 2, or 3;

[0037] R 1 For not replaced or by one or more R 1-1 Substituted 6-12 aryl groups or unsubstituted or substituted with one or more R groups 1-2The substituted 5-12-membered heteroaryl group; wherein the heteroatom in the 5-12-membered heteroaryl group is one or more of N, O and S, and the number of heteroatoms is 1, 2 or 3;

[0038] Each R 1-1 and R 1-2 Independently deuterium, halogen, cyano, amino, hydroxyl, unsubstituted, or modified by one or more R groups 1-1-1 Substituted C1-C6 alkyl groups or unsubstituted or substituted with one or more R 1-1-2 Substituted C1-C6 alkoxy groups;

[0039] Each R 1-1-1 and R 1-1-2 It can be independently deuterium, hydroxyl, cyano or halogen;

[0040] Each R 2 Independently for not replaced or by one or more R 2-1 Substituted C1-C6 alkyl, unsubstituted or substituted with one or more R 2-2 Substituted C1-C6 alkoxy groups;

[0041] Each R 2-1 and R 2-2 It can be independently deuterium, hydroxyl, cyano or halogen;

[0042] Ring B is unsubstituted or replaced by one or more R b-1 The substituted 5-12-membered fused-ring heteroaromatic ring; the heteroatoms in the 5-12-membered fused-ring heteroaromatic ring are selected from 1, 2 or 3 of N, O and S, and the number of heteroatoms is 1, 2, 3 or 4; the single heteroaromatic ring in the 5-12-membered fused-ring heteroaromatic ring is a 5-6-membered heteroaromatic ring; the number of rings in the 5-12-membered fused-ring heteroaromatic ring is 2; the heteroatoms in the 5-6-membered heteroaromatic ring are selected from 1, 2 or 3 of N, O and S, and the number of heteroatoms is 1, 2 or 3.

[0043] Each R b-1 Independently deuterium, halogen, cyano, hydroxyl, unsubstituted, or modified by one or more R groups b-1-1 Substituted C1-C6 alkyl, unsubstituted or substituted with one or more R b-1-2 Substituted C1-C6 alkoxy groups;

[0044] Each R b-1-1 and R b-1-2 Independently hydroxyl or halogen;

[0045] R 3 For not replaced or by one or more R 3-1 Substituted 3-12 membered cycloalkyl, unsubstituted or substituted with one or more R 3-2 Substituted 4-12 membered heterocyclic groups, unsubstituted or substituted with one or more R groups3-3 Substituted 4-12 membered cycloalkenyl groups, unsubstituted or with one or more R groups 3-4 Substituted 4-12 membered heterocyclic alkenyl groups, unsubstituted or with one or more R groups 3-5 Substituted 6-10 aryl groups, unsubstituted or substituted with one or more R groups 3-6 The substituted 5-10-membered heteroaryl group; the heteroatoms in the 4-12-membered heterocyclic group, 4-12-membered heterocyclic alkenyl group and 5-10-membered heteroaryl group are selected from 1, 2 or 3 of N, O and S, and the number of heteroatoms is 1, 2, 3 or 4;

[0046] Each R 3-1 R 3-2 R 3-3 and R 3-4 Independently deuterium, hydroxyl, halogen, cyano, amino, unsubstituted, or modified by one or more R groups 3-2-2 Substituted C1-C6 alkyl or C1-C6 alkoxy;

[0047] Each R 3-5 and R 3-6 Independently hydroxyl, halogen, cyano, amino, or unsubstituted or modified by one or more R groups 3-2-2 Substituted C1-C6 alkyl groups;

[0048] Each R 3-2-2 It can be independently deuterium, hydroxyl, or halogen.

[0049] In one particular scheme, R 1 In this context, the 6-12 aryl group is a 6-10 aryl group, and can also be phenyl or naphthyl, such as phenyl.

[0050] In one particular scheme, R 1 In this context, the 5-12-membered heteroaryl group is a 5-10-membered heteroaryl group, and the heteroatom in the 5-10-membered heteroaryl group can be N and / or O, and the number of heteroatoms can be 1 or 2; the 5-12-membered heteroaryl group is preferably a 5-6-membered heteroaryl group; more preferably... For example

[0051] In a certain scheme, each R 1-1 In this context, the halogen is fluorine, chlorine, bromine, or iodine, such as fluorine.

[0052] In a certain scheme, each R 1-1 In this context, the C1-C6 alkyl group is a C1-C4 alkyl group, and may also be methyl, ethyl, n-propyl, isopropyl, n-butyl, isobutyl, primary butyl, secondary butyl, or tert-butyl.

[0053] In a certain scheme, each R 1-1In this context, the C1-C6 alkoxy group is a C1-C4 alkoxy group, and may also be a methoxy group, an ethoxy group, a propoxy group, an isopropoxy group, a butoxy group, an isobutoxy group, a primary butoxy group, a secondary butoxy group, or a tert-butoxy group, such as a methoxy group.

[0054] In a certain scheme, each R 1-2 In this context, the halogen is fluorine, chlorine, bromine, or iodine, such as fluorine or chlorine.

[0055] In a certain scheme, each R 1-2 In this context, the C1-C6 alkyl group is a C1-C4 alkyl group, and may be methyl, ethyl, n-propyl, isopropyl, n-butyl, isobutyl, primary butyl, secondary butyl, or tert-butyl, with methyl being preferred.

[0056] In a certain scheme, each R 1-2 In this context, the C1-C6 alkoxy group is a C1-C4 alkoxy group, and may also be a methoxy group, an ethoxy group, a propoxy group, an isopropoxy group, a butoxy group, an isobutoxy group, a primary butoxy group, a secondary butoxy group, or a tert-butoxy group, such as a methoxy group.

[0057] In a certain scheme, each R 1-1-1 and R 1-1-2 In this context, the halogen is fluorine, chlorine, bromine, or iodine, such as fluorine.

[0058] In a certain scheme, each R 2 In this context, the C1-C6 alkyl group is a C1-C4 alkyl group, and may also be methyl, ethyl, n-propyl, isopropyl, n-butyl, isobutyl, primary butyl, secondary butyl, or tert-butyl, for example, methyl.

[0059] In a certain scheme, each R 2 In this context, the C1-C6 alkoxy group is a C1-C4 alkoxy group, and may also be a methoxy group, an ethoxy group, a propoxy group, an isopropoxy group, a butoxy group, an isobutoxy group, a primary butoxy group, a secondary butoxy group, or a tert-butoxy group.

[0060] In a certain scheme, each R 2-1 and R 2-2 In this context, the halogen is fluorine, chlorine, bromine, or iodine.

[0061] In one embodiment, the number of heteroatoms in the 5-12 membered fused-ring heteroaromatic ring B can be 1, 2, 3, or 4; the 5-20 membered fused-ring heteroaromatic ring is preferably a five- or six-membered fused-ring heteroaromatic ring, more preferably...

[0062] In one embodiment, in ring B, the 5-6 membered heteroaromatic ring is a 5-membered heteroaromatic ring, which can have 2 heteroatoms, and the heteroatoms can be one or two of N, S, or O. Preferably, the 5-6 membered heteroaromatic ring is... In a certain scheme, each R b-1 In this context, the halogen is fluorine, chlorine, bromine, or iodine, such as fluorine.

[0063] In a certain scheme, each R b-1 In this context, the C1-C6 alkyl group is a C1-C4 alkyl group, and may also be methyl, ethyl, n-propyl, isopropyl, n-butyl, isobutyl, primary butyl, secondary butyl, or tert-butyl.

[0064] In a certain scheme, each R b-1 In this context, the C1-C6 alkoxy group is a C1-C4 alkoxy group, and may also be a methoxy group, an ethoxy group, a propoxy group, an isopropoxy group, a butoxy group, an isobutoxy group, a primary butoxy group, a secondary butoxy group, or a tert-butoxy group.

[0065] In a certain scheme, each R b-1-1 and R b-1-2 In this context, the halogen is fluorine, chlorine, bromine, or iodine.

[0066] In one embodiment, in R3, the 3-12 membered cycloalkyl group is a monocyclic cycloalkyl group, such as a 3-6 membered cycloalkyl group, or cyclopropyl, cyclobutyl, cyclopentyl, or cyclohexyl.

[0067] In one embodiment, in R3, the ring in the 4-12 membered heterocyclic group can be a monocyclic heterocyclic group, and the number of heteroatoms in the 4-12 membered heterocyclic group can be 1 or 2, and the type of heteroatom can be N; preferably...

[0068] In one embodiment, in R3, the 4-12 membered cycloalkenyl group is a 4-6 membered cycloalkenyl group (e.g., a monocyclic cycloalkenyl group), and the number of carbon-carbon double bonds can be one, preferably.

[0069] In one embodiment, in R3, the 4-12 membered heterocyclic alkenyl group is a 4-6 membered heterocyclic alkenyl group (e.g., a monocyclic heterocyclic alkenyl group), and the number of heteroatoms can be 1, and the type of heteroatoms can be 1, 2, or 3 of N, O, or S, for example...

[0070] In one embodiment, in R3, the 6-10 aryl group can be aryl or naphthyl, preferably aryl.

[0071] In one embodiment, in R3, the 5-10-membered heteroaryl group is a 5-6-membered heteroaryl group, the heteroatom can be N, and the number of heteroatoms can be 1, 2, or 3, for example...

[0072] In a certain scheme, each R 3-1 R 3-2 R 3-3 and R3-4 In this context, the halogen is fluorine, chlorine, bromine, or iodine.

[0073] In a certain scheme, each R 3-1 R 3-2 R 3-3 and R 3-4 In this context, the C1-C6 alkyl group is a C1-C4 alkyl group, and may also be methyl, ethyl, n-propyl, isopropyl, n-butyl, isobutyl, primary butyl, secondary butyl, or tert-butyl.

[0074] In a certain scheme, each R 3-1 R 3-2 R 3-3 and R 3-4 In this context, the C1-C6 alkoxy group is a C1-C4 alkoxy group, and may also be a methoxy group, an ethoxy group, a propoxy group, an isopropoxy group, a butoxy group, an isobutoxy group, a primary butoxy group, a secondary butoxy group, or a tert-butoxy group.

[0075] In a certain scheme, each R 3-5 and R 3-6 In this context, the halogen is fluorine, chlorine, bromine, or iodine.

[0076] In a certain scheme, each R 3-5 and R 3-6 In this context, the C1-C6 alkyl group is a C1-C4 alkyl group, and may also be methyl, ethyl, n-propyl, isopropyl, n-butyl, isobutyl, primary butyl, secondary butyl, or tert-butyl.

[0077] In a certain scheme, each R 3-2-2 In this context, the halogen is fluorine, chlorine, bromine, or iodine.

[0078] In one particular scheme, m is 1.

[0079] In one particular scheme, R 1 For one or more R 1-2 The substituted 5-12-membered heteroaryl group; wherein the heteroatom in the 5-12-membered heteroaryl group is one or more of N, O and S, and the number of heteroatoms is 1, 2 or 3.

[0080] In a certain scheme, each R 1-2 Independently halogenated, by one or more R 1-1-1 Substituted C1-C6 alkyl or unsubstituted C1-C6 alkoxy.

[0081] In a certain scheme, each R 1-1-1 Halogens are independent of each other.

[0082] In a certain scheme, each R 2 Independently, it is an unsubstituted C1-C6 alkyl group.

[0083] In one scheme, ring B is either unsubstituted or replaced by one or more Rs. b-1 The substituted 5-12-membered fused-ring heteroaromatic ring; the heteroatoms in the 5-12-membered fused-ring heteroaromatic ring are selected from 1, 2 or 3 of N, O and S, and the number of heteroatoms is 1, 2, 3 or 4; the single heteroaromatic ring in the 5-12-membered fused-ring heteroaromatic ring is a 5-6-membered heteroaromatic ring; the number of rings in the 5-12-membered fused-ring heteroaromatic ring is 2; the heteroatoms in the 5-6-membered heteroaromatic ring are selected from 1, 2 or 3 of N, O and S, and the number of heteroatoms is 1, 2 or 3.

[0084] In a certain scheme, each R b-1 Halogens are independent of each other.

[0085] In one particular scheme, R 3 For one or more R 3-2 Substituted 4-12 membered heterocyclic groups, unsubstituted or substituted with one or more R groups 3-3 Substituted 4-12 membered cycloalkenyl groups, unsubstituted or with one or more R groups 3-6 The substituted 5-10-membered heteroaryl group; the heteroatoms in the 4-12-membered heterocyclic group and the 5-10-membered heteroaryl group are selected from one, two, or three of N, O, and S, and the number of heteroatoms is 1, 2, 3, or 4. Preferably, R 3 For not replaced or by one or more R 3-6 The substituted 5-10-membered heteroaryl group; the heteroatoms in the 4-12-membered heterocyclic group and the 5-10-membered heteroaryl group are selected from 1, 2 or 3 of N, O and S, and the number of heteroatoms is 1, 2, 3 or 4.

[0086] In a certain scheme, each R 3-2 and R 3-3 Independently hydroxyl or by one or more R 3-2-2 Substituted C1-C6 alkyl groups.

[0087] In a certain scheme, each R 3-6 It is independently a C1-C6 alkyl group.

[0088] In a certain scheme, each R 3-2-2 Independently, it is a hydroxyl group.

[0089] In one embodiment, the compound represented by Formula I,

[0090] m is 1;

[0091] R 1 For one or more R 1-2 The substituted 5-12-membered heteroaryl group; wherein the heteroatom in the 5-12-membered heteroaryl group is one or more of N, O and S, and the number of heteroatoms is 1, 2 or 3;

[0092] Each R 1-2 Independently halogenated, by one or more R 1-1-1 Substituted C1-C6 alkyl or unsubstituted C1-C6 alkoxy;

[0093] Each R 1-1-1 Halogens are independent of each other;

[0094] Each R 2 Independently, it is an unsubstituted C1-C6 alkyl group;

[0095] Ring B is unsubstituted or replaced by one or more R b-1 The substituted 5-12-membered fused-ring heteroaromatic ring; the heteroatoms in the 5-12-membered fused-ring heteroaromatic ring are selected from 1, 2 or 3 of N, O and S, and the number of heteroatoms is 1, 2, 3 or 4; the single heteroaromatic ring in the 5-12-membered fused-ring heteroaromatic ring is a 5-6-membered heteroaromatic ring; the number of rings in the 5-12-membered fused-ring heteroaromatic ring is 2; the heteroatoms in the 5-6-membered heteroaromatic ring are selected from 1, 2 or 3 of N, O and S, and the number of heteroatoms is 1, 2 or 3.

[0096] Each R b-1 Halogens are independent of each other;

[0097] R 3 For one or more R 3-2 Substituted 4-12 membered heterocyclic groups, unsubstituted or substituted with one or more R groups 3-3 Substituted 4-12 membered cycloalkenyl groups, unsubstituted or with one or more R groups 3-6 The substituted 5-10-membered heteroaryl group; the heteroatoms in the 4-12-membered heterocyclic group and the 5-10-membered heteroaryl group are selected from 1, 2 or 3 of N, O and S, and the number of heteroatoms is 1, 2, 3 or 4;

[0098] Each R 3-2 and R 3-3 Independently hydroxyl or by one or more R 3-2-2 Substituted C1-C6 alkyl groups;

[0099] Each R 3-6 Independently, it is a C1-C6 alkyl group;

[0100] Each R 3-2-2 Independently, it is a hydroxyl group.

[0101] In one embodiment, the compound represented by Formula I,

[0102] m is 1;

[0103] R 1 For one or more R 1-2The substituted 5-12-membered heteroaryl group; wherein the heteroatom in the 5-12-membered heteroaryl group is one or more of N, O and S, and the number of heteroatoms is 1, 2 or 3;

[0104] Each R 1-2 Independently halogenated, by one or more R 1-1-1 Substituted C1-C6 alkyl or unsubstituted C1-C6 alkoxy;

[0105] Each R 1-1-1 Halogens are independent of each other;

[0106] Each R 2 Independently, it is an unsubstituted C1-C6 alkyl group;

[0107] Ring B is unsubstituted or replaced by one or more R b-1 The substituted 5-12-membered fused-ring heteroaromatic ring; the heteroatoms in the 5-12-membered fused-ring heteroaromatic ring are selected from 1, 2 or 3 of N, O and S, and the number of heteroatoms is 1, 2, 3 or 4; the single heteroaromatic ring in the 5-12-membered fused-ring heteroaromatic ring is a 5-6-membered heteroaromatic ring; the number of rings in the 5-12-membered fused-ring heteroaromatic ring is 2; the heteroatoms in the 5-6-membered heteroaromatic ring are selected from 1, 2 or 3 of N, O and S, and the number of heteroatoms is 1, 2 or 3.

[0108] Each R b-1 Halogens are independent of each other;

[0109] R 3 For not replaced or by one or more R 3-6 The substituted 5-10-membered heteroaryl group; the heteroatoms in the 4-12-membered heterocyclic group and the 5-10-membered heteroaryl group are selected from 1, 2 or 3 of N, O and S, and the number of heteroatoms is 1, 2, 3 or 4;

[0110] Each R 3-6 Independently, it is a C1-C6 alkyl group;

[0111] Each R 3-2-2 Independently, it is a hydroxyl group.

[0112] In one embodiment, the compound represented by Formula I is the compound represented by Formula I-1;

[0113] Wherein, Z1 and Z2 are each independently CH, CF or N (preferably, Z1 and Z2 are not both N);

[0114] m、R 1 R 2 R b-1 and R 3 Each independently as described in any one of the present invention (e.g., R) b-1(Halogen, preferably F).

[0115] In one of the schemes, R 2 It is a methyl group.

[0116] In one of the schemes, R 1 for

[0117] In one scheme, the ring for

[0118] In one of the schemes, R 3 for

[0119] In one embodiment, the compound represented by Formula I is any of the following compounds:

[0120] In one embodiment, the drug comprises the POLQ inhibitor and pharmaceutical excipients.

[0121] The present invention provides a pharmaceutical composition, wherein the pharmaceutical composition is the above-mentioned drug.

[0122] In one embodiment, the drug is a solid composition, preferably a tablet.

[0123] In one embodiment, the content of the POLQ inhibitor is a conventional content in the art, for example, the mass ratio of the POLQ inhibitor to the drug is 0.01%-95%, such as 1%, 10%, 30%, 40%, 50%, 60%, 70%, 80% or 90%.

[0124] In one embodiment, the drug can be formulated into various suitable dosage forms depending on the route of administration, including gastrointestinal dosage forms (e.g., oral dosage forms) and non-gastrointestinal dosage forms (e.g., injection dosage forms, respiratory dosage forms, mucosal dosage forms, or cavity dosage forms), such as capsules, tablets, pills, granules, powders, or oral liquids, with tablets being preferred.

[0125] In one embodiment, the drug is administered in a manner that is conventional in the art for such drugs, such as by injection (e.g., intravenous injection, subcutaneous injection, peritoneal injection or intramuscular injection) or orally, preferably orally.

[0126] This invention provides an application of a POLQ inhibitor in the in vitro inhibition of tumor cells, wherein the tumor is as described in any embodiment of this invention;

[0127] The POLQ inhibitor is as described in any embodiment of the present invention.

[0128] In the application of the POLQ inhibitor to inhibit tumor cells in vitro, the tumors mentioned are those containing HR gene defects or TLS polymerase defects as described in any one of the present invention.

[0129] In the application of the POLQ inhibitor to inhibit tumor cells in vitro, the dosage of the POLQ inhibitor is the conventional dosage in the art, which can be adjusted as needed by those skilled in the art.

[0130] This invention provides a method for treating and / or preventing tumors, comprising administering a therapeutically effective amount of a POLQ inhibitor to a patient in need;

[0131] The POLQ inhibitor and tumor are as described in any embodiment of the present invention.

[0132] Preferably, the tumor is a tumor containing the HR gene defect or a tumor containing the TLS polymerase defect as described in any one of the present invention.

[0133] In the methods for treating and / or preventing tumors, the POLQ inhibitor may be administered at different doses depending on the condition (symptoms, number of treatments, intervals, etc.) or based on the patient's weight.

[0134] In the methods for treating and / or preventing tumors, the frequency of the POLQ inhibitor is conventional in the art (e.g., once / day, twice / day, three times / day, or once / 7 days), which can be adjusted by those skilled in the art as needed to provide optimal therapeutic effects.

[0135] In the methods for treating and / or preventing tumors, the POLQ inhibitor is administered in a manner conventional in the art, and may be administered by injection (e.g., intravenous injection, subcutaneous injection, peritoneal injection or intramuscular injection) or orally, preferably orally.

[0136] Terminology Explanation:

[0137] Unless otherwise stated, the following terms appearing in this invention have the following meanings:

[0138] The term "unsubstituted or substituted group B" means that one or more hydrogen atoms in group B are independently substituted by group A, or that B is not substituted. When multiple groups A are present, unless otherwise specified, their definitions are independent and do not affect each other. For example, "C6-C10 aryl group substituted with three halogens" means that the C6-C10 aryl group is substituted with three halogens. The definitions of the three halogens are independent and do not affect each other, including but not limited to: wait.

[0139] The term "multiple" refers to two or more, such as two, three, four, or five.

[0140] The term "pharmaceutical acceptable" means that something is relatively non-toxic, safe, and suitable for patient use.

[0141] The term "halogen" refers to fluorine, chlorine, bromine, or iodine. Halogen substitution in this invention includes, but is not limited to, substitution by one halogen, substitution by two halogens, substitution by three halogens, and typically multiple substitutions occur on a single carbon atom.

[0142] The term "aryl" refers to a cyclic, unsaturated monovalent hydrocarbon group having a specified number of carbon atoms (e.g., 6–12 or 6–10), which can be monocyclic or polycyclic (e.g., 2 or 3 rings). In polycyclic cases, the monocyclic rings share two atoms and one bond, and (at least one ring / each ring) is aromatic. The aryl group is attached to the rest of the molecule via an aromatic or non-aromatic ring. Aryl groups include, but are not limited to, phenyl, naphthyl, and others. wait.

[0143] The term "cycloalkyl" refers to a cyclic, saturated monovalent hydrocarbon group having a specified number of carbon atoms (e.g., 3 to 10 members), which is a monocyclic group. Monocyclic groups include, but are not limited to: wait.

[0144] The term "cycloalkenyl" refers to a cyclic, unsaturated monovalent hydrocarbon group having a specified number of carbon atoms (e.g., 4–12, 4–10), which can be monocyclic or polycyclic (e.g., 2 or 3, bridged, spirocyclic, fused). Monocyclic is preferred, and monocyclic cycloalkenyl groups include, but are not limited to: wait.

[0145] The term "heterocyclic group" refers to a monocyclic, bridged, spirocyclic, or fused ring (preferably monocyclic) having a specified number of ring atoms (e.g., 4–12, 4–10), a specified number of heteroatoms (e.g., 1, 2, or 3), and a specified type of heteroatom (1, 2, or 3 of N, O, and S). Monocyclic heterocyclic groups are connected to the rest of the molecule via carbon atoms or heteroatoms. Monocyclic heterocyclic groups include, but are not limited to: Etc. Spirocyclic heterocyclic groups include, but are not limited to: Etc. Bridged heterocyclic groups include, but are not limited to: etc. Cycloherocyclic groups include, but are not limited to, those containing... wait.

[0146] The term "heterocyclic alkenyl" refers to a cyclic, unsaturated monovalent group having a specified number of ring atoms (e.g., 4 to 12), a specified number of heteroatoms (e.g., 1, 2, or 3), and a specified type of heteroatom (1, 2, or 3 of N, O, and S), which can be monocyclic or polycyclic (e.g., 2 or 3, spirocyclic, preferably monocyclic). Monocyclic heterocyclic alkenyl groups include, but are not limited to: Etc. Spirocyclic heterocyclic alkenyl groups include, but are not limited to, those containing... wait.

[0147] The term "heteroaryl" refers to a cyclic, unsaturated monovalent group having a specified number of ring atoms (e.g., 5–12, 5–10, 5–6), a specified number of heteroatoms (e.g., 1, 2, or 3), and a specified type of heteroatom (1, 2, or 3 of N, O, and S). It is monocyclic or polycyclic (preferably monocyclic), with the monocyclic rings sharing two atoms and one bond, and at least one ring is aromatic. The heteroaryl group is attached to the rest of the molecule via a carbon atom or a heteroatom; the heteroaryl group is attached to the rest of the molecule via a ring with or without heteroatoms; the heteroaryl group is attached to the rest of the molecule via an aromatic ring or a non-aromatic ring. Heteroaryl groups include, but are not limited to: wait.

[0148] The term "fused heteroaryl ring" refers to a heteroaryl ring that forms a fused ring with other rings. A fused heteroaryl ring contains at least two rings, at least one of which is a heteroaryl ring. The rest of the definition of "heteroaryl ring" is the same as that of "heteroaryl group".

[0149] The term "alkyl" refers to a saturated monovalent hydrocarbon group that has a specified number of carbon atoms, is straight-chain or branched. For example, C1-C6 alkyl groups (C 1-6 Alkyl), preferably C1-C4 alkyl (C 1-4 Alkyl groups include, but are not limited to: methyl, ethyl, n-propyl, isopropyl, n-butyl, isobutyl, sec-butyl, or tert-butyl.

[0150] The term "alkoxy" refers to an oxygen atom attached to one end of an alkyl group as a connecting bond, forming an "alkyl-O-". The definition of "alkyl" is as described above.

[0151] The term "pharmaceutically acceptable salt" refers to a salt obtained by reacting a compound with a pharmaceutically acceptable acid or base. When a compound contains a relatively acidic functional group, a base addition salt can be obtained by contacting the compound with a sufficient amount of a pharmaceutically acceptable base in a suitable inert solvent. When a compound contains a relatively basic functional group, an acid addition salt can be obtained by contacting the compound with a sufficient amount of a pharmaceutically acceptable acid in a suitable inert solvent. See Handbook of Pharmaceutical Salts: Properties, Selection, and Use (P. Heinrich Stahl, Camille G. Wermuth, 2011, 2nd Revised Edition) for details.

[0152] As used in this article, “treatment” means therapeutic therapy. When a specific condition is involved, treatment means: (1) alleviating one or more biological manifestations of the disease or condition; (2) interfering with (a) one or more points in a biological cascade that causes or precipitates the condition or (b) one or more biological manifestations of the condition; (3) improving one or more symptoms, effects or side effects associated with the condition, or one or more symptoms, effects or side effects associated with the condition or its treatment; or (4) slowing the development of the condition or one or more biological manifestations of the condition.

[0153] The term "prevention" as used in this article refers to reducing the risk of developing a disease.

[0154] As used herein, the term "therapeutic effective amount" refers to an amount of pharmaceutical composition sufficient to effectively treat the disease or condition described herein when administered to a patient. The amount of pharmaceutical composition constituting a "therapeutic effective amount" will vary depending on the pharmaceutical composition, the condition and its severity, and the age of the patient to be treated, but may be adjusted as needed by those skilled in the art.

[0155] As used herein, the term "patient" refers to any animal, preferably a mammal, that is about to be or has already been administered the compound or composition according to embodiments of the invention. As used herein, the term "mammal" includes any mammal. Examples of mammals include, but are not limited to, cattle, horses, sheep, pigs, cats, dogs, mice, rats, rabbits, guinea pigs, monkeys, and humans, with humans being the most preferred.

[0156] The term "pharmaceutical excipients" as used in this article refers to excipients and additives used in the production of pharmaceuticals and the dispensing of prescriptions. It includes all substances contained in pharmaceutical preparations, excluding the active ingredient. See the Pharmacopoeia of the People's Republic of China (2020 Edition), Volume IV, or the Handbook of Pharmaceutical Excipients (Raymond C. Rowe, 2009 Sixth Edition).

[0157] Without violating common sense in the field, the above-mentioned preferred conditions can be combined arbitrarily to obtain various preferred embodiments of the present invention.

[0158] The reagents and raw materials used in this invention are all commercially available.

[0159] The positive and progressive effects of this invention are as follows: the POLQ inhibitor provided by this invention can effectively inhibit tumor cells with HR gene defects and / or TLS polymerase defects, exhibiting good activity and promising pharmaceutical prospects. The HR gene defects include, for example, BRCA2 deficiency, BRCA1 deficiency, XRCC3 deficiency, RAD51B deficiency, RAD51C deficiency, 53BP1 deficiency, FANCF deficiency, and / or REV7 deficiency. The TLS polymerase defects include, for example, REV7, POLE2, and / or POLH deficiency. Attached Figure Description

[0160] Figure 1 shows the inhibition rate curve of compound 1 against DLD1-BRCA2- / -.

[0161] Figure 2 shows the inhibition rate curve of compound 2 on DLD1-BRCA2- / -.

[0162] Figure 3 shows the inhibition rate curve of compound 3 on DLD1-BRCA2- / -.

[0163] Figure 4 shows the inhibition rate curve of compound 4 on DLD1-BRCA2- / -.

[0164] Figure 5 shows the inhibition rate curve of compound 5 against HCT116.

[0165] Figure 6 shows the inhibition rate curve of compound 5 against HCT116-BRCA2- / -.

[0166] Figure 7 shows the inhibition rate curves of compound 6 on PC3,PC3-BRCA2- / -.

[0167] Figure 8 shows the inhibition rate curve of compound 7 against LNCAP-Clone FGC.

[0168] Figure 9 shows the inhibition rate curve of compound 8 against PEO1.

[0169] Figure 10 shows the inhibition rate curves of compound 9 on SKUT1 and SKUT1-BRCA2.

[0170] Figure 11 shows the inhibition rate curve of compound 7 against MDA-MB-436.

[0171] Figure 12 shows the inhibition rate curve of compound 10 against DLD1-XRCC3.

[0172] Figure 13 shows the inhibition rate curve of compound 10 against DLD1-RAD51C.

[0173] Figure 14 shows the inhibition rate curve of compound 7 against MDA-MB-436-53BP1.

[0174] Figure 15 shows the inhibition rate curve of compound 4 on MDA-MB-436-SHLD2.

[0175] Figure 16 shows the inhibition rate curve of compound 8 against PEO1-53BP1.

[0176] Figure 17 shows the inhibitory effect of compound 4 on HCC1937.

[0177] Figure 18 shows the inhibition rate curve of compound 6 against TOV21G.

[0178] Figure 19 shows the inhibition rate curve of compound 7 against SNU-601.

[0179] Figure 20 shows the inhibition rate curve of compound 7 against KMM1.

[0180] Figure 21 shows the inhibitory effect of compound 1 on DLD1-REV7. Detailed Implementation

[0181] The present invention is further illustrated below by way of embodiments, but the invention is not limited to the scope of the embodiments described herein. Experimental methods in the following embodiments that do not specify specific conditions were performed according to conventional methods and conditions, or as selected according to the product instructions.

[0182] The cells used in the embodiments of this patent are all derived from commercially available cell lines or internally constructed knockout cell lines. Specifically, DLD1, HCT116, PC-3, MDA-MB-436, and HCC1937 are sourced from ATCC, with catalog numbers CCL-221, CCL-247, CRL-1435, HTB130, and CRL-2336, respectively. The culture methods followed the methods provided by the suppliers. The knockout cell lines on DLD1, HCT116, PC-3, and MDA-MB-436 were cultured using the same methods as the parent cell lines. LNCAPFGC cells are sourced from ATCC, catalog number CRL-1740; PEO1 cells are sourced from ECACC, catalog number 10032308; SKUT1 cells are sourced from ATCC, catalog number HTB114. All of these cells were cultured according to the methods provided by the suppliers. SNU601 and KMM1 cells were obtained from the WuXi AppTec Cell Detection Platform, and testing was performed by WuXi AppTec.

[0183] The DLD1-BRCA2, DLD1-RAD51C, DLD1-XRCC3, DLD1-REV7, PC3-BRCA2, MDA-MB-436-53BP1, and PEO1-53BP1 KO cell lines are cell lines constructed using the CRISPR KO method. These are monoclonal cell lines obtained by knockout using the single plasmid system (PX459V2). Experiments were conducted using the reference "Generation of Genomic Deletions in Mammalian Cell Lines via CRISPR / Cas9" (http: / / www.jove.com / video / 52118). The obtained monoclonal cell lines were identified as successfully knocked out using PCR, Western blot, or functional experiments.

[0184] The compounds shown in the table below can be prepared by the methods described below, or by the methods disclosed in patent application 2023114104096.

[0185] Preparation Examples

[0186] Intermediate 1: 2'-chloro-3'-fluoro-5'-methoxy-6-methyl-[4,4'-bipyridine]-3-carboxylic acid

[0187] Step 1: 2-Chloro-3-fluoro-5-methoxypyridine

[0188] 6-Chloro-5-fluoropyridin-3-ol (10 g, 67.8 mmol) was dissolved in acetonitrile (100 mL), potassium carbonate (32.7 g, 237 mmol) was added, followed by iodomethane (33.7 g, 237 mmol). The mixture was reacted at room temperature for 18 hours. After the reaction was complete, water was added, and the mixture was extracted with ethyl acetate, washed with saturated brine, dried over anhydrous sodium sulfate, and concentrated under reduced pressure. The residue was purified by column chromatography (eluent: petroleum ether / ethyl acetate, gradient: 10%-40% ethyl acetate) to give the title compound (colorless liquid, 7.1 g, yield 64.8%). LC / MS (ESI) m / z: 161.8 [M+H]+.

[0189] Step 2: 2-Chloro-3-fluoro-4-iodo-5-methoxypyridine

[0190] Under nitrogen protection, 11 g (68.1 mmol) of 2-chloro-3-fluoro-5-methoxypyridine was dissolved in 100 mL of tetrahydrofuran. The mixture was cooled to -60 °C, and a solution of n-butyllithium in n-hexane (2.5 mol / L, 40.8 mL, 102.1 mmol) was slowly added while maintaining the temperature below -60 °C. After reacting at -60 °C for half an hour, a solution of 19.0 g (74.9 mmol) of iodine in tetrahydrofuran (20 mL) was added, and the mixture was reacted at -60 °C for 1 hour. After the reaction was complete, saturated ammonium chloride was slowly added to quench the reaction. Water was added, and the mixture was extracted with ethyl acetate. The organic phase was washed with saturated brine, dried over anhydrous sodium sulfate, filtered, and the filtrate was concentrated under reduced pressure. The residue was purified by column chromatography (eluent: petroleum ether / ethyl acetate, gradient: 10%-40% ethyl acetate) to give the title compound (white solid, 12.5 g, yield 6.4%). LC / MS (ESI) m / z: 288.1 [M+H]+.

[0191] Step 3: Methyl 2'-chloro-3'-fluoro-5'-methoxy-6-methyl-[4,4'-bipyridine]-3-carboxylic acid

[0192] Under nitrogen protection, 2-chloro-3-fluoro-4-iodo-5-methoxypyridine (288 mg, 1.0 mmol), methyl 6-methyl-4-(4,4,5,5-tetramethyl-1,3,2-dioxoboro-2-yl)nicotinate (277 mg, 1.0 mmol), [1,1'-bis(diphenylphosphine)ferrocene]palladium dichloride dichloromethane complex (164 mg, 0.20 mmol), and potassium carbonate (415 mg, 3.0 mmol) were dissolved in a mixed solvent of 1,4-dioxane (10 mL) and water (2 mL). The mixture was stirred at 65 °C for 2 hours under nitrogen protection. Water was added, and the mixture was extracted with ethyl acetate. The organic phase was washed with saturated brine, dried over anhydrous sodium sulfate, filtered, and the filtrate was concentrated under reduced pressure. The residue was purified by column chromatography (eluent: petroleum ether / ethyl acetate, gradient: 20%–50% ethyl acetate) to give the title compound (white solid, 50 mg, yield 16.1%). LC / MS (ESI) m / z: 311.0 [M+H]+.

[0193] Step 4: 2'-Chloro-3'-Fluoro-5'-Methoxy-6-methyl-[4,4'-Bipyridine]-3-carboxylic acid

[0194] Methyl 2'-chloro-3'-fluoro-5'-methoxy-6-methyl-[4,4'-bipyridine]-3-carboxylic acid (500 mg, 1.60 mmol) was dissolved in a mixed solvent of tetrahydrofuran (5 mL), methanol (5 mL), and water (5 mL), and lithium hydroxide monohydrate (67 mg, 1.60 mmol) was added. The mixture was reacted at room temperature for 5 hours, and concentrated under reduced pressure to give the title compound (white solid, 445 mg, yield 93.1%). LC / MS (ESI) m / z: 296.9 [M+H]+.

[0195] Intermediate 2: 2'-chloro-5'-methoxy-6-methyl-[4,4'-bipyridine]-3-carboxylic acid

[0196] Step 1: (2-chloro-5-methoxypyridin-4-yl)boronic acid

[0197] 2-Chloro-5-methoxypyridine (10.0 g, 69.5 mmol) was dissolved in 250 mL of tetrahydrofuran and cooled to 65 °C under nitrogen protection. 2 mol / L lithium diisopropylamino (70 mL) was slowly added dropwise while maintaining the temperature below -60 °C. After the addition was complete, the reaction was maintained at this low temperature for 2 hours. Then, triisopropyl borate (26.2 g, 139 mmol) was slowly added dropwise at -65 °C, and the mixture was stirred for another hour while maintaining the temperature at -65 °C. The mixture was then heated to room temperature overnight. The reaction solution was carefully quenched by adding 100 mL of water in an ice-water bath. The resulting aqueous solution was extracted twice with ethyl acetate. The organic phase was discarded, and the pH of the aqueous phase was adjusted to 5-6 with 2 M hydrochloric acid. A large amount of solid precipitated. The solid phase was filtered under reduced pressure and dried to give the title compound (11.4 g, white solid, yield: 88.1%). LC / MS (ESI) m / z: 188.0 [M+H]+.

[0198] Step 2: Methyl 2'-chloro-5'-methoxy-6-methyl-[4,4'-bipyridine]-3-carboxylic acid

[0199] 4-Bromo-6-methylnicotinic acid methyl ester (5 g, 21.7 mmol), (2-chloro-5-methoxypyridin-4-yl)boronic acid (4.07 g, 21.7 mmol), and potassium carbonate (9 g, 65.2 mmol) were dissolved in a mixture of 180 mL of 1,4-dioxane and 36 mL of water. Under nitrogen protection, bis(triphenylphosphine)palladium dichloride (1.59 g, 2.17 mmol) was added, and the mixture was heated to 80 °C for 2 hours. The reaction solution was cooled to room temperature, filtered, and extracted separately with water and ethyl acetate. The organic phase was concentrated and purified by silica gel column chromatography (petroleum ether / ethyl acetate system: 0–50% ethyl acetate) to give the title compound (5.20 g, white solid, yield: 81.7%). LC / MS (ESI) m / z: 293.0 [M+1]+.

[0200] Step 3: 2'-Chloro-5'-methoxy-6-methyl-[4,4'-bipyridine]-3-carboxylic acid

[0201] Methyl 2'-chloro-5'-methoxy-6-methyl-[4,4'-bipyridine]-3-carboxylic acid (5.2 g, 17.7 mmol) was dissolved in 50 mL of tetrahydrofuran and 50 mL of water. Lithium hydroxide (0.64 g, 26.6 mmol) was added, and the mixture was stirred overnight at room temperature. After removing the tetrahydrofuran by concentration under reduced pressure, the pH was adjusted to 5-6 with hydrogen chloride, resulting in the precipitation of a large amount of white solid. The solid was filtered under reduced pressure, and the resulting filter cake was dried to give the title compound (4.82 g, white solid, yield: 96.7%). LC / MS (ESI) m / z: 279.1 [M+H]+.

[0202] Example 1 (Compound 2): 2'-chloro-3'-fluoro-N-(5-(4-(hydroxymethyl)piperidin-1-yl)thiazo[5,4-b]pyridin-2-yl)-5'-methoxy-6-methyl-[4,4'-bipyridine]-3-carboxamide

[0203] Step 1: N-(5-Fluorothiazol[5,4-b]pyridin-2-yl)acetamide

[0204] Acetic anhydride (2.5 mL, 26.6 mmol) and 4-dimethylaminopyridine (0.43 g, 3.55 mmol) were added to a solution of 5-fluorothiazolyl[5,4-b]pyridine-2-amine (3 g, 17.7 mmol) in 1,2-dichloroethane (50 mL). The mixture was then reacted at 80 °C for 2 h. After the reaction was complete, methanol was added and filtered, the mixture was slurried with 10% methanol, and dried to give the title compound (near-white solid, 3.6 g, yield 96.1%). LC / MS (ESI) m / z: 212.1 (M+H)+.

[0205] Step 2: N-(5-(4-(((tert-butyldimethylsilyl)oxy)methyl)piperidin-1-yl)thiazo[5,4-b]pyridin-2-yl)acetamide

[0206] To a solution of N-(5-fluorothiazolyl[5,4-b]pyridin-2-yl)acetamide (700 mg, 3.31 mmol) in N-methylpyrrolidone (5 mL), 4-(((tert-butyldimethylsilyl)oxy)methyl)piperidine (1.14 g, 4.97 mmol) and N,N-diisopropylethylamine (1.7 mL, 9.76 mmol) were added. The mixture was then stirred at 150 °C for 32 hours. After the reaction was complete, water was added, and the mixture was extracted with ethyl acetate. The organic phase was washed with saturated brine, dried over anhydrous sodium sulfate, and concentrated under reduced pressure. The residue was purified by column chromatography (eluent: petroleum ether / ethyl acetate, gradient: 0-50% ethyl acetate) to give the title compound (yellow solid, 420 mg, yield 31.2%). LC / MS (ESI) m / z: 421.2 (M+H)+.

[0207] Step 3: (1-(2-aminothiazo[5,4-b]pyridin-5-yl)piperidin-4-yl)methanol

[0208] Sodium hydroxide (399 mg, 9.98 mmol) was added to a mixture of N-(5-(4-(((tert-butyldimethylsilyl)oxy)methyl)piperidin-1-yl)thiazo[5,4-b]pyridin-2-yl)acetamide (420 mg, 0.998 mmol) in methanol (3 mL) and water (1 mL). The reaction mixture was stirred at 80 °C for 2 h. After the reaction was complete, the mixture was concentrated under reduced pressure. The residue was purified by column chromatography (eluent: dichloromethane / methanol, gradient: 0-5% methanol) to give the title compound (white solid, 150 mg, yield 56.8%). LC / MS (ESI) m / z: 265.2 (M+H)+.

[0209] Step 4: 5-(4-(((tert-butyldimethylsilyl)oxy)methyl)piperidin-1-yl)thiazo[5,4-b]pyridine-2-amine

[0210] To a solution of (1-(2-aminothiazo[5,4-b]pyridin-5-yl)piperidin-4-yl)methanol (80 mg, 0.303 mmol) in dichloromethane (5 mL), imidazole (62 mg, 0.911 mmol) and tert-butyldimethylchlorosilane (46 mg, 0.305 mmol) were added. The mixture was stirred at room temperature for 16 hours. After the reaction was complete, water was added, and the mixture was extracted with dichloromethane. The organic phase was washed with saturated brine, dried over anhydrous sodium sulfate, and concentrated under reduced pressure. The residue was purified by column chromatography (eluent: dichloromethane / methanol, gradient: 0–5% methanol) to give the title compound (white solid, 56 mg, yield 48.9%). LC / MS (ESI) m / z: 379.4 (M+H)+.

[0211] Step 5: N-(5-(4-(((tert-butyldimethylsilyl)oxy)methyl)piperidin-1-yl)thiazo[5,4-b]pyridin-2-yl)-2'-chloro-3'-fluoro-5'-methoxy-6-methyl-[4,4'-bipyridine]-3-carboxamide

[0212] To a mixture of 5-(4-(((tert-butyldimethylsilyl)oxy)methyl)piperidin-1-yl)thiazo[5,4-b]pyridine-2-amine (60 mg, 0.159 mmol) in N,N-dimethylformamide (1.2 mL) and acetonitrile (5 mL), 2'-chloro-3'-fluoro-5'-methoxy-6-methyl-[4,4'-bipyridine]-3-carboxylic acid (71 mg, 0.239 mmol) and N-methylimidazolium (39 mg, 0.475 mmol) were added sequentially. Then, N,N,N',N'-tetramethylchloroformamidine hexafluorophosphate (54 mg, 0.192 mmol) was added to the mixture at 70 °C. The mixture was stirred at 70 °C for 2 hours. After the reaction was complete, water was added, and the mixture was extracted with ethyl acetate. The organic phase was washed with saturated brine, dried over anhydrous sodium sulfate, and concentrated under reduced pressure. The residue was purified by column chromatography (eluent: dichloromethane / methanol, gradient: 0-5% methanol) to give the title compound (yellow solid, 45 mg, yield 43.1%). LC / MS (ESI) m / z: 657.4 (M+H)+.

[0213] Step 6: 2'-Chloro-3'-Fluoro-N-(5-(4-(hydroxymethyl)piperidin-1-yl)thiazo[5,4-b]pyridin-2-yl)-5'-methoxy-6-methyl-[4,4'-bipyridine]-3-carboxamide

[0214] A solution of dioxane hydrochloride (1 mL, 4 M) was added to N-(5-(4-(((tert-butyldimethylsilyl)oxy)methyl)piperidin-1-yl)thiazo[5,4-b]pyridin-2-yl)-2'-chloro-3'-fluoro-5'-methoxy-6-methyl-[4,4'-bipyridine]-3-carboxamide (45 mg, 0.069 mmol). The reaction mixture was stirred at room temperature for 1 hour. After the reaction was complete, the mixture was concentrated under reduced pressure, and the residue was purified by preparative HPLC to give the title compound (yellow solid, 20.6 mg, yield 55.4%). 1HNMR(400MHz,DMSO)δ12.81(s,1H),8.96(s,1H),8.18(s,1H),7.85(d,J=9.2Hz ,1H),7.50(s,1H),6.99(d,J=9.2Hz,1H),4.47(t,J=5.2Hz,1H),4.35-4.27(m,2 H),3.73(s,3H),3.27(t,J=5.8Hz,2H),2.88-2.76(m,2H),2.60(s,3H),1.78-1. 70(m,2H),1.68-1.55(m,1H),1.20-1.06(m,2H).LC / MS(ESI)m / z:543.3(M+H)+.

[0215] The compounds listed in the table below were synthesized using commercially available raw materials following the synthetic steps of Example 1 (Compound 2).

[0216] Example 5 (Compound 3): 2'-Chloro-N-(6-(1,4-dimethyl-1H-1,2,3-triazol-5-yl)thiazo[4,5-c]pyridin-2-yl)-5'-methoxy-6-methyl-[4,4'-bipyridine]-3-carboxamide

[0217] Step 1: 1,4-Dimethyl-5-(tributyltin)-1H-1,2,3-triazole

[0218] At -78°C, butyllithium (2.3 mL, 3.71 mmol, 1.6 mol / L) was slowly added dropwise to a tetrahydrofuran (30 mL) solution of 1,4-dimethyl-1H-1,2,3-triazole (300 mg, 3.09 mmol). The reaction mixture was stirred at -78°C for 1 hour under nitrogen protection, followed by the addition of tributyltin chloride (0.922 mL, 3.40 mmol). The reaction mixture was stirred at -78°C for 0.5 hours under nitrogen protection. After the reaction was complete, the reaction mixture was brought to room temperature, extracted with ethyl acetate, washed with saturated brine, dried over anhydrous sodium sulfate, and concentrated under reduced pressure. The residue was purified by silica gel chromatography (eluent: petroleum ether / ethyl acetate, gradient: 0-10% ethyl acetate) to give 1,4-dimethyl-5-(tributyltinyl)-1H-1,2,3-triazole (yellow oil, 1100 mg, yield: 92.2%). LC / MS (ESI) m / z: 388.2 [M+H] + .

[0219] Step 2: 6-(1,4-dimethyl-1H-1,2,3-triazol-5-yl)-2-(2,5-dimethyl-1H-pyrrolo-1-yl)thiazole[4,5-c]pyridine

[0220] To a solution of 1,4-dimethyl-5-(tributyltinyl)-1H-1,2,3-triazole (1000 mg, 2.59 mmol) and 6-chloro-2-(2,5-dimethyl-1H-pyrrolo-1-yl)thiazo[4,5-c]pyridine (683 mg, 2.59 mmol) in 1,4-dioxane (30 mL), tris(dibenzylacetone)dipalladium (355.7 mg, 0.39 mmol), tricyclohexylphosphine (217.9 mg, 0.78 mmol), and cesium carbonate (1687 mg, 5.18 mmol) were added. The reaction mixture was stirred at 110 °C for 5 hours under nitrogen protection. After the reaction was complete, it was cooled to room temperature, extracted with water and ethyl acetate, washed with saturated brine, dried over anhydrous sodium sulfate, and concentrated under reduced pressure. The residue was purified by silica gel chromatography (eluent: petroleum ether / ethyl acetate, gradient: 0-30% ethyl acetate) to give 6-(1,4-dimethyl-1H-1,2,3-triazol-5-yl)-2-(2,5-dimethyl-1H-pyrrolo-1-yl)thiazolyl[4,5-c]pyridine (yellow solid, 140 mg, yield: 16.7%). LC / MS (ESI) m / z: 325.1 [M+H] + .

[0221] Step 3: 6-(1,4-dimethyl-1H-1,2,3-triazol-5-yl)thiazolyl[4,5-c]pyridine-2-amine

[0222] Trifluoroacetic acid (5.0 mL) and water (one drop) were added to 6-(1,4-dimethyl-1H-1,2,3-triazol-5-yl)-2-(2,5-dimethyl-1H-pyrrolo-1-yl)thiazolyl[4,5-c]pyridine (140 mg, 0.432 mmol), and the mixture was stirred at 70 °C for 1 hour. After the reaction was complete, the reaction solution was concentrated under reduced pressure, and the crude product 6-(1,4-dimethyl-1H-1,2,3-triazol-5-yl)thiazolyl[4,5-c]pyridine-2-amine (yellow solid, 43.0 mg, yield: 40.5%) was used directly in the next step. LC / MS (ESI) m / z: 247.1 [M+H] + .

[0223] Step 4: 2'-Chloro-N-(6-(1,4-dimethyl-1H-1,2,3-triazol-5-yl)thiazo[4,5-c]pyridin-2-yl)-5'-methoxy-6-methyl-[4,4'-bipyridine]-3-carboxamide

[0224] 6-(1,4-dimethyl-1H-1,2,3-triazol-5-yl)thiazolyl[4,5-c]pyridine-2-amine (30.0 mg, 0.12 mmol), 2'-chloro-5'-methoxy-[3,4'-bipyridine]-4-carboxylic acid (34.0 mg, 0.12 mmol), N,N,N',N'-tetramethylchloromethamidine hexafluorophosphate (34.2 mg, 0.12 mmol) and N-methylimidazolium (30.0 mg, 0.36 mmol) were dissolved in acetonitrile (10 mL), and the reaction mixture was stirred at 70 °C for 4 hours. After the reaction was completed, the reaction solution was concentrated under reduced pressure, and the residue was purified by preparative HPLC to obtain 2'-chloro-N-(6-(1,4-dimethyl-1H-1,2,3-triazol-5-yl)thiazo[4,5-c]pyridin-2-yl)-5'-methoxy-6-methyl-[4,4'-bipyridine]-3-carboxamide (white solid, 12.2 mg, yield: 19.8%). 1 H NMR(400MHz,DMSO-d6)δ13.30(s,1H),9.20(s,1H),8.88(s,1H),8.40(s,1H),8.17(s,1H),7.61(s, 1H),7.49(s,1H),4.14(s,3H),3.60(s,3H),2.62(s,3H),2.39(s,3H).LC / MS(ESI)m / z:507.2[M+H] + .

[0225] Example 6 (Compound 4): 2'-Chloro-N-(6-(1,4-dimethyl-1H-1,2,3-triazol-5-yl)-7-fluorothiazo[4,5-c]pyridin-2-yl)-5'-methoxy-6-methyl-[4,4'-bipyridine]-3-carboxamide

[0226] Step 1: O-((9H-fluorene-9-yl)methyl)carbonate isothiocyanate

[0227] At 0 °C, a solution of chloroformate-9-fluorenyl methyl ester (50 g, 193 mmol) in ethyl acetate (160 mL) was added dropwise to a solution of potassium thiocyanate (20.7 g, 213 mmol) in ethyl acetate (160 mL). The mixture was stirred at room temperature for 16 hours. The mixture was filtered, the filtrate was concentrated under reduced pressure, and the residue was purified by silica gel column chromatography (eluent: petroleum ether / ethyl acetate = 20, V / V) to give the title compound (yellow oil, 16.0 g, yield 29.4%). ¹H NMR (400 MHz, DMSO-d6) δ 7.89 (d, J = 7.6 Hz, 2H), 7.69 (d, J = 7.6 Hz, 2H), 7.45–7.39 (m, 2H), 7.36–7.31 (m, 2H), 4.30–4.23 (m, 2H), 4.23–4.18 (m, 1H).

[0228] Step 2: (6-chloro-5-fluoropyridin-3-yl)tert-butyl carbamate

[0229] A solution of 2-chloro-3-fluoro-5-bromopyridine (8.00 g, 38.0 mmol), tert-butyl carbamate (4.90 g, 41.8 mmol), cesium carbonate (24.8 g, 76.0 mmol), and 4,5-bis(diphenylphosphine-9,9-dimethyloxanthracene) (880 mg, 1.52 mmol) in 1,4-dioxane (160 mL) was mixed with tris(dibenzylacetone)dipalladium (1.04 g, 1.14 mmol). The mixture was stirred at 85 °C for 20 hours under nitrogen protection. The mixture was concentrated under reduced pressure, and the residue was purified by silica gel column chromatography (eluent: petroleum ether / ethyl acetate = 4, V / V) to give the title compound (yellow solid, 6.10 g, yield 65.0%). LC / MS (ESI) m / z: 246.9 [M+H]+.

[0230] Step 3: (4-bromo-6-chloro-5-fluoropyridine)-3-carbamate tert-butyl ester

[0231] (6-chloro-5-fluoropyridin-3-yl)tert-butyl carbamate (6.10 g, 24.7 mmol) and N,N,N',N'-tetramethylethylenediamine (8.62 g, 74.2 mmol) were dissolved in diethyl ether (130 mL). Under nitrogen protection, the reaction mixture was cooled to -60 °C, and 1.6 M n-butyllithium (46.4 mL, 74.2 mmol) was added dropwise. After the addition was complete, the mixture was heated to -20 °C and stirred for 1.5 hours. The reaction mixture was then cooled to -60 °C, and 1,2-dibromotetrafluoroethane (19.9 g, 76.7 mmol) was added dropwise. After the addition was complete, the mixture was slowly heated to room temperature, and the reaction was quenched with 1 N HCl (92 mL). The mixture was extracted with ethyl acetate, the organic phase was washed with water and brine, dried over anhydrous sodium sulfate, and filtered. The mixture was concentrated under reduced pressure to give the title compound (yellow solid, 8.0 g, 99.4% yield). LC / MS(ESI)m / z:326.9[M+H]+.

[0232] Step 4: 4-Bromo-6-chloro-5-fluoropyridine-3-amine

[0233] (4-Bromo-6-chloro-5-fluoropyridine)-3-carbamate tert-butyl ester (8.0 g, 24.6 mmol) was dissolved in dichloromethane (100 mL) and trifluoroacetic acid (50 mL), and the reaction mixture was stirred at room temperature for one hour. The mixture was concentrated under reduced pressure to obtain a residue, which was then dissolved in ethyl acetate, neutralized with saturated aqueous sodium bicarbonate solution, and extracted with ethyl acetate. The organic phase was washed with water and brine, dried over anhydrous sodium sulfate, and filtered. The mixture was concentrated under reduced pressure to give the title compound (yellow solid, 5.7 g, yield 103%). LC / MS (ESI) m / z: 224.8 [M+H]+.

[0234] Step 5: (9H-fluorene-9-yl)methyl(6-chloro-7-fluorothiazo[4,5-c]pyridin-2-yl)carbamate hydrobromide

[0235] 4-Bromo-6-chloro-5-fluoropyridin-3-amine (5.7 g, 25.3 mmol) was dissolved in acetone (100 mL), and O-((9H-fluorene-9-yl)methyl)carbonate isothiocyanate (8.54 g, 30.3 mmol) was added. The mixture was stirred overnight at 50 °C. The reaction solution was cooled to room temperature, filtered, and the filter cake was washed with acetone. The solid was dried under reduced pressure to give the title compound (7.7 g, white solid, yield 60.1%). LC / MS (ESI) m / z: 426.0 [M+H]+.

[0236] Step 6: 6-Chloro-7-fluorothiazo[4,5-c]pyridine-2-amine

[0237] To a solution of (9H-fluorene-9-yl)methyl(6-chloro-7-fluorothiazo[4,5-c]pyridin-2-yl)carbamate hydrobromide (7.7 g, 18.1 mmol) in dichloromethane (80 mL), piperidine (17.9 mL, 181 mmol) was added, and the mixture was stirred at room temperature for one hour. Water was added to the reaction mixture, and the mixture was back-extracted with ethyl acetate and concentrated under reduced pressure in aqueous phase. The residue was purified by silica gel column chromatography (eluent: dichloromethane / methanol = 10, v / v) followed by slurrying with dichloromethane to give the title compound (white solid, 1.45 g, yield 39.4%). LC / MS (ESI) m / z: 204.1 [M+H]+.

[0238] Step 7: 6-Chloro-2-(2,5-dimethyl-1H-pyrrolo-1-yl)-7-fluorothiazo[4,5-c]pyridine

[0239] To a solution of 6-chloro-7-fluorothiazo[4,5-c]pyridin-2-amine (1.45 g, 7.12 mmol), acetone-1,65 g, 14.5 mmol, and toluene (30 mL), p-toluenesulfonic acid hydrate (0.25 g, 1.42 mmol) was added, and the mixture was refluxed at 140 °C for two hours to remove water. The reaction mixture was cooled to room temperature, ethyl acetate was added, and the mixture was washed with aqueous sodium bicarbonate solution, followed by washing with saturated brine. The mixture was concentrated under reduced pressure, and the residue was purified by silica gel column chromatography (eluent: petroleum ether / ethyl acetate = 10, V / V) to give the title compound (white solid, 1.5 g, 74.8% yield). LC / MS (ESI) m / z: 282.1 [M+H]+.

[0240] Step 8: 6-(1,4-dimethyl-1H-1,2,3-triazol-5-yl)-2-(2,5-dimethyl-1H-pyrrolo-1-yl)-7-fluorothiazo[4,5-c]pyridine

[0241] To a xylene (30 mL) solution of 6-(1,4-dimethyl-1H-1,2,3-triazol-5-yl)-2-(2,5-dimethyl-1H-pyrrolo-1-yl)-7-fluorothiazo[4,5-c]pyridine (1.5 g, 5.32 mmol) and 1,4-dimethyl-5-(tributyltinyl)-1H-1,2,3-triazole (3.08 g, 7.99 mmol), tetraphenylphosphine palladium (1.85 g, 1.60 mmol) was added, and the mixture was stirred at 150 °C for 5 hours under nitrogen protection. The mixture was concentrated under reduced pressure, and the residue was purified by silica gel column chromatography (eluent: petroleum ether / ethyl acetate = 3, V / V) to give the title compound (pale yellow solid, 1.25 g, yield 68.7%). LC / MS (ESI) m / z: 343.1 [M+H]+.

[0242] Step 9: 6-(1,4-dimethyl-1H-1,2,3-triazol-5-yl)-7-fluorothiazo[4,5-c]pyridine-2-amine hydrochloride

[0243] 6-(1,4-dimethyl-1H-1,2,3-triazol-5-yl)-2-(2,5-dimethyl-1H-pyrrolo-1-yl)-7-fluorothiazo[4,5-c]pyridine (1.25 g, 3.65 mmol) was dissolved in 2N HCl (15 mL), and the reaction mixture was stirred at 80 °C for two hours. The mixture was concentrated under reduced pressure, and the residue was slurried with acetonitrile to give the title compound (pale yellow solid, 1.0 g, 91.1% yield). LC / MS (ESI) m / z: 265.1 [M+H]+.

[0244] Step 10: 2'-Chloro-N-(6-(1,4-dimethyl-1H-1,2,3-triazol-5-yl)-7-fluorothiazo[4,5-c]pyridin-2-yl)-5'-methoxy-6-methyl-[4,4'-bipyridine]-3-carboxamide

[0245] 6-(1,4-dimethyl-1H-1,2,3-triazol-5-yl)-7-fluorothiazo[4,5-c]pyridine-2-amine hydrochloride (1.0 g, 3.78 mmol), 2'-chloro-5'-methoxy-6-methyl-[4,4'-bipyridine]-3-carboxylic acid (1.16 g, 4.16 mmol), and N-methylimidazolium (2.17 g, 26.5 mmol) were dissolved in acetonitrile (20 mL) and stirred at 70 °C for five minutes. N,N,N',N'-tetramethylchloroformamidine hexafluorophosphate (5.31 g, 18.9 mmol) was added to the reaction solution and stirred at 70 °C for two hours. Before the reactants were completely reacted, N-methylimidazole (0.62 g, 7.56 mmol) and N,N,N',N'-tetramethylchloroformamidin hexafluorophosphate (1.06 g, 3.78 mmol) were added to the reaction solution. Water was added to the reaction solution, and the mixture was extracted with ethyl acetate. The organic phase was washed with water and brine, concentrated under reduced pressure, and the residue was first subjected to silica gel column chromatography (eluent: dichloromethane / methanol = 20, v / v), and then slurried with ethyl acetate to give the title compound (834.7 mg, white solid, yield 42.0%). 1H NMR (400MHz, DMSO-d6) δ13.63(s,1H),9.14(d,J=2.0Hz,1H),8.90(s,1H),8.18(s,1H),7.62(s,1H),7. 51(s,1H),4.03(s,3H),3.61(s,3H),2.62(s,3H),2.26(d,J=1.6Hz,3H).LC / MS(ESI)m / z:525.2[M+H]+.

[0246] The compounds listed in the table below were synthesized using commercially available raw materials following the synthetic steps of Example 6 (Compound 4).

[0247] Efficacy Test Example 1: DLD1-BRCA2- / - (BRCA-mutated colon cancer cell line)

[0248] The in vitro efficacy of the inhibitor was assessed using a cell viability assay, specifically the commonly used CTG assay, with CellTiter Glo reagent (Promega, G7573). The specific assay method is as follows:

[0249] 1) Culture DLD1 and DLD1-BRCA2- / - cells. One day before the assay, digest the cells with trypsin (0.025% Trypsin-EDTA, Hyclone), centrifuge at 1000 rpm for 3 min, and collect the cells. Count the cells using a cell counter (Shanghai Mengwei Biomedical Technology Co., Ltd., SmartCell600A.SC1006), and seed them into 96-well white culture plates (WHB, catalog number WHB-96) at an appropriate seeding ratio.

[0250] 2) 24 hours after cell seeding, the drug was added, which was recorded as Day 0. The compound was added using an automated pipette (Thermo, Multidrop8). The plate was set as a 96-well plate with an initial concentration of 0.5 μM. The cells were diluted by 1 / 3 of the initial concentration, and a total of 9 detection points were set up, with 2 replicates per group. The cells were incubated at 37°C in a 5% CO2 incubator.

[0251] 3) Perform two solution changes on Day 3 and Day 7 respectively, and add the compound as in step 2);

[0252] 4) On Day 10, remove the cell culture plate and add an equal volume of CTG reagent (the CTG reagent needs to be brought back to temperature and premixed according to the reagent instructions). Gently mix for 10 minutes (speed 300) on a constant temperature mixer (Hangzhou Aosheng Instrument Co., Ltd., MSC-100). Perform chemiluminescence detection using an ELISA reader (Thermo, Varioskan LUX).

[0253] 5) Perform data analysis on the measurement results: The CV% of the control group should be less than 20%, and the z' value should be greater than 0.5. For data meeting the above quality control results, calculate cell viability (inhibition rate (%) = 100 × (control group average - experimental group) / (control group average - culture medium control group average). Use GraphPad Prism8 to perform nonlinear regression fitting of the inhibition rate curve of the compound and obtain the IC50 value.

[0254] The inhibition rate curves of compounds 1-4 against DLD1-BRCA2- / - are shown in Figure 1-4. The IC50 values ​​of compounds 1-4 are shown in Figure 1-4. 50 The concentrations were 20.73 nM, 9.68 nM, 10.10 nM, and 5.20 nM, respectively; they did not show inhibitory effects on the DLD1 cell line, and the IC50 values ​​were >50000 nM, 37106 nM, 29286 nM, and >5000 nM, respectively, with selectivity of >1000-fold.

[0255] Efficacy Test Example 2: HCT116, HCT116-BRCA2- / - (primary colon cancer cell line with BRCA mutation)

[0256] Referring to Example 1 of the efficacy test, the cell line was replaced with HCT116-BRCA2- / - cells, the inhibition rate curve of the compound was fitted, and the IC50 value was obtained.

[0257] The inhibition rate curves of compound 5 against HCT116 and HCT116-BRCA2- / - are shown in Figures 5 and 6. Among them, the compound showed good inhibition against HCT116-BRCA2- / -, with an IC50 of 9.69 nM; it showed almost no inhibitory effect on HCT116, with an IC50 > 30000 nM, and the selectivity between the two was > 1000 times.

[0258] Efficacy test case 3: PC-3-BRCA2- / - (prostate cancer line with BRCA2 mutation)

[0259] Referring to Example 1 of the efficacy test, the cell line was replaced with PC-3-BRCA2- / - cells, the inhibition rate curve of the compound was fitted, and the IC50 value was obtained.

[0260] The inhibition rate curves of compound 6 against PC3 and PC3-BRCA2- / - are shown in Figure 7. Compound 6 showed good inhibition of PC3-BRCA2- / -, with an IC50 of 132.32 nM; it showed almost no inhibitory effect on PC3, with an IC50 > 10000 nM, and the selectivity between the two was > 75 times.

[0261] Efficacy Test Case 4: LNCAP-Clone FGC (Metastatic Prostate Cancer Cell Line)

[0262] Referring to Example 1 of the efficacy test, the cell line was replaced with LNCAP-Clone FGC, and the inhibition rate curve of the compound was fitted and the IC50 value was obtained. The LNCAP-Clone FGC cell line contains the following related gene mutations: RAD51B, POLB, POLE2, POLH, MSH6, RNASEH2B, etc. Among them, RAD51B is a key protein in the HR repair pathway, POLB is a key protein in the base excision repair pathway, POLE2 and POLH are DNA polymerases in the TLS pathway, and MSH6 is a key protein in the MMR repair pathway.

[0263] The inhibition rate curve of compound 7 against LNCAP-Clone FGC is shown in Figure 8. Compound 7 has a certain killing effect on LNCAP-Clone FGC, with an IC50 of 18.8 μM.

[0264] Efficacy Test Case 5: PEO1 (BRCA2-mutated ovarian cancer cell line)

[0265] Referring to Example 1 of the efficacy test, the cell line was replaced with PEO1, the inhibition rate curve of the compound was fitted, and the IC50 value was obtained.

[0266] The inhibition rate curve of compound 8 against PEO1 is shown in Figure 9. Compound 9 has a strong killing effect on the BRCA2-mutated ovarian cancer cell line PEO1, with an IC50 of 20.6 nM.

[0267] Example 6 of efficacy testing: SKUT1, SKUT1-BRCA2 (human uterine leiomyosarcoma cell line with BRCA mutation)

[0268] Referring to Example 1 of the efficacy test, the cell lines were replaced with SKUT1 and SKUT1-BRCA2, and the inhibition rate curves of the compounds were fitted to obtain the IC50 values.

[0269] The inhibition rate curves of compound 9 against SKUT1 and SKUT1-BRCA2 are shown in Figure 10. The IC50 value of the compound against SKUT1-BRCA2 is 1.6818 μM, while it shows almost no inhibitory effect on SKUT1.

[0270] Efficacy Test Example 7: MDA-MB-436 (Breast Cancer Line with BRCA1 Mutation, TNBC)

[0271] Referring to Example 1 of the efficacy test, the cell line was replaced with MDA-MB-436, the inhibition rate curve of the compound was fitted, and the IC50 value was obtained.

[0272] The inhibition rate curve of compound 7 against MDA-MB-436 is shown in Figure 11. The compound has a strong killing effect on breast cancer cell lines with BRCA mutations, with an IC50 value of 73.07 nM.

[0273] Efficacy Test Example 8: DLD1-XRCC3, DLD1-RAD51C (HR-deficient cell lines such as XRCC3 and RAD51C)

[0274] Referring to Example 1 of the efficacy test, the cell line was replaced with DLD1, DLD1-XRCC3 or DLD1, DLD1-RAD51C, and the inhibition rate curve of the compound was fitted and the IC50 value was obtained.

[0275] Compound 10 showed virtually no inhibitory effect on DLD1. The inhibition rate curve of compound 10 against DLD1-XRCC3 is shown in Figure 12, with an IC50 value of 27.1 nM. The inhibition rate curve of compound 10 against DLD1-RAD51C is shown in Figure 13, with an IC50 value of 3.50 nM.

[0276] Efficacy Test Example 9: MDA-MB-436-53BP1, MDA-MB-436-SHLD2 (breast cancer line with BRCA1 mutation and Shieldin complex mutation, TNBC)

[0277] Referring to Example 1 of the efficacy test, the cell line was replaced with MDA-MB-436-53BP1 or MDA-MB-436-SHLD2, the detection time was adjusted to 14 days, the inhibition rate curve of the compound was fitted, and the IC50 value was obtained.

[0278] The inhibition rate curve of compound 7 against MDA-MB-436-53BP1 is shown in Figure 14, with an IC50 value of 18.47 nM.

[0279] The inhibition rate curve of compound 4 against MDA-MB-436-SHLD2 is shown in Figure 15, with an IC50 value of 23.78 nM.

[0280] Efficacy Test Case 10: PEO1-53BP1 (ovarian cancer cell line with BRCA2 mutation and Shieldin complex mutation)

[0281] Referring to Example 1 of the efficacy test, the cell line was replaced with PEO1-53BP1, the inhibition rate curve of the compound was fitted, and the IC50 value was obtained.

[0282] The inhibition rate curve of compound 8 against PEO1-53BP1 is shown in Figure 16, with an IC50 value of 2.8 nM.

[0283] Efficacy Test Example 11: HCC1937 (a breast cancer line with BRCA1 mutation and Shieldin complex mutation, TNBC)

[0284] 11.1 Cell Culture

[0285] The tumor cell lines were cultured in an incubator at 37°C and 5% CO2 according to the culture conditions shown in Table 1. Cells were passaged at 1 / 3 rate (every 3 days), and cells in the logarithmic growth phase were used for plating.

[0286] 11.2 Cell Plating

[0287] (1) Use trypan blue to stain cells and count live cells.

[0288] (2) Adjust the cell concentration to a suitable level, as shown in the table below for cell plating density:

[0289] Table 1 HCC1937 clone formation plating conditions

[0290] (3) Add 2 mL of cell suspension to each well of the culture plate.

[0291] (4) Incubate the culture plate overnight in an incubator at 37°C, 5% CO2 and 100% relative humidity.

[0292] 11.3 Combined treatment of cells with compounds

[0293] (5). Preparation of 10 mM compound 4 mother liquor:

[0294] (6) Preparation of compounds at various concentrations: Take 10 μL of the 10 mM test compound 4 and add it to 40 μL of DMSO for serial dilution in a (1 / 5) manner to obtain working solutions of 1000 μM, 200 μM, 40 μM, 8 μM, 1.6 μM and 0 μM respectively.

[0295] (7) Drug addition: Add 2 μL of the diluted compound working solution to the cell culture plate as shown (1:1000). Add 2 μL of DMSO to the blank control.

[0296] (8) Place the six-well cell plate back into the incubator for further culture.

[0297] (9) For each concentration of compound experimental group, replace the culture medium with fresh solution containing the corresponding concentration of drug on days 1, 5, 8 and 12.

[0298] 11.4 Crystal Violet Staining

[0299] (1) Remove the six-well plate from the incubator and remove the culture medium. Wash each well with 1 mL of 1×PBS and then remove the PBS.

[0300] (2). Fix cells with 100% methanol at 1 mL / well (at room temperature for 15 minutes).

[0301] (3) After fixation, remove methanol, add 1 mL of 1×PBS to each well to wash once, and then remove PBS.

[0302] (4) Weigh 0.05 g of crystal violet and prepare a 0.1% solution with 1×PBS.

[0303] (5) Stain with crystal violet from step (4) at a rate of 1 mL / well for 1 hour.

[0304] (6) After staining, wash three times with 1×PBS (5 minutes each time) until there is no background staining.

[0305] (7) Take photos and record the experimental data.

[0306] 11.5 Data Analysis

[0307] The clone area of ​​each well was calculated using ImageJ software. The inhibition rate (IR) of the detected compound was calculated using the following formula: IR(%) = ((blank control – RLU compound) / (blank control)) × 100%. The inhibition rate of different concentrations of the compound was calculated in Excel.

[0308] The test results are shown in Figure 17. Figure 17 shows the results of the colony formation experiment. When the concentration of compound 7 is 1.6 nM, the inhibition rate is >50%.

[0309] Efficacy Test Example 12: TOV21G (FANCF-mutated ovarian cancer cell line)

[0310] Referring to Example 1 of the efficacy test, the cell line was replaced with TOV21G, the inhibition rate curve of the compound was fitted, and the IC50 value was obtained.

[0311] The inhibition rate curve of compound 6 against TOV21G is shown in Figure 18. The compound can effectively kill TOV21G cells, with an IC50 value of 120.3 nM.

[0312] Efficacy Test Case 13: SNU-601 (Gastric Cancer Cell Line, POLQ High-Expression Cell Line)

[0313] Referring to Example 1 of the efficacy test, the cell line was replaced with SNU-601, the inhibition rate curve of the compound was fitted, and the IC50 value was obtained.

[0314] The inhibition rate curve of compound 7 against SNU-601 is shown in Figure 19. Compound 7 can effectively kill SNU-601 cells, with an IC50 value of 0.091 μM.

[0315] Efficacy Test Example 14: KMM1 (Multiple Myeloma Cell Line)

[0316] Referring to Example 1 of the efficacy test, the cell line was replaced with KMM1, and the inhibition rate curve of the compound was fitted to obtain the IC50 value. The KMM1 cell line is a BRCA2 CNA loss cell line with a FANCD2 driver mutation.

[0317] The inhibition rate curve of compound 7 against KMM1 is shown in Figure 20. Compound 7 can effectively kill KMM1 cells, with an IC50 value of 14.2 μM.

[0318] Efficacy Test Example 15: DLD1-REV7- / - (TLS polymerase-mutated colon cancer cell line)

[0319] Referring to Example 11 of the efficacy test, the cell line was replaced with DLD1-REV7- / -, and the inhibition rate of the fitted compound was calculated based on the ratio of the clonal area of ​​the drug-treated group to the clonal area of ​​the control group.

[0320] The test results are shown in Figure 21. Figure 21 shows the results of the colony formation experiment of the DLD1-REV7- / - cell line. The test compound 1 has a significant killing effect on DLD1-REV7- / -, and the inhibition rate is >50% when the concentration is 1.6 nM.

[0321] Effect Test Example 16

[0322] Experimental methods:

[0323] 1. Biochemical activity assay of Pol θ inhibitors (ADP Glo assay)

[0324] The N-terminus of the Polθ protein contains a helicase domain with ATPase activity, capable of hydrolyzing ATP to ADP, which can be detected by the ADP-Glo ​​assay kit (ADP-Glo). TM The Kinase Assay (Promega, V9102) was used to analyze the ATPase activity of Polθ protein and the inhibitory effect of small molecule compounds on the protein. The specific detection method is as follows:

[0325] 1) The helicase domain of the Polθ protein was purified using an insect system to obtain protein with a purity >90%. ssDNA (SEQ ID NO.1:5'-CCAGTGAATTGTTGCTCGGTACCTGCTAAC-3', Hangzhou Youkang Biotechnology Co., Ltd.) was ordered as the substrate for the Polθ protein; a reaction buffer containing 10 mM DTT (dithiothreitol), 20 mM MgCl2, Tris-HCl, and pH 7.5 was prepared.

[0326] 2) Prepare 2x ssDNA-Polθ premix and perform the reaction in a 384-well plate. Set up a control group with only buffer and add the premix to the other groups. Add the compound using an automated pipette (Thermo, Multidrop 8), starting at a concentration of 10 μM and diluting by 1 / 3. Set up a total of 9 detection sites, with 2 replicates per group.

[0327] 3) Prepare 2xATP solution, add an equal volume of 2xATP solution to all wells, and let stand at room temperature for 60 min;

[0328] 4) Following the Promega reagent instructions, add ADP-Glo ​​Detection Reagent to the reaction system and let it stand at room temperature for 60 minutes;

[0329] 5) Following the Promega reagent instructions, add Kinase Detection Reagent to the reaction system, let it stand at room temperature for 60 min, and use a microplate reader (Thermo, Varioskan LUX) to detect the chemiluminescence signal. Set the reading time interval for each well to 1000 ms.

[0330] Data analysis was performed on the measurement results: the CV% of the control group test results should be less than 10%, and the z' value should be greater than 0.5. Data meeting the above quality control results were used to calculate the inhibition rate of the compound (inhibition rate (%) = 100 * (control group average - experimental group) / (control group average - buffer group average). A nonlinear regression was used to fit the inhibition rate curve of the compound and the IC50 value was obtained.

[0331] Data list:

[0332] 2. Solubility Assay

[0333] 1) Prepare 0.1M Na2PO4 buffer (pH 7.4):

[0334] Add 11g Na2HPO4 (FW: 141.96) and 3.5g NaH2PO4_2H2O (FW: 156.03) to 1L of Mili-Q water, and adjust the pH to 7.4 with phosphoric acid or sodium hydroxide;

[0335] 2) Take 10 μL of the test compound (concentration: 10 mM in DMSO) and add it to 990 μL of the Na2PO4 buffer prepared in step 1 (final DMSO concentration: 1%).

[0336] 3) Shake the sample tube at room temperature for 2 hours (1000 rpm / min);

[0337] 4) Preparation of calibration curve:

[0338] a) Preparation of 300 μM spiking solution (SS):

[0339] Add 6 μL (10 mM in DMSO) of the test compound stock solution to 194 μL MeOH / ACN (4:1);

[0340] b) Plot the standard curve:

[0341] 5) Centrifuge the sample (10 minutes, 12,000 rpm) to precipitate undissolved particles. Filter the supernatant through a 0.22 μm filter membrane and then transfer the supernatant to a new centrifuge tube.

[0342] 6) Dilute the supernatant 10 times with 100mM buffer.

[0343] Add the supernatant (10 μL) to the buffer (100 mM, 90 μL) and dilute 10 times;

[0344] 7) Sample preparation for LC-MS / MS (API 4000) detection

[0345] Add 10 μL of sample (10-fold dilution) and standard curve sample to 400 μL of solution (MeOH:ACN = 1:1), perform instrument detection, and calculate the solubility value based on the standard curve.

[0346] Experimental results:

[0347] 4. Metabolic stability of mouse liver microsomes assay

[0348] 1) Preparation of experimental reagents

[0349] a) Phosphate-buffered saline (PBS): 100 mM, pH = 7.4 ± 0.1.

[0350] Dissolve 73.21g of dipotassium hydrogen phosphate trihydrate K2HPO4·3H2O (MW=228.22) and 10.78g of potassium dihydrogen phosphate KH2PO4 (MW=136.09) in 4000mL of pure water, and adjust the pH to 7.4±0.1 with phosphoric acid or potassium hydroxide.

[0351] b) Microparticle working solution; 1.0 mg / mL.

[0352] The purchased mouse liver microsomes (IPHASE, Cat. No.: 0121E1.01, Lot. No.: 22F001) were 20 mg / mL (10 mg, 0.5 mL). 225 μL of the solution was added to 4275 μL of PBS to prepare a 1.0 mg / mL microsome working solution.

[0353] c) NADPH regeneration system working solution: a mixture of 2 mM NADPH solution and 6 mM magnesium chloride solution

[0354] I. Weigh 6.8 mg of NADPH (MW = 833.35, 98%) powder and add it to 2000 μL of PBS solution to obtain a NADPH stock solution with a concentration of 40 mM.

[0355] II. Weigh 24.64 mg of magnesium chloride hexahydrate (MV = 203.3, 99%) solid, dissolve it in 10 mL of PBS solution to prepare a 12 mM magnesium chloride stock solution. (Note whether precipitation occurs.)

[0356] III. Take 200 μL of 40 mM NADPH stock solution, 2.0 mL of 12 mM magnesium chloride stock solution, and 1.80 mL of PBS solution to prepare a working solution for the NADPH regeneration system containing a mixture of 2 mM NADPH solution and 6 mM magnesium chloride solution.

[0357] d) Termination solution: Tolbutamide acetonitrile solution containing 200 ng / mL

[0358] I. Weigh 6.006 mg of tolbutamide (MV = 270.35, 99.9%) solid and add 30 mL of acetonitrile to prepare a 200 μg / mL tolbutamide stock solution.

[0359] II. Take 500 μL of 200 μg / mL tolbutamide stock solution and add 500 mL of acetonitrile to obtain a 200 ng / mL tolbutamide acetonitrile solution.

[0360] e) Test / Reference Working Solution: 100 μM test / reference working solution

[0361] I. Purchase 1 mL of a 1.0 mg / mL testosterone (MV = 288.42) solution and add 155.8 μL ACN to prepare a 3 mM testosterone stock solution.

[0362] II. Take 50 μL of 3 mM testosterone stock solution and add 1450 μL of acetonitrile to prepare a working solution containing 100 μM testosterone.

[0363] III. Weigh 6.98 mg propafenone hydrochloride and dissolve it in 1.8433 mL of acetonitrile; weigh 5.67 mg diclofenac sodium and dissolve it in 1.783 mL of acetonitrile. Prepare 10 mM stock solutions of propafenone and diclofenac, respectively.

[0364] IV. Take 10 μL of propafenone and diclofenac stock solutions respectively, add 990 μL of acetonitrile, and prepare working solutions of propafenone and diclofenac respectively containing 100 μM.

[0365] 2 Experimental Procedure

[0366] a) Prepare 8 incubation plates and name them T0, T5, T15, T30, T45, T60, Blank60 and NCF60 respectively.

[0367] b) Add 100 μL of microparticle working solution to each of T0, T5, T15, T30, T45, T60, Blank60 and NCF60. Add 2 μL of test sample (reference) working solution to each of T0, T5, T15, T30, T45, T60 and NCF60. Add 2 μL of acetonitrile to Blank60.

[0368] c) Place T5, T15, T30, T45, T60, and Blank60 in a 37°C water bath for pre-incubation for 10 minutes.

[0369] d) Add 600 μL of stop solution to the T0 sample to terminate the reaction; add 98 μL of phosphate buffer to NCF60, and use the NCF60 sample without the NADPH generation system as a negative control.

[0370] e) Add 98 μL of NADPH regeneration working solution to the T0 sample after the previous step is terminated. After the pre-incubation at T5, T15, T30, T45, T60, and Blank60, add 98 μL of NADPH regeneration working solution to start the reaction. Therefore, the total incubation system is 200 μL, the test sample (control) concentration is 1 μM, the microsomal concentration is 0.5 mg / mL, and the NADPH regeneration solution concentration is 1 mM.

[0371] f) Place the above T5, T15, T30, T45, T60, Blank60 and NCF60 samples into a 37℃ water bath and incubate for the corresponding times: 5min, 15min, 30min, 45min, 60min, 60min and 60min respectively.

[0372] g) After incubation for the corresponding time, 600 μL of stop solution was added to the T5, T15, T30, T45, T60, Blank60 and NCF60 samples respectively to terminate the reaction.

[0373] h) After termination, the samples T0, T5, T15, T30, T45, T60, Blank60, and NCF60 were shaken well and centrifuged at 14000 rpm for 5 min at 4℃. 100 μL of the supernatant was taken for analysis by LC-MS / MS.

[0374] 3. Data Processing:

[0375] (1) The formula for calculating half-life is as follows:

[0376] Experimental results:

[0377] In the table above, "∞" represents infinity.

[0378] 5. Mouse PK Assay

[0379] The purpose of this experiment was to conduct a pharmacokinetic study on the test compound in male ICR mice (18-22g, 4-6 weeks old, Vital River Pharmaceuticals, Beijing) after oral administration. The experimental procedure is as follows:

[0380] 1) Dissolve the compound in 5% DMSO + 40% PEG + 20% (10% TPGS) in water + 35% water to a concentration of 0.5 mg / mL, and then administer orally (5 mL / Kg);

[0381] 2) Collect blood samples: 0.03 mL at 0.25, 0.5, 1, 2, 4, 8, and 24 hours, and add EDTA-K2. CD1 mice: Collect approximately 0.03 mL of blood at each time point. Centrifuge at 4000g for 5 minutes at 4℃ to collect plasma. Aliquot plasma samples into clean polyethylene microcentrifuge tubes and store at -75±15℃ until analysis.

[0382] 3) Sample Analysis: The concentration of compounds in plasma samples will be analyzed using LC-MS / MS. Pharmacokinetic parameters will be calculated from the plasma assay results using WinNonlin (Phoenix™, version 8.3) or other similar software.

[0383] Experimental results:

[0384] While specific embodiments of the present invention have been described above, those skilled in the art should understand that these are merely illustrative examples, and various changes or modifications can be made to these embodiments without departing from the principles and essence of the present invention. Therefore, the scope of protection of the present invention is defined by the appended claims.

Claims

1. The use of a POLQ inhibitor in the preparation of drugs for treating and / or preventing tumors, characterized in that, The tumor includes one or more of the following: HR gene deficiency, TLS polymerase deficiency, and POLQ overexpression.

2. The application as described in claim 1, characterized in that, It satisfies one or more of the following conditions: (1) When the tumor includes HR gene defects, it also includes Shieldin complex defects; preferably, the Shieldin complex defects include one or more of the following target gene expression decreases and / or protein expression abnormalities: RIF1, 53BP1, SHLD2, SHLD1, SHLD3 and REV7; (2) The HR gene defect is a gene defect that can cause cells to produce HRD, preferably including one or more of the following target genes with decreased expression and / or abnormal protein function: BRCA1, BRCA2, PALB2, RAD51, RAD51C, RAD51B, RAD51D, SLX4, FANCD2, FANCF and XRCC3; preferably, the HR gene defect includes one or more of the following target genes with decreased expression and / or abnormal protein function: BRCA1, BRCA2, RAD51C and XRCC3; (3) The TLS polymerase gene defect includes one or more of the following target genes with decreased expression and / or abnormal protein function: POLI, POLH, RAD18, REV1, REV3, POLK, POLL, PCNA and REV7; preferably, the TLS polymerase gene defect includes one or two of the following target genes with decreased expression and / or abnormal protein function: REV7 and REV3; (4) The POLQ inhibitors include compounds of Formula I or pharmaceutically acceptable salts thereof; in: m can be 0, 1, 2, or 3; R 1 For not replaced or by one or more R 1-1 Substituted 6-12 aryl groups or unsubstituted or substituted with one or more R groups 1-2 The substituted 5-12-membered heteroaryl group; wherein the heteroatom in the 5-12-membered heteroaryl group is one or more of N, O and S, and the number of heteroatoms is 1, 2 or 3; Each R 1-1 and R 1-2 Independently deuterium, halogen, cyano, amino, hydroxyl, unsubstituted, or modified by one or more R groups 1-1-1 Substituted C1-C6 alkyl groups or unsubstituted or substituted with one or more R groups 1-1-2 Substituted C1-C6 alkoxy groups; Each R 1-1-1 and R 1-1-2 It can be independently deuterium, hydroxyl, cyano or halogen; Each R 2 Independently for not replaced or by one or more R 2-1 Substituted C1-C6 alkyl, unsubstituted or substituted with one or more R 2-2 Substituted C1-C6 alkoxy groups; Each R 2-1 and R 2-2 It can be independently deuterium, hydroxyl, cyano or halogen; Ring B is unsubstituted or replaced by one or more R b-1 The substituted 5-12-membered fused-ring heteroaromatic ring; the heteroatoms in the 5-12-membered fused-ring heteroaromatic ring are selected from 1, 2 or 3 of N, O and S, and the number of heteroatoms is 1, 2, 3 or 4; the single heteroaromatic ring in the 5-12-membered fused-ring heteroaromatic ring is a 5-6-membered heteroaromatic ring; the number of rings in the 5-12-membered fused-ring heteroaromatic ring is 2; the heteroatoms in the 5-6-membered heteroaromatic ring are selected from 1, 2 or 3 of N, O and S, and the number of heteroatoms is 1, 2 or 3. Each R b-1 Independently deuterium, halogen, cyano, hydroxyl, unsubstituted, or modified by one or more R groups b-1-1 Substituted C1-C6 alkyl, unsubstituted or substituted with one or more R b-1-2 Substituted C1-C6 alkoxy groups; Each R b-1-1 and R b-1-2 Independently hydroxyl or halogen; R 3 For not replaced or by one or more R 3-1 Substituted 3-12 membered cycloalkyl, unsubstituted or substituted with one or more R 3-2 Substituted 4-12 membered heterocyclic groups, unsubstituted or substituted with one or more R groups 3-3 Substituted 4-12 membered cycloalkenyl groups, unsubstituted or with one or more R groups 3-4 Substituted 4-12 membered heterocyclic alkenyl groups, unsubstituted or with one or more R groups 3-5 Substituted 6-10 aryl groups, unsubstituted or substituted with one or more R groups 3-6 The substituted 5-10-membered heteroaryl group; the heteroatoms in the 4-12-membered heterocyclic group, 4-12-membered heterocyclic alkenyl group and 5-10-membered heteroaryl group are selected from 1, 2 or 3 of N, O and S, and the number of heteroatoms is 1, 2, 3 or 4; Each R 3-1 R 3-2 R 3-3 and R 3-4 Independently deuterium, hydroxyl, halogen, cyano, amino, unsubstituted, or modified by one or more R groups 3-2-2 Substituted C1-C6 alkyl or C1-C6 alkoxy; Each R 3-5 and R 3-6 Independently hydroxyl, halogen, cyano, amino, or unsubstituted or modified by one or more R groups 3-2-2 Substituted C1-C6 alkyl groups; Each R 3-2-2 It can be independently deuterium, hydroxyl, or halogen.

3. The application as described in claim 1 or 2, characterized in that, It satisfies one or more of the following conditions: (1) The HR gene defects include one or more of the following: BRCA2 defect, BRCA1 defect, XRCC3 defect, RAD51B defect, RAD51C defect, and FANCF defect; (2) The TLS polymerase defect includes one or more of REV7 defect, POLE2 defect and POLH defect; (3) The defects in the Shieldin complex include one or more of the 53BP1 defect and the SHLD2 defect; (4) The tumors include lung cancer, breast cancer, gastric cancer, ovarian cancer, prostate cancer, colon cancer, pancreatic cancer, uterine leiomyosarcoma, multiple myeloma, cervical cancer, urinary tract cancer, skin cancer or kidney cancer; Preferred cancers include lung cancer, breast cancer, stomach cancer, ovarian cancer, prostate cancer, colon cancer, pancreatic cancer, uterine leiomyosarcoma, or multiple myeloma; More preferably, the tumor includes HR gene-deficient lung cancer, HR gene-deficient breast cancer, HR gene-deficient gastric cancer, HR gene-deficient ovarian cancer, HR gene-deficient prostate cancer, HR gene-deficient colon cancer, HR gene-deficient pancreatic cancer, HR gene-deficient uterine leiomyosarcoma, HR gene-deficient multiple myeloma, TLS polymerase-deficient lung cancer, TLS polymerase-deficient breast cancer, TLS polymerase-deficient gastric cancer, TLS polymerase-deficient ovarian cancer, TLS polymerase-deficient prostate cancer, TLS polymerase-deficient colon cancer, TLS polymerase-deficient pancreatic cancer, TLS polymerase-deficient uterine leiomyosarcoma, or TLS polymerase-deficient multiple myeloma. (5) The compound represented by Formula I is any of the following schemes: Option 1: In the compound shown in Formula I, m is 1; R 1 For one or more R 1-2 The substituted 5-12-membered heteroaryl group; wherein the heteroatom in the 5-12-membered heteroaryl group is one or more of N, O and S, and the number of heteroatoms is 1, 2 or 3; Each R 1-2 Independently halogenated, by one or more R 1-1-1 Substituted C1-C6 alkyl or unsubstituted C1-C6 alkoxy; Each R 1-1-1 Halogens are independent of each other; Each R 2 Independently, it is an unsubstituted C1-C6 alkyl group; Ring B is unsubstituted or replaced by one or more R b-1 The substituted 5-12-membered fused-ring heteroaromatic ring; the heteroatoms in the 5-12-membered fused-ring heteroaromatic ring are selected from 1, 2 or 3 of N, O and S, and the number of heteroatoms is 1, 2, 3 or 4; the single heteroaromatic ring in the 5-12-membered fused-ring heteroaromatic ring is a 5-6-membered heteroaromatic ring; the number of rings in the 5-12-membered fused-ring heteroaromatic ring is 2; the heteroatoms in the 5-6-membered heteroaromatic ring are selected from 1, 2 or 3 of N, O and S, and the number of heteroatoms is 1, 2 or 3. Each R b-1 Halogens are independent of each other; R 3 For one or more R 3-2 Substituted 4-12 membered heterocyclic groups, unsubstituted or substituted with one or more R groups 3-3 Substituted 4-12 membered cycloalkenyl groups, unsubstituted or with one or more R groups 3-6 The substituted 5-10-membered heteroaryl group; the heteroatoms in the 4-12-membered heterocyclic group and the 5-10-membered heteroaryl group are selected from 1, 2 or 3 of N, O and S, and the number of heteroatoms is 1, 2, 3 or 4; Each R 3-2 and R 3-3 Independently hydroxyl or by one or more R 3-2-2 Substituted C1-C6 alkyl groups; Each R 3-6 Independently, it is a C1-C6 alkyl group; Each R 3-2-2 Independently, it is a hydroxyl group; Option 2: In the compound shown in Formula I, m is 1; R 1 For one or more R 1-2 The substituted 5-12-membered heteroaryl group; wherein the heteroatom in the 5-12-membered heteroaryl group is one or more of N, O and S, and the number of heteroatoms is 1, 2 or 3; Each R 1-2 Independently halogenated, by one or more R 1-1-1 Substituted C1-C6 alkyl or unsubstituted C1-C6 alkoxy; Each R 1-1-1 Halogens are independent of each other; Each R 2 Independently, it is an unsubstituted C1-C6 alkyl group; Ring B is unsubstituted or replaced by one or more R b-1 The substituted 5-12-membered fused-ring heteroaromatic ring; the heteroatoms in the 5-12-membered fused-ring heteroaromatic ring are selected from 1, 2 or 3 of N, O and S, and the number of heteroatoms is 1, 2, 3 or 4; the single heteroaromatic ring in the 5-12-membered fused-ring heteroaromatic ring is a 5-6-membered heteroaromatic ring; the number of rings in the 5-12-membered fused-ring heteroaromatic ring is 2; the heteroatoms in the 5-6-membered heteroaromatic ring are selected from 1, 2 or 3 of N, O and S, and the number of heteroatoms is 1, 2 or 3. Each R b-1 Halogens are independent of each other; R 3 For not replaced or by one or more R 3-6 The substituted 5-10-membered heteroaryl group; the heteroatoms in the 4-12-membered heterocyclic group and the 5-10-membered heteroaryl group are selected from 1, 2 or 3 of N, O and S, and the number of heteroatoms is 1, 2, 3 or 4; Each R 3-6 Independently, it is a C1-C6 alkyl group; Each R 3-2-2 Independently, it is a hydroxyl group.

4. The application as described in claim 1 or 2, characterized in that, It satisfies one or more of the following conditions: (1) The tumor is BRCA2-deficient colon cancer, BRCA2-deficient primary colon cancer, BRCA2-deficient prostate cancer, RAD51B-deficient, POLE2-deficient and POLH-deficient metastatic prostate cancer, BRCA2-deficient ovarian cancer, BRCA2-deficient uterine leiomyosarcoma, BRCA-deficient breast cancer, XRCC3-deficient colon cancer, RAD51C-deficient colon cancer, 53BP1-deficient breast cancer, BRCA-deficient and SHLD2-deficient breast cancer, 53BP1-deficient and SHLD2-deficient breast cancer, BRCA-deficient and SHLD2-deficient breast cancer, FANCF-deficient ovarian cancer, POLQ-overexpressing gastric cancer, BRCA2-deficient and FANCD2-deficient multiple myeloma or TLS polymerase-deficient colon cancer. Preferably, the RAD51B-deficient, POLE2-deficient and POLH-deficient metastatic prostate cancer contains RAD51B, POLB, POLE2, POLH, MSH6 and RNASEH2B deficiencies. (2)R 1 In this context, the 6-12 aryl group is a 6-10 aryl group, and can also be phenyl or naphthyl, such as phenyl; (3)R 1 In this context, the 5-12-membered heteroaryl group is a 5-10-membered heteroaryl group, and the heteroatom in the 5-10-membered heteroaryl group can be N and / or O, and the number of heteroatoms can be 1 or 2; the 5-12-membered heteroaryl group is preferably a 5-6-membered heteroaryl group; more preferably... For example (4) Each R 1-1 In this context, the halogen is fluorine, chlorine, bromine, or iodine, such as fluorine; (5) Each R 1-1 In this context, the C1-C6 alkyl group is a C1-C4 alkyl group, and may also be methyl, ethyl, n-propyl, isopropyl, n-butyl, isobutyl, primary butyl, secondary butyl, or tert-butyl. (6) Each R 1-1 In this context, the C1-C6 alkoxy group is a C1-C4 alkoxy group, and may also be a methoxy group, an ethoxy group, a propoxy group, an isopropoxy group, a butoxy group, an isobutoxy group, a primary butoxy group, a secondary butoxy group, or a tert-butoxy group, such as a methoxy group. (7) Each R 1-2 In this context, the halogen is fluorine, chlorine, bromine, or iodine, such as fluorine or chlorine; (8) Each R 1-2 In this context, the C1-C6 alkyl group is a C1-C4 alkyl group, and may be methyl, ethyl, n-propyl, isopropyl, n-butyl, isobutyl, primary butyl, secondary butyl, or tert-butyl, with methyl being preferred; (9) Each R 1-2 In this context, the C1-C6 alkoxy group is a C1-C4 alkoxy group, and may also be a methoxy group, an ethoxy group, a propoxy group, an isopropoxy group, a butoxy group, an isobutoxy group, a primary butoxy group, a secondary butoxy group, or a tert-butoxy group, such as a methoxy group. (10) Each R 1-1-1 and R 1-1-2 In this context, the halogen is fluorine, chlorine, bromine, or iodine, such as fluorine; (11) Each R 2 In this context, the C1-C6 alkyl group is a C1-C4 alkyl group, and may be methyl, ethyl, n-propyl, isopropyl, n-butyl, isobutyl, primary butyl, secondary butyl, or tert-butyl, for example, methyl; (12) Each R 2 In this context, the C1-C6 alkoxy group is a C1-C4 alkoxy group, and may also be a methoxy group, an ethoxy group, a propoxy group, an isopropoxy group, a butoxy group, an isobutoxy group, a tert-butoxy group, a tert-butoxy group, or a methoxy group. (13) Each R 2-1 and R 2-2 In this context, the halogen is fluorine, chlorine, bromine, or iodine; (14) In ring B, the number of heteroatoms in the 5-12 membered fused-ring heteroaromatic ring can be 1, 2, 3 or 4; the 5-20 membered fused-ring heteroaromatic ring is preferably a five-membered fused-ring heteroaromatic ring, more preferably a six-membered fused-ring heteroaromatic ring. (15) In ring B, the 5-6 membered heteroaromatic ring is a 5-membered heteroaromatic ring, which may have 2 heteroatoms and may be one or two types of heteroatoms, namely N, S or O. The 5-6 membered heteroaromatic ring is preferably... (16) Each R b-1 In this context, the C1-C6 alkyl group is a C1-C4 alkyl group, and may also be methyl, ethyl, n-propyl, isopropyl, n-butyl, isobutyl, primary butyl, secondary butyl, or tert-butyl. (17) Each R b-1 In this context, the C1-C6 alkoxy group is a C1-C4 alkoxy group, and may also be a methoxy group, an ethoxy group, a propoxy group, an isopropoxy group, a butoxy group, an isobutoxy group, a tert-butoxy group, a tert-butoxy group, or a methoxy group. (18) Each R b-1-1 and R b-1-2 In this context, the halogen is fluorine, chlorine, bromine, or iodine; (19) In R3, the 3-12 membered cycloalkyl group is a monocyclic cycloalkyl group, such as a 3-6 membered cycloalkyl group, or cyclopropyl, cyclobutyl, cyclopentyl or cyclohexyl; (20) In R3, the ring in the 4-12 membered heterocyclic group can be a monocyclic heterocyclic group, the number of heteroatoms in the 4-12 membered heterocyclic group can be 1 or 2, and the type of heteroatom can be N; preferably (21) In R3, the 4-12 membered cycloalkenyl group is a 4-6 membered cycloalkenyl group (e.g., a monocyclic cycloalkenyl group), and the number of carbon-carbon double bonds can be 1, preferably 1. (22) In R3, the 4-12 membered heterocyclic alkenyl group is a 4-6 membered heterocyclic alkenyl group (e.g., a monocyclic heterocyclic alkenyl group), the number of heteroatoms can be 1, and the type of heteroatoms can be 1, 2 or 3 of N, O or S; (23) In R3, the 6-10 aryl group can be aryl or naphthyl, preferably aryl; (24) In R3, the 5-10 membered heteroaryl group is a 5-6 membered heteroaryl group, the heteroatom can be N, and the number of heteroatoms can be 1, 2 or 3, for example, (25) Each R 3-1 R 3-2 R 3-3 and R 3-4 In this context, the halogen is fluorine, chlorine, bromine, or iodine; (26) Each R 3-1 R 3-2 R 3-3 and R 3-4 In this context, the C1-C6 alkyl group is a C1-C4 alkyl group, and may also be methyl, ethyl, n-propyl, isopropyl, n-butyl, isobutyl, primary butyl, secondary butyl, or tert-butyl. (27) Each R 3-1 R 3-2 R 3-3 and R 3-4 In this context, the C1-C6 alkoxy group is a C1-C4 alkoxy group, and may also be a methoxy group, an ethoxy group, a propoxy group, an isopropoxy group, a butoxy group, an isobutoxy group, a tert-butoxy group, a tert-butoxy group, or a methoxy group. (28) Each R 3-5 and R 3-6 In this context, the halogen is fluorine, chlorine, bromine, or iodine; (29) Each R 3-5 and R 3-6 In this context, the C1-C6 alkyl group is a C1-C4 alkyl group, and may also be methyl, ethyl, n-propyl, isopropyl, n-butyl, isobutyl, primary butyl, secondary butyl, or tert-butyl. (30) Each R 3-2-2 In this context, the halogen is fluorine, chlorine, bromine, or iodine; (31) Each R b-1 In this context, the halogen is fluorine, chlorine, bromine, or iodine, such as fluorine.

5. The application as described in claim 1 or 2, characterized in that, It satisfies one or more of the following conditions: (1) m is 1; (2)R 1 For one or more R 1-2 The substituted 5-12-membered heteroaryl group; wherein the heteroatom in the 5-12-membered heteroaryl group is one or more of N, O and S, and the number of heteroatoms is 1, 2 or 3; (3) Each R 1-2 Independently halogenated, by one or more R 1-1-1 Substituted C1-C6 alkyl or unsubstituted C1-C6 alkoxy; (4) Each R 1-1-1 Halogens are independent of each other; (5) Each R 2 Independently, it is an unsubstituted C1-C6 alkyl group; (6) Ring B is unsubstituted or is replaced by one or more R b-1 The substituted 5-12-membered fused-ring heteroaromatic ring; the heteroatoms in the 5-12-membered fused-ring heteroaromatic ring are selected from 1, 2 or 3 of N, O and S, and the number of heteroatoms is 1, 2, 3 or 4; the single heteroaromatic ring in the 5-12-membered fused-ring heteroaromatic ring is a 5-6-membered heteroaromatic ring; the number of rings in the 5-12-membered fused-ring heteroaromatic ring is 2; the heteroatoms in the 5-6-membered heteroaromatic ring are selected from 1, 2 or 3 of N, O and S, and the number of heteroatoms is 1, 2 or 3. (7) Each R b-1 Halogens are independent of each other; (8)R 3 For one or more R 3-2 Substituted 4-12 membered heterocyclic groups, unsubstituted or substituted with one or more R groups 3-3 Substituted 4-12 membered cycloalkenyl groups, unsubstituted or with one or more R groups 3-6 The substituted 5-10-membered heteroaryl group; the heteroatoms in the 4-12-membered heterocyclic group and the 5-10-membered heteroaryl group are selected from one, two, or three of N, O, and S, and the number of heteroatoms is 1, 2, 3, or 4. Preferably, R 3 For not replaced or by one or more R 3-6 The substituted 5-10-membered heteroaryl group; the heteroatoms in the 4-12-membered heterocyclic group and the 5-10-membered heteroaryl group are selected from 1, 2 or 3 of N, O and S, and the number of heteroatoms is 1, 2, 3 or 4; (9) Each R 3-2 and R 3-3 Independently hydroxyl or by one or more R 3-2-2 Substituted C1-C6 alkyl groups; (10) Each R 3-6 Independently, it is a C1-C6 alkyl group; (11) Each R 3-2-2 Independently, it is a hydroxyl group.

6. The application as described in claim 1 or 2, characterized in that, The compound shown in Formula I is the same as the compound shown in Formula I-1; Wherein, Z1 and Z2 are each independently CH, CF or N; preferably, Z1 and Z2 are not both N; m、R 1 R 2 R b-1 and R 3 Each independently as described in claim 1 or 2; Alternatively, the tumor may be BRCA2-deficient colon cancer, BRCA2-deficient primary colon cancer, BRCA2-deficient prostate cancer, RAD51B-deficient, POLE2-deficient, and POLH-deficient metastatic prostate cancer, BRCA2-deficient ovarian cancer, BRCA2-deficient uterine leiomyosarcoma, BRCA1-deficient breast cancer, XRCC3-deficient colon cancer, RAD51C-deficient colon cancer, 53BP1-deficient breast cancer, BRCA1-deficient and SHLD2-deficient breast cancer, BRCA2-deficient and SHLD2-deficient breast cancer, 53BP1-deficient and SHLD2-deficient breast cancer, BRCA1-deficient and SHLD2-deficient breast cancer, FANCF-deficient ovarian cancer, POLQ-overexpressing gastric cancer, BRCA2-deficient and FANCD2-deficient multiple myeloma, or REV7-deficient colon cancer.

7. The application as described in claim 1 or 2, characterized in that, It satisfies one or more of the following conditions: (1)R 2 It is methyl; (2)R 1 for (3) Ring B is (4)R 3 for 8. The application as described in claim 1 or 2, characterized in that, The compound represented by Formula I is any of the following compounds:

9. A pharmaceutical composition, characterized in that, The pharmaceutical composition comprises the POLQ inhibitor as described in any one of claims 1-8 and pharmaceutical excipients; Preferably, the pharmaceutical composition satisfies one or more of the following conditions: (1) The pharmaceutical composition is a solid, preferably a tablet; (2) The pharmaceutical composition is the drug according to any one of claims 1-8; (3) In the pharmaceutical composition, the mass ratio of the POLQ inhibitor to the pharmaceutical composition is 0.01%-95%, for example 1%, 10%, 30%, 40%, 50%, 60%, 70%, 80% or 90%; (4) The pharmaceutical composition includes gastrointestinal and non-gastrointestinal dosage forms, such as capsules, tablets, pills, granules, powders or oral liquids, preferably tablets; (5) The drug is administered by injection or oral administration, such as intravenous injection, subcutaneous injection, peritoneal injection or intramuscular injection, preferably oral administration.

10. The use of a POLQ inhibitor in in vitro inhibition of tumor cells, wherein the tumor is as described in any one of claims 1-8; and the POLQ inhibitor is as described in any one of claims 1-8.

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