Cancer combination therapy using chk inhibitors

Abstract

The present application discloses the use of checkpoint kinase (CHK) inhibitors in combination with i) an immunotherapeutic agent, or a therapeutic agent targeting a cancer promoting / sustaining molecule, and optionally ii) a chemotherapeutic agent, e.g. gemcitabine, in the treatment of cancer.

Description

[0001] Related applications and citations are included.

[0002] This application claims priority to U.S. Provisional Application US 62 / 957,806, filed January 7, 2020.

[0003] All documents cited in or during the examination of the foregoing application (“Application References”), all documents cited or mentioned in this application (including, but not limited to, all documents, patents, and published patent applications cited herein) (“In-Application References”), all documents cited or mentioned in the In-Application References, and manufacturer’s manuals, instructions, product specifications, and product pages of any product mentioned in this application or any In-Application References are incorporated herein by reference and may be used in practicing this invention. More specifically, all referenced documents are incorporated herein by reference as if each document were specifically and individually incorporated by reference. Any Genbank sequences mentioned in this application that are those with the earliest valid filing date of this application are incorporated herein by reference. Technical Field

[0004] This application relates to the combined use of checkpoint kinase (CHK) inhibitors with immunotherapeutic agents or therapeutic agents targeting cancer-promoting / maintaining molecules, and optionally additional chemotherapeutic agents in cancer treatment. Background Technology

[0005] Cancer is the second leading cause of death worldwide, and the average 5-year survival rate for adult patients is quite low, ranging from 14% to 56% in South America.

[0006] Current traditional cancer treatment options include surgery, radiotherapy, chemotherapy, hormone therapy, and immunotherapy. Radiotherapy and chemotherapy remain the preferred treatments for many cancers due to their effectiveness in destroying cancer cells. However, these two commonly used therapies are toxic to patients because they often non-selectively destroy healthy cells, suppressing the immune system. Cancer immunotherapy involves the activation of the immune system and the expansion of the immune response, and was named "Breakthrough of the Year" by Science in 2013. Compared to other standard therapies, it offers lower toxicity and enjoys unprecedented success. However, a large proportion of patients do not respond to immunotherapy as a monotherapy, likely because cancer cells have developed mechanisms to evade immune surveillance and induce immune tolerance. Targeted therapy is a medical treatment that partially overlaps with chemotherapy and immunotherapy. Chemotherapy agents or biologics interfere with specific target molecules required for cancer formation and tumor growth, or disrupt the tissue environment that promotes cancer growth and survival. Compared to traditional cytotoxic chemotherapy, it can sometimes cause less damage to healthy cells.

[0007] Combination therapy has emerged as a promising new cancer treatment strategy because the combination of two or more therapies can target more than one cancer-inducing or cancer-maintaining pathway, thereby increasing the chances of killing cancer cells and minimizing drug resistance.

[0008] For example, low-dose gemcitabine (LDG) combined with some small molecule inhibitors of checkpoint kinase 1 (CHK1) is currently being tested in clinical trials. Furthermore, based on promising clinical results, the FDA has approved two chemotherapy regimens, carboplatin and etoposide, in combination with the immunotherapy atezolizumab for first-line treatment of extensive-stage small cell lung cancer (SCLC). Researchers have recognized the need to better understand the extent to which chemotherapy can enhance immune checkpoint blockade (ICB) responses and have begun investigating new approaches beyond cytotoxic chemotherapy, representing more optimized treatment options for immunotherapy combinations.

[0009] Checkpoint kinase

[0010] In eukaryotic cells, the cell cycle is regulated by checkpoints, which control the transition from one phase to another. The transition from S phase to G2 / M phase is regulated by checkpoint kinase 1 (CHK1) and checkpoint kinase 2 (CHK2) (in a lower ratio).

[0011] Checkpoint kinases are regulators of DNA replication and DNA damage response (DDR). Specifically, studies have shown that CHK1 is a major regulator of replication stress (RS), characterized by slowing or arrest of replication forks, primarily caused by DNA damage, replication-transcriptional collisions, and depletion of the deoxyribonucleoside triphosphate (dNTP) pool. RS can lead to genomic instability and is therefore associated with cancer cells. CHK1, in response to RS, temporarily halts the cell cycle and regulates the activation of replication initiation sites, thereby preventing excessive DNA damage and increasing the overall survival fitness of tumor cells (Kotsantis P, Petermann E, Boulton SJ. Mechanisms of oncogene-induced replication stress: jigsaw falling into place. Cancer Discov. 2018.8: 537-555).

[0012] High expression of CHK1 and other DDR proteins has been observed in some cancers, such as SCLC (Byers LA et al, Proteomic profiling identifies dysregulated pathways in small cell lung cancer and novel therapeutic targets including PARP1. Cancer Discov. 2012. 2: 798-811; Sen T et al, CHK1 inhibition in small cell lung cancer produces single-agent activity in biomarker-defined disease subsets and combination activity with cisplatin or olaparib. Cancer Res. 2017. 77: 3870-3884), and inhibition of CHK1 expression can increase the sensitivity of cancer cells to DNA damage therapy and reverse drug resistance or tolerance in cancer cells.

[0013] Combination therapy using CHK inhibitors

[0014] Despite their promising anti-tumor effects, CHK1 inhibitors may adversely affect DNA damage repair in healthy cells and may suppress the immune system to some extent. One possible way to reduce harm to the body while maintaining anti-tumor efficacy is to find combination regimens where CHK inhibitors synergize with another agent, thus providing better anti-tumor therapy using lower doses of CHK inhibitors.

[0015] Candidate drugs that can be administered in combination with CHK inhibitors can be immunotherapeutic agents or therapeutic agents that target cancer-promoting / maintaining molecules, such as PD-1 / PD-L1 inhibitors, CTLA-4 inhibitors, or ErbB2 inhibitors. CHK inhibitors can also be used in combination with chemotherapeutic agents such as gemcitabine.

[0016] Gemcitabine is known to cause dNTP depletion and replication fork arrest, even at below therapeutic concentrations. The combination of LDG and CHK1 inhibitors presents a unique approach to combining chemotherapy with targeted therapy. Compared to inducing cancer cell death using standard-dose chemotherapy, LDG can leverage its RS-inducing properties to increase cancer cell dependence on CHK1, thereby enhancing the intrinsic cytotoxic and immunostimulatory activity of CHK1 inhibitors.

[0017] Furthermore, it has been shown that CHK1 and DDR inhibition synergize with PD-L1 antibodies in vivo (Sen T et al, ibid.; Harding SM et al. Mitotic progression following DNA damage enables pattern recognition within micronuclei. Nature. 2017. 548: 466-470; Jiao S et al. PARP inhibitor upregulates PD-L1 expression and enhances cancer-associated immunosuppression. Clin Cancer Res. 2017. 23: 3711-3720). Recently, Sen et al. found that the combination therapy of SRA737 (an oral small molecule inhibitor of CHK1) and PD-L1 antibodies induced antitumor responses in multiple cancer models, and in the SCLC model, the combination of LDG with SRA737 and PD-L1 antibody / PD-1 antibody significantly increased antitumor CD8+. + Cytotoxic T cells, dendritic cells, and M1 macrophage populations (Triparna Sen et al., SRA737, and Low-Dose Gemcitabine Enhances the Effect of Programmed DeathLig and 1 Blockade by Modulating the Immune Microenvironment in SCLC. Journal of Thoracic Oncology. 2019.14(12):2152-2163).

[0018] Despite promising data from some combination therapies, it is important to note that not all therapies or specific drugs can be combined, and even fewer combination therapies work synergistically. One therapeutic agent may alter the pharmacology of a second therapeutic agent, thereby diminishing its antitumor activity. For example, in humans, one therapeutic agent may alter the conformation of a second therapeutic agent or inhibit its metabolism, leading to cumulative toxicity. For instance, in a pooled analysis of 14 phase I-III studies, 64% of patients receiving multiple doses of ipilimumab (CTLA-4 antibody) experienced immune-related adverse effects, and the incidence of adverse events was 93% in patients receiving dual therapy with PD-1 mAb and ipilimumab (Wolchok, JD, et al., Nivolumab plus ipilimumab in advanced melanoma. N. Engl. J. Med., 2013. 369: 122-33). Therefore, before concluding whether a combination regimen is appropriate or whether it can achieve maximum efficacy with minimal toxicity, a comprehensive study of the interactions between two or more antitumor agents in a combination regimen is necessary. Summary of the Invention

[0019] Surprisingly, the current inventors have discovered that compounds disclosed as CHK inhibitors in WO2009 / 092278 and WO2011 / 035077 can synergize with PD-1 antibodies to provide enhanced anti-tumor effects without increased toxicity, suggesting that these CHK inhibitors can be used in combination with immunotherapeutic agents or therapeutic agents targeting cancer-promoting / maintaining molecules for cancer treatment.

[0020] The inventors have also surprisingly discovered that this checkpoint kinase inhibitor can synergize with PD-1 antibodies and chemotherapeutic agents such as gemcitabine to provide further enhanced antitumor effects without increased toxicity, suggesting that the inhibitor can be combined with i) immunotherapeutic agents, or therapeutic agents targeting cancer-promoting / maintaining molecules, and ii) chemotherapeutic agents in cancer treatment.

[0021] Therefore, in a first aspect, this application discloses a method for treating cancer, comprising administering a therapeutically effective amount of a compound of formula I, or a pharmaceutically acceptable salt thereof, and an immunotherapeutic agent, or a therapeutic agent targeting cancer-promoting / maintaining molecules, to a subject in need.

[0022]

[0023] Where Y is NH, O, S or CH2;

[0024] R 1 Selected from:

[0025]

[0026] Where X is CH2, NH, S or O.

[0027] R 8 =-H, -NH2, -OH, -N(R) 4 R 5 ), -C(R 4 R 5 ) 1-7 NR 6 R 7 -C(R) 4 R 5 ) 1-7 OR 6 、or -N(R 4 )NR 5 R 6 , where R 4 R 5 R 6 and R 7 Independently H, C1-C6 alkyl, C3-C8 cycloalkyl with or without a core heteroatom such as O, S, and N; optionally substituted aryl, or optionally substituted heteroaryl.

[0028] R 9 R 10 R 11 R 12 and R 13 Independently H, C1-C6 alkyl, C3-C8 cycloalkyl with or without a core heteroatom such as O, S, and N; optionally substituted aryl, or optionally substituted heteroaryl.

[0029] R 2 Selected from H, OH, NH2, OR 14 NR 14 R 15 Alkyl, aryl, heteroaryl, cycloalkyl, aralkyl, heterocyclic, heterocyclic, alkenyl, and ynyl.

[0030] Where R 14 and R 15 Independently H, C1-C6 alkyl, C3-C8 cycloalkyl with or without a core heteroatom such as O, S, and N; optionally substituted aryl, or optionally substituted heteroaryl, and

[0031] R 3 It is selected from H, alkyl, aryl, heteroaryl, cycloalkyl, aralkyl, heterocyclic, heterocyclic, alkenyl, and ynyl.

[0032] In one implementation, R 3 Selected from

[0033]

[0034] Where R 16 R 17 and R 18 Independently H; F, Cl, Br, I; C1-C8 alkyl; substituted or unsubstituted C3-C8 cycloalkyl, wherein the substituent is selected from C1-C8 alkyl, C3-C8 cycloalkyl, aryl, and heteroaryl; -OR 19 ;-SR 19 ;-NR 19 R 20 ;-S(O)R 19 ;-S(O)2R 19 ;-S(O)2NR 19 R 20 ;-C(O)NR 19 R 20 ;-N(R 19 )C(O)R 20 ;-N(R 19 )S(O)2R 20 ;-N(R 19 )C(O)N(R 20 R 21 );N(R 19 )C(O)OR 20 ; optionally substituted aryl, optionally substituted heteroaryl, optionally substituted aralkyl, optionally substituted heterocyclic, optionally substituted heterocyclic; optionally substituted alkenyl, or optionally substituted alkynyl;

[0035] Where R 19 R 20 R 21 Independently, it is H, C1-C8 alkyl, C3-C8 cycloalkyl, optionally substituted aryl, optionally substituted alkylaryl, or optionally substituted heteroaryl.

[0036] Or R 16 R 17 and R 21 Independently, it is part of a fused ring containing 0-3 heteroatoms selected from N, O, and S.

[0037] In one embodiment, Y is NH. In one embodiment, Y is O. In one embodiment, Y is S.

[0038] In one embodiment, the compound of formula I is selected from...

[0039] 2-(4-fluorophenyl)-4-(3-piperidineamino)-thieno[2,3-d]pyridazine-7-carboxylic acid amide,

[0040] 2-(4-chlorophenyl)-4-(3-piperidineamino)-thieno[2,3-d]pyridazine-7-carboxylic acid amide,

[0041] 2-(4-bromophenyl)-4-(3-piperidineamino)-thieno[2,3-d]pyridazine-7-carboxylic acid amide,

[0042] 2-(4-fluorophenyl)-4-(3-tetrahydropyran-amino)-thieno[2,3-d]pyridazine-7-carboxylic acid amide,

[0043] 2-(4-chlorophenyl)-4-(3-tetrahydropyran-amino)-thieno[2,3-d]pyridazine-7-carboxylic acid amide,

[0044] 2-(4-bromophenyl)-4-(3-tetrahydropyran-amino)-thieno[2,3-d]pyridazine-7-carboxylic acid amide,

[0045] 2-(4-fluorophenyl)-4-(3-tetrahydrothiaran-amino)-thieno[2,3-d]pyridazine-7-carboxylic acid amide,

[0046] 2-(4-Chlorophenyl)-4-(3-Tetrahydrothiaran-amino)-thieno[2,3-d]pyridazine-7-carboxylic acid amide,

[0047] 2-(4-bromophenyl)-4-(3-tetrahydrothiaran-amino)-thieno[2,3-d]pyridazine-7-carboxylic acid amide,

[0048] 2-(4-chlorophenyl)-4-(2-piperidin-amino)-thieno[2,3-d]pyridazine-7-carboxylic acid amide,

[0049] 2-(4-chlorophenyl)-4-(2-piperidin-amino)-thieno[2,3-d]pyridazine-7-carboxylic acid amide,

[0050] 2-(4-chlorophenyl)-4-(S-3-piperidin-amino)-thieno[2,3-d]pyridazine-7-carboxylic acid amide,

[0051] 2-(4-chlorophenyl)-4-(R-3-piperidin-amino)-thieno[2,3-d]pyridazine-7-carboxylic acid amide,

[0052] 2-(4-chlorophenyl)-4-(3-piperidin-amino)-thieno[2,3-d]pyridazine-7-N-methylcarboxylate,

[0053] 2-(4-Chlorophenyl)-4-(3-piperidin-amino)-thieno[2,3-d]pyridazine-7-N,N-dimethylcarboxylate

[0054] 2-(4-chlorophenyl)-4-(3-tetrahydrothiaran-amino)-thieno[2,3-d]pyridazine-7-N-methylcarboxylate,

[0055] 2-(4-Chlorophenyl)-4-(3-Tetrahydrothiaran-amino)-thieno[2,3-d]pyridazine-7-N,N-dimethylcarboxylate,

[0056] 2-(4-Chlorophenyl)-4-(3-Tetrahydropyran-amino)-thieno[2,3-d]pyridazine-7-N-methylcarboxylate,

[0057] 2-(4-Chlorophenyl)-4-(3-Tetrahydropyran-amino)-thieno[2,3-d]pyridazine-7-N,N-dimethylcarboxylate,

[0058] 2-(4-fluorophenyl)-4-(3-tetrahydropyrrole-amino)-thieno[2,3-d]pyridazine-7-carboxylic acid amide,

[0059] 2-(4-chlorophenyl)-4-(3-tetrahydropyrrole-amino)-thieno[2,3-d]pyridazine-7-carboxylic acid amide,

[0060] 2-(4-bromophenyl)-4-(3-tetrahydropyrrole-amino)-thieno[2,3-d]pyridazine-7-carboxylic acid amide,

[0061] 2-(4-fluorophenyl)-4-(3-tetrahydrofuran-amino)-thieno[2,3-d]pyridazine-7-carboxylic acid amide,

[0062] 2-(4-chlorophenyl)-4-(3-tetrahydrofuran-amino)-thieno[2,3-d]pyridazine-7-carboxylic acid amide,

[0063] 2-(4-bromophenyl)-4-(3-tetrahydrofuran-amino)-thieno[2,3-d]pyridazine-7-carboxylic acid amide,

[0064] 2-(4-fluorophenyl)-4-(3-tetrahydrothiophene-amino)-thiopheno[2,3-d]pyridazine-7-carboxylic acid amide,

[0065] 2-(4-chlorophenyl)-4-(3-tetrahydrothiophene-amino)-thiopheno[2,3-d]pyridazine-7-carboxylic acid amide,

[0066] 2-(4-bromophenyl)-4-(3-tetrahydrothiophene-amino)-thiopheno[2,3-d]pyridazine-7-carboxylic acid amide,

[0067] 2-(4-Chlorophenyl)-4-(2-Tetrahydropyrrole-amino)-thieno[2,3-d]pyridazine-7-carboxylic acid amide,

[0068] 2-(4-chlorophenyl)-4-(S-3-tetrahydropyrrole-amino)-thieno[2,3-d]pyridazine-7-carboxylic acid amide,

[0069] 2-(4-chlorophenyl)-4-(R-3-tetrahydropyrrole-amino)-thieno[2,3-d]pyridazine-7-carboxylic acid amide,

[0070] 2-(4-Chlorophenyl)-4-(3-Tetrahydropyrrole-amino)-thieno[2,3-d]pyridazine-7-N-methylcarboxylate,

[0071] 2-(4-Chlorophenyl)-4-(3-Tetrahydropyrrole-amino)-thieno[2,3-d]pyridazine-7-N,N-dimethylcarboxylate,

[0072] 2-(4-chlorophenyl)-4-(3-tetrahydrothiophene-amino)-thiopheno[2,3-d]pyridazine-7-N-methylcarboxylate,

[0073] 2-(4-Chlorophenyl)-4-(3-Tetrahydrothiophene-amino)-thieno[2,3-d]pyridazine-7-N,N-dimethylcarboxylate,

[0074] 2-(4-chlorophenyl)-4-(3-tetrahydrofuran-amino)-thieno[2,3-d]pyridazine-7-N-methylcarboxylate,

[0075] 2-(4-Chlorophenyl)-4-(3-tetrahydrofuran-amino)-thieno[2,3-d]pyridazine-7-N,N-dimethylcarboxylate

[0076] 2-(4-fluorophenyl)-4-(3-pyridin-amino)-thieno[2,3-d]pyridazine-7-carboxylic acid amide,

[0077] 2-(4-chlorophenyl)-4-(3-pyridin-amino)-thieno[2,3-d]pyridazine-7-carboxylic acid amide,

[0078] 2-(4-bromophenyl)-4-(3-pyridine-amino)-thieno[2,3-d]pyridazine-7-carboxylic acid amide,

[0079] 2-(4-fluorophenyl)-4-(3-α-pyran-amino)-thieno[2,3-d]pyridazine-7-carboxylic acid amide,

[0080] 2-(4-chlorophenyl)-4-(3-α-pyran-amino)-thieno[2,3-d]pyridazine-7-carboxylic acid amide,

[0081] 2-(4-bromophenyl)-4-(3-α-pyran-amino)-thieno[2,3-d]pyridazine-7-carboxylic acid amide,

[0082] 2-(4-fluorophenyl)-4-(3-α-thiaran-amino)-thieno[2,3-d]pyridazine-7-carboxylic acid amide,

[0083] 2-(4-chlorophenyl)-4-(3-α-thiaran-amino)-thieno[2,3-d]pyridazine-7-carboxylic acid amide,

[0084] 2-(4-bromophenyl)-4-(3-α-thiaran-amino)-thieno[2,3-d]pyridazine-7-carboxylic acid amide,

[0085] 2-(4-chlorophenyl)-4-(2-pyridin-amino)-thieno[2,3-d]pyridazine-7-carboxylic acid amide,

[0086] 2-(4-chlorophenyl)-4-(4-pyridin-amino)-thieno[2,3-d]pyridazine-7-carboxylic acid amide,

[0087] 2-(4-chlorophenyl)-4-(S-3-pyridin-amino)-thieno[2,3-d]pyridazine-7-carboxylic acid amide,

[0088] 2-(4-chlorophenyl)-4-(R-3-pyridine-amino)-thieno[2,3-d]pyridazine-7-carboxylic acid amide,

[0089] 2-(4-chlorophenyl)-4-(3-pyridin-amino)-thieno[2,3-d]pyridazine-7-N-methylcarboxylate,

[0090] 2-(4-chlorophenyl)-4-(3-pyridin-amino)-thieno[2,3-d]pyridazine-7-N,N-dimethylcarboxylate

[0091] 2-(4-chlorophenyl)-4-(3-thiaran-amino)-thieno[2,3-d]pyridazine-7-N-methylcarboxylate,

[0092] 2-(4-Chlorophenyl)-4-(3-thiaran-amino)-thieno[2,3-d]pyridazine-7-N,N-dimethylcarboxylate,

[0093] 2-(4-chlorophenyl)-4-(3-pyran-amino)-thieno[2,3-d]pyridazine-7-N-methylcarboxylate,

[0094] 2-(4-chlorophenyl)-4-(3-pyran-amino)-thieno[2,3-d]pyridazine-7-N,N-dimethylcarboxylate,

[0095] 2-(4-fluorophenyl)-4-(3-pyrrolo-amino)-thieno[2,3-d]pyridazine-7-carboxylic acid amide,

[0096] 2-(4-chlorophenyl)-4-(3-pyrrolo-amino)-thieno[2,3-d]pyridazine-7-carboxylic acid amide,

[0097] 2-(4-bromophenyl)-4-(3-pyrrolo-amino)-thieno[2,3-d]pyridazine-7-carboxylic acid amide,

[0098] 2-(4-fluorophenyl)-4-(3-furan-amino)-thieno[2,3-d]pyridazine-7-carboxylic acid amide,

[0099] 2-(4-chlorophenyl)-4-(3-furan-amino)-thieno[2,3-d]pyridazine-7-carboxylic acid amide,

[0100] 2-(4-bromophenyl)-4-(3-furan-amino)-thieno[2,3-d]pyridazine-7-carboxylic acid amide,

[0101] 2-(4-fluorophenyl)-4-(3-thiophene-amino)-thiopheno[2,3-d]pyridazine-7-carboxylic acid amide,

[0102] 2-(4-chlorophenyl)-4-(3-thiophene-amino)-thiopheno[2,3-d]pyridazine-7-carboxylic acid amide,

[0103] 2-(4-bromophenyl)-4-(3-thiophene-amino)-thiopheno[2,3-d]pyridazine-7-carboxylic acid amide,

[0104] 2-(4-chlorophenyl)-4-(2-pyrrolo-amino)-thieno[2,3-d]pyridazine-7-carboxylic acid amide,

[0105] 2-(4-chlorophenyl)-4-(S-3-pyrrolo-amino)-thieno[2,3-d]pyridazine-7-carboxylic acid amide,

[0106] 2-(4-chlorophenyl)-4-(R-3-pyrrolo-amino)-thieno[2,3-d]pyridazine-7-carboxylic acid amide,

[0107] 2-(4-chlorophenyl)-4-(3-pyrrolo-amino)-thieno[2,3-d]pyridazine-7-N-methylcarboxylate,

[0108] 2-(4-Chlorophenyl)-4-(3-pyrrolo-amino)-thieno[2,3-d]pyridazine-7-N,N-dimethylcarboxylate,

[0109] 2-(4-chlorophenyl)-4-(3-thiophene-amino)-thiopheno[2,3-d]pyridazine-7-N-methylcarboxylate,

[0110] 2-(4-chlorophenyl)-4-(3-thiophene-amino)-thiopheno[2,3-d]pyridazine-7-N,N-dimethylcarboxylate,

[0111] 2-(4-chlorophenyl)-4-(3-furan-amino)-thieno[2,3-d]pyridazine-7-N-methylcarboxylate,

[0112] 2-(4-Chlorophenyl)-4-(3-furan-amino)-thieno[2,3-d]pyridazine-7-N,N-dimethylcarboxylate

[0113] 2-(3-Fluorophenyl)-4-(3-piperidinamino)-thieno[2,3-d]pyridazine-7-carboxylic acid amide,

[0114] 2-(3-Chlorophenyl)-4-(3-piperidinamino)-thieno[2,3-d]pyridazine-7-carboxylic acid amide,

[0115] 2-(3-Bromophenyl)-4-(3-piperidineamino)-thieno[2,3-d]pyridazine-7-carboxylic acid amide,

[0116] 2-(3-Chlorophenyl)-4-(3-piperidinamino)-thieno[2,3-d]pyridazine-7-N,N-carboxylic acid amide,

[0117] 2-(3-Chlorophenyl)-4-(3-Tetrahydrothiaran-amino)-thieno[2,3-d]pyridazine-7-N-methylcarboxylate,

[0118] 2-(3-Chlorophenyl)-4-(3-Tetrahydrothiaran-amino)-thieno[2,3-d]pyridazine-7-N,N-dimethylcarboxylate,

[0119] 2-(3-Chlorophenyl)-4-(3-tetrahydropyran-amino)-thieno[2,3-d]pyridazine-7-N-methylcarboxylate,

[0120] 2-(3-Chlorophenyl)-4-(3-tetrahydropyran-amino)-thieno[2,3-d]pyridazine-7-N,N-dimethylcarboxylate,

[0121] 2-(3-Fluorophenyl)-4-(3-Tetrahydropyrrole-amino)-thieno[2,3-d]pyridazine-7-carboxylic acid amide,

[0122] 2-(3-Chlorophenyl)-4-(3-Tetrahydropyrrole-amino)-thieno[2,3-d]pyridazine-7-carboxylic acid amide,

[0123] 2-(3-Bromophenyl)-4-(3-Tetrahydropyrrole-amino)-thieno[2,3-d]pyridazine-7-carboxylic acid amide,

[0124] 2-(3-Fluorophenyl)-4-(3-Tetrahydrofuran-amino)-thieno[2,3-d]pyridazine-7-carboxylic acid amide,

[0125] 2-(3-Chlorophenyl)-4-(3-tetrahydrofuran-amino)-thieno[2,3-d]pyridazine-7-carboxylic acid amide,

[0126] 2-(3-Bromophenyl)-4-(3-Tetrahydrofuran-amino)-thieno[2,3-d]pyridazine-7-carboxylic acid amide,

[0127] 2-(3-Fluorophenyl)-4-(3-Tetrahydrothiophene-amino)-thieno[2,3-d]pyridazine-7-carboxylic acid amide,

[0128] 2-(3-Chlorophenyl)-4-(3-Tetrahydrothiophene-amino)-thiopheno[2,3-d]pyridazine-7-carboxylic acid amide,

[0129] 2-(3-Bromophenyl)-4-(3-Tetrahydrothiophene-amino)-thiopheno[2,3-d]pyridazine-7-carboxylic acid amide,

[0130] 2-(3-Chlorophenyl)-4-(2-Tetrahydropyrrole-amino)-thieno[2,3-d]pyridazine-7-carboxylic acid amide,

[0131] 2-(3-Chlorophenyl)-4-(S-3-tetrahydropyrrole-amino)-thieno[2,3-d]pyridazine-7-carboxylic acid amide,

[0132] 2-(3-Chlorophenyl)-4-(R-3-tetrahydropyrrole-amino)-thieno[2,3-d]pyridazine-7-carboxylic acid amide,

[0133] 2-(3-Chlorophenyl)-4-(3-Tetrahydropyrrole-amino)-thieno[2,3-d]pyridazine-7-N-methylcarboxylate,

[0134] 2-(3-Chlorophenyl)-4-(3-Tetrahydropyrrole-amino)-thieno[2,3-d]pyridazine-7-N,N-dimethylcarboxylate,

[0135] 2-(3-fluorophenyl)-4-(3-pyridin-amino)-thieno[2,3-d]pyridazine-7-carboxylic acid amide,

[0136] 2-(3-Chlorophenyl)-4-(3-pyridin-amino)-thieno[2,3-d]pyridazine-7-carboxylic acid amide,

[0137] 2-(3-Bromophenyl)-4-(3-pyridine-amino)-thieno[2,3-d]pyridazine-7-carboxylic acid amide,

[0138] 2-(3-Fluorophenyl)-4-(3-α-pyran-amino)-thieno[2,3-d]pyridazine-7-carboxylic acid amide,

[0139] 2-(3-Chlorophenyl)-4-(3-α-pyran-amino)-thieno[2,3-d]pyridazine-7-carboxylic acid amide,

[0140] 2-(3-bromophenyl)-4-(3-α-pyran-amino)-thieno[2,3-d]pyridazine-7-carboxylic acid amide,

[0141] 2-(3-Fluorophenyl)-4-(3-α-thiaran-amino)-thieno[2,3-d]pyridazine-7-carboxylic acid amide,

[0142] 2-(3-Chlorophenyl)-4-(3-α-thiaran-amino)-thieno[2,3-d]pyridazine-7-carboxylic acid amide,

[0143] 2-(3-Bromophenyl)-4-(3-α-thiaran-amino)-thieno[2,3-d]pyridazine-7-carboxylic acid amide,

[0144] 2-(3-chlorophenyl)-4-(2-pyridin-amino)-thieno[2,3-d]pyridazine-7-carboxylic acid amide,

[0145] 2-(3-chlorophenyl)-4-(4-pyridine-amino)-thieno[2,3-d]pyridazine-7-carboxylic acid amide,

[0146] 2-(3-chlorophenyl)-4-(S-3-pyridin-amino)-thieno[2,3-d]pyridazine-7-carboxylic acid amide,

[0147] 2-(3-chlorophenyl)-4-(R-3-pyridin-amino)-thieno[2,3-d]pyridazine-7-carboxylic acid amide,

[0148] 2-(3-Chlorophenyl)-4-(3-pyridin-amino)-thieno[2,3-d]pyridazine-7-N-methylcarboxylate,

[0149] 2-(3-Chlorophenyl)-4-(3-pyridin-amino)-thieno[2,3-d]pyridazine-7-N,N-dimethylcarboxylate

[0150] 2-(3-Chlorophenyl)-4-(3-thiaran-amino)-thieno[2,3-d]pyridazine-7-N-methylcarboxylate,

[0151] 2-(3-Chlorophenyl)-4-(3-thiaran-amino)-thieno[2,3-d]pyridazine-7-N,N-dimethylcarboxylate,

[0152] 2-(3-Chlorophenyl)-4-(3-pyran-amino)-thieno[2,3-d]pyridazine-7-N-methylcarboxylate,

[0153] 2-(3-Chlorophenyl)-4-(3-pyran-amino)-thieno[2,3-d]pyridazine-7-N,N-dimethylcarboxylate,

[0154] 2-(3-Fluorophenyl)-4-(3-pyrrolo-amino)-thieno[2,3-d]pyridazine-7-carboxylic acid amide,

[0155] 2-(3-Chlorophenyl)-4-(3-pyrrolo-amino)-thieno[2,3-d]pyridazine-7-carboxylic acid amide,

[0156] 2-(3-Bromophenyl)-4-(3-pyrrolo-amino)-thieno[2,3-d]pyridazine-7-carboxylic acid amide,

[0157] 2-(3-Fluorophenyl)-4-(3-furan-amino)-thieno[2,3-d]pyridazine-7-carboxylic acid amide,

[0158] 2-(3-Chlorophenyl)-4-(3-furan-amino)-thieno[2,3-d]pyridazine-7-carboxylic acid amide,

[0159] 2-(3-Bromophenyl)-4-(3-furan-amino)-thieno[2,3-d]pyridazine-7-carboxylic acid amide,

[0160] 2-(3-fluorophenyl)-4-(3-thiophene-amino)-thiopheno[2,3-d]pyridazine-7-carboxylic acid amide,

[0161] 2-(3-Chlorophenyl)-4-(3-thiophene-amino)-thiopheno[2,3-d]pyridazine-7-carboxylic acid amide,

[0162] 2-(3-bromophenyl)-4-(3-thiophene-amino)-thiopheno[2,3-d]pyridazine-7-carboxylic acid amide,

[0163] 2-(3-chlorophenyl)-4-(2-pyrrolo-amino)-thieno[2,3-d]pyridazine-7-carboxylic acid amide,

[0164] 2-(3-chlorophenyl)-4-(S-3-pyrrolo-amino)-thieno[2,3-d]pyridazine-7-carboxylic acid amide,

[0165] 2-(3-chlorophenyl)-4-(R-3-pyrrolo-amino)-thieno[2,3-d]pyridazine-7-carboxylic acid amide,

[0166] 2-(3-Chlorophenyl)-4-(3-pyrrolo-amino)-thieno[2,3-d]pyridazine-7-N-methylcarboxylate,

[0167] 2-(3-Chlorophenyl)-4-(3-pyrrolo-amino)-thieno[2,3-d]pyridazine-7-N,N-dimethylcarboxylate,

[0168] 2-(3-Chlorophenyl)-4-(3-thiophene-amino)-thiopheno[2,3-d]pyridazine-7-N-methylcarboxylate,

[0169] 2-(3-Chlorophenyl)-4-(3-thiophene-amino)-thiopheno[2,3-d]pyridazine-7-N,N-dimethylcarboxylate,

[0170] 2-(3-Chlorophenyl)-4-(3-furan-amino)-thieno[2,3-d]pyridazine-7-N-methylcarboxylate,

[0171] 2-(3-Chlorophenyl)-4-(3-furan-amino)-thieno[2,3-d]pyridazine-7-N,N-dimethylcarboxylate

[0172] 2-(4-fluorophenyl)-4-(3-piperidinmethyl)-thieno[2,3-d]pyridazine-7-carboxamide,

[0173] 2-(4-chlorophenyl)-4-(3-piperidinmethyl)-thieno[2,3-d]pyridazine-7-carboxamide,

[0174] 2-(4-bromophenyl)-4-(3-piperidinylmethyl)-thieno[2,3-d]pyridazine-7-carboxamide,

[0175] 2-(4-fluorophenyl)-4-(3-tetrahydropyranmethyl)-thieno[2,3-d]pyridazine-7-carboxamide,

[0176] 2-(4-chlorophenyl)-4-(3-tetrahydropyranmethyl)-thieno[2,3-d]pyridazine-7-carboxamide,

[0177] 2-(4-bromophenyl)-4-(3-tetrahydropyranmethyl)-thieno[2,3-d]pyridazine-7-carboxamide,

[0178] 2-(4-fluorophenyl)-4-(3-tetrahydrothiaranmethyl)-thieno[2,3-d]pyridazine-7-carboxamide,

[0179] 2-(4-chlorophenyl)-4-(3-tetrahydrothiaranmethyl)-thieno[2,3-d]pyridazine-7-carboxamide,

[0180] 2-(4-bromophenyl)-4-(3-tetrahydrothiaranmethyl)-thieno[2,3-d]pyridazine-7-carboxamide,

[0181] 2-(4-chlorophenyl)-4-(2-piperidinmethyl)-thieno[2,3-d]pyridazine-7-carboxamide,

[0182] 2-(4-Chlorophenyl)-4-(4-piperidinmethyl)-thieno[2,3-d]pyridazine-7-carboxamide,

[0183] 2-(4-chlorophenyl)-4-(S-3-piperidinylmethyl)-thieno[2,3-d]pyridazine-7-carboxamide,

[0184] 2-(4-chlorophenyl)-4-(R-3-piperidinmethyl)-thieno[2,3-d]pyridazine-7-carboxamide,

[0185] 2-(4-chlorophenyl)-4-(3-piperidinmethyl)-thieno[2,3-d]pyridazine-7-N-methyl-formamide,

[0186] 2-(4-Chlorophenyl)-4-(3-piperidinylmethyl)-thieno[2,3-d]pyridazine-7-N,N-dimethyl-7-carboxamide,

[0187] 2-(4-chlorophenyl)-4-(3-tetrahydrothiaranmethyl)-thieno[2,3-d]pyridazine-7-N-methyl-formamide,

[0188] 2-(4-Chlorophenyl)-4-(3-Tetrahydrothiaranmethyl)-thieno[2,3-d]pyridazine-7-N,N-dimethyl-7-carboxamide,

[0189] 2-(4-chlorophenyl)-4-(3-tetrahydropyranmethyl)-thieno[2,3-d]pyridazine-7-N-methyl-formamide,

[0190] 2-(4-chlorophenyl)-4-(3-tetrahydropyranmethyl)-thieno[2,3-d]pyridazine-7-N,N-dimethyl-7-carboxamide,

[0191] 2-(4-fluorophenyl)-4-(3-pyrrolidinemethyl)-thieno[2,3-d]pyridazine-7-carboxamide,

[0192] 2-(4-chlorophenyl)-4-(3-pyrrolidinemethyl)-thieno[2,3-d]pyridazine-7-carboxamide,

[0193] 2-(4-bromophenyl)-4-(3-pyrrolidinemethyl)-thieno[2,3-d]pyridazine-7-carboxamide,

[0194] 2-(4-fluorophenyl)-4-(3-tetrahydrofuranmethyl)-thieno[2,3-d]pyridazine-7-carboxamide,

[0195] 2-(4-Chlorophenyl)-4-(3-Tetrahydrofuranmethyl)-thieno[2,3-d]pyridazine-7-carboxamide,

[0196] 2-(4-bromophenyl)-4-(3-tetrahydrofuranmethyl)-thieno[2,3-d]pyridazine-7-carboxamide,

[0197] 2-(4-fluorophenyl)-4-(3-tetrahydrothiophenemethyl)-thiopheno[2,3-d]pyridazine-7-carboxamide,

[0198] 2-(4-chlorophenyl)-4-(3-tetrahydrothiophenemethyl)-thiopheno[2,3-d]pyridazine-7-carboxamide,

[0199] 2-(4-bromophenyl)-4-(3-tetrahydrothiophenemethyl)-thiopheno[2,3-d]pyridazine-7-carboxamide,

[0200] 2-(4-chlorophenyl)-4-(3-pyrrolidinemethyl)-thieno[2,3-d]pyridazine-7-carboxamide,

[0201] 2-(4-chlorophenyl)-4-(S-3-pyrrolidinemethyl)-thieno[2,3-d]pyridazine-7-carboxamide,

[0202] 2-(4-chlorophenyl)-4-(R-3-pyrrolidinemethyl)-thieno[2,3-d]pyridazine-7-carboxamide,

[0203] 2-(4-chlorophenyl)-4-(3-pyrrolidinemethyl)-thieno[2,3-d]pyridazine-7-N-methyl-formamide,

[0204] 2-(4-Chlorophenyl)-4-(3-pyrrolidinemethyl)-thieno[2,3-d]pyridazine-7-N,N-dimethylformamide,

[0205] 2-(4-chlorophenyl)-4-(3-tetrahydrothiophenemethyl)-thiopheno[2,3-d]pyridazine-7-N-methyl-formamide,

[0206] 2-(4-chlorophenyl)-4-(3-tetrahydrothiophenemethyl)-thiopheno[2,3-d]pyridazine-7-N,N-dimethylformamide,

[0207] 2-(4-Chlorophenyl)-4-(3-Tetrahydrofuranmethyl)-thieno[2,3-d]pyridazine-7-N-methyl-formamide,

[0208] 2-(4-Chlorophenyl)-4-(3-Tetrahydrofuranmethyl)-thieno[2,3-d]pyridazine-7-N,N-dimethylformamide,

[0209] 2-(4-fluorophenyl)-4-(3-pyridylmethyl)-thieno[2,3-d]pyridazine-7-carboxamide,

[0210] 2-(4-chlorophenyl)-4-(3-pyridylmethyl)-thieno[2,3-d]pyridazine-7-carboxamide,

[0211] 2-(4-bromophenyl)-4-(3-pyridylmethyl)-thieno[2,3-d]pyridazine-7-carboxamide,

[0212] 2-(4-fluorophenyl)-4-(3-α-pyranmethyl)-thieno[2,3-d]pyridazine-7-carboxamide,

[0213] 2-(4-chlorophenyl)-4-(3-α-pyranmethyl)-thieno[2,3-d]pyridazine-7-carboxamide,

[0214] 2-(4-bromophenyl)-4-(3-α-pyranmethyl)-thieno[2,3-d]pyridazine-7-carboxamide,

[0215] 2-(4-fluorophenyl)-4-(3-α-thiamethoxymethyl)-thieno[2,3-d]pyridazine-7-carboxamide,

[0216] 2-(4-chlorophenyl)-4-(3-α-thiamethoxymethyl)-thieno[2,3-d]pyridazine-7-carboxamide,

[0217] 2-(4-bromophenyl)-4-(3-α-thiamethoxymethyl)-thieno[2,3-d]pyridazine-7-carboxamide,

[0218] 2-(4-chlorophenyl)-4-(2-pyridylmethyl)-thieno[2,3-d]pyridazine-7-carboxamide,

[0219] 2-(4-chlorophenyl)-4-(4-pyridylmethyl)-thieno[2,3-d]pyridazine-7-carboxamide,

[0220] 2-(4-chlorophenyl)-4-(3-pyridinylmethyl)-thieno[2,3-d]pyridazine-7-N-methylformamide,

[0221] 2-(4-chlorophenyl)-4-(3-pyridinylmethyl)-thieno[2,3-d]pyridazine-7-N,N-dimethylformamide

[0222] 2-(4-chlorophenyl)-4-(3-thiamethoxymethyl)-thieno[2,3-d]pyridazine-7-N-methylformamide,

[0223] 2-(4-Chlorophenyl)-4-(3-thiamethoxymethyl)-thieno[2,3-d]pyridazine-7-N,N-dimethylformamide,

[0224] 2-(4-chlorophenyl)-4-(3-pyranmethyl)-thieno[2,3-d]pyridazine-7-N-methylformamide,

[0225] 2-(4-Chlorophenyl)-4-(3-pyranmethyl)-thieno[2,3-d]pyridazine-7-N,N-dimethylformamide,

[0226] 2-(4-fluorophenyl)-4-(3-pyrrolemethyl)-thieno[2,3-d]pyridazine-7-carboxamide,

[0227] 2-(4-chlorophenyl)-4-(3-pyrrolemethyl)-thieno[2,3-d]pyridazine-7-carboxamide,

[0228] 2-(4-bromophenyl)-4-(3-pyrrolemethyl)-thieno[2,3-d]pyridazine-7-carboxamide,

[0229] 2-(4-fluorophenyl)-4-(3-furanmethyl)-thieno[2,3-d]pyridazine-7-carboxamide,

[0230] 2-(4-chlorophenyl)-4-(3-furanmethyl)-thieno[2,3-d]pyridazine-7-carboxamide,

[0231] 2-(4-bromophenyl)-4-(3-furanmethyl)-thieno[2,3-d]pyridazine-7-carboxamide,

[0232] 2-(4-fluorophenyl)-4-(3-thienmethyl)-thienro[2,3-d]pyridazine-7-carboxamide,

[0233] 2-(4-chlorophenyl)-4-(3-thienmethyl)-thienro[2,3-d]pyridazine-7-carboxamide,

[0234] 2-(4-bromophenyl)-4-(3-thienylmethyl)-thienro[2,3-d]pyridazine-7-carboxamide,

[0235] 2-(4-chlorophenyl)-4-(2-pyrrolemethyl)-thieno[2,3-d]pyridazine-7-carboxamide,

[0236] 2-(4-chlorophenyl)-4-(3-pyrrolemethyl)-thieno[2,3-d]pyridazine-7-N-methyl-formamide,

[0237] 2-(4-Chlorophenyl)-4-(3-pyrrolemethyl)-thieno[2,3-d]pyridazine-7-N,N-dimethylformamide,

[0238] 2-(4-chlorophenyl)-4-(3-thienylmethyl)-thienro[2,3-d]pyridazine-7-N-methyl-formamide,

[0239] 2-(4-chlorophenyl)-4-(3-thienylmethyl)-thienro[2,3-d]pyridazine-7-N,N-dimethylformamide,

[0240] 2-(4-Chlorophenyl)-4-(3-furanmethyl)-thieno[2,3-d]pyridazine-7-N-methylformamide,

[0241] 2-(4-chlorophenyl)-4-(3-furanmethyl)-thieno[2,3-d]pyridazine-7-N,N-dimethylformamide, and

[0242] 2-(3,5-dichlorophenyl)-4-(3-piperidinylmethyl)-thieno[2,3-d]pyridazine-7-carboxamide.

[0243] In one embodiment, the compound is selected from 2-(3-fluorophenyl)-4-(3-piperidinamino)-thieno[2,3-d]pyridazine-7-carboxylate, 2-(3-fluorophenyl)-4-(3-piperidinoxy)-thieno[2,3-d]pyridazine-7-carboxylate, 2-(3-fluorophenyl)-4-(3-piperidinthio)-thieno[2,3-d]pyridazine-7-carboxylate, and 2-(3,5-dichlorophenyl)-4-(3-piperidinmethyl)-thieno[2,3-d]pyridazine-7-carboxylate.

[0244] Immunotherapy agents or therapeutic agents targeting cancer-promoting / maintaining molecules can be inhibitors of PD-1, PD-L1, CTLA-4, HER-2, CD20, CD33, or CD52. In embodiments, immunotherapy agents or therapeutic agents targeting cancer-promoting / maintaining molecules can be antibodies targeting PD-1, PD-L1, CTLA-4, HER-2, CD20, CD33, or CD52, or antibody-drug conjugates (ADCs) or CAR-T cells targeting PD-L1, HER-2, CD20, CD33, and / or CD52.

[0245] In one implementation, the immunotherapeutic agent or the therapeutic agent targeting cancer-promoting / maintaining molecules is a PD-1 inhibitor, such as a PD-1 antibody; or a PD-L1 inhibitor, such as a PD-L1 antibody. The PD-1 antibody may be nivolumab, pembrolizumab, or toripalimab. The PD-L1 antibody may be atezolizumab, durvalumab, or avelumab.

[0246] CTLA-4 inhibitors can be CTLA-4 antibodies, such as ipilimumab. HER2 antibodies can be trastuzumab or pertuzumab. CD20 antibodies can be rituximab, tiuxetan, tositumomab, offatumumab, occrelizumab, veituzumab, or obinutuzumab. CD33 antibodies can be gemtuzumab. CD-52 antibodies can be alemtuzumab.

[0247] In one embodiment, the cancer is a solid cancer selected from lung cancer, prostate cancer, ovarian cancer, brain cancer, breast cancer, skin cancer, bladder cancer, colon cancer, gastrointestinal cancer, head and neck cancer, stomach cancer, pancreatic cancer, nerve cancer, kidney cancer, and liver cancer. In one embodiment, the cancer is colon cancer. In one embodiment, the cancer is colon adenocarcinoma.

[0248] In one implementation, the cancer is a blood cancer selected from lymphocytic leukemia, myeloid leukemia, non-Hodgkin lymphoma, and Hodgkin lymphoma.

[0249] Compounds of Formula I and immunotherapeutic agents or therapeutic agents targeting cancer-promoting / maintaining molecules can be administered simultaneously as a single composition in a pharmaceutically acceptable carrier, or as separate compositions. They can also be administered sequentially.

[0250] This application also discloses the use of the compound of formula I with immunotherapeutic agents or therapeutic agents targeting cancer-promoting / maintaining molecules in the treatment of cancer.

[0251] Exemplary combination therapy, compared to any single therapy, provides enhanced antitumor efficacy with the same or even slightly reduced toxicity, as has been demonstrated in mPD-1. - / - mPD-L1 - / -hPD-1 + / + hPD-L1 + / + This has been confirmed in transgenic mouse models.

[0252] The CHK1 inhibitor of this application synergizes with PD-1 antibodies even at doses that do not show any anti-tumor effect when used alone.

[0253] In a second aspect, this application discloses a method for treating cancer, comprising administering a therapeutically effective amount of a compound of formula I, or a pharmaceutically acceptable salt thereof, to a subject in need, and i) an immunotherapeutic agent or a therapeutic agent targeting cancer-promoting / maintaining molecules, and ii) a chemotherapeutic agent.

[0254] Immunotherapy agents or therapeutic agents targeting cancer-promoting / maintaining molecules can be inhibitors of PD-1, PD-L1, CTLA-4, HER-2, CD20, CD33, or CD52. In embodiments, immunotherapy agents or therapeutic agents targeting cancer-promoting / maintaining molecules can be antibodies targeting PD-1, PD-L1, CTLA-4, HER-2, CD20, CD33, or CD52, as well as antibody-drug conjugates (ADCs) or CAR-T cells targeting PD-L1, HER-2, CD20, CD33, and / or CD52.

[0255] In one implementation, the immunotherapeutic agent or the therapeutic agent targeting the cancer-promoting / maintaining molecule is a PD-1 inhibitor, such as a PD-1 antibody; or a PD-L1 inhibitor, such as a PD-L1 antibody. The PD-1 antibody may be nivolumab, pembrolizumab, or toripalimab. The PD-L1 antibody may be atezolizumab, durvalumab, or avelumab.

[0256] CTLA-4 inhibitors can be CTLA-4 antibodies, such as ipilimumab. HER2 antibodies can be trastuzumab or pertuzumab. CD20 antibodies can be rituximab, teimomab, tosimob, oflamb, ocletuzumab, vetuzumab, or oxotuzumab. CD33 antibodies can be gemtuzumab. CD-52 antibodies can be alemtuzumab.

[0257] The chemotherapeutic agents suitable for use in this invention may be cisplatin, pemetrexed, gemcitabine, cytarabine, hydroxyurea, temozolomide, irinotecan, cyclophosphamide, mitoxantrone, etoposide, leucovorin, fludarabine, fluorouracil, or combinations thereof.

[0258] In one embodiment, the cancer is a solid cancer selected from lung cancer, prostate cancer, ovarian cancer, brain cancer, breast cancer, skin cancer, bladder cancer, colon cancer, gastrointestinal cancer, head and neck cancer, stomach cancer, pancreatic cancer, nerve cancer, kidney cancer, and liver cancer. In one embodiment, the cancer is colon cancer. In one embodiment, the cancer is colon adenocarcinoma.

[0259] In one implementation, the cancer is a blood cancer selected from lymphocytic leukemia, myeloid leukemia, non-Hodgkin lymphoma, and Hodgkin lymphoma.

[0260] Compounds of Formula I, immunotherapeutic agents (or therapeutic agents targeting cancer-promoting molecules), and chemotherapeutic agents may be administered simultaneously as a single composition in a pharmaceutically acceptable carrier, or as separate compositions. They may also be administered sequentially.

[0261] This application also discloses the use of Formula I compounds with immunotherapeutic agents (or therapeutic agents targeting cancer-promoting molecules) and chemotherapeutic agents in the treatment of cancer.

[0262] Exemplary combination therapy, compared to any single therapy, provides enhanced antitumor efficacy with the same or even slightly reduced toxicity, as has been demonstrated in mPD-1. - / - mPD-L1 - / - hPD-1 + / + hPD-L1 + / + This has been confirmed in transgenic mouse models.

[0263] The CHK1 inhibitor described in this application can synergize with PD-1 antibodies and chemotherapeutic agents even at doses that do not show any anti-tumor effect when used alone.

[0264] Other features and advantages of this application, as expressly described and equivalents understood by those skilled in the art, will become apparent from the following drawings, detailed description and embodiments, and the claims (which should not be construed as restrictive). All disclosures, references, Genbank entries, patents and published patent applications cited throughout this application are expressly incorporated herein by reference. Attached Figure Description

[0265] The following detailed description, provided by way of example but not intended to limit the invention to the specific embodiments described, is presented in conjunction with the accompanying drawings for a better understanding.

[0266] Figure 1 This is a line graph showing the changes in mouse body weight during the test.

[0267] Figure 2 It is a line graph showing the changes in tumor size during the test.

[0268] Figure 3 This is a photo of the tumor isolated from the mouse on day 27.

[0269] Figure 4 This is a line graph showing the changes in mouse body weight during the test.

[0270] Figure 5 It is a line graph showing the changes in tumor size during the test.

[0271] Figure 6 The individual tumor sizes of mice in different groups are shown during the test.

[0272] Figure 7 This is a scatter plot showing the size of an individual tumor on day 14.

[0273] Figure 8 These are photos of tumors isolated from mice in groups 3 and 4 on day 21.

[0274] Figure 9 It is a line graph showing the changes in tumor size during the test.

[0275] Figure 10 It is a line graph showing the changes in tumor size during the test. Detailed Implementation

[0276] Before disclosing and describing specific embodiments of this application, it should be understood that this application is not limited to the specific methods and materials disclosed herein, which may vary to some extent. It should be understood that the terminology used herein is for the purpose of describing specific embodiments only and is not restrictive, and the scope of this application will be defined only by the appended claims and their equivalents.

[0277] The singular forms “a,” “a kind,” and “the” include the plural referent, unless the context clearly indicates otherwise.

[0278] Unless otherwise stated, it is assumed that any heteroatom with an unsaturated valence has a hydrogen atom sufficient to meet the valence requirement.

[0279] Throughout the specification and appended claims, the specified chemical formula or name will include all its stereoisomers, optical isomers, and racemates, if such isomers are present. Unless otherwise stated, all chiral (enantiomers and diastereomers) and racemates are within the scope of this application. Many geometric isomers of C=C double bonds, C=N double bonds, cyclic systems, etc., may also be present in the compound, and all such stable isomers are included in this application. Cis- and trans- (or E- and Z-) geometric isomers of the compounds of this application are also described, and they can be separated into mixtures of isomers or as separate isomeric forms. The present compounds can be separated in optical or racemic forms. The optical form can be prepared by the resolution of the racemate or by synthesis from optically active starting materials. All methods used to prepare the compounds of this application and the intermediates therein are considered part of this application. When preparing enantiomers and diastereomers, they can be separated by conventional methods, such as by chromatography or fractional crystallization. Depending on the process conditions, the final products of this application are obtained in free (neutral) or salt form. Both the free form and salts of these final products are within the scope of this application. If desired, one form of the compound can be converted to another. Free bases or acids can be converted to salts; salts can be converted to free compounds or another type of salt; mixtures of isomers of this application can be separated into individual isomers. The compounds of this application, in their free form and salts, can exist in various tautomer forms, wherein hydrogen atoms are transposed to other parts of the molecule, thereby rearranging the chemical bonds between the atoms of the molecule. It should be understood that all tautomer forms, where present, are included in this application.

[0280] The term "substitution" as used in this article refers to the substitution of at least one hydrogen atom with a non-hydrogen atom, as long as the normal chemical valence is maintained and the substitution results in a stable compound.

[0281] When a substituent is indicated as "optionally substituted", the substituent is selected from, for example, alkyl, cycloalkyl, aryl, heterocyclic, halogen, hydroxyl, alkoxy, oxy, alkanoyl, aryloxy, alkanoyloxy, amino, alkylamino, arylamino, arylalkylamino, disubstituted amino (wherein the two amino substituents of the amino group are selected from alkyl, aryl, or arylalkyl), alkoxyamino, arylylamino, arylalkylylamino, substituted alkanoylamino, substituted arylamino, substituted arylalkylylamino, mercapto, alkylthio, arylthio, arylalkylthio, alkylthiocarbonyl, arylthiocarbonyl, arylalkylthiocarbonyl, alkylsulfonyl, arylsulfonyl, aryl Alkylsulfonyl, sulfonamide (e.g., -SO2NH2), substituted sulfonamide, nitro, cyano, carboxyl, carbamoyl (e.g., -CONH2), substituted carbamoyl (e.g., -CONHalkyl, -CONHaryl, -CONHaralkyl, or where the two substituents on nitrogen are selected from alkyl, aryl, or aralkyl), alkoxycarbonyl, aryl, substituted aryl, guanidine, heterocyclic (e.g., indolyl, imidazolyl, furanyl, thiophene, thiazolyl, pyrrole, pyridinyl, pyrimidinyl, pyrrolidinyl, piperidinyl, morpholinyl, piperazine, homopiperazine, etc.), and substituted heterocyclic groups, unless otherwise defined.

[0282] As used herein, the terms “alkyl” or “alkylene” are intended to include branched and straight-chain saturated aliphatic hydrocarbon groups having a specific number of carbon atoms. For example, “C1-C6 alkyl” refers to an alkyl group having 1-6 carbon atoms. Exemplary alkyl groups include, but are not limited to, methyl (Me), ethyl (Et), propyl (e.g., n-propyl and isopropyl), butyl (e.g., n-butyl, isobutyl, tert-butyl), and pentyl (e.g., n-pentyl, isopentyl, neopentyl).

[0283] The term "alkenyl" refers to a straight-chain or branched hydrocarbon group containing one or more double bonds and typically having a length of 2-20 carbon atoms. For example, "C2-C8 alkenyl" contains 2-8 carbon atoms. Alkenyl groups include, but are not limited to, ethylene, propylene, butene, 1-methyl-2-but-1-ene, pentene, octene, etc.

[0284] The term "alkynyl" refers to a straight-chain or branched hydrocarbon group containing one or more triple bonds, typically with a length of 2-20 carbon atoms. For example, "C2-C8 alkynyl" contains 2-8 carbon atoms. Representative alkenyl groups include, but are not limited to, acetylene, 1-propyne, 1-butyne, heptyne, octylene, etc.

[0285] The term "alkoxy" or "alkyloxy" refers to -O-alkyl. "C1-C6 alkoxy" (or alkyloxy) is intended to include C1, C2, C3, C4, C5, and C6 alkoxy groups. Exemplary alkoxy groups include, but are not limited to, methoxy, ethoxy, propoxy (e.g., n-propoxy and isopropoxy), and tert-butoxy. Similarly, "alkylthio" or "thioalkoxy" means an alkyl group as defined above that has a specified number of carbon atoms and is linked by a sulfur bridge, such as methyl-S- and ethyl-S-.

[0286] The term "aryl," whether alone or as part of a larger group such as "aralkyl," "arylalkoxy," or "aryloxyalkyl," refers to a monocyclic, bicyclic, or tricyclic system having a total of 5-15 ring members, wherein at least one ring in the system is aromatic, and wherein each ring in the system contains 3-7 ring members. In some embodiments of this application, "aryl" refers to an aromatic ring system, including, but not limited to, phenyl, biphenyl, indanyl, 1-naphthyl, 2-naphthyl, and tetrahydronaphthyl. The term "aralkyl" or "arylalkyl" refers to an alkyl group attached to an aromatic ring. Non-limiting examples include benzyl, phenethyl, etc. Fused-ring aryl groups can be attached to another group at a suitable position on the cycloalkyl ring or aromatic ring. For example:

[0287]

[0288] An arrow coming out of the ring system indicates that the bond can be connected to any suitable ring atom.

[0289] The term "cycloalkyl" refers to a cyclic alkyl group. C3-C6 cycloalkyl is intended to include C3, C4, C5, and C6 cycloalkyl groups. Exemplary cycloalkyl groups include, but are not limited to, cyclopropyl, cyclobutyl, cyclopentyl, cyclohexyl, and norbornyl. Branched cycloalkyl groups, such as 1-methylcyclopropyl and 2-methylcyclopropyl, are included within the definition of "cycloalkyl." The term "cycloalkenyl" refers to a cyclic alkenyl group. 4-6 Cycloalkenyl is intended to include C4, C5, and C6 cycloalkenyl groups. Exemplary cycloalkenyl groups include, but are not limited to, cyclobutenyl, cyclopentenyl, and cyclohexenyl.

[0290] As used herein, the terms “heterocyclic,” “heterocyclic group,” or “heterocyclic group” refer to a stable 4-, 5-, or 6-membered monocyclic ring that is saturated, partially unsaturated, or fully unsaturated and contains a carbon atom and one, two, three, or four nitrogen, oxygen, or other non-carbon atoms.

[0291] In cases where the compounds of this application contain nitrogen atoms (e.g., amines), they can be converted into N-oxides by treatment with an oxidizing agent (e.g., mCPBA and / or hydrogen peroxide) to obtain other compounds of this application. Therefore, the nitrogen atoms shown and claimed are also considered to include the nitrogen shown and its N-oxide (N→O) derivatives.

[0292] When any variable appears more than once in any component or general formula of a compound, its definition for each occurrence is independent of its definition for every other occurrence. Thus, for example, if a group shows substitution by 0-3 R groups, that group may optionally be substituted by up to 3 R groups, and each occurrence of R is independently selected from the definition of R. Furthermore, such combinations are permitted only if the combination of substituents and / or variables results in a stable compound.

[0293] A substituent may bond to any atom in the ring when the bond on the substituent shows a cross bond with two atoms in the connecting ring. When a substituent is listed without specifying at which atom it is connected to the remaining atoms of a given general formula compound, the substituent may be connected via any atom of that substituent. Such combinations are permitted only if the combination of substituents and / or variables results in a stable compound.

[0294] This document uses the phrase "pharmaceutically acceptable" to refer to compounds, materials, compositions, and / or dosage forms that, to the extent of credible medical judgment, are suitable for use in contact with human and animal tissues without excessive toxicity, irritation, allergic reactions, and / or other problems or complications, and that meet a reasonable benefit / risk ratio. As used herein, "pharmaceutically acceptable salt" refers to a derivative of the disclosed compound, wherein the parent compound is modified by preparing its acid or base salt. Examples of pharmaceutically acceptable salts include, but are not limited to, mineral acid or inorganic acid salts with basic groups such as amines; and basic or organic salts with acidic groups such as carboxylic acids. Pharmaceutically acceptable salts include common non-toxic salts or quaternary ammonium salts of the parent compound formed from, for example, non-toxic inorganic or organic acids. For example, these common non-toxic salts include those derived from inorganic acids such as hydrochloric acid, hydrobromic acid, sulfuric acid, aminosulfonic acid, phosphoric acid, and nitric acid; and those derived from organic acids such as acetic acid, propionic acid, succinic acid, glycolic acid, stearic acid, lactic acid, malic acid, tartaric acid, citric acid, ascorbic acid, pyric acid, maleic acid, hydroxymaleic acid, phenylacetic acid, glutamic acid, benzoic acid, salicylic acid, p-aminobenzenesulfonic acid, 2-acetylsalicylic acid, fumaric acid, p-methylbenzenesulfonic acid, methanesulfonic acid, ethanedisulfonic acid, oxalic acid, and hydroxyethanesulfonic acid.

[0295] The pharmaceutically acceptable salts of the present invention can be synthesized from parent compounds containing basic or acidic groups by common chemical methods. Generally, these salts can be prepared by reacting the free acidic or basic form of these compounds with a suitable base or acid in an ideal proportion in water, in an organic solvent, or in a mixture of both; generally, non-aqueous media, such as diethyl ether, ethyl acetate, ethanol, isopropanol, or acetonitrile, are preferred. A list of suitable salts is described in Remington: The Science and Practice of Pharmacy, 22nd Edition, Allen, LV Jr., Ed.; Pharmaceutical Press, London, UK (2012), the disclosure of which is incorporated herein by reference.

[0296] As used herein, the term "effective amount" means, for example, the amount of a drug or pharmaceutical agent, i.e., the compound of this application, that elicits a biological or medical response in a tissue, system, animal, or human, as sought by an investigator or clinician. Furthermore, the term "therapeutic effective amount" means any amount that, compared to a corresponding subject who did not receive such amounts, results in improved treatment, cure, prevention, reduction of disease, symptom, or side effects, or a decrease in the rate of disease or symptom progression. Effective amounts may be administered in one or more doses, applications, or dosages and are not intended to be limited to a particular formulation or route of administration. The term also includes, within its scope, amounts that effectively enhance normal physiological function.

[0297] The term “subject” includes any human or non-human animal. The term “non-human animal” includes all vertebrates, such as mammals and non-mammals, such as non-human primates, sheep, dogs, cats, cattle, horses, chickens, amphibians, and reptiles, although mammals, such as non-human primates, sheep, dogs, cats, cattle, and horses, are preferred.

[0298] As used in this article, the term "treatment" includes any effect that causes improvement of symptoms, diseases, conditions, etc., such as reducing, decreasing, regulating, improving, or eliminating, or improving the symptoms of these conditions.

[0299] As used in this text, the term "pharmaceutical composition" refers to a combination of an active agent and a carrier (inert or active) that makes the composition particularly suitable for diagnostic or therapeutic use in vivo or in vitro.

[0300] As used in this article, “extracorporeal” includes subcutaneous, intradermal, intravenous, intramuscular, intra-articular, intra-articular, synovial, intrasternal, intracranial, intramuscular, or infusion.

[0301] Compounds of Formula I

[0302] Compounds of Formula I are disclosed as CHK inhibitors in WO2009 / 092278 and WO2011 / 035077. They exhibit direct antitumor effects and can sensitize other DNA-damaging drugs. The synthetic methods and functional tests of these compounds are also detailed in these two international patent applications.

[0303] Exemplary compounds include (2-(3-fluorophenyl)-4-(3-piperidinamino)-thieno[2,3-d]pyridazine-7-carboxylate, referred to as compound 6 in Examples 1-5), (2-(3-fluorophenyl)-4-(3-piperidinoxy)-thieno[2,3-d]pyridazine-7-carboxylic acid amide, referred to as compound 6-1 in Example 2), (2-(3-fluorophenyl)-4-(3-piperidinthio)-thieno[2,3-d]pyridazine-7-carboxylic acid amide, referred to as compound 6-2 in Example 2), (2-(3,5-dichlorophenyl)-4-(3-piperidinmethyl)-thieno[2,3-d]pyridazine-7-carboxamide, referred to as compound 6-3 in Example 2).

[0304] Immunotherapy agents or therapeutic agents targeting cancer-promoting molecules

[0305] The term "immunotherapy agent" in this article refers to therapeutic agents that participate in the activation of the immune system and / or amplification of the immune response. Immunotherapy agents include, but are not limited to, PD-1 inhibitors, PD-L1 inhibitors, CTLA-4 inhibitors, and CD33 inhibitors. PD-1 inhibitors, PD-L1 inhibitors, CTLA-4 inhibitors, and CD33 inhibitors include, but are not limited to, PD-1 antibodies, PD-L1 antibodies, CTLA-4 antibodies, CD33 antibodies, PD-L1 antibody-drug conjugates, CD33 antibody-drug conjugates, CAR-T cells targeting PD-L1, and CAR-T cells targeting CD33.

[0306] In this article, therapeutic agents targeting cancer-promoting molecules refer to those that target molecules required for cancer cell growth, which partially overlaps with immunotherapies. Such therapeutic agents include, but are not limited to, PD-1 inhibitors, PD-L1 inhibitors, CTLA-4 inhibitors, HER-2 inhibitors, CD20 inhibitors, CD33 inhibitors, or CD52 inhibitors. These inhibitors can be antibodies targeting PD-1, PD-L1, CTLA-4, HER-2, CD20, and / or CD33, antibody-drug conjugates targeting PD-L1, HER-2, CD20, and / or CD33, or CAR-T cells.

[0307] Among these targets, PD-1, PD-L1, and CTLA-4 are immune checkpoints.

[0308] PD-1 is a protein on the cell surface that regulates the immune system's response to human cells by downregulating the immune system and promotes self-tolerance by suppressing T-cell inflammatory activity. PD-1 binds to two ligands, PD-L1 and PD-L2. The binding of PD-L1 to PD-1 transmits an inhibitory signal, which reduces the proliferation of antigen-specific T cells in lymph nodes while simultaneously reducing apoptosis of regulatory T cells (anti-inflammatory and suppressor T cells). PD-1 and PD-L1 inhibitors activate the immune system to attack tumors and have been used to treat certain types of cancer, making them among the most intensively studied cancer therapeutics. However, only less than approximately 20% of patients respond to PD-1 / PD-L1 inhibitors.

[0309] CTLA-4, also known as cytotoxic T-lymphocyte antigen 4, is a transmembrane protein on the surface of T cells. When it binds to B7 on antigen-presenting cells, it inhibits T cell activation. This is believed to occur during the "promotion" phase of the immune system in lymph nodes. The overall effect is immune downregulation. Therefore, CTLA-4 is described as an immune checkpoint "shut-off." Two fully human CTLA-4 monoclonal antibodies have been developed and are being tested in phase III clinical trials for the treatment of metastatic melanoma: ipilimumab and tremelimumab. Ipilimumab has been approved by the FDA for the treatment of metastatic melanoma, adjuvant melanoma, and renal cell carcinoma.

[0310] Other therapeutic agents are known to play a role in promoting cancer formation and development.

[0311] The PD-1 inhibitors that can be used in this invention include, but are not limited to, nivolumab, pembrolizumab, cimiplimab injection (Cemiplimab-rwlc), toripalimab, sintilimab, camrelizumab, and tislelizumab.

[0312] The PD-L1 inhibitors that can be used in this invention include, but are not limited to, atezolizumab, avelumab, and durvalumab.

[0313] The CTLA-4 inhibitors that can be used in this invention include, but are not limited to, ipilimumab, trimemumab, zalifrelimab, AGEN-1181, KN-044, BCD-145, abatacept, BMS-986249, IO-102, ONC-392, REGN-4659, HBM-4003, RG2077, and YH-001.

[0314] Certain bispecific molecules targeting PD-1 and CTLA-4 can also be used in the methods of this invention, including, but not limited to, BCD-217, AK104, PSB205, and MEDI-5752.

[0315] Certain bispecific molecules targeting PD-L1 and CTLA-4 can also be used in the methods of this invention, including, but not limited to, KN-046.

[0316] Chemotherapy drugs

[0317] Chemotherapy agents, as discussed in this article, are powerful chemicals that kill rapidly growing cells in the body. Such preparations are commonly used to treat cancers, such as cancer cells that grow and divide faster than other cells.

[0318] Chemotherapy agents used for cancer treatment include, but are not limited to, cisplatin, pemetrexed, gemcitabine, cytarabine, hydroxyurea, temozolomide, irinotecan, cyclophosphamide, mitoxantrone, etoposide, leucovorin, fludarabine, and fluorouracil.

[0319] Gemcitabine, a chemotherapy drug used to treat various types of cancer, is a ribonucleoside reductase inhibitor that causes dNTP depletion and replication fork arrest, blocking the formation of new DNA. First approved for medical use in 1995, it is now used alone as a first-line treatment for pancreatic cancer and in combination with cisplatin for the first-line treatment of advanced or metastatic bladder cancer and advanced or metastatic non-small cell lung cancer. It is also used in combination with carboplatin as a second-line treatment for ovarian cancer and in combination with paclitaxel as a second-line treatment for metastatic or unremovable breast cancer. Use of gemcitabine may cause side effects such as bone marrow suppression, liver and kidney problems, nausea, fever, and hair loss.

[0320] Cisplatin is another chemotherapy drug commonly used to treat various cancers. It was discovered in 1845 and put into medical use in 1978. It works by binding to DNA and thus inhibiting DNA replication, and is used to treat sarcomas, SCLC, ovarian cancer, and other cancers.

[0321] combination therapy

[0322] Compounds of Formula I, as CHK inhibitors, can be used in combination with i) immunotherapeutic agents or therapeutic agents targeting cancer-promoting molecules, and optionally (ii) chemotherapeutic agents, to achieve better anticancer effects and / or lower toxicity to the human body.

[0323] Compounds of Formula I, together with immunotherapeutic agents or therapeutic agents targeting cancer-promoting / maintaining molecules, can be administered as a single composition in a pharmaceutically acceptable carrier, or as separate compositions. They can also be administered sequentially.

[0324] In other embodiments, the Formula I compound, the chemotherapeutic agent, and the immunotherapeutic agent (therapeutic agent targeting cancer-promoting molecules) may be administered as a single composition in a pharmaceutically acceptable carrier, or as separate compositions. They may also be administered sequentially.

[0325] The combination therapy of this application can be used to treat cancers, such as solid cancers selected from lung cancer, prostate cancer, ovarian cancer, brain cancer, breast cancer, skin cancer, bladder cancer, colon cancer, gastrointestinal cancer, head and neck cancer, stomach cancer, pancreatic cancer, neurogenic cancer, kidney cancer, and liver cancer, or hematologic cancers selected from lymphocytic leukemia, myeloid leukemia, non-Hodgkin lymphoma, and Hodgkin lymphoma. In one embodiment, the cancer is colon cancer. In one embodiment, the cancer is colon adenocarcinoma.

[0326] The combination therapy of this application can be applied to animals, preferably mammals (e.g., domestic animals, cats, dogs, mice, rats), and more preferably humans. Any method of administration can be used to deliver the compound of Formula I, the immunotherapy agent (or the therapeutic agent targeting a cancer-promoting molecule), and / or the chemotherapeutic agent to a subject in need. In some embodiments, the compound of Formula I, the chemotherapeutic agent, and / or the immunotherapy agent (or the therapeutic agent targeting a cancer-promoting molecule) is administered orally. In other embodiments, the compound of Formula I, the chemotherapeutic agent, and / or the immunotherapy agent (or the therapeutic agent targeting a cancer-promoting molecule) is administered parenterally.

[0327] One or more other agents or treatments, such as immune enhancers, immunosuppressants, antitumor vaccines, cytokine therapies (e.g., IL2 and GM-CSF), and / or tyrosine kinase inhibitors, may optionally be used in combination with the combination therapy of this application. Other agents may be combined with the combination therapy of this application in a single dosage form, or these agents may be administered as separate dosage forms, simultaneously or sequentially.

[0328] Pharmaceutical composition and administration

[0329] This application also provides pharmaceutically acceptable compositions comprising a therapeutically effective amount of one or more Formula I compounds formulated with one or more pharmaceutically acceptable carriers (additives) and / or diluents, a therapeutically effective amount of the aforementioned immunotherapeutic agent or therapeutic agent targeting cancer-promoting molecules formulated with one or more pharmaceutically acceptable carriers (additives) and / or diluents, optionally a therapeutically effective amount of the aforementioned chemotherapeutic agent formulated with one or more pharmaceutically acceptable carriers (additives) and / or diluents, and optionally one or more other desired therapeutic agents. The compounds of this application may be administered in any suitable manner, for example, as pills, capsules (each comprising a sustained-release or time-release formulation), tablets, powders, granules, aqueous solutions, tinctures, suspensions (including nanosuspensions, microsuspensions, spray-dried dispersants), syrups, and emulsions, via oral administration; sublingual administration; buccal administration; parenteral administration, such as by subcutaneous, intravenous, intramuscular, or intrasternal injection, or by infusion techniques (e.g., as sterile injectable aqueous or non-aqueous solutions or suspensions); nasal administration, including, for example, administration to the nasal membrane via inhalation spray; topical administration, such as in the form of ointments or creams; or rectal administration, such as in the form of suppositories. Chemotherapeutic agents of this application may be administered in any suitable manner with a pharmaceutically acceptable carrier. Immunotherapy agents or therapeutic agents targeting cancer-promoting molecules may be administered in any suitable manner with a pharmaceutically acceptable carrier. The pharmaceutical compositions of this application may also be prepared as liposomes and nanoparticles.

[0330] The term "pharmaceutically acceptable" is used in this article to refer to compounds, materials, compositions, and / or dosage forms that, to the extent of credible medical judgment, are suitable for contact with human and animal tissues without causing excessive toxicity, irritation, allergic reactions, and / or other problems or complications, and that meet a reasonable benefit / risk ratio. "Pharmaceutically acceptable carriers" refers to media generally accepted in the field for the delivery of bioactive agents to animals, particularly mammals, including, i.e., adjuvants, excipients, or carriers such as diluents, preservatives, fillers, flow conditioners, disintegrants, wetting agents, emulsifiers, suspensions, sweeteners, flavoring agents, fragrances, antibacterial agents, antifungal agents, lubricants, and dispersants, depending on the inherent characteristics of the route of administration and the dosage form.

[0331] The dosing regimens of the compounds, chemotherapeutic agents, and / or immunotherapeutic agents, or therapeutic agents targeting cancer-promoting molecules of the present invention will, of course, vary based on known factors, such as the pharmacodynamic properties of each specific drug, and its mode of administration and route of administration; the recipient's species, age, sex, health status, disease condition, and weight; the characteristics and severity of symptoms; the type of concurrent treatment; the frequency of treatment; the route of administration; the patient's renal and hepatic function; and the desired effect. Generally, the guidelines are that, when used for the specified effect, the oral daily dose of each active ingredient is about 0.001 to about 5000 mg per day, preferably about 0.01 to about 1000 mg per day, and most preferably about 0.1 to about 250 mg per day. For intravenous administration, during a constant-rate infusion, the most preferred dose is about 0.01 to about 10 mg / kg / minute.

[0332] The compounds, immunotherapeutic agents, or therapeutic agents targeting cancer-promoting molecules, and optionally chemotherapeutic agents of this application, may be administered in a single daily dose, or the total daily dose may be administered in divided doses twice, three, or four times a day.

[0333] Example

[0334] Example 1. The combination of CHK inhibitor and PD-1 antibody has a synergistic anti-tumor effect.

[0335] CT-26WT cells were kept at 37°C and 5% CO2 in RPMI 1640 medium (10-040-CV, Coming cellgro) supplemented with 10% FBS (10270-106, GIBCO) and used before 10 passages.

[0336] On day 0, 45 female BALB / c mice, aged 4-5 weeks, were subcutaneously injected with approximately 200.0 μL of a solution containing approximately 5 × 10⁻⁶ ppm of sodium hydroxide. 5 CT-26WT cell culture medium.

[0337] On day 5, the 32 tumor-bearing mice were randomly divided into 4 groups of 8 mice each, based on their body weight. The remaining mice were euthanized.

[0338] Four groups of animals were administered 1.0 mg / mL of anti-m-PD-1 antibody (BE0146, BioXcell) in PBS (21-040-CVR, BioXcell), 3.5 mg / mL of compound 6 in saline, anti-m-PD-1 antibody + compound 6, and control solvent (saline), respectively. The dosages are shown in Table 1 below.

[0339] Table 1. Dosing Regimen

[0340]

[0341] The mice's physiological condition and activity level were observed daily, and their body weight and tumor volume were measured every Tuesday, Thursday, and Sunday. Tumor volume (V) was calculated as (length × width). 2 ) / 2.

[0342] On day 27, mice were euthanized, and tumors were collected, weighed, and photographed. Mouse body weight, tumor size, and tumor weight were analyzed using t-tests in SPSS. A p-value below 0.05 was considered statistically significant between groups.

[0343] Tumor growth inhibition (TGI) is calculated using the following two formulas.

[0344] Tumor growth inhibition = (average tumor size in the solvent group - average tumor size in the drug treatment group) / average tumor size in the solvent group × 100%

[0345] Tumor growth inhibition = (average tumor weight in the solvent group - average tumor weight in the drug treatment group) / average tumor weight in the solvent group × 100% Furthermore, using the formula Q = E A+B / (E A +E B -E A ×E B The Q-value was calculated to assess the combined effect of the PD-1 antibody and compound 6, where E... A+B E A and E B The Q values ​​refer to the TGI for combination therapy, PD-1 administration, and compound 6 administration, respectively. A Q value higher than 1.15 indicates a synergistic or additive effect.

[0346] During the experiment, the animals in the solvent group appeared vigorous, and no mouse deaths were observed in any of the four groups. The mean mouse body weight and mean tumor size from day 9 to day 27 are shown in the figures. Figure 1 and Figure 2 Furthermore, individual tumors isolated from mice on day 27 were displayed on... Figure 3 .

[0347] like Figure 1 As shown, from day 9 to day 27, the overall body weight of mice in all groups increased, partly due to tumor growth. After deducting tumor weight, the average body weight of mice in the solvent group, PD-1 antibody group, CHK inhibitor group, and combination therapy group on day 27 were 17.71 g, 19.68 g, 18.41 g, and 18.60 g, respectively. It can be seen that mice in the CHK inhibitor group or combination therapy group were lighter than mice in the PD-1 antibody group, indicating that CHK inhibitors may be more toxic than PD-1 antibodies, while the combination therapy did not increase toxicity. In fact, mice in the combination therapy group were even slightly heavier than mice in the CHK inhibitor group.

[0348] according to Figure 2 and Figure 3 In the solvent group, tumors continued to grow, while in the PD-1 antibody group, tumor size began to decrease on day 27. Tumors in the CHK inhibitor group stopped growing from day 15, and tumors in the combination therapy group began to shrink on day 15 and were significantly smaller and lighter than those in other groups by day 27. One mouse in the combination therapy group experienced almost complete tumor regression. All of this indicates that CHK inhibitors act on tumors very rapidly and have very strong anti-tumor activity; the addition of PD-1 antibodies further enhances the anti-tumor effect without introducing additional toxicity.

[0349] The tumor growth inhibition rate was calculated based on the tumor size and weight on day 27 and is summarized in Table 2 below.

[0350] Table 2. Tumor growth inhibition rate

[0351] Grouping TGI - Tumor Size TGI - Tumor Weight solvent Anti-m-PD-1 antibody 30.93％ 33.23％ Compound 6 72.74％ 77.53％ Anti-m-PD-1 antibody + compound 6 94.34％ 94.59％

[0352] The average Q values ​​based on tumor size-related TGI and tumor weight-related TGI were 1.16 and 1.11, respectively, which were higher or slightly lower than 1.15.

[0353] The above data indicate that 1) CHK inhibitor compound 6 acts rapidly on tumors and has a relatively strong effect; and 2) CHK inhibitors and PD-1 antibodies work synergistically to achieve enhanced anti-tumor effects, with comparable or slightly lower toxicity.

[0354] Example 2. Combination therapy using low-dose CHK inhibitors has good anti-tumor effects.

[0355] In this embodiment, the combined antitumor effect of low-dose compound 6 with PD-1 antibody was investigated. Furthermore, the antitumor effects of three compounds obtained by structurally modifying compound 6—compound 6-1, compound 6-2, and compound 6-3—were tested, either alone or in combination with PD-1 antibody.

[0356] On day 0, 105 female BALB / c mice, 4-5 weeks old, were subcutaneously injected with approximately 200.0 μL of a solution containing approximately 5 × 10⁻⁶ ppm of sodium hydroxide. 5 CT-26WT cell culture medium.

[0357] On day 5, the 80 tumor-bearing mice were randomly divided into 10 groups of 8 mice each, based on their body weight. The remaining mice were euthanized.

[0358] Ten groups of animals were administered 1.0 mg / mL of anti-m-PD-1 antibody (BE0146, BioXcell) in PBS (21-040-CVR, BioXcell), 1.5 mg / mL of compound 6 in saline, 1.5 mg / mL of compound 6-1 in saline, 1.5 mg / mL of compound 6-2 in saline, 1.5 mg / mL of compound 6-3 in saline, anti-m-PD-1 antibody + compound 6, anti-m-PD-1 antibody + compound 6-1, anti-m-PD-1 antibody + compound 6-2, anti-m-PD-1 antibody + compound 6-3, and control solvent (saline), respectively. The dosages are shown in Table 3 below.

[0359] The mice's physiological condition and activity level were observed daily, and their body weight and tumor volume were measured every Tuesday, Thursday, and Sunday. Tumor volume (V) was calculated as (length × width). 2 ) / 2.

[0360] On day 23, mice were euthanized, and tumors were collected, weighed, and photographed. Mouse body weight, tumor size, and tumor weight were analyzed using t-tests in SPSS. A statistically significant difference between groups was considered to be below 0.05. Tumor growth inhibition (TGI) based on tumor weight and the Kim Jong-jun Q-value were also calculated.

[0361] Table 3. Dosing Regimen

[0362]

[0363] The average mouse body weight and average tumor size from day 9 to day 23 are summarized in Tables 4 and 5. The average tumor weight, TGI, and Kim Jong-jun Q value for individuals and groups at day 23 are shown in Tables 6 and 7.

[0364] The animals in the solvent group appeared very active, and the physiological state and activity levels of the other groups were also normal. As shown in Table 4, from day 9 to day 23, the body weight of the mice in each group generally increased, partly due to tumor growth. There were no statistically significant differences between the solvent group and the drug administration groups, between the PD-1 antibody group and the combination groups, between each compound group and the corresponding combination group, or between compound 6 and compound 6-1, 6-2, or 6-3 groups.

[0365] Table 4. Mean mouse body weight in solvent group and drug administration group

[0366]

[0367] After deducting tumor weight, the average mouse weights in the 10 groups on day 23 were 18.19g, 18.16g, 17.63g, 17.77g, 18.04g, 18.16g, 17.76g, 17.27g, 17.51g, and 17.67g, respectively. It can be seen that the mouse weights in the solvent group, the anti-m-PD-1 antibody group, and the compound 6-3 group appeared to be similar and higher than the other groups; compound 6, even at lower doses, may still be toxic, but its combination therapy with the PD-1 antibody did not increase toxicity.

[0368] Table 5. Mean tumor size in mice in the solvent group and the drug administration group

[0369]

[0370] According to Table 5, all four compounds, alone or in combination with the PD-1 antibody, inhibited tumor growth. In most groups, tumors continued to grow until day 23, while in the compound 6 group, tumor size began to decrease on day 23, and in the PD-1 antibody + compound 6 group, it began to decrease on day 22.

[0371] At day 23, there was no significant difference in tumor size between the compound 6 group and the compound 6-1, 6-2, or 6-3 groups, or between the compound 6 + PD-1 antibody group and the PD-1 antibody + compound 6-1 group, or the PD-1 antibody + compound 6-2 group. Compared to PD-1 antibodies alone, the combination of compounds 6-1, 6-2, or 6-3 with PD-1 antibodies provides better anti-tumor efficacy.

[0372] Specifically, tumors in the PD-1 antibody + compound 6 group began to grow slowly on day 16 and began to decrease in size from day 22. The average tumor size in this group on day 23 was significantly smaller than that in the compound 6 group, the PD-1 antibody + compound 6-3 group, and the PD-1 antibody group.

[0373] Table 6 shows the antitumor effects of the four compounds, alone or in combination with the anti-m-PD-1 antibody, as measured by tumor weight, with each combination therapy group providing significantly better effects than their respective individual compound groups. Furthermore, although no difference was found between the compound 6 group and the compound 6-1 / 6-2 / 6-3 groups, mice treated with anti-m-PD-1 antibody + compound 6 had significantly lighter tumors compared to the combination of anti-m-PD-1 antibody with compound 6-1, compound 6-2, or compound 6-3. The anti-m-PD-1 antibody inhibited tumor growth in some mice but was ineffective in most. In the anti-m-PD-1 antibody + compound 6 group, tumors almost disappeared in three mice (mice NO.: 2, 7, and 8), and the tumors were quite small in two other mice (mice NO.: 3 and 6).

[0374] Table 6. Individual and group mean tumor weight in mice in the solvent and drug administration groups

[0375]

[0376] According to Table 7, all four compounds synergistically interacted with the anti-m-PD-1 antibody, and the anti-m-PD-1 antibody + compound 6 group had the highest TGI among all groups.

[0377] Table 7. TGI and Jinzhengjun Q-value based on tumor weight in the control and treatment groups

[0378]

[0379] The data above indicate that: 1) when used in combination with PD-1 antibodies, low-dose compound 6 provides good anti-tumor effects, resulting in enhanced anti-tumor efficacy, while having comparable or slightly lower toxicity; 2) modified compounds derived from compound 6, namely compounds 6-1, 6-2, and 6-3, also synergize with PD-1 antibodies; and 3) when modified compounds derived from compound 6 are administered together with PD-1 antibodies, their efficacy in inhibiting tumor growth is lower than that of compound 6.

[0380] Example 3. In mPD-1 - / - mPD-L1 - / - hPD-1 + / + hPD-L1 + / + Combination therapy in transgenic mouse models

[0381] MC38 cells were kept at 37°C and 5% CO2 in 10-cm Piper dishes supplemented with 10% FBS (10270-106, GIBCO, ThermoFisher Scientific), 100 U / ml penicillin, and 100 U / ml streptomycin.

[0382] For female C57BL / 6-Pdcd1 em1(hPDCD1)Cd274 em1(hPD-L1) / Smoc Mice, 20-25g (Shanghai Southern Model Biotechnology), were subcutaneously injected with approximately 3×10 5 MC38 cells.

[0383] Table 8. Dosing Regimen

[0384]

[0385] Q2D: Every two days

[0386] When the tumor grows to 50-70mm 3 These mice were randomly divided into 4 groups of 8 mice each, and this day was designated as day 0. Starting from day 0, according to the dosing regimen in Table 8 above, the animals were administered 40.0 mg / mL of PD-1 antibody (teriprelimab, Shanghai Junshi Biosciences Co., Ltd., China), 3.3 mg / mL of hIgG4 (Novoprotein, NovoPro Bioscience Inc.) in saline, 8 mg / mL of compound 6+ PD-1 antibody in saline, and compound 6+ hIgG4, respectively.

[0387] The mice's physiological condition and activity level were observed daily, and their body weight and tumor volume were measured twice a week. Tumor volume (V) was calculated as (length × width). 2 ) / 2.

[0388] Mice in groups 1 and 2 were euthanized on day 14, and mice in groups 3 and 4 were euthanized on day 21. Tumors were collected and photographed. Mouse body weight and tumor size were analyzed using t-tests in SPSS. A p-value below 0.05 was considered statistically significant between groups. TGI and Kim Jung-kyun Q-value were calculated.

[0389] All animals appeared to be active, and no mice died during the experiment. The average mouse weights for each group are shown below. Figure 4 The group median and individual tumor volume from day 1 to day 14 are shown in the figure. Figure 5 and Figure 6 Individual tumor sizes in all groups on day 14 are shown. Figure 7 Tumors in groups 3 and 4 were observed on day 21. Figure 8 .

[0390] like Figure 4 As shown, from day 1 to day 14, the body weight of mice in all groups generally increased, partly due to tumor growth. There were no significant differences among the four groups, but the average weight of group 2 tended to be higher.

[0391] according to Figure 5 and Figure 6In most mice, tumors grew from day 1 to day 14, while some mice in group 4 maintained smaller tumor sizes. The median tumor volume in group 2 was no different from group 1, while the median tumor volume in group 4 was lower than the other three groups on day 14, further demonstrating... Figure 7 All of this indicates that compound 6, at a dose so low that it provides no antitumor activity when used in combination with hIgG1, can provide a better antitumor effect when used in combination with a PD-1 antibody.

[0392] In addition, such as Figure 8 As shown, mice in group 4 had smaller tumors on day 21 than mice in group 3.

[0393] The mean tumor size on day 0 and day 14, and the TGI on day 14, are summarized in Table 9 below. The Kim Jong-jun Q value calculated based on TGI is 1.25.

[0394] Table 9. Tumor size and tumor growth inhibition rate

[0395]

[0396] The data above indicate that compound 6, even at doses that do not show anti-tumor effects on its own, synergizes with PD-1 antibodies.

[0397] Example 4. Combination therapy in a transgenic mouse model

[0398] Using a low dose of PD-1 antibody, i.e., 2.5 mg / kg of PD-1 antibody, the test in Example 3 was repeated in the same transgenic mouse model, with some other minor variations.

[0399] Specifically, for female C57BL / 6-Pdcd1 em1(hPDCD1) Cd274 em1(hPD-L1) / Smoc Mice (Shanghai Southern Model Biotechnology Co., Ltd.), 5-7 weeks old, were subcutaneously injected with approximately 3×10 5 MC38 cells.

[0400] When the tumor grows to 50-100mm 3 These mice were randomly divided into 4 groups of 8 mice each, and this day was designated as day 0. Starting from day 0, the animals were administered PD-1 antibody (teriprelimab, Shanghai Junshi Biosciences Co., Ltd., China) at 40 mg / mL in 5% glucose solution, hIgG4 (Novoprotein, NovoPro Bioscience Inc.) at 3.3 mg / mL in 5% glucose solution, compound 6+ PD-1 antibody at 8 mg / mL in 5% glucose solution, and compound 6+ hIgG4, respectively, according to the dosing regimens shown in Table 10 below.

[0401] Table 10. Dosing Regimen

[0402]

[0403] BIW: Twice a week

[0404] The tumor size growth curve was obtained and is shown in... Figure 9 The TGI values ​​of the groups treated with compound 6+hIgG4, PD-1 antibody, and compound 6+PD-1 antibody were 4.30%, 46.95%, and 70.53%, respectively, with a Jinzhengjun Q value of 1.43.

[0405] The data and results were similar to those of Example 3. Compound 6, when used at a dose that did not show antitumor effects when used alone, synergized with the PD-1 antibody.

[0406] Example 5. The combination of CHK inhibitor, PD-1 antibody and gemcitabine has a synergistic anti-tumor effect.

[0407] MC38 cells were maintained at 37°C and 5% CO2 in RPMI 1640 medium (10-040-CV, Corning Cellgro) supplemented with 10% FBS (10270-106, GIBCO) and used before 10 passages.

[0408] For female C57BL / 6-Pdcdl em1(hPDCD1) Cd274 em1(hPD-L1) / Smoc Mice (Shanghai Southern Model Biotechnology), 20-25g, were subcutaneously injected with approximately 5×10⁻⁶ mg / L. 5 MC38 cells.

[0409] When the tumor grows to 50-70mm 3 The mice were randomly divided into 8 groups of 8 mice each, and this day was designated as day 9. Starting from day 9, the animals were administered the following medications according to the dosing regimens listed in Table 11: 20 mg / mL gemcitabine (GEM) in saline, 40.0 mg / mL PD-1 antibody (teriprelimab, Shanghai Junshi Biosciences Co., Ltd., China) in saline, 8 mg / mL compound 6 in saline, compound 6 + gemcitabine, gemcitabine + PD-1 antibody, compound 6 + PD-1 antibody, compound 6 + gemcitabine + PD-1 antibody, and control solvent (5% GS).

[0410] Observe the mice's physiological condition and activity level daily, and measure their weight and tumor volume every other day. Tumor volume (V) is calculated as (length × width). 2 ) / 2.

[0411] Table 11. Dosing Regimen

[0412]

[0413] QW: Weekly; Q5D: Every 5 Days

[0414] Mouse body weight and tumor size were analyzed using t-tests in SPSS. A p-value below 0.05 was considered statistically significant between groups.

[0415] Tumor growth inhibition (TGI) is calculated using the following formula.

[0416] Tumor growth inhibition = (average tumor size in the solvent group - average tumor size in the drug treatment group) / average tumor size in the solvent group × 100%

[0417] Calculate the Q value of Kim Jong-jun as described above.

[0418] The animals in the solvent group appeared vigorous, and no mouse deaths were observed in any group during the experiment. The average tumor size from day 9 to day 23 in different groups is shown below. Figure 10 It can be seen that the tumor size of mice in groups 1 to 4 generally increased from day 9 to day 23, while in group 5, the tumor size stopped growing at day 22 and thereafter. Tumor growth in groups 6 to 8 was under control. The data indicate that compound 6, in combination with GEM, PD-1 antibody, or both, exhibits superior antitumor activity compared to single therapy or the GEM+PD-1 antibody combination. Among the three groups with the best antitumor effects, group 8 had the smallest average tumor size, significantly smaller than the other two groups, indicating the good efficacy of the three-drug combination.

[0419] Table 12. Tumor growth inhibition and Jin Zhengjun Q value

[0420]

[0421] *Due to the limitations of evaluative methods, combination therapy with three compounds was evaluated using different permutations and combinations.

[0422] Based on tumor size on day 23, the TGI for groups 2 through 8 were determined to be 30.02%, 17.05%, 52.53%, 67.75%, 37.74%, 80.83%, and 92.22%, respectively.

[0423] The Q value of Kim Jong-jun was calculated and summarized in Table 12 above.

[0424] It can be seen that the combination of compound 6 with PD-1 antibody, or PD-1 antibody with low-dose gemcitabine, has a synergistic effect, which was not observed in other combinations.

Claims

1. The use of a compound of formula I or a pharmaceutically acceptable salt thereof, and an immunotherapeutic agent or a therapeutic agent targeting a cancer-promoting molecule, or a compound of formula I or a pharmaceutically acceptable salt thereof, an immunotherapeutic agent or a therapeutic agent targeting a cancer-promoting molecule, and a chemotherapeutic agent in the preparation of a medicament for treating cancer. Among them, immunotherapeutic agents or therapeutic agents targeting cancer-promoting molecules are antibodies targeting PD-1 or PD-L1. Where Y is NH, O, S or CH2; R 1 Selected from: ， Where X is CH2 or NH. R 8 For H, R 2 It is H or C1-C6 alkyl, and R 3 for , Where R 16 and R 17 One of them is F, Cl, Br, or I, and the other is H, F, Cl, Br, or I. The cancers selected are lung cancer, breast cancer, skin cancer, bladder cancer, colon cancer, head and neck cancer, stomach cancer, kidney cancer, liver cancer, and Hodgkin's lymphoma.

2. The use according to claim 1, wherein Y is NH.

3. The use according to claim 1, wherein Y is O.

4. The use according to claim 1, wherein Y is S.

5. The use according to claim 1, wherein Y is CH2.

6. The use according to claim 1, wherein the compound of formula I is selected from: 2-(3-Fluorophenyl)-4-(3-piperidinamino)-thieno[2,3-d]pyridazine-7-carboxylic acid amide, 2-(3-Fluorophenyl)-4-(3-piperidinoxy)-thieno[2,3-d]pyridazine-7-carboxylic acid amide, 2-(3-Fluorophenyl)-4-(3-piperidinthio)-thieno[2,3-d]pyridazine-7-carboxylic acid amide, 2-(3,5-Dichlorophenyl)-4-(3-piperidinmethyl)-thieno[2,3-d]pyridazine-7-carboxamide, 2-(4-fluorophenyl)-4-(3-piperidineamino)-thieno[2,3-d]pyridazine-7-carboxylic acid amide, 2-(4-chlorophenyl)-4-(3-piperidineamino)-thieno[2,3-d]pyridazine-7-carboxylic acid amide, 2-(4-bromophenyl)-4-(3-piperidineamino)-thieno[2,3-d]pyridazine-7-carboxylic acid amide, 2-(4-chlorophenyl)-4-(2-piperidin-amino)-thieno[2,3-d]pyridazine-7-carboxylic acid amide, 2-(4-chlorophenyl)-4-(S-3-piperidin-amino)-thieno[2,3-d]pyridazine-7-carboxylic acid amide, 2-(4-chlorophenyl)-4-(R-3-piperidin-amino)-thieno[2,3-d]pyridazine-7-carboxylic acid amide, 2-(4-chlorophenyl)-4-(3-piperidin-amino)-thieno[2,3-d]pyridazine-7-N-methylcarboxylate, 2-(4-fluorophenyl)-4-(3-pyridine-amino)-thieno[2,3-d]pyridazine-7-carboxylic acid amide, 2-(4-chlorophenyl)-4-(3-pyridin-amino)-thieno[2,3-d]pyridazine-7-carboxylic acid amide, 2-(4-bromophenyl)-4-(3-pyridine-amino)-thieno[2,3-d]pyridazine-7-carboxylic acid amide, 2-(4-chlorophenyl)-4-(2-pyridin-amino)-thieno[2,3-d]pyridazine-7-carboxylic acid amide, 2-(4-chlorophenyl)-4-(4-pyridin-amino)-thieno[2,3-d]pyridazine-7-carboxylic acid amide, 2-(4-chlorophenyl)-4-(S-3-pyridin-amino)-thieno[2,3-d]pyridazine-7-carboxylic acid amide, 2-(4-chlorophenyl)-4-(R-3-pyridin-amino)-thieno[2,3-d]pyridazine-7-carboxylic acid amide, 2-(4-chlorophenyl)-4-(3-pyridin-amino)-thieno[2,3-d]pyridazine-7-N-methylcarboxylate 2-(3-Chlorophenyl)-4-(3-piperidinamino)-thieno[2,3-d]pyridazine-7-carboxylic acid amide, 2-(3-Bromophenyl)-4-(3-piperidineamino)-thieno[2,3-d]pyridazine-7-carboxylic acid amide, 2-(3-fluorophenyl)-4-(3-pyridin-amino)-thieno[2,3-d]pyridazine-7-carboxylic acid amide, 2-(3-Chlorophenyl)-4-(3-pyridine-amino)-thieno[2,3-d]pyridazine-7-carboxylic acid amide, 2-(3-Bromophenyl)-4-(3-pyridine-amino)-thieno[2,3-d]pyridazine-7-carboxylic acid amide, 2-(3-chlorophenyl)-4-(2-pyridine-amino)-thieno[2,3-d]pyridazine-7-carboxylic acid amide, 2-(3-chlorophenyl)-4-(4-pyridine-amino)-thieno[2,3-d]pyridazine-7-carboxylic acid amide, 2-(3-chlorophenyl)-4-(S-3-pyridine-amino)-thieno[2,3-d]pyridazine-7-carboxylic acid amide, 2-(3-chlorophenyl)-4-(R-3-pyridine-amino)-thieno[2,3-d]pyridazine-7-carboxylic acid amide, 2-(3-Chlorophenyl)-4-(3-pyridin-amino)-thieno[2,3-d]pyridazine-7-N-methylcarboxylate, 2-(4-fluorophenyl)-4-(3-piperidinmethyl)-thieno[2,3-d]pyridazine-7-carboxamide, 2-(4-chlorophenyl)-4-(3-piperidinmethyl)-thieno[2,3-d]pyridazine-7-carboxamide, 2-(4-bromophenyl)-4-(3-piperidinmethyl)-thieno[2,3-d]pyridazine-7-carboxamide, 2-(4-chlorophenyl)-4-(2-piperidinmethyl)-thieno[2,3-d]pyridazine-7-carboxamide, 2-(4-chlorophenyl)-4-(4-piperidinmethyl)-thieno[2,3-d]pyridazine-7-carboxamide, 2-(4-chlorophenyl)-4-(S-3-piperidinmethyl)-thieno[2,3-d]pyridazine-7-carboxamide, 2-(4-chlorophenyl)-4-(R-3-piperidinmethyl)-thieno[2,3-d]pyridazine-7-carboxamide, 2-(4-chlorophenyl)-4-(3-piperidinylmethyl)-thieno[2,3-d]pyridazine-7-N-methyl-formamide, 2-(4-fluorophenyl)-4-(3-pyridylmethyl)-thieno[2,3-d]pyridazine-7-carboxamide, 2-(4-chlorophenyl)-4-(3-pyridylmethyl)-thieno[2,3-d]pyridazine-7-carboxamide, 2-(4-bromophenyl)-4-(3-pyridylmethyl)-thieno[2,3-d]pyridazine-7-carboxamide, 2-(4-chlorophenyl)-4-(2-pyridylmethyl)-thieno[2,3-d]pyridazine-7-carboxamide, 2-(4-chlorophenyl)-4-(4-pyridylmethyl)-thieno[2,3-d]pyridazine-7-carboxamide, and 2-(4-chlorophenyl)-4-(3-pyridylmethyl)-thieno[2,3-d]pyridazine-7-N-methylformamide.

7. The use according to claim 6, wherein the compound of formula I is selected from: 2-(3-Fluorophenyl)-4-(3-piperidinamino)-thieno[2,3-d]pyridazine-7-carboxylic acid amide, 2-(3-Fluorophenyl)-4-(3-piperidinoxy)-thieno[2,3-d]pyridazine-7-carboxylic acid amide, 2-(3-fluorophenyl)-4-(3-piperidinthio)-thieno[2,3-d]pyridazine-7-carboxylate, and 2-(3,5-Dichlorophenyl)-4-(3-piperidinylmethyl)-thieno[2,3-d]pyridazine-7-carboxamide.

8. The use according to claim 1, wherein the antibody targeting PD-1 is selected from nivolumab, pembrolizumab, and toripalimab.

9. The use according to claim 1, wherein the antibody targeting PD-L1 is selected from atezolizumab, durvalumab, and averumab.

10. The use according to claim 1, wherein the chemotherapeutic agent is selected from cisplatin, pemetrexed, gemcitabine, cytarabine, hydroxyurea, temozolomide, irinotecan, cyclophosphamide, mitoxantrone, etoposide, leucovorin, fludarabine, and fluorouracil.

11. The use according to claim 10, wherein the chemotherapeutic agent is gemcitabine.

12. A pharmaceutically acceptable composition comprising a compound of formula I or a pharmaceutically acceptable salt thereof, and an immunotherapeutic agent or a therapeutic agent targeting a cancer-promoting molecule. Among them, immunotherapeutic agents or therapeutic agents targeting cancer-promoting molecules are antibodies targeting PD-1 or PD-L1. Where Y is NH, O, S or CH2; R 1 Selected from: ， Where X is CH2 or NH. R 8 For H, R 2 It is H or C1-C6 alkyl, and R 3 for , Where R 16 and R 17 One of them is F, Cl, Br or I, and the other is H, F, Cl, Br or I.

13. The composition according to claim 12, wherein Y is NH.

14. The composition according to claim 12, wherein Y is O.

15. The composition according to claim 12, wherein Y is S.

16. The composition according to claim 12, wherein Y is CH2.

17. The composition according to claim 12, wherein the compound of formula I is selected from: 2-(3-Fluorophenyl)-4-(3-piperidinamino)-thieno[2,3-d]pyridazine-7-carboxylic acid amide, 2-(3-Fluorophenyl)-4-(3-piperidinoxy)-thieno[2,3-d]pyridazine-7-carboxylic acid amide, 2-(3-Fluorophenyl)-4-(3-piperidinthio)-thieno[2,3-d]pyridazine-7-carboxylic acid amide, 2-(3,5-Dichlorophenyl)-4-(3-piperidinmethyl)-thieno[2,3-d]pyridazine-7-carboxamide, 2-(4-fluorophenyl)-4-(3-piperidineamino)-thieno[2,3-d]pyridazine-7-carboxylic acid amide, 2-(4-chlorophenyl)-4-(3-piperidineamino)-thieno[2,3-d]pyridazine-7-carboxylic acid amide, 2-(4-bromophenyl)-4-(3-piperidineamino)-thieno[2,3-d]pyridazine-7-carboxylic acid amide, 2-(4-chlorophenyl)-4-(2-piperidin-amino)-thieno[2,3-d]pyridazine-7-carboxylic acid amide, 2-(4-chlorophenyl)-4-(S-3-piperidin-amino)-thieno[2,3-d]pyridazine-7-carboxylic acid amide, 2-(4-chlorophenyl)-4-(R-3-piperidin-amino)-thieno[2,3-d]pyridazine-7-carboxylic acid amide, 2-(4-chlorophenyl)-4-(3-piperidin-amino)-thieno[2,3-d]pyridazine-7-N-methylcarboxylate, 2-(4-fluorophenyl)-4-(3-pyridine-amino)-thieno[2,3-d]pyridazine-7-carboxylic acid amide, 2-(4-chlorophenyl)-4-(3-pyridin-amino)-thieno[2,3-d]pyridazine-7-carboxylic acid amide, 2-(4-bromophenyl)-4-(3-pyridine-amino)-thieno[2,3-d]pyridazine-7-carboxylic acid amide, 2-(4-chlorophenyl)-4-(2-pyridin-amino)-thieno[2,3-d]pyridazine-7-carboxylic acid amide, 2-(4-chlorophenyl)-4-(4-pyridin-amino)-thieno[2,3-d]pyridazine-7-carboxylic acid amide, 2-(4-chlorophenyl)-4-(S-3-pyridin-amino)-thieno[2,3-d]pyridazine-7-carboxylic acid amide, 2-(4-chlorophenyl)-4-(R-3-pyridin-amino)-thieno[2,3-d]pyridazine-7-carboxylic acid amide, 2-(4-chlorophenyl)-4-(3-pyridin-amino)-thieno[2,3-d]pyridazine-7-N-methylcarboxylate 2-(3-Chlorophenyl)-4-(3-piperidinamino)-thieno[2,3-d]pyridazine-7-carboxylic acid amide, 2-(3-Bromophenyl)-4-(3-piperidineamino)-thieno[2,3-d]pyridazine-7-carboxylic acid amide, 2-(3-fluorophenyl)-4-(3-pyridin-amino)-thieno[2,3-d]pyridazine-7-carboxylic acid amide, 2-(3-Chlorophenyl)-4-(3-pyridine-amino)-thieno[2,3-d]pyridazine-7-carboxylic acid amide, 2-(3-Bromophenyl)-4-(3-pyridine-amino)-thieno[2,3-d]pyridazine-7-carboxylic acid amide, 2-(3-chlorophenyl)-4-(2-pyridine-amino)-thieno[2,3-d]pyridazine-7-carboxylic acid amide, 2-(3-chlorophenyl)-4-(4-pyridine-amino)-thieno[2,3-d]pyridazine-7-carboxylic acid amide, 2-(3-chlorophenyl)-4-(S-3-pyridine-amino)-thieno[2,3-d]pyridazine-7-carboxylic acid amide, 2-(3-chlorophenyl)-4-(R-3-pyridine-amino)-thieno[2,3-d]pyridazine-7-carboxylic acid amide, 2-(3-Chlorophenyl)-4-(3-pyridin-amino)-thieno[2,3-d]pyridazine-7-N-methylcarboxylate, 2-(4-fluorophenyl)-4-(3-piperidinmethyl)-thieno[2,3-d]pyridazine-7-carboxamide, 2-(4-chlorophenyl)-4-(3-piperidinmethyl)-thieno[2,3-d]pyridazine-7-carboxamide, 2-(4-bromophenyl)-4-(3-piperidinmethyl)-thieno[2,3-d]pyridazine-7-carboxamide, 2-(4-chlorophenyl)-4-(2-piperidinmethyl)-thieno[2,3-d]pyridazine-7-carboxamide, 2-(4-chlorophenyl)-4-(4-piperidinmethyl)-thieno[2,3-d]pyridazine-7-carboxamide, 2-(4-chlorophenyl)-4-(S-3-piperidinmethyl)-thieno[2,3-d]pyridazine-7-carboxamide, 2-(4-chlorophenyl)-4-(R-3-piperidinmethyl)-thieno[2,3-d]pyridazine-7-carboxamide, 2-(4-chlorophenyl)-4-(3-piperidinylmethyl)-thieno[2,3-d]pyridazine-7-N-methyl-formamide, 2-(4-fluorophenyl)-4-(3-pyridylmethyl)-thieno[2,3-d]pyridazine-7-carboxamide, 2-(4-chlorophenyl)-4-(3-pyridylmethyl)-thieno[2,3-d]pyridazine-7-carboxamide, 2-(4-bromophenyl)-4-(3-pyridylmethyl)-thieno[2,3-d]pyridazine-7-carboxamide, 2-(4-chlorophenyl)-4-(2-pyridylmethyl)-thieno[2,3-d]pyridazine-7-carboxamide, 2-(4-chlorophenyl)-4-(4-pyridylmethyl)-thieno[2,3-d]pyridazine-7-carboxamide, and 2-(4-chlorophenyl)-4-(3-pyridylmethyl)-thieno[2,3-d]pyridazine-7-N-methylformamide.

18. The composition according to claim 17, wherein the compound of formula I is selected from: 2-(3-Fluorophenyl)-4-(3-piperidinamino)-thieno[2,3-d]pyridazine-7-carboxylic acid amide, 2-(3-Fluorophenyl)-4-(3-piperidinoxy)-thieno[2,3-d]pyridazine-7-carboxylic acid amide, 2-(3-fluorophenyl)-4-(3-piperidinthio)-thieno[2,3-d]pyridazine-7-carboxylate, and 2-(3,5-Dichlorophenyl)-4-(3-piperidinylmethyl)-thieno[2,3-d]pyridazine-7-carboxamide.

19. The composition of claim 12, wherein the antibody targeting PD-1 is selected from nivolumab, pembrolizumab, and toripalimab.

20. The composition of claim 12, wherein the antibody targeting PD-L1 is selected from atezolizumab, durvalumab, and averumab.

21. The composition of claim 12, further comprising a chemotherapeutic agent.

22. The composition of claim 21, wherein the chemotherapeutic agent is selected from cisplatin, pemetrexed, gemcitabine, cytarabine, hydroxyurea, temozolomide, irinotecan, cyclophosphamide, mitoxantrone, etoposide, leucovorin, fludarabine, and fluorouracil.

23. The composition according to claim 21 or 22, wherein the chemotherapeutic agent is gemcitabine.

24. The composition according to claim 21 or 22, wherein the chemotherapeutic agent is cisplatin.

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