Crystal form of ENPP1 inhibitors

Abstract

The present invention provides a compound represented by formula (I) in crystalline form, its specific crystalline form, a pharmaceutical composition containing the same, and its use. The compound represented by formula (I) in crystalline form and its specific crystalline form have good crystallinity and chemical purity, are easy to manufacture, have high yields, and have the potential to become excellent pharmaceuticals. [Formula 1] JPEG2026519885000016.jpg47170

Description

[Technical Field]

[0001] This application claims priority to the prior application No. 202310706966.6, filed with the China National Intellectual Property Administration on June 15, 2023, with the title of the invention being "Crystal Form of ENPP1 Inhibitor." The full text of the said application is incorporated into this application by reference.

[0002] This invention belongs to the field of medicinal chemistry and discloses crystalline forms of ENPP1 inhibitors, and also includes the application of these crystalline forms in the manufacture of pharmaceuticals for the treatment of diseases related to ENPP1. [Background technology]

[0003] ENPP1 is a type II transmembrane glycoprotein with nucleotide pyrophosphatase and phosphodiesterase activity, belonging to the extracellular nucleotide pyrophosphatase / phosphodiesterase (Enpp) family, which consists of seven proteins with different functions (1-7). The ENPP1 structure has two N-terminal SMB domains (SMBENPP11 and SMB2), two linker regions (L1 and L2), one catalytic domain, and one nuclease-like domain. ENPP1 exhibits different expression in immune cells, being expressed at low levels in NK cells, dendritic cells (DCs), and macrophages, and at high levels in neutrophils. ENPP1 is also expressed in a small subset of B cells, which may be involved in regulating T cell activity. ENPP1 expression is elevated in M2 subtype macrophages, which play a role in tumorigenesis. ENPP1 expression is increased in astrocytic cell tumors, breast cancer, and head and neck tumors. ENPP1 expression varies significantly among different tumor tissues. ENPP1 is specifically overexpressed in some tumors and may be a driving factor for tumor immune evasion and metastasis.

[0004] ENPP1 plays a crucial role in immune responses to various stimuli via the STING pathway, selectively activating the STING signaling pathway in tumor cells and other cells in the tumor microenvironment, achieving high selectivity. ENPP1 also catalyzes the hydrolysis of ATP into PPi and AMP, promoting the production of adenosine, which has a potent immunosuppressive effect. Inhibition of ENPP1 can relieve tumor immunosuppression by inhibiting ATP hydrolysis and reducing adenosine production.

[0005] STING agonists non-selectively activate STING in cancer cells and host cells, while ENPP1 restricts the scope of STING activation to tumor tissue and the tumor microenvironment, thereby enhancing anti-tumor immunity. Furthermore, ENPP1 is selectively elevated in metastatic and chromosomally unstable tumor cells, and systemic administration of ENPP1 inhibitors can hinder the ability of spreading tumor cells to evade immune surveillance, thus avoiding the technical difficulties associated with intratumoral administration of STING agonists.

[0006] The combination of ENPP1 inhibitors and radiotherapy has shown favorable effects in animal models, and has also demonstrated some synergistic effects when combined with multiple antitumor drugs such as immune checkpoint inhibitors like PD-1 and PARP inhibitors. Currently, all ENPP1 inhibitor development is in the preclinical research and development stage, but several pharmaceutical companies have already published patents related to ENPP1 inhibitors, such as WO2021061803A1, WO2021158829A1, WO2020190912A1, WO2019177971A1, and WO2019046778A1.

[0007] While several small molecules of ENPP1 inhibitors have been publicly released using existing technologies, no clinically-grade small molecule drugs have yet reached the market. Therefore, the development of new compounds with market potential and superior efficacy and pharmacokinetic results remains a pressing need. This invention designs a series of compounds with a novel structure represented by the general formula, discovers that compounds with such structures exhibit superior efficacy and action, and demonstrates their significant potential for the development of ENPP1 inhibitors. [Overview of the Initiative]

[0008] According to the first aspect, the present invention relates to the following formula (I): [ka] We provide a compound with a crystalline form represented by formula (I).

[0009] In a preferred embodiment of the present invention, the compound represented by formula (I) in the crystalline form is crystalline form A, which has diffraction peaks at 2θ angles of 24.9±0.2°, 25.1±0.2°, and 25.7±0.2° in an X-ray powder diffraction pattern using Cu-Kα rays.

[0010] In some embodiments of the present invention, crystalline form A of the compound of formula (I) has diffraction peaks at 2θ angles of 12.8±0.2°, 14.0±0.2°, 16.0±0.2°, 18.5±0.2°, 24.9±0.2°, 25.1±0.2°, and 25.7±0.2° in an X-ray powder diffraction pattern using Cu-Kα rays.

[0011] In some embodiments of the present invention, crystalline form A of the compound of formula (I) has diffraction peaks in the X-ray powder diffraction pattern using Cu-Kα rays at 2θ angles of 12.8±0.2°, 14.0±0.2°, 16.0±0.2°, 18.5±0.2°, 24.9±0.2°, 25.1±0.2°, 25.7±0.2°, 26.9±0.2°, and 28.5±0.2°.

[0012] In some embodiments of the present invention, crystalline form A of the compound of formula (I) has diffraction peaks in the X-ray powder diffraction pattern using Cu-Kα rays at 2θ angles of 12.8±0.2°, 14.0±0.2°, 16.0±0.2°, 18.5±0.2°, 24.9±0.2°, 25.1±0.2°, 25.7±0.2°, 26.9±0.2°, 28.5±0.2°, and 29.8±0.2°.

[0013] In some embodiments of the present invention, crystalline form A of the compound of formula (I) has diffraction peaks in the X-ray powder diffraction pattern using Cu-Kα rays at 2θ angles of 12.8±0.2°, 14.0±0.2°, 16.0±0.2°, 17.0±0.2°, 18.5±0.2°, 20.5±0.2°, 24.9±0.2°, 25.1±0.2°, 25.7±0.2°, 26.9±0.2°, 28.5±0.2°, and 29.8±0.2°.

[0014] In some embodiments of the present invention, the crystalline form A of the compound of formula (I) is shown in Figure 1 or Figure 7 as an XRPD pattern using Cu-Kα radiation. In some embodiments of the present invention, the crystalline form A of the compound of formula (I) is shown in Table 1 or Table 2, based on XRPD pattern analysis using Cu-Kα radiation. In some embodiments of the present invention, the A crystalline form of the compound of formula (I) has an endothermic peak at 233.23±4°C in the differential scanning calorimetry curve.

[0015] In some embodiments of the present invention, the A crystalline form of the compound of formula (I) has an exothermic peak at 234.3 ± 4°C in the differential scanning calorimetry curve. In some embodiments of the present invention, the A crystalline form of the compound of formula (I) has a differential scanning calorimetry (DSC) pattern substantially as shown in Figure 2. In some embodiments of the present invention, the thermogravimetric analysis curve of the A crystal form of the compound of formula (I) above shows a weight loss (weight loss rate) of 13.50 ± 1%, for example, 13.5111%, between approximately 105 and 254°C.

[0016] In some embodiments of the present invention, the A crystalline form of the compound of formula (I) exhibits a weight loss of 13.50±1%, for example 13.5111%, at 254.00±3℃ in the thermogravimetric analysis curve. In some embodiments of the present invention, the A crystalline form of the compound of formula (I) has a thermogravimetric analysis (TGA) pattern substantially as shown in Figure 3. The present invention also provides crystalline form B of the compound of formula (I), which has diffraction peaks at 2θ angles of 19.19±0.2°, 27.18±0.2°, 13.44±0.2°, and 22.55±0.2° in its X-ray powder diffraction pattern using Cu-Kα rays.

[0017] In some embodiments of the present invention, crystalline form B of the compound of formula (I) has diffraction peaks in the X-ray powder diffraction pattern using Cu-Kα rays at 2θ angles of 26.19±0.2°, 20.76±0.2°, 4.42±0.2°, 22.84±0.2°, 19.19±0.2°, 27.18±0.2°, 13.44±0.2°, and 22.55±0.2°. In some embodiments of the present invention, the crystalline form B of the compound of formula (I) is shown in Figure 4 as an XRPD pattern using Cu-Kα radiation. In some embodiments of the present invention, the crystalline form B of the compound of formula (I) is shown in Table 3, based on XRPD pattern analysis using Cu-Kα radiation.

[0018] In some embodiments of the present invention, crystalline form B of the compound of formula (I) is the DMF solvate of the compound of formula (I). In some embodiments of the present invention, the ratio of the compound of formula (I) to the DMF solvent in crystalline form B of the compound of formula (I) is 1:1. The present invention also provides crystalline form B of the compound of formula (I), which has diffraction peaks at 2θ angles of 22.11±0.2°, 22.55±0.2°, 20.96±0.2°, and 26.63±0.2° in its X-ray powder diffraction pattern using Cu-Kα rays.

[0019] In some embodiments of the present invention, the crystalline form C of the compound of formula (I) has diffraction peaks at 2θ angles of 22.11±0.2°, 22.55±0.2°, 20.96±0.2°, 26.63±0.2°, 15.01±0.2°, 25.68±0.2°, 26.38±0.2°, 28.54±0.2°, 13.22±0.2°, 18.68±0.2° in the X-ray powder diffraction pattern using Cu-Kα radiation.

[0020] In some embodiments of the present invention, the XRPD pattern of the crystalline form C of the compound of formula (I) using Cu-Kα radiation is shown in FIG. 5. In some embodiments of the present invention, the analysis of the XRPD pattern of the crystalline form C of the compound of formula (I) using Cu-Kα radiation is shown in Table 4. In some embodiments of the present invention, the crystalline form C of the compound of formula (I) is a DMSO solvate of the compound of formula (I).

[0021] The present invention also provides a crystalline form D of the compound of formula (I) in crystalline form, which has diffraction peaks at 2θ angles of 22.71±0.2°, 18.92±0.2°, 22.96±0.2°, 26.4±0.2°, 24.44±0.2°, 16.62±0.2°, 13.15±0.2°, 17.79±0.2, 27.12±0.2° in the X-ray powder diffraction pattern using Cu-Kα radiation.

[0022] In some embodiments of the present invention, the crystalline form D of the compound of formula (I) has diffraction peaks at 2θ angles of 22.71±0.2°, 18.92±0.2°, 22.96±0.2°, 26.4±0.2°, 24.44±0.2°, 16.62±0.2°, 13.15±0.2°, 17.79±0.2°, 27.12±{0.2°}, 24.17±0.2°, 28.81±0.2°, 19.95±0.2°, 28.42±0.2°, 24.85±0.2°, 14.78±0.2°, 17.36±0.2°, 29.16±0.2° in the X-ray powder diffraction pattern using Cu-Kα radiation.

[0023] In some embodiments of the present invention, the crystalline form D of the compound of formula (I) is shown in Figure 6 as an XRPD pattern using Cu-Kα radiation. In some embodiments of the present invention, the crystalline form D of the compound of formula (I) is shown in Table 5, based on XRPD pattern analysis using Cu-Kα radiation. In some embodiments of the present invention, the crystalline form D of the compound of formula (I) is the NMP solvate of the compound of formula (I).

[0024] In some embodiments of the present invention, the ratio of the compound of formula (I) to the NMP solvent in crystalline form D of the compound of formula (I) is 1:1.

[0025] According to a second aspect, the present invention provides a method for producing a compound of formula (I) in the crystalline form described in the first aspect, the production method comprising method 1, in which the compound of formula I is dissolved in an organic solvent A, and the solution is further added to an organic solvent B to precipitate crystals.

[0026] The organic solvent A is selected from one or more of DMF (N,N-dimethylformamide), DMSO (dimethyl sulfoxide), and NMP (N-methylpyrrolidone), and the organic solvent B is selected from one or more of water, methanol, ethanol, MTBE, toluene, and dichloromethane.

[0027] In some embodiments of the present invention, when the compound of formula (I) is crystalline form A, in method 1, organic solvent A is selected from DMF and organic solvent B is selected one or two from water and ethanol, or organic solvent A is selected from DMSO and organic solvent B is selected one or two from toluene and methanol.

[0028] In some embodiments of the present invention, when the compound of formula (I) is crystalline form B, in method 1 above, organic solvent A is selected from DMF, and organic solvent B is selected from one or two of MTBE and toluene.

[0029] In some embodiments of the present invention, when the compound of formula (I) is in crystalline form D, in method 1 above, organic solvent A is selected from NMP, and organic solvent B is selected from one or more of water, dichloromethane, and MTBE.

[0030] The method for producing the compound of formula (I) in the crystalline form includes method 2, in which the compound of formula I is dissolved in an organic solvent C, and then an organic solvent D is added to precipitate crystals. The organic solvent C is selected from one or more of DMF, DMSO, and NMP, and the organic solvent D is selected from one or more of acetone, ethylene glycol methyl ether, water, dichloromethane, ethyl acetate, MTBE, toluene, tetrahydrofuran, dioxane, ethanol, trifluoroethanol, acetonitrile, ethylene glycol dimethyl ether, isopropanol, and methyl ethyl ketone.

[0031] In some embodiments of the present invention, when the compound of formula (I) is crystalline form A, in method 1 above, organic solvent C is selected from DMF and organic solvent D is selected one or more from acetone, ethylene glycol methyl ether, water, and dichloromethane; or organic solvent C is selected from DMSO and organic solvent D is selected one or more from ethanol, trifluoroethanol, ethylene glycol methyl ether, acetonitrile, dioxane, ethylene glycol dimethyl ether, and methyl ethyl ketone; or organic solvent C is selected from NMP and organic solvent D is selected one or more from water, acetonitrile, dichloromethane, and ethylene glycol dimethyl ether.

[0032] In some embodiments of the present invention, when the compound of formula (I) is in crystalline form B, in method 2 above, organic solvent C is selected from DMF, and organic solvent D is selected from one or more of ethyl acetate, MTBE, toluene, tetrahydrofuran, dioxane, and methyl ethyl ketone.

[0033] In some embodiments of the present invention, when the compound of formula (I) is crystalline form C, in method 2 above, the organic solvent C is selected from DMSO, and the organic solvent D is selected one or more from ethyl acetate, water, and toluene.

[0034] In some embodiments of the present invention, when the compound of formula (I) is in crystalline form D, in method 2 above, the organic solvent C is selected from NMP, and the organic solvent D is selected from one or more of isopropanol, MTBE, toluene, tetrahydrofuran, and methyl ethyl ketone.

[0035] The method for producing the compound of formula (I) in the crystalline form is as follows: Method 3 includes suspending the compound of formula I in an organic solvent E, forming a slurry, and filtering to obtain crystals.

[0036] The organic solvent E mentioned above is selected from one or more of the following: methanol, ethanol, isopropanol, ethyl acetate, n-heptane, MTBE, ethylene glycol methyl ether, water, acetonitrile, toluene, dichloromethane, tetrahydrofuran, dioxane, ethylene glycol dimethyl ether, methyl ethyl ketone, dimethylformamide, dimethyl sulfoxide, N-methylpyrrolidone, and acetone.

[0037] In some embodiments of the present invention, when the compound of formula (I) is in crystalline form A, the organic solvent E in method 3 above is selected from one or more of methanol, ethanol, isopropanol, ethyl acetate, n-heptane, MTBE, ethylene glycol methyl ether, water, acetonitrile, toluene, dichloromethane, tetrahydrofuran, dioxane, ethylene glycol dimethyl ether, and methyl ethyl ketone.

[0038] In some embodiments of the present invention, when the compound of formula (I) is in crystalline form B, the organic solvent E in method 3 above is selected from DMF. In some embodiments of the present invention, when the compound of formula (I) is in crystalline form C, the organic solvent E in method 3 above is selected from DMSO. In some embodiments of the present invention, when the compound of formula (I) is of crystalline form D, the organic solvent E in method 3 above is selected from NMP.

[0039] According to a third aspect, the present invention provides a crystalline composition comprising a compound of formula (I) in the crystalline form described in the first aspect. Preferably, in the crystalline composition, one or more compounds of formula (I) in the crystalline form are selected from crystalline form A, crystalline form B, crystalline form C, and crystalline form D.

[0040] In some embodiments of the present invention, the compound of formula (I) in the crystalline form described above accounts for 10% or more, 20% or more, 30% or more, 40% or more, 50% or more, 60% or more, 70% or more, 80% or more, 90% or more, 91% or more, 92% or more, 93% or more, 94% or more, 95% or more, 96% or more, 97% or more, 98% or more, 99% or more, or 99.5% or more of the weight of the crystalline composition.

[0041] Preferably, the compound of formula (I) in the above crystalline form accounts for 50% or more, 60% or more, 70% or more, 80% or more, 90% or more, 91% or more, 92% or more, 93% or more, 94% or more, 95% or more, 96% or more, 97% or more, 98% or more, 99% or more, or 99.5% or more of the weight of the above crystalline composition.

[0042] In some embodiments of the present invention, the crystalline composition comprises crystalline form A of the compound of formula (I). In some embodiments of the present invention, the compound of formula (I) in the crystalline form described above accounts for 10% or more, 20% or more, 30% or more, 40% or more, 50% or more, 60% or more, 70% or more, 80% or more, 90% or more, 91% or more, 92% or more, 93% or more, 94% or more, 95% or more, 96% or more, 97% or more, 98% or more, 99% or more, or 99.5% or more of the weight of the crystalline composition.

[0043] Preferably, the above-mentioned crystal form A accounts for 50% or more, 60% or more, 70% or more, 80% or more, 90% or more, 91% or more, 92% or more, 93% or more, 94% or more, 95% or more, 96% or more, 97% or more, 98% or more, 99% or more, or 99.5% or more of the weight of the above-mentioned crystal composition.

[0044] According to a fourth aspect, the present invention provides a pharmaceutical composition comprising a compound represented by formula (I) in crystalline form and an optional pharmaceutically acceptable carrier. Preferably, in the pharmaceutical composition, the compound of formula (I) in crystalline form is selected from one or more crystalline forms A, B, C, and D. In some embodiments of the present invention, the pharmaceutical composition comprises a crystalline form A of the compound of formula (I) and an optional pharmaceutically acceptable carrier.

[0045] According to a fifth aspect, the present invention provides a pharmaceutical composition comprising the crystallization composition described in the third aspect and an optional pharmaceutically acceptable carrier.

[0046] According to the sixth aspect, the present invention provides a pharmacopoeia for the use of a compound of formula (I) in the crystalline form described in the first aspect, a crystalline composition described in the third aspect, or a pharmaceutical composition described in the fourth aspect, or for use in the manufacture of a pharmaceutical.

[0047] In some embodiments of the present invention, the pharmaceutical agent is used to prevent and / or treat ENPP1-mediated diseases. In some embodiments of the present invention, the pharmaceutical is used to prevent and / or treat cancer or tumor-related diseases, or cardiovascular diseases. In some embodiments of the present invention, the pharmaceutical composition further comprises one, two, or three or more other pharmaceuticals for treating cancer or tumors.

[0048] In some embodiments of the present invention, the pharmaceutical composition is used in combination with one, two, or three or more other pharmaceuticals or therapeutic means for treating cancer or tumors.

[0049] Preferably, the antitumor agent is a chemotherapeutic agent, a targeted therapy agent, or an immunotherapy agent, and preferably, the pharmaceutical used for the prevention and / or treatment of the cancer or tumor is a cell signaling inhibitor, chlorambucil, melphalan, cyclophosphamide, ifosfamide, busulfan, carmustine, lomustine, streptozotocin, cisplatin, carboplatin, oxaliplatin, dacarbazine, temozolomide, procarbazine, methotrexate, fluorouracil, cytarabine, gemcitabine, methotrexate Lucaptopurine, fludarabine, vinblastine, vincristine, vinorelbine, paclitaxel, docetaxel, topotecan, irinotecan, etoposide, trabectedin, dactinomycin, doxorubicin, epirubicin, daunorubicin, mitoxantrone, bleomycin, mitomycin C, isapillone, tamoxifen, flutamide, gonadorelin analog, megestrol, prednisone, dexamethoxazole Metazone, methylprednisolone, thalidomide, interferon α, calcium leucovorate, sirolimus, sirolimus lipids, everolimus, afatinib, alisertib, amuvatinib, apatinib, axitinib, bortezomib, bosutinib, brivanib, cabozantinib, cediranib, crenolanib, crizotinib, dacomitine Dacomitinib, Danusertib, Dasatinib, Dovitinib, Erlotinib, Foretinib, Ganetespib, Gefitinib, Ibrutinib, Icotinib, Imatinib, Iniparib, Lapatinib, Lenvatinib, Linifanib, Linsitinib, Masitinib, Momerotinib, Motesanib,Neratinib, nilotinib, niraparib, oprozomib, olapanib, pazopanib, pictilisib, ponatinib, quizartinib, regorafenib, rigosertib, rucaparib, ruxolitinib, saracatinib, salidegib idegib), sorafenib, sunitinib, telatinib, tivantinib, tivozanib, tofacitinib, trametinib, vandetanib, veliparib, vemurafenib, bismodegib, volasertib, alemtuzumab, bevacizumab, brentuximab The preferred therapeutic agents include, but are not limited to, vedotin, catumaxomab, cetuximab, denosumab, gemtuzumab, ipilimumab, nimotuzumab, ofatumumab, panitumumab, rituximab, tocitumumab, trastuzumab, PI3K inhibitors, CSF1R inhibitors, A2A and / or A2B receptor antagonists, IDO inhibitors, anti-PD-1 antibodies, anti-PD-L1 antibodies, LAG3 antibodies, TIM-3 antibodies, TIGIT antibodies, CD47 antibodies, CLAUDIN 18.2 antibodies, anti-CTLA-4 antibodies, or any combination thereof. Preferred therapeutic agents are radiotherapy or surgery.

[0050] In some embodiments of the present invention, the drug is used to treat skin cancer, bladder cancer, ovarian cancer, breast cancer, gastric cancer, pancreatic cancer, prostate cancer, colon cancer, lung cancer, bone cancer, brain cancer, neurocytoma, rectal cancer, colon cancer, familial adenomatous polyposis, hereditary nonpolyposis colorectal cancer, esophageal cancer, lip cancer, laryngeal cancer, hypopharyngeal cancer, tongue cancer, salivary gland cancer, adenocarcinoma, medullary thyroid cancer, papillary thyroid cancer, kidney cancer, renal parenchymal cancer, cervical cancer, uterine cancer, endometrial cancer, choriocarcinoma, testicular cancer, urinary tract cancer, melanoma, brain tumors (e.g., glioblastoma, astrocytoma, meningioma, medulloblastoma), peripheral ectodermal tumors, Hodgkin lymphoma, non-Hodgkin lymphoma, Burkitt It is used to prevent and / or treat diseases such as trilymphoma, leukemia, acute lymphoblastic leukemia (ALL), chronic lymphocytic leukemia (CLL), acute myeloid leukemia (AML), chronic myeloid leukemia (CML), adult T-cell leukemia / lymphoma, diffuse large B-cell lymphoma (DLBCL), hepatocellular carcinoma, gallbladder cancer, bronchial cancer, small cell lung cancer, non-small cell lung cancer, multiple myeloma, basal cell tumor, teratoma, retinoblastoma, choroidal melanoma, seminomas, rhabdomyosarcoma, craniopharyngioma, osteosarcoma, chondrosarcoma, myosarcoma, liposarcoma, fibrosarcoma, Ewing's sarcoma, and plasmacytoma.

[0051] In some embodiments of the present invention, cardiovascular diseases targeted for treatment and / or prevention include myocardial infarction (MI), heart failure (HF), cardiac trauma, abnormal scar formation of cardiac tissue, cardiomyopathy, cardiomyocyte death, acceleration of cardiac repair, and release of pro-inflammatory molecules from cardiomyocytes.

[0052] [Definition and explanation] Unless otherwise specified, the following terms and expressions in this invention have the following meanings. Unless otherwise specified, any particular term or expression is not considered uncertain or unclear and should be understood as having a general meaning.

[0053] In this invention, "a compound of formula (I) in crystalline form (i.e., a compound represented by formula (I) in crystalline form)" refers to a compound represented by formula (I) that exhibits a crystalline form, and includes the anhydrous, solvent-free form, hydrate form, and solvate form of the compound represented by formula (I). The crystalline form is preferably the anhydrous, solvent-free form or the hydrate form, and more preferably the anhydrous, solvent-free form.

[0054] The term "solvide" or "solvate" refers to an aggregate formed between a solvent molecule in a stoichiometric or non-stoichiometric ratio and a compound represented by formula (I) according to the present invention, and includes aggregates containing water molecules and one or more other solvent molecules simultaneously, and aggregates containing only one or more other solvent molecules.

[0055] The term "hydrate" refers to an aggregate formed between water molecules in stoichiometric or non-stoichiometric ratios and the compound represented by formula (I) according to the present invention.

[0056] "Anhydrous, solvent-free form" refers to a form in which water molecules or solvent molecules are not contained within the unit cell or crystal lattice, or a form in which water molecules or solvent molecules coexist with the compound represented by formula (I) via non-molecular force bonding (e.g., adsorption).

[0057] The term "crystalline composition" refers to a solid form containing crystalline form A as referred to herein. Furthermore, in addition to the crystalline form according to the present invention, the crystalline composition may optionally contain compounds represented by formula (I) in other crystalline or amorphous forms, or salts thereof, or impurities other than these substances. Those skilled in the art should understand that the sum of the content of each component in the crystalline composition should be 100%.

[0058] The "room temperature" mentioned above refers to the temperature of room temperature in the general sense in this field, and is generally between 10 and 30°C, preferably 25°C ± 5°C.

[0059] In X-ray powder diffraction patterns, the terms "substantially" or "substantially as shown in the figure" mean that a substantially pure particular crystal form accounts for at least 50%, or at least 60%, or at least 70%, or at least 80%, or at least 90%, or at least 95%, or at least 96%, or at least 97%, or at least 98%, or at least 99% of the peaks in that X-ray powder diffraction pattern. Furthermore, if the content of a certain crystal form in the product gradually decreases, the diffraction peaks belonging to some of those crystal forms in the X-ray powder diffraction pattern may decrease due to factors related to the detection sensitivity of the instrument. It is also well known in the field of crystallography that there may be a small error in the position of the peaks for any given crystal form. For example, changes in temperature during sample analysis, sample movement, or instrument calibration can cause the position of the peaks to shift, and the measurement error of the 2θ value is approximately ±0.3°, and may typically be around ±0.2°. Therefore, when identifying various crystal structures, such errors should be taken into consideration, and the terms "substantially" or "substantially as shown in the figure" are intended to include such differences in diffraction peak positions, where such differences are ±0.3°, preferably ±0.2°, and preferably ±0.1°.

[0060] In DSC patterns, the term "substantially" or "substantially as shown in the figure" means that, for the same crystalline form of the same compound, in a series of analyses, the error in the thermal transition onset temperature, endothermic peak temperature, exothermic peak temperature, melting point, etc., is typically within approximately 8°C, usually within approximately 5°C, and usually within approximately 3°C. If a compound has a specific thermal transition onset temperature, endothermic peak temperature, exothermic peak temperature, melting point, etc., then this refers to a temperature within ±5°C of that temperature.

[0061] As used herein, the term “prevention” means, when used for a disease or disorder (also called “symptom”) (e.g., a viral disease), that the compound or drug (e.g., a combination product for which protection is claimed in this invention) can reduce the frequency of symptoms of a medical disorder or delay the onset of symptoms in a subject compared to a subject who is not administered the compound or drug.

[0062] As used herein, the term “treatment” means reducing, alleviating or improving the symptoms of a disease or disorder, improving underlying metabolic symptoms, inhibiting a disease or disorder, for example, halting the progression of a disease or disorder, alleviating a disease or disorder, reducing a disease or disorder, mitigating the symptoms of a disease or disorder, or preventing the symptoms of a disease or disorder. The pharmaceutical composition according to the present invention can be manufactured using conventional methods in the art.

[0063] In the context of this specification, the terms “pharmaceutically acceptable carrier” or “excipient” or “pharmaceutically acceptable excipient” refer to an excipient (auxiliary agent) that does not have an apparent irritant effect on an organism and does not impair the biological activity and performance of the active compound. The term “pharmaceutically acceptable excipient” includes solvents, propellants, solubilizers, co-solvents, emulsifiers, colorants, binders, disintegrants, fillers, lubricants, wetting agents, osmotic pressure regulators, stabilizers, fluidizers, flavoring agents, preservatives, suspending agents, coatings, fragrances, anti-stickers, antioxidants, chelating agents, penetration enhancers, pH adjusters, buffers, plasticizers, surfactants, foaming agents, defoamers, thickeners, inclusion agents, humectants, absorbents, diluents, flocculants and dispersants (anti-flocculants), filtration aids, release retarders, etc. Those skilled in the art can select specific pharmaceutically acceptable excipients as needed. Knowledge of excipients is well known to those skilled in the art; for example, one can refer to "Pharmacy" (supervised by Cui Fu-de, 5th edition, People's Health Press, 2003).

[0064] The terms "comprise" and their English variations (e.g., "comprises" and "comprising") should all be interpreted in an open-ended, inclusive sense, meaning "to include, but not limited to,..."

[0065] Within the scope of the present invention, many different embodiments can be constructed by combining various options of any feature with various options of other features. The present invention is intended to include all possible embodiments consisting of various options of all technical features.

[0066] The intermediate compounds according to the present invention can be produced by a variety of synthesis methods well known to those skilled in the art, including the specific embodiments described below and embodiments combined with other chemical synthesis methods, as well as equivalent forms well known to those skilled in the art. Preferred embodiments include, but are not limited to, the examples of the present invention.

[0067] The chemical reactions of specific embodiments of the present invention are achieved in a suitable solvent, which must be compatible with the chemical changes of the present invention and the desired reagents and materials. Those skilled in the art may need to modify or select synthesis procedures or reaction schemes based on existing embodiments to obtain the compounds of the present invention.

[0068] The present invention will be described in detail below with reference to examples, but these examples do not imply any limitation to the present invention.

[0069] All reagents and solvents used in this invention are commercially available and can be used without the need for further purification.

[0070] [Technical effects] The compound of formula (I) of the present invention possesses good PK properties and oral bioavailability, a stable crystalline form, and good pharmaceutical prospects. The compound of formula (I) exhibits good inhibitory activity against ENPP1, and the compound of formula (I) shows good pharmacokinetic indicators.

[0071] The compound represented by formula (I) in crystalline form according to the present invention, and the specific crystalline form, have at least one of the following beneficial effects: (1) The compound represented by formula (I) in crystalline form has good properties that make it convenient for weighing, transporting, separating, purifying, and storing; (2) The compound represented by formula (I) in crystalline form and the specific crystalline form have good crystallinity; (3) The compound represented by formula (I) in crystalline form and the specific crystalline form have excellent operability, such as being easy to purify, filter, and separate, and in particular, crystalline form A is easy to manufacture and has a high yield; and (4) The crystalline form has excellent physical and chemical stability, and in particular, crystalline form A has high purity and can be used directly as a pharmaceutical raw material, and has excellent prospects for medicinal use.

[0072] 1. Equipment and Analytical Methods 1.1 Powder X-ray Diffraction (X-ray powder diffractometer: XRPD) Equipment Model: D8 Advance Examination conditions: General rules 0451 of Part 4 of the Chinese Pharmacopoeia X-ray generator: copper target Tube voltage: 40KV Scattering slit: 0.6 mm Detector slit: 10.5 mm Scatter prevention slit: 2.5° Scan range: 3-50° Step duration: 0.1 seconds / step Step size: 0.02°

[0073] 1.2 Differential Scanning Calorimeter (DSC) Device model: DSC 3 Test conditions: The temperature is raised to 300°C at a starting temperature of 30°C and a heating rate of 10°C / min. The purge gas flow rate is 50 mL / min, the dry gas flow rate is 150 mL / min, the purge gas is N2, and the crucible is an aluminum crucible.

[0074] 1.3 Thermal Gravimetric Analyzer (TGA) Device model: TGA 2 Test conditions: The mixture is heated to 350°C at a starting temperature of 30°C and a heating rate of 10°C / min. The balance protection gas flow rate is 20 mL / min, the reaction gas flow rate is 50 mL / min, the purge gas is N2, and the crucible is an aluminum oxide crucible.

[0075] 1.4 Dynamic Gas Adsorption System (DVS) Equipment model: DVS Intrinsic Test method: 25℃ 0%RH-90%RH-0%RH, refer to General Rules 0103 of Part 4 of the Chinese Pharmacopoeia. Balance dm / dt: 0.002 RH (%) Measurement gradient: 10% / step RH (%) Measurement gradient range: 0%~90%

[0076] Description of hygroscopic characteristics and definition of boundary values ​​for hygroscopic weight increase (hygroscopic weight increase rate): JPEG2026519885000003.jpg44170

[0077] 1.5 Crystalline Stability Test in Water and Physical Media The preparation process for the biological medium is as shown in the table below. Crystalline samples were added to the biological medium and water, shaken at a constant temperature of 37°C for 24 hours, and samples were taken at 1 hour, 4 hours, and 24 hours. The resulting solutions were filtered through a 0.22 μm aqueous filtration membrane (filter). Some samples with high concentrations were diluted as needed with a diluent, and the signal peak area of ​​the solution was measured using HPLC. Finally, the concentration of the compound in the solution was calculated based on the peak area, the HPLC standard curve of the raw materials, and the dilution factor. Furthermore, pH values ​​were measured for samples taken at different time points, and XRPD tests were performed on the residual solids.

[0078] JPEG2026519885000004.jpg100170 [Brief explanation of the drawing]

[0079] [Figure 1] This is the XRPD pattern of crystalline form A of the compound of formula (I) produced in Example 2. [Figure 2] This figure shows the DSC pattern of the A crystal form of compound (I). [Figure 3] This figure shows the TGA pattern of the A crystal form of compound (I). [Figure 4] This figure shows the XRPD pattern of crystalline form B of the compound of formula (I) produced in Example 4. [Figure 5] This figure shows the XRPD pattern of crystalline form C of the compound of formula (I) produced in Example 5. [Figure 6] This figure shows the XRPD pattern of crystalline form D of the compound of formula (I) produced in Example 6. [Figure 7] This figure shows the XRPD pattern of crystalline form A of the compound of formula (I) produced in Example 3. [Figure 8] The figure shows indicators of cardiac function and changes in body weight in a mouse model of acute myocardial ischemia induced by the compound of formula (I), specifically A) left ventricular ejection fraction, B) left ventricular short-axis shortening ratio, and C) changes in body weight. [Figure 9]The diagram shows the changes in myocardial fibrosis in a mouse model of acute myocardial ischemia induced by the compound of formula (I), with D) Masson-stained pathological sections of mouse cardiac tissue and E) the degree of myocardial fibrosis in mice. [Figure 10] This figure shows indicators of cardiac function and changes in body weight in a rat acute myocardial ischemia model induced by the compound of formula (I), specifically a) left ventricular short-axis shortening ratio in rats, b) left ventricular ejection fraction in rats, and c) changes in rat body weight. [Figure 11] This figure shows the changes in myocardial fibrosis in a rat model of acute myocardial ischemia induced by the compound of formula (I), with d) Masson-stained pathological sections of rat cardiac tissue and e) the degree of myocardial fibrosis in rats. [Figure 12] This figure shows a comparison of the XRPD patterns of crystal form A before and after stability experiments. Note: "Compound 1" shown in the figure is the compound of formula (I). [Modes for carrying out the invention]

[0080] The present invention will be further described below with reference to specific examples in order to better understand its contents, but these specific examples are not intended to limit the scope of the present invention.

[0081] [Example 1] Synthesis of compound (I) [ka]

[0082] Step 1: Preparation of 4-chloro-7-methoxy-1,8-naphthyridine-3-carboxylate ethyl ester 12 g, 22.7 mmol, 1 equivalent of 2-(((6-methoxypyridine-2-yl)amino)methylene)malonic acid 1,3-diethyl ester was dissolved in phosphorus oxychloride (60 mL), and the reaction system was carried out under a nitrogen atmosphere at 110 °C for 4 hours with stirring. After monitoring the completion of the reaction by LC-MS, the solvent was removed by distillation under reduced pressure, and the resulting residue was dissolved in ethyl acetate (60 mL) for clarification. Water (20 mL) was added for rapid washing, and the organic phase was dried over anhydrous sodium sulfate, filtered, and the filtrate was spin-dried. The resulting crude product was separated and purified by flash chromatography (Silica gel, DCM:EA=1:1) to obtain the target compound (1.5 g, yield 12%). LC-MS(ESI)[M+H] + = 266.0.

[0083] Step 2: Preparation of 4-((4-bromo-2,6-difluorobenzyl)amino)-7-methoxy-1,8-naphthyridine-3-carboxylate ethyl Ethyl 4-chloro-7-methoxy-1,8-naphthyridine-3-carboxylate (3.0 g, 18.75 mmol, 1 equivalent) was dissolved in acetonitrile (80 mL), and potassium carbonate (5.2 g, 37.5 mmol, 2.0 equivalents) and (4-bromo-2,6-difluorophenyl)methylamine (4.37 g, 19.69 mmol, 1.05 equivalents) were added. The reaction was carried out under a nitrogen atmosphere at 40°C for 20 hours. After monitoring for reaction completion by LC-MS, the reaction mixture was concentrated, diluted with ethyl acetate, washed with water, and the organic phase was dried over anhydrous sodium sulfate. The filtrate was filtered and spin-dried. The resulting crude product was separated and purified by flash chromatography (Silica gel, DCM:EA=4:1) to obtain the target compound (3.5 g, yield 69.2%). LC-MS(ESI)[M+H] + = 452.0.

[0084] Step 3: Preparation of (4-((4-bromo-2,6-difluorobenzyl)amino)-7-methoxy-1,8-naphthyrizin-3-yl)methanol 4-((4-bromo-2,6-difluorobenzyl)amino)-7-methoxy-1,8-naphthyridine-3-carboxylate ethyl (120 mg, 22.7 mmol, 1 equivalent) was dissolved in ethanol (10 mL), and sodium borohydride (134 mg, 5 equivalents) was gradually added. The reaction system was carried out under a nitrogen atmosphere at 50°C for 16 hours with stirring. After monitoring the completion of the reaction by LC-MS, the solvent was spin-dried, and the resulting residue was quenched with saturated sodium chloride solution (5 mL), extracted with ethyl acetate (10 mL), the organic phase was dried over anhydrous sodium sulfate, filtered, and the filtrate was spin-dried. The resulting crude product was separated and purified by flash chromatography (Silica gel, DCM:MeOH=2:1) ​​to obtain the target compound (120 mg, yield 38%).

[0085] LCMS (ESI) [M+H] + = 410.0; 1 H NMR (400 MHz, CDCl3) δ 8.43 (s, 1H), 8.35 (d, J = 9.2 Hz, 1H), 7.09 (d, J = 6.8 Hz, 2H), 6.90 (d, J = 9.2 Hz, 1H), 4.75 (s, 2H), 4.69 (s, 2H), 4.12 (s, 3H).

[0086] Step 4: Preparation of 1-(4-bromo-2,6-difluorobenzyl)-8-methoxy-1,4-dihydro-2H-[1,3]oxazino[5,4-c][1,8]naphthyridine-2-one (4-((4-bromo-2,6-difluorobenzyl)amino)-7-methoxy-1,8-naphthyridine-3-yl)methanol (300 mg, 0.73 mmol, 1 equivalent) was dissolved in dichloromethane (15 mL), and N,N-diisopropylethylamine (473 mg, 3.66 mmol, 5 equivalents) and triphosgene (651 mg, 2.19 mmol, 3 equivalents) were added, and the mixture was reacted at 0°C for 2 hours.

[0087] The reaction was monitored for completion using LC-MS. The reaction mixture was washed with water, dried over anhydrous sodium sulfate, filtered, and the filtrate was spin-dried. The resulting crude product was separated and purified by flash silica gel column chromatography (Silica gel, MeOH:DCM=5:95) to obtain the target compound (250 mg, yield 78.51%). LC-MS(ESI)[M+H] + =436.0.

[0088] Step 5: Preparation of 1-(2,6-difluoro-4-((4-methoxybenzyl)thio)benzyl)-8-methoxy-1,4-dihydro-2H-[1,3]oxazino[5,4-c][1,8]naphthyridine-2-one 1-(4-bromo-2,6-difluorobenzyl)-8-methoxy-1,4-dihydro-2H-[1,3]oxazino[5,4-c][1,8]naphthyridine-2-one (200 mg, 0.46 mmol, 1 equivalent) was dissolved in dioxane (10 mL), and (4-methoxyphenyl)methanethiol (141 mg, 0.92 mmol, 2 equivalents), [5-(diphenylphosphin)-9,9-dimethyl-9H-xanthogen-4-yl]diphenylphosphine (53 mg, 0.09 mmol, 0.2 equivalents), tris(1,5-diphenylpenta-1,4-dien-3-one)dipalladium (84 mg, 0.09 mmol, 0.2 equivalents), and N,N-diisopropylethylamine (178 mg, 1.38 mmol, 3 equivalents) were added. The reaction was carried out at 90°C for 2 hours, and the completion of the reaction was monitored by LC-MS. The reaction mixture was concentrated and spin-dried, and then separated and purified by flash silica gel column chromatography (Silica gel, MeOH:DCM=5:95) to obtain the target compound (110 mg, yield 47.09%). LC-MS(ESI)[M+H] + = 510.1.

[0089] Step 6: Preparation of 3,5-difluoro-4-((8-methoxy-2-oxo-2H-[1,3]oxazino[5,4-c][1,8]naphthyridine-1(4H)-yl)methyl)benzenesulfonamide 1-(2,6-difluoro-4-((4-methoxybenzyl)thio)benzyl)-8-methoxy-1,4-dihydro-2H-[1,3]oxazino[5,4-c][1,8]naphthyridine-2-one (100 mg, 0.2 mmol, 1.0 equivalent), acetic acid (83 mg, 1.37 mmol, 7.0 equivalents), and water (50 mg, 2.75 mmol, 14.0 equivalents) were dissolved in tetrahydrofuran (15 mL), and dichlorohydantoin (116 mg, 0.59 mmol, 3 equivalents) was added at 0°C. The mixture was reacted at 25°C for 1 hour. Ammonia water (0.5 mL) was added, and the mixture was reacted at 25°C for 10 minutes. The completion of the reaction of the raw materials was monitored by LC-MS. After concentrating the reaction mixture, it was purified by preparative HPLC to obtain the target compound (14.6 mg, yield 17.05%).

[0090] LCMS (ESI) [M+H] + = 437.1; 1 HNMR (400MHz, MeOD-d4) δ 8.71 (s, 1H), 8.59 (d, J = 9.2 Hz, 1H), 7.46 (d, J = 7.6 Hz, 2H), 7.13 (d, J = 9.6 Hz, 1H), 5.57 (s, 2H), 5.30 (s, 2H), 4.12 (s, 3H).

[0091] [Example 2] Preparation of the A crystalline form of compound (I) The compound of formula (I) (100 mg) and dimethyl sulfoxide (0.3 mL) prepared in Example 1 were sequentially added to a reaction flask, gradually heated and dissolved to clarify, and then gradually added to water (1 mL) at 80°C. After gradually cooling to room temperature, the mixture was stirred for 1 hour, filtered, the filtered cake was washed with an appropriate amount of water, and the filtered cake was dried under reduced pressure at 65°C to obtain the A crystalline form of the compound of formula (I).

[0092] JPEG2026519885000006.jpg43170

[0093] [Example 3] Preparation of the A crystalline form of compound (I) The compound of formula (I) (100 mg) prepared in Example 1 was added to a reaction flask, acetone (1 mL) was added, and the mixture was stirred at room temperature for 24 hours to form a suspension slurry. The mixture was filtered, and the filtered cake was dried under reduced pressure at 45°C to obtain the A crystalline form of the compound of formula (I).

[0094] JPEG2026519885000007.jpg39170

[0095] [Example 4] Preparation of crystalline form B of compound (I) 260 mg of the sample was weighed and dissolved in 10 mL of DMF at room temperature. 0.8 mL of the solution was taken, and approximately 8 mL of methyl tert-butyl ether was added dropwise. After stirring at room temperature for a certain period of time, the system from which the solid precipitated was centrifuged, and the solid was vacuum-dried at room temperature to obtain crystal form B.

[0096] JPEG2026519885000008.jpg43170

[0097] [Example 5] Preparation of crystalline form C of compound (I) 260 mg of the sample was weighed and dissolved in 4 mL of DMSO at room temperature. 0.3 mL of the solution was taken, and approximately 3 mL of toluene was added dropwise. After stirring at room temperature for a certain period of time, the system from which the solid precipitated was centrifuged, and the solid was vacuum-dried at room temperature to obtain crystalline form C.

[0098] JPEG2026519885000009.jpg43170

[0099] [Example 6] Preparation of crystalline form D of compound (I) 260 mg of the sample was weighed and dissolved in 10 mL of NMP at room temperature. 0.8 mL of the solution was taken, and approximately 8 mL of methyl tert-butyl ether was added dropwise. After stirring at room temperature for a certain period of time, the system from which the solid precipitated was centrifuged, and the solid was vacuum-dried at room temperature to obtain crystalline form D.

[0100] JPEG2026519885000010.jpg77170

[0101] [Test Example 1] Study of hygroscopic properties of crystalline forms Experimental equipment: DVS Intrinsic Experimental method: 25-35 mg of crystalline form A of the compound of formula (I) prepared in Example 2 was taken and tested on a DVS sample plate.

[0102] Experimental conclusion: The crystalline form A of compound (I) showed a weight increase W = 0.4461% at 25°C and 80% humidity, indicating slight hygroscopicity. After DVS testing, the crystalline form remained unchanged.

[0103] [Test Example 2] Solid Stability Study 20 mg of crystalline form A of the compound of formula (I) prepared in Example 2 was weighed into a vial and left open for 7 days at high temperature (60°C), long-term (25°C / 60% RH), and accelerated (40°C / 75% RH). After that, samples were taken and purity detection and X-ray powder diffraction analysis were performed to investigate the stability of Example 2 (crystalline form A) under different conditions. The results are shown in Table 6 and Figure 12.

[0104] JPEG2026519885000011.jpg39170

[0105] The data shows that Example 2 (crystal form A) can maintain chemical stability and crystal form stability in solid stability tests.

[0106] [Test Example 3] Stability test of crystalline form A of compound (I) in a biological solvent medium A sample of Example 2 (crystal form A) was taken and stability tests were performed in three biological media (FaSSIF, FeSSIF, and FaSSGF) and water. After 24 hours, the residual sample was taken and subjected to X-ray powder diffraction analysis. The results are shown in Table 7.

[0107] JPEG2026519885000012.jpg37170

[0108] The data shows that Example 2 (crystalline form A) can maintain a stable form in a biological solvent medium.

[0109] [Test Example 4] In vitro ENPP1 enzyme inhibition test ENPP1 is a transmembrane glycoprotein capable of hydrolyzing nucleotides and derivatives having a nucleotide-5'-monophosphate structure. ENPP1 can hydrolyze artificially synthesized 5'-monophosphate-p-nitrophenyl ester (TMP-pNP) to produce nucleotide-5'-monophosphate and the chromogenic product p-nitrophenol. The amount of p-nitrophenol produced can be directly measured by absorbance at 405 nm, and this is proportional to the enzyme activity.

[0110] Experimental Procedure A 100-fold serially diluted compound was prepared using DMSO, starting at 1 mM and followed by a 4-fold serial dilution to establish 10 concentration points. Using Echo, 300 nL of the compound gradient was transferred to a 384-well detection plate (the final concentration starting at 10 μM and resulting in 10 concentration points through 4-fold serial gradient dilution). First, 15 μL of 0.2 ng / μL of hENPP1 enzyme (2x final concentration) prepared in experimental buffer (250 mM NaCl, 50 mM Tris, pH 9.5) was added to each well. Next, 15 μL of 400 μM TMP-pNP (2x final concentration) prepared in experimental buffer was added, and after incubation at 37°C for 0.5 hours, the OD 405 nm value was read using a microplate reader. The inhibition rate (%) was calculated using the formula: % inhibition rate = (OD high signal control - OD sample well) / (OD high signal control - OD low signal control) × 100. The IC50 value was calculated by 4-parameter fitting. High signal control: DMSO group without inhibitor, low signal control group: blank control group. The data for the compound of formula (I) prepared in Example 1 are shown in Table 8.

[0111] JPEG2026519885000013.jpg20170

[0112] [Test Example 5] Pharmacokinetic study of compound concentrations in mice using LC-MS / MS Testing principle: Using LC-MS / MS, the drug concentration in plasma of the target drug at different time points is measured, and the pharmacokinetic curve of the target compound in vivo is plotted.

[0113] Test method: The test compound (compound of formula (I) prepared in Example 1) was dissolved in DMSO to prepare a stock solution with a final concentration of 20 mg / mL. The compound was then dissolved in 5% DMSO (Sigma-Aldrich, SHBJ2847), 45% PEG400 (Sigma-Aldrich, BCCC0015), and 50% dd H2O solvent to a concentration of 1 mg / mL. The mouse source was CD-1 male (JH Laboratory Animal Co. LTD). Nine mice were collected for each compound administration group and administered orally (PO) at a dose of 10 mg / kg. Cross-collections of blood were performed at 0.25 hours, 0.5 hours, 1 hour, 2 hours, 4 hours, 8 hours, and 24 hours after administration. Three samples were collected at each time point, and 110 μL of whole blood (K2EDTA anticoagulant) was collected. The samples were immediately centrifuged at 4°C and 2,000 g for 5 minutes to recover the serum, which was then stored at -70°C. This process involves measuring blood concentrations using a triple-quadrupole MS system (SCIEX), creating standard curves, preparing quality control solutions, and preparing samples.

[0114] Standard curve preparation and quality control solution preparation: Prepare a working solution by diluting with MeOH:H2O (1:1), and add 3 μL of the above standard curve preparation and quality control working solution to 57 μL of blank plasma.

[0115] Sample preparation: Take 30 μL of plasma sample, add 200 μL of internal standard solution (Propranolol, 40 ng / mL), mix uniformly for 1 minute, centrifuge at 5800 rpm for 10 minutes, take 100 μL of supernatant, transfer to a new plate, and analyze. Chromatographic conditions, including mobile phase composition, elution gradient conditions, flow rate, and retention time, are optimized according to the sample. A Waters BEH C18 (2.1 × 50 mm, 1.7 μm) chromatography column is used, with an injection volume of 1 μL. Mass spectrometry is performed using an electrospray ionization source (TuREo spray), selecting multi-channel reaction monitoring (MRM) mode in cation detection mode for secondary mass spectrometry. Based on drug concentration-time data, pharmacokinetic parameters including peak concentration Cmax (maximum blood concentration), peak time Tmax (time to reach peak concentration), area under the drug-time curve AUC, and elimination half-life t1 / 2 are calculated using a non-compartmental model with WinNonlin 8.2 software. The AUC is calculated using the linear trapezoidal curve (linear up log down).

[0116] JPEG2026519885000014.jpg38170

[0117] The test results indicate that the compound of formula (I) according to the present invention has good in vivo pharmacokinetics and has the potential to become a pharmaceutical product.

[0118] [Test Example 6] Evaluation of the efficacy of compound (I) in the mouse pancreatic cancer Pan02 model. 1. Experimental Objective: This study evaluates the in vivo efficacy of the investigational drug in a mouse pancreatic cancer Pan02 cell subcutaneous allogeneic transplant tumor model.

[0119] 2. Test method: 2.1 Cell culture: Mouse pancreatic cancer Pan02 cells were cultured in vitro as a monolayer. The culture conditions were as follows: 10% fetal bovine serum, 0.01 mg / mL sinsulin, 1% penicillin / streptomycin / amphotericin B in DMEM medium, and cultured in a 5% CO2 incubator at 37°C. When the cell saturation reached 80 - 90% and the number of cells reached the required number, the cells were harvested, counted, and seeded.

[0120] 2.2 Animals: C57BL / 6 mice, female, 6 - 8 weeks old, weighing 18 - 22 g. They were provided by Shanghai Xipu'er-Bikai Laboratory Animal Co., Ltd. or other suppliers.

[0121] 2.3 Tumor seeding and animal grouping·administration: 0.1 mL (5×10 6 cells) of Pan02 cells were subcutaneously seeded on the right side of the back of each mouse. When the average tumor volume reached about 100 - 200 mm 3 , the animals were grouped and administration was started. Before administration, the animal body weights were weighed and the tumor volumes were measured. The animals were randomly grouped according to the tumor volume, with 8 animals in each group. The control group was administered with the solvent (5% DMSO + 45% PEG400 + 50% H20)(1% CMC-Na), and the experimental group was administered with a dose of 20 mg / kg. According to the mouse body weight, the administration volume was 10 μL / g, and oral administration (intragastric administration) was performed twice a day. The health status of the animals was observed daily. If the tumor volume exceeded 3,000 mm 3 , there were severe diseases or pain, or the weight loss exceeded 20% and continued to deteriorate, euthanasia must be performed.

[0122] 2.4 Tumor growth inhibition rate: The diameter of the tumor was measured with a vernier caliper twice a week. The formula for calculating the tumor volume was V = 0.5a×b 2 , where a and b represent the long diameter and short diameter of the tumor, respectively. The tumor inhibitory effect of the compound: TGI(%) = [(1 - (average tumor volume after administration in the treatment group - average tumor volume at the start of administration in the treatment group)) / (average tumor volume before treatment in the solvent control group - average tumor volume after treatment in the solvent control group)]×100%. For data analysis, a t-test was used for comparison between two groups, and p < 0.05 was judged to be significantly different.

[0123] 3. Experimental results: The compound of formula (I) produced in Example 1 of the present invention showed activity in a mouse pancreatic cancer Pan02 model. When the compound of formula (I) according to the present invention was administered for 16 days, tumor growth was significantly suppressed with a TGI of 43.4%, showing a statistically significant difference (p<0.05) compared to the control group, and did not suppress the body weight of the mice, indicating good safety. The crystalline form of the compound of formula (I) (e.g., crystalline form A of the compound of formula (I)) has essentially the same pharmacokinetic properties.

[0124] [Test Example 7] Pharmacodynamic evaluation of compounds in a mouse myocardial ischemia model 1. Experimental Objective The purpose of this study is to evaluate the efficacy of the compound in a C57 BL / 6 mouse myocardial ischemia model.

[0125] 2. Test Method After the animals had been acclimatized for a week upon arrival at the facility, they were divided into four groups according to their weight the day before surgery: a sham group of 6 animals and a model group of 8 animals. The following day, they underwent myocardial ischemia surgery (sham group mice were only sutured through thoracotomy, without ligation) to create a model. The model was created using the left anterior descending coronary artery (LAD) ligation method, and after modeling, penicillin (20,000 units / mouse im) was administered for 3 days, and disinfection was performed to prevent infection.

[0126] One hour before surgery, each group of animals was administered the corresponding compound. The control group (vehicle) was administered the solvent: 5% DMSO + 45% PEG400 + 50% H2O (1% CMC-Na), while the experimental groups were administered 4 mg / kg and 20 mg / kg, respectively, by oral intragastric administration twice daily for 28 consecutive days. After administration, the animals' body weight was monitored daily, and cardiac ultrasound imaging parameters were measured on days 1, 7, and 28. The primary echocardiographic indicators were left ventricular short axis shortening (FS) and left ventricular ejection fraction (LVEF). After the echocardiogram on day 28, cardiac tissue was collected, placed in tissue fixative, and subjected to pathological analysis using Masson's tricolor staining.

[0127] 3. Experimental Results Compound (I) described in Example 1 of the present invention showed good cardiac function improving activity in a mouse myocardial infarction model. The compound of formula (I) according to the present invention has a protective effect on cardiac function after pre-administration the day before modeling. As shown in Figure 5, on day 1, the 4mpk and 20mpk dose groups increased the left ventricular ejection fraction (LVEF) by 15% and 10%, respectively (Figure A), and the left ventricular short axis shortening ratio (FS) by 7% and 5%, respectively (Figure B), compared to the model solvent control group. On day 7, the left ventricular ejection fraction (LVEF) increased by 23.4% and 23.3%, respectively, and the left ventricular short axis shortening ratio (FS) increased by 12.2% and 12.3%, respectively. On day 28, the LVEF and FS of the model solvent control group worsened further, while compound (I) was able to further maintain the left ventricular LVEF and FS of mice, and no significant effect on body weight was observed (Figure C). Statistical significance was analyzed using two-way ANOVA / one-way ANOVA / t-test analysis, and p<0.05 was considered statistically significant.

[0128] The compound of formula (I) produced in Example 1 of the present invention showed an ameliorative effect on myocardial fibrosis (myocardial fibrosis) in mice with myocardial infarction. As shown in Figure 6, Figure D is an electron micrograph of a Masson-stained pathological section of mouse cardiac tissue. Figure E shows the statistical results regarding the degree of fibrosis in the mouse cardiac tissue. As shown in the figure, in the sham surgery group, the red myocardial fibers of the mice were neatly arranged, and blue collagen fibers were hardly observed. In the MI model control group, cardiomyocytes were necrotic, the myocardial tissue structure was disordered, and a large area of ​​blue collagen fibers was observed in the infarcted area. In the group administered with compound (I), the myocardial tissue structure of the mice was improved to a certain extent, with red cardiomyocytes and blue collagen alternately distributed in the infarcted area, and the collagen fibers were significantly reduced. This indicates that after 28 days of administration of compound (I), the degree of myocardial fibrosis after myocardial infarction (MI) could be reduced, and the degree of fibrosis could be reduced by 8.8% and 9.6%, respectively, compared to the model solvent control group (Figure E). The crystalline form A of the compound of formula (I) has substantially the same medicinal properties.

[0129] [Test Example 8] Pharmacodynamic evaluation of compounds in a rat myocardial ischemia model 1. Experimental Objective: The purpose of this study is to evaluate the efficacy of the compound in a rat myocardial ischemia model.

[0130] 2. Test Method After the animals had been acclimatized for one week upon arrival at the facility, the left anterior descending coronary artery (LAD) was ligated to create a model. Penicillin (80,000 units / rat im) was administered for three days after the model was created, and disinfection was performed to prevent infection.

[0131] One day after model establishment, the modeled animals were randomly divided into four groups based on baseline body weight and cardiac ultrasound imaging parameters. Each group consisted of four animals in the sham surgery group and six animals in the myocardial infarction model group (MI). Drug administration was initiated. The control group was administered a blank solvent, Vehicle: 5% DMSO + 45% PEG400 + 50% H2O (1% CMC-Na). The experimental groups received doses of 30 mg / kg and 75 mg / kg, respectively, administered orally and intragastricly twice daily for 28 consecutive days. After administration, the animals' body weight was monitored daily, and cardiac ultrasound imaging parameters were measured on days 7 and 28. The primary echocardiographic indicators were left ventricular short axis shortening (FS) and left ventricular ejection fraction (LVEF). After the echocardiogram on day 28, cardiac tissue was collected, placed in tissue fixative, and subjected to pathological analysis using Masson's tricolor staining.

[0132] 3. Experimental Results The compound of formula (I) prepared in Example 1 of the present invention showed good cardiac function-improving activity in a rat myocardial infarction model (Figure 7a, b). The compound of formula (I) according to the present invention significantly increased the left ventricular ejection fraction (LVEF) and left ventricular short axis shortening index (FS) in rats on day 7 of administration. In the 30mpk and 75mpk administration groups, LVEF increased by 11.43% and 18.85%, respectively, and the left ventricular short axis shortening index (FS) increased by 9.73% and 12.62%, respectively. On day 28, in the 30mpk and 75mpk administration groups, LVEF increased by 18.75% and 23.8%, respectively, and the left ventricular short axis shortening index (FS) increased by 13.01% and 15.88%, respectively. Furthermore, a dose-dependent trend was observed, and no significant effect on body weight was observed (Figure c). Statistical significance was analyzed using a t-test, and p<0.05 was considered to indicate a significant difference.

[0133] The compound of formula (I) produced in Example 1 of the present invention showed an ameliorative effect on myocardial fibrosis (myocardial fibrosis) in rats with myocardial infarction. Figure d is an electron micrograph of a Masson-stained pathological section of rat cardiac tissue. Figure e shows the statistical results regarding the degree of fibrosis in rat cardiac tissue. As shown in Figure 8, in the sham surgery group, the red myocardial fibers of the rats were neatly arranged, and blue collagen fibers were hardly observed. In the MI model control group, cardiomyocytes were necrotic, the myocardial tissue structure was disordered, and a large area of ​​blue collagen fibers was observed in the infarcted area. In the group administered with compound (I), the myocardial tissue structure of the rats was improved to a certain extent, with red cardiomyocytes and blue collagen alternately distributed in the infarcted area, and collagen fibers were significantly reduced. This indicates that after 28 days of administration of compound (I), the degree of myocardial fibrosis after myocardial infarction (MI) could be reduced, and the degree of fibrosis could be reduced by 5.2% and 6.5%, respectively, compared to the model solvent control group (Figure e). The crystalline form A of the compound of formula (I) has substantially the same medicinal properties.

Claims

1. The following formula: 【Chemistry 1】 A compound with the crystalline form represented by formula (I).

2. The compound represented by formula (I) of the crystalline form is crystalline form A, and in the X-ray powder diffraction pattern using Cu-Kα rays, diffraction peaks are found at 2θ angles of 24.9±0.2°, 25.1±0.2°, and 25.7±0.2°. Alternatively, in an X-ray powder diffraction pattern using Cu-Kα rays, diffraction peaks are found at 2θ angles of 12.8±0.2°, 14.0±0.2°, 16.0±0.2°, 18.5±0.2°, 24.9±0.2°, 25.1±0.2°, and 25.7±0.2°. Alternatively, in an X-ray powder diffraction pattern using Cu-Kα rays, diffraction peaks are found at 2θ angles of 12.8±0.2°, 14.0±0.2°, 16.0±0.2°, 18.5±0.2°, 24.9±0.2°, 25.1±0.2°, 25.7±0.2°, 26.9±0.2°, and 28.5±0.2°. Alternatively, in an X-ray powder diffraction pattern using Cu-Kα rays, diffraction peaks are found at 2θ angles of 12.8±0.2°, 14.0±0.2°, 16.0±0.2°, 18.5±0.2°, 24.9±0.2°, 25.1±0.2°, 25.7±0.2°, 26.9±0.2°, 28.5±0.2°, and 29.8±0.2°. Alternatively, in an X-ray powder diffraction pattern using Cu-Kα rays, diffraction peaks are found at 2θ angles of 12.8±0.2°, 14.0±0.2°, 16.0±0.2°, 17.0±0.2°, 18.5±0.2°, 20.5±0.2°, 24.9±0.2°, 25.1±0.2°, 25.7±0.2°, 26.9±0.2°, 28.5±0.2°, and 29.8±0.2°. Alternatively, having an X-ray powder diffraction pattern substantially as shown in Figure 1 or Figure 7, Alternatively, the analysis of the XRPD pattern using Cu-Kα radiation is shown in Table 1 or Table 2. Alternatively, the differential scanning calorimetry curve has an endothermic peak at 233.23 ± 4°C. Alternatively, the differential scanning calorimetry curve has an exothermic peak at 234.3 ± 4°C. Alternatively, it may have a differential scanning calorimetry (DSC) pattern essentially as shown in Figure 2, Alternatively, the thermogravimetric analysis curve shows a weight loss of 13.50 ± 1%, for example, 13.5111%, between approximately 105 and 254°C. Alternatively, in the thermogravimetric analysis curve, the weight loss reaches 13.50 ± 1% at 254.00 ± 3°C, for example, 13.5111%. Alternatively, the A crystal form of the compound of formula (I) has substantially the thermogravimetric analysis (TGA) pattern shown in Figure 3, Alternatively, the compound represented by formula (I) of the crystalline form is crystalline form B, and in the X-ray powder diffraction pattern using Cu-Kα rays, diffraction peaks are found at 2θ angles of 19.19±0.2°, 27.18±0.2°, 13.44±0.2°, and 22.55±0.2°. Alternatively, the crystal form B has diffraction peaks at 2θ angles of 26.19±0.2°, 20.76±0.2°, 4.42±0.2°, 22.84±0.2°, 19.19±0.2°, 27.18±0.2°, 13.44±0.2°, and 22.55±0.2° in an X-ray powder diffraction pattern using CuKα rays. Alternatively, the crystal form B is shown in Figure 4 as an XRPD pattern using Cu-Kα radiation. Alternatively, the analysis of the XRPD pattern using Cu-Kα radiation for the aforementioned crystal form B is shown in Table 3. Alternatively, the crystalline form B is the DMF solvate of the compound of formula (I), Alternatively, the compound represented by formula (I) of the crystalline form is crystalline form C, and in an X-ray powder diffraction pattern using Cu-Kα rays, it has diffraction peaks at 2θ angles of 22.11±0.2°, 22.55±0.2°, 20.96±0.2°, and 26.63±0.2°. Alternatively, the crystalline form C has diffraction peaks at 2θ angles of 22.11±0.2°, 22.55±0.2°, 20.96±0.2°, 26.63±0.2°, 15.01±0.2°, 25.68±0.2°, 26.38±0.2°, 28.54±0.2°, 13.22±0.2°, and 18.68±0.2° in an X-ray powder diffraction pattern using CuKα rays. Alternatively, the crystal form C is shown in Figure 5 by an XRPD pattern using Cu-Kα radiation. Alternatively, the crystal form C is shown in Table 4, based on the analysis of the XRPD pattern using Cu-Kα radiation. Alternatively, the crystalline form C is the DMSO solvate of the compound of formula (I), Alternatively, the compound represented by formula (I) of the crystalline form is crystalline form C, and in an X-ray powder diffraction pattern using Cu-Kα rays, diffraction peaks are found at 2θ angles of 22.71±0.2°, 18.92±0.2°, 22.96±0.2°, 26.4±0.2°, 24.44±0.2°, 16.62±0.2°, 13.15±0.2°, 17.79±0.2°, and 27.12±0.2°. Alternatively, the crystal form D has diffraction peaks at 2θ angles of 22.71±0.2°, 18.92±0.2°, 22.96±0.2°, 26.4±0.2°, 24.44±0.2°, 16.62±0.2°, 13.15±0.2°, 17.79±0.2°, 27.12±0.2°, 24.17±0.2°, 28.81±0.2°, 19.95±0.2°, 28.42±0.2°, 24.85±0.2°, 14.78±0.2°, 17.36±0.2°, and 29.16±0.2° in the X-ray powder diffraction pattern using Cu-Kα rays. Alternatively, the crystal form D is shown in Figure 6 by an XRPD pattern using Cu-Kα radiation. Alternatively, the crystal form D is shown in Table 5, based on the analysis of the XRPD pattern using Cu-Kα radiation. Alternatively, the crystalline form D is the NMP solvate of the compound of formula (I). A compound represented by formula (I) in the crystalline form described in claim 1, characterized by the features described above.

3. Method 1 includes adding the compound of formula I to organic solvent A to dissolve it, and then adding the solution to organic solvent B to precipitate crystals. The organic solvent A is selected from one or more of DMF (N,N-dimethylformamide), DMSO (dimethyl sulfoxide), and NMP (N-methylpyrrolidone), and the organic solvent B is selected from one or more of water, methanol, ethanol, MTBE, toluene, and dichloromethane. In some embodiments of the present invention, when the compound of formula (I) is crystalline form A, in method 1, organic solvent A is selected from DMF and organic solvent B is selected one or two from water and ethanol, or organic solvent A is selected from DMSO and organic solvent B is selected one or two from toluene and methanol. In some embodiments of the present invention, when the compound of formula (I) is crystalline form B, in method 1, organic solvent A is selected from DMF, and organic solvent B is selected from one or two of MTBE and Truet. In some embodiments of the present invention, when the compound of formula (I) is in crystalline form D, in method 1, organic solvent A is selected from NMP, and organic solvent B is selected from one or more of water, dichloromethane, and MTBE. or, Method 2 includes dissolving the compound of formula I in an organic solvent C, and then adding an organic solvent D to precipitate crystals. The organic solvent C is selected from one or more of DMF, DMSO, and NMP, and the organic solvent D is selected from one or more of acetone, ethylene glycol methyl ether, water, dichloromethane, ethyl acetate, MTBE, toluene, tetrahydrofuran, dioxane, ethanol, trifluoroethanol, acetonitrile, ethylene glycol dimethyl ether, isopropanol, and methyl ethyl ketone. Preferably, when the compound of formula (I) is crystalline form A, in method 1, organic solvent C is selected from DMF and organic solvent D is selected one or more from acetone, ethylene glycol methyl ether, water, and dichloromethane; or organic solvent C is selected from DMSO and organic solvent D is selected one or more from ethanol, trifluoroethanol, ethylene glycol methyl ether, acetonitrile, dioxane, ethylene glycol dimethyl ether, and methyl ethyl ketone; or organic solvent C is selected from NMP and organic solvent D is selected one or more from water, acetonitrile, dichloromethane, and ethylene glycol dimethyl ether. Preferably, when the compound of formula (I) is in crystalline form B, in method 2, organic solvent C is selected from DMF, and organic solvent D is selected from ethyl acetate, MTBE, toluene, tetrahydrofuran, dioxane, and methyl ethyl ketone, Preferably, when the compound of formula (I) is in crystalline form C, in method 2, the organic solvent C is selected from DMSO, and the organic solvent D is selected from one or more of ethyl acetate, water, and toluene. Preferably, when the compound of formula (I) is in crystalline form D, in method 2, the organic solvent C is selected from NMP, and the organic solvent D is selected from one or more of isopropanol, MTBE, toluene, tetrahydrofuran, and methyl ethyl ketone. or, Method 3 includes suspending the compound of formula I in an organic solvent E, forming a slurry, and filtering to obtain crystals. The organic solvent E is selected from one or more of the following: methanol, ethanol, isopropanol, ethyl acetate, n-heptane, MTBE, ethylene glycol methyl ether, water, acetonitrile, toluene, dichloromethane, tetrahydrofuran, dioxane, ethylene glycol dimethyl ether, methyl ethyl ketone, dimethylformamide, dimethyl sulfoxide, N-methylpyrrolidone, and acetone. Preferably, when the compound of formula (I) is in crystalline form A, the organic solvent E in method 3 is selected from one or more of methanol, ethanol, isopropanol, ethyl acetate, n-heptane, MTBE, ethylene glycol methyl ether, water, acetonitrile, toluene, dichloromethane, tetrahydrofuran, dioxane, ethylene glycol dimethyl ether, and methyl ethyl ketone. Preferably, when the compound of formula (I) is in crystalline form B, the organic solvent E in method 3 is selected from DMF, Preferably, when the compound of formula (I) is in crystalline form C, the organic solvent E in method 3 is selected from DMSO. Preferably, when the compound of formula (I) is in crystalline form D, the organic solvent E in method 3 is selected from NMP. A method for producing a compound of formula (I) in the crystalline form described in claim 1 or 2.

4. A crystalline composition comprising a compound of formula (I) in a crystalline form as described in any one of claims 1 to 3, wherein the compound of formula (I) in a crystalline form is selected from one or more of crystalline forms A, B, C, and D. Preferably, the compound of formula (I) in the crystalline form accounts for 10% or more, 20% or more, 30% or more, 40% or more, 50% or more, 60% or more, 70% or more, 80% or more, 90% or more, 91% or more, 92% or more, 93% or more, 94% or more, 95% or more, 96% or more, 97% or more, 98% or more, 99% or more, or 99.5% or more of the weight of the crystalline composition. Preferably, the compound of formula (I) in the crystalline form accounts for 50% or more, 60% or more, 70% or more, 80% or more, 90% or more, 91% or more, 92% or more, 93% or more, 94% or more, 95% or more, 96% or more, 97% or more, 98% or more, 99% or more, or 99.5% or more of the weight of the crystalline composition. Crystalline composition.

5. A pharmaceutical composition comprising a compound of formula (I) in the crystalline form described in claim 1 or 2, or a crystalline composition described in claim 4, and an optionally pharmaceutically acceptable carrier.

6. The pharmaceutical composition according to claim 5, comprising one, two, or three or more other pharmaceuticals for treating cancer or tumors.

7. The compound of formula (I) in crystalline form according to claim 1 or 2, the crystalline composition according to claim 4, or the pharmaceutical composition according to claim 5 or 6, used as a pharmacopoeia for the prevention and / or treatment of ENPP1-mediated diseases, or used in the manufacture of such pharmacopoeia.

8. The compound of formula (I) in crystalline form according to claim 1 or 2, the crystalline composition according to claim 4, or the pharmaceutical composition according to claim 5 or 6, used as a pharmacopoeia for the prevention and / or treatment of cancer or tumor-related diseases or cardiovascular diseases, or used in the manufacture of such pharmacopoeia. Preferably, the compound of formula (I) in crystalline form according to claim 1 or 2, the crystalline composition according to claim 4, or the pharmaceutical composition according to claim 5 or 6 is used in combination with one, two, or three or more other pharmaceuticals or therapeutic means for treating cancer or tumors.

9. The aforementioned antitumor drug is a chemotherapy drug, a targeted therapy drug, or an immunotherapy drug. Preferably, the pharmaceuticals used for the prevention and / or treatment of the cancer or tumor include cell signaling inhibitors, chlorambucil, melphalan, cyclophosphamide, ifosfamide, busulfan, carmustine, lomustine, streptozotocin, cisplatin, carboplatin, oxaliplatin, dacarbazine, temozolomide, procarbazine, methotrexate, fluorouracil, cytarabine, gemcitabine, mercaptopurine, fludarabine, vinblastine, vincristine, and vinorelvidone. Paclitaxel, Docetaxel, Topotecan, Irinotecan, Etoposide, Trabectedin, Dactinomycin, Doxorubicin, Epirubicin, Daunorubicin, Mitoxantrone, Bleomycin, Mitomycin C, Isapirone, Tamoxifen, Flutamide, Gonadorelin analog, Megestrol, Prednisone, Dexamethasone, Methylprednisolone, Thalidomide, Interferon α, Leucovo Calcium phosphate, sirolimus, sirolimus lipids, everolimus, afatinib, alicertib, amvatinib, apatinib, axitinib, bortezomib, bosutinib, brivanib, cabozantinib, cediranib, crenolanib, crizotinib, dacomitinib, danucertib, dasatinib Dovitinib, Erlotinib, Foretinib, Ganetespib, Gefitinib, Ibrutinib, Icotinib, Imatinib, Iniparib, Lapatinib, Lenvatinib, Linifanib, Linsitinib, Masitinib, Momerotinib, Motesanib, Neratinib, Nilotinib, Niraparib,Oprozomib, Olapanib, Pazopanib, Pictilisib, Ponatinib, Quizartinib, Regorafenib, Rigosertib, Rucaparib, Ruxolitinib, Saracatinib, Saridigib, Sorafenib enib), sunitinib, telatinib, tivantinib, tivozanib, tofacitinib, trametinib, vandetanib, veliparib, vemurafenib, vismodegib, volasertib, alemtuzumab, bevacizumab, brentuximab Vedotin, Catumaxomab, Cetuximab, Denosumab, Gemtuzumab, Ipilimumab, Nimotuzumab, Ofatumumab, Panitumumab, Rituximab, Tocitumumab, Trastuzumab, PI3K inhibitors, CSF1R inhibitors, A2A and / or A2B receptor antagonists, IDO inhibitors, Anti-PD-1 antibodies, Anti-PD-L1 antibodies, LAG3 antibodies, TIM-3 antibodies, TIGIT antibodies, CD47 antibodies, CLAUDIN 18. The therapeutic means is selected from the group consisting of 18.2 antibody, anti-CTLA-4 antibody, or any combination thereof, and preferably, the therapeutic means is radiotherapy or surgery. The pharmaceutical composition according to claim 6, or the use according to claim 8.

10. The aforementioned cancers or tumors include skin cancer, bladder cancer, ovarian cancer, breast cancer, gastric cancer, pancreatic cancer, prostate cancer, colon cancer, lung cancer, bone cancer, brain cancer, neurocytoma, rectal cancer, colon cancer, familial adenomatous polyposis, hereditary nonpolyposis colorectal cancer, esophageal cancer, lip cancer, laryngeal cancer, hypopharyngeal cancer, tongue cancer, salivary gland cancer, adenocarcinoma, medullary thyroid cancer, papillary thyroid cancer, kidney cancer, renal parenchymal cancer, cervical cancer, endometrial cancer, choriocarcinoma, testicular cancer, urinary tract cancer, melanoma, brain tumors (e.g., glioblastoma, astrocytoma, meningioma, medulloblastoma), peripheral ectodermal tumors, Hodgkin lymphoma, non-Hodgkin lymphoma, and Burke The group consists of T-cell lymphoma, leukemia, acute lymphoblastic leukemia (ALL), chronic lymphocytic leukemia (CLL), acute myeloid leukemia (AML), chronic myeloid leukemia (CML), adult T-cell leukemia / lymphoma, diffuse large B-cell lymphoma (DLBCL), hepatocellular carcinoma, gallbladder cancer, bronchial cancer, small cell lung cancer, non-small cell lung cancer, multiple myeloma, basal cell tumor, teratoma, retinoblastoma, choroidal melanoma, seminomas, rhabdomyosarcoma, craniopharyngioma, osteosarcoma, chondrosarcoma, myosarcoma, liposarcoma, fibrosarcoma, Ewing's sarcoma, and plasmacytoma. The cardiovascular disease is characterized by being selected from the group consisting of myocardial infarction (MI), heart failure (CF), cardiac trauma, abnormal scar formation of cardiac tissue, cardiomyopathy, cardiomyocyte death, promotion of cardiac repair, and release of pro-inflammatory molecules from cardiomyocytes. The use described in claim 8.

Images