Salt, crystal form and preparation method of methionine adenylyltransferase 2A heterocyclic inhibitor

Abstract

A salt of a methionine adenosine transferase 2A heterocyclic inhibitor having a structure represented by formula (I), a crystal form, and a preparation method and use thereof, the salt crystal form has high purity and stable chemical properties, and has important significance for developing drugs which are suitable for industrial production and have good biological activity.

Description

Salt, crystal form and preparation method of methionine adenosyltransferase 2A heterocyclic inhibitor

[0001] This application claims priority to Chinese patent application No. 202310705474.5, filed on June 14, 2023, and cites the full text of the aforementioned Chinese patent application. Technical Field

[0002] The present invention belongs to the field of biomedicine, and specifically relates to salts, crystal forms, preparation methods and uses of methionine adenosyltransferase 2A heterocyclic inhibitors. Background Art

[0003] Methionine adenosyltransferase (MAT) (also known as S-adenosylmethionine synthetase) is a cellular enzyme that catalyzes the synthesis of S-adenosylmethionine (SAM or AdoMet) from methionine and ATP and is considered to be the rate-limiting step of the methionine cycle. SAM is the propylamino donor in polyamine biosynthesis and the major methyl donor for DNA methylation, and is involved in gene transcription and cell proliferation, as well as the production of secondary metabolites. The three human isoenzymes of MAT include MAT1, MAT2, and MAT3, of which MAT1 and MAT3 are expressed in liver tissue. Methionine adenosyltransferase 2A (MAT2A) is a subtype of MAT2 that is ubiquitously expressed in human cell types and is the predominant form in human cancers, and it also plays a key role in the production of SAM.

[0004] MTAP (methylthioadenosine phosphorylase) is an enzyme ubiquitously expressed in normal tissues that catalyzes the conversion of methylthioadenosine (MTA) to adenine and 5-methylthioribose-1-phosphate, adenine to adenosine monophosphate, and 5-methylthioribose-1-phosphate to methionine and formate. MTA can be used as an alternative purine source when purine synthesis is blocked, for example by antimetabolites.

[0005] The gene encoding MTAP is located in a region on chromosome 9 and is frequently deleted in cancer patients from cells of the central nervous system, pancreas, esophagus, bladder, and lung. Loss of MTAP leads to the accumulation of MTA, making MTAP-deficient cells more dependent on SAM production and, therefore, MAT2A activity, compared to cells expressing MTAP. In a screen of approximately 400 cancer cell lines, MAT2A knockdown resulted in a greater percentage of MTAP-deficient cells losing viability than cells normally expressing MTAP. Furthermore, inducible knockdown of the MAT2A protein reduced tumor growth in vivo. These results suggest that MAT2A inhibitors may provide a novel therapeutic approach for patients with tumors that lack MTAP.

[0006] PCT / CN2022 / 140380 provides a novel heterocyclic methionine adenosyltransferase 2A inhibitor. The crystalline structure of a pharmaceutically active ingredient often affects the chemical stability of the drug. Differences in crystal form, preparation method, and storage conditions can lead to variations in the compound's crystalline structure, sometimes accompanied by the formation of alternative crystalline forms. Therefore, in-depth research on the polymorphic forms of compounds and the acquisition of highly pure and chemically stable salts and crystalline forms are of great significance for the development of drugs suitable for industrial production and exhibiting excellent biological activity.

[0007] Summary of the Invention

[0008] All contents involved in patent PCT / CN2022 / 140380 are added to the present invention by reference.

[0009] The present invention provides an acid salt of the compound represented by the following formula I:

[0010] Where X is selected from CR 3 or N; Y is selected from CR 4 or N; Z is selected from CR 5 or N; W is selected from CR 6 or N;

[0011] where R 3 、R 4 、R 5 and R 6 are each independently selected from hydrogen, cyano, C2-C6 alkynyl, halogen, hydroxyl, NH2, (C1-C6 alkyl)-NR 7 -, (C1-C6 alkyl)-O-, (C1-C6 alkyl)-S-, C1-C6 alkyl, C3-C6 cycloalkyl, 6-10 membered aryl, C2-C6 alkenyl or C3-C6 cycloalkenyl, said C1-C6 alkyl, C2-C6 alkenyl, C3-C6 cycloalkyl or C3-C6 cycloalkenyl itself or as part of another group is optionally substituted by halogen, cyano, hydroxyl, -NR 7 R 8 , C1-C3 alkyl, C1-C3 alkoxy, C2-C6 alkenyl or C2-C6 alkynyl, the 6-10 membered aryl is optionally substituted by halogen, hydroxyl, cyano, -NR 7 R 8 , NO2, C1-C3 alkyl, C1-C3 alkoxy, C2-C6 alkenyl or C2-C6 alkynyl substituted, wherein the C1-C3 alkyl, C1-C3 alkoxy, C2-C6 alkenyl or C2-C6 alkynyl is optionally substituted with halogen, hydroxy, cyano, -NR 7 R 8 or NO2 replacement;

[0012] R1 and R 2 Each is independently selected from 6-10 membered aryl or 9-18 membered benzoheterocyclyl, wherein the 6-10 membered aryl or 9-18 membered benzoheterocyclyl is optionally substituted by halogen, hydroxyl, cyano, -NR 7 R 8 、NO2、-NR 9 C(O)R 10 , C1-C6 alkyl, (C1-C6 alkyl)-O-, -C(O)NR 9 R 10 or 5-7 membered heteroaryl, wherein the C1-C6 alkyl itself or as part of another group or the 5-7 membered heteroaryl is optionally substituted with halogen, cyano, hydroxy, C1-C3 alkyl, (C1-C3 alkyl)-O- or -NR 7 R 8 replace;

[0013] R 7 、R 8 、R 9 and R 10 Each independently selected from H or C1-C6 alkyl;

[0014] The condition is: at most 2 of W, X, Y and Z are N at the same time;

[0015] The acid in the acid salt is an inorganic acid or an organic acid; the inorganic acid includes hydrochloric acid, sulfuric acid, nitric acid, hydrobromic acid, hydrofluoric acid, hydroiodic acid or phosphoric acid; the organic acid includes formic acid, acetic acid, propionic acid, dichloroacetic acid, trichloroacetic acid, acetohydroxamic acid, adipic acid, benzenesulfonic acid, benzoic acid, phthalic acid, p-hydroxybenzoic acid, capric acid, hexanoic acid, caprylic acid, cinnamic acid, citric acid, succinic acid, cyclohexanesulfamic acid, camphorsulfonic acid, aspartic acid, camphoric acid, D-gluconic acid, glucuronic acid, L-glutamic acid, L-ascorbic acid, lactic acid, flat peach acid, pyroglutamic acid, L-tartaric acid, lauryl sulfuric acid, dibenzoyltartaric acid, ethanesulfonic acid, formic acid, fumaric acid, galactosonic acid, gentisic acid, glutaric acid, glycolic acid, hippuric acid, isethionic acid, lactobionic acid, ascorbic acid, aspartic acid, lauric acid, camphoric acid, maleic acid, malonic acid, methanesulfonic acid, niacin, stearic acid, oleic acid, orotic acid, oxalic acid, palmitic acid, pamoic acid, propionic acid, salicylic acid, sebacic acid, succinic acid, thiocyanic acid, undecylenic acid, trifluoroacetic acid, benzenesulfonic acid, p-toluenesulfonic acid, or L-malic acid.

[0016] The present invention also provides an acid salt of a compound of formula I, wherein the compound of formula I has a structure as shown in formula II,

[0017] Among them, R 1 、R 2 、R 3 、R4 and R 5 The definition of is as defined in the compound of formula I.

[0018] In one embodiment of the present invention, R 3 、R 4 、R 5 and R 6 Each is independently selected from hydrogen, halogen, hydroxy, NH2, (C1-C6 alkyl)-NR 7 -, (C1-C6 alkyl)-O-, C1-C6 alkyl, C3-C6 cycloalkyl or 6-10 membered aryl, wherein the C1-C6 alkyl itself or as part of another group or C3-C6 cycloalkyl is optionally substituted by halogen, cyano, hydroxy or -NR 7 R 8 Substituted, the 6-10 membered aryl group is optionally substituted by a halogen-substituted C1-C3 alkoxy group;

[0019] Preferably, R 3 、R 4 、R 5 and R 6 Each is independently selected from hydrogen, halogen, hydroxy, NH2, (C1-C6 alkyl)-NR 7 -, (C1-C6 alkyl)-O-, C1-C6 alkyl, C3-C6 cycloalkyl or 6-10 membered aryl, wherein the C1-C6 alkyl itself or as part of another group or C3-C6 cycloalkyl is optionally substituted with halogen, and the 6-10 membered aryl is optionally substituted with halogen-substituted C1-C3 alkoxy;

[0020] More preferably, R 3 、R 4 、R 5 and R 6 Each is independently selected from hydrogen, halogen, hydroxy, NH2, (C1-C6 alkyl)-NR 7 -, (C1-C6 alkyl)-O-, C1-C6 alkyl, C3-C6 cycloalkyl or 6-10 membered aryl, wherein the C1-C6 alkyl itself or as part of another group or the C3-C6 cycloalkyl is optionally substituted with fluorine, and the 6-10 membered aryl is optionally substituted with a fluorine-substituted C1-C3 alkoxy group.

[0021] In one embodiment of the present invention, R 3 is selected from hydrogen, C1-C6 alkyl or C3-C6 cycloalkyl; preferably, R 3 is selected from hydrogen or C1-C6 alkyl; more preferably, R 3 is selected from hydrogen, methyl or cyclopropyl; further preferably, R 3 is selected from hydrogen or methyl; most preferably, R 3 Selected from hydrogen.

[0022] In one embodiment of the present invention, R 4 is selected from hydrogen or C1-C6 alkyl; preferably, R 4 is selected from hydrogen or methyl; more preferably, R 4 Selected from hydrogen.

[0023] In one embodiment of the present invention, R 5 Selected from hydrogen, halogen, hydroxy, (C1-C6 alkyl)-NR 7 -, (C1-C6 alkyl) -O-, C3-C6 cycloalkyl or 6-10 membered aryl, wherein the C1-C6 alkyl itself or as part of another group or C3-C6 cycloalkyl is optionally substituted with fluorine, and the 6-10 membered aryl is optionally substituted with fluorine-substituted C1-C3 alkoxy; preferably, R 5 is selected from hydrogen, chlorine, hydroxyl, cyclopropyl, CF3CH2O-, CHF2O-, CF3CH2NH-, 4-difluoromethoxyphenyl or CH3CH2O-; more preferably, R 5 is selected from cyclopropyl, CF3CH2O-, CF3CH2NH- or CH3CH2O-; further preferably, R 5 is selected from cyclopropyl, CF3CH2O- or CF3CH2NH-; most preferably, R 5 Selected from cyclopropyl.

[0024] In one embodiment of the present invention, R 1 and R 2 Each independently selected from phenyl, wherein the groups are optionally substituted by halogen, hydroxy, cyano, -NR 7 R 8 、NO2、-NR 9 C(O)R 10 , C1-C6 alkyl, (C1-C6 alkyl)-O-, -C(O)NR 9 R 10 or 5-7 membered heteroaryl, wherein the C1-C6 alkyl itself or as part of another group or the 5-7 membered heteroaryl is optionally substituted with halogen, cyano, hydroxy, C1-C3 alkyl, (C1-C3 alkyl)-O- or -NR 7 R 8 substituted; preferably, R 1 and R 2 Each independently selected from phenyl, wherein the group is optionally substituted by C1-C6 alkyl or (C1-C6 alkyl)-O-, wherein the C1-C6 alkyl itself or as part of another group is optionally substituted by halogen; preferably, R 7、R 8 、R 9 and R 10 are each independently selected from H.

[0025] In another embodiment of the present invention, R 1 Selected from phenyl, wherein the group is optionally substituted by C1-C6 alkyl or (C1-C6 alkyl)-O-, wherein the C1-C6 alkyl itself or as part of another group is optionally substituted by halogen; preferably, R 1 Selected from More preferably, R 1 Selected from

[0026] In one embodiment of the present invention, R 2 Selected from phenyl or wherein the group is optionally substituted by (C1-C6 alkyl)-O-, and the C1-C6 alkyl is optionally substituted by halogen; preferably, R 2 Selected from More preferably, R 2 Selected from

[0027] In one embodiment of the present invention, the acid in the acid salt is selected from hydrochloric acid, sulfuric acid, maleic acid, phosphoric acid, acetic acid, fumaric acid, L-malic acid, citric acid, succinic acid, adipic acid, L-tartaric acid, L-ascorbic acid, hippuric acid, gentisic acid, malonic acid, methanesulfonic acid, benzoic acid, p-toluenesulfonic acid, phthalic acid, p-hydroxybenzoic acid, propionic acid, oxalic acid, L-glutamic acid, stearic acid or D-gluconic acid.

[0028] In one embodiment of the present invention, the acid salt is selected from hydrochloride, sulfate, phosphate, maleate, citric acid, adipate, methanesulfonate or oxalate.

[0029] The present invention also provides acid salts of the following compounds:

[0030] In one embodiment of the present invention, an acid salt of the compound 2-cyclopropyl-9-[4-(difluoromethoxy)phenyl]-7-(2-methyl-2H-indazol-5-yl)-8H-pyrimido[1,2-b]pyridazin-8-one is provided, and the structure of the compound is as follows:

[0031] In one embodiment of the present invention, an acid salt of the compound 2-cyclopropyl-9-[4-(difluoromethoxy)phenyl]-7-(2-methyl-2H-indazol-5-yl)-8H-pyrimido[1,2-b]pyridazin-8-one is provided, wherein the acid in the acid salt is an inorganic acid or an organic acid; the inorganic acid is selected from hydrochloric acid, sulfuric acid or phosphoric acid; the organic acid is selected from maleic acid, acetic acid, citric acid, succinic acid, adipic acid, L-tartaric acid, L-ascorbic acid, hippuric acid, gentisic acid, malonic acid, methanesulfonic acid, benzoic acid, p-toluenesulfonic acid, phthalic acid, p-hydroxybenzoic acid, propionic acid, oxalic acid, L-glutamic acid, stearic acid or D-gluconic acid.

[0032] In one embodiment of the present invention, an acid salt of the compound 2-cyclopropyl-9-[4-(difluoromethoxy)phenyl]-7-(2-methyl-2H-indazol-5-yl)-8H-pyrimido[1,2-b]pyridazin-8-one is provided, wherein the acid in the acid salt is an inorganic acid or an organic acid; the inorganic acid is selected from hydrochloric acid, sulfuric acid or phosphoric acid; the organic acid is selected from maleic acid, citric acid, adipic acid, methanesulfonic acid or oxalic acid.

[0033] On the other hand, the present invention also provides an acid salt crystalline form of the compound represented by the above formula I.

[0034] In one embodiment of the present invention, a crystalline form of an acid salt of the compound 2-cyclopropyl-9-[4-(difluoromethoxy)phenyl]-7-(2-methyl-2H-indazol-5-yl)-8H-pyrimido[1,2-b]pyridazin-8-one is provided, wherein the acid in the acid salt is an inorganic acid or an organic acid; the inorganic acid is selected from hydrochloric acid, sulfuric acid or phosphoric acid; the organic acid is selected from maleic acid, citric acid, adipic acid, methanesulfonic acid or oxalic acid.

[0035] In one embodiment of the present invention, an acid salt crystalline form of the compound 2-cyclopropyl-9-[4-(difluoromethoxy)phenyl]-7-(2-methyl-2H-indazol-5-yl)-8H-pyrimido[1,2-b]pyridazin-8-one is provided, wherein the acid salt crystalline form is selected from a hydrochloride crystalline form, a sulfate crystalline form, a phosphate crystalline form, a maleic acid crystalline form, a citric acid crystalline form, an adipate crystalline form, a methanesulfonate crystalline form or an oxalate crystalline form.

[0036] In one embodiment of the present invention, the compound 2-cyclopropyl-9-[4-(difluoromethoxy)phenyl]-7-(2-methyl-2H-indazol-5-yl)-8H-pyrimido[1,2-b]pyridazin-8-one is provided as hydrochloride salt form A, sulfate salt form A, phosphate salt form A, maleate salt form A, citrate salt form A, adipate salt form A, adipate salt form B, methanesulfonate salt form A or oxalate salt form A.

[0037] In one embodiment of the present invention, a hydrochloride crystalline form A of the compound 2-cyclopropyl-9-[4-(difluoromethoxy)phenyl]-7-(2-methyl-2H-indazol-5-yl)-8H-pyrimido[1,2-b]pyridazin-8-one is provided, and its X-ray powder diffraction pattern has diffraction peaks at 2θ of 11.5°±0.2°, 17.0°±0.2°, 17.8°±0.2°, 22.4°±0.2°, 23.3°±0.2° and 27.1°±0.2°.

[0038] Preferably, the hydrochloride salt form A has an X-ray powder diffraction pattern having diffraction peaks at 2θ of 6.3°±0.2°, 11.5°±0.2°, 17.0°±0.2°, 17.8°±0.2°, 22.4°±0.2°, 23.3°±0.2°, 24.2°±0.2° and 27.1°±0.2°.

[0039] More preferably, the hydrochloride salt form A has an X-ray powder diffraction pattern having diffraction peaks at 2θ of 4.1°±0.2°, 6.3°±0.2°, 11.5°±0.2°, 17.0°±0.2°, 17.8°±0.2°, 22.4°±0.2°, 22.8°±0.2°, 23.3°±0.2°, 24.2°±0.2° and 27.1°±0.2°.

[0040] Further preferably, the hydrochloride salt form A has an X-ray powder diffraction pattern having diffraction peaks at 2θ of 4.1°±0.2°, 6.3°±0.2°, 8.3°±0.2°, 11.5°±0.2°, 17.0°±0.2°, 17.8°±0.2°, 22.4°±0.2°, 22.8°±0.2°, 23.3°±0.2°, 24.2°±0.2°, 27.1°±0.2° and 27.5°±0.2°.

[0041] In one embodiment, the hydrochloride salt form A has an X-ray powder diffraction pattern of 2θ as shown in the following table:

[0042] Table 1 X-ray powder diffraction pattern data of hydrochloride crystal form A

[0043] In one embodiment, the X-ray powder diffraction pattern of the hydrochloride salt form A expressed in 2θ angles has a pattern as shown in FIG14 .

[0044] In one embodiment, the hydrochloride salt form A has the 1 H NMR spectrum.

[0045] In one embodiment of the present invention, a sulfate salt crystalline form A of the compound 2-cyclopropyl-9-[4-(difluoromethoxy)phenyl]-7-(2-methyl-2H-indazol-5-yl)-8H-pyrimido[1,2-b]pyridazin-8-one is provided, and its X-ray powder diffraction pattern has diffraction peaks at 2θ of 3.4°±0.2°, 15.6°±0.2°, 17.1°±0.2°, 20.6°±0.2°, 23.4°±0.2° and 24.1°±0.2°.

[0046] Preferably, the sulfate crystalline form A has an X-ray powder diffraction pattern having diffraction peaks at 2θ of 3.4°±0.2°, 15.6°±0.2°, 17.1°±0.2°, 20.3°±0.2°, 20.6°±0.2°, 23.4°±0.2°, 24.1°±0.2° and 24.5°±0.2°.

[0047] More preferably, the sulfate crystalline form A has an X-ray powder diffraction pattern having diffraction peaks at 2θ of 3.4°±0.2°, 6.8°±0.2°, 15.6°±0.2°, 17.1°±0.2°, 20.3°±0.2°, 20.6°±0.2°, 23.4°±0.2°, 24.1°±0.2°, 24.5°±0.2° and 26.6°±0.2°.

[0048] Further preferably, the sulfate crystalline form A has an X-ray powder diffraction pattern having diffraction peaks at 2θ of 3.4°±0.2°, 6.8°±0.2°, 15.6°±0.2°, 17.1°±0.2°, 20.3°±0.2°, 20.6°±0.2°, 23.4°±0.2°, 24.1°±0.2°, 24.5°±0.2°, 26.6°±0.2°, 27.7°±0.2° and 28.0°±0.2°.

[0049] In one embodiment, the sulfate salt crystalline form A has an X-ray powder diffraction pattern of 2θ as shown in the following table:

[0050] Table 2 X-ray powder diffraction pattern data of sulfate crystal form A

[0051] In one embodiment, the X-ray powder diffraction of the sulfate salt crystalline form A expressed in 2θ angle has a pattern as shown in FIG16 .

[0052] In one embodiment, the sulfate salt crystalline form A has a hydrogen spectrum as shown in FIG17 .

[0053] In one embodiment of the present invention, a phosphate crystalline form A of the compound 2-cyclopropyl-9-[4-(difluoromethoxy)phenyl]-7-(2-methyl-2H-indazol-5-yl)-8H-pyrimido[1,2-b]pyridazin-8-one is provided, and its X-ray powder diffraction pattern has diffraction peaks at 2θ of 12.2°±0.2°, 16.1°±0.2°, 19.9°±0.2°, 22.4°±0.2°, 23.2°±0.2° and 28.7°±0.2°.

[0054] Preferably, the phosphate crystal form A has an X-ray powder diffraction pattern having diffraction peaks at 2θ of 12.2°±0.2°, 16.1°±0.2°, 19.4°±0.2°, 19.9°±0.2°, 21.5°±0.2°, 22.4°±0.2°, 23.2°±0.2° and 28.7°±0.2°.

[0055] More preferably, the phosphate crystalline form A has an X-ray powder diffraction pattern having diffraction peaks at 2θ of 12.2°±0.2°, 16.1°±0.2°, 18.1°±0.2°, 19.4°±0.2°, 19.9°±0.2°, 21.5°±0.2°, 22.4°±0.2°, 23.2°±0.2°, 25.9°±0.2° and 28.7°±0.2°.

[0056] Further preferably, the phosphate crystalline form A has an X-ray powder diffraction pattern having diffraction peaks at 2θ of 12.2°±0.2°, 14.9°±0.2°, 16.1°±0.2°, 18.1°±0.2°, 19.4°±0.2°, 19.9°±0.2°, 20.4°±0.2°, 21.5°±0.2°, 22.4°±0.2°, 23.2°±0.2°, 25.9°±0.2° and 28.7°±0.2°.

[0057] In one embodiment, the phosphate crystal form A has an X-ray powder diffraction pattern of 2θ as shown in the following table:

[0058] Table 3 X-ray powder diffraction pattern data of phosphate crystal form A

[0059] In one embodiment, the X-ray powder diffraction of the phosphate crystal form A expressed in 2θ angle has a pattern as shown in FIG18 .

[0060] In one embodiment of the present invention, a maleate salt form A of the compound 2-cyclopropyl-9-[4-(difluoromethoxy)phenyl]-7-(2-methyl-2H-indazol-5-yl)-8H-pyrimido[1,2-b]pyridazin-8-one is provided, and its X-ray powder diffraction pattern has diffraction peaks at 2θ of 6.7°±0.2°, 7.3°±0.2°, 14.7°±0.2°, 16.7°±0.2°, 20.2°±0.2° and 22.0°±0.2°.

[0061] Preferably, the maleate salt crystalline form A has an X-ray powder diffraction pattern having diffraction peaks at 2θ of 6.7°±0.2°, 7.3°±0.2°, 13.4°±0.2°, 14.7°±0.2°, 16.7°±0.2°, 20.2°±0.2°, 22.0°±0.2° and 26.9°±0.2°.

[0062] More preferably, the maleate salt form A has an X-ray powder diffraction pattern having diffraction peaks at 2θ of 6.7°±0.2°, 7.3°±0.2°, 13.4°±0.2°, 14.7°±0.2°, 16.7°±0.2°, 18.7°±0.2°, 20.2°±0.2°, 22.0°±0.2°, 24.6°±0.2° and 26.9°±0.2°.

[0063] Further preferably, the maleate salt form A has an X-ray powder diffraction pattern having diffraction peaks at 2θ of 6.7°±0.2°, 7.3°±0.2°, 8.0°±0.2°, 13.4°±0.2°, 14.7°±0.2°, 16.7°±0.2°, 18.7°±0.2°, 20.2°±0.2°, 22.0°±0.2°, 24.1°±0.2°, 24.6°±0.2° and 26.9°±0.2°.

[0064] In one embodiment, the maleate salt crystalline form A has an X-ray powder diffraction pattern of 2θ as shown in the following table:

[0065] Table 4 X-ray powder diffraction pattern data of maleate salt form A

[0066] In one embodiment, the X-ray powder diffraction of the maleate salt form A expressed in 2θ angle has a pattern as shown in FIG19 .

[0067] In one embodiment, the maleate salt crystalline form A has a hydrogen spectrum as shown in Figure 20.

[0068] In one embodiment of the present invention, a citrate crystalline form A of the compound 2-cyclopropyl-9-[4-(difluoromethoxy)phenyl]-7-(2-methyl-2H-indazol-5-yl)-8H-pyrimido[1,2-b]pyridazin-8-one is provided, and its X-ray powder diffraction pattern has diffraction peaks at 2θ of 4.4°±0.2°, 9.2°±0.2°, 13.9°±0.2°, 17.5°±0.2°, 18.0°±0.2° and 21.6°±0.2°.

[0069] Preferably, the citrate salt crystalline form A has an X-ray powder diffraction pattern having diffraction peaks at 2θ of 4.4°±0.2°, 9.2°±0.2°, 13.9°±0.2°, 14.3°±0.2°, 15.3°±0.2°, 17.5°±0.2°, 18.0°±0.2° and 21.6°±0.2°.

[0070] More preferably, the citrate salt form A has an X-ray powder diffraction pattern having diffraction peaks at 2θ of 4.4°±0.2°, 9.2°±0.2°, 13.4°±0.2°, 13.9°±0.2°, 14.3°±0.2°, 15.3°±0.2°, 17.5°±0.2°, 18.0°±0.2°, 21.6°±0.2° and 22.6°±0.2°.

[0071] Further preferably, the citrate salt form A has an X-ray powder diffraction pattern having diffraction peaks at 2θ of 4.4°±0.2°, 7.6°±0.2°, 9.2°±0.2°, 12.2°±0.2°, 13.4°±0.2°, 13.9°±0.2°, 14.3°±0.2°, 15.3°±0.2°, 17.5°±0.2°, 18.0°±0.2°, 21.6°±0.2° and 22.6°±0.2°.

[0072] In one embodiment, the citrate salt crystalline form A has an X-ray powder diffraction pattern of 2θ as shown in the following table:

[0073] Table 5 X-ray powder diffraction pattern data of citrate crystal form A

[0074] In one embodiment, the X-ray powder diffraction of the citrate salt form A expressed in 2θ angle has a pattern as shown in FIG21 .

[0075] In one embodiment, the citrate salt crystalline form A has a hydrogen spectrum as shown in FIG22 .

[0076] In one embodiment of the present invention, a crystalline form A of the adipate salt of the compound 2-cyclopropyl-9-[4-(difluoromethoxy)phenyl]-7-(2-methyl-2H-indazol-5-yl)-8H-pyrimido[1,2-b]pyridazin-8-one is provided, and its X-ray powder diffraction pattern has diffraction peaks at 2θ of 15.2°±0.2°, 17.6°±0.2°, 19.7°±0.2°, 21.8°±0.2°, 23.8°±0.2° and 24.6°±0.2°.

[0077] Preferably, the adipate salt crystalline form A has an X-ray powder diffraction pattern having diffraction peaks at 2θ of 9.2°±0.2°, 15.2°±0.2°, 17.6°±0.2°, 19.7°±0.2°, 21.8°±0.2°, 22.6°±0.2°, 23.8°±0.2° and 24.6°±0.2°.

[0078] More preferably, the adipate salt crystalline form A has an X-ray powder diffraction pattern having diffraction peaks at 2θ of 9.2°±0.2°, 14.2°±0.2°, 15.2°±0.2°, 17.6°±0.2°, 19.7°±0.2°, 21.8°±0.2°, 22.6°±0.2°, 23.8°±0.2°, 24.6°±0.2° and 25.4°±0.2°.

[0079] Further preferably, the adipate salt crystalline form A has an X-ray powder diffraction pattern having diffraction peaks at 2θ of 9.2°±0.2°, 13.0°±0.2°, 14.2°±0.2°, 15.2°±0.2°, 17.6°±0.2°, 19.7°±0.2°, 21.8°±0.2°, 22.6°±0.2°, 23.1°±0.2°, 23.8°±0.2°, 24.6°±0.2° and 25.4°±0.2°.

[0080] Further preferably, the adipate salt crystalline form A has an X-ray powder diffraction pattern having diffraction peaks at 2θ of 9.2°±0.2°, 11.3°±0.2°, 13.0°±0.2°, 14.2°±0.2°, 15.2°±0.2°, 17.6°±0.2°, 19.7°±0.2°, 21.8°±0.2°, 22.6°±0.2°, 23.1°±0.2°, 23.8°±0.2°, 24.6°±0.2°, 25.4°±0.2° and 27.1°±0.2°.

[0081] In one embodiment, the 2θ of the X-ray powder diffraction pattern of the adipate salt form A is shown in the following table:

[0082] Table 6 X-ray powder diffraction pattern data of adipate salt form A

[0083] In one embodiment, the X-ray powder diffraction of the adipate salt form A expressed in 2θ angle has a pattern as shown in FIG23 .

[0084] In one embodiment of the present invention, a crystalline form B of the adipate salt of the compound 2-cyclopropyl-9-[4-(difluoromethoxy)phenyl]-7-(2-methyl-2H-indazol-5-yl)-8H-pyrimido[1,2-b]pyridazin-8-one is provided, and its X-ray powder diffraction pattern has diffraction peaks at 2θ of 8.7°±0.2°, 9.2°±0.2°, 13.7°±0.2°, 17.5°±0.2°, 17.9°±0.2° and 19.3°±0.2°.

[0085] Preferably, the adipate salt crystalline form B has an X-ray powder diffraction pattern having diffraction peaks at 2θ of 8.7°±0.2°, 9.2°±0.2°, 13.7°±0.2°, 17.5°±0.2°, 17.9°±0.2°, 19.3°±0.2°, 23.4°±0.2° and 24.0°±0.2°.

[0086] More preferably, the adipate salt form B has an X-ray powder diffraction pattern having diffraction peaks at 2θ of 5.9°±0.2°, 8.7°±0.2°, 9.2°±0.2°, 13.7°±0.2°, 16.8°±0.2°, 17.5°±0.2°, 17.9°±0.2°, 19.3°±0.2°, 23.4°±0.2° and 24.0°±0.2°.

[0087] Further preferably, the adipate salt form B has an X-ray powder diffraction pattern having diffraction peaks at 2θ of 5.9°±0.2°, 8.7°±0.2°, 9.2°±0.2°, 13.7°±0.2°, 15.4°±0.2°, 16.8°±0.2°, 17.5°±0.2°, 17.9°±0.2°, 19.3°±0.2°, 23.4°±0.2°, 24.0°±0.2° and 25.4°±0.2°.

[0088] In one embodiment, the adipate salt crystalline form B has an X-ray powder diffraction pattern of 2θ as shown in the following table:

[0089] Table 7 X-ray powder diffraction pattern data of adipate salt form B

[0090] In one embodiment, the X-ray powder diffraction of the adipate salt form B expressed in 2θ angle has a pattern as shown in FIG24 .

[0091] In one embodiment, the adipate salt form B has a hydrogen spectrum as shown in FIG25 .

[0092] In one embodiment of the present invention, a crystalline form A of the methanesulfonate of the compound 2-cyclopropyl-9-[4-(difluoromethoxy)phenyl]-7-(2-methyl-2H-indazol-5-yl)-8H-pyrimido[1,2-b]pyridazin-8-one is provided, and its X-ray powder diffraction pattern has diffraction peaks at 2θ of 11.2°±0.2°, 13.0°±0.2°, 18.1°±0.2°, 20.3°±0.2°, 22.6°±0.2° and 25.9°±0.2°.

[0093] Preferably, the mesylate salt form A has an X-ray powder diffraction pattern having diffraction peaks at 2θ of 11.2°±0.2°, 13.0°±0.2°, 18.1°±0.2°, 20.3°±0.2°, 21.2°±0.2°, 21.4°±0.2°, 22.6°±0.2° and 25.9°±0.2°.

[0094] More preferably, the mesylate salt form A has an X-ray powder diffraction pattern at 2θ of 11.2°±0.2°, 13.0°±0.2°, 18.1°±0.2°, 20.3°±0.2°, 21.2°±0.2°, 21.4°±0.2°, 22.6°±0.2°, 23.9°±0.2°, 24.5°±0.2° and 25.9°±0.2°.

[0095] Further preferably, the mesylate salt form A has an X-ray powder diffraction pattern at 2θ of 11.2°±0.2°, 13.0°±0.2°, 17.2°±0.2°, 18.1°±0.2°, 20.3°±0.2°, 21.2°±0.2°, 21.4°±0.2°, 22.6°±0.2°, 23.5°±0.2°, 23.9°±0.2°, 24.5°±0.2° and 25.9°±0.2° having diffraction peaks.

[0096] In one embodiment, the 2θ of the X-ray powder diffraction pattern of the mesylate salt form A is shown in the following table:

[0097] Table 8 X-ray powder diffraction pattern data of mesylate salt form A

[0098] In one embodiment, the X-ray powder diffraction pattern of the mesylate salt form A expressed in 2θ angles is as shown in FIG26 .

[0099] In one embodiment, the mesylate salt Form A has a hydrogen spectrum as shown in FIG27 .

[0100] In one embodiment of the present invention, there is provided a crystalline form A of the oxalate salt of the compound 2-cyclopropyl-9-[4-(difluoromethoxy)phenyl]-7-(2-methyl-2H-indazol-5-yl)-8H-pyrimido[1,2-b]pyridazin-8-one, which has an X-ray powder diffraction pattern having diffraction peaks at 2θ of 10.7°±0.2°, 12.7°±0.2°, 15.4°±0.2°, 22.4°±0.2°, 24.6°±0.2° and 24.9°±0.2°.

[0101] Preferably, the oxalate crystalline form A has an X-ray powder diffraction pattern having diffraction peaks at 2θ of 7.5°±0.2°, 10.7°±0.2°, 12.7°±0.2°, 15.4°±0.2°, 17.6°±0.2°, 22.4°±0.2°, 24.6°±0.2° and 24.9°±0.2°.

[0102] More preferably, the oxalate salt form A has an X-ray powder diffraction pattern having diffraction peaks at 2θ of 7.5°±0.2°, 10.7°±0.2°, 12.7°±0.2°, 14.8°±0.2°, 15.4°±0.2°, 16.4°±0.2°, 17.6°±0.2°, 22.4°±0.2°, 24.6°±0.2° and 24.9°±0.2°.

[0103] Further preferably, the oxalate salt form A has an X-ray powder diffraction pattern having diffraction peaks at 2θ of 7.5°±0.2°, 10.7°±0.2°, 12.7°±0.2°, 14.8°±0.2°, 15.4°±0.2°, 16.4°±0.2°, 17.6°±0.2°, 18.8°±0.2°, 22.4°±0.2°, 24.6°±0.2°, 24.9°±0.2° and 25.5°±0.2°.

[0104] In one embodiment, the oxalate crystalline form A has an X-ray powder diffraction pattern of 2θ as shown in the following table:

[0105] Table 9 X-ray powder diffraction pattern data of oxalate crystal form A

[0106] In one embodiment, the X-ray powder diffraction of the oxalate salt form A expressed in 2θ angle has a pattern as shown in FIG28 .

[0107] In one embodiment, the oxalate salt form A has a hydrogen spectrum as shown in Figure 29.

[0108] In a preferred embodiment of the present invention, the acid salt crystalline form of the compound is a solvent-containing or solvent-free crystalline form, wherein the solvent is selected from water, methanol, ethanol, n-propanol, isopropanol, tert-butanol, n-butanol, isobutanol, acetone, 2-butanone, 3-pentanone, dichloromethane, chloroform (chloroform), ethyl formate, ethyl acetate, acetonitrile, tetrahydrofuran, 2-methyl-tetrahydrofuran, 1,4-dioxane, benzene, toluene, N,N-dimethylformamide, N,N-dimethylacetamide, N-methylpyrrolidone, dimethyl sulfoxide, n-heptane, heptane, isopropyl acetate, cyclohexane, methyl tert-butyl ether and one or more of isopropyl ether.

[0109] In a preferred embodiment of the present invention, the acid salt crystalline form of the compound 2-cyclopropyl-9-[4-(difluoromethoxy)phenyl]-7-(2-methyl-2H-indazol-5-yl)-8H-pyrimido[1,2-b]pyridazin-8-one is a solvent-containing or solvent-free crystalline form, and the solvent is selected from one or more of water, ethanol, acetone, dichloromethane, chloroform, 2-butanone, tetrahydrofuran and N,N-dimethylformamide.

[0110] In a further preferred embodiment of the present invention, the crystalline form of the acid salt of the compound 2-cyclopropyl-9-[4-(difluoromethoxy)phenyl]-7-(2-methyl-2H-indazol-5-yl)-8H-pyrimido[1,2-b]pyridazin-8-one is an anhydrous crystalline form.

[0111] On the other hand, the present invention also provides a method for preparing an acid salt or a crystalline form of an acid salt of a compound represented by formula I, comprising the step of forming a salt from the compound represented by formula I with a corresponding acid.

[0112] On the other hand, the present invention also provides a method for preparing an acid salt crystalline form of a compound represented by formula I, comprising the step of forming a salt by reacting a compound of formula I with a corresponding acid in a reaction solvent; the reaction solvent is selected from one or more of water, methanol, ethanol, n-propanol, isopropanol, tert-butanol, n-butanol, isobutanol, acetone, 2-butanone, 3-pentanone, dichloromethane, chloroform (chloroform), ethyl formate, ethyl acetate, acetonitrile, tetrahydrofuran, 2-methyl-tetrahydrofuran, 1,4-dioxane, benzene, toluene, N,N-dimethylformamide, N,N-dimethylacetamide, N-methylpyrrolidone, dimethyl sulfoxide, n-heptane, heptane, isopropyl acetate, cyclohexane, methyl tert-butyl ether and isopropyl ether.

[0113] In some embodiments of the present invention, the reaction solvent is selected from one or more of water, ethanol, acetone, 2-butanone or chloroform.

[0114] In some embodiments of the present invention, the molar ratio of the compound of formula I to the corresponding acid in the preparation method is 1:0.1-3; preferably 1:0.1-1.5; more preferably 1:0.3-1.

[0115] In some embodiments of the present invention, the mass volume ratio of the compound of formula I to the reaction solvent in the preparation method is 1:10-50; preferably 1:20-30; more preferably 1:25-30.

[0116] On the other hand, the present invention also provides a pharmaceutical composition containing a therapeutically effective amount of the acid salt or acid salt crystalline form of the compound represented by Formula I.

[0117] On the other hand, the present invention also provides a pharmaceutical composition comprising a therapeutically effective amount of the acid salt or acid salt crystalline form of the compound represented by Formula I and a pharmaceutically acceptable carrier.

[0118] The pharmaceutical compositions of the present invention can be administered by any suitable route or method, such as oral or parenteral (e.g., intravenous) administration. The therapeutically effective amount of the acid salt or crystalline form of the aforementioned compound (calculated as the free base of the compound) is from about 1 mg to 1 g / kg body weight / day.

[0119] For oral administration, the pharmaceutical compositions of the present invention are typically provided in the form of tablets, capsules, or solutions. Tablets may comprise an acid salt or a crystalline form of the compound of the present invention and a pharmaceutically acceptable carrier. The carrier may include, but is not limited to, a diluent, a disintegrant, a binder, and a lubricant.

[0120] For parenteral administration, the pharmaceutical composition of the present invention can be administered by intravenous injection, intramuscular injection or subcutaneous injection. It is usually provided in a sterile aqueous solution or suspension or lyophilized powder, and the appropriate pH and isotonicity are adjusted.

[0121] On the other hand, the present invention also provides the use of the acid salt or acid salt crystalline form of the aforementioned compound or a pharmaceutical composition thereof in the preparation of a medicament for preventing and / or treating a disease or disease state mediated by MAT2A.

[0122] On the other hand, the present invention also provides a method for preventing and / or treating MAT2A-mediated diseases or disease states, comprising administering an effective amount of the acid salt or acid salt crystalline form of the aforementioned compound or a pharmaceutical composition thereof to an individual in need.

[0123] In another aspect, the present invention further provides an acid salt or acid salt crystalline form of the aforementioned compound of the present invention or a pharmaceutical composition of the present invention for use in preventing and / or treating a disease or condition mediated by MAT2A. Examples of the MAT2A-mediated disease or condition include colorectal cancer, etc.

[0124] In some embodiments, the MAT2A-mediated disease is a disease mediated by MAT2A overexpression.

[0125] Related definitions

[0126] Unless otherwise specified, the following terms used in the specification and claims have the following meanings:

[0127] The term "optional" or "optionally" means that the subsequently described event or circumstance may or may not occur, and that the description includes both occurring and not occurring.

[0128] As used herein, numerical ranges refer to the individual integers within the given range. For example, "C1-C6" means that the group can have 1 carbon atom, 2 carbon atoms, 3 carbon atoms, 4 carbon atoms, 5 carbon atoms, or 6 carbon atoms; "C3-C6" means that the group can have 3 carbon atoms, 4 carbon atoms, 5 carbon atoms, or 6 carbon atoms.

[0129] The term "membered" refers to the number of skeletal atoms or groups of atoms that make up the ring. For example, "5-7 membered" means that the number of skeletal atoms or groups of atoms that make up the ring is 5, 6, or 7. Thus, for example, pyridine, piperidine, piperazine, and benzene are six-membered rings, while thiophene and pyrrole are five-membered rings.

[0130] The term "substituted" means that any one or more hydrogen atoms on a specified group are replaced by a substituent, as long as the valence state of the specified group is normal and the compound after the substitution is stable. For example, "substituted with halogen" means that any one or more hydrogen atoms on a specified group are replaced by a halogen, as long as the valence state of the specified group is normal and the compound after the substitution is stable.

[0131] The term "alkyl" refers to a saturated aliphatic hydrocarbon group, including straight-chain or branched saturated hydrocarbon groups, having the indicated number of carbon atoms. For example, the term "C1-C6 alkyl" includes C1 alkyl, C2 alkyl, C3 alkyl, C4 alkyl, C5 alkyl, or C6 alkyl. Examples include, but are not limited to, methyl, ethyl, n-propyl, isopropyl, n-butyl, isobutyl, tert-butyl, n-pentyl, 2-pentyl, 3-pentyl, n-hexyl, 2-hexyl, or 3-hexyl.

[0132] The term "cycloalkyl" refers to a monocyclic saturated hydrocarbon system with no heteroatoms and no double bonds. Examples of the term "3-6 membered cycloalkyl" include, but are not limited to, cyclopropyl, cyclobutyl, cyclopentyl or cyclohexyl.

[0133] The term "halogen" refers to fluorine, chlorine, bromine and iodine.

[0134] The term "aryl" refers to an all-carbon monocyclic or fused bicyclic aromatic ring group having a conjugated π-electron system, derived by removing a single hydrogen atom from a single carbon atom of a parent aromatic ring system. This includes bicyclic groups fused to saturated, partially unsaturated, or aromatic carbocyclic rings; examples include, but are not limited to, phenyl, naphthyl, anthracenyl, indene, indane, 1,2-dihydronaphthalene, or 1,2,3,4-tetrahydronaphthalene.

[0135] The term "heteroaryl" refers to a monovalent aromatic group containing at least one heteroatom independently selected from nitrogen, oxygen and sulfur. Examples of "5-7 membered heteroaryl" include, but are not limited to, pyridyl, thienyl, imidazolyl, pyrimidinyl, pyridyl, furyl, pyrazinyl or thiazolyl.

[0136] The term "9-18 membered benzoheterocyclyl" refers to a ring system having 9-18 ring atoms or ring atom groups formed by the fusion of a benzene ring and a heterocycle, wherein the benzene ring and the heterocycle share a pair of adjacent ring atoms, and the connection site with the parent core structure is located in the benzene ring portion. The heterocycle portion is a 5-12 membered saturated, partially unsaturated or fully unsaturated ring system having ring carbon atoms and 1 to 4 ring heteroatoms or heteroatoms, and the heteroatoms or heteroatoms are independently selected from nitrogen, sulfur, oxygen, sulfoxide, sulfone, Heterocycles can be monocyclic, bicyclic, or tricyclic ring systems, in which two or more rings are present in fused, spiro, or bridged forms. Examples include, but are not limited to,

[0137] in Refers to the chemical bond connection. When it appears in a bicyclic or polycyclic ring When the connection position is uncertain, it means that the connection site is limited to Any atom on the monocyclic ring where the valence permits. For example, It means that the connection sites are located only on any carbon atom of the benzene ring in the bicyclic ring and must meet the requirements of atomic valence bonds.

[0138] The "X-ray powder diffraction pattern" in the present invention is obtained by measuring using Cu-Kα radiation. It should be noted that in the X-ray powder diffraction spectrum (XRD), the diffraction spectrum obtained by the crystalline compound is often characteristic for a specific crystal, wherein the relative intensity of the bands (especially at low angles) may vary due to the dominant orientation effect caused by differences in crystallization conditions, particle size and other measurement conditions. Therefore, the relative intensity of the diffraction peak is not characteristic for the crystal targeted. When judging whether it is the same as a known crystal, it is more important to pay attention to the relative position of the peak rather than their relative intensity. In addition, for any given crystal, there may be slight errors in the position of the peak, which is also well known in the field of crystallography. For example, due to changes in temperature during sample analysis, sample movement, or instrument calibration, the position of the peak can move, and the measurement error of the 2θ value is sometimes about ±0.5° and sometimes about ±0.2°. Therefore, this error should be taken into account when determining each crystal structure. When the 2θ deviation of the key characteristic peak shift is around ±0.5°, especially around ±0.2°, they can all be identified as the same crystal form.

[0139] Differential Scanning Calorimetry (DSC) measures the transition temperatures of a crystal when heat is absorbed or released due to changes in its crystal structure or melting. For the same crystalline form of the same compound, the thermal transition temperatures and melting points typically agree within about 5°C, usually about 3°C, in consecutive analyses. When a compound is described as having a given DSC peak or melting point, this refers to the DSC peak or melting point ±5°C. DSC provides an auxiliary method for distinguishing between different crystalline forms. Different crystalline forms can be identified by their distinct transition temperature signatures. It should be noted that for mixtures, the DSC peak or melting point may fluctuate over a wider range. Furthermore, because decomposition occurs during melting, the melting temperature is related to the heating rate.

[0140] Thermogravimetric analysis (TGA) is a thermal analysis technique that measures the relationship between the mass of a sample and temperature changes under a controlled temperature. When the substance being tested sublimes or vaporizes during heating, releasing gases or losing water of crystallization, the mass of the substance changes. In this case, the thermogravimetric curve is not a straight line but rather shows a downward trend. By analyzing the thermogravimetric curve, we can determine the temperature at which the change occurs and, based on the weight loss, calculate the amount of mass lost.

[0141] The term "as shown" when referring to, for example, an XRD pattern, a DSC pattern, or a TGA pattern includes patterns that are not necessarily identical to those depicted herein, but that fall within the limits of experimental error when considered by one skilled in the art.

[0142] The term "effective amount" or "therapeutically effective amount" refers to a non-toxic but sufficient amount of a drug or pharmaceutical agent to achieve the desired effect.

[0143] The term "pharmaceutically acceptable carrier" refers to any carrier that is non-irritating to the body and does not impair the biological activity and properties of the active compound. This includes, but is not limited to, any diluent, disintegrant, binder, glidant, or wetting agent approved by the State Food and Drug Administration for use in humans or animals.

[0144] The abbreviations used in the claims and description have the following meanings:

[0145] M:mol / L;

[0146] mM: mmol / L;

[0147] μM: μmol / L;

[0148] nM: nmol / L;

[0149] LCMS: liquid chromatography-mass spectrometry;

[0150] Brij35: lauryl polyoxyethylene ether;

[0151] BSA: bovine serum albumin;

[0152] DMSO: dimethyl sulfoxide;

[0153] rpm: revolutions per minute;

[0154] Tris-HCl: tris(hydroxymethyl)aminomethane hydrochloride;

[0155] OD 620 : Absorbance at 620nm wavelength;

[0156] SAM: S-adenosylmethionine or S-adenosylmethionine;

[0157] MeOH: methanol;

[0158] 1,4-Dioxane:1,4-dioxane;

[0159] EtOH: ethanol;

[0160] ACN: acetonitrile;

[0161] DMF: N,N-dimethylformamide;

[0162] DCM: dichloromethane;

[0163] CHCl3: chloroform;

[0164] Acetone: acetone;

[0165] MEK: 2-butanone. BRIEF DESCRIPTION OF THE DRAWINGS

[0166] Figure 1 shows the HCT116 MTAP in Test Example 3 - / - Tumor inhibition curve of transplanted tumor model.

[0167] Figure 2 shows the HCT116MTAP in test case 3. - / - Results of intratumoral SAM inhibition in the transplanted tumor model.

[0168] FIG3 is the tumor inhibition curve of the KP-4 transplanted tumor model in Test Example 4.

[0169] FIG4 shows the results of intratumoral SAM inhibition in the KP-4 transplanted tumor model in Test Example 4.

[0170] Figure 5 is an X-ray powder diffraction pattern of Form A of the compound 2-cyclopropyl-9-[4-(difluoromethoxy)phenyl]-7-(2-methyl-2H-indazol-5-yl)-8H-pyrimido[1,2-b]pyridazin-8-one.

[0171] Figure 6 is a TGA spectrum of compound 2-cyclopropyl-9-[4-(difluoromethoxy)phenyl]-7-(2-methyl-2H-indazol-5-yl)-8H-pyrimido[1,2-b]pyridazin-8-one Form A.

[0172] Figure 7 is the DSC spectrum of compound 2-cyclopropyl-9-[4-(difluoromethoxy)phenyl]-7-(2-methyl-2H-indazol-5-yl)-8H-pyrimido[1,2-b]pyridazin-8-one Form A.

[0173] Figure 8 is an X-ray powder diffraction pattern of Form B of the compound 2-cyclopropyl-9-[4-(difluoromethoxy)phenyl]-7-(2-methyl-2H-indazol-5-yl)-8H-pyrimido[1,2-b]pyridazin-8-one.

[0174] Figure 9 is a TGA spectrum of compound 2-cyclopropyl-9-[4-(difluoromethoxy)phenyl]-7-(2-methyl-2H-indazol-5-yl)-8H-pyrimido[1,2-b]pyridazin-8-one Form B.

[0175] Figure 10 is the DSC spectrum of the compound 2-cyclopropyl-9-[4-(difluoromethoxy)phenyl]-7-(2-methyl-2H-indazol-5-yl)-8H-pyrimido[1,2-b]pyridazin-8-one Form B.

[0176] Figure 11 is an X-ray powder diffraction pattern of the compound 2-cyclopropyl-9-[4-(difluoromethoxy)phenyl]-7-(2-methyl-2H-indazol-5-yl)-8H-pyrimido[1,2-b]pyridazin-8-one Form G.

[0177] Figure 12 is a TGA spectrum of compound 2-cyclopropyl-9-[4-(difluoromethoxy)phenyl]-7-(2-methyl-2H-indazol-5-yl)-8H-pyrimido[1,2-b]pyridazin-8-one Form G.

[0178] Figure 13 is the DSC spectrum of compound 2-cyclopropyl-9-[4-(difluoromethoxy)phenyl]-7-(2-methyl-2H-indazol-5-yl)-8H-pyrimido[1,2-b]pyridazin-8-one Form G.

[0179] Figure 14 is an X-ray powder diffraction pattern of the compound 2-cyclopropyl-9-[4-(difluoromethoxy)phenyl]-7-(2-methyl-2H-indazol-5-yl)-8H-pyrimido[1,2-b]pyridazin-8-one hydrochloride Form A.

[0180] Figure 15 is a crystalline form A of the compound 2-cyclopropyl-9-[4-(difluoromethoxy)phenyl]-7-(2-methyl-2H-indazol-5-yl)-8H-pyrimido[1,2-b]pyridazin-8-one hydrochloride 1 H NMR spectrum.

[0181] Figure 16 is an X-ray powder diffraction pattern of compound 2-cyclopropyl-9-[4-(difluoromethoxy)phenyl]-7-(2-methyl-2H-indazol-5-yl)-8H-pyrimido[1,2-b]pyridazin-8-one sulfate crystal form A.

[0182] Figure 17 is a crystalline form A of the compound 2-cyclopropyl-9-[4-(difluoromethoxy)phenyl]-7-(2-methyl-2H-indazol-5-yl)-8H-pyrimido[1,2-b]pyridazin-8-one sulfate 1 H NMR spectrum.

[0183] Figure 18 is an X-ray powder diffraction pattern of compound 2-cyclopropyl-9-[4-(difluoromethoxy)phenyl]-7-(2-methyl-2H-indazol-5-yl)-8H-pyrimido[1,2-b]pyridazin-8-one phosphate crystal form A.

[0184] Figure 19 is an X-ray powder diffraction pattern of Form A of 2-cyclopropyl-9-[4-(difluoromethoxy)phenyl]-7-(2-methyl-2H-indazol-5-yl)-8H-pyrimido[1,2-b]pyridazin-8-one maleate salt.

[0185] Figure 20 is a crystal of the maleate salt of compound 2-cyclopropyl-9-[4-(difluoromethoxy)phenyl]-7-(2-methyl-2H-indazol-5-yl)-8H-pyrimido[1,2-b]pyridazin-8-one Form A. 1 H NMR spectrum.

[0186] Figure 21 is an X-ray powder diffraction pattern of Form A of 2-cyclopropyl-9-[4-(difluoromethoxy)phenyl]-7-(2-methyl-2H-indazol-5-yl)-8H-pyrimido[1,2-b]pyridazin-8-one citrate salt.

[0187] Figure 22 shows the crystal structure of the compound 2-cyclopropyl-9-[4-(difluoromethoxy)phenyl]-7-(2-methyl-2H-indazol-5-yl)-8H-pyrimido[1,2-b]pyridazin-8-one citrate salt Form A. 1 H NMR spectrum.

[0188] Figure 23 is an X-ray powder diffraction pattern of compound 2-cyclopropyl-9-[4-(difluoromethoxy)phenyl]-7-(2-methyl-2H-indazol-5-yl)-8H-pyrimido[1,2-b]pyridazin-8-one adipate salt form A.

[0189] Figure 24 is an X-ray powder diffraction pattern of compound 2-cyclopropyl-9-[4-(difluoromethoxy)phenyl]-7-(2-methyl-2H-indazol-5-yl)-8H-pyrimido[1,2-b]pyridazin-8-one adipate salt Form B.

[0190] Figure 25 is a diagram of the compound 2-cyclopropyl-9-[4-(difluoromethoxy)phenyl]-7-(2-methyl-2H-indazol-5-yl)-8H-pyrimido[1,2-b]pyridazin-8-one adipate salt crystal form B 1 H NMR spectrum.

[0191] Figure 26 is an X-ray powder diffraction pattern of Form A of 2-cyclopropyl-9-[4-(difluoromethoxy)phenyl]-7-(2-methyl-2H-indazol-5-yl)-8H-pyrimido[1,2-b]pyridazin-8-one methanesulfonate.

[0192] Figure 27 is a crystal of the compound 2-cyclopropyl-9-[4-(difluoromethoxy)phenyl]-7-(2-methyl-2H-indazol-5-yl)-8H-pyrimido[1,2-b]pyridazin-8-one methanesulfonate Form A 1 H NMR spectrum.

[0193] Figure 28 is an X-ray powder diffraction pattern of compound 2-cyclopropyl-9-[4-(difluoromethoxy)phenyl]-7-(2-methyl-2H-indazol-5-yl)-8H-pyrimido[1,2-b]pyridazin-8-one oxalate salt Form A.

[0194] Figure 29 shows the crystal structure of 2-cyclopropyl-9-[4-(difluoromethoxy)phenyl]-7-(2-methyl-2H-indazol-5-yl)-8H-pyrimido[1,2-b]pyridazin-8-one oxalate salt Form A. 1 H NMR spectrum. DETAILED DESCRIPTION

[0195] The preparation method of compound of the present invention is described in more detail below, but these specific preparation methods do not constitute any restriction to the scope of the present invention.In addition, reaction conditions such as the amount of reactant, solvent, alkali, compound used, reaction temperature, reaction times etc. are not limited to the following example.

[0196] The compounds of the present invention can also be conveniently prepared by optionally combining various synthetic methods described in this specification or known in the art. Such a combination can be easily performed by those skilled in the art.

[0197] Example 1: 2-cyclopropyl-7,9-bis[4-(difluoromethoxy)phenyl]-8H-pyrimido[1,2-b]pyridazin-8-one

[0198] a) Preparation of N-(5-methoxypyridazin-3-yl)acetamide

[0199] To the reaction flask were added 3-chloro-5-methoxypyridazine (1 g), acetamide (0.61 g), trisdibenzylideneacetone dipalladium (0.32 g), 4,5-bisdiphenylphosphino-9,9-dimethylxanthene (0.40 g), and cesium carbonate (6.76 g) in sequence, followed by 1,4-dioxane (100 mL). The mixture was stirred at 100° C. for 3 hours under nitrogen protection and concentrated under reduced pressure to obtain the residue, which was evaporated to dryness. The residue was extracted with ethyl acetate (3×50 mL), and the combined organic phases were washed with saturated brine (1×50 mL), dried over anhydrous sodium sulfate, filtered, and the filtrate was concentrated under reduced pressure. The title compound (780 mg) was obtained.

[0200] LCMS m / z=168.05[M+1] + .

[0201] b) Preparation of 6-aminopyridazin-4(1H)-one

[0202] N-(5-methoxypyridazin-3-yl)acetamide (500 mg) and aqueous hydrogen bromide solution (15 mL) were added to a microwave tube, stirred at 140°C for 5 hours under microwave irradiation, and concentrated under reduced pressure to obtain 360 mg of the title compound.

[0203] LCMS m / z=111.95[M+1] + .

[0204] c) Preparation of 6-amino-3,5-dibromopyridazin-4(1H)-one

[0205] 6-Aminopyridazin-4(1H)-one (360 mg) and N-bromosuccinimide (1.73 g) were added to a reaction flask in N,N-dimethylformamide (4 mL) and stirred at room temperature for 2 hours. The residue was concentrated under reduced pressure and evaporated to dryness. The residue was washed with acetonitrile (3 x 5 mL) to obtain 360 mg of the title compound. d) Preparation of 6-amino-3,5-dibromo-1-[(1E)-3-cyclopropyl-3-oxoprop-1-en-1-yl]pyridazin-4(1H)-one

[0206] To a reaction flask, 6-amino-3,5-dibromopyridazin-4(1H)-one (430 mg), (2E)-3-chloro-1-cyclopropylprop-2-en-1-one (167.0 mg) and potassium carbonate (663.0 mg) were added and stirred at 25°C overnight in N,N-dimethylformamide (43 mL). The mixture was concentrated under reduced pressure to obtain a mixture. The pH of the mixture was adjusted to 2-3 with 1 M hydrochloric acid, and the mixture was washed with water (3 x 10 mL) to obtain 335 mg of the title compound.

[0207] e) Preparation of 7,9-dibromo-2-cyclopropyl-8H-pyrimido[1,2-b]pyridazin-8-one

[0208] To the reaction flask was added 6-amino-3,5-dibromo-1-[(1E)-3-cyclopropyl-3-oxoprop-1-en-1-yl]pyridazin-4(1H)-one (355 mg) and a 2 mol / L solution of hydrogen chloride in 1,4-dioxane (10 mL). The mixture was stirred at 25°C for 1 h. The resulting residue was concentrated under reduced pressure and evaporated to dryness. The residue was washed with aqueous sodium bicarbonate solution (1 x 10 mL). The filter cake was collected by filtration and washed with water (3 x 5 mL) to give 320 mg of the title compound.

[0209] f) Preparation of 2-cyclopropyl-7,9-bis[4-(difluoromethoxy)phenyl]-8H-pyrimido[1,2-b]pyridazin-8-one

[0210] To the reaction flask were added 7,9-dibromo-2-cyclopropyl-8H-pyrimido[1,2-b]pyridazin-8-one (60 mg), 4-(difluoromethoxy)phenylboronic acid (98.1 mg), water (0.4 mL), 1,4-dioxane (2 mL), potassium phosphate (184.6 mg) and [1,1'-bis(diphenylphosphino)ferrocene]dichloropalladium dichloromethane complex (14.17 mg). Under nitrogen protection, the mixture was stirred at 80°C for 1 hour, and the mixture was concentrated under reduced pressure to dryness. The title compound (37 mg) was obtained by preparative separation. Preparation and separation conditions: Chromatographic column: XBridge PrepOBD C18 column, 30*150 mm, 5 μm; column temperature: 25°C; mobile phase A: water (10 mmol / L NH4HCO3), mobile phase B: acetonitrile; flow rate: 60 mL / min; elution gradient: 45% B to 85% B from 0 to 10 min, and elution at 85% B after 10 min; detection wavelength: UV 220 nm; retention time (min): 7.32.

[0211] 1 H NMR(400MHz, DMSO-d6)δ8.72(d,J=7.2Hz,1H),8.27-8.21(m,2H),7.64-7.58(m,2H),7.56-7.26(m,3H),7.2 1-7.11(m,3H),7.07(d,J=7.2Hz,1H),2.20(tt,J=8.1,4.5Hz,1H),1.18-1.10(m,2H),1.01(p,J=3.8Hz,2H).

[0212] LCMS m / z=472[M+H] + .

[0213] Example 2: 7,9-bis(benzo[d][1,3]dioxol-5-yl)-2-cyclopropyl-8H-pyrimido[1,2-b]pyridazin-8-one

[0214] Prepared according to the preparation method of Example 1, replacing 4-(difluoromethoxy)phenylboronic acid in step f) with benzo[d][1,3]dioxol-5-ylboronic acid That's it.

[0215] 1H NMR (400MHz, DMSO-d6) δ8.69(d,J=7.2Hz,1H),7.88(dd,J=8.3,1.7Hz,1H),7.76(d,J=1.7Hz,1H),7.09-6.99(m,4H),6.93( d,J=8.0Hz,1H),6.11(s,2H),6.04(s,2H),2.18(td,J=8.0,4.1Hz,1H),1.12(dt,J=6.7,3.4Hz,2H),1.01(t,J=3.8Hz,2H).

[0216] LCMS m / z=428[M+1] + .

[0217] Example 3: 2-cyclopropyl-9-[4-(difluoromethoxy)phenyl]-7-(2-methyl-2H-indazol-5-yl)-8H-pyrimido[1,2-b]pyridazin-8-one

[0218] a) Preparation of 9-bromo-2-cyclopropyl-7-(2-methyl-2H-indazol-5-yl)-8H-pyrimido[1,2-b]pyridazin-8-one

[0219] To the reaction flask were added 7,9-dibromo-2-cyclopropyl-8H-pyrimido[1,2-b]pyridazin-8-one (50 mg), 2-methylindazol-5-ylboronic acid (28.06 mg), potassium carbonate (60.09 mg), [1,1'-bis(diphenylphosphino)ferrocene]dichloropalladium dichloromethane complex (11.81 mg), 1,4-dioxane (0.5 mL), and water (0.1 mL) in sequence. Under nitrogen protection, the mixture was stirred at 60°C for 1 h. The reactants were concentrated under reduced pressure and purified by silica gel column chromatography (mobile phase: ethyl acetate / petroleum ether = 4 / 1 (V / V)) to give 40 mg of the title compound.

[0220] 1 H NMR (400MHz, DMSO) δ8.98(s,1H),8.81(d,J=7.1Hz,1H),8.54(s,1H),7.95(d,J=8.9Hz,1H) ,7.66(d,J=9.1Hz,1H),7.14(d,J=7.1Hz,1H),4.20(s,3H),2.34(s,1H),1.46-1.10(m,4H).

[0221] LCMS m / z=396[M+1] + .

[0222] b) Preparation of 2-cyclopropyl-9-[4-(difluoromethoxy)phenyl]-7-(2-methyl-2H-indazol-5-yl)-8H-pyrimido[1,2-b]pyridazin-8-one

[0223] To the reaction flask were added 9-bromo-2-cyclopropyl-7-(2-methyl-2H-indazol-5-yl)-8H-pyrimido[1,2-b]pyridazin-8-one (35 mg), 4-(difluoromethoxy)phenylboronic acid (24.90 mg), potassium carbonate (36.62 mg), [1,1'-bis(diphenylphosphino)ferrocene]dichloropalladium dichloromethane complex (7.20 mg), 1,4-dioxane (1 mL), and water (0.2 mL) in sequence. Under nitrogen protection, the mixture was stirred at 80°C for 1 h. The reactants were concentrated under reduced pressure to obtain 14.4 mg of the title compound. Preparation and separation conditions: Chromatographic column: XBridge PrepOBD C18 column, 30*150 mm, 5 μm; column temperature: 25°C; mobile phase A: water (10 mmol / L NH4HCO3), mobile phase B: acetonitrile; flow rate: 60 mL / min; elution gradient: 25% B to 70% B at 0 to 10 min, and 85% B after 10 min; detection wavelength: UV 220 nm; retention time (min): 6.8.

[0224] 1 H NMR (400MHz, DMSO-d6)8.97(s,1H),8.76(d,J=7.2Hz,1H),8.52(s,1H),7.97(dd,J=9.2,1.6Hz,1H),7.68-7.59(m,3H),7.31(t,J=74.4Hz,1H ),7.22-7.16(m,2H),7.06(d,J=7.2Hz,1H),4.20(s,3H),2.19(dq,J=8.1,4.6,4.0Hz,1H),1.13(dd,J=7.8,3.5Hz,2H),1.02(t,J=3.8Hz,2H).

[0225] LCMS m / z=460[M+1] + .

[0226] Test Example 1: Biological Activity Test

[0227] 1. MAT2A Enzyme Test Method

[0228] 1. Experimental Procedure

[0229] a) First, prepare 5× MAT2A assay buffer (250 mM Tris-HCl, pH 8.0; 250 mM KCl; 75 mM MgCl2; 0.025% BSA; 0.05% Brij35; 1.5 mM EDTA) and partially dilute to 1× for later use;

[0230] b) Preparation and addition of MAT2A enzyme (BPS, 71401): MAT2A enzyme was prepared to 3.674 ng / μL (1.67×, final concentration 2.20 ng / μL) in 1× MAT2A assay buffer. Using a BioTek (MultiFlo FX) automated dispenser, 15 μL of 1.67× MAT2A enzyme solution was added to each of the compound test wells and negative control wells. Simultaneously, 15 μL of 1× MAT2A assay buffer was added to the blank control well.

[0231] c) Compound Preparation and Addition: Dilute the test compound from a 10 mM stock solution to 100 μM using DMSO. Dilute the positive drug AGI-24512 under the same conditions. Use a Tecan compound titrator (D300e) to automatically dispense the compound into each well according to a pre-set concentration gradient. Negligible amounts of the injected volume should be considered. The concentration gradient starts at 1 μM and is divided into 1 / 2 log dilutions, with a total of eight steps. Centrifuge at 2500 rpm for 30 seconds and incubate at 25°C for 30 minutes.

[0232] d) ATP preparation: dilute 10 mM ATP (Sigma, A7699) to 700 μM using 1× MAT2A assay buffer;

[0233] e) Preparation and addition of substrate and ATP mixture: 5× MAT2A assay buffer, 3 μL / well; 750 μM L-methionine (Adamas, 01100469), 2.5 μL / well; 700 μM ATP, 2.5 μL / well; double-distilled water, 2 μL / well. Prepare the required total volume of mixture based on the number of wells tested. Add 10 μL to each well using a BioTek (MultiFlo FX) automated dispenser. Centrifuge at 2500 rpm for 30 seconds and incubate at 25°C for 150 minutes.

[0234] f) Addition of Biomol Green detection reagent: 50 μL of Biomol Green (Enzo, BML-AK111) was added to each well using a BioTek (MultiFlo FX) automated dispenser. The cells were centrifuged at 2500 rpm for 30 seconds and incubated at 25°C for 20 minutes.

[0235] g) After the reaction, the OD was read using a Perkin Elmer (Envision 2105) multi-function plate reader. 620 value.

[0236] 2. Data Analysis

[0237] The inhibition rate calculation formula is as follows:

[0238] in,

[0239] OD sample: OD of the sample well 620 value;

[0240] ODmin: represents the OD of the blank control well without enzyme or test compound 620 mean;

[0241] ODmax: represents the OD of the negative control well with enzyme and no compound 620 Mean.

[0242] GraphPad Prism 5 software was then used to fit the dose-effect curve using log(inhibitor) vs. response-Variable slope to obtain the IC of the compound for MAT2A enzyme inhibition. 50 value.

[0243] 2. Cell Testing Methods

[0244] 1. Experimental Procedure

[0245] HCT116MTAP- / - cells (purchased from Horizon Discovery): A human colorectal cancer cell line lacking the MTAP gene, cultured in RPMI 1640 medium supplemented with 10% FBS (fetal bovine serum). On day 0, cells in logarithmic growth phase were adjusted to a viable cell density of 5000 cells / mL and seeded into 96-well plates at 100 μL / well. A blank control was also set up in parallel. The seeded plates were incubated overnight at 37°C in a 5% CO2 incubator.

[0246] On day 1 of the experiment, the cell plates were removed from the overnight cultures, the supernatant discarded, and 80 μL of serum-free RPMI 1640 medium was added to each well. The cells were then starved for 4 hours in an incubator. The test compound was dissolved in DMSO (dimethyl sulfoxide) to prepare a 10 mM stock solution. After the starvation period, the cell plates were removed and 80 μL of RPMI 1640 + 20% FBS medium was added to each well. The cell plates were placed on a D300e automated dosing system (Tecan) and the dosing program was set to: test compound at a maximum concentration of 30 μM, with a three-fold serial dilution using DMSO for a total of 10 concentrations, with two replicates per well. The final DMSO concentration in each well of the 96-well plate was 0.3% (v / v). The pre-prepared 10 mM stock solution of the test compound was removed and the dosing protocol described above was repeated. After dosing, the cell plates were incubated in an incubator for 120 hours.

[0247] On the 6th day of the experiment, the cell plate was removed and 50 μL was added to each well. (purchased from Promega), and the fluorescence signal was measured on Envision (PerkinElmer) according to the operating procedures of the instruction manual.

[0248] 2. Data Analysis

[0249] GraphPad Prism 5 software was used to fit the dose-effect curve: log (inhibitor) vs. response-Variable slope to obtain the IC value of the compound for cell proliferation inhibition. 50 The inhibition rate calculation formula is:

[0250] in:

[0251] Test substance signal value: mean fluorescence signal of cell+medium+compound group;

[0252] Signal value of blank group: mean fluorescence signal of culture medium group;

[0253] Signal value of negative control group: mean fluorescence signal of cells+medium group.

[0254] 3. Experimental results:

[0255] The IC of AGI-24512 for inhibiting MAT2A was determined according to the above experimental method. 50 It is 26.79nM.

[0256] AGI-24512 structure:

[0257] The experimental results of the compounds of the present invention are shown in Table 10 below:

[0258] Table 10

[0259] Test Example 2: In vivo pharmacokinetic study in ICR mice

[0260] 1. Experimental Procedure

[0261] Male ICR mice (6-10 weeks old, Weitong Lihua Laboratory Animal Technology Co., Ltd.) were housed in a SPF animal room at a temperature of 20-25°C, a relative humidity of 40%-70%, and a 12-hour light and dark cycle. The animals had free access to water and food. After at least 5 days of normal feeding and good physical condition after veterinary examination, mice were enrolled in this experiment. Each mouse was numbered at the tail.

[0262] The day before the experiment, mice were fasted overnight with free access to water. Four hours after dosing, the compounds were prepared into 20 mg / mL stock solutions in DMSO. An appropriate volume of the 20 mg / mL stock solution was accurately pipetted into a glass vial. An appropriate volume of PEG400 was added, mixed, and then propylene glycol (PG) was added. The final formulation had a solvent ratio of DMSO:PEG400:PG (v / v / v) of 5:65:30, resulting in a dosing solution of each test compound at a concentration of 1 mg / mL.

[0263] After weighing the mice, calculate the theoretical dosing volume for each mouse using the following formula. The actual dosing volume for each mouse and the time of blood sample collection should be recorded in detail in the corresponding table.

[0264] On the day of the experiment, mice in each group were gavage-administered with a 10 mg / kg dose of the test compound. At various time points after dosing, approximately 40 μL of blood was collected from the mice via orbital orbital examination and placed in EDTA-K2 anticoagulant tubes. Whole blood samples were centrifuged at 1500–1600 g for 10 minutes, and the resulting plasma was stored at -40–-20°C for biological sample analysis.

[0265] 2. Data Analysis

[0266] The concentrations of the compounds in the biological samples were determined by LC-MS / MS analysis, and the pharmacokinetic parameters were calculated using a non-compartmental model in Pharsight Phoenix 7.0.

[0267] 3. Experimental Results

[0268] The pharmacokinetic parameter calculation results are shown in Table 11 below.

[0269] Table 11

[0270] Test Example 3: HCT116MTAP - / - In vivo drug efficacy experiment on subcutaneous xenograft tumors in nude mice

[0271] 1. Experimental Procedure

[0272] Female Nu / Nu nude mice (6-8 weeks old, Beijing Weitonglihua Laboratory Animal Technology Co., Ltd.) were housed in an SPF animal room at a temperature of 20-25°C, a relative humidity of 40%-70%, and a 12-h light and dark cycle. The animals had free access to water and food. Animals were acclimated to the environment before the experiment.

[0273] HCT116MTAP - / - Cells (Horizon) were cultured and expanded in vitro. Cells in the logarithmic growth phase were collected and resuspended in serum-free RPMI-1640 medium. The cell concentration was adjusted to 6.0×10 7 cells / mL; use a 1mL syringe to inject the cell suspension into the subcutaneous tissue of the right axilla of nude mice, inject 100μL into each animal, and observe the animal status and monitor the growth of the transplanted tumor regularly.

[0274] When the tumor volume reaches 100-300 mm 3 Animals with tumors that were excessively large, too small, or uncertain in size were eliminated. Tumor-bearing mice in good health with tumors of similar size were selected and grouped using a randomized block method. The treatment group received daily oral administration (AG-270: 50 mg / kg, structure shown below; test compound: 5 mg / kg), while the control group received the same volume of blank vehicle daily. Tumor diameters were measured twice weekly during the treatment period, and tumor volumes were calculated. Animal weights were also recorded.

[0275] SAM detection in transplanted tumors: At the end of the experiment, animals were euthanized using CO2. Tumor tissue was removed, washed with cold PBS, weighed, snap-frozen in liquid nitrogen, and stored at -80°C until use. The frozen tumor tissue was removed, thawed in an ice bath, and then homogenized with 80% methanol-water solution (containing 1 M formic acid) at a ratio of 1:10 (w / v). The homogenate was collected and processed for SAM (S-adenosylmethionine) analysis using LC-MS / MS.

[0276] AG-270 structure:

[0277] 2. Data Analysis

[0278] The calculation formula for tumor volume (TV) is: TV = 1 / 2 × a × b 2 ; Where a represents the long diameter of the tumor and b represents the short diameter of the tumor.

[0279] The formula for calculating relative tumor volume (RTV) is: RTV = TV t / TV initial Among them, TV initial is the tumor volume measured when the group was administered, TV t The tumor volume was measured at each time during the drug administration period.

[0280] The calculation formula of relative tumor proliferation rate (T / C (%)) is: T / C% = (RTV T / RTV C )×100%; among which, RTV T Relative tumor volume of the treatment group, RTV C Relative tumor volume to that of the vehicle control group is shown.

[0281] The calculation formula of tumor growth inhibition (TGI (%)) is: TGI = [1-(TV t(T) -TV initial(T) ) / (TV t(C) -TV initial(C) )]×100%; among which, TV t(T) represents the tumor volume of each treatment group, TV initial(T) Indicates the tumor volume of the treatment group at the time of drug administration, TV t(C) represents the tumor volume of the solvent control group at each measurement, TV initial(C) It represents the tumor volume of the solvent control group at the time of group administration.

[0282] The formula for calculating the animal weight loss rate is: Animal weight loss rate = 100% × (BW initial -BW t ) / BW initial Among them, BW t Indicates the animal body weight measured each time during the dosing period, BW initial Indicates the body weight of animals at the time of grouping and dosing.

[0283] The calculation formula of tumor weight inhibition rate IR (%) is: IR = 100% × (W C -W T ) / W C Among them, W C represents the tumor weight of the control group, W T Indicates tumor weight in treatment group.

[0284] The experimental data were calculated and statistically processed using Microsoft Office Excel 2007. Unless otherwise specified, the data were expressed as mean ± standard error (mean ± SE), and the t-test was used for comparison between the two groups.

[0285] 3. Experimental Results

[0286] HCT116MTAP - / - The tumor inhibition curve of the transplanted tumor model and the results of intratumoral SAM inhibition are shown in Figures 1 and 2, respectively.

[0287] Test Example 4: In vivo efficacy experiment on KP-4 subcutaneous xenograft tumors in mice

[0288] 1. Experimental Procedure

[0289] Female NOD SCID mice (6-8 weeks old, Beijing Weitonglihua Laboratory Animal Technology Co., Ltd.) were housed in a SPF animal room at a temperature of 20-25°C, a relative humidity of 40%-70%, and a 12-hour light and dark cycle. The animals had free access to water and food. Animals were acclimated to the feeding regimen before the experiment.

[0290] KP-4 cells (Nanjing Kebai Biotechnology Co., Ltd.) were cultured and expanded in vitro. Cells in the logarithmic growth phase were collected and resuspended in serum-free RPMI-1640 medium. The cell concentration was adjusted to 1.0 × 10 8 cells / mL; use a 1mL syringe to inject the cell suspension into the subcutaneous tissue of the right axilla of nude mice, inject 100μL into each animal, and observe the animal status and monitor the growth of the transplanted tumor regularly.

[0291] When the tumor volume reaches 80-100 mm 3 Animals with tumors that were excessively large, small, or of uncertain size were eliminated. Tumor-bearing mice in good health with tumors of similar size were selected and grouped using a randomized block method. The treatment group received daily oral administration (AG-270: 100 mg / kg, test compound: 2.5 mg / kg), while the control group received the same volume of blank vehicle daily. Tumor diameters were measured twice weekly during the treatment period, and tumor volumes were calculated. Animal weights were also recorded.

[0292] SAM detection in transplanted tumors: At the end of the experiment, animals were euthanized using CO2. Tumor tissue was removed, washed with cold PBS, weighed, snap-frozen in liquid nitrogen, and stored at -80°C until use. The frozen tumor tissue was removed, thawed in an ice bath, and then homogenized with 80% methanol-water solution (containing 1 M formic acid) at a ratio of 1:10 (w / v). The homogenate was collected and processed for SAM (S-adenosylmethionine) analysis using LC-MS / MS.

[0293] 2. Data Analysis

[0294] The calculation formula for tumor volume (TV) is: TV = 1 / 2 × a × b 2 ; Where a represents the long diameter of the tumor and b represents the short diameter of the tumor.

[0295] The formula for calculating relative tumor volume (RTV) is: RTV = TV t / TV initial ; Among them, TV initial is the tumor volume measured when the group was administered, TV t The tumor volume was measured at each time during the drug administration period.

[0296] The calculation formula of relative tumor proliferation rate (T / C (%)) is: T / C% = (RTV T / RTV C )×100%; among which, RTV T Relative tumor volume of the treatment group, RTV C Relative tumor volume to that of the vehicle control group is shown.

[0297] The calculation formula of tumor growth inhibition (TGI (%)) is: TGI = [1-(TV t(T) -TV initial(T) ) / (TV t(C) -TV initial(C) )]×100%; among which, TV t(T) represents the tumor volume of each treatment group, TV initial(T) Indicates the tumor volume of the treatment group at the time of drug administration, TV t(C) represents the tumor volume of the solvent control group at each measurement, TV initial(C) It represents the tumor volume of the solvent control group at the time of group administration.

[0298] The formula for calculating the animal weight loss rate is: Animal weight loss rate = 100% × (BWinitial -BW t ) / BW initial Among them, BW t Indicates the animal body weight measured each time during the dosing period, BW initial Indicates the body weight of animals at the time of grouping and dosing.

[0299] The calculation formula of tumor weight inhibition rate IR (%) is: IR = 100% × (W C -W T ) / W C Among them, W C represents the tumor weight of the control group, W T Indicates tumor weight in treatment group.

[0300] The experimental data were calculated and statistically processed using Microsoft Office Excel 2007. Unless otherwise specified, the data were expressed as mean ± standard error (mean ± SE), and the t-test was used for comparison between the two groups.

[0301] 3. Experimental Results

[0302] The tumor inhibition curve of the KP-4 transplanted tumor model and the intratumoral SAM inhibition results are shown in Figures 3 and 4, respectively.

[0303] Test Example 5: Polymorph Study

[0304] It is well known to those skilled in the art that when the above-mentioned compounds are shown to have a good MAT2A inhibitory effect, their crystal forms often have the same pharmacological and pharmacodynamic activity. Based on this, the inventors further studied the physicochemical properties of the corresponding crystal forms of the compounds. However, the preparation and characterization of the following specific crystal forms do not limit the scope of protection of the present invention. Based on this invention, those skilled in the art can obtain more crystals of the compounds of the present invention, and these crystals are all protected by the present invention. The details are as follows:

[0305] 1. Instrument test information

[0306] Table 12. X-ray powder diffractometer parameters

[0307] Table 13. TGA and DSC instrument test parameters

[0308] Table 14. DVS instrument test parameters

[0309] 2. Preparation of Buffer

[0310] 2.1 pH 1.0 hydrochloric acid solution: Take 9.00mL of the specified amount of hydrochloric acid, dilute it to 1000mL with water, and shake well.

[0311] 2.2 Phosphate buffer:

[0312] 0.2 mol / L potassium dihydrogen phosphate solution: Take 27.22 g of potassium dihydrogen phosphate, dissolve it in water and dilute to 1000 mL.

[0313] 0.2 mol / L sodium hydroxide solution: Take 8.00 g of sodium hydroxide, dissolve it in water and dilute to 1000 mL.

[0314] Take 250mL of 0.2mol / L potassium dihydrogen phosphate solution and mix it with the amount of 0.2mol / L sodium hydroxide solution specified in the table below, then add water to dilute to 1000mL and shake well to obtain the product.

[0315] Table 15 Phosphate buffer

[0316] 3. Research on salt and crystal form

[0317] Preparation of salts and crystalline forms of 2-cyclopropyl-9-[4-(difluoromethoxy)phenyl]-7-(2-methyl-2H-indazol-5-yl)-8H-pyrimido[1,2-b]pyridazin-8-one.

[0318] 3.1 Salt screening

[0319] About 10-20 mg of free base and an equal molar amount of ligand acid were stirred in 0.5 mL of solvent at room temperature for a period of time. The system with solid residue was centrifuged and the solid was vacuum dried at 50°C and characterized by XRD.

[0320] Table 16 Compound salt formation results

[0321] The results showed that hydrochloride, sulfate, phosphate, maleic acid, citrate, adipate, methanesulfonate and oxalate crystal forms were obtained.

[0322] 3.2 Crystal preparation

[0323] 3.2.1 Preparation of Form A

[0324] Preparation of 2-cyclopropyl-9-[4-(difluoromethoxy)phenyl]-7-(2-methyl-2H-indazol-5-yl)-8H-pyrimido[1,2-b]pyridazin-8-one: 9-bromo-2-cyclopropyl-7-(2-methyl-2H-indazol-5-yl)-8H-pyrimido[1,2-b]pyridazin-8-one (3.8 g), 4-(difluoromethoxy)phenylboronic acid (3.6 g), potassium carbonate (3.98 g), [1,1'-bis(diphenylphosphine)] Ferrocene] palladium dichloride dichloromethane complex (0.78 g), 1,4-dioxane (40 mL), and water (8 mL) were stirred at 80°C under nitrogen for 2 hours. The reactant was concentrated under reduced pressure and purified by column chromatography (DCM:EA = 0% to 100%) to obtain a crude product (1.5 g). The crude product was slurried with DMF / MeCN (5 mL:5 mL), filtered, and the filter cake was washed twice with MeCN (1 mL) and dried in vacuo at 70°C to obtain Form A (535.1 mg). Analysis and testing revealed XRD results as shown in Figure 5; TGA results as shown in Figure 6 showed no significant weight loss when the sample was heated to 120°C; and DSC results as shown in Figure 7 showed a sharp endothermic peak at 262.42°C (onset temperature). Comprehensive characterization results indicated that Form A was an anhydrous crystalline form.

[0325] 3.2.2 Preparation of Form B

[0326] A 20 mg sample of Form G was slurried in 0.6 mL of methanol (or ethyl acetate) at room temperature for 3 days, dried under vacuum at 50°C for 4 hours, and collected to yield Form B. Analysis and testing revealed XRD results, as shown in Figure 8; TGA results, as shown in Figure 9, showed no significant weight loss upon heating to 150°C; and DSC results, as shown in Figure 10, revealed two endothermic peaks at 219.76°C and 266.19°C (onset temperature). Comprehensive characterization results indicated that Form B was an anhydrous crystalline form.

[0327] 3.2.3 Preparation of Form G

[0328] 9-Bromo-2-cyclopropyl-7-(2-methyl-2H-indazol-5-yl)-8H-pyrimido[1,2-b]pyridazin-8-one (24 g), 4-(difluoromethoxy)phenylboronic acid (19.17 g), potassium carbonate (24.96 g), [1,1'-bis(diphenylphosphino)ferrocene]dichloropalladium dichloromethane complex (4.32 g), 1,4-dioxane (480 mL), and water (96 mL) were added to the reaction flask. Under nitrogen protection, the reaction was stirred at 72°C for 2 hours. The reactants were concentrated under reduced pressure at 50°C to obtain a black solid. 400 mL of water was added and stirred for 40 minutes. The mixture was filtered to obtain a black filter cake. The filter cake was mixed with 400 mL of anhydrous ethanol, heated to 72°C, and filtered while hot to obtain a filter cake. The filter cake was mixed with 450 mL of ethanol. The mixture was stirred with DMSO for 40 minutes, filtered, and the filtrate collected. 300 mL of water was added to the filtrate, filtered, and the filter cake collected. The filter cake was mixed with 150 mL of DMSO, heated to 82°C, stirred at this temperature for 50 minutes, cooled to room temperature, filtered, and the filter cake collected. The filter cake was mixed with 300 mL of anhydrous ethanol, stirred at room temperature for 1 hour, and filtered to collect the product. Analysis and testing revealed XRD results as shown in Figure 11; TGA results as shown in Figure 12, showing a weight loss of 0.71% upon heating to 150°C; and DSC results as shown in Figure 13. Comprehensive characterization results indicated that Form G was an anhydrous crystalline form.

[0329] 3.3 Salt and crystal preparation

[0330] 3.3.1 Hydrochloride

[0331] 20 mg of Form B and an equimolar amount of hydrochloric acid (3.63 μL concentrated hydrochloric acid) were slurried in 0.5 mL of 2-butanone at room temperature for 3 days, vacuum dried at 40°C for 3.5 hours, and the product was collected to obtain Form A hydrochloride. After detection and analysis, the XRD results are shown in Figure 14. 1 The H NMR results are shown in 15.

[0332] 3.3.2 Sulfate

[0333] 5 mL of ethanol was added to a 20-mL glass vial, and 24.2 μL (1.0 equivalent) of concentrated sulfuric acid was added and mixed evenly. 200 mg of free Form G was added to the above solution, stirred at room temperature overnight, filtered, and the filter cake was dried under reduced pressure at 40°C for 4 hours to collect 212.5 mg of the product, which was the sulfate Form A. After testing and analysis, the XRD results are shown in Figure 16. 1 The H NMR results are shown in Figure 17. Comprehensive characterization results showed that sulfate crystal form A was anhydrous crystalline form.

[0334] 3.3.3 Phosphate

[0335] 300 mg of Form G sample and 0.34 equivalents of sulfuric acid (14.9 μL, 85% phosphoric acid) were slurried in 7.5 mL of ethanol at room temperature for 2 days, filtered, and the filter cake was vacuum dried at 40°C for 4 hours to collect 281.7 mg of product, obtaining Form A phosphate. The XRD results are shown in Figure 18.

[0336] 3.3.4 Maleate

[0337] 20 mg of Form B and 5.1 mg of maleic acid were slurried in 0.5 mL of chloroform at room temperature for 3 days, dried under vacuum at 40°C for 3.5 hours, and the product was collected to obtain Form A maleate. After testing and analysis, the XRD results are shown in Figure 19; 1 H NMR results ( FIG. 20 ) showed that the molar ratio of the compound to maleic acid in the maleate salt Form A sample was 1:1.0.

[0338] 3.3.5 Citrate

[0339] 20 mg of Form B and 8.5 mg of anhydrous citric acid were slurried in 0.5 mL of chloroform at room temperature for 3 days, vacuum dried at 40°C for 3.5 hours, and the product was collected to obtain citrate Form A. After detection and analysis, the XRD results are shown in Figure 21. 1 The H NMR results are shown in FIG22 .

[0340] 3.3.6 Adipate salt form A

[0341] 10 mg of Form A and 3.2 mg of adipic acid were slurried in 0.5 mL of acetone / water (3:1, v:v) at room temperature for 3 days, dried under vacuum at 50°C for 4 hours, and the product was collected to obtain Form A adipate. The XRD results are shown in Figure 23.

[0342] 3.3.7 Adipate salt form B

[0343] 300 mg of compound Form G and 95.7 mg of adipic acid were added to a 20-mL glass vial; 8.5 mL of 2-butanone was added and stirred at room temperature for 2 days; the mixture was filtered; the filter cake was dried under reduced pressure at 40°C for 4 hours, and 332.8 mg of the product was collected to obtain Form B of the adipate salt. The XRD results are shown in Figure 24. 1 H NMR results showed that the molar ratio of the compound to adipic acid in the adipate salt form B sample was 1:0.5. Comprehensive characterization results showed that the adipate salt form B was an anhydrous crystalline form.

[0344] 3.3.8 Methanesulfonate

[0345] 300 mg of compound Form G was added to a 20-mL glass bottle; 7.5 mL of 2-butanone was added; 63.4 mg of methanesulfonic acid was added to the suspension, stirred at room temperature for 2 days, filtered, and dried under reduced pressure at 40°C for 4 hours to collect 340 mg of the product to obtain Form A of the methanesulfonate salt. The XRD results are shown in Figure 26. 1 H NMR results showed that the molar ratio of the compound to methanesulfonic acid in the mesylate Form A sample was 1:1.0. Comprehensive characterization results showed that the mesylate Form A was an anhydrous crystalline form.

[0346] 3.3.9 Oxalate

[0347] Form B and 5.6 mg of oxalic acid dihydrate were slurried in 0.5 mL of 2-butanone (or chloroform) at room temperature for 3 days and vacuum dried at 40°C for 3.5 hours to obtain oxalate form A. After detection and analysis, the XRD results are shown in Figure 28. 1 The H NMR results are shown in FIG29 .

[0348] 4. Solubility Experiment

[0349] The solubility of the sample was measured at a 10 mg / mL feed concentration at 37°C in four solvent systems: pH 1.0, pH 4.5, and pH 6.8 buffers, and water, for 1, 4, and 24 hours using a shaking system (400 rpm). Samples were centrifuged (10,000 rpm, 2 minutes) and filtered (through a 0.22 μm PTFE filter, discarding the pre-liquid). The filtrate concentration was determined by HPLC, and the remaining solid was analyzed by XRD. The solubility test results are summarized in Table 17.

[0350] Table 17 Summary of dynamic solubility test results

[0351] The results showed that the solubility of sulfate crystal form A in pH 1.0 buffer for 1 hour and pH 6.8 buffer for 24 hours was higher than that of free base, and the solubility of adipate crystal form B in pH 4.5 buffer for 4 hours was higher than that of free base.

[0352] 5. Stability test

[0353] The sulfate salt Form A, adipate salt Form B, and methanesulfonate salt Form A samples were stored at 25°C / 60% RH for one week, and then the physicochemical stability of the samples was tested using XRD and HPLC. The test data are listed in Table 18.

[0354] Table 18 Stability study results

[0355] The results showed that the crystal form and purity of the three salts did not change significantly after being placed under 25℃ / 60%RH and 40℃ / 75%RH conditions for one week.

[0356] 6.PK experiments

[0357] Adipate salt form B, mesylate salt form A, and form B were selected for a comparative PK study. The samples were uniformly dispersed in a mixed solvent (prepared as: 6.7% w / w hydroxypropyl methylcellulose acetate succinate-MF, 1% w / w povidone K30, 2% w / w d-α tocopheryl succinate polyethylene glycol ester, 0.1% w / w dimethicone) and administered by gavage. The study was conducted in male BALB / c mice. The dose of each sample was 10 mg / kg (the amount of the compound was converted to the same amount of the free base). Orbital blood was collected at 0.25, 0.5, 2, 4, 8, 24, and 48 hours after administration and detected by LCMS. Each group was studied in triplicate. The results are shown in the table below.

[0358] Table 19 Instruments and test conditions

[0359] Table 20 Results of pharmacokinetic parameters study in mice

[0360] According to the research results, compared with Form B, the exposure of Form B of the adipate salt increased by 12 times, and the exposure of Form A of the methanesulfonate salt increased by 7 times; the absorption rate of the two salt forms was also significantly faster than that of the free base.

Claims

1. An acid salt of a compound, characterized in that The specific structure of the compound is as follows: The acid in the acid salt is an inorganic acid or an organic acid; the inorganic acid includes hydrochloric acid, sulfuric acid, nitric acid, hydrobromic acid, hydrofluoric acid, hydroiodic acid or phosphoric acid; the organic acid includes formic acid, acetic acid, propionic acid, dichloroacetic acid, trichloroacetic acid, acetohydroxamic acid, adipic acid, benzenesulfonic acid, benzoic acid, phthalic acid, p-hydroxybenzoic acid, capric acid, caproic acid, caprylic acid, cinnamic acid, citric acid, succinic acid, cyclohexanesulfamic acid, camphorsulfonic acid, Aspartic acid, camphoric acid, D-gluconic acid, glucuronic acid, L-glutamic acid, L-ascorbic acid, lactic acid, mandelic acid, pyroglutamic acid, L-tartaric acid, dodecyl sulfate, dibenzoyltartaric acid, ethanesulfonic acid, formic acid, fumaric acid, galactosyl acid, gentisic acid, glutaric acid, glycolic acid, hippuric acid, isethionic acid, lactobionic acid, ascorbic acid, aspartic acid, lauric acid, camphoric acid, maleic acid, malonic acid, methanesulfonic acid, nicotinic acid, stearic acid, oleic acid, orotic acid, oxalic acid, palmitic acid, pamoic acid, propionic acid, salicylic acid, sebacic acid, succinic acid, thiocyanic acid, undecylenic acid, trifluoroacetic acid, benzenesulfonic acid, p-toluenesulfonic acid or L-malic acid; preferably, the acid in the acid salt is an inorganic acid or an organic acid; the inorganic acid is selected from hydrochloric acid, sulfuric acid or phosphoric acid; the organic acid is selected from maleic acid, acetic acid, citric acid, succinic acid, adipic acid, L-tartaric acid, L-ascorbic acid, hippuric acid, gentisic acid, malonic acid, methanesulfonic acid, benzoic acid, p-toluenesulfonic acid, phthalic acid, p-hydroxybenzoic acid, propionic acid, oxalic acid, L-glutamic acid, stearic acid or D-gluconic acid; more preferably, the acid in the acid salt is an inorganic acid or an organic acid; the inorganic acid is selected from hydrochloric acid, sulfuric acid or phosphoric acid; the organic acid is selected from maleic acid, citric acid, adipic acid, methanesulfonic acid or oxalic acid.

2. The acid salt according to claim 1, characterized in that The compound is 2-cyclopropyl-9-[4-(difluoromethoxy)phenyl]-7-(2-methyl-2H-indazol-5-yl)-8H-pyrimido[1,2-b]pyridazin-8-one, and its specific structural formula is as follows:

3. A crystalline acid salt of a compound, characterized in that: The compound is 2-cyclopropyl-9-[4-(difluoromethoxy)phenyl]-7-(2-methyl-2H-indazol-5-yl)-8H-pyrimido[1,2-b]pyridazin-8-one, and the specific structural formula is as follows: The acid in the acid salt is an inorganic acid or an organic acid; the inorganic acid is selected from hydrochloric acid, sulfuric acid or phosphoric acid; the organic acid is selected from maleic acid, acetic acid, citric acid, succinic acid, adipic acid, L-tartaric acid, L-ascorbic acid, hippuric acid, gentisic acid, malonic acid, methanesulfonic acid, benzoic acid, p-toluenesulfonic acid, phthalic acid, p-hydroxybenzoic acid, propionic acid, oxalic acid, L-glutamic acid, stearic acid or D-gluconic acid; preferably, the acid in the acid salt is an inorganic acid or an organic acid; the inorganic acid is selected from hydrochloric acid, sulfuric acid or phosphoric acid; the organic acid is selected from maleic acid, citric acid, adipic acid, methanesulfonic acid or oxalic acid; more preferably, the acid salt crystal form is selected from hydrochloride crystal form, sulfate crystal form, phosphate crystal form, maleic acid crystal form, citric acid crystal form, adipate crystal form, methanesulfonate crystal form or oxalate crystal form.

4. The acid salt crystalline form according to claim 3, characterized in that: The acid salt crystalline form is the hydrochloride crystalline form A of the compound 2-cyclopropyl-9-[4-(difluoromethoxy)phenyl]-7-(2-methyl-2H-indazol-5-yl)-8H-pyrimido[1,2-b]pyridazin-8-one, and its X-ray powder diffraction pattern at 2θ is 11.5°±0.2°, 17.0°±0.2°, 17.8°±0.2°, 22.4°±0.2°, 23.3°±0.2° and 27.1°±0.2° Preferably, the hydrochloride salt form A has an X-ray powder diffraction pattern at 2θ of 6.3°±0.2°, 11.5°±0.2°, 17.0°±0.2°, 17.8°±0.2°, 22.4°±0.2°, 23.3°±0.2°, 24.2°±0.2° and 27.1°±0.2°; More preferably, the hydrochloride salt form A has an X-ray powder diffraction pattern at 2θ of 4.1°±0.2°. The hydrochloride salt form A has diffraction peaks at 2θ of 4.1°±0.2°, 6.3°±0.2°, 11.5°±0.2°, 17.0°±0.2°, 17.8°±0.2°, 22.4°±0.2°, 22.8°±0.2°, 23.3°±0.2°, 24.2°±0.2° and 27.1°±0.2°; further preferably, the hydrochloride salt form A has an X-ray powder diffraction pattern at 2θ of 4.1°±0.2°, 6.3°±0.2°, 8.3°±0.2°, 11.5°±0.2°, 17.0°±0.2°, 17.8°±0.2°, 22.4°±0.2°, 22.8°±0.2°, 23.3°±0.2°, 24.2°±0.2°, 27.1°±0.2° and 27.5°±0.2° have diffraction peaks; further preferably, the X-ray powder diffraction of the hydrochloride salt form A expressed in 2θ angles has a spectrum as shown in Figure 14; Preferably, the hydrochloride crystal form A has the 1 H NMR spectrum; Or the acid salt crystalline form is the sulfate crystalline form A of the compound 2-cyclopropyl-9-[4-(difluoromethoxy)phenyl]-7-(2-methyl-2H-indazol-5-yl)-8H-pyrimido[1,2-b]pyridazin-8-one, and its X-ray powder diffraction pattern has diffraction peaks at 2θ of 3.4°±0.2°, 15.6°±0.2°, 17.1°±0.2°, 20.6°±0.2°, 23.4°±0.2° and 24.1°±0.2°; preferably, the sulfate crystalline form A, its X-ray powder diffraction pattern has diffraction peaks at 2θ of 3.4°±0.2°, 15.6°±0.2°, 17.1°±0.2°, 20.3°±0.2°, 20.6°±0.2°, 23.4°±0.2°, 24.1°±0.2° and 24.5°±0.2° have diffraction peaks; more preferably, the sulfate crystalline form A has an X-ray powder diffraction pattern with 2θ of 3.4°±0.2°, 6.8°±0.2°, 15.6°±0.2°, 17.1°±0.2°, 20.3°±0.2°, 20.6°±0.2°, 23.4°±0.2°, 24.1°±0.2°, 24.5°±0.2° and 26.6°±0.2° having diffraction peaks; further preferably, the sulfate crystalline form A has an X-ray powder diffraction pattern with 2θ of 3.4°±0.2°, 6.8°±0.2°, 15.6°±0.2°, 17.1°±0.2°, 20.3°±0.2°, 20.6°±0.2°, 23.4°±0.2°, 24.1°±0.2°, 24.5°±0.2° and 26.6°±0.2° having diffraction peaks; further preferably, the sulfate crystalline form A, whose X-ray powder diffraction pattern has diffraction peaks at 2θ of 3.4°±0.2°, 6.8°±0.2°, 15.6°±0.2°, 17.1°±0.2°, 20.3°±0.2°, 20.6°±0.2°, 23.4°±0.2°, 24.1°±0.2°, 24.5°±0.2°, 26.6°±0.2°, 27.7°±0.2° and 28.0°±0.2°; further preferably, the X-ray powder diffraction of the sulfate crystalline form A expressed in 2θ angles has a spectrum as shown in Figure 16; Preferably, the sulfate crystal form A has a hydrogen spectrum as shown in FIG17 ; Or the acid salt crystalline form is the phosphate crystalline form A of the compound 2-cyclopropyl-9-[4-(difluoromethoxy)phenyl]-7-(2-methyl-2H-indazol-5-yl)-8H-pyrimido[1,2-b]pyridazin-8-one, and its X-ray powder diffraction pattern is at 2θ of 12.2°±0.2°, 16.1°±0.2°, 19.9°±0.2°, 22.4°±0.2°, 23.2°±0.2° and 28.7°±0.2° Preferably, the phosphate crystal form A has an X-ray powder diffraction pattern with diffraction peaks at 2θ of 12.2°±0.2°, 16.1°±0.2°, 19.4°±0.2°, 19.9°±0.2°, 21.5°±0.2°, 22.4°±0.2°, 23.2°±0.2° and 28.7°±0.2°; More preferably, the phosphate crystal form A has an X-ray powder diffraction pattern with diffraction peaks at 2θ of 12.2°±0 .2°, 16.1°±0.2°, 18.1°±0.2°, 19.4°±0.2°, 19.9°±0.2°, 21.5°±0.2°, 22.4°±0.2°, 23.2°±0.2°, 25.9°±0.2° and 28.7°±0.2°; further preferably, the phosphate crystal form A has an X-ray powder diffraction pattern at 2θ of 12.2°±0.2°, 14.9°±0.2°, and 28.7°±0.2°. , 16.1°±0.2°, 18.1°±0.2°, 19.4°±0.2°, 19.9°±0.2°, 20.4°±0.2°, 21.5°±0.2°, 22.4°±0.2°, 23.2°±0.2°, 25.9°±0.2° and 28.7°±0.2° have diffraction peaks; further preferably, the X-ray powder diffraction of the phosphate crystal form A expressed in 2θ angles has a spectrum as shown in Figure 18; Or the acid salt crystalline form is the maleate crystalline form A of the compound 2-cyclopropyl-9-[4-(difluoromethoxy)phenyl]-7-(2-methyl-2H-indazol-5-yl)-8H-pyrimido[1,2-b]pyridazin-8-one, whose X-ray powder diffraction pattern has 2θ of 6.7°±0.2°, 7.3°±0.2°, 14.7°±0.2°, 16.7°±0.2°, 20.2°±0.2° and 22.0°±0.2° Preferably, the maleate salt form A has an X-ray powder diffraction pattern with diffraction peaks at 2θ of 6.7°±0.2°, 7.3°±0.2°, 13.4°±0.2°, 14.7°±0.2°, 16.7°±0.2°, 20.2°±0.2°, 22.0°±0.2° and 26.9°±0.2°; More preferably, the maleate salt form A has an X-ray powder diffraction pattern with diffraction peaks at 2θ of 6.7°±0.2°, 7.3°±0.2°, 13.4°±0.2°, 14.7°±0.2°, 16.7°±0.2°, 20.2°±0.2°, 22.0°±0.2° and 26.9°±0.2°. .2°, 7.3°±0.2°, 13.4°±0.2°, 14.7°±0.2°, 16.7°±0.2°, 18.7°±0.2°, 20.2°±0.2°, 22.0°±0.2°, 24.6°±0.2° and 26.9°±0.2°; further preferably, the maleate salt form A has an X-ray powder diffraction pattern at 2θ of 6.7°±0.2°, 7.3°±0.2°, 13.4°±0.2°, 14.7°±0.2°, 16.7°±0.2°, 18.7°±0.2°, 20.2°±0.2°, 22.0°±0.2°, 24.6°±0.2° and 26.9°±0.2°. 8.0°±0.2°, 13.4°±0.2°, 14.7°±0.2°, 16.7°±0.2°, 18.7°±0.2°, 20.2°±0.2°, 22.0°±0.2°, 24.1°±0.2°, 24.6°±0.2° and 26.9°±0.2° have diffraction peaks; further preferably, the X-ray powder diffraction of the maleate salt form A expressed in 2θ angles has a spectrum as shown in Figure 19; Preferably, the maleate salt form A has a hydrogen spectrum as shown in Figure 20; Or the acid salt crystalline form is the citrate crystalline form A of the compound 2-cyclopropyl-9-[4-(difluoromethoxy)phenyl]-7-(2-methyl-2H-indazol-5-yl)-8H-pyrimido[1,2-b]pyridazin-8-one, and its X-ray powder diffraction pattern is 4.4°±0.2°, 9.2°±0.2°, 13.9°±0.2°, 17.5°±0.2°, 18.0°±0.2° and 21.6°±0.2° at 2θ Preferably, the citrate salt form A has an X-ray powder diffraction pattern with diffraction peaks at 2θ of 4.4°±0.2°, 9.2°±0.2°, 13.9°±0.2°, 14.3°±0.2°, 15.3°±0.2°, 17.5°±0.2°, 18.0°±0.2° and 21.6°±0.2°; More preferably, the citrate salt form A has an X-ray powder diffraction pattern with diffraction peaks at 2θ of 4.4°±0 .2°, 9.2°±0.2°, 13.4°±0.2°, 13.9°±0.2°, 14.3°±0.2°, 15.3°±0.2°, 17.5°±0.2°, 18.0°±0.2°, 21.6°±0.2° and 22.6°±0.2°; further preferably, the citrate salt form A has an X-ray powder diffraction pattern at 2θ of 4.4°±0.2°, 7.6°±0.2°, 9.2°±0.2°, 12.2°±0.2°, 13.4°±0.2°, 13.9°±0.2°, 14.3°±0.2°, 15.3°±0.2°, 17.5°±0.2°, 18.0°±0.2°, 21.6°±0.2° and 22.6°±0.2° have diffraction peaks; further preferably, the X-ray powder diffraction of the citrate salt form A expressed in 2θ angles has a spectrum as shown in Figure 21; Preferably, the citrate salt form A has a hydrogen spectrum as shown in FIG22 .

5. The acid salt crystalline form according to claim 3, characterized in that: The acid salt crystalline form is the adipate crystalline form A of the compound 2-cyclopropyl-9-[4-(difluoromethoxy)phenyl]-7-(2-methyl-2H-indazol-5-yl)-8H-pyrimido[1,2-b]pyridazin-8-one, and its X-ray powder diffraction pattern has diffraction peaks at 2θ of 15.2°±0.2°, 17.6°±0.2°, 19.7°±0.2°, 21.8°±0.2°, 23.8°±0.2° and 24.6°±0.2°; preferably, the adipate crystalline form A has an X-ray powder diffraction pattern at 2θ of 9.2°±0.2°, 15.2° More preferably, the adipate salt crystalline form A has diffraction peaks at 2θ of 9.2°±0.2°, 14.2°±0.2°, 15.2°±0.2°, 17.6°±0.2°, 19.7°±0.2°, 21.8°±0.2°, 22.6°±0.2°, 23.8°±0.2°, 24.6°±0.2° and 25. 4°±0.2°; further preferably, the adipate salt form A has a diffraction peak at 2θ of 9.2°±0.2°, 13.0°±0.2°, 14.2°±0.2°, 15.2°±0.2°, 17.6°±0.2°, 19.7°±0.2°, 21.8°±0.2°, 22.6°±0.2°, 23.1°±0.2°, 23.8°±0.2°, 24.6°±0.2° and 25.4°±0.2°; further preferably, the adipate salt form A has an X-ray powder diffraction pattern at 2θ of 9.2°±0.2°, 13.0°±0.2°, 14.2°±0.2°, 15.2°±0.2°, 17.6°±0.2°, 19.7°±0.2°, 21.8°±0.2°, 22.6°±0.2°, 23.1°±0.2°, 23.8°±0.2°, 24.6°±0.2° and 25.4°±0.2°. Figure has diffraction peaks at 2θ of 9.2°±0.2°, 11.3°±0.2°, 13.0°±0.2°, 14.2°±0.2°, 15.2°±0.2°, 17.6°±0.2°, 19.7°±0.2°, 21.8°±0.2°, 22.6°±0.2°, 23.1°±0.2°, 23.8°±0.2°, 24.6°±0.2°, 25.4°±0.2° and 27.1°±0.2°; further preferably, the X-ray powder diffraction of the adipate salt form A expressed in 2θ angles has a spectrum as shown in Figure 23; Or the acid salt crystalline form is the adipate crystalline form B of the compound 2-cyclopropyl-9-[4-(difluoromethoxy)phenyl]-7-(2-methyl-2H-indazol-5-yl)-8H-pyrimido[1,2-b]pyridazin-8-one, whose X-ray powder diffraction pattern at 2θ is 8.7°±0.2°, 9.2°±0.2°, 13.7°±0.2°, 17.5°±0.2°, 17.9°±0.2° and 1 Preferably, the adipate salt crystalline form B has a diffraction peak at 2θ of 8.7°±0.2°, 9.2°±0.2°, 13.7°±0.2°, 17.5°±0.2°, 17.9°±0.2°, 19.3°±0.2°, 23.4°±0.2° and 24.0°±0.2°; More preferably, the adipate salt crystalline form B, whose X-ray powder diffraction pattern has diffraction peaks at 2θ of 5.9°±0.2°, 8.7°±0.2°, 9.2°±0.2°, 13.7°±0.2°, 16.8°±0.2°, 17.5°±0.2°, 17.9°±0.2°, 19.3°±0.2°, 23.4°±0.2° and 24.0°±0.2°; further preferably, the adipate salt crystalline form B, whose X-ray powder diffraction pattern has diffraction peaks at 2θ of 5.9°±0.2°, 8.7°±0.2°, 9.2°±0.2°, 13.7°±0.2°, 15.4°±0.2°, 16.8°±0.2°, 17.5°±0.2°, 17.9°±0.2°, 19.3°±0.2°, 23.4°±0.2°, 24.0°±0.2° and 25.4°±0.2° have diffraction peaks; further preferably, the X-ray powder diffraction of the adipate salt form B expressed in 2θ angles has a spectrum as shown in Figure 24; Preferably, the adipate salt form B has a hydrogen spectrum as shown in Figure 25; Or the acid salt crystalline form is the mesylate crystalline form A of the compound 2-cyclopropyl-9-[4-(difluoromethoxy)phenyl]-7-(2-methyl-2H-indazol-5-yl)-8H-pyrimido[1,2-b]pyridazin-8-one, whose X-ray powder diffraction pattern has 2θ of 11.2°±0.2°, 13.0°±0.2°, 18.1°±0.2°, 20.3°±0.2°, 22.6°±0.2° and 25.9°±0.2° Preferably, the mesylate salt form A has an X-ray powder diffraction pattern at 2θ of 11.2°±0.2°, 13.0°±0.2°, 18.1°±0.2°, 20.3°±0.2°, 21.2°±0.2°, 21.4°±0.2°, 22.6°±0.2° and 25.9°±0.2°; More preferably, the mesylate salt form A has an X-ray powder diffraction pattern at 2θ of 11.2°±0.2°, 13.0°±0.2°, 18.1°±0.2°, 20.3°±0.2°, 21.2°±0.2°, 21.4°±0.2°, 22.6°±0.2° and 25.9°±0.2°. The diffraction peaks are as follows: 0.2°, 13.0°±0.2°, 18.1°±0.2°, 20.3°±0.2°, 21.2°±0.2°, 21.4°±0.2°, 22.6°±0.2°, 23.9°±0.2°, 24.5°±0.2° and 25.9°±0.2°; further preferably, the mesylate salt form A has an X-ray powder diffraction pattern at 2θ of 11.2°±0.2°, 13.0°±0.2°, 18.1°±0.2°, 20.3°±0.2°, 21.2°±0.2°, 21.4°±0.2°, 22.6°±0.2°, 23.9°±0.2°, 24.5°±0.2° and 25.9°±0.2°. , 17.2°±0.2°, 18.1°±0.2°, 20.3°±0.2°, 21.2°±0.2°, 21.4°±0.2°, 22.6°±0.2°, 23.5°±0.2°, 23.9°±0.2°, 24.5°±0.2° and 25.9°±0.2° have diffraction peaks; further preferably, the X-ray powder diffraction of the mesylate salt form A expressed in 2θ angles has a spectrum as shown in Figure 26; Preferably, the mesylate salt form A has a hydrogen spectrum as shown in Figure 27; Or the acid salt crystalline form is the oxalate crystalline form A of the compound 2-cyclopropyl-9-[4-(difluoromethoxy)phenyl]-7-(2-methyl-2H-indazol-5-yl)-8H-pyrimido[1,2-b]pyridazin-8-one, and its X-ray powder diffraction pattern at 2θ is 10.7°±0.2°, 12.7°±0.2°, 15.4°±0.2°, 22.4°±0.2°, 24.6°±0.2° and 24.9°±0.2° Preferably, the oxalate crystalline form A has an X-ray powder diffraction pattern with diffraction peaks at 2θ of 7.5°±0.2°, 10.7°±0.2°, 12.7°±0.2°, 15.4°±0.2°, 17.6°±0.2°, 22.4°±0.2°, 24.6°±0.2° and 24.9°±0.2°; More preferably, the oxalate crystalline form A has an X-ray powder diffraction pattern with diffraction peaks at 2θ of 7.5°±0.2°, 10.7°±0.2°, 12.7°±0.2°, 15.4°±0.2°, 17.6°±0.2°, 22.4°±0.2°, 24.6°±0.2° and 24.9°±0.2°. .2°, 10.7°±0.2°, 12.7°±0.2°, 14.8°±0.2°, 15.4°±0.2°, 16.4°±0.2°, 17.6°±0.2°, 22.4°±0.2°, 24.6°±0.2° and 24.9°±0.2° have diffraction peaks; further preferably, the oxalate crystalline form A has an X-ray powder diffraction pattern at 2θ of 7.5°±0.2°, 10.7°±0.2°, There are diffraction peaks at 12.7°±0.2°, 14.8°±0.2°, 15.4°±0.2°, 16.4°±0.2°, 17.6°±0.2°, 18.8°±0.2°, 22.4°±0.2°, 24.6°±0.2°, 24.9°±0.2° and 25.5°±0.2°; further preferably, the X-ray powder diffraction of the oxalate crystalline form A expressed in 2θ angles has a spectrum as shown in Figure 28; Preferably, the oxalate crystal form A has a hydrogen spectrum as shown in Figure 29.

6. The acid salt crystalline form according to claim 3, characterized in that: The acid salt crystalline form is a solvent-containing or solvent-free crystalline form, wherein the solvent is selected from one or more of water, methanol, ethanol, n-propanol, isopropanol, tert-butanol, n-butanol, isobutanol, acetone, 2-butanone, 3-pentanone, dichloromethane, chloroform, ethyl formate, ethyl acetate, acetonitrile, tetrahydrofuran, 2-methyl-tetrahydrofuran, 1,4-dioxane, benzene, toluene, N,N-dimethylformamide, N,N-dimethylacetamide, N-methylpyrrolidone, dimethyl sulfoxide, n-heptane, heptane, isopropyl acetate, cyclohexane, methyl tert-butyl ether and isopropyl ether; preferably, the solvent is selected from one or more of water, ethanol, acetone, dichloromethane, chloroform, 2-butanone, tetrahydrofuran and N,N-dimethylformamide.

7. A method for preparing the acid salt or acid salt crystalline form according to any one of claims 1 to 6, characterized in that: The method comprises the step of forming a salt of the compound with a corresponding acid.

8. The preparation method according to claim 7, characterized in that: The method comprises the step of forming a salt with a corresponding acid in a reaction solvent; the reaction solvent is selected from one or more of water, methanol, ethanol, n-propanol, isopropanol, tert-butanol, n-butanol, isobutanol, acetone, 2-butanone, 3-pentanone, dichloromethane, chloroform, ethyl formate, ethyl acetate, acetonitrile, tetrahydrofuran, 2-methyl-tetrahydrofuran, 1,4-dioxane, benzene, toluene, N,N-dimethylformamide, N,N-dimethylacetamide, N-methylpyrrolidone, dimethyl sulfoxide, n-heptane, heptane, isopropyl acetate, cyclohexane, methyl tert-butyl ether and isopropyl ether; preferably one or more of water, ethanol, acetone, 2-butanone or chloroform; The molar ratio of the compound to the corresponding acid is 1:0.1-3; preferably 1:0.1-1.5; more preferably 1:0.3-1; the mass volume ratio of the compound to the reaction solvent is 1:10-50; preferably 1:20-30; more preferably 1:25-30.

9. A pharmaceutical composition comprising a therapeutically effective amount of the acid salt or acid salt crystalline form according to any one of claims 1 to 6 and a pharmaceutically acceptable carrier.

10. Use of the acid salt or acid salt crystalline form according to any one of claims 1 to 6 or the pharmaceutical composition according to claim 9 in the preparation of a medicament for preventing and / or treating a disease or disease state mediated by MAT2A.

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