A polymorphic form of a p-toluenesulfonic acid salt of a multicyclic compound, pharmaceutical compositions, and uses thereof

Abstract

The application belongs to the field of pharmaceutical chemistry, and relates to different p-toluenesulfonate crystal forms of a polycyclic compound (E)-2-(3-(2-cyano-2-(6-methoxy-3H-imidazo[4,5-c]pyridin-2-yl)vinyl)-2,5-dimethyl-1H-pyrrol-1-yl)-5-methylthiophene-3-carbonitrile, pharmaceutical compositions containing the crystal forms and purposes. The purity, water solubility, chemical stability and crystal form stability of the p-toluenesulfonate crystal form are all better than those of other crystal forms.

Description

TECHNICAL FIELD

[0001] The present application belongs to the field of pharmaceutical chemistry, and relates to different p-toluenesulfonic acid salt crystal forms of a polycyclic compound, namely (E)-2-(3-(2-cyano-2-(6-methoxy-3H-imidazo[4,5-c]pyridin-2-yl)vinyl)-2,5-dimethyl-1H-pyrrol-1-yl)-5-methylthiophene-3-carbonitrile (hereinafter referred to as "compound of formula (I)"), as well as to pharmaceutical compositions comprising the crystal forms and medical uses. BACKGROUND

[0002] DHX33 belongs to the DEAD / H-box containing RNA helicase protein family. Among them, DEAD / H represents the abbreviation of amino acids Asp-Glu-Ala-Asp / His, which appears in the protein sequence of the members of the RNA helicase family, and is highly involved in nucleic acid substrate binding and ATP hydrolysis. Although these family members share these same sequences, each RNA helicase has its own specific specificity and unique biological function. The molecular weight of human DHX33 protein is 72 kDa, which has the function of unwinding nucleic acids, and uses the biological energy released by ATP hydrolysis to drive the conformational change of RNA and protein complexes, and is involved in various RNA metabolic activities, specifically from RNA transcription, splicing, editing, translation to degradation, etc. The function of DHX33 is not limited to the modification of RNA molecules, studies have shown that in addition to unwinding RNA double strands, DHX33 protein is also involved in DNA metabolism. Specifically, DHX33 protein can unwind the double-stranded structure of DNA and plays an important role in gene expression.

[0003] Studies have shown that DHX33 affects the methylation state of DNA by binding to the promoters of various cancer-related genes, thereby regulating the expression of various cancer genes and tumor development-related signaling pathways at the genome level, and plays a crucial role in cell growth, proliferation, migration, apoptosis, metabolism and other cell activities. In addition, it is found that DHX33 can sense the invasion of foreign double-stranded RNA molecules and play an important role in the innate immunity of cells. As a very important cell growth regulatory gene, DHX33 is highly expressed in various cancers, such as lung cancer, lymphoma, glioblastoma, breast cancer, colon cancer, liver cancer, etc. The occurrence and development of various cancers depend on the high expression of DHX33 protein. Genetic knockout of DHX33 can significantly inhibit the occurrence and development of RAS oncogene-driven lung cancer; in vivo and in vitro experiments have confirmed that the occurrence and development of various cancers such as breast cancer, colon cancer, brain glioma, lymphoma, etc. are significantly inhibited after inhibiting DHX33 protein. Since the protein function of DHX33 depends on its helicase activity, the helicase activity mutant of DHX33 does not have the function of DHX33 protein, and thus cannot replace the protein function encoded by the wild-type DHX33 gene.

[0004] The applicant has found various compounds that can inhibit the RNA helicase activity of DHX33 (for example, (E)-2-(3-(2-cyano-2-(6-methoxy-3H-imidazo[4,5-c]pyridin-2-yl)vinyl)-2,5-dimethyl-1H-pyrrol-1-yl)-5-methylthiophene-3-carbonitrile), and verified that these compounds can significantly inhibit the growth and proliferation of cancer cells in vitro and in vivo. SUMMARY

[0005] The present application aims to provide a different p-toluenesulfonic acid salt crystal form of a polycyclic compound, namely (E)-2-(3-(2-cyano-2-(6-methoxy-3H-imidazo[4,5-c]pyridin-2-yl)vinyl)-2,5-dimethyl-1H-pyrrol-1-yl)-5-methylthiophene-3-carbonitrile (hereinafter referred to as "compound of formula (I)"), a pharmaceutical composition comprising the same, and uses thereof.

[0006] The structural formula of the compound of formula (I) is as follows:

[0007]

[0008] In a first aspect, the present application provides a p-toluenesulfonic acid salt crystal form 1 of the compound of formula (I), which has characteristic diffraction peaks at the following 2θ angles in the X-ray powder diffraction (XRPD) pattern: 4.6±0.2° and 18.3±0.2°.

[0009] In some embodiments, the X-ray powder diffraction pattern of the p-toluenesulfonic acid salt Form 1 of the compound of Formula (I) further comprises one or more of the following characteristic diffraction peaks: 21.2 ± 0.2°, 25.9 ± 0.2°, and 28.0 ± 0.2°.

[0010] In some embodiments, the X-ray powder diffraction pattern of the p-toluenesulfonic acid salt Form 1 of the compound of Formula (I) further comprises one or more of the following characteristic diffraction peaks: 4.1 ± 0.2°, 19.8 ± 0.2°, 24.1 ± 0.2°, and 26.3 ± 0.2°.

[0011] The present application also provides a method for preparing the p-toluenesulfonic acid salt Form 1 of the compound of Formula (I), comprising: adding an organic solvent to the free form of the compound of Formula (I), then adding p-toluenesulfonic acid, stirring at room temperature for about 2-5 days, collecting the solid and drying to obtain the p-toluenesulfonic acid salt Form 1. In a specific embodiment of the present application, the organic solvent is selected from isopropyl alcohol, acetone.

[0012] In an embodiment of the present application, in the above preparation method, the stirring can be carried out at room temperature for about 1-4 days, for example, about 1, 1.5, 2, 2.5, 3, 3.5, 4 days.

[0013] In an embodiment of the present application, in the above preparation method, the drying can be carried out at 40-95°C under vacuum for about 8-16 hours. In a specific embodiment, the drying can be carried out at 45, 50, 55, 60, 65, 70, 75, 80, 85, 90, 95°C under vacuum for about 8, 8.5, 9, 9.5, 10, 10.5, 11, 11.5, 12, 12.5, 13, 13.5, 14, 14.5, 15, 15.5, 16 hours.

[0014] In a third aspect, the present application provides a pharmaceutical composition comprising at least one of the above-mentioned crystal forms and one or more pharmaceutically acceptable carriers. The composition can be used for treating and / or preventing a disease or disorder mediated at least in part by DHX33, such as cancer, viral infection and inflammation, which can be, for example, breast cancer, colon cancer, brain glioma, lymphoma, etc.

[0015] In a fourth aspect, the present application provides use of the above-mentioned crystal form or the above-mentioned pharmaceutical composition in the preparation of a medicament for preventing and / or treating a disease or disorder mediated at least in part by DHX33.

[0016] In a fifth aspect, the present application provides a method for preventing and / or treating a disease or disorder mediated at least in part by DHX33, comprising the step of: administering to an individual in need thereof a prophylactically and / or therapeutically effective amount of the above-mentioned crystalline form or the above-mentioned pharmaceutical composition; preferably, the disease is selected from the group consisting of DHX33-mediated cancer, viral infection and inflammation.

[0017] The present application is not limited to the particular embodiments described herein; it should also be understood that the terminology used herein is for the purpose of describing particular embodiments only and is not intended to be limiting.

[0018] Definitions of Terms

[0019] Unless defined otherwise, technical and scientific terms used herein have the same meaning as commonly understood by one of ordinary skill in the art to which this application belongs. If there is a conflict between the definitions provided in this application and those provided in the art to which this application pertains, the definitions provided in this application control. When referring to a range, preferred range or preferred upper and lower limits of a value, concentration or other quantity or parameter, it is understood that the disclosure specifically contemplates the inclusion of any range formed by combining an upper limit of any range or preferred value with a lower limit of any range or preferred value, regardless of whether that range is specifically disclosed. Unless otherwise stated, numerical ranges expressed in the format "from X to Y" are understood to include X and Y. This is contrary to the normal practice of reciting the end values of a range. It is also understood that the endpoints of the ranges given herein are preferred endpoints. Also, it is intended that all transitional phrases such as "comprising," "including," "carrying," "having," "containing," "involving," "holding," "composed of," and the like are to be understood to be open-ended, i.e., to mean including but not limited to. Only the transitional phrases "consisting of and "consisting essentially of shall be construed as closed or semi-closed transitional phrases, respectively, meaning that the transitional phrases "consisting of and "consisting essentially of shall be inapplicable to the claim element following such phrases.

[0020] The term "about" when used before a numerical designation, typically means a value that is within 10% of the indicated value, unless otherwise clear from the context.

[0021] The recitation of a list of elements does not preclude additional such elements being present nor preclude such elements not being present unless specifically recited. The terms "comprising," "including," "containing," and "having" and the like are inclusive and open-ended and specify the presence of stated elements or integers or steps but do not preclude the presence or addition of one or more other elements, integers, steps, or groups thereof. The term "consisting of" is a closed term, and specifies a total list of elements, integers, or steps that are present. The term "consisting essentially of" is a semi-closed term, and specifies that additional elements, integers, or steps that do not materially affect the basic and novel characteristics of the claimed subject matter can be present. It is understood that the term "comprising" encompasses "consisting of" and "consisting essentially of."

[0022] The term "X-ray powder diffraction pattern (XRPD pattern)," refers to an experimentally observed diffraction pattern or parameters, data or values derived therefrom. An XRPD pattern is typically characterized by peak positions (abscissa) and / or peak intensities (ordinate).

[0023] The term "diffraction angle" or "2Θ" refers to the peak position in degrees (°) set in an X-ray diffraction experiment and is typically the unit of abscissa in a diffraction pattern. If a reflection is diffracted when the incident beam forms an angle of Θ with a certain lattice plane, the experimental setup requires that the reflected beam is recorded at an angle of 2Θ. It should be understood that the specific 2Θ values mentioned herein for a particular crystalline form are intended to represent the 2Θ values (in degrees) measured using the X-ray diffraction experimental conditions described herein.

[0024] It should be noted that in powder X-ray diffraction spectra, the position of a peak or the relative intensity of a peak can vary due to the measuring instrument, the measuring method / conditions, and the like. For any particular crystalline form, the position of a peak can be in error by ±0.2° in 2Θ. Therefore, in determining each crystalline form, this error should be taken into account, and within the error, it is also within the scope of the present application.

[0025] For the same crystalline form, the position of an endothermic peak of DSC can vary due to the measuring instrument, the measuring method / conditions, and the like. For any particular crystalline form, the position of an endothermic peak can be in error by ±5°C, and can be by ±3°C. Therefore, in determining each crystalline form, this error should be taken into account, and within the error, it is also within the scope of the present application.

[0026] For the same crystalline form, the position of a weight loss temperature of TGA can vary due to the measuring instrument, the measuring method / conditions, and the like. For any particular crystalline form, the position of a weight loss temperature can be in error by ±5°C, and can be by ±3°C. Therefore, in determining each crystalline form, this error should be taken into account, and within the error, it is also within the scope of the present application.

[0027] It should be understood that different types of equipment or different test conditions can give slightly different XRPD patterns and characteristic peaks or different DSC patterns and characteristic peaks. The specific numerical values provided should not be taken as absolute values.

[0028] The term "room temperature" refers to 20 °C ± 5 °C.

[0029] The term "prevention" refers to prophylactic administration to reduce the likelihood of the onset of a disease or symptoms, or to delay the onset of the disease or symptoms.

[0030] The term "treatment" is intended to refer to the alleviation or eradication of the disease state or condition being addressed. It is also to be appreciated that the treatment of a disease state or condition includes a partial but significant therapeutic effect in addition to complete treatment.

[0031] The term "pharmaceutically acceptable" refers to those compounds, materials, compositions and / or crystalline forms that are suitable for use in contact with human and animal tissues, within the scope of sound medical judgment, without excessive toxicity, irritation, allergic reaction or other problems or complications, and are commensurate with a reasonable benefit / risk ratio. "Pharmaceutically acceptable carrier" refers to an inert substance that is administered together with the active ingredient and facilitates the administration of the active ingredient, including but not limited to any glidant, sweetener, diluent, preservative, dye / colorant, flavor enhancer, surfactant, wetting agent, dispersant, disintegrant, suspending agent, stabilizer, isotonic agent, solvent or emulsifier that is acceptable for use in humans or animals (e.g., livestock) as approved by the State Food and Drug Administration.

[0032] Above-mentioned pharmaceutical composition can act systemically and / or act topically.For this purpose, they can be administered by suitable approach, for example by parenteral, local, intravenous, oral, subcutaneous, intraarterial, intradermal, percutaneous, rectal, intracranial, intraperitoneal, intranasal, intramuscular approach or as inhalant administration.

[0033] The above-mentioned route of administration can be achieved through suitable dosage forms. The dosage forms that can be used in the present invention include, but are not limited to, tablets, capsules, lozenges, hard candies, powders, sprays, creams, ointments, suppositories, gels, pastes, lotions, ointments, aqueous suspensions, injectable solutions, elixirs, syrups, etc.

[0034] When administered orally, the pharmaceutical composition can be prepared into any orally acceptable preparation form, including but not limited to tablets, capsules, aqueous solutions, aqueous suspensions, and the like.

[0035] The term "disease or condition mediated at least in part by DHX33" refers to a disease whose pathogenesis at least partially involves factors related to DHX33, such as cancer, viral infection, and inflammation.

[0036] The term "effective amount" refers to a dose that can induce a biological or medical response in cells, tissues, organs or organisms (eg, individuals) and is sufficient to achieve the desired preventive and / or therapeutic effect.

[0037] The therapeutically effective amount of the crystalline form described herein is from about 0.0001-20 mg / Kg body weight / day, for example, from 0.001-10 mg / Kg body weight / day.

[0038] The dosage frequency of the crystalline forms described herein can be determined by a physician based on the needs of the individual patient, for example, once or twice a day, or more times a day. Administration can be intermittent, for example, wherein the patient receives a daily dose of the crystalline form over a period of several days, followed by a period of several or more days in which the patient does not receive a daily dose of the crystalline form. BRIEF DESCRIPTION OF THE DRAWINGS

[0039] Figure 1 is an X-ray powder diffraction (XRPD) pattern of Form 1 sulfate salt of the compound of Formula (I).

[0040] Figure 2 is an X-ray powder diffraction (XRPD) pattern of Form 1 p-toluenesulfonic acid salt of the compound of Formula (I). 1 HNMR pattern.

[0041] Figure 3 is an X-ray powder diffraction (XRPD) pattern of Form 1 sulfate salt of the compound of Formula (I).

[0042] Figure 4 is an X-ray powder diffraction (XRPD) pattern of Form 2 sulfate salt of the compound of Formula (I).

[0043] Figure 5 is an X-ray powder diffraction (XRPD) pattern of Form 4 sulfate salt of the compound of Formula (I).

[0044] Figure 6 is an X-ray powder diffraction (XRPD) pattern of Form 4 sulfate salt of the compound of Formula (I). 1 HNMR pattern.

[0045] Figure 7 is an X-ray powder diffraction (XRPD) pattern of Form 1 p-toluenesulfonic acid salt of the compound of Formula (I).

[0046] Figure 8 is an X-ray powder diffraction (XRPD) pattern of Form 2 p-toluenesulfonic acid salt of the compound of Formula (I).

[0047] Figure 9 is a thermogravimetric analysis (TGA) pattern of Form 1 p-toluenesulfonic acid salt of the compound of Formula (I).

[0048] Figure 10 is a differential scanning calorimetry (DSC) pattern of Form 1 p-toluenesulfonic acid salt of the compound of Formula (I).

[0049] Figure 11 is an X-ray powder diffraction (XRPD) pattern of Form 1 p-toluenesulfonic acid salt of the compound of Formula (I). 1 HNMR pattern.

[0050] Figure 12 is an XRPD comparison pattern of Form 1 p-toluenesulfonic acid salt of the compound of Formula (I) before and after stability testing.

[0051] Figure 13 is a moisture uptake DVS test pattern of Form 1 p-toluenesulfonic acid salt of the compound of Formula (I).

[0052] Figure 14is the XRPD comparison chart of p-toluenesulfonic acid salt Form 1DVS of the compound of formula (I) before and after the test. DETAILED DESCRIPTION

[0053] The intermediate compounds of the present application can be prepared by a variety of synthetic methods well known to those skilled in the art, including the specific embodiments set forth below, embodiments formed by a combination of the specific embodiments with other chemical synthetic methods well known to those skilled in the art, and equivalents thereof as appreciated by those skilled in the art, preferred embodiments include but are not limited to the examples of the present application.

[0054] The chemical reactions of the specific embodiments of the present application are performed in solvents appropriate to the reagents and materials employed and suitable for the chemical changes involved. In some instances, the protecting groups can need to be removed using procedures typical for one skilled in the art. In order to obtain the compounds of the present application, it can be necessary to make modifications or selections of the synthetic sequences or reaction routes on the basis of the existing embodiments by one skilled in the art.

[0055] The present application will be described in detail below with the examples, which are not meant to limit the present application in any way.

[0056] X-ray powder diffraction (XRPD)

[0057] XRPD test parameters

[0058]

[0059] Thermogravimetric analysis (TGA) and differential scanning calorimetry (DSC)

[0060] TGA and DSC test parameters

[0061]

[0062]

[0063] H NMR spectrum ( 1 HNMR)

[0064] 1 H NMR test parameters

[0065]

[0066] Ion chromatography (IC)

[0067] IC test parameters

[0068]

[0069] High performance liquid chromatography (HPLC) HPLC test parameters

[0070]

[0071]

[0072] Dynamic vapor sorption analysis (DVS)

[0073] DVS test parameters

[0074]

[0075] Example 1: Synthesis of compound of formula (I) and its characterization

[0076] 1.1 The synthesis route is as follows:

[0077]

[0078] wherein R is

[0079] 1.2 Synthesis method:

[0080] 1.2.1 Preparation of compound 2 (6-methoxypyridine-3,4-diamine)

[0081]

[0082] Compound 1 (2-methoxy-5-nitropyridin-4-amine) (3.0 g, 17.74 mmol, 1.0 eq.) was dissolved in methanol (30 mL), and carbon supported palladium catalyst (300 mg, 0.1 wt%) was added. The mixture was stirred at room temperature for 16 h in the presence of hydrogen. After filtration and concentration, compound 2 (6-methoxypyridine-3,4-diamine) (2.6 g, yield: 100%) was obtained as a brown solid. MS (ESI) m / z: 140 [M+H]+; TLC: DCM:MeOH (10:1); Rf(compound 2) = 0.5. f :Rf(compound 1) = 0.7; Rf(compound 3) = 0.5. f :Rf(compound 1) = 0.7; Rf(compound 3) = 0.5.

[0083] 1.2.2 Preparation of compound 4 (2-(6-methoxy-3H-imidazo[4,5-c]pyridin-2- yl)acetonitrile)

[0084]

[0085] Compound 2 (6-methoxy-pyridine-3,4-diamine) (1.5 g, 10.2 mmol, 1.0 eq.) and compound 3 (ethyl cyanoacetate) (3.5 g, 30.6 mmol, 3.0 eq.) were dissolved in dimethylformamide (6 mL) and stirred at 180 °C for 5 h. After cooling, the solvent was removed. The residue was purified by flash column chromatography (dimethylformamide:methanol = 200:1 to 50:1) to give compound 4 (2-(6-methoxy-3H-imidazo[4,5-c]pyridin-2-yl)acetonitrile) (500 mg, yield: 26.0%) as a brown solid powder. MS (ESI) m / z: 188 [M+H]+; TLC: petroleum ether / ethyl acetate (1:1); Rf (compound 4): 0.2. f :Rf (compound 1) = 0.5; Rf (compound 3) = 0.2. f :Rf (compound 1) = 0.5; Rf (compound 3) = 0.2.

[0086] 1.2.3, Synthesis of (E)-2-(3-(2-cyano-2-(6-methoxy-3H-imidazo[4,5-c]pyridin-2-yl)vinyl)-2,5-dimethyl-1H-pyrrol-1-yl)-methylthiophene-3-carbonitrile (Compound A)

[0087]

[0088] Compound 4 (2-(6-methoxy-3H-imidazo[4,5-c]pyridin-2-yl)acetonitrile) (80 mg, 0.42 mmol, 1.0 eq.) was dissolved in 1 mL of ethanol, and compound 5 (2-(3-formyl-2,5-dimethyl-1H-pyrrol-1-yl)-methylthiophene-3-carbonitrile) (103 mg, 0.42 mmol, 1.0 eq.) and piperidine (36 mg, 0.42 mmol, 1.0 eq.) were added. After heating the mixture to reflux and stirring for 1 h, the reaction was completed. The reaction was cooled to room temperature and filtered. Compound AB24288 ((E)-2-(3-(2-cyano-2-(6-methoxy-3H-imidazo[4,5-c]pyridin-2-yl)vinyl)-2,5-dimethyl-1H-pyrrol-1-yl)-methylthiophene-3-carbonitrile) (95 mg, yield: 13.6%) was obtained as a yellow powder after drying the collected solid.

[0089] 1 1H NMR (400 MHz, DMSO-d6) d 8.53 (s, 1H), 8.16 (s, 1H), 7.31 (s, 1H), 6.96 (s, 1H), 6.89 (s, 1H), 3.89 (s, 3H), 2.51 (s, 3H), 2.30 (s, 3H), 2.08 (s, 3H).

[0090] The free form of the compound of formula (I) is in amorphous state, having an XRPD pattern substantially as shown in Figure 1

[0091] The free form of the compound of formula (I) 1 The H NMR pattern is as shown in Figure 2

[0092] In view of the poor aqueous solubility of the free form of the weak base compound of formula (I), in order to improve the water solubility of the compound and facilitate the improvement of its bioavailability, the present application uses two strong acids, sulfuric acid and p-toluenesulfonic acid, to carry out salt formation reaction, aiming to improve the physicochemical properties of the compound and facilitate the later formulation development.

[0093] Example 2: Sulfate salt crystal form of the compound of formula (I) and a preparation method thereof

[0094] Process for preparing a sulfate salt crystalline form of a compound of Formula (I)

[0095] Pilot test: about 20 mg of the free form of the compound of formula (I) and a certain molar amount of counterion (1.1 eq.) were weighed into a 3.0 mL vial, 1.0 mL of solvent (isopropyl alcohol, acetone and ethyl acetate were selected) was added, and magnetic stirring was carried out at room temperature. After stirring for 2 days, the obtained solid was separated and dried at 50°C under blast, and then XRPD was measured. Three sulfate salt crystal forms, i.e. sulfate salt crystal form 1 / 2 / 3 (weak crystal state), were obtained, wherein crystal form 1 was obtained by using isopropyl alcohol as the solvent ( Figure 3 ); crystal form 2 was obtained by using acetone as the solvent ( Figure 4 ); and crystal form 3 was obtained by using ethyl acetate as the solvent, and no XRPD pattern was provided due to poor crystallinity. From the XRPD pattern, the purity of the sulfate salt crystal form 1 is poor, so acetone is selected as the solvent for the later scale-up synthesis of the sulfate salt crystal form 2 of the compound of formula (I).

[0096] Scale-up synthesis: 199.92 mg of the free form of the compound of formula (I) was weighed into 5 mL of acetone to form a suspension; 10 mL of acetone solution containing 29.0 μL of sulfuric acid (1.1 eq.) was added under stirring at room temperature; the system was turbid after stirring for 4 days under cyclic temperature rising and falling, and then centrifugation was carried out, and the solid was dried at 50°C under blast for 2 hours; the solid was crushed, and the drying was continued at 50°C under blast overnight. The separated solid was tested by XRPD, and the crystal form 2 obtained in the pilot test was converted into the sulfate salt crystal form 4.

[0097] The sulfate salt crystal form 4 has an XRPD pattern substantially as shown in Figure 5

[0098] The XRPD pattern of the sulfate salt crystal form 4 has characteristic diffraction peaks at the following 2θ angles, as shown in Table 1:

[0099] Table 1

[0100] ​​​

[0101] Sulfate Form 4 1 H-NMR spectrum Figure 6 As shown. Figure 6 It can be seen that the structure of this compound is no different from that of the free base compound.

[0102] The results of high performance liquid chromatography (HPLC) showed that the HPLC purity of sulfate decreased significantly during the drying process and the solid state stability was poor. The HPLC purity data are shown in Table 2:

[0103] Table 2

[0104] Solid form HPLC purity Free base (amorphous) 96.20 Sulfate salt crystalline form 4 75.10

[0105] Example 3: Preparation and Characterization of the Crystalline Form of the Compound of Formula (I) p-Toluenesulfonate

[0106] Small test: weigh about 20 mg of the free form of the compound of formula (I) and a certain molar amount of the counterion (1.1 eq.) into a 3.0 mL vial, add 1.0 mL of solvent (selected from isopropanol, acetone and ethyl acetate), and stir magnetically at room temperature. After stirring for 2 days, separate the resulting solid and air-dry at 50°C before XRPD analysis. Two p-toluenesulfonate salt forms 1 / 2 were obtained, of which Form 1 was obtained using isopropanol and acetone as solvents. Form 1 of the p-toluenesulfonate salt has substantially the same Figure 7 The XRPD pattern shown; Form 2 was obtained by using ethyl acetate as solvent ( Figure 8 From the XRPD pattern, the crystallinity of the p-toluenesulfonate salt Form 1 was slightly better. Acetone was subsequently selected as the solvent to amplify the synthesis of the p-toluenesulfonate salt Form 1 of the compound of formula (I).

[0107] Scale-up synthesis: 200.16 mg of the free form was weighed and suspended in 5 mL of acetone. With stirring at room temperature, 5 mL of an acetone solution containing 90.99 mg of p-toluenesulfonic acid (1.1 eq.) was added. Cyclic heating and stirring for 4 days resulted in turbidity. The solution was centrifuged and air-dried at 50°C for 2 hours. The solid was crushed and air-dried at 50°C overnight. (Cyclic heating and cooling program: 40°C (120 min) -> 40°C -> 5°C (0.1°C / min) -> 5°C (120 min) -> 40°C (0.1°C / min) -> 40°C (120 min) -> 5°C (0.1°C / min) -> 5°C (120 min), two cycles, followed by raising the temperature to 25°C with constant stirring.

[0108] The p-toluenesulfonate salt form 1 has substantially the following Figure 7 The XRPD pattern of the crystal form 1 has characteristic diffraction peaks at the following 2θ angles, as shown in Table 3.

[0109] Table 3

[0110]

[0111]

[0112] like Figure 9 As shown in the thermogravimetric analysis (TGA) curve of the p-toluenesulfonate salt form 1, the sample loses about 1.23% of its weight when heated to 120°C, indicating thermal stability. The p-toluenesulfonate salt form 1 is an anhydrate.

[0113] like Figure 10 As shown, the differential scanning calorimetry (DSC) curve of p-toluenesulfonate salt form 1 has an endothermic peak at 258.54°C.

[0114] In the p-toluenesulfonate salt form 1, the molar ratio of the compound of formula (I) to p-toluenesulfonic acid is about 1:1, and it is a monosalt.

[0115] p-Toluenesulfonate Form 1 1 H-NMR spectrum Figure 11 As shown in Figure 2, NMR analysis shows that the structure of this crystalline compound is identical to that of the free base. The base-acid ratio is approximately 1:1, and no significant organic solvent residue is observed.

[0116] Example 3: 24-hour equilibrium solubility experiment in water

[0117] The 24-hour equilibrium solubility of the free base compound of formula (I) and the p-toluenesulfonate salt form 1 was tested. The specific steps are as follows: 10 mg of each sample was weighed and added to 1 mL of the corresponding solvent to prepare a suspension, magnetically stirred at 37±2°C, sampled after 24 hours, centrifuged, and the supernatant was filtered and then subjected to solubility testing. The supernatant was filtered through a 0.22 μm PTFE filter membrane and then subjected to solubility testing. The results are shown in Table 4. The water solubility of the free base compound of formula (I) is approximately 16.2 μg / mL, and the solubility of the p-toluenesulfonate salt form 1 is 28.43 μg / mL.

[0118] Table 4

[0119] Solid form Equilibrium solubility in water at 37°C (pg / mL) Free base (amorphous) 16.2 p-toluenesulfonate salt crystalline form 1 28.43

[0120] Example 4: One-week stability experiment

[0121] The free base and p-methanesulfonate crystal forms 1 and 2 were placed under 25°C / 60%RH (long term) and 40°C / 75%RH (accelerated) conditions for one week, and then the HPLC purity and crystal form changes were tested ( Figure 12). The results are shown in Table 5. After one week of storage under the two test conditions, the HPLC purity and crystal form of the p-toluenesulfonate salt Form 1 did not change significantly, and the chemical stability and crystal form stability of the p-toluenesulfonate salt Form 1 were both good. The comparison of the spectra is shown in Figure 12

[0122] Table 5

[0123]

[0124] Example 5: Hygroscopicity

[0125] The stability risk of the sample at 25°C with humidity change was evaluated, and the p-toluenesulfonate salt Form 1 with good solid-state stability was subjected to DVS testing Figure 13 ), and the solid sample after the test was collected for XRPD testing Figure 14 ).

[0126] The results are summarized in Table 6 and shown in Figure 14 The p-toluenesulfonate salt Form 1 was slightly hygroscopic. After the test, the crystal form of the sample did not change.

[0127] Table 6 Summary of Hygroscopicity Evaluation Results

[0128]

[0129] *: Hygroscopicity definition in the 2020 edition of the Chinese Pharmacopoeia: under the condition of 25°C / 80% RH, the weight gain of the sample is 0.2-2% (less than 2% but not less than 0.2%) for slight hygroscopicity.​

Claims

1. A crystalline form of a p-toluenesulfonate salt of a polycyclic compound, wherein the structural formula of the compound is as shown in formula (I): in, The p-toluenesulfonate crystalline form is p-toluenesulfonate crystalline form 1, and its X-ray powder diffraction pattern has characteristic diffraction peaks at the following 2θ angles: 3.8±0.2°, 12.7±0.2°, 23.0±0.2°, 25.3±0.2° and 27.2±0.2°.

2. The crystal form according to claim 1, wherein Its X-ray powder diffraction pattern also has characteristic diffraction peaks at one or more of the following 2θ angles: 13.4±0.2°, 18.8±0.2°, 20.3±0.2° and 25.2±0.2°.

3. The crystal form according to claim 1, wherein The p-toluenesulfonate salt Form 1 has an XRPD pattern substantially as shown in FIG5 .

4. A pharmaceutical composition comprising the crystal form according to any one of claims 1 to 3 and a pharmaceutically acceptable carrier.

5. Use of the crystalline form according to any one of claims 1 to 3 or the pharmaceutical composition according to claim 4 in the preparation of a medicament for preventing and / or treating a disease or condition at least partially mediated by DHX33.

6. The use according to claim 5, wherein The disease is selected from the group consisting of DHX33-mediated cancer, viral infection, and inflammation.

7. The method for preparing the p-toluenesulfonate crystalline form 1 according to claim 1, comprising: An organic solvent is added to the free state of the compound of formula (I), followed by adding p-toluenesulfonic acid, stirring, collecting the solid, and then drying to obtain the p-toluene methanesulfonate salt form 1, wherein the organic solvent is selected from isopropanol and acetone.

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