CN115710257A

CN115710257A - Sulfate crystal form of amide compound, pharmaceutical composition and application

Info

Publication number: CN115710257A
Application number: CN202211486081.1A
Authority: CN
Inventors: 张严冬
Original assignee: Shenzhen Keye Health Co ltd
Current assignee: Shenzhen Keye Health Co ltd
Priority date: 2022-11-24
Filing date: 2022-11-24
Publication date: 2023-02-24

Abstract

The invention belongs to the field of pharmaceutical chemistry, and relates to different sulfate crystal forms of an amide compound, namely 1- (3-cyano-5-methylthiophene-2-yl) -N- (6-methoxy-1H-benzo [ d ] imidazole-2-yl) -2, 5-dimethyl-1H-pyrrole-3-carboxamide, a pharmaceutical composition containing the crystal forms, and application of the pharmaceutical composition. The chemical stability and the crystal form stability of the crystal form of the sulfate are better than those of the crystal form of free alkali.

Description

Sulfate crystal form of amide compound, pharmaceutical composition and application

Technical Field

The invention belongs to the field of pharmaceutical chemistry, and relates to different sulfate crystal forms of an amide compound, namely 1- (3-cyano-5-methylthiophene-2-yl) -N- (6-methoxy-1H-benzo [ d ] imidazole-2-yl) -2, 5-dimethyl-1H-pyrrole-3-carboxamide (hereinafter referred to as a' compound of formula (1)), and a pharmaceutical composition containing the crystal forms and medical application.

Background

DHX33 belongs to the family of RNA helicase proteins that contain a DEAD/H cassette. Among them, DEAD/H stands for the amino acid abbreviation Asp-Glu-Ala-Asp/His, which is present in the protein sequences of members of the RNA helicase family, together with a number of other conserved amino acid sequences, 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 the human DHX33 protein is 72kDa, has the function of unwinding nucleic acid, utilizes the bioenergy released by ATP hydrolysis to drive the change of the conformation of RNA and protein complexes, and further participates in a series of biological processes of RNA transcription, shearing, editing, translation, degradation and the like. The function of DHX33 is not limited to the modification of RNA molecules, and studies have shown that DHX33 proteins are involved in DNA metabolism in addition to unwinding RNA duplexes. In particular, DHX33 protein can unravel the double-stranded structure of DNA and play an important role in the gene expression process.

Research shows that DHX33 is combined with gene promoters related to various cancers to influence the methylation state of DNA, so that the expression of various cancer genes and signal paths related to tumor development are regulated and controlled at the genome level, and the DHX plays a vital role in various cell activities such as cell growth, proliferation, migration, apoptosis, metabolism and the like. In addition, DHX33 was found to sense the invasion of foreign double stranded RNA molecules and play an important role in the innate immunity of cells. DHX33 is used as an important cell growth regulation gene and is highly expressed in various cancers, such as lung cancer, lymphoma, glioblastoma, breast cancer, colon cancer, liver cancer and the like. The development of various cancers is dependent on the high expression of DHX33 protein. The genetic knockout of DHX33 can obviously inhibit the generation and development of the lung cancer driven by RAS oncogene; in vivo and in vitro experiments prove that after DHX33 protein is inhibited, the occurrence and the development of various cancers such as breast cancer, colon cancer, brain glioma, lymphoma and the like are obviously inhibited. Because the protein function of DHX33 depends on the helicase activity of the DHX33, the mutant with the deletion of the helicase activity of DHX33 does not have the function of DHX33 protein, and can not replace the protein function coded by the wild DHX33 gene.

Applicants have discovered that a variety of compounds that inhibit the RNA helicase activity of DHX33 (e.g., 1- (3-cyano-5-methylthiophen-2-yl) -N- (6-methoxy-1H-benzo [ d ] imidazol-2-yl) -2, 5-dimethyl-1H-pyrrole-3-carboxamide) and demonstrated that these compounds significantly inhibit the growth and proliferation of cancer cells both in vitro and in vivo.

Disclosure of Invention

The present invention aims to provide different crystal forms of the sulfate salt of an amide compound, i.e. 1- (3-cyano-5-methylthiophen-2-yl) -N- (6-methoxy-1H-benzo [ d ] imidazol-2-yl) -2, 5-dimethyl-1H-pyrrole-3-carboxamide (hereinafter referred to as "compound of formula (1)"), pharmaceutical compositions comprising the crystal forms and uses thereof.

The structural formula of the compound of formula (1) is as follows:

in a first aspect, the present invention provides a crystalline form a of the sulphate salt of the compound of formula (1) having an X-ray powder diffraction (XRPD) pattern with characteristic diffraction peaks at the following 2 Θ angles: 5.5 + -0.2 deg. and 11.0 + -0.2 deg..

In some embodiments, the X-ray powder diffraction pattern of the sulfate salt form a of the compound of formula (1) also has characteristic diffraction peaks at one or more of the following 2 Θ angles: 21.5 +/-0.2 degrees, 22.0 +/-0.2 degrees and 23.6 +/-0.2 degrees.

In some embodiments, the X-ray powder diffraction pattern of the sulfate salt form a of the compound of formula (1) also has characteristic diffraction peaks at one or more of the following 2 Θ angles: 10.2 + -0.2 deg., 13.0 + -0.2 deg. and 18.8 + -0.2 deg..

The present invention also provides a process for the preparation of the sulfate salt form a of the compound of formula (1) comprising: adding an organic solvent into free crystal form A of the compound shown in the formula (1), then adding sulfuric acid, stirring for about 2-5 days at room temperature, collecting a solid, and drying to obtain the sulfate crystal form A. In a specific embodiment of the present invention, the organic solvent is selected from one or both of acetone and isopropanol.

In a second aspect, the present invention provides crystalline form B of the sulphate salt of the compound of formula (1) having an X-ray powder diffraction (XRPD) pattern with characteristic diffraction peaks at the following 2 Θ angles: 5.5 +/-0.2 degrees, 16.1 +/-0.2 degrees and 21.0 +/-0.2 degrees.

In some embodiments, the X-ray powder diffraction pattern of the crystalline form B sulfate salt of the compound of formula (1) also has characteristic diffraction peaks at one or more of the following 2 Θ angles: 11.8 +/-0.2 degrees, 25.2 +/-0.2 degrees and 26.8 +/-0.2 degrees.

In some embodiments, the X-ray powder diffraction pattern of the mesylate salt form B of the compound of formula (1) also has characteristic diffraction peaks at one or more of the following 2 Θ angles: 15.0 +/-0.2 degrees, 19.2 +/-0.2 degrees and 23.8 +/-0.2 degrees.

The present invention also provides a process for the preparation of the crystalline form B of the sulfate salt of the compound of formula (1), comprising: adding an organic solvent into the free crystal form A of the compound shown in the formula (1), then adding sulfuric acid, stirring for about 2-5 days at room temperature, collecting a solid, and drying to obtain the sulfate crystal form B. In a particular embodiment of the invention, the organic solvent is acetonitrile.

In an embodiment of the present invention, in the above preparation method, the stirring may be performed at room temperature for about 1 to 4 days, for example, about 1, 1.5, 2, 2.5, 3, 3.5, 4 days.

In an embodiment of the present invention, in the above preparation method, the drying may be performed at 40 to 95 ℃ for about 8 to 16 hours under vacuum. In particular embodiments, drying may be performed at 45, 50, 55, 60, 65, 70, 75, 80, 85, 90, 95 ℃ 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.

In a third aspect, the present invention provides a pharmaceutical composition comprising at least one of the above crystalline forms and one or more pharmaceutically acceptable carriers. The compositions can be used to treat and/or prevent diseases or disorders mediated at least in part by DHX33, such as cancer, viral infections and inflammation, such as breast cancer, colon cancer, brain glioma, lymphoma and the like.

In a fourth aspect, the present invention provides the use of the crystalline form as described above or the pharmaceutical composition as described above for the manufacture of a medicament for the prevention and/or treatment of a disease or condition mediated at least in part by DHX 33.

In a fifth aspect, the present invention provides a method for the prevention and/or treatment of a disease or condition mediated at least in part by DHX33, comprising the steps of: administering a prophylactically and/or therapeutically effective amount of the crystalline form or the pharmaceutical composition described above to an individual in need thereof; preferably, the disease is selected from the group consisting of cancer, viral infection and inflammation mediated by DHX 33.

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

Definition of terms

Unless defined otherwise, all technical and scientific terms used herein have the same meaning as commonly understood by one of ordinary skill in the art to which this invention belongs. In case of conflict, the present application will control. When an amount, concentration, or other value or parameter is expressed in terms of a range, preferred range, or upper preferable numerical value and lower preferable numerical value, it is understood that any range defined by any pair of upper range limits or preferred numerical values in combination with any lower range limits or preferred numerical values is specifically disclosed, regardless of whether the range is specifically disclosed. Unless otherwise indicated, numerical ranges set forth herein are intended to include the endpoints of the ranges and all integers and fractions (decimal) within the range.

The term "about," when used in conjunction with a numerical variable, generally means that the value of the variable and all values of the variable are within experimental error (e.g., within 95% confidence interval for the mean) or within ± 10% of the specified value, or more.

The expressions "comprising" or similar expressions "including", "containing" and "having" and the like which are synonymous are open-ended and do not exclude additional, unrecited elements, steps or components. The expression "consisting of 8230comprises" excludes any element, step or ingredient not specified. The expression "consisting essentially of 8230comprises means that the scope is limited to the specified elements, steps or components, plus optional elements, steps or components that do not materially affect the basic and novel characteristics of the claimed subject matter. It is to be understood that the expression "comprising" covers the expressions "consisting essentially of and" consisting of 823030303030303030the expression "comprises" or "comprises" is used.

The term "X-ray powder diffraction pattern (XRPD pattern)" refers to an experimentally observed diffraction pattern or a parameter, data or value derived therefrom. XRPD patterns are generally characterized by peak position (abscissa) and/or peak intensity (ordinate).

The term "diffraction angle" or "2 θ" refers to the peak position in degrees (°) set based on X-ray diffraction experiments, and is typically the abscissa unit in the diffraction pattern. If the reflection is diffracted when the incident beam makes an angle theta with a certain lattice plane, the experimental setup requires recording the reflected beam at an angle of 2 theta. It should be understood that reference herein to particular 2 theta values for particular crystalline forms is intended to refer to 2 theta values (in degrees) measured using the X-ray diffraction experimental conditions described herein.

In the powder X-ray diffraction spectrum, the position of a peak or the relative intensity of the peak may vary depending on the measurement instrument, the measurement method, the conditions, and the like. For any particular crystal form, there may be errors in the position of the peaks, and the error in the determination of the 2 θ values may be ± 0.2 °. Therefore, this error should be taken into account when determining each crystal form, and is within the scope of this application.

For the same crystal form, the position of the endothermic peak of DSC may vary depending on the measurement apparatus, measurement method/conditions, and the like. For any particular crystal form, there may be an error in the position of the endothermic peak, which may be ± 5 ℃ and may be ± 3 ℃. Therefore, this error should be taken into account when determining each crystal form, and is within the scope of this application.

For the same crystal form, the appearance position of the weight loss temperature of the TGA may vary due to factors such as the measurement instrument, the measurement method/conditions, and the like. For any particular crystal form, there may be an error in the position of the weight loss temperature, which may be ± 5 ℃ and may be ± 3 ℃. Therefore, this error should be taken into account when determining each crystal form, and is within the scope of this application.

It will be appreciated that different types of equipment or with different test conditions may give slightly different XRPD patterns and characteristic peaks or different DSC patterns and characteristic peaks. The specific values provided are not to be taken as absolute values.

The term "room temperature" means 20 ℃. + -. 5 ℃.

The term "preventing" refers to prophylactic administration to reduce the likelihood of or delay the onset of a disease or condition.

The term "treatment" is intended to alleviate or eliminate the disease state or disorder to which it is directed. It is also understood that treatment of the disease state or condition described includes not only complete treatment, but also less than complete treatment, but achieves some biologically or medically relevant result.

The term "pharmaceutically acceptable" is intended to refer to those compounds, materials, compositions, and/or crystalline forms which are, within the scope of sound medical judgment, suitable for use in contact with the tissues of human beings and animals without excessive toxicity, irritation, allergic response, or other problem or complication, commensurate with a reasonable benefit/risk ratio. By "pharmaceutically acceptable carrier" is meant an inert substance which facilitates administration of the active ingredient in conjunction with 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, isotonizing agent, solvent, or emulsifier acceptable for use in humans or animals (e.g., livestock) as permitted by the national food and drug administration.

The pharmaceutical compositions described above may act systemically and/or locally. For this purpose, they may be administered by a suitable route, for example by parenteral, topical, intravenous, oral, subcutaneous, intraarterial, intradermal, transdermal, rectal, intracranial, intraperitoneal, intranasal, intramuscular routes or as an inhalant.

The above administration route can be achieved by a suitable dosage form. Dosage forms that may be used in the present invention include, but are not limited to: tablets, capsules, troches, hard candies, powders, sprays, creams, ointments, suppositories, gels, pastes, lotions, ointments, aqueous suspensions, injectable solutions, elixirs, syrups, and the like.

When administered orally, the above pharmaceutical compositions may be formulated into any orally acceptable dosage form including, but not limited to, tablets, capsules, aqueous solutions, aqueous suspensions, and the like.

The term "a disease or condition mediated at least in part by DHX 33" refers to a disease whose pathogenesis includes at least a portion of the factors associated with DHX33, such as cancer, viral infection, and inflammation.

The term "effective amount" refers to a dose that is capable of inducing a biological or medical response in a cell, tissue, organ or organism (e.g., an individual) and is sufficient to achieve a desired prophylactic and/or therapeutic effect.

A therapeutically effective amount of the crystalline form described herein is from about 0.0001 to 20mg/Kg body weight/day, for example from 0.001 to 10mg/Kg body weight/day.

The dosage frequency of the crystalline forms described herein may be determined by a physician, for example, 1 or 2 times per day, or more times per day, depending on the individual needs of the patient. Administration may 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 days or more during which the patient does not receive a daily dose of the crystalline form.

Drawings

Figure 1 is an X-ray powder diffraction (XRPD) pattern of free form a of the compound of formula (1).

Figure 2 is a thermogravimetric analysis (TGA) plot of free crystalline form a of the compound of formula (1).

FIG. 3 is a Differential Scanning Calorimetry (DSC) profile of free form A of the compound of formula (1).

FIG. 4 is a free crystalline form A of the compound of formula (1) ¹ HNMR map.

Figure 5 is an X-ray powder diffraction (XRPD) pattern of the sulfate salt form a of the compound of formula (1).

Figure 6 is a thermogravimetric analysis (TGA) plot of the sulfate salt form a of the compound of formula (1).

FIG. 7 is a Differential Scanning Calorimetry (DSC) profile of the sulfate salt form A of the compound of formula (1).

FIG. 8 is a crystalline form A of the sulfate salt of the compound of formula (1) ¹ HNMR map.

Figure 9 is an X-ray powder diffraction (XRPD) pattern of the sulfate salt form B of the compound of formula (1).

Figure 10 is a thermogravimetric analysis (TGA) plot of the crystalline form B of the sulfate salt of the compound of formula (1).

Figure 11 is a Differential Scanning Calorimetry (DSC) plot of the sulfate salt form B of the compound of formula (1).

FIG. 12 is a crystalline form B of the sulfate salt of the compound of formula (1) ¹ HNMR picture.

Figure 13 is an XRPD comparison of the solid before and after equilibrium solubility testing of form a of the sulfate salt of the compound of formula (1).

Figure 14 is an XRPD comparison of the stability of the sulfate form a of the compound of formula (1) before and after standing.

Figure 15 is a DVS test plot of the sulfate salt form a of the compound of formula (1).

Figure 16 is an XRPD comparison of form a of the sulphate salt of the compound of formula (1) before and after DVS testing.

Figure 17 is a XRPD comparison of the free base form a of the compound of formula (1) before and after stable standing.

Detailed Description

The intermediate compounds of the present application may be prepared by a variety of synthetic methods well known to those skilled in the art, including the specific embodiments listed below, embodiments formed by combinations thereof with other chemical synthetic methods, and equivalents thereof well known to those skilled in the art, with preferred embodiments including, but not limited to, the examples of the present application.

The chemical reactions of the embodiments herein are carried out in a suitable solvent that is compatible with the chemical changes herein and the reagents and materials required therefor. In order to obtain the compounds of the present application, it is sometimes necessary for a person skilled in the art to modify or select a synthesis procedure or a reaction scheme based on the existing embodiments.

The present application will now be described in detail by way of examples, which are not intended to limit the present application in any way.

X-ray powder diffraction (XRPD)

XRPD test parameters

Thermogravimetric analysis (TGA) and Differential Scanning Calorimetry (DSC) TGA and DSC test parameters

Nuclear magnetic hydrogen spectrum ( ¹ HNMR)

¹ HNMR test parameters

Ion Chromatography (IC)

IC test parameters

High Performance Liquid Chromatography (HPLC)

HPLC test parameters

Dynamic moisture sorption analysis (DVS)

DVS test parameters

Example 1: free crystalline form A of the compound of formula (I) (1- (3-cyano-5-methylthiophen-2-yl) -N- (6-methoxy-1H-benzo [ d ] imidazol-2-yl) -2, 5-dimethyl-1H-pyrrole-3-carboxamide) and a process for its preparation

(1) Preparation method of compound 3 (2-acetyl-4-ethyl pentanoate)

Compound 1 (ethyl acetoacetate) (5 g,38.42mmol,1.0 eq) was dissolved in triethylamine (75 mL), and compound 2 (chloroacetone) (3.5 g,38.42mmol,1.0 eq) was added. The reactants were reacted at 110 ℃ for 2 hours under nitrogen protection. After concentration, the residue was dissolved in water (100 mL) and then extracted twice with dichloromethane (50 mL each). The organic layer was washed with brine, dried over anhydrous sodium sulfate, filtered and concentrated. The residue was purified by flash column chromatography (petroleum ether/ethyl acetate =100/1 to 50/1 to 20/1) to obtain compound 3 (ethyl 2-acetyl-4-pentanoate) as a colorless oil (1.3 g, yield: 18.3%). MS (ESI) m/z:187[ m ] +H ⁺ ]。TLC：PE/EA(2/1)；R _f (compound 1) =0.6; r is _f (compound 3) =0.4.

(2) Preparation method of compound 5 (1- (3-cyano-5-methylthiophene-2-yl) -2, 5-dimethyl-1H-pyrrole-3-carboxylic acid ethyl ester)

Compound 3 (ethyl 2-acetyl-4-pentanoate) (1g, 7.23mmol, 1.0eq) was dissolved in toluene (20 mL), and compound 4 (2-amino-3-cyano-5-methylthiophene) (1.6 g,8.68mmol, 1.2eq) and p-toluenesulfonic acid (9mg, 1.45mmol, 0.2eq) were added. The reaction was stirred at 110 ℃ for 16 h. The solid was filtered and concentrated. The residue was purified by flash column chromatography (petroleum ether/ethyl acetate =50/1 to 30/1) to obtain compound 5 (1- (3-cyano-5-methylthiophen-2-yl) -2, 5-dimethyl-1H-pyrrole-3-carboxylic acid ethyl ester) as a yellow oil (860 mg, yield: 41%). MS (ESI) m/z [ alpha ], [ M ] +H ] ⁺ ]. TLC: petroleum ether/ethyl acetate (10/1); r is _f (compound 3) =0.2; r _f (compound 5) =0.4.

(3) Process for the preparation of free crystalline form A of the compound of formula (I)

To the compound 5 (1- (3-cyano-5-methylthiophen-2-yl) -2, 5-dimethyl-1H-pyrrole-3-carboxylic acid ethyl ester) (50mg, 0.306mmol, 1.0eq) and the compound 7 (5-methoxy-1H-benzimidazol-2-amine) (88mg, 0.306mmol, 1.0eq) dissolved in 1mL of toluene was added trimethylaluminum (0.15mL, 0.306mmol,1.0eq,2M dissolved in toluene). The reaction was stirred at 100 ℃ for 16 hours. The mixture was cooled to room temperature, quenched with methanol (10 mL), and then adjusted to pH 3 with 3M hydrochloric acid. The mixture was diluted with water (30 mL) and extracted three times with 20mL portions of ethyl acetate. The organic layer was washed with brine, dried over anhydrous sodium sulfate, filtered and concentrated. The residue was subjected to preparative high pressure liquid phase Prep-HPLC (acetonitrile/water (containing 0.1% formic acid)) to obtain the compound of formula (I) as a free form a (5 mg, yield: 4%). MS (ESI) m/z:406[ 2 ], [ M ] +H ⁺ ]。 ¹ H NMR(400MHz,DMSO-d ₆ ):δ12.04(s,1H),11.20(s,1H),7.32(s,1H),7.29(s,1H),6.98(s,1H),6.89(s,1H),6.69(d,J＝7.2Hz,1H),3.73(s,3H),2.47(s,3H)，2.38(s,3H)，2.04(s,3H)。

Form a in free form has an XRPD pattern substantially as shown in figure 1.

The XRPD pattern of free form a has characteristic diffraction peaks at the following 2 θ angles, as shown in table 1:

TABLE 1

As shown in fig. 2, the thermogravimetric analysis (TGA) curve of free form a lost about 1.02% of weight upon warming to about 200 ℃.

As shown in fig. 3, the Differential Scanning Calorimetry (DSC) curve of free form a has an endothermic peak at about 163 ℃.

Process for preparing a compound of formula (1) in free crystalline form ¹ The HNMR map is shown in FIG. 4.

Example 2: crystal forms of sulfate salts of compounds of formula (I) and methods for their preparation

Process for the preparation of the sulphate crystal form a of the compound of formula (I)

Weighing about 20mg of free form A into a 3mL glass vial, adding 1.2 equivalents (5.78 mg) of sulfuric acid, adding 0.5mL of acetone, ultrasonically promoting the sample to dissolve, stirring at room temperature for about 2 days, centrifuging to obtain a solid, and vacuum drying at 60 ℃ for 12 hours. And (5) separating and testing XRPD, namely obtaining the sulfate crystal form A. The method replaces acetone with isopropanol, and can also prepare the sulfate crystal form A.

The sulfate form a has an XRPD pattern substantially as shown in figure 5.

The XRPD pattern of sulfate form a has characteristic diffraction peaks at the following 2 θ angles, as shown in table 2-1:

TABLE 2-1

As shown in fig. 6, the thermogravimetric analysis (TGA) curve of the sulfate form a lost about 0.3390% of weight when warmed to about 150 ℃. The sulfate form a is anhydrous.

As shown in fig. 7, the Differential Scanning Calorimetry (DSC) curve of the sulfate form a has an endothermic peak at about 233.42 ℃.

Of the sulfate form A ¹ The H-NMR spectrum is shown in FIG. 8. It can be seen from FIG. 8 that the structure of the compound is not different from that of the free base compound.

Process for the preparation of the sulphate crystal form B of the compound of formula (I)

Weighing about 20mg of free form A into a 3mL glass vial, adding 1.2 equivalents (5.78 mg) of sulfuric acid, adding 0.5mL of acetonitrile, ultrasonically promoting the sample to dissolve, stirring at room temperature for about 2 days, centrifuging to obtain a solid, and vacuum drying at 60 ℃ for 12 hours. And (5) separating and testing XRPD, namely obtaining the sulfate crystal form B.

The sulfate form B has an XRPD pattern substantially as shown in figure 9. The XRPD pattern of sulfate form B has characteristic diffraction peaks at the following 2 theta angles, as shown in table 2-2.

Tables 2 to 2

As shown in fig. 10, the thermogravimetric analysis (TGA) curve of sulfate form B lost about 4.7791% of weight upon warming to about 140 ℃. The sulfate crystal form B is a hydrate or an anhydrate, but water is adsorbed on the surface.

As shown in fig. 11, the Differential Scanning Calorimetry (DSC) curve of form B of the sulfate salt has multiple endothermic peaks at about 31-114 ℃ and melts at 205.98 ℃.

The molar ratio of compound of formula (I) to sulfuric acid in form B of sulfate salt is about 1.

Of the crystal form B of the sulfate ¹ The H-NMR spectrum is shown in FIG. 12. The crystal form of the compound has no difference from the structure of the free base in view of nuclear magnetism.

Example 3: 24-hour equilibrium solubility in Water experiment

The free base form a and the sulfate form a were tested for 24 hours equilibrium solubility. The method comprises the following specific steps: respectively weighing 10mg of free base crystal form A and sulfate crystal form A into 1mL of water to prepare suspension, magnetically stirring at 37 +/-2 ℃, sampling after 24 hours, centrifugally separating the sample, using the obtained solid for XRPD test, and filtering the supernatant through a 0.22 mu m PTFE filter membrane for solubility test. As a result, the solubility of the free base form A was about 1.92. Mu.g/mL and the solubility of the sulfate form A was 0.56. Mu.g/mL, as shown in Table 3.

TABLE 3

Solid form	Equilibrium solubility in Water (μ g/mL) at 37 ℃
		Free base crystalline form A	1.92
Sulfates of sulfuric acidCrystal form A	0.56

XRPD comparison of solid before and after sulfate form a equilibrium solubility test, as shown in figure 13.

Example 4: one week stability test

HPLC purity and crystal form change were tested after taking free base form a and sulfate form a samples for one week at 25 ℃/60% rh (long term) and 40 ℃/75% rh (accelerated) respectively. The results are shown in table 4, after the sulfate form a was left under the two test conditions for one week, neither HPLC purity nor crystal form was significantly changed, and the sulfate form a was better in chemical stability and crystal form stability, wherein the chemical stability was better than that of the free base. The XRPD pattern of the stability samples before and after placement is shown in fig. 14.

TABLE 4

Example 5: hygroscopicity test

Evaluating the stability risk of the sample at 25 ℃ along with the change of humidity, carrying out DVS (digital visual inspection) test on the sulfate crystal form A, and collecting the tested solid sample for XRPD (X-ray diffraction pattern) test. The results are shown in Table 5.

The sulphate form a absorbs <0.5% water at 0-30% rh, the sample is slightly hygroscopic.

The sulfate form A absorbs water to 1.5-2% under 80-90% RH.

After DVS testing, no significant change occurred in the sulfate form a sample form (the difference in XRPD was primarily due to preferred orientation and crystallinity).

The DVS and XRPD test results are shown in fig. 15-16. An XRPD comparison of the free base form a of the compound of formula (1) before and after standing is shown in figure 17.

TABLE 5

Claims

1. A crystalline form of a sulfate salt of an amide compound, wherein the amide compound has a structural formula as shown in formula (1):

2. the crystalline form of claim 1 which is form a sulfate salt having an X-ray powder diffraction pattern with characteristic diffraction peaks at the following 2 Θ angles: 5.5 +/-0.2 degrees and 11.0 +/-0.2 degrees;

preferably, the X-ray powder diffraction pattern thereof also has characteristic diffraction peaks at one or more of the following 2 θ angles: 21.5 +/-0.2 degrees, 22.0 +/-0.2 degrees and 23.6 +/-0.2 degrees;

preferably, the X-ray powder diffraction pattern thereof also has characteristic diffraction peaks at one or more of the following 2 θ angles: 10.2 + -0.2 deg., 13.0 + -0.2 deg. and 18.8 + -0.2 deg..

Preferably, the sulfate form a has an XRPD pattern substantially as shown in figure 5.

3. The crystalline form of claim 1 which is form B sulfate having an X-ray powder diffraction pattern with characteristic diffraction peaks at the following 2 Θ angles: 5.5 +/-0.2 degrees, 16.1 +/-0.2 degrees and 21.0 +/-0.2 degrees;

preferably, the X-ray powder diffraction pattern thereof also has characteristic diffraction peaks at one or more of the following 2 θ angles: 11.8 +/-0.2 degrees, 25.2 +/-0.2 degrees and 26.8 +/-0.2 degrees;

preferably, the X-ray powder diffraction pattern thereof also has characteristic diffraction peaks at one or more of the following 2 θ angles: 15.0 +/-0.2 degrees, 19.2 +/-0.2 degrees and 23.8 +/-0.2 degrees;

preferably, the sulfate form B has an XRPD pattern substantially as shown in figure 9.

4. A pharmaceutical composition comprising the crystalline form of any one of claims 1-3 and a pharmaceutically acceptable carrier.

5. Use of a crystalline form according to any one of claims 1-3 or a pharmaceutical composition according to claim 4 in the manufacture of a medicament for the prevention and/or treatment of a disease or condition mediated at least in part by DHX 33; preferably, the disease is selected from the group consisting of cancer, viral infection and inflammation mediated by DHX 33.

6. A process for preparing the crystalline form a of sulfate salt of claim 2, comprising: adding an organic solvent into the free crystal form A of the compound shown in the formula (1), then adding sulfuric acid, stirring, collecting solids, and drying to obtain the sulfate crystal form A.

7. The method according to claim 6, wherein the organic solvent is selected from one or both of acetone and isopropanol.

8. A process for preparing form B of the sulfate salt of claim 3, comprising: adding an organic solvent into the free crystal form A of the compound shown in the formula (1), then adding sulfuric acid, stirring, collecting solids, and drying to obtain the sulfate crystal form B.

9. The method according to claim 8, wherein the organic solvent is selected from acetonitrile.