Crystal forms of thienopyrimidine compound and preparation method therefor

Abstract

Disclosed in the present invention are three crystal forms of thienopyrimidine compound A and a preparation method therefor, wherein crystal form I is of a trihydrochloride salt, and crystal forms II and III are of dihydrochloride salts. The three crystal forms all have good stability and solubility, with a solubility of ≥4 mg / mL in a 5% aqueous glucose solution, so that the basic requirements for the solubility of a compound to be prepared into a lyophilized preparation for injection are met. The three crystal forms of thienopyrimidine compound A prepared in the present invention have good stability and solubility, can be prepared into a lyophilized preparation for clinical injection, and can be used for the effective treatment of patients with advanced, recurrent, etc. lymphoma, myeloma and lymphocytic leukemia, or drug-resistant patients.

Description

Crystal form of thiophene pyrimidine compound and preparation method thereof

[0001] The present invention claims priority to Chinese patent application No. 2024100134315, filed with the Patent Office of China on January 4, 2024, entitled “Crystal Form of Thiophene Pyrimidine Compounds and Preparation Method Thereof,” the entire contents of which are incorporated herein by reference. Technical Field

[0002] The present invention belongs to the field of pharmaceutical technology, and particularly relates to a crystal form of a thiophene pyrimidine compound and a preparation method thereof. Background Art

[0003] The incidence of hematologic malignancies is high. In the United States alone, an estimated 184,720 new cases of hematologic malignancies will be diagnosed in 2023. New cases of leukemia, lymphoma, and myeloma will account for approximately 9.4% of all new cancer diagnoses in the United States in 2023. Leukemia accounts for 32%, lymphoma for 48%, and myeloma for 19%. An estimated 1,629,474 people in the United States are living with or recovering from hematologic malignancies, including leukemia, lymphoma, myeloma, myelodysplastic syndrome, and myeloproliferative neoplasms (Leukemia and Lymphoma Society 2023). Statistics from the China Anti-Cancer Association show that approximately 84,000 new cases of lymphoma and over 47,000 deaths occur in my country each year, with an annual increase of 5%. Additionally, approximately 10,000 to 15,000 new cases of myeloma and over 20,000 new cases of acute and chronic lymphocytic leukemia are diagnosed in my country each year. Currently, there is a lack of effective treatments for patients with late-stage, relapsed, or drug-resistant lymphomas, myeloma, and lymphocytic leukemia.

[0004] Phosphoinositide 3-kinase (PI3K) and histone deacetylase (HDAC) are important targets for tumor cell survival. HDAC inhibitors inhibit multiple tumor cell messenger targets through epigenetic regulation. PI3K and HDAC inhibitors have significant anti-cancer effects and have been clinically validated (Ho, T et al., Journal of Medicinal Chemistry 63, 12460-12484, 2020; Zhang, M et al., Chemical Science 11, 5855-5865, 2020; Vanhaesebroeck, B et al., Nature Reviews Drug Discovery 20, 741-769, 2021). Several known phosphoinositide 3-kinase and histone deacetylase inhibitors, including Copanisib, Alpelisib, Idelalisib, Vorinostat, Belinostat, etc., have been approved by the US FDA for marketing. However, these drugs cannot inhibit phosphoinositide 3-kinase and histone deacetylase simultaneously, and have poor effects on refractory or recurrent blood tumors, and cannot meet the increasing clinical needs.

[0005] Thiophene pyrimidine compounds (Compound A) are dual-target inhibitors of HDAC (histone deacetylase) / PI3K (phosphoinositide 3-kinase). By selectively inhibiting the synergistic tumor cell messenger core protein kinase target PI3K and the epigenetic target HDAC, they disrupt the tumor cell messenger network, thereby exerting a powerful killing effect on various tumor cells. This inhibitor can effectively inhibit tumor growth in multiple blood and solid xenograft tumor animal models, with particularly significant effects in various hematological B-cell malignancies. Safety evaluation experiments have also shown a good safety profile. It can be used to effectively treat patients with late-stage relapse or drug-resistant lymphomas, myeloma, and lymphocytic leukemia. Its structural formula is as follows:

[0006] However, the solubility of this compound is low, and the drug may easily precipitate when it is prepared into a clinical preparation for injection. Summary of the Invention

[0007] Based on this, the object of the present invention is to provide a stable crystal form of a thiophene pyrimidine compound to improve its solubility.

[0008] In order to achieve the above-mentioned object of the invention, the present invention includes the following technical solutions.

[0009] In a first aspect, the present invention provides a crystalline form I of a thiophene pyrimidine compound, which has characteristic peaks at 4.77°, 9.52°, 14.28°, 21.12°, 23.66°, 25.21°, and 28.67° in an X-ray powder diffraction pattern expressed in 2θ angles, with an error of ±0.2°;

[0010] The structural formula of the thiophene pyrimidine compound is as follows:

[0011] In some embodiments, the crystalline form I of the thiophene pyrimidine compound has characteristic peaks at 4.77°, 7.06°, 7.32°, 9.52°, 10.58°, 14.28°, 18.45°, 18.84°, 20.86°, 21.12°, 21.70°, 22.40°, 23.66°, 25.21°, 25.92°, 26.88°, 28.17°, 28.67°, 28.99°, 29.78°, 30.49° and 32.87° in the X-ray powder diffraction pattern, with an error of ±0.2°;

[0012] In some of the embodiments, the characteristic peaks of the crystalline form I of the thiophene pyrimidine compound in the X-ray powder diffraction pattern and the relative intensities of the characteristic peaks are as follows, expressed in 2θ angles with an error of ±0.2°:

[0013] In some embodiments, the X-ray powder diffraction pattern of the crystalline form I of the thiophene pyrimidine compound is shown in Figure 1-1.

[0014] In some embodiments, the differential scanning calorimetry curve of the crystalline form I of the thiophene pyrimidine compound includes an exothermic peak at 220.9±0.5°C.

[0015] In some of the embodiments, the differential scanning calorimetry curve of the crystalline form I of the thiophene pyrimidine compound is shown in FIG2 .

[0016] In a second aspect, the present invention provides a crystalline form II of a thiophene pyrimidine compound, which has characteristic peaks at 5.57°, 19.28°, 21.85°, 22.51°, 23.02°, 24.10°, 24.88°, 26.17°, 27.98°, and 29.50° in its X-ray powder diffraction pattern, expressed in 2θ angles, with an error of ±0.2°;

[0017] The structural formula of the thiophene pyrimidine compound is as follows:

[0018] In some embodiments, expressed in 2θ angles, the crystalline form II of the thiophene pyrimidine compound has characteristic peaks at 5.57°, 10.73°, 12.33°, 14.77°, 17.53°, 19.28°, 21.29°, 21.85°, 22.51°, 23.02°, 23.69°, 24.10°, 24.88°, 25.48°, 26.17°, 27.68°, 27.98°, 29.50° and 29.93° in the X-ray powder diffraction pattern, with an error of ±0.2°.

[0019] In some of the embodiments, the characteristic peaks of the crystalline form II of the thiophene pyrimidine compound in the X-ray powder diffraction pattern and the relative intensities of the characteristic peaks are as follows, expressed in 2θ angles with an error of ±0.2°:

[0020] In some of the embodiments, the X-ray powder diffraction pattern of the crystal form II of the thiophene pyrimidine compound is shown in FIG3 .

[0021] In some embodiments, the differential scanning calorimetry curve of the crystalline form II of the thiophene pyrimidine compound includes endothermic peaks at 79.5±0.5°C, 141.9±0.5°C and 207.7±0.5°C.

[0022] In some of the embodiments, the differential scanning calorimetry curve of the crystal form II of the thiophene pyrimidine compound is shown in FIG4 .

[0023] In a third aspect, the present invention provides a crystalline form III of a thiophene pyrimidine compound, which has characteristic peaks at 10.47°, 15.67°, 21.23°, 21.44°, 22.45°, 23.06°, 25.73°, and 29.63° in its X-ray powder diffraction pattern, expressed in 2θ angles, with an error of ±0.2°;

[0024] The structural formula of the thiophene pyrimidine compound is as follows:

[0025] In some embodiments, expressed in 2θ angles, the crystalline form III of the thiophene pyrimidine compound has characteristic peaks at 10.47°, 14.72°, 15.28°, 15.67°, 16.95°, 18.79°, 20.55°, 21.23°, 21.44°, 22.45°, 22.71°, 23.06°, 24.73°, 25.73°, 27.77°, 27.99°, 29.63°, 30.14° and 31.25° in the X-ray powder diffraction pattern, with an error of ±0.2°.

[0026] In some of the embodiments, the characteristic peaks of the crystalline form III of the thiophene pyrimidine compound in the X-ray powder diffraction pattern and the relative intensities of the characteristic peaks are as follows, expressed in 2θ angles with an error of ±0.2°:

[0027] In some of the embodiments, the X-ray powder diffraction pattern of the crystal form III of the thiophene pyrimidine compound is shown in FIG5 .

[0028] In some embodiments, the differential scanning calorimetry curve of the crystalline form III of the thiophene pyrimidine compound includes endothermic peaks at 96.1±0.5°C and 213.4±0.5°C.

[0029] In some of the embodiments, the differential scanning calorimetry curve of the crystal form III of the thiophene pyrimidine compound is shown in FIG6 .

[0030] In a fourth aspect, the present invention provides a method for preparing the crystalline form I of the thiophene pyrimidine compound, comprising the following steps:

[0031] The mixture of methanol, the purified compound A and water is stirred uniformly, and the hydrogen chloride / methyl tert-butyl ether solution is slowly added at a temperature of 0°C-10°C, followed by stirring at a temperature of 0°C-10°C for 15 hours-25 hours, filtering, washing, and drying to obtain the crystalline form I of the thiophene pyrimidine compound;

[0032] The time for slowly adding the hydrogen chloride / methyl tert-butyl ether solution is 2 hours to 3 hours;

[0033] The concentration of hydrogen chloride in the hydrogen chloride / methyl tert-butyl ether solution is 4 mol / L-6 mol / L;

[0034] The structural formula of the compound A is as follows:

[0035] In some embodiments, the method for preparing the crystalline form I of the thiophene pyrimidine compound comprises the following steps:

[0036] A mixture of methanol, purified compound A and water is stirred at 20°C-30°C for 1.5 hours to 2.5 hours, and a hydrogen chloride / methyl tert-butyl ether solution is slowly added at a temperature of 4°C-6°C, followed by stirring at a temperature of 4°C-6°C for 18 hours to 22 hours, filtering, washing, and drying to obtain the crystalline form I of the thiophene pyrimidine compound.

[0037] In some embodiments, the mass ratio of methanol to water is 100-130:1, preferably 105-125:1; preferably 110-120:1; preferably 114-118:1.

[0038] In some embodiments, the mass ratio of the purified compound A to methanol is 1:11-14.

[0039] In some embodiments, the mass ratio of the purified compound A to methanol is 1:12-13.

[0040] In some embodiments, the mass ratio of the purified compound A to the hydrogen chloride / methyl tert-butyl ether solution is 1:2-5.

[0041] In some embodiments, the mass ratio of the purified compound A to the hydrogen chloride / methyl tert-butyl ether solution is 1:3-4.

[0042] In some embodiments, the method for purifying the purified compound A comprises the following steps:

[0043] Dimethyl sulfoxide and compound A are stirred at a temperature of 45°C-55°C for 1.5 hours-2.5 hours, methanol and compound A seed crystals are added for the first time at a temperature of 45°C-55°C, stirred for 1.5 hours-2.5 hours, methanol is added for the second time, and stirred for 1.5 hours-2.5 hours; the temperature is lowered to 20°C-30°C, stirred for 1.5 hours-2.5 hours, filtered, washed, and dried to obtain purified compound A.

[0044] In some embodiments, the purification method of the purified compound A is as follows:

[0045] Dimethyl sulfoxide and compound A are stirred at a temperature of 48°C-52°C for 1.8 hours-2.2 hours, methanol and compound A seed crystals are added for the first time at a temperature of 48°C-52°C, stirred for 1.8 hours-2.2 hours, methanol is added for the second time, and stirred for 1.8 hours-2.2 hours; the temperature is lowered to 22°C-27°C, stirred for 1.8 hours-2.2 hours, filtered, washed, and dried to obtain purified compound A.

[0046] In some embodiments, the mass ratio of dimethyl sulfoxide to compound A is 4-9:1, preferably 5-8:1; preferably 6-7:1.

[0047] In some embodiments, the mass ratio of the first added methanol to the mass of compound A is 1-2:1.

[0048] In some embodiments, the mass of the added compound A seed crystals is 0.4%-0.5% of the mass of the compound A.

[0049] In some embodiments, the mass ratio of the second added methanol to the mass ratio of compound A is 12-20:1, preferably 14-18:1, and preferably 15-17:1.

[0050] In a fifth aspect, the present invention provides a method for preparing the crystal form II of the thiophene pyrimidine compound, comprising the following steps:

[0051] The crystal form I of the thiophene pyrimidine compound is added to a mixed solution of ethanol and water, stirred at a temperature of 20° C.-30° C. for 72 hours-120 hours, filtered, washed, and dried to obtain the crystal form II of the thiophene pyrimidine compound.

[0052] In some embodiments, the stirring time is 84 hours to 108 hours, preferably 90 hours to 102 hours, preferably 94 hours to 98 hours, and preferably 95 hours to 97 hours.

[0053] In some embodiments, the volume ratio of ethanol to water is 2-4:1, preferably 2.5-3.5:1.

[0054] In some embodiments, the ratio of the mixed solution of ethanol and water to the crystalline form I of the thiophene pyrimidine compound is 8-12 mL:1 g, preferably 9-11 mL:1 g.

[0055] In a sixth aspect, the present invention provides a method for preparing the crystalline form III of the thiophene pyrimidine compound, comprising the following steps:

[0056] Methanol is added to the crystalline form I of the thiophene pyrimidine compound, stirred at a temperature of 20°C-30°C for 8 minutes-12 minutes, then stirred at a temperature of 45°C-55°C for 25 minutes-35 minutes, cooled to 0°C-10°C, kept stirring for 2 hours-4 hours, filtered, washed, and dried to obtain the crystalline form III of the thiophene pyrimidine compound.

[0057] In some embodiments, the method for preparing the crystalline form III of the thiophene pyrimidine compound comprises the following steps:

[0058] Methanol is added to the crystalline form I of the thiophene pyrimidine compound, stirred at a temperature of 22°C-27°C for 9 minutes-11 minutes, then stirred at a temperature of 48°C-52°C for 28 minutes-32 minutes, cooled to 4°C-6°C, kept stirring for 2.5 hours-3.5 hours, filtered, washed, and dried to obtain the crystalline form III of the thiophene pyrimidine compound.

[0059] In some embodiments, the ratio of the crystalline form I of the thiophene pyrimidine compound and methanol is 1 g:16-25 mL, preferably 1 g:18-22 mL, and preferably 1 g:19-21 mL.

[0060] In some embodiments, the cooling rate to 0°C-10°C is 0.08-0.12°C / min.

[0061] The present invention prepares three crystal forms of thiophene pyrimidine compound A through process optimization, wherein crystal form I is a trihydrochloride salt, and crystal forms II and III are dihydrochlorides. All three crystal forms have good stability and solubility, with a solubility of ≥4 mg / mL in a 5% glucose aqueous solution, meeting the basic solubility requirements for preparing lyophilized preparations for injection. Among them, the solubility of crystal forms I and III in a 5% glucose solution is much higher than that of crystal form II, which can increase the dissolution rate and improve the storage stability of the lyophilized preparation for injection before lyophilization. The drug solution will not precipitate drug after long-term storage, making it more suitable for preparation of lyophilized preparations for injection and a more optimal crystal form.

[0062] The three crystal forms of the thiophene pyrimidine compound A prepared by the present invention have good stability and solubility, and can be prepared into freeze-dried preparations for clinical injection for the effective treatment of patients with late-stage relapse or drug resistance of lymphoma, myeloma and lymphatic leukemia.

[0063] The crystalline form I of the present invention can be prepared from the free base of compound A in a methanol / water / hydrogen chloride / methyl tert-butyl ether solution system, the crystalline form II can be prepared from the crystalline form I by slurrying in EtOH / water, and the crystalline form III can be prepared from the crystalline form I by slowly cooling in MeOH; the preparation processes of the three crystalline forms are simple and suitable for industrial production. BRIEF DESCRIPTION OF THE DRAWINGS

[0064] Figure 1-1 is the XRPD pattern of Form I of Compound (I) prepared by the method of Example 1.

[0065] Figure 1-2 is the XRPD spectrum of Form I of Compound (I) prepared by the method of Operation 1 in Comparative Example 1.

[0066] Figure 1-3 is the XRPD spectrum of the solid of compound (I) prepared by method 2 of comparative example 1.

[0067] FIG2 is a DSC spectrum of Compound (I) Form I prepared by the method of Example 1.

[0068] Figure 3 is the XRPD pattern of Form II of Compound (I) prepared by the method of Example 2.

[0069] FIG4 is a DSC spectrum of Form II of Compound (I) prepared by the method of Example 2.

[0070] Figure 5 is the XRPD pattern of Form III of Compound (I) prepared by the method of Example 3.

[0071] FIG6 is a DSC spectrum of Form III of Compound (I) prepared by the method of Example 3. DETAILED DESCRIPTION

[0072] The technical solutions of the present invention are further illustrated by specific examples below. Those skilled in the art should understand that the examples are only for helping to understand the present invention and should not be regarded as specific limitations of the present invention.

[0073] Unless otherwise defined, all technical and scientific terms used in the present invention have the same meaning as those commonly understood by those skilled in the art. The terms used in the specification of the present invention are only for the purpose of describing specific embodiments and are not intended to limit the present invention.

[0074] The terms "comprise," "comprising," and "having," and any variations thereof, are intended to cover non-exclusive inclusions. For example, a process, method, apparatus, product, or device comprising a series of steps is not limited to the listed steps or modules but may optionally include steps not listed, or other steps inherent to the process, method, product, or device.

[0075] In this application, "plurality" refers to two or more. "And / or" describes the relationship between related objects, indicating that three possible relationships exist. For example, "A and / or B" can mean: A exists alone, A and B exist simultaneously, or B exists alone. The character " / " generally indicates that the related objects are in an "or" relationship.

[0076] The following are specific examples.

[0077] In the following examples, the operating parameters of the X-ray powder diffraction (XRPD) analysis were set as follows:

[0078] Tube:Cu:K-Alpha

[0079] Generator: Voltage: 40kV; Current: 40mA.

[0080] Scan Scope: 3 to 40deg;

[0081] Sample rotation speed:15rpm.

[0082] Scanningrate:10deg / min or others.

[0083] The differential scanning calorimetry (DSC) method used was as follows:

[0084] The samples were tested in perforated aluminum pans under a 50 mL / min nitrogen atmosphere and heated at 10°C / min from 25°C to 250°C.

[0085] The method for determining the chloride ion content is as follows:

[0086] Adopting the general rules of the Chinese Pharmacopoeia 2020 edition <0701> The chlorine content of each crystalline form sample of Compound (I) was determined by potentiometric titration and the chlorine content was calculated based on the results corrected by weight content.

[0087] Example 1. Preparation of Compound (I) Crystalline Form I

[0088] Compound (I) can be prepared from compound A, which is prepared according to the synthesis method in Chinese invention patent CN104292242B.

[0089] Purification of Compound A: Dimethyl sulfoxide (56.8 kg) and Compound A (8.7 kg) were added to a reactor, the reactor temperature was adjusted to 50°C and stirred for 2 hours until the materials were completely dissolved. The solution was transferred to a crystallization kettle through a pipeline filter. Methanol (13 kg) and Compound A seed crystals (0.04 kg) were added to the crystallization kettle at 50°C and stirred for 2 hours. Methanol (139 kg) was added at a rate of no more than 30 kg / hour over 5 hours and stirred for 2 hours. The reactor temperature was cooled to 25°C at a rate of 5°C / hour over 5 hours, stirred for 2 hours, and filtered to obtain a solid. The solid was washed three times with methanol and dried to obtain 8.39 kg of purified Compound A.

[0090] Preparation of Compound (I) Form I: Methanol (104.4 kg), the dry product of Compound A obtained above (8.39 kg), and water for injection (0.9 kg) were added to a reactor, the reactor temperature was adjusted to 25°C, and the mixture was stirred for 2 hours. The reactor temperature was adjusted to 5°C, and a hydrogen chloride / methyl tert-butyl ether solution (5 mol / L concentration, 31 kg) was slowly added (approximately 12 kg / hour) and stirred at 5°C for 20 hours. The solid was filtered to obtain a solid, which was washed three times with methyl tert-butyl ether and dried to obtain 9.14 kg of Compound (I) Form I solid, with a molar yield of 89.7% and a chlorine content of 17%.

[0091] The XRPD pattern of Compound (I) Form I is shown in Figure 1-1, and its peak positions are listed in Table 1; the DSC pattern is shown in Figure 2.

[0092] Table 1

[0093] Comparative Example 1 Preparation of the Crystalline Form of Compound (I) (Crystalline Form is Not Constant)

[0094] Procedure 1: After cooling a suspension of purified Compound A (8.78 kg) in methanol (126 kg) and dichloromethane (290 kg) to 0-10°C, add 2.2 M hydrochloric acid / methanol solution (32 kg) at below 10°C. Stir until the solid is completely dissolved, filter, and transfer the filtrate to a clean room. Methyl tert-butyl ether (140 kg) is added dropwise at 5-10°C with stirring. Stirring is continued at this temperature for 7 hours. After filtration, the resulting solid is rinsed with methyl tert-butyl ether (44 kg) to obtain a wet product (11.34 kg). This wet product is vacuum-dried at 40-50°C for 20 hours, then at 50-60°C for 30 hours until the moisture content is ≤5%, yielding a pale yellow solid Compound (I) (10.04 kg, 94.1% yield, 16% chloride content). The XRPD pattern of the obtained product is shown in Figures 1-2, indicating Form I.

[0095] Operation 2: The purified compound A (600 g) was taken and the preparation method of operation 1 was repeated to obtain a light yellow solid compound (I) (594 g, yield 78.2%, chlorine content 16%). Its XRPD spectrum is shown in Figures 1-3, which is an unknown crystal form.

[0096] Repeat the preparation method of Example 1 and Comparative Example 1, and perform impurity content and crystal form detection on multiple batches of crystalline solids prepared therefrom. The impurity content detection method is as follows:

[0097] The purity, content and related substances of Compound I were determined by reversed-phase high performance liquid chromatography (HPLC), and the content was calculated by the external standard method. The content of individual impurities was calculated by the main component external standard method with the addition of a correction factor.

[0098] 1. Chromatographic conditions

[0099] Chromatographic column: Agilent Zorbax SB-C18 (150 mm × 4.6 mm, 3.5 μm); mobile phase A: trifluoroacetic acid-water (0.05:100), mobile phase B: trifluoroacetic acid-acetonitrile (0.05:100); detection wavelength: 250 nm; flow rate: 1.5 ml / min; injection volume: 5 μl; column temperature: 30°C; injection plate temperature: 15°C; run time: 50 min.

[0100] Elution gradient:

[0101] Diluent:

[0102] Diluent 1: acetonitrile-water (50:50); Diluent 2: acetonitrile; Diluent 3: pure water.

[0103] Injection washing agent: acetonitrile-water (50:50).

[0104] 2. Solution Preparation

[0105] (1) Diluent 1 / Blank Solution: Acetonitrile-Water (50:50): Thoroughly mix 500 ml of acetonitrile and 500 ml of water. Label this solution "BLK."

[0106] Diluent 2: Acetonitrile

[0107] Diluent 3: pure water

[0108] (2) Preparation of the reference solution (0.44 mg / ml Compound I solution)

[0109] Prepare two replicates: accurately weigh approximately 22 mg of Compound I reference solution into a 50 mL volumetric flask. Dissolve the solution in 25 mL of Diluent 2, sonicate for 5 minutes, and then bring to volume with Diluent 3. Mix thoroughly, sonicate, and cool to room temperature. Label these solutions as Reference Solution 1 (A-STD-1) and Reference Solution 2 (A-STD-2).

[0110] (3) Related substance reference solution (0.022 mg / ml Compound I solution)

[0111] Prepare two parallel batches. Pipette 5 ml of each Assay Reference Solution (A-STD-1) into a 100 ml volumetric flask. Dissolve and dilute to volume with Diluent 1, then mix thoroughly. Label these solutions as Impurity Reference Solution 1 (R-STD-1) and Impurity Reference Solution 2 (R-STD-2).

[0112] (4) Related substance test solution (2.2 mg / ml solution of Compound I)

[0113] Prepare two test sample solutions in parallel. Accurately weigh approximately 55 mg of the test sample into a 25 mL volumetric flask. Dissolve the sample in 12.5 mL of Diluent 2. Ultrasonicate for 5 minutes. Allow to stand at room temperature, then bring the solution to volume with Diluent 3. Mix thoroughly, ultrasonicate, and cool to room temperature. Label these solutions as Related Substance Test Solution 1 (R-SPL-1) and Related Substance Test Solution 2 (R-SPL-2).

[0114] (5) Assay solution (0.44 mg / ml Compound I solution)

[0115] Prepare two test sample solutions in parallel. Accurately pipette 5 ml of R-SPL-1 test sample solution into a 25 ml volumetric flask. Dissolve and dilute to the mark with Diluent 1, then mix thoroughly. Label the two aliquots as A-SPL-1 and A-SPL-2.

[0116] 3. Calculation method:

[0117] content:

[0118] Calculate the content of the test sample according to the following formula: Content (%, w / w) = (V_SPL × A_SPL) / (W_SPL × RF) × 100%

[0119] Where:

[0120] ASPL represents the peak area of ​​compound I in the test solution

[0121] WSPL represents the sample weight of compound I in the test solution (mg)

[0122] VSPL represents the dilution volume of the test solution (ml)

[0123] RF represents the response factor of compound I

[0124] The weight content of the free base of the test compound I was calculated according to the following formula: Weight content of free base (%, w / w) = weight content (%, w / w) × M_F / M_S

[0125] Where:

[0126] ASPL represents the peak area of ​​compound I in the test solution

[0127] WSPL represents the sample weight of compound I in the test solution (mg)

[0128] VSPL represents the dilution volume of the test solution (ml)

[0129] RF represents the response factor of compound I

[0130] MS represents the molecular weight of compound I, 616.95

[0131] MF represents the molecular weight of compound A, 507.57

[0132] Calculate the weight content of the test sample (on a dry basis) according to the following formula: Weight content (on a dry basis) (%, w / w) = (V_SPL × A_SPL) / (W_SPL × RF × (100% - KF - RS)) × 100%

[0133] Where:

[0134] ASPL represents the peak area of ​​compound I in the test solution

[0135] WSPL represents the sample weight of compound I in the test solution (mg)

[0136] VSPL represents the dilution volume of the test solution (ml)

[0137] RF represents the response factor of compound I

[0138] KF represents the water content of the test sample of compound I (%)

[0139] RS represents the total residual solvent of the test sample of Compound I (the calculated result is not less than the LOQ of the solvent) (%)

[0140] The free base content (based on dry product) of the test sample was calculated according to the following formula: Free base content (based on dry product) (%, w / w) = (free base content (%, w / w)) / ((100% - KF-RS)) × 100%

[0141] Where:

[0142] ASPL represents the peak area of ​​compound I in the test solution

[0143] WSPL represents the sample weight of compound I in the test solution (mg)

[0144] VSPL represents the dilution volume of the test solution (ml)

[0145] RF represents the response factor of compound I

[0146] KF represents the water content of the test sample of compound I (%)

[0147] RS represents the total residual solvent of the test sample of Compound I (the calculated result is not less than the LOQ of the solvent) (%)

[0148] (1) Impurity content

[0149] Compare the sample chromatogram with the blank chromatogram, integrate the peaks with an area of ​​not less than 0.02% (LOD), and calculate the single impurity according to the following formula: Single impurity = (V_S × A_S) / (W_S × RF_AVE × RRF) × 100%

[0150] Where:

[0151] AS represents the peak area of ​​the impurity in the test solution of the related substance

[0152] WS represents the weight of the test sample of the related substance (mg)

[0153] RFAVE represents the average response factor of compound I in 6 consecutive injections of R-STD-1 solution

[0154] VS represents the dilution volume of the test solution of the related substance (ml)

[0155] RRF represents the relative response factor of each impurity, which is 1.23 for K-acid, 1.16 for K-ester, and 1.00 for other unspecified impurities.

[0156] (2) Total impurity content

[0157] All individual impurities with a minimum concentration of 0.05% are summed.

[0158] (3) Purity

[0159] Purity of Compound I = 100% - total impurities

[0160] The test results are compared in Table 2 below. The process of Comparative Example 1 involves adding a hydrochloric acid / methanol solution to a methanol / dichloromethane suspension of Compound A at 0-10°C, followed by dropwise addition of a methyl tert-butyl ether solution for crystallization. The crystal form of the compound obtained using this process is difficult to control, the crystal forms obtained from different batches are inconsistent, and the impurity content is relatively high. The improved preparation process of Example 1 involves reacting Compound A with a hydrogen chloride / methyl tert-butyl ether solution in the presence of methanol and water for crystallization. This method can effectively control the impurity content and maintain consistent crystal forms across multiple batches.

[0161] Table 2

[0162] Example 2. Preparation of Compound (I) Crystalline Form II

[0163] To a 2 L reactor, 900 mL of EtOH was added, followed by 300 mL of purified water and 120 g of Compound (I) Form I. The mixture was stirred at 25° C. for 96 hours, filtered, and the filter cake was rinsed with 200 mL of EtOH. The mixture was vacuum dried at 50° C. for 20 hours to obtain 108 g of Compound (I) Form II solid with a molar yield of 90% and a chlorine content of 12%.

[0164] The XRPD pattern of Compound (I) Form II is shown in FIG3 , and the peak positions are listed in Table 3 ; the DSC pattern is shown in FIG4 .

[0165] Table 3

[0166] Example 3. Preparation of Compound (I) Crystalline Form III

[0167] To a 500 ml reactor, 10 g of Form I of Compound (I) and 200 ml of MeOH were added, and the mixture was stirred at 25° C. for 10 minutes, then at 50° C. for 30 minutes. The temperature was slowly decreased from 50° C. to 5° C. at a rate of 0.1° C. / min and the mixture was kept stirring for 3 hours. The mixture was filtered, and 20 mL of MeOH was added to rinse the filter cake. The mixture was vacuum dried at 50° C. for 20 hours to obtain 9.2 g of Form III of Compound (I) with a molar yield of 92% and a chlorine content of 12%.

[0168] The XRPD pattern of Compound (I) Form III is shown in FIG5 , and the peak positions are listed in Table 4 ; the DSC pattern is shown in FIG6 .

[0169] Table 4

[0170] Example 4. Comparison of chloride ion content and solubility of crystal forms I, II, and III

[0171] The solubility determination method is as follows: weigh 5 mg of the crystalline form sample to be tested into a liquid phase vial and add 1 mL of 5% glucose solution. If the compound is substantially dissolved, add an appropriate amount of solid compound; otherwise, proceed directly to the next step. Add a stirrer and stir at 25°C / 700 rpm for 24 hours. Transfer the sample to a filter centrifuge tube and centrifuge at 14,000 rpm for 10 minutes. After diluting 100-fold, analyze the sample content in the liquid phase and calculate the solubility. The content determination method is described in Example 1.

[0172] Chloride ion content was determined using the general rules of the Chinese Pharmacopoeia 2020 edition. <0701> The chlorine content of the crystalline sample was determined by potentiometric titration and calculated based on the result corrected by weight content. The weight content determination method is shown in the content determination method in Example 1.

[0173] The results are shown in Table 5 below: Form I is the trihydrochloride salt, and Forms II and III are the dihydrochloride salts. All three forms have a solubility of ≥4 mg / mL in a 5% glucose aqueous solution, meeting the basic solubility requirement for preparing a lyophilized injection preparation (the minimum requirement is 4 mg / mL). If the solubility is too low, it is easy to cause the compound to dissolve for a long time during the preparation process, and the drug will precipitate when the liquid is left for a long time, or even fail to dissolve completely, which is not conducive to the preparation of a lyophilized injection preparation. Among the three forms, Form I and Form III have a much higher solubility in 5% glucose solution than Form II, making it easier to prepare a lyophilized injection preparation and a more optimal form.

[0174] Table 5

[0175] Example 5. Stability of Compound (I) Form I Sample

[0176] The compound (I) crystalline form I sample was placed in a double-layer medicinal low-density polyethylene bag, each layer was tied with a cable tie, silica gel desiccant (product / desiccant: 5 / 1) was added, and the bag was placed in a single-layer aluminum foil bag and heat-sealed. Finally, the bag was placed in a plastic barrel and placed under different stability test conditions for a certain period of time. The impurity content of the sample was detected and compared with the test results of the 0-day sample.

[0177] The experimental results are shown in Table 6 below: Compound (I) Form I has good stability.

[0178] Table 6

[0179] The technical features of the above-mentioned embodiments can be combined arbitrarily. In order to make the description concise, not all possible combinations of the technical features in the above-mentioned embodiments are described. However, as long as there is no contradiction in the combination of these technical features, they should be considered to be within the scope of this specification.

[0180] The above-described embodiments merely illustrate several implementations of the present invention, and while their descriptions are relatively specific and detailed, they should not be construed as limiting the scope of the patent. It should be noted that a person skilled in the art would be able to make numerous variations and improvements without departing from the spirit of the present invention, all of which fall within the scope of protection of the present invention. Therefore, the scope of protection of the patent for this invention shall be determined by the appended claims.

Claims

1. Crystal form I of a thienopyrimidine compound, characterized in that, Expressed in terms of the 2θ angle, it has characteristic peaks at 4.77°, 9.52°, 14.28°, 21.12°, 23.66°, 25.21° and 28.67° in the X-ray powder diffraction pattern, with an error of ±0.2°. The structural formula of the said pyrimidine compounds is as follows:

2. The polymorphic form I of the thiophenopyrimidine compound according to claim 1, wherein Expressed in terms of the 2θ angle, it has characteristic peaks at 4.77°, 7.06°, 7.32°, 9.52°, 10.58°, 14.28°, 18.45°, 18.84°, 20.86°, 21.12°, 21.70°, 22.40°, 23.66°, 25.21°, 25.92°, 26.88°, 28.17°, 28.67°, 28.99°, 29.78°, 30.49° and 32.87° in the X-ray powder diffraction pattern, with an error of ±0.2°.

3. The crystalline form I of the thiophenopyrimidine compound according to claim 2, characterized in that, Expressed in terms of the 2θ angle, the error is ±0.2°, and the characteristic peaks in the X-ray powder diffraction pattern and the relative intensities of the respective characteristic peaks are as follows:

4. The polymorphic form I of the thiophenopyrimidine compound according to claim 1, characterized in that, Its X-ray powder diffraction pattern is shown in Figure 1-1.

5. The polymorphic form I of the thiophenopyrimidine compound according to any one of claims 1-4, characterized in that, Its differential scanning calorimetry curve includes an exothermic peak at 220.9 ± 0.5 °C.

6. The crystalline form I of the thiophene pyrimidine compound according to claim 5, characterized in that, Its differential scanning calorimetry curve is shown in Figure 2.

7. Polymorph II of a thiophene pyrimidine compound, characterized in that, Expressed in terms of the 2θ angle, it has characteristic peaks at 5.57°, 19.28°, 21.85°, 22.51°, 23.02°, 24.10°, 24.88°, 26.17°, 27.98° and 29.50° in the X-ray powder diffraction pattern, with an error of ±0.2°. The structural formula of the said pyrimidine thione compounds is as follows:

8. The polymorphic form II of the thiophenopyrimidine compound according to claim 7, characterized in that, Expressed in terms of the 2θ angle, it has characteristic peaks at 5.57°, 10.73°, 12.33°, 14.77°, 17.53°, 19.28°, 21.29°, 21.85°, 22.51°, 23.02°, 23.69°, 24.10°, 24.88°, 25.48°, 26.17°, 27.68°, 27.98°, 29.50° and 29.93° in the X-ray powder diffraction pattern, with an error of ±0.2°.

9. The polymorphic form II of the thiophenopyrimidine compound according to claim 8, characterized in that, Expressed in terms of the 2θ angle, the error is ±0.2°, and the characteristic peaks in the X-ray powder diffraction pattern and the relative intensities of the respective characteristic peaks are as follows:

10. The polymorphic form II of the thiophenopyrimidine compound according to claim 7, characterized in that, Its X-ray powder diffraction pattern is shown in Figure 3.

11. The polymorphic form II of the thiophenopyrimidine compound according to any one of claims 7-10, characterized in that, Its differential scanning calorimetry curve includes endothermic peaks at 79.5 ± 0.5 °C, 141.9 ± 0.5 °C and 207.7 ± 0.5 °C.

12. The crystalline form II of the thiophenopyrimidine compound according to claim 11, characterized in that, Its differential scanning calorimetry curve is shown in Figure 4.

13. Polymorph III of a thiophenopyrimidine compound, characterized in that, Expressed in terms of the 2θ angle, it has characteristic peaks at 10.47°, 15.67°, 21.23°, 21.44°, 22.45°, 23.06°, 25.73°, 29.63° in the X-ray powder diffraction pattern, with an error of ±0.2°. The structural formula of the said pyrimidine-thiophene compounds is as follows:

14. The polymorphic form III of the thiophenopyrimidine compound according to claim 13, wherein Expressed in terms of the 2θ angle, it has characteristic peaks at 10.47°, 14.72°, 15.28°, 15.67°, 16.95°, 18.79°, 20.55°, 21.23°, 21.44°, 22.45°, 22.71°, 23.06°, 24.73°, 25.73°, 27.77°, 27.99°, 29.63°, 30.14° and 31.25° in the X-ray powder diffraction pattern, with an error of ±0.2°.

15. The crystalline form III of the thiophenopyrimidine compound according to claim 14, characterized in that, Expressed in terms of the 2θ angle, the error is ±0.2°, and the characteristic peaks in the X-ray powder diffraction pattern and the relative intensities of the respective characteristic peaks are as follows:

16. The crystalline form III of the thiophenopyrimidine compound according to claim 13, characterized in that, Its X-ray powder diffraction pattern is shown in Figure 5.

17. The crystalline form III of the thiophenopyrimidine compound according to any one of claims 13-16, characterized in that, Its differential scanning calorimetry curve includes endothermic peaks at 96.1 ± 0.5 °C and 213.4 ± 0.5 °C.

18. The crystalline form III of the thiophenopyrimidine compound according to claim 17, characterized in that, Its differential scanning calorimetry curve is shown in Figure 6.

19. A method for preparing polymorphic form I of the thiophenopyrimidine compound according to any one of claims 1-6, characterized in that, It includes the following steps: Stir the mixture of methanol, purified compound A and water evenly, slowly add the hydrogen chloride / methyl tert-butyl ether solution at a temperature of 0°C - 10°C, then stir at a temperature of 0°C - 10°C for 15 - 25 hours, filter, wash, and dry to obtain the crystalline form I of the said thiophene pyrimidine compound; The time for slowly adding the hydrogen chloride / methyl tert-butyl ether solution is 2 - 3 hours; The hydrogen chloride concentration in the hydrogen chloride / methyl tert-butyl ether solution is 4 mol / L - 6 mol / L; The structural formula of the said Compound A is as follows:

20. The preparation method of polymorphic form I of the thiophenopyrimidine compound according to claim 19, characterized in that, It includes the following steps: Stir the mixture of methanol, purified compound A and water at 20°C - 30°C for 1.5 - 2.5 hours, slowly add the hydrogen chloride / methyl tert-butyl ether solution at a temperature of 4°C - 6°C, then stir at a temperature of 4°C - 6°C for 18 - 22 hours, filter, wash, and dry to obtain the crystalline form I of the said thiophene pyrimidine compound.

21. A method for preparing polymorphic form I of a thiophenopyrimidine compound according to claim 19 or 20, characterized in that, The mass ratio of methanol to water is 100 - 130:1, preferably 105 - 125:1; preferably 110 - 120:1; preferably 114 - 118:1; and / or, The mass ratio of the purified compound A to methanol is 1:11 - 14, preferably 1:12 - 13; and / or, The mass ratio of the purified compound A to the hydrogen chloride / methyl tert-butyl ether solution is 1:2 - 5, preferably 1:3 - 4.

22. The preparation method of polymorphic form I of the thiophenopyrimidine compound according to claim 19 or 20, characterized in that, The purification method of the purified compound A includes the following steps: Stir dimethyl sulfoxide and compound A at a temperature of 45°C - 55°C for 1.5 - 2.5 hours, first add methanol and compound A crystal seeds at a temperature of 45°C - 55°C, stir for 1.5 - 2.5 hours, then add methanol for the second time and stir for 1.5 - 2.5 hours; cool down to 20°C - 30°C, stir for 1.5 - 2.5 hours, filter, wash, and dry to obtain the purified compound A.

23. The preparation method of polymorphic form I of the thiophenopyrimidine compound according to claim 22, characterized in that, The mass ratio of dimethyl sulfoxide to compound A is 4 - 9:1, preferably 5 - 8:1; preferably 6 - 7:1; and / or, The mass ratio of the methanol added for the first time to compound A is 1 - 2:1; and / or The mass of the compound A crystal seeds added is 0.4% - 0.5% of the mass of the said compound A; and / or, The mass ratio of the methanol added for the second time to compound A is 12 - 20:1, preferably 14 - 18:1, preferably 15 - 17:

1.

24. A method for preparing polymorph II of the thiophenopyrimidine compound according to any one of claims 7-12, characterized in that, It includes the following steps: Add the crystalline form I of the thiophene pyrimidine compound to the mixed solution of ethanol and water, stir at a temperature of 20°C - 30°C for 72 - 120 hours, filter, wash, and dry to obtain the crystalline form II of the said thiophene pyrimidine compound; The crystalline form I of the said thiophene pyrimidine compound is the crystalline form I of the thiophene pyrimidine compound according to any one of claims 1 - 6.

25. The method for preparing polymorph II of the thiophenopyrimidine compound according to claim 24, characterized in that, The stirring time is 84 - 108 hours, preferably 90 - 102 hours, preferably 94 - 98 hours, preferably 95 - 97 hours; and / or, The volume ratio of ethanol to water is 2 - 4:1, preferably 2.5 - 3.5:1; and / or, The ratio of the mixed solution of ethanol and water to polymorph I of the thiophenopyrimidine compound is 8 - 12 mL:1 g, preferably 9 - 11 mL:1 g.

26. A method for preparing polymorph III of the thiophenopyrimidine compound according to any one of claims 13-18, characterized in that, It includes the following steps: Add methanol to polymorph I of the thiophenopyrimidine compound, stir at a temperature of 20°C - 30°C for 8 minutes - 12 minutes, then stir at a temperature of 45°C - 55°C for 25 minutes - 35 minutes, cool down to 0°C - 10°C, keep warm and stir for 2 hours - 4 hours, filter, wash, and dry to obtain polymorph III of the thiophenopyrimidine compound; The polymorph I of the thiophenopyrimidine compound is the polymorph I of the thiophenopyrimidine compound according to any one of claims 1 - 6.

27. The method for preparing the crystalline form III of the thiophenopyrimidine compound according to claim 26, wherein The ratio of polymorph I of the thiophenopyrimidine compound to methanol is 1 g:16 - 25 mL, preferably 1 g:18 - 22 mL, preferably 1 g:19 - 21 mL; and / or, The cooling rate for cooling down to 0°C - 10°C is 0.08 - 0.12 °C / minute.

Images