CN117865979A

CN117865979A - Crystal form of thiophene pyrimidine compound and preparation method thereof

Info

Publication number: CN117865979A
Application number: CN202410013431.5A
Authority: CN
Inventors: 何劼; 王堃林; 王丽; 蔡雄
Original assignee: Guangzhou Bebet Pharmaceutical Co ltd
Current assignee: Guangzhou Bebet Pharmaceutical Co ltd
Priority date: 2024-01-04
Filing date: 2024-01-04
Publication date: 2024-04-12

Abstract

The invention discloses three crystal forms of a thiophene pyrimidine compound A and a preparation method thereof, wherein the crystal form I is 3 hydrochloride, the crystal forms II and III are 2 hydrochloride, the three crystal forms have good stability and solubility, and the solubility in 5% glucose aqueous solution is more than or equal to 4mg/mL, so that the basic requirement of the prepared freeze-dried preparation for injection on the solubility of the compound is met. The three crystal forms of the thiophene pyrimidine compound A prepared by the invention have good stability and solubility, can be prepared into freeze-dried preparations for clinical injection, and are used for effectively treating lymphoma, myeloma, lymphocytic leukemia late recurrence and other or drug-resistant patients.

Description

Crystal form of thiophene pyrimidine compound and preparation method thereof

Technical Field

The invention belongs to the technical field of medicines, and particularly relates to a crystal form of a thiophene pyrimidine compound and a preparation method thereof.

Background

The incidence of hematological neoplasms is high, and about 184,720 new hematological neoplasms are available in 2023 in the united states alone. The newly increased leukemia, lymphoma and myeloma patients account for about 9.4% of the newly identified total cancers in 2023 in the united states. Wherein: leukemia 32%, lymphoma 48%, myeloma 19%. It is estimated that a total of 1,629,474 people in the united states suffer from hematological tumors or are in convalescence, and disease types include leukemia, lymphoma, myeloma, myelodysplastic syndrome, and myeloproliferative neoplasms (Leukemia and Lymphoma Society 2023). Statistics of the Chinese anticancer Association show that about 8.4 thousands of new lymphoma patients in China annually, the death number exceeds 4.7 thousands, and the rate of increase is 5% annually. In addition, the new myeloma patients in China are about 1 to 1.5 ten thousand cases and the acute and chronic lymphocytic leukemia is about 2 ten thousand cases each year. Currently, patients with advanced relapse or drug resistance of lymphomas, myelomas, and lymphoblastic leukemias lack effective drugs.

Phosphoinositide 3-kinase (PI 3K) and Histone Deacetylase (HDAC) are important targets for tumor cell survival, and HDAC inhibitors can inhibit multiple targets of tumor cell messengers through epigenetic regulation and control mechanisms. PI3K inhibitors and HDAC inhibitors have significant anticancer 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; vanhaesbroeck, B et al Nature Reviews Drug Discovery 20,741-769,2021). Several known inhibitors of phosphoinositide 3-kinase and histone deacetylase, including copanib, alpelinib, idelalisib, vorinostat, belinostat, etc. have been marketed by the FDA in the united states, but these drugs cannot inhibit phosphoinositide 3-kinase and histone deacetylase simultaneously, have poor effects on refractory or recurrent blood tumors, and cannot meet increasing clinical demands.

Thiophene pyrimidine compounds (compound A) are HDAC (histone deacetylase)/PI 3K (phosphoinositide 3-kinase) double-target inhibitors, and can damage a tumor cell messenger network by selectively inhibiting a tumor cell messenger core protein kinase target PI3K and an epigenetic target HDAC with synergistic action, so that a strong killing effect on various tumor cells is exerted. The inhibitor can strongly and effectively inhibit tumor growth in animal models of various blood and solid xenograft tumors, has obvious effect in various blood B-cell malignant tumors, and has good safety as shown by safety evaluation experiments. Can be used for effectively treating lymphoma, myeloma, and lymphocytic leukemia late recurrence or drug-resistant patients. The structural formula is as follows:

however, the compound has low solubility, and the problem that the medicine is easy to separate out when the compound is used for preparing clinical preparations for injection.

Disclosure of Invention

Based on this, the object of the present invention is to provide stable crystalline forms of thiophenopyrimidines to increase their solubility.

In order to achieve the above object, the present invention includes the following technical solutions.

In one aspect, the invention provides a crystal form I of a thiophene pyrimidine compound, which is expressed by a 2 theta angle and has characteristic peaks at 4.77 degrees, 9.52 degrees, 14.28 degrees, 21.12 degrees, 23.66 degrees, 25.21 degrees and 28.67 degrees in an X-ray powder diffraction pattern, wherein the error is +/-0.2 degrees;

the structural formula of the thiophene pyrimidine compound is as follows:

in some of these embodiments, form I of the thiophene pyrimidine compound has characteristic peaks in the X-ray powder diffraction pattern 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 terms of 2θ degrees, with an error of ±0.2 °;

in some of these embodiments, expressed in terms of 2 theta angles, the error is + -0.2 deg., the characteristic peaks and the relative intensities of the characteristic peaks in the X-ray powder diffraction pattern of form I of the thiophene pyrimidine compound are as follows:

in some of these embodiments, the crystalline form I of the thiophene pyrimidine compound has an X-ray powder diffraction pattern as shown in fig. 1-1.

In some of these embodiments, the differential scanning calorimetry curve of crystal form I of the thiophene pyrimidine compound comprises an exotherm peak at 220.9±0.5 ℃.

In some of these embodiments, the differential scanning calorimetry curve of form I of the thiophene pyrimidine compound is shown in figure 2.

In a second aspect, the present invention provides a crystalline form II of a thiophene pyrimidine compound, expressed as 2θ angles, having 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 an X-ray powder diffraction pattern, with an error of ±0.2 °;

the structural formula of the thiophene pyrimidine compound is as follows:

in some of these embodiments, the crystalline form II of the thiophene pyrimidine compound has characteristic peaks in the X-ray powder diffraction pattern 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 terms of 2θ degrees, with an error of ±0.2 °.

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

in some embodiments, the X-ray powder diffraction pattern of crystalline form II of the thiophene pyrimidine compound is shown in fig. 3.

In some of these embodiments, the differential scanning calorimetry curve of crystal form II of the thiophene pyrimidine compound comprises endothermic peaks at 79.5±0.5 ℃,141.9±0.5 ℃ and 207.7±0.5 ℃.

In some embodiments, the differential scanning calorimetry curve of crystal form II of the thiophene pyrimidine compound is shown in fig. 4.

In a third aspect, the invention provides a crystal form III of a thiophene pyrimidine compound, which is expressed by a 2 theta angle and has characteristic peaks at 10.47 degrees, 15.67 degrees, 21.23 degrees, 21.44 degrees, 22.45 degrees, 23.06 degrees, 25.73 degrees and 29.63 degrees in an X-ray powder diffraction pattern, wherein the error is +/-0.2 degrees;

the structural formula of the thiophene pyrimidine compound is as follows:

in some embodiments, the crystalline form III of the thiophene pyrimidine compound has characteristic peaks in the X-ray powder diffraction pattern 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 terms of 2θ, and an error of ±0.2 °.

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

in some embodiments, the X-ray powder diffraction pattern of crystalline form III of the thiophene pyrimidine compound is shown in fig. 5.

In some embodiments, the differential scanning calorimetry curve of crystal form III of the thiophene pyrimidine compound comprises endothermic peaks at 96.1±0.5 ℃ and 213.4±0.5 ℃.

In some embodiments, the differential scanning calorimetry curve of crystal form III of the thiophene pyrimidine compound is shown in fig. 6.

In a fourth aspect, the invention provides a preparation method of the crystal form I of the thiophene pyrimidine compound, which comprises the following steps:

uniformly stirring a mixture of methanol, a purified compound A and water, slowly adding a hydrogen chloride/methyl tertiary butyl ether solution at the temperature of 0-10 ℃, stirring for 15-25 hours at the temperature of 0-10 ℃, filtering, washing and drying to obtain a crystal form I of the thiophene pyrimidine compound;

the time for slowly adding the hydrogen chloride/methyl tertiary butyl ether solution is 2-3 hours;

the concentration of hydrogen chloride in the hydrogen chloride/methyl tertiary butyl ether solution is 4mol/L-6mol/L;

the structural formula of the compound A is as follows:

in some embodiments, the preparation method of the crystal form I of the thiophene pyrimidine compound comprises the following steps:

stirring the mixture of methanol, the purified compound A and water at 20-30 ℃ for 1.5-2.5 hours, slowly adding hydrogen chloride/methyl tertiary butyl ether solution at 4-6 ℃, stirring at 4-6 ℃ for 18-22 hours, filtering, washing and drying to obtain the crystal form I of the thiophene pyrimidine compound.

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

in some of these embodiments, the mass ratio of purified compound a to methanol is 1:11-14.

In some of these embodiments, the mass ratio of purified compound a to methanol is 1:12-13.

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

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

In some of these embodiments, the purification method of purified compound a comprises the steps of:

stirring dimethyl sulfoxide and a compound A at the temperature of 45-55 ℃ for 1.5 hours to 2.5 hours, adding methanol and a compound A seed crystal at the temperature of 45-55 ℃ for the first time, stirring for 1.5 hours to 2.5 hours, adding methanol for the second time, and stirring for 1.5 hours to 2.5 hours; cooling to 20-30 ℃, stirring for 1.5-2.5 hours, filtering, washing and drying to obtain the purified compound A.

In some of these embodiments, the purification method of the purified compound a is as follows:

stirring dimethyl sulfoxide and a compound A at 48-52 ℃ for 1.8-2.2 hours, adding methanol and a compound A seed crystal at 48-52 ℃ for the first time, stirring for 1.8-2.2 hours, adding methanol for the second time, and stirring for 1.8-2.2 hours; cooling to 22-27 ℃, stirring for 1.8-2.2 hours, filtering, washing and drying to obtain the purified compound A.

In some of these embodiments, the mass ratio of dimethyl sulfoxide to compound a is 4-9:1, preferably 5-8:1, a step of; preferably 6-7:1.

in some of these embodiments, the mass ratio of methanol first added to compound a is 1-2:1.

in some of these embodiments, the mass of compound a seed crystals added is 0.4% to 0.5% of the mass of compound a.

In some of these embodiments, the mass ratio of methanol to compound a added for the second time is 12-20:1, preferably 14-18:1, preferably 15-17:1.

in a fourth aspect, the invention provides a preparation method of a crystal form II of the thiophene pyrimidine compound, which comprises the following steps:

adding the crystal form I of the thiophene pyrimidine compound into a mixed solution of ethanol and water, stirring for 72-120 hours at the temperature of 20-30 ℃, filtering, washing and drying to obtain the crystal form II of the thiophene pyrimidine compound.

In some of these embodiments, the stirring time is 84 hours to 108 hours, preferably 90 hours to 102 hours, preferably 94 hours to 98 hours, preferably 95 hours to 97 hours.

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

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

In a sixth aspect, the invention provides a preparation method of a crystal form III of a thiophene pyrimidine compound, which comprises the following steps:

adding methanol into the crystal form I of the thiophene pyrimidine compound, stirring for 8-12 minutes at the temperature of 20-30 ℃, then stirring for 25-35 minutes at the temperature of 45-55 ℃, cooling to 0-10 ℃, preserving heat, stirring for 2-4 hours, filtering, washing and drying to obtain the crystal form III of the thiophene pyrimidine compound.

In some embodiments, the preparation method of the crystal form III of the thiophene pyrimidine compound comprises the following steps:

adding methanol into the crystal form I of the thiophene pyrimidine compound, stirring for 9-11 minutes at the temperature of 22-27 ℃, then stirring for 28-32 minutes at the temperature of 48-52 ℃, cooling to 4-6 ℃, preserving heat, stirring for 2.5-3.5 hours, filtering, washing and drying to obtain the crystal form III of the thiophene pyrimidine compound.

In some embodiments, the ratio of the crystal form I of the thiophene pyrimidine compound to methanol is 1g:16-25mL, preferably 1g:18-22mL, preferably 1g:19-21mL.

In some of these embodiments, the cooling rate to 0 ℃ to 10 ℃ is 0.08 ℃ to 0.12 ℃/min.

According to the preparation method, three crystal forms of the thiophene pyrimidine compound A are prepared through process optimization, wherein the crystal form I is 3 hydrochloride, the crystal forms II and III are 2 hydrochloride, the three crystal forms have good stability and solubility, and the solubility in a 5% glucose aqueous solution is more than or equal to 4mg/mL, so that the basic requirement of the prepared freeze-dried preparation for injection on the solubility is met. The solubility of the crystal form I and the crystal form III in 5% glucose solution is far higher than that of the crystal form II, so that the dissolution rate can be improved, the storage stability of liquid medicine before freeze-drying of the freeze-dried preparation for injection is improved, the phenomenon of drug precipitation can not occur when the liquid medicine is placed for a long time, and the freeze-dried preparation for injection is more suitable for being prepared into the freeze-dried preparation for injection and is a more optimal crystal form.

The three crystal forms of the thiophene pyrimidine compound A prepared by the invention have good stability and solubility, can be prepared into freeze-dried preparations for clinical injection, and are used for effectively treating lymphoma, myeloma, lymphocytic leukemia late recurrence and other or drug-resistant patients.

The crystal form I can be prepared from the free base of the compound A in a methanol/water/hydrogen chloride/methyl tertiary butyl ether solution system, the crystal form II can be prepared from the crystal form I by pulping in EtOH/water, and the crystal form III can be prepared from the crystal form I by slowly cooling in MeOH; the preparation process of the three crystal forms is very simple, and is suitable for industrial production.

Drawings

FIG. 1-1 is an XRPD pattern for form I of compound (I) prepared by the method of example 1.

FIGS. 1-2 are XRPD patterns of form I of compound (I) prepared by the procedure of comparative example 1, procedure 1.

FIGS. 1-3 are XRPD patterns of the compound (I) solid prepared by the procedure of comparative example 1, procedure 2.

FIG. 2 is a DSC chart of form I of compound (I) prepared by the method of example 1.

FIG. 3 is an XRPD pattern for Compound (I) form II prepared by the method of example 2.

FIG. 4 is a DSC chart of form II of compound (I) prepared by the method of example 2.

FIG. 5 is an XRPD pattern for form III of compound (I) prepared by the method of example 3.

FIG. 6 is a DSC chart of form III of compound (I) prepared by the method of example 3.

Detailed Description

The technical scheme of the invention is further described by the following specific examples. It will be apparent to those skilled in the art that the examples are merely to aid in understanding the invention and are not to be construed as a specific limitation thereof.

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. The terminology used in the description of the invention herein is for the purpose of describing particular embodiments only and is not intended to be limiting of the invention.

The terms "comprising" and "having" and any variations thereof, are intended to cover a non-exclusive inclusion. For example, a process, method, apparatus, article, or device that comprises a list of steps is not limited to the elements or modules listed but may alternatively include additional steps not listed or inherent to such process, method, article, or device.

In the present invention, the term "plurality" means two or more. "and/or", describes an association relationship of an association object, and indicates that there may be three relationships, for example, a and/or B, and may indicate: a exists alone, A and B exist together, and B exists alone. The character "/" generally indicates that the context-dependent object is an "or" relationship.

The following are specific examples.

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

Tube:Cu:K-Alpha

Generator:Voltage:40kV；Current:40mA.

Scan Scope:3to 40deg；

Sample rotation speed:15rpm.

Scanning rate:10deg/min or others。

the method used for Differential Scanning Calorimeter (DSC) is as follows:

samples in a pricked aluminum pot were tested under 50mL/min nitrogen protection with 10 ℃/min heating from 25 ℃ to 250 ℃.

The method for measuring the chloride ion content comprises the following steps:

and (3) determining the chlorine content of each crystal form sample of the compound (I) by adopting the general rule <0701> of the 2020 edition of Chinese pharmacopoeia, and calculating by using the corrected result of the weight content.

EXAMPLE 1 preparation of Compound (I) form I

The compound (I) can be prepared from a compound A, wherein the compound A is prepared according to the synthesis method in Chinese patent No. CN 104292242B.

Purification of compound a: dimethyl sulfoxide (56.8 kg) and compound A (8.7 kg) were added to the reaction vessel, the temperature of the reaction vessel was adjusted to 50℃and stirred for 2 hours until the material was completely dissolved. The solution was transferred to a crystallization kettle via a pipe filter. Methanol (13 kg) and compound A seed crystals (0.04 kg) were added to the crystallization kettle at 50℃and stirred for 2 hours, and methanol (139 kg) was added at a rate of not more than 30 kg/hour over 5 hours and stirred for 2 hours. The reaction vessel temperature was lowered to 25℃at 5℃per hour over 5 hours, stirred for 2 hours, filtered to give a solid, which was washed three times with methanol and dried to give 8.39kg of purified compound A.

Preparation of compound (I) crystalline form I: methanol (104.4 kg), the dry product of compound A (8.39 kg) obtained above and water for injection (0.9 kg) were added to a reaction vessel, and the temperature of the reaction vessel was adjusted to 25℃and stirred for 2 hours. The temperature of the reaction vessel was adjusted to 5 ℃, a hydrogen chloride/methyl tert-butyl ether solution (5 mol/L concentration, 31 kg) was slowly (about 12 kg/hr) added, and stirred at 5 ℃ for 20 hours, the solid was obtained by filtration, washed three times with methyl tert-butyl ether, and dried to obtain 9.14kg of compound (I) as a crystalline form I solid, 89.7% in molar yield, and 17% in chlorine content.

The XRPD patterns of compound (I) form I are shown in fig. 1-1, and the list of peak positions is shown in table 1; the DSC spectrum is shown in figure 2.

TABLE 1

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Comparative example 1 preparation of Compound (I) in crystalline form (crystalline form is not constant)

Operation 1: after cooling the purified suspension of Compound A (8.78 kg) in methanol (126 kg) and methylene chloride (290 kg) to 0-10deg.C, a 2.2M hydrochloric acid/methanol solution (32 kg) was added at a temperature below 10deg.C. After the addition was completed, the mixture was stirred until the solid was completely dissolved, and then filtered, and the filtrate was transferred to a clean room. Methyl tert-butyl ether (140 kg) was added dropwise at 5-10℃under stirring, and stirring was continued at this temperature for 7 hours after completion of the addition. After filtration, the resulting solid was rinsed with methyl tert-butyl ether (44 kg) to give a wet product (11.34 kg) which was dried under vacuum at 40-50 ℃ for 20 hours and then dried under vacuum at 50-60 ℃ for 30 hours until the moisture was less than or equal to 5% to give compound (I) as a pale yellow solid (10.04 kg, yield 94.1%, chlorine content 16%) whose XRPD pattern is shown in fig. 1-2 as crystalline form I.

Operation 2: the preparation of procedure 1 was repeated except for taking purified compound a (600 g) to give compound (I) (594 g, yield 78.2%, chlorine content 16%) as a pale yellow solid whose XRPD pattern is shown in fig. 1-3 as an unknown crystalline form.

The preparation method of example 1 and comparative example 1 was repeated, and impurity content and crystal form detection were performed on the batches of crystal form solids prepared therefrom. The method for detecting the impurity content comprises the following steps:

the purity, content and related substances of the compound I are measured by adopting a reversed phase High Performance Liquid Chromatography (HPLC), the content is calculated according to an external standard method, and the content of single impurities is calculated according to a main component external standard method added with correction factors.

1. Chromatographic conditions

Chromatographic column: agilent Zorbax SB-C18 (150 mm. Times.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: 250nm; flow rate: 1.5ml/min; sample injection amount: 5 μl; chromatographic column temperature: 30 ℃; sample introduction disc temperature: 15 ℃; run time: 50min.

Elution gradient:

a diluent:

diluent 1: acetonitrile-water (50:50); diluent 2: acetonitrile; diluent 3: pure water.

Washing injection: acetonitrile-water (50:50).

2. Solution preparation

(1) Diluent 1/blank solution: acetonitrile-water (50:50): 500ml of acetonitrile and 500ml of water were thoroughly mixed. The solution was labeled "BLK".

Diluent 2: acetonitrile

Diluent 3: pure water

(2) Preparation of content control solution (0.44 mg/ml Compound I solution)

Two portions were prepared in parallel, and about 22mg of compound I control was precisely weighed and placed in a 50ml volumetric flask. Firstly, 25ml of diluent 2 is added for dissolution, ultrasound is carried out for 5 minutes, after the room temperature is placed, the diluent 3 is used for fixing the volume to the scale, the mixture is evenly mixed, the ultrasound dissolution is carried out, and the room temperature is cooled. The label is a reference solution-1 (A-STD-1) and a reference solution-2 (A-STD-2).

(3) Reference substance solution (0.022 mg/ml Compound I solution)

Two portions were prepared in parallel and 5ml of the content control solution (A-STD-1) was removed to a 100ml volumetric flask. Dissolving with diluent 1, diluting to scale, and mixing. The mark is impurity reference substance solution-1 (R-STD-1) and impurity reference substance solution-2 (R-STD-2).

(4) Test solutions of the substances of interest (2.2 mg/ml Compound I solution)

Two test solutions were prepared in parallel. Precisely weighing about 55mg of the sample, placing in a 25ml volumetric flask, adding 12.5ml of diluent 2 for dissolving, performing ultrasonic treatment for 5 minutes, standing at room temperature, fixing the volume to a scale with diluent 3, mixing uniformly, performing ultrasonic dissolution, and cooling to room temperature. The labeling is the related substance sample solution-1 (R-SPL-1) and the related substance sample solution-2 (R-SPL-2).

(5) Content test sample solution (0.44 mg/ml Compound I solution)

Two sample solutions were prepared in parallel, and 5ml of the R-SPL-1 sample solution was precisely removed and placed in a 25ml volumetric flask. Dissolving with diluent 1, diluting to scale, and mixing. The labeling is content of test sample solution-1 (A-SPL-1) and content of test sample solution-2 (A-SPL-2).

3. The calculation method comprises the following steps:

the content is as follows:

the content of the test sample was calculated according to the following formula:

content (%, W/W) = (v_splx a_spl)/(w_splx RF) ×100%

Wherein:

ASPL represents the peak area of Compound I in the test sample solution

WSPL represents the sample size (mg) of Compound I in the test solution

VSPL represents the dilution volume (ml) of the test sample solution

RF represents the response factor of Compound I

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

Wherein:

ASPL represents the peak area of Compound I in the test sample solution

WSPL represents the sample size (mg) of Compound I in the test solution

VSPL represents the dilution volume (ml) of the test sample solution

RF represents the response factor of Compound I

MS represents the molecular weight of Compound I, 616.95

MF represents the molecular weight of compound a, 507.57

The weight content of the test sample (in terms of dry product) was calculated according to the following formula:

weight content (in dry product) (%, W/W) = (v_spl×a_spl)/(w_spl×rf× (100% -KF-RS)) ×100%

Wherein:

ASPL represents the peak area of Compound I in the test sample solution

WSPL represents the sample size (mg) of Compound I in the test solution

VSPL represents the dilution volume (ml) of the test sample solution

RF represents the response factor of Compound I

KF represents the moisture content (%)

RS represents total solvent residue (solvent with calculation result not less than LOQ) (%) of compound I test sample

The free base weight content of the test sample (on a dry basis) was calculated according to the following formula:

free base weight content (in dry form) (%, w/w) = (free base weight content (%, w/w))/(100% -KF-RS)) ×100%

Wherein:

ASPL represents the peak area of Compound I in the test sample solution

WSPL represents the sample size (mg) of Compound I in the test solution

VSPL represents the dilution volume (ml) of the test sample solution

RF represents the response factor of Compound I

KF represents the moisture content (%)

(1) Impurity content

Comparing the chromatogram of the sample with the blank chromatogram, integrating peaks with an area of not less than 0.02% (LOD), and calculating single impurity according to the following formula:

mono-hetero= (v_s×a_s)/(w_s×rf_ave×rrf) ×100%

Wherein:

AS represents the peak area of impurities in the sample solution of the relevant substance

WS stands for weight of test sample of the substance (mg)

RFAVE represents the mean value of the response factors of Compound I in the continuous 6-needle R-STD-1 solution

VS represents the dilution volume (ml) of the test solution of the substance of interest

RRF represents the relative response factor of each impurity, K-acid) was 1.23, K-ester was 1.16, and the other unspecified impurities were 1.00.

(2) Total impurity content

All individual impurities not less than 0.05% are summed.

(3) Purity of

Purity of compound I = 100% -total impurities

The test results are compared as shown in table 2 below: the process of comparative example 1 is that a methanol/methylene dichloride suspension of a compound A is added with a hydrochloric acid/methanol solution at 0-10 ℃, then a methyl tertiary butyl ether solution is added dropwise for crystallization, the crystal form of the compound obtained by the process is not easy to control, the crystal forms prepared from different batches are not constant, and the impurity content is higher. The improved preparation process of the example 1 is characterized in that the hydrogen chloride/methyl tertiary butyl ether solution is used for reacting and crystallizing the compound A in the presence of methanol and water, so that the impurity content can be effectively controlled, and the crystal forms of multiple batches of operation are kept consistent.

TABLE 2

EXAMPLE 2 preparation of Compound (I) form II

900mL of EtOH is added into a 2L reaction kettle, 300mL of purified water is continuously added, 120g of compound (I) crystal form I is added, stirring is carried out for 96 hours at 25 ℃, filtering is carried out, 200mL of EtOH is added to wash a filter cake, vacuum drying is carried out for 20 hours at 50 ℃, 108g of compound (I) crystal form II solid is obtained, the molar yield is 90%, and the chlorine content is 12%.

The XRPD pattern of compound (I) form II is shown in fig. 3, and the list of peak positions is shown in table 3; the DSC pattern is shown in FIG. 4.

TABLE 3 Table 3

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EXAMPLE 3 preparation of Compound (I) form III

10g of the crystalline form I of the compound (I), 200ml of MeOH, stirring at 25 ℃ for 10 minutes, then stirring at 50 ℃ for 30 minutes, slowly cooling from 50 ℃ to 5 ℃ at 0.1 ℃/minute, maintaining the temperature and stirring for 3 hours, filtering, adding 20 mM of MeOH to rinse the filter cake, and drying in vacuo at 50 ℃ for 20 hours to obtain 9.2g of the crystalline form III of the compound (I) with a molar yield of 92% and a chlorine content of 12%.

The XRPD pattern of compound (I) form III is shown in fig. 5, and the list of peak positions is shown in table 4; the DSC spectrum is shown in FIG. 6.

TABLE 4 Table 4

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EXAMPLE 4 chloride ion content and solubility comparison of crystalline forms I, II, III

The method for measuring the solubility is as follows: weighing 5mg of the crystal form sample to be measured into a liquid phase small bottle, adding 1mL of 5% glucose solution, and supplementing a proper amount of compound solid if the compound is basically dissolved, otherwise, directly carrying out the next operation. Adding a stirrer, stirring for 24 hours at 25 ℃/700rpm, transferring the sample into a filtering centrifuge tube, centrifuging for 10min at 14000rpm, diluting for 100 times, and performing liquid phase analysis on the content of the sample and calculating to obtain the solubility. The content determination method is shown in the method for determining the content in the example 1.

The chlorine ion content is measured by adopting the general rule <0701> of the 2020 edition of Chinese pharmacopoeia to determine the chlorine content of the crystal form sample by potentiometric titration, and the chlorine content is calculated by the result of weight content correction, wherein the weight content measuring method is the measuring method of the content in the example 1.

The results are shown in Table 5 below: the crystal form I is 3 hydrochloride, the crystal forms II and III are 2 hydrochloride, the three crystal forms have solubility of more than or equal to 4mg/mL in 5% glucose aqueous solution, the basic requirement (minimum requirement is 4 mg/mL) on the solubility of the freeze-dried preparation for injection is met, if the solubility is low, the compound dissolution time in the preparation liquid preparation process is long, the medicine is separated out after being placed for a long time, even the phenomenon of incomplete dissolution is avoided, and the preparation of the freeze-dried preparation for injection is not facilitated. Among the three crystal forms, the solubility of the crystal form I and the crystal form III in 5% glucose solution is far higher than that of the crystal form II, so that the crystal form I and the crystal form III are easier to prepare freeze-dried preparation for injection, and are more optimal crystal forms.

TABLE 5

Crystal form	Chloride ion content (%)	Solubility (mg/ml)	Hydrochloride salt
				Crystal form I	17	9	3 hydrochloride salt
Crystal form II	12	4	2 hydrochloride salt
				Form III	12	7	2 hydrochloride salt

Example 5 stability of Compound (I) form I sample

Putting a compound (I) crystal form I sample into a double-layer medicinal low-density polyethylene bag, tying each layer by using a binding belt, adding a silica gel desiccant (product/desiccant: 5/1), putting into a single-layer aluminum foil bag for heat sealing, finally putting into a plastic barrel, inspecting impurity content of the sample after a certain time under different stability inspection conditions, and comparing with a 0-day sample inspection result.

The experimental results are shown in table 6 below: the compound (I) crystal form I has good stability.

TABLE 6

	k-acid impurity%	k-ester impurity%	Total impurity%
				Day 0	0.18	0.17	0.35
25 ℃/60% RH,1 month	0.20	0.17	0.37
				5 ℃ +/-3, 1 month	0.18	0.17	0.35
-20 ℃ +/-5 ℃ for 1 month	0.17	0.17	0.34

The technical features of the above-described embodiments may be arbitrarily combined, and all possible combinations of the technical features in the above-described embodiments are not described for brevity of description, however, as long as there is no contradiction between the combinations of the technical features, they should be considered as the scope of the description.

The above examples illustrate only a few embodiments of the invention, which are described in detail and are not to be construed as limiting the scope of the invention. It should be noted that it will be apparent to those skilled in the art that several variations and modifications can be made without departing from the spirit of the invention, which are all within the scope of the invention. Accordingly, the scope of protection of the present invention is to be determined by the appended claims.

Claims

1. A crystalline form I of a thiophene pyrimidine compound, characterized by having 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 terms of 2θ, with an error of ±0.2 °;

the structural formula of the thiophene pyrimidine compound is as follows:

2. form I of the thiophene pyrimidine according to claim 1, characterized in that it has characteristic peaks in the X-ray powder diffraction pattern expressed in terms of 2Θ angles 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 ° with an error of ±0.2 °.

3. Form I of the thiophenopyrimidine compound of claim 2, wherein the error is ± 0.2 ° in terms of 2Θ angle, the characteristic peaks in the X-ray powder diffractogram and the relative intensities of the characteristic peaks are as follows:

4. form I of the thiophenopyrimidines according to claim 1, wherein the X-ray powder diffraction pattern is as shown in fig. 1-1.

5. Form I of the thiophenopyrimidines according to any one of claims 1 to 4, wherein the differential scanning calorimetry curve comprises an exotherm peak at 220.9 ± 0.5 ℃.

6. Form I of the thiophene pyrimidine compound according to claim 5, wherein the differential scanning calorimetry curve is shown in fig. 2.

7. A crystalline form II of a thiophene pyrimidine compound, characterized by having characteristic peaks in the X-ray powder diffraction pattern at 5.57 °, 19.28 °, 21.85 °, 22.51 °, 23.02 °, 24.10 °, 24.88 °, 26.17 °, 27.98 ° and 29.50 ° in terms of 2θ, with an error of ±0.2 °;

the structural formula of the thiophene pyrimidine compound is as follows:

8. the crystalline form II of the thiophenopyrimidine compound of claim 7, wherein said crystalline form II 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 an X-ray powder diffractogram, as expressed in terms of 2Θ angle, and an error of ± 0.2 °.

9. The crystalline form II of the thiophenopyrimidine compound of claim 8, wherein the error is ± 0.2 ° in terms of 2Θ angle, and the characteristic peaks and the relative intensities of the characteristic peaks in the X-ray powder diffraction pattern are as follows:

10. form II of the thiophenopyrimidines according to claim 7, wherein the X-ray powder diffraction pattern is as shown in figure 3.

11. Form II of the thiophene pyrimidine compound according to any one of claims 7 to 10, wherein its differential scanning calorimetry curve comprises endothermic peaks at 79.5±0.5 ℃,141.9±0.5 ℃ and 207.7±0.5 ℃.

12. The crystalline form II of the thiophene pyrimidine compound according to claim 11, wherein the differential scanning calorimetry curve is shown in fig. 4.

13. A crystalline form III of a thiophene pyrimidine compound, characterized by having characteristic peaks at 10.47 °, 15.67 °, 21.23 °, 21.44 °, 22.45 °, 23.06 °, 25.73 °, 29.63 ° in an X-ray powder diffraction pattern, expressed as 2θ, with an error of ±0.2 °;

the structural formula of the thiophene pyrimidine compound is as follows:

14. the crystalline form III of the thiophene pyrimidine compound according to claim 13, characterized by characteristic peaks in the X-ray powder diffraction pattern 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 terms of 2Θ angles, with an error of ± 0.2 °.

15. The crystalline form III of the thiophene pyrimidine compound according to claim 14, wherein the error is ± 0.2 ° in terms of 2Θ angle, and the characteristic peaks and the relative intensities of the characteristic peaks in the X-ray powder diffraction pattern are as follows:

16. the crystalline form III of the thiophene pyrimidine compound according to claim 13, wherein the X-ray powder diffraction pattern thereof is shown in fig. 5.

17. Form III of thiophenopyrimidine compound according to any of claims 13 to 16, wherein its differential scanning calorimetry curve comprises endothermic peaks at 96.1 ± 0.5 ℃ and 213.4 ± 0.5 ℃.

18. The crystalline form III of the thiophene pyrimidine compound according to claim 17, wherein the differential scanning calorimetry curve is shown in fig. 6.

19. A process for the preparation of crystalline form I of a thiophene pyrimidine compound according to any one of claims 1 to 6, comprising the steps of:

the structural formula of the compound A is as follows:

20. the method for preparing the crystal form I of the thiophene pyrimidine compound according to claim 19, comprising the following steps:

21. The method for preparing the crystal form I of the thiophene pyrimidine compound according to claim 19 or 20, wherein the mass ratio of the methanol to the water is 100-130:1, preferably 105-125:1, a step of; preferably 110-120:1, a step of; preferably 114-118:1, a step of; and/or the number of the groups of groups,

the mass ratio of the purified compound A to the methanol is 1:11-14, preferably 1:12-13; and/or the number of the groups of groups,

the mass ratio of the purified compound A to the hydrogen chloride/methyl tertiary butyl ether solution is 1:2-5, preferably 1:3-4.

22. The process for the preparation of crystalline form I of a thiophene pyrimidine compound according to claim 19 or 20, wherein the purification process of the purified compound a comprises the steps of:

23. The preparation method of the crystal form I of the thiophene pyrimidine compound according to claim 22, wherein the mass ratio of the dimethyl sulfoxide to the compound A is 4-9:1, preferably 5-8:1, a step of; preferably 6-7:1, a step of; and/or the number of the groups of groups,

the mass ratio of the methanol added for the first time to the compound A is 1-2:1, a step of; and/or

The mass of the seed crystal added into the compound A is 0.4-0.5% of the mass of the compound A; and/or the number of the groups of groups,

the mass ratio of the methanol added for the second time to the compound A is 12-20:1, preferably 14-18:1, preferably 15-17:1.

24. a process for the preparation of crystalline form II of a thiophene pyrimidine compound according to any one of claims 7 to 12, comprising the steps of:

adding a crystal form I of a thiophene pyrimidine compound into a mixed solution of ethanol and water, stirring at a temperature of 20-30 ℃ for 72-120 hours, filtering, washing and drying to obtain a crystal form II of the thiophene pyrimidine compound;

the crystal form I of the thiophene pyrimidine compound is the crystal form I of the thiophene pyrimidine compound according to any one of claims 1 to 6.

25. The process for the preparation of crystalline form II of a thiophene pyrimidine compound according to claim 24, wherein the stirring time is 84 hours to 108 hours, preferably 90 hours to 102 hours, preferably 94 hours to 98 hours, preferably 95 hours to 97 hours; and/or the number of the groups of groups,

the volume ratio of the ethanol to the water is 2-4:1, preferably 2.5-3.5:1, a step of; and/or the number of the groups of groups,

the ratio of the mixed solution of ethanol and water to the crystal form I of the thiophene pyrimidine compound is 8-12mL:1g, preferably 9-11mL:1g.

26. A process for the preparation of crystalline form III of a thiophene pyrimidine compound according to any one of claims 13 to 18, comprising the steps of:

adding methanol into a crystal form I of a thiophene pyrimidine compound, stirring for 8-12 minutes at the temperature of 20-30 ℃, then stirring for 25-35 minutes at the temperature of 45-55 ℃, cooling to 0-10 ℃, preserving heat, stirring for 2-4 hours, filtering, washing and drying to obtain a crystal form III of the thiophene pyrimidine compound;

27. The method for preparing the crystal form III of the thiophene pyrimidine compound according to claim 26, wherein the ratio of the crystal form I of the thiophene pyrimidine compound to methanol is 1g:16-25mL, preferably 1g:18-22mL, preferably 1g:19-21mL; and/or the number of the groups of groups,

the cooling speed of cooling to 0-10 ℃ is 0.08-0.12 ℃/min.