WO2021121146A1 - Crystal form a of aminopyrimidine mesylate compound, preparation method therefor, and application thereof - Google Patents

Abstract

Disclosed is a crystal form A of an aminopyrimidine mesylate compound represented by formula I, a preparation method therefor, and an application thereof. The crystal form A has excellent solubility, high bioavailability in animals, and can be used to treat or prevent diseases or medical conditions mediated by epidermal growth factor (EGFR) mutations (activated or drug-resistant) in mammals.

Description

Crystal form A of aminopyrimidine compound methanesulfonate and its preparation method and application Technical field

The invention belongs to the field of drug synthesis, and specifically relates to the crystal form A of aminopyrimidine compound methanesulfonate and its preparation method and application.

Background technique

Epidermal growth factor receptor (EGFR) is a type of transmembrane receptor tyrosine kinase in the human body. The activation (ie phosphorylation) of this kinase is of great significance to the inhibition of tumor cell proliferation, angiogenesis, tumor invasion, metastasis and apoptosis. EGFR kinase is involved in the disease process of most cancers, and these receptors are overexpressed in many major human tumors. Overexpression, mutations, or high expression of ligands associated with these family members can lead to some tumor diseases, such as non-small cell lung cancer, colorectal cancer, breast cancer, head and neck cancer, cervical cancer, bladder cancer, and thyroid. Cancer, stomach cancer, kidney cancer, etc.

(吉非替尼)来用于治疗晚期非小细胞肺癌(NSCLC)，掀开了第一代EGFR抑制剂的开发。众多临床试验证实，对于EGFR阳性突变非小细胞肺癌患者而言，分子靶向药物治疗效果显著优于传统化疗。 In recent years, epidermal growth factor receptor tyrosine kinase has become one of the most attractive targets in current anti-tumor drug research. In 2003, the US FDA approved the first epidermal growth receptor tyrosine kinase inhibitor (EGFR-TKI) drug

(Gefitinib) for the treatment of advanced non-small cell lung cancer (NSCLC), opening the development of the first generation of EGFR inhibitors. Numerous clinical trials have confirmed that, for patients with EGFR-positive non-small cell lung cancer, the therapeutic effect of molecular targeted drugs is significantly better than traditional chemotherapy.

Although the first-generation EGFR inhibitor targeted drugs responded well to the initial treatment of many non-small cell lung cancer (NSCLC) patients, most patients will eventually develop disease progression due to drug resistance (such as EGFR secondary T790M mutation). The emergence of drug resistance is caused by various mechanisms based on the original EGFR pathway activity mutations. In the research on resistance of the first generation of EGFR inhibitors, the research frontier is the irreversible third generation of EFGR inhibitors.

开发成功，尚无其它有效针对T790M耐药突变患者的药物获批供临床治疗使用；若干针对T790M突变的候选药物正处于临床开发阶段。这类第三代EGFR抑制剂化学结构与第一代截然不同。与第一代EGFR抑制 剂的主要不同点在于它们均使用高选择性核心结构替代第一、二代EGFR-TKI的低选择性氨基喹啉核心结构。相对于野生型EGFR，这些第三代化合物均对EGFR阳性耐药后T790M突变具有高度特异选择性。 But so far, in the third generation of EGFR inhibitors worldwide, except for AstraZeneca’s osimertinib

The development is successful, and no other drugs that effectively target patients with T790M resistance mutations have been approved for clinical treatment; several drug candidates for T790M mutations are in the clinical development stage. The chemical structure of this third-generation EGFR inhibitor is completely different from that of the first-generation. The main difference from the first-generation EGFR inhibitors is that they both use a highly selective core structure to replace the low-selectivity aminoquinoline core structure of the first and second-generation EGFR-TKIs. Compared with wild-type EGFR, these third-generation compounds are highly specific and selective for the T790M mutation after EGFR positive resistance.

Chinese Patent Application No. CN201580067776.8 discloses a compound with the following formula I, which also belongs to the third-generation EGFR-TKI class of small molecule targeted drugs. The compound has a high inhibitory effect on non-small cell lung cancer (NSCLC) cells with single active mutations and T790M double mutant EGFR, and its effective inhibitory concentration is significantly lower than the concentration required to inhibit the activity of wild-type EGFR tyrosine kinase. It has the characteristics of good sex, low side effects and good safety.

Chinese Patent Application No. CN201780050034.3 also discloses various salts and corresponding crystal forms of the compound of the above formula I structure. Example 2 thereof discloses two crystal forms of the methanesulfonate of the compound of formula I, 2A and 2B, respectively.

Summary of the invention

The purpose of the present invention is to provide a new crystal form of aminopyrimidine compound methanesulfonate and its preparation method and application. The present invention therefore relates to a substantially pure new crystalline form of the mesylate salt of the compound of formula I.

Compared with the crystals 2A and 2B disclosed in Example 2 of Chinese Patent Application No. CN201780050034.3 (CN109937043A), the crystal form of this application is different from the number, position and intensity of XPRD characteristic peaks, and the melting point and melting range are also significantly different. This application The crystal form A obtained under the preparation process has higher purity and more stable properties, which is more conducive to its application in the field of medicine.

In order to achieve the above objectives, the present invention adopts the following technical solutions:

The present invention provides the crystal form A of the methanesulfonate salt of an aminopyrimidine compound. The aminopyrimidine compound is a compound of formula I. The XRPD pattern of this crystal form is 11.06, 12.57, 13.74, 14.65 in 2θ (±0.2°), There are diffraction peaks at 15.48, 16.58, 17.83, 19.20, 19.79, 20.88, 22.05, 23.06, 24.23, 25.10, 25.71, 26.15, 27.37, 27.42 (relative intensity greater than 10%),

Preferably, the XRPD pattern of the crystal form A is shown in FIG. 3.

Preferably, the infrared spectrum of the crystal form A is shown in FIG. 6.

Preferably, the crystal form A has a melting point of 242-244°C.

Preferably, the crystalline form A has a thermal weight loss of -0.035% in the range of 33.4-120°C in a thermogravimetric analysis (TGA) measurement.

The present invention also provides a method for preparing the crystal form A of the aminopyrimidine compound methanesulfonate, which comprises: dissolving the methanesulfonate of the compound of formula I with an organic solvent, stirring and crystallization, filtering, and drying to obtain the target crystal form A, The organic solvent includes dimethyl sulfoxide, tetrahydrofuran, N-methylpyrrolidone, N,N-dimethylformamide and N,N-dimethylacetamide.

Preferably, it further comprises: preparing the methanesulfonate of the compound of formula I and methanesulfonic acid in a molar ratio of 1.05-0.97, preferably equimolar ratio.

Preferably, an anti-solvent is added for stirring and crystallization, and the anti-solvent includes methyl tert-butyl ether, isopropyl acetate, ethyl acetate, methyl isobutyl ketone, acetonitrile, toluene, acetone and water.

Preferably, normal pressure or vacuum drying is performed at 30-70°C (more preferably 40-50°C) after filtration.

The present invention also provides a pharmaceutical composition comprising the crystal form A of the aminopyrimidine compound methanesulfonate and a pharmaceutically acceptable carrier.

The present invention also provides the crystal form A of the aminopyrimidine compound methanesulfonate, and the application of the above-mentioned pharmaceutical composition or combination drug in the preparation of antineoplastic drugs.

The present invention additionally provides a method for treating or preventing diseases or medical conditions, preferably tumors, mediated by EGFR mutations in mammals, especially humans, comprising administering a therapeutically effective amount of an aminopyrimidine compound to an individual in need thereof The crystalline form A of a sulfonate or a pharmaceutical composition comprising the crystalline form A of an aminopyrimidine compound methanesulfonate and a pharmaceutically acceptable carrier.

Preferably, the tumor is lung cancer.

The beneficial effects of the present invention are as follows:

It is found through experiments that the crystal form A of the aminopyrimidine compound methanesulfonate salt of the present invention has high purity, stable properties, good solubility, and high bioavailability in animals. It can be used to treat or prevent diseases or medical conditions mediated by epidermal growth factor (EGFR) mutations (activation or drug resistance) in mammals, especially humans, especially tumors or cancers, especially non-small cell lung cancer; and can be made Pharmaceutical composition, and application in combination medicine. It can be confirmed by the experiments in this application that the crystalline form A of the aminopyrimidine compound mesylate salt of the present invention has the characteristics of the mesylate salt forms 2A and 2B disclosed in Example 2 of Chinese Patent Application No. CN201780050034.3. More favorable physical and chemical properties. Clinical trials have confirmed that compared with the marketed drug osimertinib, the crystal form A of the present invention has a higher disease control rate and disease remission rate for patients with non-small cell lung cancer, and the incidence of adverse reactions (proportion and type) Significantly lower.

Description of the drawings

Figure 1 is a ¹ H NMR spectrum of the compound of formula I.

^{Figure 2 is the 1} H NMR spectrum of the crystal form A.

Figure 3 is the XRPD spectrum of Form A and a table recording the data of each peak.

Figure 4 is a differential scanning calorimetry (DSC) spectrum of Form A.

Figure 5 is a thermogravimetric analysis (TGA) spectrum of Form A.

Fig. 6 is the infrared spectrum (IR) spectrum of crystal form A.

Fig. 7 is a comparison chart of powder X-ray diffraction patterns (XPRD) of crystal form A obtained by different preparation methods.

Figure 8 is a graph showing the blood concentration-time curve of SD rats after a single administration.

Fig. 9 is a graph showing the tumor volume change-time curve of human lung cancer H1975 subcutaneous xenografted tumor in nude mice under single and combined administration of the methanesulfonate salt of the compound of formula (I) prepared in Example 2 (ie crystal form A) .

Fig. 10 is a graph showing the body weight change-time curve of human lung cancer H1975 nude mice under single and combined administration of the methanesulfonate salt of the compound of formula (I) prepared in Example 2 (ie, crystal form A).

Detailed ways

The following examples are used to further describe the beneficial effects of the present invention. The examples are only for illustrative purposes and do not limit the scope of the present invention. At the same time, obvious changes and modifications made by those skilled in the art according to the present invention also include Within the scope of the present invention.

In the following examples, the "room temperature" can be 15-25°C.

(1) N-(2-(2-(Dimethylamino)ethoxy)-4-methoxy-5-((4-(1-methyl-1H-indol-3-yl)pyrimidine -2-yl)amino)phenyl)acrylamide (compound of formula I)

Known (for example, see CN201580067776.8) N-(2-(2-(dimethylamino)ethoxy)-4-methoxy-5-((4-(1-methyl-1H- Indol-3-yl)pyrimidin-2-yl)amino)phenyl)acrylamide (compound of formula I) can be prepared by the following synthetic route:

Step 1-Preparation of Intermediate J:

Preparation: In a 10L reaction flask, add 6L of anhydrous tetrahydrofuran solvent, protected by nitrogen, and cool to 0°C. While stirring, slowly add 101 g of sodium hydride (101 g, 2.52 mol), and the internal temperature does not exceed 10°C, and add 234 g of dimethylaminoethanol (234 g, 2.62 mol). After the addition, the temperature is adjusted to room temperature to prepare a sodium alkoxide solution.

In a 30L reaction flask, add N-(4-fluoro-2-methoxy-5-nitrophenyl)-4-(1-methyl-1H-indol-3-yl)-2-pyrimidinamine ( Starting material B) (430g, 1.10mol), then add 9L of tetrahydrofuran, turn on the stirring, dissolve it, control the temperature at 10±10°C, and slowly add the prepared sodium alkoxide solution dropwise. Control the temperature at 10±10℃ and keep it for 5.0h. When the raw material content is ≤0.5%, the reaction ends. Control the temperature at 10±10°C, slowly add 3% hydrochloric acid solution dropwise, adjust the pH of the solution to 6-7, stir for 1.5h and then stand to separate the layers, separate the organic phase, and concentrate to 15-20L. After cooling to 20±5°C, 4.3 kg of water was slowly added dropwise, filtered, and dried to obtain 497 g of yellow powder intermediate J with a yield of 98.0% and an HPLC purity of 99.3%. MS m/z: 463.2[M+1].

Nuclear magnetic data: ¹ HNMR (d ₆ -DMSO): δ ppm: 8.78 (s, 1H); 8.42-8.28 (m, 3H); 8.16 (s, 1H); 7.53 (d, 1H, J = 8.28); 7.29- 7.20 (m, 2H); 7.13-7.07 (m, 1H); 7.01 (s, 1H); 4.33 (t, 2H, J = 5.65); 4.02 (s, 3H); 3.88 (s, 3H); 2.71 ( t, 2H, J = 5.77); 2.27 (s, 6H).

Step 2-Preparation of Intermediate K:

Preparation: Add 5L of tetrahydrofuran and intermediate J (350g, 108mmol) to a 10L hydrogenation reactor, add 17.5g of wet palladium charcoal, replace the hydrogenation reactor with hydrogen, adjust the pressure value to 0.2MPa, control the temperature at 25°C, and keep the reaction warm. At 9h, HPLC monitors the progress of the reaction, and stops the reaction when the substrate is ≤0.5%. Filter, concentrate the filtrate under reduced pressure until the solvent volume is about 2L, adjust the internal temperature to room temperature, slowly add 4L n-heptane dropwise within 4-7 hours, filter and dry the solid under reduced pressure to obtain 285g of white powder intermediate K The yield was 86%, and the HPLC purity was 99.60%. MS m/z: 433.3 [M+1].

Nuclear magnetic data: ¹ HNMR (CDCl ₃ ): δ ppm: 8.42 (d, 1H, J = 7.78), 8.28 (s, 1H), 8.26 to 8.23 (m, 1H), 7.78 (s, 1H), 7.51 (d, 1H,J=8.28),7.41(s,1H),7.26-7.23(m,1H),7.19- 7.11(m,2H),6.72(s,1H), 4.38(br,2H),4.06(t, 2H,J=5.77), 3.88(s,3H), 3.75(s,3H), 2.63(t,2H,J=5.77), 2.26(s,6H).

Step 3-Preparation of compound of formula I:

Add 250 mL of anhydrous tetrahydrofuran solvent and Intermediate K (14 g, 32 mmol) to the reaction flask and stir, cool to 0-5° C., add 10% hydrochloric acid (12 ml), and stir for 20 minutes. At 0-5°C, slowly drop 3-chloropropionyl chloride (5.6 g, 45 mmol) into the reaction flask. Stir for 3 hours, after sampling test (K/(U+K)≤0.5%) is qualified, add 36% potassium hydroxide aqueous solution (75ml, 480mmol), heat to 23-25°C, and stir for 12 hours. Raise the temperature to 50-60°C and stir for 4 hours. After the sampling test (U/(U+L)≤0.1%) is qualified, stand still for liquid separation. Separate the organic phase, wash with 10% brine three times, dry, filter, and concentrate the organic phase to 150 ml. The temperature was raised to 40° C., 150 ml of n-heptane was slowly added dropwise, and the temperature was lowered to room temperature to precipitate crystals. Filtered and dried to obtain 10.71 g of light brown solid (compound of formula I), yield 68%, HPLC purity: 99.8% (all single impurities do not exceed 0.15%). MS m/z: 487.3[M+1].

Nuclear magnetic data (Figure 1): ¹ HNMR (d ₆ -DMSO): δppm: 9.84 (s, 1H), 8.90 ~ 8.82 (m, 1H), 8.32-8.25 (m, 2H), 7.89 (s, 1H) ,7.51(d,1H,J＝8.25), 7.27～7.10(m,1H), 6.94(s,1H), 6.49(dd,1H,J＝16.88,10.13), 6.25(dd,1H,J＝16.95 ,1.81),5.80～5.75(m,1H),4.19(t,2H,J＝5.57),3.88(d,6H,J＝14.63,6H),3.34(s,3H),2.58(d,2H, J=5.5), 2.28 (s, 6H).

(2) N-(2-(2-(Dimethylamino)ethoxy)-4-methoxy-5-((4-(1-methyl-1H-indol-3-yl)pyrimidine -2-yl)amino)phenyl)acrylamide methanesulfonate (Form A) preparation

Example 1

The compound of formula I (3 g, 6.1 mmol) was dissolved in 24 ml of dimethyl sulfoxide DMSO solvent, the temperature was raised to 65° C., and the mixture was stirred to clear. Add an equivalent amount of methanesulfonic acid (0.59 g, 6.1 mmol) to the system. The temperature was lowered to 50°C, and 12ml of isopropyl acetate IPAc was slowly added. Stir at 50°C for 1 hour, then lower the temperature to 15°C. Add 21ml IPAc in 4 hours. The solution was stirred and crystallized at 15°C, filtered under reduced pressure, the filter cake was washed with isopropyl acetate, and washed with acetone to reduce the residual DMSO solvent. Blow drying at 50°C (or vacuum drying at 50°C) to obtain 3.16 g of light yellow solid (crystal form A). HPLC purity is 100%, yield is 88%, DMSO: <100ppm; IPAc: <100ppm. MS m/z: 487.2 [M+1-MsOH]. Melting point: 242-244°C.

Nuclear magnetic data (figure 2): ¹ HNMR(d ₆ -DMSO): δppm: 9.57(brs,1H), 9.40(s,1H), 8.71(s,1H), 8.48(s,1H), 8.32(d , 1H, J = 7.9), 8.29 (d, 1H, J = 5.3), 7.96 (s, 1H), 7.51 (d, 1H, J = 8.2), 7.23 (ddd, 1H, J = 7.9, 7.1, 0.8 ), 7.19 (d, 1H, J = 5.4), 7.15 (ddd, 1H, J = 7.8, 7.3, 0.5), 6.94 (s, 1H), 6.67 (dd, 1H, J = 16.9, 10.2), 6.27 ( dd, 1H, J = 16.9, 1.8), 5.57 (dd, 1H, J = 16.9, 1.7), 4.44 (t, 2H, J = 4.6), 3.89 (s, 3H), 3.88 (s, 3H), 3.58 (t, 2H, J=4.6), 2.93 (s, 6H), 2.39 (s, 3H).

After testing, the powder X-ray diffraction pattern of crystal form A obtained in this example has diffraction angle 2θ values of 11.06±0.2°, 12.57±0.2°, 13.74±0.2°, 14.65±0.2°, 15.48±0.2°, 16.58±0.2°, 17.83±0.2°, 19.20±0.2°, 19.79±0.2°, 20.88±0.2°, 22.05±0.2°, 23.06±0.2°, 24.23±0.2°, 25.10±0.2°, 25.71±0.2°, 26.15±0.2°, 27.37±0.2°, 27.42±0.2° has a characteristic peak; its XRPD spectrum is shown in Figure 3 and the attached table, DSC diagram is shown in Figure 4, TGA diagram is shown in Figure 5, and infrared spectrum IR diagram is shown in Figure 6. Show.

Example 2

The compound of formula I (28.25 g, 58.1 mmol) was dissolved in 224 ml of dimethyl sulfoxide DMSO solvent, the temperature was raised to 15-35° C., and the mixture was stirred to clear. 0.97 equivalents of methanesulfonic acid (5.4 g, 0.97 mmol) were added to the system in batches. Slowly add 448 ml of methyl isobutyl ketone (MIBK). Stir for 1 hour, then lower the temperature to 10-15°C. The solution was reacted with salt formation at 10-15°C, samples were taken, and the residue of the compound of formula I in the mother liquor was detected by HPLC (≤0.4%). After the reaction was completed, vacuum filtration was performed to obtain 32 g of the crude methanesulfonate of the compound of formula I.

Add 3g of the crude methanesulfonate of the compound of formula I into 24ml of dimethyl sulfoxide DMSO solvent, stir to clear at 65°C, cool down, slowly add 48ml of methyl isobutyl ketone (MIBK) dropwise, stir and crystallize 6-8 After hours, vacuum filtration, drying at 60° C. (or 60° C. vacuum and reduced pressure drying) to obtain the target crystal form A. Melting point: 242-244°C. The XRPD pattern of the crystal form is consistent with Figure 3 (Figure 7), and all characteristic peaks are within the error range.

Example 3

Add 3 g of the crude methanesulfonate salt of formula I (prepared in Reference Example 2) into 24 ml of dimethyl sulfoxide DMSO solvent, stir to clear at 65°C, cool down, slowly add 48 ml of ethyl acetate dropwise, and stir to crystallize 6 -8 hours, vacuum filtration, washing the filter cake with ethyl acetate, and air drying at 70°C (or 60°C vacuum drying under reduced pressure) to obtain the target 2.8g crystal form A. Melting point: 242-244°C. The XRPD pattern of the crystal form is consistent with Figure 3, and all the characteristic peaks are within the error range.

Example 4

Add 3g of the crude methanesulfonate of the compound of formula I into 24ml of dimethyl sulfoxide DMSO solvent, stir to clear at 65°C, cool down, slowly add 12ml of methyl tert-butyl ether (MTBK) dropwise, and stir to crystallize 6-8 After hours, vacuum suction filtration, and blowing drying at 50°C (or 30°C vacuum drying under reduced pressure) to obtain the target crystal form A. Melting point: 242-244°C. The XRPD pattern of the crystal form is consistent with Figure 3, and all the characteristic peaks are within the error range.

Example 5

Add 3 g of the crude methanesulfonate salt of the compound of formula I into 24 ml of dimethyl sulfoxide DMSO solvent, stir and dissolve at 65° C., cool down, slowly add 48 ml of acetonitrile (ACN) dropwise, stir and crystallize for 6-8 hours, and pump under reduced pressure. Filter and dry by blowing at 50°C (or 60°C under reduced pressure under vacuum) to obtain the target crystal form A. Melting point: 242-244°C. The XRPD pattern of the crystal form is consistent with Figure 3, and all the characteristic peaks are within the error range.

Example 6

Add 3 g of the crude methanesulfonate salt of the compound of formula I into 24 ml of dimethyl sulfoxide DMSO solvent, stir and dissolve at 65°C, then cool down, slowly add 48 ml of toluene (PhMe) dropwise, stir and crystallize for 6-8 hours, and pump under reduced pressure. Filter and dry by blowing at 50°C (or drying under vacuum at 50°C) to obtain the target crystal form A. Melting point: 242-244°C. The XRPD pattern of the crystal form is consistent with Figure 3, and all the characteristic peaks are within the error range.

Example 7

Add 10 g of the crude mesylate salt of the compound of formula I to 100 ml of tetrahydrofuran THF solvent, stir to clear at 65°C, cool to room temperature, stir and crystallize for 6-8 hours, filter, and dry at 50°C to obtain 8.8 g of target crystal form A. Melting point: 242-244°C. The XRPD pattern of the crystal form is consistent with Figure 3, and all the characteristic peaks are within the error range.

Example 8

Add 2g of the crude methanesulfonate salt of the compound of formula I to 20ml of N-methylpyrrolidone NMP solvent, stir and dissolve at 65°C, cool to room temperature, stir and crystallize for 6-8 hours, filter under reduced pressure, and blow at 50°C. Drying (or vacuum drying at 50°C) to obtain the target crystal form A. Melting point: 242-244°C. The XRPD pattern of the crystal form is consistent with Figure 3, and all the characteristic peaks are within the error range.

Example 9

Add 2g of the crude methanesulfonate salt of the compound of formula I into 30ml N,N-dimethylformamide DMF solvent, stir and dissolve at 65°C, cool to room temperature, stir and crystallize for 6-8 hours, and filter under reduced pressure. Blow drying at °C (or 60 °C vacuum drying under reduced pressure) to obtain the target crystal form A. Melting point: 242-244°C. The XRPD pattern of the crystal form is consistent with Figure 3, and all the characteristic peaks are within the error range.

Example 10

Add 2g of the crude methanesulfonate salt of the compound of formula I into 50ml N,N-dimethylacetamide DMAc solvent, stir and dissolve at 65°C, cool to room temperature, stir and crystallize for 6-8 hours, filter under reduced pressure, 60 Blow drying at °C (or 60 °C vacuum drying under reduced pressure) to obtain the target crystal form A. Melting point: 242-244°C. The XRPD pattern of the crystal form is consistent with Figure 3, and all the characteristic peaks are within the error range.

Example 11

Add 2g of the crude methanesulfonate salt of the compound of formula I into 50ml of acetone/water mixed solvent (acetone/water=1/1v/v), raise the temperature to 50°C and stir to make slurry, cool to room temperature, and stir and crystallize for 6-8 hours. Pressure suction filtration, and blowing drying at 50°C (or 50°C vacuum drying under reduced pressure) to obtain the target crystal form A. Melting point: 242-244°C. The XRPD pattern of the crystal form is consistent with Figure 3, and all the characteristic peaks are within the error range.

(3) N-(2-(2-(Dimethylamino)ethoxy)-4-methoxy-5-((4-(1-methyl-1H-indol-3-yl)pyrimidine -2-yl)amino)phenyl)acrylamide methanesulfonate (Form A) structure confirmation

The crystal form A (purity greater than 99.80%) prepared by the method of Example 2 was used for the structure determination test of the crystal form A using the following example method.

Test case 1

The elemental analysis method was used to test the percentages of C, H, N, and S elements contained in the crystal form A, and the test was performed twice in parallel, and the average value was taken.

CHN test equipment: Elementar Vario EL III element analyzer (C, H, N determination);

CHN test method: the sample is decomposed by combustion, quantitatively converted, tested, and then processed by data to obtain the percentages of C, H, and N;

S titration method: accurately weigh the crystal form A samples, respectively wrap them tightly with ash-free filter paper, decompose them by combustion method, and _{absorb them with pure water and 30% H 2} O ₂ as the absorption liquid. After the absorption is complete, transfer to a 250mL volumetric flask, rinse the combustion flask with ethanol, and transfer the rinsing fluid to the 250mL volumetric flask. Then add a certain amount of known concentration of HNO ₃ and chlorophosphonazo III indicator, and then titrate with a known concentration of Ba(ClO ₄ ) ₂ ·3H ₂ O until the color of the solution changes from purple to blue. Record the consumption volume of Ba(ClO ₄ ) ₂ ·3H ₂ O and calculate the percentage of S.

Table 1. Elemental analysis test results of crystal form A

The measurement results show that _{the difference between the actual and theoretical values of C, H, N, and S content of the crystal form A (C 27} H ₃₀ N ₆ O ₃ ·CH ₄ O ₃ S) sample is less than 0.3%, indicating that the actual measured value is different from the theoretical value. The theoretical value is consistent, it is a salt with 1 molecule of methanesulfonic acid. It is consistent with the data reported in Example 2 of Chinese Patent Application No. CN201780050034.3.

Test case 2

Purpose: To test the specific crystal structure of Form A by powder X-ray diffraction analysis.

Instrument: Bruker D8 Advance powder X-ray diffractometer

Table 2. Test condition parameter table

Measurement results and analysis: From the powder X-ray diffraction pattern of crystal form A (Figure 3 and the attached table), it can be seen that this compound has a specific crystal structure. Its 2θ angle (°) has strong peaks at 11.06, 12.57, 13.74, 14.65, 15.48, 16.58, 17.83, 19.20, 19.79, 20.88, 22.05, 23.06, 24.23, 25.10, 25.71, 26.15, 27.37, 27.42 (relative intensity is greater than 10%).

Table 3. Comparison table with Example 2 of Chinese patent application number CN201780050034.3

Test case 3

Purpose: Use the differential thermal analysis DSC method to test the melting point of crystal form A.

Instrument: TA Q2000 Differential Calorimetry Scanner

Parameters: heating range: 30～300℃; heating rate: 10℃/min

Measurement results and analysis: It can be seen from the DSC graph (Figure 4) that the melting point of the crystal form A is 242-244°C.

Table 4. Comparison with Chinese Patent Application No. CN201780050034.3 Example 2 Methanesulfonate

Different crystal forms of mesylate Melting point Crystal Form A 242-244°C 2A 233.3-238.1℃ 2B 157.3-180.2℃

Test case 4

Purpose: Use thermogravimetric analysis to test whether crystal form A contains water or solvent compounds.

Instrument: TA Q5000 Thermogravimetric Analyzer

Parameters: heating range: 30～300℃; heating rate: 10℃/min

Measurement results and analysis: It can be seen from the TSA graph (Figure 5) that the weight loss of crystal form A is very small at 33.40°C to 120.00°C. Combined with DSC and elemental analysis, it is shown that crystal form A does not contain water or solvent.

Table 5. Comparison with Chinese Patent Application No. CN201780050034.3 Example 2 Methanesulfonate

Different crystal forms of mesylate Thermal weight loss (weight loss wt%) Crystal Form A -0.035% (33.4-120℃) 2A -0.435％(32.8-210.0℃) 2B -8.063% (29.7-193.4℃)

Test case 5

Purpose: To test the infrared spectrum of crystal form A by infrared absorption spectroscopy.

Instrument: Nicolet 380 FT-IR infrared spectrometer, YP-2 tablet press

Method: KBr tablet method

Measurement results and analysis: It can be seen from the infrared absorption spectrum (Figure 6) that the structure of this product contains aromatic rings, aromatic heterocycles, alkenyl groups, -CH3, -CH2-, amine groups, amine salts, amides, ether groups, and Sulfonate structure, etc. The above infrared IR spectrum results are consistent with the structure of crystal form A.

Test case 6

Purpose: To investigate the material stability of the crystal form A compound prepared in Example 2 of the method of this patent.

Method: Refer to the requirements of the Chinese Pharmacopoeia (2015 Edition) for the stability test of APIs, using influencing factors and accelerated/long-term stability inspection methods to test the material stability data of crystal form A (the test design is shown in Table 6 below).

Instrument: climatic condition box; light box

Table 6. Stability test design

*Remarks: In the experiment of influencing factors, samples at 40°C are tested only when the 60°C test result does not meet the acceptance criteria. Only when the 25°C/90%RH test result does not meet the acceptance criteria, the 25°C/75%RH sample is tested.

Table 7. Experimental results of influencing factors-high temperature test

Table 8. Experimental results of influencing factors-high humidity test

Table 9. Experimental results of influencing factors-light test

Table 10. Experimental results of influencing factors-light test

Table 11. Experimental results of influencing factors—long-term test

Measurement results and analysis: Influencing factors, long-term and accelerated stability research data show that after long-term storage of crystal form A for 36 months, there is no significant change in the detection indicators compared with 0 days. All the detection indicators meet the established standards and there is no obvious defect. trend. It shows that the crystal form A prepared by this process has good stability under the existing packaging conditions, and various test indexes can meet the requirements of quality standards.

(4) Study on the druggability of crystal form A

Test example 1

Purpose: To evaluate the solubility of crystal form A in different biological media.

Experimental design: Weigh approximately 2.5 mg of the compound of formula I and crystal form A (methanesulfonate of the compound of formula I) into three glass bottles, and add 200 μL of SGF, FaSSIF and FeSSIF to the bottles respectively. Stir the suspension for about 30 minutes at 37°C. (1) If it becomes a clear solution, continue to add the compound until it becomes a suspension; (2) If it is not, continue to stir the suspension for 24 hours. In 24h, it is necessary to measure the pH value and visually inspect its appearance. If it is still a suspension after 24 hours, centrifuge the solids, and dilute the supernatant with 50% acetonitrile aqueous solution, and then use HPLC to determine the concentration of the diluted solution for content detection. The results are shown in the table below.

Table 12. Comparison results of the solubility of the compound of formula I and crystal form A in different biological media

Conclusion: The crystalline form A of the compound of formula I methanesulfonate has a better solubility in bio-related media than that of the compound of formula I, especially in the simulated gastric juice SGF medium. It is suggested that the crystal form A is suitable for the development of oral gastric-dissolved fast-release solid preparations.

Test example 2

Objective: To evaluate the comparison of the pharmacokinetic parameters of SD rats after a single oral administration of crystal form A and formula I compound

Experimental design: Healthy SD rats were randomly divided into formula I compound group (free base) and formula I compound methanesulfonate crystal form A group. Both groups were male, with 21 rats in each group, weighing 190-210g. Each test compound was administered orally at a single dose of 60 mg/kg under fasting conditions. Whole blood was collected intravenously and plasma was separated at different time points 0.5, 1, 2, 4, 8, 12 and 24 hours after administration. The liquid phase was used. Chromatography/tandem mass spectrometry (LC-MS/MS) detects the drug concentration of the crystal form A or formula I compound in the plasma, and uses a non-compartmental model to calculate the relevant pharmacokinetic parameters. The main pharmacokinetic parameters are shown in the following table:

Table 13. The main pharmacokinetic parameters of the compound group of formula I and the mesylate crystal form group A of the compound of formula I

Pharmacokinetic parameters Free base Crystal Form A C _max (ng/mL) 496 622 T _max (h) 4.00 2.00 T _1/2 (h) 5.02 3.31 AUC _0-24 (ng·h/ml) 5899 6754

AUC _0-∞ (ng·h/ml) 6236 6846 MRT _0-24 (h) 8.29 7.46 MRT _0-∞ (h) 9.53 7.75

C _max : the highest concentration of the drug in the plasma; T _max : the peak time of the highest concentration of the drug; T _1/2 : elimination half-life; AUC: the area under the plasma concentration-time curve; MRT: the average residence time of the drug

It can be concluded from the above table and Figure 8 that the bioavailability of crystal form A is significantly better than that of free alkali.

Test example 3

Objective: To evaluate the drug activity test of crystal form A as a lung cancer drug

1) Inhibition of specific cell growth and selection specific experiments

Methods: Fluorescence cell proliferation technology and cell microscopic visualization detection technology (Alma blue method) were used to detect the deletion of crystal form A on human epidermal cancer cells (A431, wild-type EGFR) and human lung cancer cells (HCC827, EGFR exon 19) Type activating mutation), whether human lung cancer cells (H1975, EGFR L858R/T790M drug-resistant mutation) have anti-proliferative activity.

Experimental design: Cells in the logarithmic growth phase were seeded in a 96-well plate (the cell concentration was 2000 cells/well; cell suspension was 200 μl/well), and _{cultured at 30°C and 5% CO 2 for} 24 hours to make the cells adherent. The DMSO solution of the test compound was diluted with the cell culture medium solution from high to low to different concentrations, and each dilution was 3 times to obtain a total of 9 different concentrations. The test drug solutions of different concentrations were added to the 96-well cell plate containing the above-mentioned cancer cells, with 3 replicate wells for each concentration. At the same time, set up a cell control well containing only DMSO cell culture medium solution. After continuing the culture at 37°C and 5% CO ₂ for 72 hours, the culture was terminated.

Cell proliferation was detected after staining with Alamar Blue reagent (resazurin). Resazurin (resazurin) is reduced by cells to resorufin and becomes a fluorescent substance (excitation at 544nm, color development at 612nm), and the fluorescence intensity is proportional to the number of cells. The resazurin reagent was dissolved in PBS solution to prepare a stock solution with a concentration of 440 μM. Add 40μl of 440μM resazurin stock solution to the cell culture plate, incubate for 5 hours, restore the cell culture plate to normal culture conditions, and use Cytation3 multi-mode microplate reader (Biotek) to read the fluorescence measurement after 72 hours .

Fluorescence detection was normalized with cell-free (background) well readings, and the average of total growth in 72 hours and vehicle control wells (n=3) was determined. The results are shown in the table below:

Table 14. Experimental results of specific cell growth inhibition and selection of crystal form A and osimertinib

*Data source: Table 4, P21, PHARMACOLOGY REVIEW(S), APPLICATION NUMBER:208065Orig1s000, CDER of FDA; https://www.accessdata.fda.gov/drugsatfda_docs/nda/2015/208065Orig1s000PharmR. pdf

The results showed that the crystal form A compound showed excellent activity to inhibit the proliferation and growth of cancer cells for H1975 and HCC827, but did not significantly inhibit the growth of wild-type cells, which was equivalent to the foreign marketed product (osimertinib); but the crystal form A showed Shows a superior inhibitory selectivity for H1975 and HCC827 cells (compared to wild-type A431 cancer cells).

2) Inhibition selectivity for different kinase targets

Kinase inhibitors are used as drugs for clinical treatment. In addition to targeted kinases, it is necessary to avoid that inhibitors also inhibit other non-targeted kinases, and may cause some cytotoxic reactions (the so-called "off-target effects"). In this experiment, high-throughput screening (kinase profiling) of 97 kinases (mutation site kinase zymograms related to human diseases) was carried out by KINOMEscan's in vitro competitive binding analysis method ^*1.

Table 15. Results of high-throughput screening of 97 kinase zymograms by crystal form A and osimertinib

^*1 Refer to Fabian, MA et al. A small molecule-kinase interaction map for clinical kinase inhibitors. Nat. Biotechnol. 23, 329-336 (2005) for detailed introduction of in vitro binding analysis method.

^*2 Data source: P15, P43, PHARMACOLOGY REVIEW(S), APPLICATION NUMBER:208065Orig1s000, CDER of FDA;

https://www.accessdata.fda.gov/drugsatfda_docs/nda/2015/208065Orig1s000PharmR. pdf

The results show that the selectivity of the inhibitory activity of the non-target kinase of the crystal form A compound is better than that of the marketed drug osimertinib.

Drug Safety Evaluation of Test Example 4 Crystal Form A

1) Safety pharmacology test

OBJECTIVE: To simulate the clinical route of administration, to observe the effects on the central nervous system of SD rats, the respiratory system of SD rats and the cardiovascular system of Beagle dogs after a single oral administration of crystal form A preparations. The purpose is to study the potential undesirable potential adverse effects on physiology (central nervous system, respiratory system and cardiovascular system) when the crystalline form A compound is in the therapeutic range or above the therapeutic range.

method:

[Central Nervous System Experiment Design] Take 7-week-old SD rats, half male and half male, weighing between 170-235g, a total of 40 SD rats (20 male and female), randomly divided into 4 groups, 5 male and female in each group. Animals are given a single oral dose of vehicle (0.5% (w/v) methyl cellulose), 10, 40 or 160 mg/kg of crystalline form A compound dissolved in 1% (w/v) methyl cellulose (vehicle) For the suspension, control animals were given vehicle only. The administration volume for all animals was 10 mL/kg. Use the Functional Observation Combination (FOB), which includes tests on the animal's motor function, behavioral changes, coordination function, sensory/motor reflex and body temperature, to evaluate the effects of different doses of test products on the function of the animal's central nervous system. FOB observations were performed at about 2 hours, 4 hours and 24 hours before the experiment and after the administration.

[Respiratory system experiment design] Take 7-8 weeks old SD rats, half male and half female, weighing between 168-287g, a total of 40 rats (20 male and female) rats are randomly divided into 4 groups, each group has 5 males and 5 males only. Animals are given a single oral dose of vehicle (0.5% (w/v) methyl cellulose), 10, 40 or 160 mg/kg of crystalline form A compound dissolved in 0.5% (w/v) methyl cellulose (vehicle) For the suspension, control animals were given vehicle only. The administration volume for all animals was 10 mL/kg. On the day of dosing, the animals should be placed in the "outcrops" plethysmograph for at least 5 minutes before the data collection. After the adaptation, the respiratory data (respiratory rate, tidal volume, and minute ventilation) are collected immediately. The collection time is Before administration, about 2 hours (±10min), 4 hours (±10min) and 24 hours after administration, the duration of each collection is about 15 minutes continuously. Record the changes in tidal volume, respiratory volume and respiratory frequency before and after administration. Set 0h, 2h, 4h and 24h time points. The indexes after administration were compared with their own controls, and statistical analysis was performed.

[Experimental Design of Cardiovascular System] Take 22-24 months old beagle dogs, half male and half female, weighing between 9.3-10.5kg, a total of 6 beagle dogs, and implant remote sensing sub-devices in advance. According to the design of dose escalation, each dog was given 1 vehicle and 10, 30, 100 mg/kg of crystalline form A compound suspension dissolved in 0.5% (w/v) methylcellulose (vehicle), control animals Give only the vehicle. The interval between the two doses is 3 days (except for controls, which are 2 days apart). The dosage volume is 5ml/kg, and the actual dosage is based on the animal's weight recently weighed. Using the telemetry method, the ECG data before the administration, 2 hours, 4 hours, 6 hours, 8 hours and 24 hours after the administration were collected, and the obtained electrocardiogram data were submitted to the ECG expert veterinarian for evaluation.

Results: Oral crystal form A has low toxicity. The test results of the central nervous system, respiratory system and cardiovascular system of experimental animals are all negative, indicating that the adverse reactions are small and have high safety.

2) Genotoxicity test

Purpose: Genotoxicity test can be used to detect somatic cell mutagens, germ cell mutagens and potential carcinogens. Using a combination of in vitro and in vivo genetic toxicity tests, selected bacteria (including Salmonella typhimurium and Escherichia coli) reverse mutation in vitro test (Ames test), Chinese hamster ovary cell chromosomal aberration test in vitro, and intragastric administration of bone marrow to rats The micronucleus in vivo test is a three-part test to comprehensively evaluate the potential genotoxicity of crystal form A formulations.

method:

[Ames experimental design] Four kinds of histidine-deficient Salmonella typhimurium were induced by detecting different doses of crystalline form A compounds with and without exogenous metabolic activation system (rat liver S9 induced by Aroclor1254) (TA98, TA100, TA1535 and TA1537) and tryptophan-deficient Escherichia coli Escherichia Coli WP2 uvrA to produce back mutations in order to evaluate its potential mutagenicity. A negative/solvent control group and a positive control group are also set up at the same time. The experimental dosage is set as: 10, 25, 50, 100, 250 and 1000μg/dish for the addition and not adding S9 series for TA98, TA100, TA1535 and TA1537; 100, 250, 500 for the addition and not adding S9 series for WP2uvrA , 1000, 2500 and 5000μg/dish.

[Chromosome aberration experiment design] By detecting different doses of crystal form A compounds with and without exogenous metabolic activation system (rat liver S9 induced by Aroclor1254), Chinese hamster ovary cells (CHO-WBL) were induced to produce chromosomes Ability to distort. In the S9 activation experimental system, the cells were treated for 3 hours; without the S9 experimental system, the cells were treated for 3 hours and 22 hours. After culturing for 22 hours after administration, all cells were harvested. A negative/solvent control group and a positive control group are also set up at the same time. The experimental dosage is set as follows: the activation system with S9 is 2, 5, 12 and 14μg/ml; the non-activated series without S9 is 1, 3 and 6μg/ml.

[Design of bone marrow micronucleus experiment] Take 7-8 week old rats, weighing between 241-206g, a total of 46 male rats were randomly divided into 7 groups, each group of 6 animals (of which 5 and 6 groups are 8 ). The vehicle (0.5% (w/v) methyl cellulose) (groups 1 and 2) was given a single oral dose, 500, 1000, 2000 or 2000 mg/kg dissolved in 0.5% (w/v) methyl cellulose ( Vehicle) crystal form A compound suspension (groups 3-6), and 20 mg/kg of the positive control drug (physiological saline solution of cyclophosphamide monohydrate, administered via a single intraperitoneal injection) (group 7). The administration volume for all animals was 10 mL/kg. Animals in groups 1, 3, 4, 5, and 7 were sacrificed approximately 24 hours after administration. Animals in groups 2 and 6 were sacrificed approximately 48 hours after administration. Micronucleus-containing polychromatic erythrocytes ( MN-PCE) formation rate.

Results: No genotoxicity (Ames test, mouse micronucleus test and chromosome aberration test) was found in all genotoxicity studies of the crystal form A compound. The results were all negative. It can be inferred that the crystal form A does not have mutagenicity and obvious genetic harm. It shows that the crystal form A has no obvious genotoxicity and mutagenic effects.

Clinical Application Example 1 Human clinical trial of crystal form A

OBJECTIVE: To evaluate the safety and efficacy of oral preparations prepared by crystal form A in Chinese patients with locally advanced or metastatic non-small cell lung cancer who have previously received EGFR-TKI treatment.

Materials and methods: Capsules prepared with crystal form A (excipients are microcrystalline cellulose, lactose, croscarmellose sodium, colloidal silicon dioxide, magnesium stearate; after mixing according to the required drug dosage Directly filling and filling No. 3 gelatin hollow capsules), a total of 128 patients with non-small cell lung cancer were evaluated for safety and efficacy.

Results: The clinical safety studies after single and multiple oral administrations in 6 dose groups of 30, 60, 120, 180, 240, 300 mg have been completed, and the results show that the dose of 30 mg-300 mg is safe.

The mid-term efficacy analysis of the clinical trial showed that QD from 30-300mg once a day, the lowest starting dose of 30mg showed efficacy (disease control rate DCR80.0%), the recommended phase II dose of 180mg disease remission rate ORR=73.1%, higher than the marketed The drug osimertinib (66.1%); the disease remission rate DCR is 96.2%, which is also higher than the listed drug osimertinib (91%). The specific efficacy evaluation is as follows:

Table 16. Efficacy evaluation results of different dose groups

The precise interpretation of the terms involved above is: CR: complete remission, PR: partial remission, SD: stable disease, PD: disease progression, ORR: objective response rate, DCR: disease control rate.

The mid-term efficacy analysis of the clinical trial showed that from 30-300mg QD once a day, the incidence of adverse reactions (proportion and type) was significantly lower than that of the marketed drug Osimertinib. The specific safety evaluation is as follows:

Table 17. Comparison table of adverse reaction AE incidence between crystal form A and osimertinib

Table 18. Comparison list of TEAE types of adverse events greater than 10% of crystal form A and osimertinib

*Data source: Page 4, Instructions for Use of Osimertinib Mesylate Tablets and Page 89, MEDICAL&STATISTICAL REVIEW(S), APPLICATION NUMBER:208065Orig1s000, CDER of FDA;

https://www.accessdata.fda.gov/drugsatfda_docs/nda/2015/208065Orig1s000MedR.p df

Clinical Application Example 2 Application of Combination Study of Crystal Form A

Purpose: To observe the crystal form A prepared in Example 2 of the present invention and the compound BPI-7722 (N-(5-((4-ethylpiperazin-1-yl)methyl)pyridin-2-yl)-5- Fluoro-4-(3-isopropyl-2-methyl-2H-indazol-5-yl)pyrimidin-2-amine hydrochloride) as a single agent or combination therapy on human lung cancer H1975 subcutaneously transplanted tumors in nude mice Inhibition.

Materials and Method:

The free base form of compound BPI-7722 can be synthesized from Example 1 of Chinese Patent Application No. CN201580047916.5 (Publication No. CN106687454A) and has the following structure:

Cell culture: H1975 (human lung adenocarcinoma) was purchased from the Cell Bank of the Chinese Academy of Sciences, and the cells were cultured in a CO ₂ constant temperature incubator. The cell culture medium is modified RPMI; 10% fetal bovine serum; 100U/ml penicillin and 100μg/ml streptomycin. The cells were digested with 0.25% trypsin and passaged every other day. After the cells are expanded to the required number of cells and the cells are in the logarithmic growth phase, the cell count is collected for inoculation.

Experimental animals: BALB/c nude male nude mice, 24, 6-7 weeks, 16-18g, 6 in each group, purchased from Shanghai Lingchang Biological Technology Co., Ltd. Twenty-one days after the nude mice were inoculated with H1975 cells, ^{24 mice with a tumor volume of 80-254 mm 3} were selected and randomly divided into 4 groups according to the tumor volume and body weight, with 6 animals in each group. Respectively: Group 1: 0.5% sodium carboxymethyl cellulose solvent control group; Group 2: Example 2 crystal form A (5mg/kg) single-drug group; Group 3: BPI-7722 (100mg/kg) single-drug group ; Group 4: Crystal Form A (5mg/kg)/BPI-7722 (25mg/kg) combination medication group.

Experimental method: Human lung cancer H1975 cell line (5×10 ⁶ cells/mouse) was inoculated into experimental mice under the skin of the right back. The tumor growth was regularly observed, and the diameter of the transplanted tumor was measured with a vernier caliper. (80～254mm ³ ) according to tumor size and mouse body weight. Each group was administered intragastrically at the above dose, and the dose was calculated according to the latest body weight of the mice, and the dose was 10ml/kg. It is administered once a day for 21 consecutive days. Throughout the experiment, the body weight and tumor size of the mice were measured three times a week to observe whether there was a toxic reaction. Tumor volume calculation formula: tumor volume TV (mm ³ )=0.5×(tumor long diameter×tumor short diameter ² ). The tumor growth curve of each experimental group is shown in the table below and Figure 9, and the body weight curve is shown in the table below and Figure 10.

Table 19. Summary of the relationship between average tumor volume and time in each dose group

*The animals in the blank group have been euthanized due to excessive tumor burden, so no data collection was performed on the 21st day.

Table 20. Summary table of the relationship between average body weight and time in each dose group

*The animals in the blank group have been euthanized due to excessive tumor burden, so no data collection was performed on the 21st day.

Conclusion: The crystalline form A compound of Example 2 of the present invention has a good inhibitory effect on the growth of human lung cancer H1975 nude mice subcutaneously transplanted tumors; the inhibitory effect after combined with BPI-7722 is significantly better than that of the single drug in inhibiting tumor growth ; And show good safety.

Although the present invention has been described to a certain extent through its implementation and with reference to the accompanying drawings, it is worth noting that after reading the above disclosure of this application, those skilled in the art may not need to deviate from the spirit and scope of the present invention. Various modifications, changes or modifications are made to the present invention, and such changes and modifications should be included in the scope of the appended claims of the present invention.

Claims (11)

The crystal form A of the aminopyrimidine compound mesylate salt is characterized in that the aminopyrimidine compound is a compound of formula I, and the XRPD pattern of the crystal form A has 12.57, 13.74, 14.65 in 2θ (±0.2°) , 15.48, 16.58, 17.83, 19.20, 19.79, 22.05, 24.23, 25.10, 26.15, 27.37, 27.42 diffraction peaks,

The crystalline form A of the aminopyrimidine compound methanesulfonate according to claim 1, wherein the XRPD pattern of the crystalline form A is shown in FIG. 3. The crystalline form A of the aminopyrimidine compound methanesulfonate according to claim 1, wherein the infrared spectrum of the crystalline form A is shown in FIG. 6. The preparation method of the crystalline form A of the methanesulfonate salt of the aminopyrimidine compound according to any one of claims 1 to 3, comprising: dissolving the methanesulfonate salt of the compound of formula I in an organic solvent, stirring, crystallization, filtering, and drying to obtain The target crystal form A, the organic solvent includes dimethyl sulfoxide, tetrahydrofuran, N-methylpyrrolidone, N,N-dimethylformamide and N,N-dimethylacetamide. The preparation method according to claim 4, further comprising: preparing the methanesulfonate of the compound of formula I by mixing the compound of formula I and methanesulfonic acid in a molar ratio of 1.05-0.97. The preparation method according to claim 4, characterized in that an anti-solvent is added for stirring and crystallization, and the anti-solvent includes methyl tert-butyl ether, isopropyl acetate, ethyl acetate, methyl isobutyl ketone, Acetonitrile, toluene, acetone and water. The preparation method according to claim 4, characterized in that, after filtering, drying is performed under normal pressure or vacuum at 30-70°C. A pharmaceutical composition comprising the crystalline form A of the methanesulfonate salt of the aminopyrimidine compound described in any one of claims 1 to 3 and a pharmaceutically acceptable carrier. The crystalline form A of the methanesulfonate of the aminopyrimidine compound according to any one of claims 1-3, the pharmaceutical composition according to claim 8, or the methanesulfonate of the aminopyrimidine compound according to any one of claims 1-3 The application of the combination of the crystal form A of the acid salt and BPI-7722 in the preparation of anti-tumor drugs. The application according to claim 9, wherein the tumor is lung cancer, preferably non-small cell lung cancer. The method for treating or preventing diseases or medical conditions mediated by EGFR mutations in mammals, especially humans, preferably tumors, comprises administering a therapeutically effective amount of a crystal form of an aminopyrimidine compound mesylate to an individual in need A or a pharmaceutical composition comprising a crystalline form A of an aminopyrimidine compound mesylate and a pharmaceutically acceptable carrier.

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