Crystal form, preparation method and use of arylphosphorus oxide compounds

Abstract

The present invention relates to a polymorph of an aromatic phosphorus oxide compound and a dihydrate crystal form thereof, a preparation method and uses thereof. The crystalline aromatic phosphorus oxide compound and its dihydrate crystal can be used as an EGFR and / or ALK inhibitor, and the dihydrate crystal has good stability and is not hygroscopic.

Description

Technical Field

[0001] The present invention belongs to the field of pharmaceutical chemistry, and specifically relates to a medicinal arylphosphorus oxide compound and a crystal form of its dihydrate, a preparation method thereof and uses thereof. Background Art

[0002] Protein kinases represent a large family of proteins that play an important role in the regulation of a variety of cellular processes and in maintaining control of cellular functions. They include proliferation, apoptosis, cytoskeletal rearrangement, differentiation, development, immune response, nervous system function and conduction. In addition, many diseases and / or disorders are associated with abnormal, aberrant or dysregulated activity of one or more kinases.

[0003] Lung cancer is one of the most common malignant tumors. It is generally divided into two categories: small cell lung cancer (SCLC) and non-small cell lung cancer (NSCLC). However, whether in China or the world, lung cancer ranks first. Among them, non-small cell lung cancer (NSCLC) accounts for more than 80% of all lung cancers, which seriously threatens human health (Chinese Journal of Lung Cancer [J], 2012 Feb 20; 15 (2): 106-111).

[0004] EGFR (epidermal growth factor receptor, EGFR) is the full name of epidermal growth factor receptor, which is a transmembrane glycoprotein with tyrosine kinase activity widely distributed on the cell membranes of various tissues in the human body. The mutation and abnormal activation of EGFR are closely related to the occurrence, development, malignancy and metastasis of various tumors such as non-small cell lung cancer, breast cancer, esophageal cancer, etc. Most patients with non-small-cell lung cancer (NSCLC) have overexpression of EGFR. Among Asian non-small cell lung cancer populations (especially Chinese populations), about 40-50% have EGFR mutations. Therefore, inhibition of EGFR can significantly improve the survival of NSCLC patients. Common mutations of EGFR can be divided into two categories. One is drug-sensitive mutations, that is, anti-tumor targeted drugs can be used after mutation, such as exon 19 deletion and exon 21 L858R mutation; the other is drug-resistant mutations, that is, resistance to certain anti-tumor targeted drugs after mutation, such as T790M mutation and C797S mutation. The first-generation EGFR small molecule inhibitor drugs gefitinib, erlotinib and icotinib have achieved significant clinical efficacy and prolonged survival in patients with EGFR sensitive mutations. However, most of the patients who benefited from the drug will develop drug resistance after using the drug for a few months. Among them, more than 50% of the resistant patients are resistant due to the T790M mutation of EGFR. Although the second-generation EGFR irreversible inhibitor drugs afatinib and neratinib have achieved good results in preclinical studies, they lack selectivity for wild-type EGFR (EGFRWT) and have side effects such as greater skin toxicity. Osimertinib (AZD9291), a third-generation irreversible inhibitor that overcomes EGFR T790M resistance, can effectively treat patients with advanced non-small cell lung cancer with epidermal growth factor receptor T790M mutation or resistance to other EGFR inhibitors in clinical practice. Although Osimertinib has achieved great success in the clinical treatment of non-small cell lung cancer with EGFR T790M mutation, some patients who benefited from it developed drug resistance after 9 to 14 months of treatment (Nature Medicine 2015, 21(6), 560-562). Studies have found that up to 40% of drug-resistant patients are resistant to Osimertinib due to the (EGFR) C797S point mutation. Further mechanism studies have shown that the (EGFR) C797S point mutation converts the cysteine ​​at position 797 to serine, resulting in Osimertinib's inability to form a covalent bond with the target protein, ultimately causing drug resistance. Currently, there is no clinically available drug that is an effective EGFR inhibitor for the new mutation (C797S).Therefore, new types of highly selective EGFR inhibitors are urgently needed to address problems such as drug resistance caused by (EGFR) C797S point mutation.

[0005] Anaplastic lymphoma kinase (ALK), also known as ALK tyrosine kinase receptor or CD246, is an active enzyme encoded by the ALK gene in the human body. Fusion genes formed by ALK are closely related to the occurrence and development of various tumors such as non-small cell lung cancer. In patients with non-small cell lung cancer, fusion oncogenes such as EML4-ALK (echinoderm microtubule-associated protein 4 and anaplastic lymphoma kinase fusion gene) account for about 3-7%. Therefore, the development of protein kinase inhibitors for ALK fusion gene positive has important clinical value. On the other hand, with the increase in non-small cell lung cancer patients and the popularization of second-generation sequencing technology (deep sequencing) in recent years, researchers have continuously found that EGFR mutants and ALK gene fusion can occur simultaneously in some non-small cell lung cancer patients. Therefore, new types of highly selective protein kinase inhibitors are urgently needed to solve problems such as drug resistance caused by (EGFR) C797S point mutations, while also solving ALK gene fusions and mutations. Summary of the invention

[0006] One or more embodiments provide a crystal of (2-((5-bromo-2-((2-methoxy-5-methyl-4-(4-(4-methylpiperazin-1-yl)piperidin-1-yl)phenyl)amino)pyrimidin-4-yl)amino)-5-methylphenyl)dimethylphosphine oxide dihydrate having a crystalline form 1, using Cu-Ka radiation and expressed in 2θ angles, the X-ray powder diffraction pattern of the crystalline form 1 having the following characteristic peaks: 5.54±0.2°, 11.06°±0.2, 19.12°±0.2 and 27.90±0.2°.

[0007] In one or more embodiments, using Cu-Ka radiation and expressed in 2θ angles, the X-ray powder diffraction pattern of Form 1 has the following characteristic peaks: 5.54±0.2°, 11.06±0.2°, 16.15±0.2°, 16.73±0.2°, 17.10±0.2°, 19.12±0.2°, 21.18±0.2°, 24.16±0.2°, 26.27±0.2°, 26.48±0.2°, 27.90±0.2°, and 33.64±0.2°.

[0008] In one or more embodiments, using Cu-Ka radiation and expressed in 2θ angles, the X-ray powder diffraction pattern of Form 1 has the following characteristic peaks: 5.54±0.2°, 8.50±0.2°, 10.53±0.2°, 11.06±0.2°, 11.48±0.2°, 13.00±0.2°, 16.15±0.2°, 16.73±0.2°, 17.10±0.2°, 19.12±0.2°, 19.47±0.2°, 21.18±0.2°, 24.16±0.2°, 26.27±0.2°, 26.48±0.2°, 27.90±0.2°, and 33.64±0.2°.

[0009] In one or more embodiments, using Cu-Ka radiation and expressed in 2θ angles, the X-ray powder diffraction pattern of Form 1 has the following characteristic peaks: 5.54±0.2°, 8.50±0.2°, 10.53±0.2°, 11.06±0.2°, 11.48±0.2°, 13.00±0.2°, 16.15±0.2°, 16.73±0.2°, 17.10±0.2°, 18.34±0.2°, 19.12±0.2°, 11.20±0.2°, 11.30±0.2°, 11.40±0.2°, 11.60±0.2°, 11.70±0.2°, 11.80±0.2°, 11.90±0.2°, 12.1 ... 9.47±0.2°, 21.18±0.2°, 22.21±0.2°, 22.65±0.2°, 23.10±0.2°, 23.49±0.2°, 24.16±0.2°, 26.27±0.2°, 26.48±0.2°, 26.93±0.2°, 27.35±0.2°, 27.90±0.2°, 29.35±0.2°, 32.22±0.2°, 33.64±0.2°, 34.17±0.2°.

[0010] In one or more embodiments, using Cu-Ka radiation and expressed in 2θ angles, the X-ray powder diffraction pattern of Form 1 is as follows: Figure 1 shown.

[0011] One or more embodiments provide anhydrous crystals of (2-((5-bromo-2-((2-methoxy-5-methyl-4-(4-(4-methylpiperazin-1-yl)piperidin-1-yl)phenyl)amino)pyrimidin-4-yl)amino)-5-methylphenyl)dimethylphosphine oxide, which has a crystalline form 2, using Cu-Ka radiation and expressed in 2θ angles, the X-ray powder diffraction pattern of the crystalline form 2 has the following characteristic peaks: 5.72±0.2°, 10.24±0.2° , 11.44±0.2°, 15.07±0.2°, 15.09±0.2°, 15.92±0.2°, 16.41±0.2°, 17.07±0.2°, 17.38±0.2°, 18.49±0.2°, 19.49±0.2°, 20.07±0.2°, 21.59±0.2°, 22.41±0.2°, 23.25±0.2°, 23.55±0.2° and 25.19±0.2°.

[0012] In one or more embodiments, using Cu-Ka radiation and expressed in 2θ angles, the X-ray powder diffraction pattern of Form 2 has the following characteristic peaks: 5.72±0.2°, 10.24±0.2°, 11.44±0.2°, 15.07±0.2°, 15.09±0.2°, 15.92±0.2°, 16.41±0.2°, 17.07±0.2°, 17.38±0.2°, 18.49±0.2°, 18.71±0.2°, 0.2°, 19.49±0.2°, 20.07±0.2°, 21.59±0.2°, 22.41±0.2°, 23.05±0.2°, 23.25±0.2°, 23.55±0.2°, 24.71±0.2°, 25.19±0.2°, 26.36±0.2°, 27.22±0.2°, 27.51±0.2°, 28.45±0.2°, 29.49±0.2° and 29.95±0.2°.

[0013] In one or more embodiments, using Cu-Ka radiation and expressed in 2θ angles, the X-ray powder diffraction pattern of Form 2 is as follows: Figure 2 shown.

[0014] One or more embodiments provide anhydrous crystals of (2-((5-bromo-2-((2-methoxy-5-methyl-4-(4-(4-methylpiperazin-1-yl)piperidin-1-yl)phenyl)amino)pyrimidin-4-yl)amino)-5-methylphenyl)dimethylphosphine oxide, which has a crystalline form 3, using Cu-Ka radiation and expressed in 2θ angles, the X-ray powder diffraction pattern of the crystalline form 3 has the following characteristic peaks: 4.86±0.2°, 5.62±0.2°, 6.88 ±0.2°, 8.16±0.2°, 10.54±0.2°, 12.31±0.2°, 15.90±0.2°, 16.22±0.2°, 17.21±0.2°, 18.30±0.2°, 19.73±0.2°, 20.67±0.2°, 21.43±0.2°, 22.97±0.2°, 23.69±0.2°, 25.51±0.2°, 28.77±0.2° and 30.39±0.2°.

[0015] In one or more embodiments, using Cu-Ka radiation and expressed in 2θ angle, the X-ray powder diffraction pattern of the crystalline form 3 has the following characteristic peaks: 4.86±0.2°, 5.62±0.2°, 6.88±0.2°, 8.16±0.2°, 10.54±0.2°, 12.31±0.2°, 14.32±0.2°, 15.90±0.2°, 16.22±0.2°, 17.21±0. 2°, 18.30±0.2°, 19.73±0.2°, 20.67±0.2°, 21.43±0.2°, 22.97±0.2°, 23.69±0.2°, 25.51±0.2°, 27.59±0.2°, 28.07±0.2°, 28.77±0.2°, 29.09±0.2°, 30.39±0.2°, 34.55±0.2° and 35.32±0.2°.

[0016] In one or more embodiments, using Cu-Ka radiation and expressed in 2θ angles, the X-ray powder diffraction pattern of Form 3 is as follows: Figure 3 shown.

[0017] One or more embodiments provide a pharmaceutical composition comprising a dihydrate crystal having Form 1, an anhydrous crystal having Form 2, or an anhydrous crystal having Form 3 of the present application and a pharmaceutically acceptable carrier.

[0018] One or more embodiments provide a pharmaceutical composition comprising dihydrate crystals having crystalline form 1, anhydrous crystals having crystalline form 2, or anhydrous crystals having crystalline form 3 of the present application and other anticancer drugs or antitumor drugs.

[0019] In one or more embodiments, the anticancer drug or antitumor drug is one or more of a cytotoxic drug, a hormone drug, an antimetabolite drug, a tumor-targeted drug, a PARP inhibitor drug, an adjuvant therapy drug or an antitumor biological drug.

[0020] In one or more embodiments, the cytotoxic drug is one or more of carboplatin, cisplatin, irinotecan, paclitaxel, fluorouracil, cytarabine, lenalidomide, and retinoic acid.

[0021] In one or more embodiments, the hormone drug is one or more of dexamethasone, fulvestrant, and tamoxifen.

[0022] In one or more embodiments, the antimetabolite drug is one or more of fluorouracil, methotrexate, furanylfluorouracil, and cytarabine.

[0023] In one or more embodiments, the tumor-targeted drug is one or more of imatinib, erlotinib, and lapatinib.

[0024] In one or more embodiments, the PARP inhibitor drug is one or more of Olaparib, Rubraca, and Zejula.

[0025] In one or more embodiments, the adjuvant therapy drug is one or more of recombinant human granulocyte colony stimulating factor, erythropoietin, pamidronate disodium, and zoledronic acid.

[0026] In one or more embodiments, the anti-tumor biological drug is one or more of Keytruda, Opdivo, Tecentriq, Imfinzi, and Bavencio.

[0027] One or more embodiments provide use of the pharmaceutical composition of the present application having dihydrate crystals of crystal form 1, anhydrous crystals of crystal form 2, or anhydrous crystals of crystal form 3 in the preparation of a medicament for preventing and / or treating cancer.

[0028] In one or more embodiments, the cancer is plasmacytoma, mantle cell tumor, multiple myeloma, melanoma, breast cancer, liver cancer, cervical cancer, lung cancer, lymphoma, leukemia, ovarian cancer, kidney cancer, gastric cancer, nasopharyngeal cancer, thyroid cancer, pancreatic cancer, prostate cancer, adenocarcinoma, oral cancer, esophageal cancer, squamous cell carcinoma, or colon cancer.

[0029] One or more embodiments provide the use of the pharmaceutical composition of the present application having dihydrate crystals of Form 1, anhydrous crystals of Form 2, or anhydrous crystals of Form 3 in the preparation of EGFR inhibitors or ALK inhibitors or EGFR and ALK inhibitors or protein kinase inhibitors.

[0030] In one or more embodiments, the EGFR inhibitor or ALK inhibitor or EGFR and ALK inhibitor or protein kinase inhibitor is used to treat plasmacytoma, mantle cell tumor, multiple myeloma, melanoma, breast cancer, liver cancer, cervical cancer, lung cancer, lymphoma, leukemia, ovarian cancer, kidney cancer, gastric cancer, nasopharyngeal cancer, thyroid cancer, pancreatic cancer, prostate cancer, adenocarcinoma, oral cancer, esophageal cancer, squamous cell carcinoma or colon cancer.

[0031] In one or more embodiments, the EGFR has one or more mutations selected from the group consisting of L858R mutation, Del19 mutation, T790M mutation, and C797S mutation.

[0032] In one or more embodiments, the EGFR has a C797S mutation.

[0033] In one or more embodiments, the ALK has an EML-4-ALK fusion and / or an EML-4-ALK-L1196M mutation.

[0034] One or more embodiments provide a method for preparing a crystal of the dihydrate of the present application, comprising slurrying the compound (2-((5-bromo-2-((2-methoxy-5-methyl-4-(4-(4-methylpiperazin-1-yl)piperidin-1-yl)phenyl)amino)pyrimidin-4-yl)amino)-5-methylphenyl)dimethylphosphine oxide in a mixture of ethyl acetate and water at 10-30°C (e.g., 10°C, 15°C, 20°C, 25°C or 30°C), filtering, washing the filter cake with ethyl acetate, and then vacuum drying the filter cake to a moisture content of 5% to 6% by weight (based on the total weight of the filter cake) to obtain a crystalline substance.

[0035] In one or more embodiments, the weight ratio of ethyl acetate to water is 5:1-10:1, for example 5:1, 6:1, 7:1, 8:1, 9:1 or 10:1.

[0036] In one or more embodiments, the beating time is 30-120 minutes (eg, 30 minutes, 40 minutes, 50 minutes, 60 minutes, 70 minutes, 80 minutes, 90 minutes, 100 minutes, 110 minutes, or 120 minutes).

[0037] In one or more embodiments, the beating time is 50-70 minutes.

[0038] In one or more embodiments, vacuum drying is performed at a temperature of 15°C to 35°C (15°C, 20°C, 25°C, 30°C, or 35°C) and a pressure of -0.05 to -0.1 MPa (e.g., -0.05 MPa, -0.06 MPa, -0.07 MPa, -0.08 MPa, -0.09 MPa, or -0.1 MPa).

[0039] In one or more embodiments, vacuum drying is performed at a temperature of 20° C. to 30° C. and a pressure of −0.08 to −0.1 MPa.

[0040] In one or more embodiments, vacuum drying is performed to a moisture content of 4% to 7% (eg, 4%, 5%, 6% or 7%).

[0041] In one or more embodiments, vacuum drying is performed to a moisture content of 5% to 6%.

[0042] One or more embodiments provide a method for preparing the dihydrate crystal of the present application, which comprises a volatile crystallization method, a slurry crystallization method, an antisolvent crystallization method, and a cooling crystallization method.

[0043] In one or more embodiments, in the volatile crystallization method, the compound (2-((5-bromo-2-((2-methoxy-5-methyl-4-(4-(4-methylpiperazin-1-yl)piperidin-1-yl)phenyl)amino)pyrimidin-4-yl)amino)-5-methylphenyl)dimethylphosphine oxide is dissolved in a first solvent and / or a second solvent at a temperature of 10-40°C (e.g., 10°C, 15°C, 20°C, 25°C, 30°C, 35°C or 40°C), filtered, and the filtrate is evaporated to dryness to obtain a crystalline substance.

[0044] In one or more embodiments, in the volatile crystallization method, the temperature, the first solvent, the second solvent and their volume ratio are as shown in the following table:

[0045] temperature First solvent Secondary solvent Volume ratio of the first solvent to the second solvent 40℃ Methanol - - 10-30℃ Ethanol - - 40℃ Isopropyl alcohol - - 10-30℃ acetone - - 40℃ Ethyl acetate - - 10-30℃ Tetrahydrofuran - - 10-30℃ Chloroform - - 10-30℃ Methanol water 2.0 / 0.1 10-30℃ Ethanol water 3.0 / 0.1 10-30℃ acetone water 5.0 / 0.1 10-30℃ Tetrahydrofuran water 5.0 / 0.1 40℃ Methanol Ethyl acetate 2.0 / 1.0 10-30℃ Ethanol n-Heptane 3.0 / 1.0 10-30℃ acetone Methyl tert-butyl ether 9.0 / 3.0 10-30℃ Butanone Acetonitrile 8.0 / 3.0 10-30℃ Tetrahydrofuran n-Heptane 5.0 / 2.0 40℃ Chloroform Methylcyclohexane 1.0 / 1.0

[0046] In one or more embodiments, in the slurry crystallization method, the compound (2-((5-bromo-2-((2-methoxy-5-methyl-4-(4-(4-methylpiperazin-1-yl)piperidin-1-yl)phenyl)amino)pyrimidin-4-yl)amino)-5-methylphenyl)dimethylphosphine oxide is heated at a temperature of 4-40°C (4°C, 5°C, 6°C, 7°C, 8°C, 9°C, 10°C, 15°C, 20°C, 25°C, 30°C, 35°C or 40°C). , forming a suspension in the first solvent and / or the second solvent, and after 1-3 days (e.g., 1, 2 or 3 days) of crystal slurry, centrifuging, and drying the solid at a temperature of 20°C to 30°C (20°C, 21°C, 22°C, 23°C, 24°C, 25°C, 26°C, 27°C, 28°C, 29°C or 30°C) and a pressure of -0.08 to -0.1 MPa (e.g., -0.08 MPa, -0.09 MPa or -0.1 MPa) to obtain a crystalline substance.

[0047] In one or more embodiments, in the slurry crystallization method, the temperature, the first solvent, the second solvent and their volume ratio are as shown in the following table:

[0048]

[0049] In one or more embodiments, in the antisolvent crystallization method, the compound (2-((5-bromo-2-((2-methoxy-5-methyl-4-(4-(4-methylpiperazin-1-yl)piperidin-1-yl)phenyl)amino)pyrimidin-4-yl)amino)-5-methylphenyl)dimethylphosphine oxide is dissolved in a first solvent and then added to a second solvent at a temperature of 10°C to 30°C (e.g., 10°C, 15°C, 20°C, 25°C or 30°C), Alternatively, after being dissolved in the first solvent, the second solvent is added to the first solvent, and after the solid is precipitated, it is stirred and centrifuged. The solid is dried at a temperature of 20°C to 30°C (for example, 20°C, 21°C, 22°C, 23°C, 24°C, 25°C, 26°C, 27°C, 28°C, 29°C or 30°C) and a pressure of -0.08 to -0.1MPa (for example, -0.08MPa, -0.09MPa or -0.1MPa) to obtain a crystalline substance.

[0050] In one or more embodiments, in the antisolvent crystallization method, the first solvent, the second solvent and their volume ratio are as shown in the following table:

[0051]

[0052] In one or more embodiments, in the cooling crystallization method, the compound (2-((5-bromo-2-((2-methoxy-5-methyl-4-(4-(4-methylpiperazin-1-yl)piperidin-1-yl)phenyl)amino)pyrimidin-4-yl)amino)-5-methylphenyl)dimethylphosphine oxide is dissolved in a first solvent or a mixed solvent of a first solvent and a second solvent at an initial temperature, cooled to room temperature or 4°C, and centrifuged after solid precipitation. The solid is dried at a temperature of 20°C to 30°C (e.g., 20°C, 21°C, 22°C, 23°C, 24°C, 25°C, 26°C, 27°C, 28°C, 29°C or 30°C) and a pressure of -0.08 to -0.1 MPa (e.g., -0.08 MPa, -0.09 MPa or -0.1 MPa) to obtain a crystalline substance;

[0053] More preferably, the starting temperature, cooling method, and solvent are as described in the following table:

[0054]

[0055] The compound (2-((5-bromo-2-((2-methoxy-5-methyl-4-(4-(4-methylpiperazin-1-yl)piperidin-1-yl)phenyl)amino)pyrimidin-4-yl)amino)-5-methylphenyl)dimethylphosphine oxide has the following structure: It is a better EGFR and / or ALK inhibitor.

[0056] The dihydrate of the compound (2-((5-bromo-2-((2-methoxy-5-methyl-4-(4-(4-methylpiperazin-1-yl)piperidin-1-yl)phenyl)amino)pyrimidin-4-yl)amino)-5-methylphenyl)dimethylphosphine oxide has the following structure: It has good stability and solubility, is not hygroscopic, and can achieve a balance between various properties to meet the requirements of pharmaceutical crystal form. BRIEF DESCRIPTION OF THE DRAWINGS

[0057] Figure 1 This is the XRPD pattern of Form 1.

[0058] Figure 2 This is the XRPD pattern of Form 2.

[0059] Figure 3 This is the XRPD pattern of Form 3.

[0060] Figure 4 This is a purity detection diagram of Form 1.

[0061] Figure 5a This is the TGA spectrum of Form 1.

[0062] Figure 5b This is the TGA spectrum of Form 1.

[0063] Figure 6 This is the DSC spectrum of Form 1.

[0064] Figure 7 This is the FT-IR spectrum of Form 1.

[0065] Figure 8 This is the temperature-dependent XRPD pattern of Form 1.

[0066] Fig. 9 This is the XRPD pattern of Form 1 under accelerated test conditions for 6 months. DETAILED DESCRIPTION

[0067] Example 1-1: Preparation of (2-((5-bromo-2-((2-methoxy-5-methyl-4-(4-(4-methylpiperazin-1-yl)piperidin-1-yl)phenyl)amino)pyrimidin-4-yl)amino)-5-methylphenyl)dimethylphosphine oxide

[0068] Step 1: Preparation of DA01

[0069]

[0070] Into a 1.0L three-necked flask, add 2-iodo-4-methylaniline (DASM01, 96.0g, 0.412mol, 1.0eq), K2CO3 (79.68g, 0.577mol, 1.4eq), Xantphos (6.24g, 0.0165mol, 0.04eq) and DMF 576mL, replace with nitrogen twice, then add Pd(OAc)2 (3.7g, 0.0165mol, 0.04eq) and dimethylphosphine oxide (DASM02, 38.58g, 0.494mol, 1.2eq) under nitrogen bubbling, and continue to bubble nitrogen for 15min after the addition. After bubbling, heat to 100℃ in a nitrogen atmosphere for reaction until the reaction is complete (about 2-3h) by TLC detection. After the reaction is completed, the reaction solution is cooled to room temperature, 1.72L of water is added dropwise, and the mixture is stirred at room temperature for 30min, then filtered, and the filtrate is collected. 86.0g of sodium chloride is added to the filtrate and stirred to dissolve, and then extracted twice with 1.0L of dichloromethane, and the aqueous phase is extracted once with 1.0L of DCM / MeOH (10:1) mixed solvent. The organic phases are combined, dried over anhydrous sodium sulfate, and concentrated under reduced pressure to obtain an oily substance. 48mL of ethyl acetate is added to the oily substance and stirred evenly, and then 144mL of petroleum ether or n-heptane is added to slurry for 3h, filtered, and the filter cake is washed with 100mL of a mixed solvent of petroleum ether / ethyl acetate (1:1). The solid filter cake is then placed at 50°C and air-dried to obtain a total of 66.80g of DA01 product with a yield of 88.5%.1 H NMR (400MHz, DMSO-d6) δ=9.21 (brs, 2H), 7.57 (d, J=13.6, 1H), 7.43 (d, J=8.4, 1H), 7.26-7.23 (m, 1H), 2.33 (s, 3H), 1.85 (s, 3H), 1.82 (s, 3H).

[0071] Step 2: Preparation of DA02

[0072]

[0073] Add DA01 (50.0 g, 0.273 mol, 1.0 eq), K2CO3 (45.33 g, 0.328 mol, 1.2 eq) and 300 mL of DMSO into a 500 mL three-necked flask, stir evenly and add 5-bromo-2,4-dichloropyrimidine (DASM03, 68.36 g, 0.30 mol, 1.1 eq), and then place at 60°C to react until the reaction is complete as detected by TLC (about 3 hours of reaction). After the reaction is completed, cool the reaction solution to room temperature, add 900 mL of water dropwise, and a large amount of solid product is precipitated. Stir at room temperature for 2 hours, filter, wash the filter cake twice with 200 mL of water, collect the filter cake and place it at 60°C to dry with air, and obtain 94.0 g of DA02 product, with a yield of 92.00%. 1 H NMR (400MHz, CDCl3) δ=11.19 (s, 1H), 8.43 (dd, J=8.4, 4.8Hz, 1H), 8.32 (s, 1H ), 7.41 (d, J=8.8Hz, 1H), 7.08 (m, 1H), 2.38 (s, 3H), 1.86 (s, 3H), 1.83 (s, 3H). MS-ESI(m / z):395.9642(M+Na) + .

[0074] Step 3: Preparation of Compound DA

[0075]

[0076] Compound DA02 (32.08 g, 85.64 mmol), compound DASM04 (30 g, 94.20 mmol, 1.1 eq), trifluoroacetic acid (24.41 g, 214.09 mmol, 2.5 eq), and 256 mL of ethylene glycol monomethyl ether were added to a reaction bottle, and then the temperature was raised to 100 ° C for reaction until the reaction was completed by TLC detection (about 6 h of reaction). After the reaction, the reaction solution was cooled to room temperature, 256 mL of saturated sodium bicarbonate aqueous solution and 512 mL of water were added, and the mixture was stirred at room temperature for 1 h, then filtered and washed with water to obtain The filter cake was stirred with 150 mL of a mixed solvent of DCM / EA = 2 / 1 at room temperature for about 1 h, then filtered, and the filter cake was dissolved in 230 mL of methanol under reflux, stirred at room temperature for about 4 h, filtered, and the filter cake was washed with a small amount of methanol. The filter cake was then dried in vacuo at 40°C to 50°C to obtain a dry product of compound DA (i.e., (2-((5-bromo-2-((2-methoxy-5-methyl-4-(4-(4-methylpiperazin-1-yl)piperidin-1-yl)phenyl)amino)pyrimidin-4-yl)amino)-5-methylphenyl)dimethylphosphine oxide, 37.0 g, yield 65.8%). 1 H NMR (400MHz, CDCl3) δ = 10.29 (s, 1H), 8.32 (dd, J = 8.8, 4.8Hz, 1H), 8.18 (s, 1H) , 8.01 (s, 1H), 7.44-7.30 (m, 2H), 7.11 (d, J = 14.0Hz, 1H), 6.61 (s, 1H), 3.85 (s, 3H), 3.15 (d, J=11.6Hz, 2H), 2.80-2.52 (m, 11H), 2.38 (s, 3H), 2.35 (s, 3H), 2.1 4 (s, 3H), 1.96 (d, J=11.0Hz, 2H), 1.85 (s, 3H), 1.82 (s, 3H), 1.78-1.68 (m, 2H). MS-ESI(m / z):656.2459(M+H) + .

[0077] Example 1-2: Preparation of the crystalline form of the compound (2-((5-bromo-2-((2-methoxy-5-methyl-4-(4-(4-methylpiperazin-1-yl)piperidin-1-yl)phenyl)amino)pyrimidin-4-yl)amino)-5-methylphenyl)dimethylphosphine oxide dihydrate

[0078] Ethyl acetate (22.19 kg) and water (2.72 kg) (9:1) were added to a 30 L reactor, followed by the addition of compound DA (i.e., (2-((5-bromo-2-((2-methoxy-5-methyl-4-(4-(4-methylpiperazin-1-yl)piperidin-1-yl)phenyl)amino)pyrimidin-4-yl)amino)-5-methylphenyl)dimethylphosphine oxide, 1.88 kg), and slurrying was performed at 10-30 degrees Celsius for 50-70 minutes, followed by suction filtration, and the filter cake was washed with 3.07 kg of ethyl acetate. After washing, The wet filter cake was transferred to a vacuum drying oven and dried to a moisture content of 5% to 6% at a temperature of 20°C to 30°C and a pressure of -0.08 to -0.1 MPa. The material was collected to obtain a crystalline compound, i.e., (2-((5-bromo-2-((2-methoxy-5-methyl-4-(4-(4-methylpiperazin-1-yl)piperidin-1-yl)phenyl)amino)pyrimidin-4-yl)amino)-5-methylphenyl)dimethylphosphine oxide dihydrate, a total of 1.84 kg, purity: 99.66%, single impurity ≤ 0.06% (see the purity detection spectrum). Figure 4 ), melting point: 213℃~217℃.

[0079] Example 2: XRPD determination of (2-((5-bromo-2-((2-methoxy-5-methyl-4-(4-(4-methylpiperazin-1-yl)piperidin-1-yl)phenyl)amino)pyrimidin-4-yl)amino)-5-methylphenyl)dimethylphosphine oxide dihydrate crystal form:

[0080] Determine according to the Chinese Pharmacopoeia (2020 Edition) - Part IV - General Rule 0451 - X-ray Diffraction - Method 2 - Powder X-ray Diffraction:

[0081] Table 1. Powder XRPD test parameters

[0082] Device Name X-ray diffractometer (XRPD) model Bruker D8 Advance Diffractometer Scan Type Locked Coupled Scanning method Continuous Scan detector SSD160-2 X-ray Source Cu-Ka Diverging slits 0.6mm Voltage / Current 40KV, 40mA Step Length 0.02°2θ Scan speed 0.2sec / step Starting angle 3°

[0083] The XRPD pattern of (2-((5-bromo-2-((2-methoxy-5-methyl-4-(4-(4-methylpiperazin-1-yl)piperidin-1-yl)phenyl)amino)pyrimidin-4-yl)amino)-5-methylphenyl)dimethylphosphine oxide dihydrate crystalline form is as follows Figure 1 As shown, the XRPD data are shown in Table 2, and the crystal form is named as Crystal Form 1.

[0084] Table 2 XRPD data of Form 1

[0085]

[0086] Example 3: TGA determination of Form 1

[0087] Table 3. TGA test parameters:

[0088]

[0089] The TGA detection spectrum of Form 1 is as follows Figure 5a and Figure 5b shown.

[0090] The test results show that there is an obvious water loss step (weight loss step) between about 50-80°C in Form 1, and the weight loss is about 5.7%.

[0091] Example 4: DSC determination of Form 1

[0092] Table 4.DSC test parameters:

[0093]

[0094] The DSC detection spectrum of Form 1 is as follows Figure 6 The DSC spectrum shows that Form 1 has an obvious endothermic peak between about 50 and 80°C.

[0095] Example 5: Fourier transform infrared spectroscopy (FT-IR) determination of Form 1

[0096] Table 5. FT-IR test parameters:

[0097]

[0098] The FT-IR detection spectrum of Form 1 is as follows Figure 7 shown.

[0099] Example 6: Preparation of Form 2 of (2-((5-bromo-2-((2-methoxy-5-methyl-4-(4-(4-methylpiperazin-1-yl)piperidin-1-yl)phenyl)amino)pyrimidin-4-yl)amino)-5-methylphenyl)dimethylphosphine oxide

[0100] Take 30 mg of compound DA, add 1.0 mL of dichloromethane, dissolve, and then concentrate to dryness under reduced pressure on a rotary evaporator at 30°C to obtain a crystalline substance. Then, an X-ray powder diffraction (XRPD) test was immediately performed, and the XRPD spectrum is shown in Figure 2 , XRPD data are shown in Table 6, named as Form 2.

[0101] Table 6 XRPD data of Form 2

[0102] serial number 2θ strength serial number 2θ strength serial number 2θ strength 1 5.72 100％ 11 18.71 9.5％ 21 26.36 6.9％ 2 10.24 35.4％ 12 19.49 27.4％ 22 27.22 6％ 3 11.44 36.2％ 13 20.07 23.5％ 23 27.51 6.6％ 4 15.07 5.1％ 14 21.59 15.5％ 24 28.45 7％ 5 15.09 4.8％ 15 22.41 15.8％ 25 28.75 5.8％ 6 15.92 16.1％ 16 23.05 7.2％ 26 29.49 6.4％ 7 16.41 57.5％ 17 23.25 13.7％ 27 29.95 7.4％ 8 17.07 15.7％ 18 23.55 10.9％ 28 31.93 5.2％ 9 17.38 26.1％ 19 24.71 7.2％ 29 36.71 4％ 10 18.49 16.7％ 20 25.19 18.1％

[0103] Example 7: Preparation of Form 3 of (2-((5-bromo-2-((2-methoxy-5-methyl-4-(4-(4-methylpiperazin-1-yl)piperidin-1-yl)phenyl)amino)pyrimidin-4-yl)amino)-5-methylphenyl)dimethylphosphine oxide

[0104] Take 30 mg of compound DA, add 1.0 mL of trifluoroethanol, dissolve, and then concentrate to dryness under reduced pressure on a rotary evaporator at 30°C to obtain a crystalline substance. Then, an X-ray powder diffraction (XRPD) test was immediately performed, and the XRPD spectrum is shown in Figure 3 , XRPD data are shown in Table 7, named as Form 3.

[0105] Table 7 XRPD data of Form 3

[0106] serial number 2θ strength serial number 2θ strength serial number 2θ strength 1 4.86 17.4 10 17.21 100 19 28.07 10.3 2 5.62 11.1 11 18.3 31.5 20 28.77 16.7 3 6.88 12.2 12 19.73 59.9 21 29.09 11.1 4 8.16 11.6 13 20.67 34.7 22 30.39 15.6 5 10.54 21.9 14 21.43 59.6 23 30.79 7.6 6 12.31 13.6 15 22.97 38.3 24 32.95 9.3 7 14.32 8.3 16 23.69 26.9 25 34.55 10 8 15.92 32.1 17 25.51 20.4 26 35.32 10.3 9 16.22 24.1 18 27.59 10.2

[0107] Example 8: Determination of moisture content of Form 1 (Fischer method)

[0108] According to the "Chinese Pharmacopoeia" (2020 edition) Part IV - General Rules - 0832 - Method 1 Operation and determination:

[0109] Experimental instruments and equipment

[0110] Electronic balance: Model Sartorius BSA 224S

[0111] Moisture titrator: Model Metrohm 888 Titrando

[0112] Experimental Materials

[0113] Karl Fischer reagent: Shanghai MacLean Biochemical Technology Co., Ltd., batch number: C12156591

[0114] Solvent: Anhydrous methanol, purchased from Tedia, batch number: 710901

[0115] Test product information

[0116] Crystal form 1: prepared in Example 1-2

[0117] The measurement results are shown in Table 8:

[0118] Table 8

[0119]

[0120] According to Part IV of the Chinese Pharmacopoeia (2020 Edition) - General Rules - 0832 - Method 1, the water content of Form 1 was measured to be 5.40%, which corresponds to the TGA water loss in Example 3, both confirming that Form 1 contains 2 molecules of water.

[0121] Example 9: Determination of moisture content of Form 2 and Form 3 (Fischer method)

[0122] The determination was carried out in accordance with Part IV - General Rules - 0832 - Method 1 of the Chinese Pharmacopoeia (2020 edition).

[0123] Experimental instruments and equipment

[0124] Electronic balance: Model Sartorius BSA 224S

[0125] Moisture titrator: Model Metrohm888 Titrando

[0126] Experimental Materials

[0127] Karl Fischer reagent: Shanghai MacLean Biochemical Technology Co., Ltd., batch number: C12156591

[0128] Solvent: Anhydrous methanol, purchased from Tedia, batch number: 710901

[0129] Test product information

[0130] Form 2: prepared in Example 6

[0131] Form 3: prepared in Example 7

[0132] The measurement results are shown in Table 9:

[0133] Table 9

[0134]

[0135] According to Part IV - General Rules - 0832 - Method 1 of the Chinese Pharmacopoeia (2020 Edition), the water content of Form 1 was measured to be 1.18% and 0.51% for Form 2 and Form 3, respectively.

[0136] Example 10: Variable Temperature XRPD of Form 1

[0137] The crystal form 1 was placed on a hot stage XRPD detection instrument to test the XRPD spectra under different temperature conditions. Figure 8 .

[0138] When the temperature on the hot stage XRPD detection instrument is 50°C, the crystal form of Form 1 remains unchanged. When the temperature reaches 80°C and 120°C, Form 1 loses two crystal waters and becomes an unstable free-state crystal form that can only be monitored on the hot stage XRPD detection instrument. Once the temperature is cooled to room temperature, the unstable free-state crystal form is transformed into Form 1 again, confirming that the free-state crystal form has poor stability and is easily transformed into Form 1 at room temperature.

[0139] Example 11: Other preparation methods of Form 1

[0140] (1) Volatile crystallization method

[0141] Experimental operation: For the compound DA sample, solvent 1 or a mixed solvent of solvent 1 and solvent 2 was added at the temperature shown in Table 10, and the solid sample was ultrasonically dissolved, then filtered, and the filtrate was exposed to evaporate to dryness at the corresponding temperature, and the solid was taken for characterization. The characterization results are shown in Table 10.

[0142] Table 10

[0143]

[0144] Note: The room temperature here refers to 10-30℃, such as 20℃, 25℃.

[0145] (2) Slurry crystallization method

[0146] Experimental operation: Take about 30 mg of compound DA sample, add the corresponding solvent 1 or a mixed solvent of solvent 1 / solvent 2 at the temperature shown in Table 11 to obtain a suspension. After 3 days of crystallization, centrifuge and dry the solid overnight at a temperature of 20°C to 30°C and a pressure of -0.08 to -0.1 MPa. Then take the solid for XRPD characterization. The characterization results are shown in Table 11.

[0147] Table 11

[0148]

[0149]

[0150] Note: The room temperature here refers to 10-30℃, such as 20℃, 25℃.

[0151] (3) Antisolvent crystallization method

[0152] Experimental operation: Take about 20 mg of compound DA sample and add solvent 1 to obtain a clear solution. Slowly add solvent 2 to the clear solution of solvent 1 under stirring (positive addition), or add the clear solution of solvent 1 to solvent 2 (reverse addition). After solids are precipitated, stir overnight, centrifuge, vacuum dry overnight at room temperature, and take solids for XRPD characterization; if no crystals are precipitated, transfer to 4°C and stir overnight. After solids are precipitated, centrifuge. Under the conditions of temperature of 20°C to 30°C and pressure of -0.08 to -0.1MPa, dry the solids overnight and perform XRPD characterization. The characterization results are shown in 12.

[0153] Table 12

[0154]

[0155] Note: The room temperature here refers to 10-30℃, such as 20℃, 25℃.

[0156] (4) Cooling crystallization method

[0157] Experimental operation: Take about 20 mg of compound DA sample, add the corresponding solvent 1 or a mixed solvent of solvent 1 / solvent 2, form a solution at the starting temperature (60°C), then stir at the ending temperature, stir overnight after the solid precipitates, centrifuge, dry the solid overnight at a temperature of 20°C to 30°C and a pressure of -0.08 to -0.1 MPa, and take the solid for XRPD characterization. The results are shown in Table 13.

[0158] Table 13

[0159]

[0160] Example 12: Moisture absorption comparison experiment

[0161] According to the "Guidelines for Hygroscopicity of Drugs" in Part 4 of the "Chinese Pharmacopoeia" (2020 Edition), hygroscopicity tests were carried out on compound DA, Form 1 (prepared in Example 1-2), Form 2 and Form 3 obtained in the above examples.

[0162] The hygroscopic characteristics and hygroscopic weight gain are defined as follows:

[0163] Deliquesce: Absorbs enough water to form a liquid;

[0164] Highly hygroscopic: weight gain due to moisture absorption is not less than 15%;

[0165] Hygroscopic: weight gain due to moisture absorption is less than 15% but not less than 2%;

[0166] Slightly hygroscopic: weight gain due to moisture absorption is less than 2% but not less than 0.2%;

[0167] No or almost no hygroscopicity: weight gain due to moisture is less than 0.2%.

[0168] The measurement results are shown in Table 14:

[0169] Table 14

[0170]

[0171] Conclusion: Compound DA is hygroscopic, Form 2 and Form 3 are slightly hygroscopic, and Form 1 is non- or almost non-hygroscopic. Hygroscopicity will affect the mixing uniformity and content uniformity of the formulation process development. Form 1 can maintain better fluidity, compressibility and uniformity during the formulation process compared to Form 2, Form 3 and Compound DA, and is more conducive to formulation development.

[0172] Example 13: Vacuum drying stability test

[0173] After the crystal form 1 obtained in the above Example 1-2 was vacuum dried at 35°C, it was found that the crystal form 1 did not undergo transformation. The water content of the crystal form 1 after vacuum drying at 35°C for 24 hours was measured to be 5.26%.

[0174] Example 14: Stability Test

[0175] (1) Stability Test 1 of Form 1 (Prepared in Example 1-2)

[0176] Experimental operation: About 30 mg of Form 1 solid sample was taken, placed in a centrifuge tube, and left open at room temperature in different solvent atmospheres. After a period of time (4th day and 6th day), the solid was taken for XRPD characterization, and the results are shown in Table 15. From Table 15, it can be seen that the crystal form of Form 1 remained unchanged after being left in different solvent atmospheres for a period of time.

[0177] Table 15

[0178]

[0179] (2) Stability test 2 of crystal form 1

[0180] About 20 mg of Form 1 (prepared in Example 1-2) was placed in a sample bottle, placed under different temperature and humidity conditions for a period of time, and the solid was taken for XRPD characterization. The results are shown in Table 16. As can be seen from Table 16, Form 1 maintains the same crystal form under different humidity (low humidity and high humidity) and different temperature (low temperature and high temperature) conditions.

[0181] Table 16

[0182] condition Day 4 Results Analysis Day 6 Results Analysis Day 11 Results Analysis RT-12%RH Crystal form 1 Crystal form 1 Crystal form 1 RT-58%RH Crystal form 1 Crystal form 1 Crystal form 1 RT-97%RH Crystal form 1 Crystal form 1 Crystal form 1 25℃-60%RH Crystal form 1 Crystal form 1 Crystal form 1 40℃-75%RH Crystal form 1 Crystal form 1 Crystal form 1

[0183] (3) Stability test 3 of crystal form 1

[0184] The stability of Form 1 (prepared in Example 1-2) was investigated under long-term and accelerated conditions. The results showed that Form 1 remained unchanged after being placed under long-term and accelerated conditions for 10 days and 6 months, as shown in Table 17 and Table 17-1. Fig. 9 .

[0185] Table 17

[0186]

[0187] Table 17-1

[0188]

[0189] According to the 2020 edition of the Chinese Pharmacopoeia and the ICHQ1A stability study guidelines, the crystal form 1 obtained in Example 1-2 was placed under long-term (25°C ± 2°C, 65% RH ± 10% RH) and accelerated (40°C ± 2°C, 75% RH ± 10% RH) conditions for 6 months, and the properties, maximum single impurities and total impurities of the crystal form 1 were detected. The results showed that the crystal form 1 was placed under long-term and accelerated conditions for 6 months, and the stability was good, the properties did not change, the maximum single impurity was ≤0.07%, which was much smaller than the quality control limit of 0.15% required by the Chinese Pharmacopoeia and ICH guidelines. At the same time, the total impurities did not increase compared with day 0 after 6 months of placement, meeting the stability requirements of the pharmaceutical crystal form.

[0190] At the same time, after being placed under long-term conditions for 30 days, the properties of Form 2 and Form 3 changed significantly, and did not meet the requirements for pharmaceutical crystal forms (according to the Chinese Pharmacopoeia 2020 edition and ICHQ1A stability study guidelines, Form 2 and Form 3 were placed under long-term conditions (25℃±2℃, 65%RH±10%RH) for only 1 month, and their properties changed from off-white solid powder to light yellow solid powder, which was a significant change), see Table 17-2.

[0191] Table 17-2

[0192]

[0193] (4) High temperature and light stability test of crystal forms 1-3

[0194] The stability of Form 1 (prepared in Example 1-2), Form 2, and Form 3 was investigated under illumination conditions. Under illumination conditions, the properties of Form 2 and Form 3 changed significantly, both changing from off-white solid powders to light yellow solid powders, while the properties of Form 1 did not change. The test results are shown in Table 18-1.

[0195] Table 18-1

[0196]

[0197] The results show that the crystal stability of Form 1 under light conditions is better than that of Form 2 and Form 3.

[0198] The crystal stability of Form 1 under high temperature and light conditions was investigated, and the results are shown in Table 18-2, indicating that Form 1 has good crystal stability after being placed under high temperature and light conditions for 30 days, and no significant changes were observed in properties, single impurities and total impurities. The crystal stability of Forms 2 and 3 under high temperature, light and long-term conditions was investigated, and the results are shown in Tables 18-3 and 18-4. The experimental results show that Forms 2 and 3 are placed under high temperature and light conditions for 10 days or 30 days, and the properties change significantly from off-white solid powder to light yellow solid powder. In addition, the impurity growth of Forms 2 and 3 is particularly fast under high temperature and light conditions (Form 2 grows from 0.63% on the 0th day to 2.0% on the 30th day under light conditions, and Form 3 grows from 0.56% on the 0th day to 3.05% on the 30th day under light conditions), indicating that their stability is poor.

[0199] Table 18-2

[0200]

[0201] Table 18-3

[0202]

[0203] Table 18-4

[0204]

[0205] Example 15: Solubility Test

[0206] Weigh the finely powdered compound DA and Form 1 (prepared in Example 1-2), Form 2 and Form 3 samples, add them to a certain volume of solvent at 25°C±2°C, shake vigorously for 30 seconds every 5 minutes, and test the solubility for 24 hours.

[0207] The solubility test showed that in a solution with a pH of 6.8, compared with the amorphous compound DA, crystal form 2 and crystal form 3, the solubility of crystal form 1 is better, see Table 18-5.

[0208] Table 18-5

[0209]

[0210] Taking into account the above three properties of hygroscopicity, stability and solubility, compared with compound DA, Form 1 is superior in both hygroscopicity and solubility, which is an unexpected effect; compared with Forms 2 and 3, Form 1 has better hygroscopicity, stability and solubility, and is therefore the dominant form.

[0211] Example 16: Preparation of control compounds

[0212] Preparation of reference compound 1

[0213] Reference compound 1A (intermediate):

[0214]

[0215] In a 250mL three-necked flask, add 2-amino-6-methylphenyl-dimethylphosphine oxide (4.52g, 24.67mmol), 2,4,5-trichloropyrimidine (4.98g, 27.14mmol), K2CO3 (4.11g, 29.60mmol), 27mL DMSO, and heat to 60°C for about 3.5 hours until the reaction is complete by TLC detection. The reaction is cooled to room temperature, 90mL H2O is added, and a large amount of yellow solid is precipitated. Stirring is continued for about 0.5h and then filtered. The filter cake is slurried with 25mL petroleum ether / ethyl acetate = 4 / l at room temperature for 4h and then filtered and dried to obtain compound 1A (6.60g, yield 81.0%). MS-ESI (m / z): 332.1483 (M+H) + .

[0216] Control compound 1:

[0217]

[0218] Compound 1A (600 mg, 1.82 mmol), compound 2-methoxy-5-methyl-4-(4-(1-methylpiperidin-4-yl)piperazine-1-yl)aniline (636 mg, 2.00 mmol), 15% hydrogen chloride ethanol solution (1.33 g, 5.46 mmol), and 9.0 mL of ethylene glycol monomethyl ether were added to a sealed tube, followed by reaction at 100°C for 7-8 h. After completion of the reaction as detected by TLC, the reaction solution was cooled to room temperature, and then 18 mL of water and 9.0 mL of saturated NaHCO3 aqueous solution were added to precipitate a large amount of solid. The solid was allowed to stand for 0.5 hour and then filtered. The filter cake was washed with water, dried, and then slurried and purified with a mixed solvent of 4 ml EA and 1.0 mL methanol to obtain reference compound 1 (500 mg, yield 45.5%). 1H NMR (400 MHz, methanol-d4) δ = 8.02 (s, 1H), 7.93 (dd, J = 8.0, 4.0 Hz, 1H), 7.64 (s, 1H), 7.42 (t, J = 8.0 Hz, 1H), 7.15 (dd, J = 7.6, 3.6 Hz, 1H), 6.69 (s, 1H), 3.84 (s, 3H), 3.11 (d, J=11.6Hz, 2H), 3.02-2.41 (m, 13H), 2.32 (m, 4H), 2.04 (s, 3H), 2.00 (d, J=12. 6Hz, 2H), 1.94 (s, 3H), 1.90 (s, 3H), 1.68 (m, 2H). MS-ESI (m / z): 612.2981 (M+H)+.

[0219] Preparation of reference compound 2

[0220] Reference compound 2A (intermediate):

[0221]

[0222] In a 100mL three-necked flask, (2-aminophenyl)-dimethylphosphine oxide (2.5g, 15.00mmol), 2,4,5-trichloropyrimidine (2.70g, 15.00mmol), K2CO3 (2.45g, 67.75mmol), nBu4NHSO4 (0.5g, 1.50mmol), 50mL DMF were added, and the temperature was raised to 65°C for about 4.5 hours until the reaction was complete by TLC detection. The reaction solution was then cooled to room temperature, 200mLH2O was added, and a large amount of yellow solid precipitated. After stirring for about 0.5h, the mixture was filtered, and the filter cake was washed with 100mL H2O and dried to obtain the reference compound 2A (2.84g, yield 60.0%). MS-ESI (m / z): 316.0178 (M+H) + .

[0223] Control compound 2:

[0224]

[0225] The reference compound 2A (335 mg, 1.06 mmol), 2-methoxy-5-methyl-4-(4-(1-methylpiperidin-4-yl)piperazine-1-yl)aniline (406 mg, 1.27 mmol), 15% hydrogen chloride in ethanol (774 mg, 3.18 mmol) and 4.5 mL of ethylene glycol monomethyl ether were added to the reaction bottle, and then the tube was sealed and reacted at 120°C for 5-6 hours. After the reaction was completed by TLC detection, the reaction solution was cooled to room temperature, and then 15 mL of saturated NaHCO3 aqueous solution was added to precipitate a large amount of solid. The mixture was stirred for 0.5 hours and then filtered. The filter cake was washed with water and the filter cake was purified by slurrying with 9 mL of EtOH / H2O=1 / 2 mixed solvent to obtain the reference compound 2 (320 mg, yield 50.5%). 1 H NMR (400MHz, CDCl3) δ = 10.80 (s, 1H), 8.63 (dd, J = 8.4, 4.4Hz, 1H), 8.10 (s, 1H), 8.04 (s, 1H), 7.51 (m, 1H), 7.38-7.26 (m, 2H), 7.13 (m, 1H), 6.63 (s, 1H), 3.86 (s, 3H), 3.16 (d, J=12.0Hz, 2H), 2.61 (m, 9H), 2.32 (s, 3H), 2.18 (s, 3H), 2.06 (m, 2H), 1.96 (m, 2H), 1.87 (s, 3H), 1.83 (s, 3H), 1.72 (m, 2H). MS-ESI (m / z): 598.2826 (M+H) + .

[0226] Preparation of reference compound 3

[0227] Reference compound 3A (intermediate):

[0228]

[0229] 5-Fluoro-2-nitroanisole (2.0 g, 11.69 mmol), 1-methyl-4-(4-piperidinyl)piperazine (2.57 g, 14.02 mmol), potassium carbonate (3.25 g, 23.37 mmol), and 30 mL of DMF were added to a 100 mL round-bottom flask, and the temperature was raised to 120° C. for about 3 hours until the reaction was complete as detected by TLC. The reaction solution was cooled to room temperature, 30 mL of water was added, and extracted with 30 mL×3 ethyl acetate, the organic phases were combined, washed with 30 mL of saturated sodium chloride aqueous solution, dried over anhydrous sodium sulfate, filtered, and concentrated to obtain the control compound 3A (3.57 g, yield 91.3%).

[0230] Reference compound 3B (intermediate):

[0231]

[0232] The reference compound 3A (3.57 g, 10.68 mmol), 0.36 g 5% Pd / C, 72 mL methanol, hydrogen replacement 2-3 times, stirred at room temperature under hydrogen atmosphere (normal pressure) overnight until the reaction was completed by TLC detection, then filtered with diatomaceous earth, the filtrate was vacuum concentrated, and column chromatography was used for separation and purification to obtain the reference compound 3B (2.17 g, yield 66.8%). MS-ESI (m / z): 305.2315 (M+H) + . Reference compound 3C (intermediate):

[0233]

[0234] In a 500mL three-necked flask, 2-iodo-4-methylaniline (10g, 42.9mmol), K3PO4 (10.9g, 51.5mmol), Xantphos (2.48g, 4.3mmol), Pd(OAc)2 (0.96g, 4.3mmol), dimethylphosphine (5g, 64.4mmol), and 100mL DMF were added, and the temperature was raised to 120°C for about 3h until the reaction was complete by TLC detection. The reaction solution was cooled to room temperature, filtered, and 600mL H2O was added to precipitate a large amount of yellow solid, which was filtered, and the filtrate was extracted with 500mL×3 ethyl acetate. The organic phases were combined, washed with 250mL×2 saturated sodium chloride aqueous solution, dried over anhydrous sodium sulfate, filtered, and the filtrate was concentrated to obtain compound 3C, which was continued to the next step without purification. 1H NMR (400MHz, DMSO-d6) δ=9.21 (brs, 2H), 7.57 (d, J=13.6, 1H), 7.43 (d, J=8.4, 1H), 7.26-7.23 (m, 1H), 2.33 (s, 3H), 1.85 (s, 3H), 1.82 (s, 3H).

[0235] Reference compound 3D (intermediate):

[0236]

[0237] In a 500mL three-necked flask, add the reference compound 3C obtained in the previous step, 2,4,5-trichloropyrimidine (11.80g, 64.3mmol), K2CO3 (7.76g, 128.73mmol), nBu4NHSO4 (1.45g, 4.29mmol), 100mL DMF, and heat to 65°C for about 4.5 hours until the reaction is complete as detected by TLC. The reaction is cooled to room temperature, 200mL H2O is added, and a large amount of yellow solid is precipitated. Stirring is continued for about 0.5h, and then filtered, the filter cake is washed with 100mL H2O, and dried to obtain compound 3D (8.57g, total yield of two steps 60.5%). 1H NMR (400MHz, DMSO-d6) δ=11.62 (s, 1H), 8.40 (s, 1H), 8.27 (dd, J=8.4, 4.4Hz, 1H), 7.44 (m, 2H), 2.33 (s, 3H), 1.81 (s, 3H), 1.77 (s, 3H).

[0238] Control compound 3:

[0239]

[0240] The reference compound 3D (400 mg, 1.21 mmol), the reference compound 3B (443 mg, 1.45 mmol), 15% hydrogen chloride ethanol solution (884 mg, 3.63 mmol), and 6 mL of ethylene glycol monomethyl ether were added to the reaction bottle, and the tube was sealed and reacted at 120°C for 5-6 h until the reaction was completed by TLC detection. The reaction solution was then cooled to room temperature, 6 mL of saturated NaHCO3 aqueous solution and 6 mL of H2O were added, and extracted with 30 mL×3 dichloromethane, and the organic phases were combined and washed with 20 mL of saturated sodium chloride aqueous solution, dried over anhydrous sodium sulfate, filtered, concentrated, and purified by column chromatography (DCM / MeOH=8 / 1), and 5 mL of PE / EA=4 / 1 mixed solvent was used to obtain the reference compound 3 (257 mg, yield 35.4%). 1 H NMR (400MHz, CDCl3) δ=10.63 (s, 1H), 8.49 (dd, J=8.8, 4.8Hz, 1H), 8.19-7.99 (m, 2H), 7.31 (d, J=8.8Hz, 1H), 7.25 (s, 1H), 7.07 (d, J=14.0Hz, 1H), 6.61-6. 44(m, 2H), 3.87(s, 3H), 3.66(d, J=12.0Hz, 2H), 2.66(m, 9H), 2.38(s, 3H), 2. 32(s, 3H), 2.21(s, 2H), 1.97(m, 2H), 1.85(s, 3H), 1.81(s, 3H), 1.73(m, 2H). MS-ESI (m / z): 598.2804 (M+H)+ .

[0241] Example 17: In vitro activity test of compound DA

[0242] Experimental Example 1 Cell Antiproliferation Experiment 1

[0243] 1. Experimental purpose: To detect the effect of the test compound on the proliferation of BaF3 cell lines containing EGFR mutations

[0244] 2. Experimental information:

[0245] 2.1. Cell information: BaF3 cell line with EGFR mutation is from WuXi AppTec:

[0246] BaF3-expressing EGFR mutation cell line

[0247] EGFR-T790M / Del19

[0248] EGFR-C797S / Del19

[0249] EGFR-T790M / L858R

[0250] EGFR-C797S / L858R

[0251] EGFR-C797S / T790M / L858R

[0252] EGFR-C797S / T790M / Del19

[0253] EGFR-WT wild-type cell line

[0254] Experimental methods

[0255] Day 0: Seeding

[0256] a. Turn on the UV light of the biosafety cabinet and start the countdown for 30 minutes.

[0257] b. Preheat the culture medium in a 37°C water bath.

[0258] c. After UV irradiation, open the biosafety cabinet. Wipe the preheated culture medium, PBS, etc. with alcohol and place them in the biosafety cabinet.

[0259] d. Take the cells out of the incubator and pipette them into a uniform cell suspension in a biosafety cabinet before counting.

[0260] e. According to the cell counting results, adjust the cell suspension density to 3000 cells per well and seed 50 μl per well in a 384-well plate.

[0261] f. Incubate the seeded cells in a 37°C, 5% CO2 incubator for 3 hours.

[0262] g. Add compounds to the cell plate using Tecan (liquid handling station).

[0263] Day 3: Place the cell plates and CTG (CellTiter-Glo chemiluminescent cell viability detection reagent) with the added compounds at room temperature for equilibrium, then add 25ul of CTG to each well, centrifuge at 1000 rpm for 1min, and shake for 1-2min. Centrifuge again at 1000 rpm for 1min, let stand for 10min, and then detect the signal value in Envision. XL-Fit analysis software was used to calculate the IC value of each compound by computer fitting. 50 (Read the concentration of the compound corresponding to the 50% inhibition rate to get the IC value of the compound on cell activity inhibition 50 ). Inhibition rate % = (reading value of control group without drug addition - reading value of sample) / reading value of control group without drug addition × 100

[0264] 3. Experimental results: IC values ​​of compound DA, control compound and the main drugs for the treatment of non-small cell lung cancer on the market, brigatinib and osimertinib, on the activity inhibition of Ba / F3 cells expressing EGFR-T790M / Del19, EGFR-C797S / Del19, EGFR-T790M / L858R, EGFR-C797S / L858R, EGFR-C797S / T790M / L858R, EGFR-C797S / T790M / Del19 and EGFR-WT 50 The values ​​are shown in Tables 19 and 20.

[0265] 4. Conclusion: From Tables 19 and 20, we can see that compound DA has good selectivity for the activity of Ba / F3 cells expressing EGFR wild-type (EGFR-WT), and has a good inhibitory effect on the proliferation of Ba / F3 cells expressing EGFR double mutation and triple mutation (such as EGFR-T790M / Del19, EGFR-C797S / Del19, EGFR-T790M / L858R, EGFR-C797S / L858R, EGFR-C797S / T790M / L858R, EGFR-C797s / T790M / Del19), especially the inhibitory activity on Ba / F3 cell lines expressing EGFR double mutation and triple mutation is much better than that of control compounds 1, 2, 3 and the marketed drugs brigatinib and osimertinib.

[0266] Table 19

[0267]

[0268] Table 20

[0269]

[0270] Experimental Example 2 Cell Anti-Proliferation Experiment 2

[0271] 1. Experimental purpose: To detect the effect of the test compound on the proliferation of PC9 cell lines containing EGFR mutations

[0272] 2. Experimental information:

[0273] 2.1. Cell information: PC9 cells carry EGFRDel19 mutation, and other mutant cells were constructed from PC9 cells according to the commonly used stable cell line construction method.

[0274] PC9-cell line expressing EGFR mutation

[0275] EGFR-Del19

[0276] EGFR-T790M / De119

[0277] EGFR-C797S / T790M / Del19

[0278] 2.2 Experimental methods:

[0279] The test compound was diluted 5 times at an appropriate concentration as the experimental starting concentration, and a total of 6 concentration gradients were obtained. Brigatinib (purchased from Selleck) was diluted 3 times at a starting concentration of 5 μM, and a total of 6 concentration gradients were obtained. The test compound and brigatinib were added to the above cells, incubated at 37°C and 5% CO2 for 72 hours, and the SRB (Sulforhodamine B) detection method was used to read the optical density value of each well at a wavelength of 490 nm using an enzyme reader.

[0280] The cell optical density value at 0 drug action is set as the Tz value, which represents the value of the cell when the drug is added. The cell optical density value of the solvent control DMSO for 72 hours is set as the C value; the cell optical density value of the test compound for 72 hours is set as the Ti value. The cell response to the drug is calculated according to the method proposed by the US NIH-NCI (National Institutes of Health-National Cancer Institute): when Ti ≥ Tz, it is [(Ti-Tz) / (C-Tz)]×100; when Ti<Tz, it is [(Ti-Tz) / Tz]×100. Based on the above calculations, the 4ParameterLogistic Model in the XLfit software was used to calculate GI 50 The concentration of the test compound required to inhibit cell growth by 50% was expressed as follows:

[0281] 3. Experimental results: GI effects of compound DA, control compound and brigatinib, the main drug for the treatment of non-small cell lung cancer on PC9 cells expressing EGFR-Del19, EGFR-T790M / Del19, and EGFR-C797S / T790M / Del19 50 The values ​​are shown in Table 21.

[0282] 4. Conclusion: From Table 21, we can see that compound DA has a good inhibitory effect on the proliferation activity of PC9 cells expressing EGFR single mutation, double mutation and triple mutation (such as EGFR-Del19, EGFR-T790M / Del19, EGFR-C797S / T790M / Del19), and the inhibitory activity is much better than that of control compounds 1, 2, 3 and the marketed drug brigatinib.

[0283] Table 21

[0284]

[0285] Experimental Example 3 Cell Antiproliferation Experiment 3

[0286] 1. Experimental purpose: To detect the effect of the test compound on the proliferation of BaF3 cell lines containing ALK mutations

[0287] 2. Experimental information:

[0288] 2.1. Cell information: BaF3 cells with ALK mutation were obtained from WuXi AppTec

[0289] Ba / F3-cell lines expressing ALK gene fusions and mutations

[0290] Ba / F3-EML-4-ALK-WT

[0291] Ba / F3-EML-4-ALK-L1196M

[0292] Experimental methods

[0293] Day 0: Seeding

[0294] a. Turn on the UV light of the biosafety cabinet and start the countdown for 30 minutes.

[0295] b. Preheat the culture medium in a 37°C water bath.

[0296] c. After UV irradiation, open the biosafety cabinet. Wipe the preheated culture medium, PBS, etc. with alcohol and place them in the biosafety cabinet.

[0297] d. Take the cells out of the incubator and pipette them into a uniform cell suspension in a biosafety cabinet before counting.

[0298] e. According to the cell counting results, adjust the cell suspension density to 3000 cells per well and seed 50 μl per well in a 384-well plate.

[0299] f. Incubate the seeded cells in a 37°C, 5% CO2 incubator for 3 hours.

[0300] g. Add compounds to the cell plate using Tecan (liquid handling station).

[0301] Day 3: The cell plates and CTG with added compounds were placed at room temperature for equilibrium, and then 25ul of CTG was added to each well. The plates were centrifuged at 1000 rpm for 1min and then shaken for 1-2min. The plates were centrifuged at 1000 rpm for 1min and then left to stand for 10min before the signal value was detected in Envision. The IC values ​​of each compound were calculated by computer fitting using XL-Fit analysis software. 50 (Read the concentration of the compound corresponding to the 50% inhibition rate to get the IC value of the compound on cell activity inhibition 50 ). Inhibition rate % = (reading value of control group without drug addition - reading value of sample) / reading value of control group without drug addition × 100

[0302] 3. Experimental results: IC values ​​of compound DA and control compound on BaF3 cells of EML-4-ALK-WT and EML-4-ALK-L1196M 50 The values ​​are shown in Table 22.

[0303] 4. Conclusion: From Table 22, we can see that compound DA has a very good inhibitory effect on the proliferation of BaF3 cells expressing EML-4-ALK-WT and EML-4-ALK-L1196M, and its inhibitory activity is better than that of control compounds 1, 2, and 3.

[0304] Table 22

[0305]

[0306] Experimental Example 4 Cell Antiproliferation Experiment 4

[0307] 1. Experimental purpose: To detect the effect of the test compound on the proliferation of BaF3 cell lines containing ALK mutations

[0308] 2. Experimental Materials

[0309] BaF3 EML-4-ALK-L1196M cell line, Brand: Kangyuan Bochuang, Catalog No.: KC-0102

[0310] Cell Counting Kit-8 (CCK-8), Brand: Targetmol, Catalog No.: C0005

[0311] Multifunctional microplate reader POLARstar Omega, brand: BMG LABTECH

[0312] 3. Experimental Methods

[0313] Day 0: Seeding

[0314] a. Take the cells out of the incubator and pipette them into a uniform cell suspension in a clean bench before counting.

[0315] b. According to the cell counting results, the cell suspension density was adjusted and 110 μL per well (containing 8,000 cells) was inoculated in a 96-well plate.

[0316] c. Add 10 μL of the serially diluted compound to the cell plate to make the final concentration of the compound 500, 166.67, 55.56, 18.52, 6.17, 2.06, 0.69, 0.23, and 0.08 nM.

[0317] d. Incubate the cell plate in a 37°C, 5% CO2 incubator for 72 h.

[0318] Day 3: After CCK8 was equilibrated at room temperature, 10 μL of CCK8 was added to each well. The cell plate was incubated in an incubator at 37°C and 5% CO2 for 2 h, and the signal value OD was detected in the multifunctional microplate reader POLARstar Omega. 450 The IC values ​​of each compound were calculated by computer fitting using GraphPadprism analysis software. 50 (Read the concentration of the compound corresponding to the 50% inhibition rate to get the IC value of the compound on cell activity inhibition 50 ). Inhibition rate % = (reading value of control group without drug addition - reading value of sample) / (reading value of control group without drug addition - reading value of control group with only culture medium) × 100

[0319] 4. Experimental results: IC of compound DA and compound 1 of patent WO2021073498A1 on BaF3 cells expressing EML-4-ALK-L1196M 50 The values ​​are shown in Table 23.

[0320] Conclusion: From Table 23, we can see that compound DA has a very good inhibitory effect on the proliferation of BaF3 cells expressing EML-4-ALK-L1196M, and its inhibitory activity is better than that of compound 1 of patent WO2021073498A1.

[0321] Table 23: Inhibitory effects of compound DA and compound 1 of patent WO2021073498A1 on BaF3 EML-4-ALK-L1196M cells

[0322] Compound <![CDATA[First test IC 50 (nM)]]> <![CDATA[Second trial IC 50 (nM)]]> Compound DA 15.14 10.38 Patent WO2021073498A1 Compound 1 81.45 74.61

[0323] Example 18: In vivo pharmacodynamics experiment of compound DA crystal form 1

[0324] The model used in this experiment is an engineered BaF3 EML-4-ALK-L1196M cell BALB / c nude mouse subcutaneous transplant tumor model. Engineered BaF3 cells were cultured in vitro in suspension. BaF3 EML-4-ALK-L1196M culture conditions were GIBCO1640 medium with 10% fetal bovine serum and cultured in a 37°C 5% CO2 incubator. The cells were processed and passaged twice a week. When the number of cells reached the requirement, the cells were collected, counted, and diluted with PBS to a concentration of 1×10 7 / ml tumor cell suspension was placed in an ice box and taken to the animal room for direct injection. Use a 1ml syringe to take the above prepared concentration of 1×10 7 / ml tumor cell suspension was injected subcutaneously under the axilla on the left forelimb of the mouse, with 0.25ml injected at each inoculation site. When the average tumor volume reached about 160mm 3 The drug administration started at 5 pm, with 15 mice in the vehicle group and 10 mice in each of the other drug administration groups.

[0325] Dosage method and frequency: Oral gavage, administration volume 10ml / kg, the model group was given an equal volume of solvent. Administer once a day for 14 consecutive days. The dose of Form 1 (prepared in Example 1-2) is 30mg / kg and 40mg / kg. At the same molar dose, the dose of Compound 1 of Patent WO2021073498A1 is 30.2mg / kg and 40.2mg / kg.

[0326] The general conditions of the mice, such as spirit, activity, and eating, were observed every day, and their body weight was measured three times a week. The short diameter (a) and long diameter (b) of the tumor of each mouse were measured with a vernier caliper three times a week, and the tumor volume was calculated according to the formula (a2×b) / 2. The relative tumor volume (RTV) was calculated based on the measured and calculated tumor volume, RTV=Vt / V0. V0 is the tumor volume at the time of random grouping (i.e., d0), and Vt is the tumor volume at each measurement (i.e., dn). The relative tumor proliferation rate, an indicator for evaluating antitumor activity, was calculated according to the following formula: relative tumor proliferation rate T / C (%):

[0327] (Note: TRTV: RTV of treatment group; CRTV: RTV of model control group. According to the evaluation standard of efficacy in the "Technical Guidelines for Non-clinical Research of Cytotoxic Anti-tumor Drugs" issued by the China National Drug Administration, T / C% ≤ 40% is effective).

[0328] Table 24: Average tumor volume of each group at different time points in the BaF3 EML-4-ALK-L1196M nude mouse transplant tumor model

[0329]

[0330] Note: a. Days after drug administration;

[0331] Table 25: Relative tumor proliferation rate T / C (%) of BaF3 EML-4-ALK-L1196M nude mouse transplant tumor model

[0332]

[0333] ▲: original RTV value compared with the RTV value of the vehicle control group, p < 0.05, ▲▲: p < 0.01;

[0334] ★: original RTV value compared with the same molar dose of crystal form 1 group p < 0.05, ★★: p < 0.01;

[0335] The above test results show that:

[0336] In this experiment, the doses of Form 1 at 30 mg / kg and 40 mg / kg could significantly inhibit the growth of BaF3 EML-4-ALK-L1196M transplanted tumors, and there was a dose-effect relationship between the dose groups. At the end of the D14 experiment, the average tumor volume of the tumor-bearing mice in the solvent control group was 1947 mm 3 The T / C% of the crystalline form 1 at doses of 30 mg / kg and 40 mg / kg were both less than 40%, 4.85% and 2.34% respectively, and the tumor volumes were 101 mm 3 and 49mm 3 (p values ​​compared with the vehicle control group were: <0.0001 and <0.0001, respectively).

[0337] The doses of 30.2 mg / kg and 40.2 mg / kg of compound 1 of patent WO2021073498A1 had no significant inhibitory effect on the growth of BaF3 EML-4-ALK-L1196M transplanted tumors; at the end of the D14 experiment, their T / C% were 126.04% and 80.71%, respectively, both of which did not meet the effective standard; their tumor volumes were 2547 mm 3 and 1605mm 3 .

[0338] The inhibitory effect of crystalline form 1 at doses of 30 mg / kg and 40 mg / kg on the growth of BaF3 EML-4-ALK-L1196M transplanted tumors was significantly stronger than that of the patented WO2021073498A1 compound 1 at doses of 30.2 mg / kg and 40.2 mg / kg at the same molar dose (T / C% of D14: 4.85% vs. 126.04%, p<0.0001; 2.34% vs. 80.71%, p<0.0001).

Claims

1. A crystal of (2-((5-bromo-2-((2-methoxy-5-methyl-4-(4-(4-methylpiperazin-1-yl)piperidin-1-yl)phenyl)amino)pyrimidin-4-yl)amino)-5-methylphenyl)dimethylphosphine oxide dihydrate, having a crystalline form 1, wherein the X-ray powder diffraction pattern of the crystalline form 1 has the following characteristic peaks using Cu-Ka radiation and expressed in 2θ angles: 5.54±0.2°, 11.06°±0.2, 19.12°±0.2 and 27.90±0.2°.

2. The crystal of claim 1, wherein the X-ray powder diffraction pattern of the crystalline form 1 using Cu-Ka radiation and expressed in 2θ angles has the following characteristic peaks: 5.54±0.2°, 11.06±0.2°, 16.15±0.2°, 16.73±0.2°, 17.10±0.2°, 19.12±0.2°, 21.18±0.2°, 24.16±0.2°, 26.27±0.2°, 26.48±0.2°, 27.90±0.2° and 33.64±0.2°.

3. The crystal according to claim 1, wherein the X-ray powder diffraction pattern of the crystalline form 1 has the following characteristic peaks using Cu-Ka radiation and expressed in 2θ angles: 5.54±0.2°, 8.50±0.2°, 10.53±0.2°, 11.06±0.2°, 11.48±0.2°, 13.00±0.2°, 16.15±0.2°, 16.73±0.2°, 17.10±0.2°, 19.12±0.2°, 19.47±0.2°, 21.18±0.2°, 24.16±0.2°, 26.27±0.2°, 26.48±0.2°, 27.90±0.2° and 33.64±0.2°.

4. The crystal according to claim 1, wherein the X-ray powder diffraction pattern of the crystalline form 1 has the following characteristic peaks using Cu-Ka radiation and expressed in 2θ angles: 5.54±0.2°, 8.50±0.2°, 10.53±0.2°, 11.06±0.2°, 11.48±0.2°, 13.00±0.2°, 16.15±0.2°, 16.73±0.2°, 17.10±0.2°, 18.34±0.2°, 19.12±0.2° , 19.47±0.2°, 21.18±0.2°, 22.21±0.2°, 22.65±0.2°, 23.10±0.2°, 23.49±0.2°, 24.16±0.2°, 26.27±0.2°, 26.48±0.2°, 26.93±0.2°, 27.35±0.2°, 27.90±0.2°, 29.35±0.2°, 32.22±0.2°, 33.64±0.2°, and 34.17±0.2°.

5. The crystal according to claim 1, wherein Cu-Ka radiation is used and expressed in 2θ angles, and the X-ray powder diffraction pattern of the crystal form 1 is shown in Figure 1.

6. A pharmaceutical composition comprising the dihydrate crystal according to any one of claims 1 to 5 and a pharmaceutically acceptable carrier.

7. A pharmaceutical composition comprising the dihydrate crystal according to any one of claims 1 to 5 and other anti-tumor drugs.

8. The pharmaceutical composition according to claim 7, wherein the anti-tumor drug is one or more of a cytotoxic drug, a hormone drug, an antimetabolite drug, a tumor-targeted drug, a PARP inhibitor drug, or an anti-tumor biological drug.

9. The pharmaceutical composition according to claim 7, wherein the anti-tumor drug is an adjuvant therapy drug.

10. The pharmaceutical composition according to claim 8, wherein the cytotoxic drug is one or more of carboplatin, cisplatin, irinotecan, paclitaxel, fluorouracil, cytarabine, lenalidomide, and retinoic acid; and the hormone drug is one or more of dexamethasone, fulvestrant, and tamoxifen.

11. The pharmaceutical composition according to claim 8, wherein the antimetabolite drug is one or more of fluorouracil, methotrexate, furanylfluorouracil, and cytarabine.

12. The pharmaceutical composition according to claim 8, wherein the tumor targeting drug is one or more of imatinib, erlotinib, and lapatinib.

13. The pharmaceutical composition according to claim 8, wherein the PARP inhibitor drug is one or more of Olaparib, Rubraca, and Zejula.

14. The pharmaceutical composition according to claim 9, wherein the adjuvant therapy drug is one or more of recombinant human granulocyte colony stimulating factor, erythropoietin, pamidronate disodium, and zoledronic acid.

15. The pharmaceutical composition according to claim 8, wherein the anti-tumor biological drug is one or more of Keytruda, Opdivo, Tecentriq, Imfinzi, and Bavencio.

16. Use of the dihydrate crystal according to any one of claims 1 to 5 or the pharmaceutical composition according to any one of claims 6 to 15 in the preparation of a medicament for preventing and / or treating cancer.

17. The method of claim 16, wherein the cancer is plasmacytoma, mantle cell tumor, multiple myeloma, melanoma, breast cancer, liver cancer, cervical cancer, lung cancer, lymphoma, leukemia, ovarian cancer, kidney cancer, gastric cancer, nasopharyngeal cancer, thyroid cancer, pancreatic cancer, prostate cancer, adenocarcinoma, oral cancer, esophageal cancer, squamous cell carcinoma or colon cancer.

18. Use of the dihydrate crystal according to any one of claims 1 to 5 or the pharmaceutical composition according to any one of claims 6 to 15 in the preparation of an EGFR inhibitor or an ALK inhibitor or an EGFR and ALK inhibitor.

19. The use of claim 18, wherein the EGFR inhibitor or ALK inhibitor or EGFR and ALK inhibitor is used to treat plasmacytoma, mantle cell tumor, multiple myeloma, melanoma, breast cancer, liver cancer, cervical cancer, lung cancer, lymphoma, leukemia, ovarian cancer, kidney cancer, gastric cancer, nasopharyngeal cancer, thyroid cancer, pancreatic cancer, prostate cancer, adenocarcinoma, oral cancer, esophageal cancer, squamous cell carcinoma or colon cancer.

20. The use according to claim 18, wherein the EGFR has one or more mutations selected from the group consisting of L858R mutation, Del19 mutation, T790M mutation and C797S mutation.

21. The use according to claim 20, wherein the EGFR has a C797S mutation.

22. The use according to claim 18, wherein the ALK has EML-4-ALK fusion and / or EML-4-ALK-L1196M mutation.

23. A method for preparing a crystal of the dihydrate of any one of claims 1 to 5, comprising slurrying the compound (2-((5-bromo-2-((2-methoxy-5-methyl-4-(4-(4-methylpiperazin-1-yl)piperidin-1-yl)phenyl)amino)pyrimidin-4-yl)amino)-5-methylphenyl)dimethylphosphine oxide in a mixture of ethyl acetate and water at 10-30° C., filtering, washing the filter cake with ethyl acetate, and then vacuum drying the filter cake to a moisture content of 5% to 6% to obtain a crystalline substance; The weight ratio of ethyl acetate to water is 5:1-10:1; The beating time is 30-120 minutes; vacuum drying at a temperature of 15°C to 35°C and a pressure of -0.05 to -0.1 MPa; Vacuum dry to 4% to 7% moisture.

24. The method of claim 23, wherein the weight ratio of ethyl acetate to water is 9:

1.

25. The method of claim 23, wherein the beating time is 50-70 minutes.

26. The method of claim 23, wherein the drying is performed under vacuum at a temperature of 20°C to 30°C and a pressure of -0.08 to -0.1 MPa.

27. The method of claim 23, wherein the mixture is vacuum dried to a moisture content of 5% to 6%.

28. A method for preparing the dihydrate crystals according to any one of claims 1 to 5, comprising a volatilization crystallization method, a slurry crystallization method, an antisolvent crystallization method, and a cooling crystallization method; In the volatile crystallization method, the compound (2-((5-bromo-2-((2-methoxy-5-methyl-4-(4-(4-methylpiperazin-1-yl)piperidin-1-yl)phenyl)amino)pyrimidin-4-yl)amino)-5-methylphenyl)dimethylphosphine oxide is dissolved in a first solvent and / or a second solvent at a temperature of 10-40° C., filtered, and the filtrate is evaporated to dryness to obtain a crystalline substance; The temperature, the first solvent, the second solvent and their volume ratio are as shown in the following table: In the slurry crystallization method, the compound (2-((5-bromo-2-((2-methoxy-5-methyl-4-(4-(4-methylpiperazin-1-yl)piperidin-1-yl)phenyl)amino)pyrimidin-4-yl)amino)-5-methylphenyl)dimethylphosphine oxide is suspended in a first solvent and / or a second solvent at a temperature of 4-40° C. After slurrying for 1-3 days, the suspension is centrifuged, and the solid is dried at a temperature of 20° C. to 30° C. and a pressure of -0.08 to -0.1 MPa to obtain a crystalline substance; The temperature, the first solvent, the second solvent and their volume ratio are as shown in the following table: In the antisolvent crystallization method, the compound (2-((5-bromo-2-((2-methoxy-5-methyl-4-(4-(4-methylpiperazin-1-yl)piperidin-1-yl)phenyl)amino)pyrimidin-4-yl)amino)-5-methylphenyl)dimethylphosphine oxide is dissolved in a first solvent at a temperature of 10°C to 30°C and then added to a second solvent, or the compound (2-((5-bromo-2-((2-methoxy-5-methyl-4-(4-(4-methylpiperazin-1-yl)piperidin-1-yl)phenyl)amino)pyrimidin-4-yl)amino)-5-methylphenyl)dimethylphosphine oxide is dissolved in a first solvent ... first solvent, and after a solid is precipitated, the solid is stirred and centrifuged, and the solid is dried at a temperature of 20°C to 30°C and a pressure of -0.08 to -0.1 MPa to obtain a crystalline substance; The first solvent, the second solvent and their volume ratio are as shown in the following table: In the cooling crystallization method, the compound (2-((5-bromo-2-((2-methoxy-5-methyl-4-(4-(4-methylpiperazin-1-yl)piperidin-1-yl)phenyl)amino)pyrimidin-4-yl)amino)-5-methylphenyl)dimethylphosphine oxide is dissolved in a first solvent or a mixed solvent of a first solvent and a second solvent at an initial temperature, cooled to room temperature or 4° C., solids are precipitated and then centrifuged, and the solids are dried at a temperature of 20° C. to 30° C. and a pressure of -0.08 to -0.1 MPa to obtain a crystalline substance; The starting temperature, cooling method, and solvent are as follows:

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