Crystal form of benzothiophene substituted fused ring compound and preparation method thereof

Abstract

The invention relates to a crystal form of a benzothiophene substituted fused ring compound and a preparation method thereof. Specifically, the invention provides a crystal form A and a crystal form D of 2-amino-7-fluoro-4-((5S, 5aS, 6S, 9R)-1-fluoro-12-((1-((4-(fluoromethylsubunit) piperidine-1-yl) methyl) cyclopropyl) methoxyl)-5-methyl-5a, 6, 7, 8, 9, 10-hexahydro-5H-4-oxa-3, 10a, 11, 13, 14-pentaaza-6, 9-methylbridged naphtho [1, 8-ab] heptano-2-yl) benzo [b] thiophene-3-formonitrile, and the crystal form A and the crystal form D have the advantages that the crystal form A and the crystal form D are good in stability; and the medicine can be better used for clinical treatment. # imgabs0 #

Description

Technical Field

[0001] The present invention belongs to the field of medical technology and relates to a crystalline form of a benzothiophene-substituted condensed ring compound and a preparation method thereof. Background Art

[0002] RAS is one of the oncogenes with the highest mutation rate in tumors, and about 30% of human malignant tumors are related to mutations in the RAS gene. The RAS family includes KRAS, NRAS, and HRAS, among which KRAS mutations are the most common, accounting for about 85%. KRAS mutations are common in solid tumors, and high-frequency mutations exist in the three major fatal human cancers: lung cancer (17%), colorectal cancer (33%), and pancreatic cancer (61%). Among the KRAS gene mutations, 97% are mutations in the 12th or 13th amino acid residues, among which G12D is an important mutation. Data analysis of European and American populations shows that in pancreatic cancer, colorectal cancer, and non-small cell lung cancer, G12D mutations account for 36%, 12%, and 4% of patients, respectively.

[0003] After KRAS is activated, it regulates cell proliferation, survival, migration, metabolism and other functions through numerous downstream signaling pathways represented by RAF-MEK-ERK, PI3K-AKT-mTOR and TIAM1-RAc. After the KRAS gene mutates, the protein remains in an activated state, resulting in the continuous activation of downstream signaling pathways and promoting tumorigenesis.

[0004] PCT / CN2023 / 109598 provides a KRAS G12D inhibitor, whose chemical name is 2-amino-7-fluoro-4-((5S,5aS,6S,9R)-1-fluoro-12-((1-((4-(fluoromethylidene)piperidin-1-yl)methyl)cyclopropyl)methoxy)-5-methyl-5a,6,7,8,9,10-hexahydro-5H-4-oxa-3,10a,11,13,14-pentaaza-6,9-methylnaphtho[1,8-ab]heptylcyclo-2-yl)benzo[b]thiophene-3-carbonitrile, having a structure shown in Formula 1,

[0005]

[0006] The crystal form of the active pharmaceutical ingredient often affects the chemical stability of the drug. Different crystallization conditions and storage conditions may lead to changes in the crystal structure of the compound, and sometimes other forms of crystals may be produced. Generally speaking, amorphous drug products do not have a regular crystal structure and often have other defects, such as poor product stability, fine crystallization, difficult filtration, easy agglomeration, poor fluidity, etc. The polymorphic form of the drug has different requirements for product storage, production and scale-up. Therefore, it is necessary to conduct in-depth research on the crystal form of the aforementioned compound and improve the various properties of the aforementioned compound. Summary of the invention

[0007] The present disclosure provides a new crystal form of a compound represented by Formula 1, which has good stability and can be better applied in clinical practice.

[0008]

[0009] The crystalline form A of the compound represented by Formula 1 provided in the present disclosure has an X-ray powder diffraction pattern represented by a diffraction angle 2θ, and has characteristic peaks at 8.984, 12.800, 14.133, 14.675, 18.977, and 21.018.

[0010] In some embodiments, the crystalline form A of the compound represented by Formula 1 has an X-ray powder diffraction pattern represented by a diffraction angle 2θ, with characteristic peaks at 8.984, 9.837, 12.800, 14.133, 14.675, 15.944, 16.635, 17.595, 18.977, 21.018, and 26.006.

[0011] In some embodiments, Form A of the compound represented by Formula 1 has an X-ray powder diffraction pattern represented by a diffraction angle 2θ, with characteristic peaks at 8.984, 9.837, 12.800, 14.133, 14.675, 15.944, 16.635, 17.595, 18.977, 21.018, 22.089, 23.026, 26.006, and 29.233.

[0012] In some embodiments, the crystalline form A of the compound represented by Formula 1 has an X-ray powder diffraction pattern represented by a diffraction angle 2θ as shown in Figure 1 shown.

[0013] The present disclosure also provides a method for preparing the crystalline form A of the compound represented by Formula 1, wherein the method is selected from any one of the following methods:

[0014] Method 1: dissolving the compound of formula 1 in solvent I and volatilizing the solvent, wherein the solvent I is selected from methanol, 10% water / methanol, and 50% acetonitrile / methanol;

[0015] Method 2: Add the compound of formula 1 into solvent II and stir, wherein the solvent II is selected from water, acetonitrile, n-heptane, cyclohexane, isopropyl ether, 7% water / ethanol, and 50% water / methanol.

[0016] The crystalline form B of the compound represented by Formula 1 provided in the present disclosure has an X-ray powder diffraction pattern represented by a diffraction angle 2θ, and has characteristic peaks at 4.651, 8.536, 11.909, 14.077, 15.584, and 20.969.

[0017] In some embodiments, Form B of the compound represented by Formula 1 has an X-ray powder diffraction pattern represented by a diffraction angle 2θ, with characteristic peaks at 4.651, 8.536, 9.290, 11.909, 14.077, 15.584, 17.365, 18.878, 20.969, and 22.281.

[0018] In some embodiments, the crystalline form B of the compound represented by Formula 1 has an X-ray powder diffraction pattern represented by a diffraction angle 2θ as shown in Figure 2 shown.

[0019] The present disclosure also provides a method for preparing a crystalline form B of the compound of Formula 1, the method comprising the steps of dissolving the compound of Formula 1 in tetrahydrofuran / ethanol (2:1), and heating and cooling at 40-5°C.

[0020] The present disclosure also provides a method for preparing Form B of the compound of Formula 1, the method comprising the steps of dissolving the compound of Formula 1 in ethyl acetate / ethanol (1:1) and volatilizing the solvent.

[0021] The present disclosure also provides a method for preparing the crystalline form B of the compound of Formula 1, the method comprising the steps of adding the amorphous form of the compound of Formula 1 to ethanol and stirring.

[0022] The crystalline form C of the compound represented by Formula 1 provided in the present disclosure has an X-ray powder diffraction pattern represented by a diffraction angle 2θ, and has characteristic peaks at 8.666, 14.121, 18.947, 20.764, and 29.128.

[0023] In some embodiments, the crystalline form C of the compound represented by Formula 1 has an X-ray powder diffraction pattern represented by a diffraction angle 2θ, with characteristic peaks at 8.666, 9.348, 12.575, 14.121, 16.942, 18.947, 20.764, 29.128, and 33.304.

[0024] In some embodiments, Form C of the compound represented by Formula 1 has an X-ray powder diffraction pattern represented by a diffraction angle 2θ, with characteristic peaks at 8.666, 9.348, 12.575, 14.121, 16.942, 18.947, 20.764, 22.780, 23.932, 25.469, 29.128, and 33.304.

[0025] In some embodiments, the crystalline form C of the compound represented by Formula 1 has an X-ray powder diffraction pattern represented by a diffraction angle 2θ as shown in Figure 3 shown.

[0026] The present disclosure also provides a method for preparing a crystalline form C of a compound of Formula 1, the method comprising the steps of adding the compound of Formula 1 to an alcohol solvent I and stirring, wherein the alcohol solvent I is selected from ethanol, isopropanol, and n-propanol.

[0027] The crystalline form D of the compound represented by Formula 1 provided in the present disclosure has characteristic peaks at 7.229, 9.137, 11.664, 16.506 and 20.682 in an X-ray powder diffraction pattern represented by a diffraction angle 2θ.

[0028] In some embodiments, the crystalline form D of the compound represented by Formula 1 has an X-ray powder diffraction pattern represented by a diffraction angle 2θ, with characteristic peaks at 5.265, 7.229, 9.137, 11.664, 16.506, 17.396, 20.682, 21.849, and 23.001.

[0029] In some embodiments, the crystalline form D of the compound represented by Formula 1 has an X-ray powder diffraction pattern represented by a diffraction angle 2θ as shown in Figure 4 shown.

[0030] The present disclosure also provides a method for preparing the crystalline form D of the compound represented by Formula 1, wherein the method is selected from any one of the following methods:

[0031] Method 1: Dissolve the compound of formula 1 in 2-methyltetrahydrofuran, add methyl tert-butyl ether, and stir;

[0032] Method 2: Add the compound of formula 1 into methyl tert-butyl ether or 50% ethyl acetate / n-heptane and stir.

[0033] The crystalline form E of the compound represented by Formula 1 provided in the present disclosure has an X-ray powder diffraction pattern represented by a diffraction angle 2θ, and has characteristic peaks at 5.144, 8.390, 10.312, 13.575, and 20.130.

[0034] In some embodiments, the crystalline form E of the compound represented by Formula 1 has an X-ray powder diffraction pattern represented by a diffraction angle 2θ, with characteristic peaks at 5.144, 8.390, 10.312, 13.575, 14.845, 15.568, 16.293, 17.776, 18.488, and 20.130.

[0035] In some embodiments, the crystalline form E of the compound represented by Formula 1 has an X-ray powder diffraction pattern represented by a diffraction angle 2θ, and has characteristic peaks at 5.144, 8.390, 10.312, 13.575, 14.845, 15.568, 16.293, 17.776, 18.488, 20.130, 21.350, 22.029, 23.036, and 31.408.

[0036] In some embodiments, the crystalline form E of the compound represented by Formula 1 has an X-ray powder diffraction pattern represented by a diffraction angle 2θ as shown in Figure 5 shown.

[0037] The present disclosure also provides a method for preparing the crystalline form E of the compound represented by Formula 1, wherein the method is selected from any one of the following methods:

[0038] Method 1: dissolving the compound of formula 1 in solvent II, adding solvent III, and stirring, wherein the solvent II is selected from acetone, ethyl acetate, tetrahydrofuran / ethanol (2:1), and the solvent III is selected from MTBE and n-heptane;

[0039] Method 2: Add the compound of formula 1 into 50% ethyl acetate / n-heptane or acetone and stir;

[0040] Method three: dissolving the compound of formula 1 in solvent IV and stirring, wherein the solvent IV is selected from ethyl acetate / methyl tert-butyl ether (v / v=1:5), 2-methyltetrahydrofuran, and dichloromethane.

[0041] The crystalline form F of the compound represented by Formula 1 provided in the present disclosure has an X-ray powder diffraction pattern represented by a diffraction angle 2θ, and has characteristic peaks at 4.782, 5.122, 9.989, 15.495, and 20.134.

[0042] In some embodiments, the crystalline form F of the compound represented by Formula 1 has an X-ray powder diffraction pattern represented by a diffraction angle 2θ, with characteristic peaks at 4.782, 5.122, 9.989, 13.135, 14.171, 15.495, 16.552, 17.396, and 20.134.

[0043] In some embodiments, the crystalline form F of the compound represented by Formula 1 has an X-ray powder diffraction pattern represented by a diffraction angle 2θ as shown in Figure 6 shown.

[0044] The present disclosure also provides a method for preparing a crystalline form F of a compound of Formula 1, the method comprising the steps of dissolving the compound of Formula 1 in dichloromethane, adding n-heptane, and stirring.

[0045] The crystalline form G of the compound represented by Formula 1 provided in the present disclosure has an X-ray powder diffraction pattern represented by a diffraction angle 2θ, and has characteristic peaks at 10.692, 16.093, 16.447, 17.922, 24.567, and 33.426.

[0046] In some embodiments, the crystalline form G of the compound represented by Formula 1 has an X-ray powder diffraction pattern represented by a diffraction angle 2θ, with characteristic peaks at 7.724, 8.701, 10.692, 12.719, 13.325, 16.093, 16.447, 17.922, 19.340, 20.746, 21.219, 24.567, and 33.426.

[0047] In some embodiments, the crystalline form G of the compound represented by Formula 1 has an X-ray powder diffraction pattern represented by a diffraction angle 2θ, and has characteristic peaks at 7.724, 8.701, 10.692, 12.719, 13.325, 16.093, 16.447, 17.922, 19.340, 19.891, 20.746, 21.219, 22.886, 23.577, 24.567, 25.386, 26.330, and 33.426.

[0048] In some embodiments, the crystalline form G of the compound represented by Formula 1 has an X-ray powder diffraction pattern represented by a diffraction angle 2θ as shown in Figure 7 shown.

[0049] The present disclosure also provides a method for preparing the crystalline form G of the compound of Formula 1, the method comprising the steps of adding the amorphous form of the compound of Formula 1 into methanol and stirring.

[0050] The crystalline form H of the compound represented by Formula 1 provided in the present disclosure has an X-ray powder diffraction pattern represented by a diffraction angle 2θ, and has characteristic peaks at 5.099, 7.096, 9.872, 15.784, and 19.440.

[0051] In some embodiments, the crystalline form H of the compound represented by Formula 1 has an X-ray powder diffraction pattern represented by a diffraction angle 2θ, with characteristic peaks at 5.099, 7.096, 8.528, 9.872, 11.100, 13.212, 15.784, 19.440, 20.621, 21.120, and 22.425.

[0052] Formula 1

[0053] In some embodiments, the crystalline form H of the compound represented by Formula 1 has an X-ray powder diffraction pattern represented by a diffraction angle 2θ as shown in Figure 8 shown.

[0054] The present disclosure also provides a method for preparing the crystalline form H of the compound represented by Formula 1, wherein the method is selected from any one of the following methods:

[0055] Method 1: Add the amorphous compound of formula 1 into solvent V and stir, wherein the solvent V is selected from p-xylene, methyl tert-butyl ether, and ethyl acetate;

[0056] Method 2: Dissolve the amorphous compound of formula 1 in dichloromethane, add it into n-heptane and stir.

[0057] Method 3: adding the amorphous compound of formula 1 to ethyl acetate, and then adding a ligand, followed by stirring, wherein the ligand is selected from citric acid ethanol solution, tartaric acid ethanol solution, and fumaric acid.

[0058] The crystalline form I of the compound represented by Formula 1 provided in the present disclosure has an X-ray powder diffraction pattern represented by a diffraction angle 2θ, and has characteristic peaks at 4.992, 7.006, 7.552, 15.706, 17.272, 17.696, and 21.247.

[0059] In some embodiments, the crystalline form I of the compound represented by Formula 1 has an X-ray powder diffraction pattern represented by a diffraction angle 2θ, with characteristic peaks at 4.992, 7.006, 7.552, 8.416, 10.072, 13.212, 14.363, 15.706, 17.272, 17.696, and 21.247.

[0060] In some embodiments, the crystalline form I of the compound represented by Formula 1 has an X-ray powder diffraction pattern represented by a diffraction angle 2θ, and has characteristic peaks at 4.992, 7.006, 7.552, 8.416, 10.072, 11.177, 13.212, 14.363, 15.706, 16.515, 17.272, 17.696, 19.361, 19.968, 21.247, 22.809, and 23.500.

[0061] In some embodiments, the crystalline form I of the compound represented by Formula 1 has an X-ray powder diffraction pattern represented by a diffraction angle 2θ as shown in Fig. 9 shown.

[0062] The present disclosure also provides a method for preparing the crystalline form I of the compound of formula 1, the method comprising adding the compound of formula 1 to ethyl acetate, then adding an acetic acid ethanol solution, and stirring.

[0063] The crystal form J of the compound represented by Formula 1 provided in the present disclosure has an X-ray powder diffraction pattern represented by a diffraction angle 2θ, and has characteristic peaks at 7.645, 14.939, 18.701, 20.045, and 23.539.

[0064] In some embodiments, the crystalline form J of the compound represented by Formula 1 has an X-ray powder diffraction pattern represented by a diffraction angle 2θ, with characteristic peaks at 7.645, 8.682, 14.939, 17.243, 18.701, 20.045, 23.539, 24.498, and 27.032.

[0065] In some embodiments, the crystalline form J of the compound represented by Formula 1 has an X-ray powder diffraction pattern represented by a diffraction angle 2θ as shown in Fig.10 shown.

[0066] The present disclosure also provides a method for preparing Form J of the compound of Formula 1, the method comprising heating Form I of the compound of Formula 1 to 215°C.

[0067] In certain embodiments, the preparation method described in the present disclosure further comprises any step of crystallization, centrifugation (filtration), washing or drying.

[0068] The crystallization method of the present disclosure includes but is not limited to stirring crystallization, static crystallization or volatile crystallization. In some embodiments, the crystallization is stirring crystallization. In some embodiments, the crystallization is static crystallization.

[0069] The present disclosure also provides a pharmaceutical composition, which contains the aforementioned crystal form A, crystal form B, crystal form C, crystal form D, crystal form E, crystal form F, crystal form G, crystal form H, crystal form I or crystal form J, and a pharmaceutical excipient selected from pharmaceutically acceptable excipients.

[0070] The present disclosure also provides a pharmaceutical composition, which is prepared from the aforementioned crystal form A, crystal form B, crystal form C, crystal form D, crystal form E, crystal form F, crystal form G, crystal form H, crystal form I or crystal form J, and an optional pharmaceutically acceptable excipient.

[0071] The present disclosure also provides a method for preparing a pharmaceutical composition, comprising the step of mixing the aforementioned crystal form A, crystal form B, crystal form C, crystal form D, crystal form E, crystal form F, crystal form G, crystal form H, crystal form I or crystal form J with a pharmaceutically acceptable excipient.

[0072] The present disclosure also provides the use of the aforementioned crystalline form A, crystalline form B, crystalline form C, crystalline form D, crystalline form E, crystalline form F, crystalline form G, crystalline form H, crystalline form I or crystalline form J or the aforementioned composition in the preparation of a composition for preventing and / or treating cancer.

[0073] The use disclosed herein, wherein the cancer is selected from breast cancer, ovarian cancer, pancreatic cancer, prostate cancer, gastric cancer, colorectal cancer, lung cancer, kidney cancer, liver cancer, cervical cancer, endometrial cancer, epithelial cancer, esophageal cancer, neuroblastoma, glioma, bone cancer, nasopharyngeal cancer, oral cancer, thyroid cancer, skin cancer, squamous cell carcinoma; preferably, the cancer is selected from breast cancer, ovarian cancer, pancreatic cancer, prostate cancer, gastric cancer, colorectal cancer and lung cancer.

[0074] The "2θ or 2θ angle" mentioned in the present disclosure refers to the diffraction angle, θ is the Bragg angle, and the unit is ° or degree; the error range of each characteristic peak 2θ is ±0.20 (including the case where the number exceeding 1 decimal place is rounded off), specifically -0.20, -0.19, -0.18, -0.17, -0.16, -0.15, -0.14, -0.13, -0.12, -0.11, -0.10, -0.09, -0.08, -0.07, -0.06, -0.05, -0.04, -0.03, -0.02, -0.01, 0.00, 0.01, 0.02, 0.03, 0.04, 0.05, 0.06, 0.07, 0.08, 0.09, 0.10, 0.11, 0.12, 0.13, 0.14, 0.15, 0.16, 0.17, 0.18, 0.19, 0.20.

[0075] The numerical values ​​in the present disclosure, such as the data of the measured and calculated content of the relevant substances, inevitably have a certain degree of error. Generally speaking, ±10% is within the reasonable error range. There is a certain degree of error variation depending on the context where it is used, and the error variation does not exceed ±10%, and can be ±9%, ±8%, ±7%, ±6%, ±5%, ±4%, ±3%, ±2% or ±1%, preferably ±5%.

[0076] The starting material used in the crystal preparation method of the present invention can be a compound in any form, and specific forms include but are not limited to: amorphous, any crystal form, hydrate, solvate, etc.

[0077] The drying temperature in the present disclosure is generally 25° C.-100° C., preferably 40° C.-70° C., and the drying can be performed under normal pressure or reduced pressure.

[0078] The crystallization methods described in the present disclosure include room temperature crystallization, cooling crystallization, volatile solvent crystallization, adding seed crystals to induce crystallization, etc. The cooling temperature is selected from below 65°C, preferably -10°C to 60°C, and stirring can also be performed during the crystallization process.

[0079] The "differential scanning calorimetry or DSC" described in the present disclosure refers to measuring the temperature difference and heat flow difference between a sample and a reference object during the process of heating or maintaining a constant temperature of the sample to characterize all physical and chemical changes related to thermal effects and obtain phase change information of the sample.

[0080] According to the description of hygroscopic characteristics and the definition of hygroscopic weight gain in the "9103 Drug Hygroscopicity Guidelines" in the fourth volume of the 2015 edition of the Chinese Pharmacopoeia,

[0081] Deliquesce: Absorbs enough water to form a liquid;

[0082] Highly hygroscopic: weight gain due to moisture absorption is not less than 15%;

[0083] Hygroscopic: weight gain due to moisture absorption is less than 15% but not less than 2%;

[0084] Slightly hygroscopic: weight gain due to moisture absorption is less than 2% but not less than 0.2%;

[0085] No or almost no hygroscopicity: weight gain due to moisture is less than 0.2%.

[0086] The "excipients" described in the present disclosure include, but are not limited to, any adjuvant, carrier, glidant, sweetener, diluent, preservative, dye / colorant, flavoring agent, surfactant, wetting agent, dispersant, suspending agent, stabilizer, isotonic agent or emulsifier approved by the U.S. Food and Drug Administration for use by humans or livestock animals. BRIEF DESCRIPTION OF THE DRAWINGS

[0087] Figure 1 This is the XRPD spectrum of Compound 1 Form A.

[0088] Figure 2 This is the XRPD spectrum of Compound 1 Form B.

[0089] Figure 3 This is the XRPD spectrum of Compound 1 Form C.

[0090] Figure 4 This is the XRPD spectrum of Compound 1 Form D.

[0091] Figure 5 This is the XRPD spectrum of Compound 1 Form E.

[0092] Figure 6 This is the XRPD spectrum of Compound 1 Form F.

[0093] Figure 7 This is the XRPD spectrum of Compound 1 Form G.

[0094] Figure 8 This is the XRPD spectrum of Compound 1 Form H.

[0095] Fig. 9 This is the XRPD spectrum of Compound 1 Form I.

[0096] Fig.10 This is the XRPD spectrum of Compound 1 Form J.

[0097] Fig.11 This is the XRPD spectrum of the amorphous form of compound 1. DETAILED DESCRIPTION

[0098] The present disclosure will be explained in more detail below in conjunction with embodiments or experimental examples. The embodiments or experimental examples in the present disclosure are only used to illustrate the technical solutions in the present disclosure, and are not intended to limit the essence and scope of the present disclosure.

[0099] Test conditions of the instruments used in the experiment:

[0100] The structures of the compounds were determined by nuclear magnetic resonance (NMR) and / or mass spectrometry (MS). NMR shifts (δ) were expressed in 10 -6 The unit of (ppm) is given. NMR measurements were performed using a Bruker AVANCE NEO 500M NMR spectrometer, with deuterated dimethyl sulfoxide (DMSO-d6), deuterated chloroform (CDCl3), deuterated methanol (CD3OD) as the measurement solvent, and tetramethylsilane (TMS) as the internal standard.

[0101] MS was measured using Agilent 1200 / 1290DAD-6110 / 6120 Quadrupole MS LC / MS (manufacturer: Agilent, MS model: 6110 / 6120 Quadrupole MS), Waters ACQuity UPLC-QD / SQD (manufacturer: waters, MS model: waters ACQuity Qda Detector / waters SQ Detector), and THERMO Ultimate 3000-Q Exactive (manufacturer: THERMO, MS model: THERMO Q Exactive).

[0102] High performance liquid chromatography (HPLC) analysis was performed using Agilent HPLC 1200DAD, Agilent HPLC 1200VWD and Waters HPLC e2695-2489.

[0103] High performance liquid chromatography was performed using Waters 2767, Waters 2767-SQ Detecor2, Shimadzu LC-20AP and Gilson-281 preparative chromatographs.

[0104] Silica gel column chromatography generally uses Yantai Huanghai Silica Gel 200-300 mesh silica gel as the carrier.

[0105] XRPD is X-ray powder diffraction detection: the measurement is carried out using a BRUKER D8 X-ray diffractometer, and the specific collection information is: Cu anode (40kV, 40mA), Cu-Kα1 ray Kα2 rays Kβ rays Scanning mode: θ / 2θ, scanning range (2θ range): 5°~45°.

[0106] DSC is differential scanning calorimetry: the measurement was performed using a METTLER TOLEDO DSC 3+ differential scanning calorimeter, with a heating rate of 10°C / min, the specific temperature range refers to the corresponding spectrum (mostly 25-250°C or 300°C or 350°C), and a nitrogen purge rate of 50 mL / min.

[0107] TGA is thermogravimetric analysis: the test was performed using a METTLER TOLEDO TGA2 thermogravimetric analyzer, with a heating rate of 10°C / min, the specific temperature range refers to the corresponding spectrum (mostly 30-350°C), and a nitrogen purge rate of 50mL / min.

[0108] DVS stands for dynamic moisture adsorption: SMSDVS Advantage is used for detection. At 25°C, the humidity changes from 50% to 95% to 0% to 95% to 50%, with a step of 10% (the last step is 5%) (the specific range of humidity is subject to the corresponding graph, and most of the methods listed here are used). The judgment standard is that dm / dt is not greater than 0.002%.

[0109] The known starting materials disclosed herein can be synthesized by methods known in the art, or can be purchased from ABCR GmbH & Co. KG, Acros Organics, Aldrich Chemical Company, AccelaChemBio Inc, Darui Chemicals, and other companies.

[0110] Unless otherwise specified in the examples, all reactions can be carried out under an argon atmosphere or a nitrogen atmosphere.

[0111] Argon atmosphere or nitrogen atmosphere means that the reaction bottle is connected to an argon or nitrogen balloon with a capacity of about 1L.

[0112] The reaction progress in the embodiment is monitored by thin layer chromatography (TLC), the developing solvent used in the reaction, the eluent system of column chromatography used for purifying the compound and the developing solvent system of thin layer chromatography include: A: n-hexane / ethyl acetate system, B: dichloromethane / methanol system, the volume ratio of the solvent is adjusted according to the polarity of the compound, and a small amount of alkaline or acidic reagents such as triethylamine and acetic acid can also be added for adjustment.

[0113] Example 1 Preparation of the compound of formula 1

[0114] 2-amino-7-fluoro-4-((5S,5aS,6S,9R)-1-fluoro-12-((1-((4-(fluoromethylidene)piperidin-1-yl)methyl)cyclopropyl)methoxy)-5-methyl-5a,6,7,8,9,10-hexahydro-5H-4-oxa-3,10a,11,13,14-pentaaza-6,9-methano[1,8-ab]heptyl-2-yl)benzo

[0115] [b]Thiophene-3-carbonitrile 1

[0116]

[0117]

[0118] first step

[0119] tert-Butyl 4-(fluoromethylidene)piperidin-1-carboxylate 1b

[0120] 2-((Fluoromethyl)sulfonyl)pyridine (4.2 g, 23.97 mmol) was dissolved in tetrahydrofuran (50 mL), and a 1 M solution of potassium bis(trimethylsilyl)amide in tetrahydrofuran (30 mL) was added at -78°C. The temperature was maintained for reaction for 30 minutes, and then N-tert-butoxycarbonyl-4-piperidone 1a (5 g, 25.09 mmol, Shanghai Shaoyuan) was added. The temperature was maintained for reaction for 3 hours, and then the mixture was returned to room temperature for reaction for 1 hour. Saturated ammonium chloride was added to the reaction solution for quenching, and 3N hydrochloric acid (100 mL) was added. After stirring for 1 hour, the mixture was extracted with ethyl acetate (50 mL×3). The organic phases were combined, washed with saturated sodium chloride solution, dried over anhydrous sodium sulfate, filtered to remove the desiccant, and concentrated under reduced pressure. The residue was purified by silica gel column chromatography with eluent system B to obtain the title compound 1b (2 g, yield: 37%).

[0121] Step 2

[0122] 4-(Fluoromethylidene)piperidine hydrochloride 1c

[0123] Compound 1b (1 g, 4.64 mmol) was dissolved in 4 M hydrogen chloride solution in 1,4-dioxane (20 mL), and the mixture was stirred for 1 hour. The reaction solution was concentrated under reduced pressure to give the crude title compound 1c (700 mg), which was used directly in the next step without purification.

[0124] MS m / z(ESI):116.1[M+1].

[0125] Step 3

[0126] 2,5,7-Trichloro-8-fluoropyrido[4,3-d]pyrimidin-4-ol 1e

[0127] The crude compound 1d (2 g, 8 mmol, prepared according to the method in PCT / CN2023 / 109598) was dissolved in phosphorus oxychloride (25 mL), N,N-diisopropylethylamine (5.16 g, 40 mmol) was added, and the mixture was stirred at 110°C for 14 hours. The reaction solution was cooled to room temperature and concentrated under reduced pressure. The residue was dissolved in 1,4-dioxane, and 20% potassium carbonate solution was added dropwise to adjust the pH to 2-3. After stirring for 2 hours, the mixture was filtered, and the filter cake was washed with water and dried to obtain the crude title compound 1e (1.5 g). The product was used directly in the next step without purification. MS m / z (ESI): 267.8 [M+1].

[0128] Step 4

[0129] (1S,2S,5R)-2-((S)-1-((2,7-dichloro-8-fluoro-4-hydroxypyrido[4,3-d]pyrimidin-5-yl)oxy)ethyl)-3,8-diazabicyclo

[0130] [3.2.1] Octane-8-carboxylic acid tert-butyl ester 1g

[0131] (1S,2S,5R)-2-((S)-1-hydroxyethyl)-3,8-diazabicyclo[3.2.1]octane-8-carboxylic acid tert-butyl ester 1f (370 mg, 1.44 mmol, prepared by the method disclosed in Intermediate 29 on page 164 of the specification of the patent application "WO2022173678A1") was dissolved in tetrahydrofuran (10 mL), and sodium hydride (201 mg, 5.2 mmol, 60% purity) was added under ice bath. After reacting for 30 minutes, compound 1e (353 mg, 1.31 mmol) was added, and the reaction was stirred for 2 hours. Water was added to the reaction solution to quench the reaction and then concentrated under reduced pressure to obtain the crude title compound 1g (600 mg). The product was not purified and used directly in the next step.

[0132] MS m / z(ESI):488.2[M+1].

[0133] Step 5

[0134] (5S,5aS,6S,9R)-2,12-Dichloro-1-fluoro-5-methyl-5a,6,7,8,9,10-hexahydro-5H-4-oxa-3,10a,11,13,14-pentaaza

[0135] -6,9-methylnaphtho[1,8-ab]heptyl-14-carboxylic acid tert-butyl ester 1h

[0136] Compound 1g (78 mg, 159.7 μmol) was dissolved in dichloromethane (2 mL), and N,N-diisopropylethylamine (61.9 mg, 478.9 μmol) and phosphorus oxychloride (122.4 mg, 798.2 μmol) were added under ice bath, and the reaction was stirred for 2 hours. Saturated sodium bicarbonate solution was added to the reaction solution to quench, and dichloromethane (10 mL×2) was used. The organic phases were combined and dried over anhydrous sodium sulfate. The desiccant was removed by filtration and concentrated under reduced pressure to obtain the crude title compound 1h (75 mg). The product was directly used in the next step without purification.

[0137] MS m / z(ESI):470.2[M+1].

[0138] Step 6

[0139] tert-Butyl (5S,5aS,6S,9R)-12-((1-((tert-Butyldimethylsilyl)oxy)methyl)cyclopropyl)methoxy)-2-chloro-1-fluoro-5-methyl-5a,6,7,8,9,10-hexahydro-5H-4-oxa-3,10a,11,13,14-pentaaza-6,9-methanaphtho[1,8-ab]heptyl-14-carboxylate

[0140] Ester 1j

[0141] (1-(((tert-Butyldimethylsilyl)oxy)methyl)cyclopropyl)methanol 1i (1.4 g, 6.4 mmol) was dissolved in tetrahydrofuran (15 mL), and a 2M solution of sodium bis(trimethylsilyl)amide in tetrahydrofuran was added under ice bath. The temperature was maintained and stirred for 30 minutes. Then, a solution of crude compound 1h (2.3 g, 4.9 mmol) in tetrahydrofuran (20 mL) was added under ice bath. The temperature was maintained and stirred for 1 hour. Saturated ammonium chloride solution was added to the reaction solution for quenching, and the solution was extracted with ethyl acetate (30 mL×2). The organic phases were combined and dried over anhydrous sodium sulfate. The desiccant was filtered out and concentrated under reduced pressure. The residue was purified by silica gel column chromatography with eluent system B to give the title compound 1j (2 g, yield: 62.8%).

[0142] MS m / z(ESI):650.2[M+1].

[0143] Step 7

[0144] (5S,5aS,6S,9R)-2-Chloro-1-fluoro-12-((1-(hydroxymethyl)cyclopropyl)methoxy)-5-methyl-5a,6,7,8,9,10-hexahydro-5H-4-oxapropane

[0145] -3,10a,11,13,14-pentaaza-6,9-methanaphtho[1,8-ab]heptyl-4-carboxylic acid tert-butyl ester 1k

[0146] Compound 1j (100 mg, 153.8 μmol) was dissolved in tetrahydrofuran (4 mL), and a 1M tetrabutylammonium fluoride solution in tetrahydrofuran (187 μL) was added. The mixture was stirred for 2 hours, and a saturated aqueous solution of ammonium chloride was added to the reaction solution for quenching. The mixture was extracted with ethyl acetate (15 mL×3). The organic phases were combined, washed with water and a saturated sodium chloride solution in sequence, and dried over anhydrous sodium sulfate. After filtering to remove the desiccant, the filtrate was concentrated under reduced pressure to obtain a crude title compound 1k (82 mg). The product was used directly in the next step without purification. MSm / z(ESI):536.2[M+1].

[0147] Step 8

[0148] (5S,5aS,6S,9R)-2-Chloro-1-fluoro-5-methyl-12-((1-((methylsulfonyl)oxy)methyl)cyclopropyl)methoxy)-5a,6,7,8,9,10-hexahydro-5H-4-oxa-3,10a,11,13,14-pentaaza-6,9-methanaphtho[1,8-ab]heptyl-14carboxylic acid tert-butyl ester

[0149] 1l

[0150] The crude compound 1k (83 mg, 154.9 μmol) and N,N-diisopropylethylamine (60 mg, 464.2 μmol) were dissolved in dichloromethane (3 mL), and methanesulfonyl chloride (25 mg, 218.2 μmol) was added under ice bath. The reaction mixture was naturally restored to room temperature for 30 minutes. Saturated aqueous ammonium chloride was added to the reaction solution for quenching, and the mixture was extracted with ethyl acetate (10 mL×3). The organic phases were combined, washed with water and saturated sodium chloride solution in turn, and dried over anhydrous sodium sulfate. The desiccant was filtered out and the filtrate was concentrated under reduced pressure to obtain the crude title compound 1l (95 mg), which was used directly in the next step without purification.

[0151] MS m / z(ESI):614.2[M+1].

[0152] Step 9

[0153] tert-Butyl (5S,5aS,6S,9R)-2-chloro-1-fluoro-12-((1-((4-(fluoromethylidene)piperidin-1-yl)methyl)cyclopropyl)methoxy)-5-methyl-5a,6,7,8,9,10-hexahydro-5H-4-oxa-3,10a,11,13,14-pentaaza-6,9-methanaphtho[1,8-ab]heptyl-14-carboxylate

[0154] Ester 1m

[0155] The crude compound 1l (95 mg, 154.7 μmol) and compound 1c (35.5 mg, 234.5 μmol) were dissolved in acetonitrile (4 mL), and anhydrous potassium carbonate (64 mg, 463 μmol) and sodium iodide (70 mg, 467 μmol) were added. The mixture was stirred at 80°C for 2 hours. The reaction solution was concentrated under reduced pressure and the residue was purified by silica gel column chromatography with eluent system A to give the title compound 1m (80 mg, yield: 81.6%).

[0156] MS m / z(ESI):633.2[M+1].

[0157] Step 10: (5S,5aS,6S,9R)-2-(2-((tert-butoxycarbonyl)amino)-3-cyano-7-fluorobenzo[b]thiophen-4-yl)-1-fluoro-12-((1-((4-(fluoromethylidene)piperidin-1-yl)methyl)cyclopropyl)methoxy)-5-methyl-5a,6,7,8,9,10-hexahydro-5H-4-oxa-3,10a,11,13,14-pentaaza-6,9-methano[1,8-ab]heptyl-14-carboxylic acid tert-butyl ester 1o

[0158] Compound 1m (20 mg, 31.6 μmol), tert-butyl (3-cyano-4-(5,5-dimethyl-1,3,2-dioxaborohexan-2-yl)-7-fluorobenzo[b]thiophen-2-yl)carbamate 1n (19 mg, 47 μmol, prepared by the method disclosed in preparation 23 on page 39 of the specification of patent application "WO2022261154A1"), tetrakis(triphenylphosphine)palladium (7 mg, 6.1 μmol), and cesium carbonate (30 mg, 92 μmol) were mixed in N,N-dimethylformamide (1 mL), replaced with nitrogen, and reacted at 90°C for 2 hours. After the reaction solution was cooled to room temperature, ethyl acetate was added to dilute it, dried over anhydrous sodium sulfate, filtered to remove the desiccant, and concentrated under reduced pressure to obtain the crude title compound 1o (15 mg), which was used directly in the next step without purification.

[0159] MS m / z(ESI):889.2[M+1].

[0160] Step 11

[0161] 2-amino-7-fluoro-4-((5S,5aS,6S,9R)-1-fluoro-12-((1-((4-(fluoromethylidene)piperidin-1-yl)methyl)cyclopropyl)methoxy)-5-methyl-5a,6,7,8,9,10-hexahydro-5H-4-oxa-3,10a,11,13,14-pentaaza-6,9-methano[1,8-ab]heptyl-2-yl)benzo

[0162] [b]Thiophene-3-carbonitrile 1

[0163] The crude compound 1o (15 mg, 16.9 μmol) was dissolved in acetonitrile (1 mL), and 4 M hydrochloric acid 1,4-dioxane solution (0.5 mL) was added under ice bath, and the temperature was maintained for 0.5 hours, and then the reaction was carried out at room temperature for 2 hours. The mixture was concentrated under reduced pressure, and the residue was purified by high performance liquid preparative chromatography (Waters-2545, chromatographic column: ODS-BIO C18, 30*150 mm, 5 μm; mobile phase: aqueous phase (10 mmol / L ammonium bicarbonate) and acetonitrile, gradient ratio: acetonitrile 54%-69% 15 min, flow rate: 30 mL / min, column temperature: room temperature) to obtain the title compound 1 (4 mg, yield: 34.4%). The product was free amorphous as detected by X-ray powder diffraction, and the XRPD spectrum was as shown in FIG. Fig.11 .

[0164] MS m / z(ESI):687.3[M-1].

[0165] 1 H NMR (500MHz, CD3OD): δ7.39(dd,1H),7.04(t,1H),6.51(d,1H),5.41(dd,1H),4.5–4.46(m,2H),4.40(d,1H),4.10(d,1H),3.7 0(d,1H),3.60(d,1H),3.19(s,1H),2.62–2.31(m,8H),2.09(d,3H),1.92–1.75(m,3H),1.61(d,3H),0.75(s,2H),0.52(s,2H).

[0166] Test Example 1: Biological evaluation of GP2d and AGS cell 3D proliferation inhibition experiment

[0167] Experimental methods

[0168] GP2d cells were cultured with complete medium, i.e., DMEM / high glucose medium (Hyclone, SH30243.01) containing 10% fetal bovine serum (Corning, 35-076-CV). On the first day of the experiment, GP2d cells were seeded in a 96-well low-adhesion plate (Corning, CLS7007-24EA) at a density of 1000 cells / well using complete medium, with 90 μL of cell suspension per well, centrifuged at 2000 rpm at room temperature for 5 minutes, and then placed in a 37°C, 5% CO2 cell culture incubator for overnight culture.

[0169] AGS cells were cultured with complete medium, i.e., RPMI1640 medium (Hyclone, SH30809.01) containing 10% fetal bovine serum (Corning, 35-076-CV). On the first day of the experiment, AGS cells were seeded in a 96-well low-adhesion plate (Corning, CLS7007-24EA) at a density of 1000 cells / well using complete medium, with 90 μL of cell suspension per well, centrifuged at 2000 rpm for 5 minutes at room temperature, and then placed in a 37°C, 5% CO2 cell culture incubator for overnight culture.

[0170] On the second day, 10 μL of the compound to be tested prepared with complete medium was added to each well. The final concentration of the compound for GP2d cells was 9 concentration points of 5-fold gradient dilution starting from 1 μM, and the final concentration of the compound for AGS cells was 9 concentration points of 5-fold gradient dilution starting from 10 μM. A blank control containing 0.5% DMSO was set. The well plate was placed in a cell culture incubator at 37°C and 5% CO2 for 5 days. On the seventh day, the 96-well cell culture plate was removed and 50 μL of CellTiter- 3D Cell Viability Assay reagent (Promega, G9682) was shaken at room temperature in the dark for 25 minutes, then pipetted to mix and 100 μL was taken out from each well and transferred to a white opaque 96-well plate (PerkinElmer, 6005290), and the luminescent signal value was read using a multi-function microplate reader (PerkinElmer, EnVision2105).

[0171] The IC of the inhibitory activity of the compounds was calculated using Graphpad Prism software 50 The results are shown in Table 1 below.

[0172] Table 1 AGS, GP2d cell 3D proliferation inhibitory activity data

[0173]

[0174] Conclusion: The disclosed compounds have a good inhibitory effect on the 3D proliferation of AGS and GP2d cells.

[0175] Test Example 2: Biological evaluation of AsPC-1 cell 3D proliferation inhibition experiment

[0176] On the first day of the experiment, AsPC-1 cells that grew well and reached 70%-80% confluence were digested and resuspended in RPMI 1640 (Hyclone, SH30809.01) medium containing 10% FBS, and the cell density was adjusted to the required level. 90 μL of cell suspension was added to each well of a U-shaped low-adsorption 96-well plate (Corning, CLS7007-24EA), and the cell density was 1500 cells / well. After the cell plate was centrifuged at 2500 rpm for 5 minutes, it was placed in a 37°C, 5% CO2 incubator and cultured overnight. On the second day, the 20mM test compound dissolved in DMSO was diluted with DMSO to a first concentration of 2mM, and then diluted 5-fold gradients for a total of 9 concentration points, and the control well was DMSO. The gradient diluted compound was then further diluted 20 times with culture medium. 10 μL of the test compound diluted with culture medium was added to each well of the cell plate, and the final concentration of the compound was the first concentration of 10 μM and the 9 concentration points of the 5-fold gradient dilution. The cell wells containing 0.5% DMSO were set as the solvent control wells, and the wells containing only the culture medium and 0.5% DMSO were set as blank control wells. The compound concentrations and control wells were set in duplicate, and the final DMSO concentration in each well was 0.5%. After centrifugation at 2500 rpm for 3 minutes, the cell plate was placed in a 37°C, 5% CO2 incubator for 5 days. On the seventh day, the 96-well cell culture plate was removed and 50 μL of the luminescent cell activity detection reagent CellTiter- 3D Cell Viability Assay (Promega, G9683), shake at room temperature in the dark for 25 minutes, pipette up and down to mix, and transfer 100 μL per well to the white opaque OptiPlate TM -96-well plate (PerkinElmer, 6005290), and the luminescent signal value was read using a multi-function microplate reader (PerkinElmer, EnVision2105).

[0177] The inhibition rate was calculated using the following formula: Inhibition rate = (luminescence value 溶媒对照孔 - Luminescence value 受试化合物 ) / (luminous value 溶媒对照孔 - Luminescence value 空白对照孔 )×100%. GraphPadPrism software was used to draw a curve based on the concentration of each compound and the corresponding inhibition rate, and the IC of the compound was calculated. 50 value.

[0178] Table 2. AsPC-1 cell 3D proliferation inhibition activity data

[0179]

[0180] Conclusion: The disclosed compounds have a good inhibitory effect on the 3D proliferation of AsPC-1 cells.

[0181] Test Example 3: Pharmacokinetic Evaluation

[0182] 1. Summary

[0183] Balb / c nude mice were used as test animals, and the drug concentration in plasma at different times after gavage (ig) of the example compounds to balb / c nude mice was determined by LC / MS / MS method. The pharmacokinetic behavior of the disclosed compounds in balb / c nude mice was studied, and their pharmacokinetic characteristics were evaluated.

[0184] 2. Experimental plan

[0185] 2.1. Experimental Drugs

[0186] Compound 1.

[0187] 2.2 Experimental Animals

[0188] 9 female BALB / c nude mice were provided by Weitonglihua Laboratory Animal Technology Co., Ltd., with production license SCXK(Zhejiang)2019-0001.

[0189] 2.3 Drug preparation

[0190] A certain amount of the test compound was weighed respectively, and 5% DMF+45% PG+50% (10% HS15-pH7.4 buffer)+400 mpk SNAC was added to prepare a 4 mg / mL colorless clear solution.

[0191] 2.4 Drug administration

[0192] The dosage was 40.0 mg / kg and the administration volume was 10 mL / kg.

[0193] 3. Operation

[0194] Before administration and 0.25, 0.5, 1.0, 2.0, 4.0, 6.0, 8.0, 11.0, and 24.0 hours after administration, 0.1 mL of blood was collected from the eye sockets and placed in EDTA-K2 anticoagulant tubes. The blood was centrifuged at 10,000 rpm for 1 minute (4°C), and the plasma was separated within 1 hour and stored at -20°C for testing. The blood collection and centrifugation process was performed under ice bath conditions.

[0195] Determine the content of the test compound in the plasma of balb / c nude mice after administration of different concentrations of the drug: Take 25 μL of balb / c nude mouse plasma samples at each time after administration, 200 μL of acetonitrile containing internal standard (Compound 1: internal standard is verapamil 100 ng / mL), vortex mix for 5 minutes, and centrifuge at 3700 rpm for 20 minutes, and mix the supernatant with water (1:1). Take 0.5-2 μL of the supernatant for LC / MS / MS analysis.

[0196] 4. Pharmacokinetic parameter results

[0197] Table 3. Pharmacokinetic parameters of the disclosed compounds in balb / c nude mice

[0198]

[0199] Conclusion: The disclosed compounds have good pharmacokinetic advantages in balb / c nude mice.

[0200] Example 2 Preparation of Free Form A

[0201] 5 mg of the compound shown in Formula 1 was dissolved in 0.175 mL of methanol, and the solvent was evaporated at room temperature to obtain a solid. The product was defined as free crystal form A by X-ray powder diffraction detection, and the XRPD spectrum was as follows: Figure 1 The characteristic peak positions are shown in Table 4. The DSC spectrum shows that the endothermic peaks are 68.18°C, 106.18°C and 160.02°C. The TGA spectrum shows that the weight loss is 3.92% at 31°C-115°C.

[0202] Table 4

[0203]

[0204]

[0205] Example 3 Preparation of Free Form A

[0206] 5 mg of the compound shown in Formula 1 was dissolved in the solvent in Table 5, and the solvent was evaporated at room temperature to obtain a solid. The product was free crystal form A as determined by X-ray powder diffraction.

[0207] Table 5

[0208] Solvent Solvent volume (mL) result 10% water / methanol 0.275 Crystalline Form A Acetonitrile / methanol (1:1) 0.25 Crystalline Form A

[0209] Example 4 Preparation of Free Form A

[0210] 5 mg of the compound represented by Formula 1 was added with 0.5 mL of a solvent as shown in Table 6 below, and stirred at room temperature for 4 days to obtain the title product.

[0211] Table 6

[0212] Solvent Crystal form water A Acetonitrile A n-Heptane A 7% water / ethanol A 50% methanol / water A Cyclohexane A Isopropyl ether A

[0213] Example 5 Preparation of Free Form B

[0214] 5 mg of the compound shown in Formula 1 was dissolved in 0.025 mL of tetrahydrofuran / ethanol (2:1), and the temperature was raised and lowered at 40-5°C for 3 cycles, with a heating / lowering rate of 1°C / min. Solid was precipitated, centrifuged, and the solid was vacuum dried at 40°C to obtain the title product. The product was defined as free form B by X-ray powder diffraction detection, and the XRPD spectrum was as follows: Figure 2 The characteristic peak positions are shown in Table 7. The DSC spectrum shows that the endothermic peaks are 127.5°C and 161.52°C. The TGA spectrum shows that the weight loss is 2.72% at 30°C-143°C.

[0215] Table 7

[0216]

[0217]

[0218] Example 6 Preparation of Free Form B

[0219] 5 mg of the compound of formula 1 was dissolved in 0.425 mL of ethyl acetate / ethanol (1:1), and the solvent was evaporated at room temperature to obtain a solid. The product was free crystal form B as determined by X-ray powder diffraction.

[0220] Example 7 Preparation of Free Form B

[0221] 0.075 mL of ethanol was added to 5 mg of the amorphous compound of Formula 1, and the mixture was stirred at room temperature for crystallization to obtain a solid. The product was free-state crystalline form B as determined by X-ray powder diffraction.

[0222] Example 8 Preparation of Free Form C

[0223] 5 mg of the compound shown in Formula 1 was added with 0.5 mL of ethanol, stirred at room temperature for 4 days, centrifuged, and the solid was dried in vacuum at 40°C to obtain the title product. The product was defined as free form C by X-ray powder diffraction detection, and the XRPD spectrum was as follows: Figure 3 The characteristic peak positions are shown in Table 8. The DSC spectrum shows that the endothermic peak is 155.84°C. The TGA spectrum shows that the weight loss is 4.88% at 30°C-150°C.

[0224] Table 8

[0225]

[0226]

[0227] Example 9 Preparation of Free Form C

[0228] 5 mg of the compound of Formula 1 was added with 0.5 mL of solvent as shown in Table 9 below, stirred at room temperature for 4 days, centrifuged, and the solid was vacuum dried at 40°C. X-ray powder diffraction analysis showed that the product was free crystalline Form C.

[0229] Table 9

[0230] Solvent Crystal form Isopropyl alcohol C n-Propanol C

[0231] Example 10 Preparation of Free Form D

[0232] 5 mg of the compound shown in Formula 1 was dissolved in 0.1 mL of 2-methyltetrahydrofuran, and 0.5 mL of methyl tert-butyl ether was added. The mixture was stirred at room temperature to precipitate, centrifuged, and the solid was dried in vacuo at 40°C to obtain the title product. The product was defined as free crystal form D by X-ray powder diffraction detection. The XRPD spectrum is shown in Figure 4 The characteristic peak positions are shown in Table 10. The DSC spectrum shows that the endothermic peak has a peak value of 195.22°C. The TGA spectrum shows that the weight loss is 0.49% from 30°C to 132°C.

[0233] Table 10

[0234]

[0235] Example 11 Preparation of Free Form D

[0236] 5 mg of the compound represented by Formula 1 was added with 0.5 mL of solvent as shown in Table 11 below, stirred at room temperature for 4 days, centrifuged, and the solid was dried under vacuum at 40° C. to obtain the title product.

[0237] Table 11

[0238] Starting crystal form Solvent Crystal form Crystalline Form A Methyl tert-butyl ether D Amorphous Ethyl acetate / n-heptane (1:1) D

[0239] Example 12 Preparation of Free Form D

[0240] 5 mg of the amorphous compound of Formula 1 was dissolved in 0.4 mL of methyl tert-butyl ether, stirred to precipitate, centrifuged, and the solid was dried in vacuo at 40° C. to obtain the title product.

[0241] Example 13 Preparation of Free Form E

[0242] 5 mg of the compound shown in Formula 1 was dissolved in 0.1 mL of acetone, and 0.5 mL of methyl tert-butyl ether was added. The mixture was stirred at room temperature to precipitate, centrifuged, and the solid was dried in vacuum at 40°C to obtain the title product. The product was defined as free crystal form E by X-ray powder diffraction detection. The XRPD spectrum is shown in Figure 5 The characteristic peak positions are shown in Table 12. The DSC spectrum shows that the endothermic peak has a peak value of 188.85°C. The TGA spectrum shows that the weight loss is 2.37% from 32°C to 160°C.

[0243] Table 12

[0244]

[0245] Example 14 Preparation of Free Form E

[0246] 5 mg of the compound of formula 1 was dissolved in solvent I in Table 13, and solvent II in Table 13 was added, stirred at room temperature to precipitate, centrifuged, and the solid was vacuum dried at 40°C to obtain the title product. The product was free crystalline form E as determined by X-ray powder diffraction.

[0247] Table 13

[0248]

[0249]

[0250] Example 15 Preparation of Free Form E

[0251] 5 mg of the compound represented by Formula 1 was added with 0.5 mL of solvent as shown in Table 14 below, stirred at room temperature for 4 days, centrifuged, and the solid was dried under vacuum at 40° C. to obtain the title product.

[0252] Table 14

[0253] Solvent Crystal form Ethyl acetate / n-heptane (1:1) E acetone E

[0254] Example 16 Preparation of Free Form E

[0255] 5 mg of the compound represented by Formula 1 was dissolved in 1.2 mL of ethyl acetate / methyl tert-butyl ether (v / v=1:5), and the solvent was evaporated at room temperature to obtain the title product.

[0256] Example 17 Preparation of Free Form E

[0257] 5 mg of the amorphous compound of Formula 1 was dissolved in 0.05 mL of a solvent as shown in Table 15 below, stirred to precipitate, centrifuged, and the solid was dried in vacuo at 40° C. to obtain the title product.

[0258] Table 15

[0259] Solvent Crystal form 2-Methyltetrahydrofuran E Dichloromethane E

[0260] Example 18 Preparation of Free Form F

[0261] 5 mg of the compound of formula 1 was dissolved in 0.15 mL of dichloromethane, and 0.75 mL of n-heptane was added. The mixture was stirred at room temperature to precipitate, centrifuged, and the solid was dried under vacuum at 40°C to obtain the title product. The product was defined as free crystal form F by X-ray powder diffraction detection. The XRPD spectrum is shown in Figure 6 The characteristic peak positions are shown in Table 16. The DSC spectrum shows that the endothermic peak is 163.53°C. The TGA spectrum shows that the weight loss is 1.70% at 32°C-102°C.

[0262] Table 16

[0263]

[0264]

[0265] Example 19 Preparation of Free Form G

[0266] 5 mg of the amorphous compound of Formula 1 was added with 0.05 mL of methanol, stirred at room temperature for crystallization, centrifuged, and the solid was dried in vacuo at 40°C to obtain the title product. The product was defined as free crystal form G by X-ray powder diffraction detection, and the XRPD spectrum was as follows: Figure 7 The characteristic peak positions are shown in Table 17. The DSC spectrum shows that the endothermic peak is 215.63°C. The TGA spectrum shows that the weight loss is 0.87% from 31°C to 105°C.

[0267] Table 17

[0268]

[0269] Example 20 Preparation of Free Form H

[0270] 5 mg of the amorphous compound of Formula 1 was added with 0.5 mL of p-xylene, stirred at room temperature for 3 days, centrifuged, and the solid was vacuum dried at 40°C to obtain the title product. The product was defined as free crystalline form H by X-ray powder diffraction detection, and the XRPD spectrum was as follows: Figure 8 The characteristic peak positions are shown in Table 18. The DSC spectrum shows that the endothermic peak value is 154.82°C. The TGA spectrum shows that the weight loss is 5.82% from 31°C to 170°C.

[0271] Table 18

[0272]

[0273] Example 21 Preparation of Free Form H

[0274] The amorphous compound of Formula 1 was added with 1 mL of solvent as shown in Table 19 below, stirred at room temperature overnight, centrifuged, and the solid was dried under vacuum at 40° C. to obtain the title product.

[0275] Table 19

[0276] Amount of the compound shown in formula 1 (mg) Solvent Solvent volume (mL) Crystal form 80 Methyl tert-butyl ether 1 H 112 Ethyl acetate 1 H

[0277] Example 22 Preparation of Free Form H

[0278] 78 mg of the amorphous compound of Formula 1 was dissolved in 0.8 mL of DCM and added to 4 mL of n-heptane. Solid precipitated and stirred at room temperature overnight. The solid was centrifuged and dried in vacuo at 40° C. to obtain the title product.

[0279] Example 23 Preparation of Free Form H

[0280] 7 mg of the amorphous compound of Formula 1 was added with 0.1 mL of ethyl acetate and 10.7 μL of 2M ethanolic citric acid solution (the ligand may be 10.7 μL of 2M ethanolic tartaric acid solution or 2.5 mg of fumaric acid), stirred for crystallization, centrifuged, and the solid was dried under vacuum at 40° C. to obtain the title product.

[0281] Example 24 Preparation of Free Form I

[0282] 7 mg of the amorphous compound of Formula 1 was added with 0.1 mL of ethyl acetate and 10.7 μL of 2M acetic acid ethanol solution, stirred to precipitate, centrifuged, and the solid was vacuum dried to obtain the title product. The product was defined as free form I by X-ray powder diffraction detection, and the XRPD spectrum was as follows: Fig. 9 The characteristic peak positions are shown in Table 20. The DSC spectrum shows that the endothermic peaks are 162.18°C and 224.15°C. The TGA spectrum shows that the weight loss is 4.27% at 33°C-140°C.

[0283] Table 20

[0284]

[0285] Example 25 Preparation of Free Form J

[0286] 5 mg of the amorphous compound of Formula 1 and the crystalline form I of the compound were heated to 215°C to obtain the title product. The product was defined as the free crystalline form J by X-ray powder diffraction detection. The XRPD spectrum is as follows: Fig.10 , and the positions of its characteristic peaks are shown in Table 21.

[0287] Table 21

[0288]

[0289] Test Example 4: Stability of Influencing Factors

[0290] The aforementioned crystal forms A, D, E, G, and H were laid out openly, and the stability of the samples was investigated under high temperature (40° C., 60° C.) and high humidity (RH75%, RH92.5%) conditions, respectively. The sampling investigation period was 30 days.

[0291] Table 22

[0292]

[0293]

[0294]

[0295] Conclusion: The influencing factor experiment shows that the free crystal forms A, D, E, G, and H have good physical and chemical stability under the influencing factor high humidity (75% RH, 92.5% RH) conditions for 30 days; they have good physical stability under light conditions for 30 days, and the chemical purity is reduced. The free crystal forms A, D, E, and G have good physical stability under the influencing factor high temperature (40℃, 60℃) conditions for 30 days.

[0296] Test Example 5: Long-term accelerated test stability

[0297] The aforementioned crystalline forms A, D, E, G, and H were placed at 25°C / 60% RH and 40°C / 75% RH, respectively, to investigate their stability.

[0298] Table 23

[0299]

[0300]

[0301] Conclusion: Long-term accelerated experiments show that the free crystal forms A and H have good physical and chemical stability under the conditions of 25℃ / 60%RH and 40℃ / 75%RH for 1 month; the free crystal forms D, E and G have good physical and chemical stability under the conditions of 25℃ / 60%RH and 40℃ / 75%RH for 2 months.

Claims

1. A crystalline form A of the compound represented by formula 1, characterized in that: The X-ray powder diffraction pattern represented by the diffraction angle 2θ has characteristic peaks at 8.984, 12.800, 14.133, 14.675, 18.977, and 21.018, preferably at 8.984, 9.837, 12.800, 14.133, 14.675, 15.944, 16.635, 17.595, 18. 977, 21.018, 26.006, more preferably 8.984, 9.837, 12.800, 14.133, 14.675, 15.944, 16.635, 17.595, 18.977, 21.018, 22.089, 23.026, 26.006, 29.233, 2. The crystalline form A according to claim 1, characterized in that The X-ray powder diffraction pattern expressed in terms of diffraction angle 2θ is shown in FIG1 .

3. A method for preparing the crystalline form A according to claim 1 or 2, wherein the method is selected from any one of the following methods: Method 1: dissolving the compound of formula 1 in solvent I and volatilizing the solvent, wherein the solvent I is selected from methanol, 10% water / methanol, and 50% acetonitrile / methanol; Method 2: Add the compound of formula 1 into solvent II and stir, wherein the solvent II is selected from water, acetonitrile, n-heptane, cyclohexane, isopropyl ether, 7% water / ethanol, and 50% water / methanol.

4. A crystalline form D of the compound represented by formula 1, characterized in that: The X-ray powder diffraction pattern represented by the diffraction angle 2θ has characteristic peaks at 7.229, 9.137, 11.664, 16.506, and 20.682, and preferably has characteristic peaks at 5.265, 7.229, 9.137, 11.664, 16.506, 17.396, 20.682, 21.849, and 23.

001.

5. The crystal form D according to claim 4, characterized in that: The X-ray powder diffraction pattern expressed in terms of diffraction angle 2θ is shown in FIG4 .

6. A method for preparing the crystalline form D according to claim 4 or 5, wherein the method is selected from any one of the following methods: Method 1: Dissolve the compound of formula 1 in 2-methyltetrahydrofuran, add methyl tert-butyl ether, and stir; Method 2: Add the compound of formula 1 into methyl tert-butyl ether or 50% ethyl acetate / n-heptane and stir.

7. According to the crystal form according to any one of claims 1-2, 4-5, the 2θ angle error range is ±0.

20.

8. A pharmaceutical composition comprising the crystal form according to any one of claims 1-2, 4-5 and optionally a pharmaceutically acceptable excipient.

9. A method for preparing a pharmaceutical composition, comprising the step of mixing the crystal form according to any one of claims 1-2, 4-5 and a pharmaceutically acceptable excipient.

10. Use of the crystal form according to any one of claims 1-2, 4-5, or the pharmaceutical composition according to claim 8 in the preparation of a KRAS inhibitor.

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