A crystal form of a pyrimido five-membered nitrogen heterocyclic derivative and a preparation method thereof

Abstract

This disclosure relates to a crystalline form of a pyrimido-five-membered nitrogen heterocyclic derivative and a method for preparing the same. Specifically, this disclosure relates to different crystalline forms of a compound represented by formula (I) and a method for preparing the same. The crystalline form of the compound of formula (I) provided herein exhibits excellent stability and is therefore more suitable for clinical treatment. #imgabs0#

Description

[0001] This application claims priority to Chinese patent application No. 202011563824.1, filed on December 25, 2020. This application incorporates the entire text of the aforementioned Chinese patent application. Technical Field

[0002] The present disclosure relates to a crystal form of a pyrimido five-membered nitrogen heterocyclic derivative, a preparation method thereof, and medical uses thereof, and belongs to the field of pharmaceuticals. Background Art

[0003] Src homology domain 2-containing tyrosine phosphatase-2 (SHP2) is an evolutionarily conserved non-receptor protein tyrosine phosphatase (PTP) encoded by the PTPN11 gene. It is composed of two SH2 domains (N-SH2 and C-SH2) and a PTP catalytic domain. It is widely expressed in human tissues and plays a crucial role in maintaining tissue development and cellular homeostasis. SHP2 is involved in signaling through the Ras-mitogen-activated protein kinase, JAK-STAT, or phosphoinositide 3-kinase AKT pathways. Mutations in the PTPN11 gene, and subsequently in SHP2, have been identified in a variety of human diseases, including Noonan syndrome, Leopard syndrome, juvenile myelomonocytic leukemia, neuroblastoma, melanoma, acute myeloid leukemia, and breast, lung, and colon cancers (same as Claim19). Therefore, SHP2 represents a highly attractive target for the development of new therapeutics to treat various diseases.

[0004] The patent applications for related research on the SHP2 target that have been disclosed include WO2018136264A, WO2015003094A, WO2018160731A, WO2018130928A1, WO2018136265A, WO2018172984A, WO2018081091, WO2016203405, WO2017211303A, WO2018013597A, etc. Currently, Novartis' SHP2 inhibitor TN0155 and JACOBIO's SHP2 inhibitor JAB-3068 are both in Phase I clinical trials, and there are no products targeting this target on the market. Therefore, it is still necessary to continue to develop more efficient new SHP2 inhibitors in order to provide patients with new and effective anti-cancer drugs.

[0005] Patent application WO2020259679A discloses a compound represented by formula (I). To meet the demand for medication, it is necessary to study its crystal form.

[0006] Summary of the Invention

[0007] The present disclosure provides a crystal form A of a compound represented by formula (I),

[0008]

[0009] The X-ray powder diffraction pattern expressed in diffraction angle 2θ degrees has characteristic peaks at 4.847, 9.801, 13.778, 14.770, 15.444, and 26.077.

[0010] In an optional embodiment, the crystal form A of the compound represented by formula (I) provided by the present disclosure has an X-ray powder diffraction pattern expressed as a diffraction angle 2θ, with characteristic peaks at 4.847, 9.801, 13.138, 13.778, 14.770, 15.444, 18.363, 19.856, 21.092, 23.371, 26.077, and 28.130.

[0011] In an optional embodiment, the crystal form A of the compound represented by formula (I) provided by the present disclosure has an X-ray powder diffraction pattern expressed as a diffraction angle 2θ, and has characteristic peaks at 4.847, 9.801, 13.138, 13.778, 14.770, 15.444, 18.363, 19.856, 21.092, 22.034, 23.371, 24.460, 26.077, 28.130, 28.970, 31.894, 32.920, 33.916, and 38.924.

[0012] In an optional embodiment, the present disclosure provides a crystal form A of the compound represented by formula (I), whose X-ray powder diffraction spectrum is as shown in the attached Figure 2 shown.

[0013] The present disclosure provides a method for preparing crystal form A of the compound represented by formula (I), which is selected from:

[0014] a) mixing the compound represented by formula (I) with solvent I, dissolving it, and crystallizing it, wherein the solvent I is selected from

[0015] At least one of dichloromethane and chloroform; or

[0016] b) mixing the compound represented by formula (I) with solvent II to crystallize, wherein the solvent II is at least one selected from tetrahydrofuran, ethyl acetate, toluene, acetone, methanol, ethanol, acetonitrile, methyl tert-butyl ether, water, isopropyl ether, butanone, and n-hexane; or

[0017] c) mixing the compound represented by formula (I) with solvent III to obtain a clear solution, mixing the clear solution with solvent IV to precipitate crystals, wherein the solvent III is selected from at least one of methanol, N,N-dimethylformamide, and N-methylpyrrolidone; and the solvent IV is selected from at least one of acetonitrile, ethyl acetate, isopropyl acetate, methyl tert-butyl ether, and isopropyl ether.

[0018] The present disclosure provides a crystal form B of a compound represented by formula (I), which has an X-ray powder diffraction pattern represented by a diffraction angle 2θ, with characteristic peaks at 4.606, 9.110, 11.423, 13.745, 16.006, and 22.973.

[0019] In an optional embodiment, the B crystal form of the compound represented by formula (I) provided by the present disclosure has an X-ray powder diffraction pattern expressed as a diffraction angle of 2θ, with characteristic peaks at 4.606, 9.110, 11.423, 13.745, 16.006, 18.349, 22.973, 25.285, 27.505, and 29.262.

[0020] In an optional embodiment, the B crystal form of the compound represented by formula (I) provided by the present disclosure has an X-ray powder diffraction pattern expressed as a diffraction angle 2θ, with characteristic peaks at 4.606, 9.110, 11.423, 13.745, 16.006, 18.349, 19.767, 22.973, 24.700, 25.285, 27.505, 29.262, 31.451, 32.407, 34.072, 35.983, 37.216, and 38.388.

[0021] A method for preparing crystal form B of a compound represented by formula (I) comprises dissolving the compound represented by formula (I) in methanol, filtering the solution, mixing the solution with methyl tert-butyl ether, and crystallizing the solution.

[0022] The present disclosure provides a crystal form C of a compound represented by formula (I), which has an X-ray powder diffraction pattern represented by a diffraction angle 2θ, with characteristic peaks at 8.905, 12.920, 13.428, 14.074, 18.458, and 22.519.

[0023] In an optional embodiment, the C form of the compound represented by formula (I) provided by the present disclosure has an X-ray powder diffraction pattern expressed as a diffraction angle 2θ, with characteristic peaks at 8.905, 12.920, 13.428, 14.074, 16.104, 17.996, 18.458, 18.965, 20.580, 22.519, 23.949, 26.395, 28.795, and 31.748.

[0024] In an optional embodiment, the present disclosure provides a crystal form C of the compound represented by formula (I), with an X-ray powder diffraction pattern expressed as a diffraction angle of 2θ, having characteristic peaks at 8.905, 12.920, 13.428, 14.074, 16.104, 17.996, 18.458, 18.965, 20.580, 22.519, 23.949, 25.011, 26.395, 27.226, 28.379, 28.795, 30.041, 31.748, 32.487, 35.578, 37.978, and 41.393.

[0025] A method for preparing crystal form C of a compound represented by formula (I), comprising mixing the compound represented by formula (I) with at least one solvent selected from dioxane, N-methylpyrrolidone, and N,N-dimethylformamide to obtain a clear solution, and mixing the clear solution with at least one solvent selected from methyl tert-butyl ether and isopropyl alcohol to precipitate crystals.

[0026] The present disclosure provides a D-type crystal of a compound represented by formula (I), which has an X-ray powder diffraction pattern represented by a diffraction angle 2θ, with characteristic peaks at 4.766, 9.594, 14.089, 14.471, 18.981, 19.609, and 25.987.

[0027] In an optional embodiment, the D form of the compound represented by formula (I) provided by the present disclosure has an X-ray powder diffraction pattern expressed as a diffraction angle 2θ, with characteristic peaks at 4.766, 9.594, 13.461, 14.089, 14.471, 16.602, 18.981, 19.609, 20.238, 22.616, 24.187, 24.770, 25.264, 25.987, 29.123, 30.380, 32.714, 34.688, 39.042, 39.535, and 44.382.

[0028] A method for preparing the D crystal form of the compound represented by formula (I) is selected from the following methods:

[0029] a) mixing the compound represented by formula (I) with methanol to obtain a clear solution, and mixing the clear solution with isopropyl ether to obtain crystals;

[0030] b) mixing the compound represented by formula (I) with ethanol-water, dissolving the mixture, and volatilizing the mixture to separate crystals.

[0031] The present disclosure provides a crystalline form E of a compound represented by formula (I), which has an X-ray powder diffraction pattern represented by a diffraction angle 2θ, with characteristic peaks at 4.603, 9.209, 13.920, 15.097, 19.700, and 25.454.

[0032] In an optional embodiment, the E crystal form of the compound represented by formula (I) provided by the present disclosure has an X-ray powder diffraction pattern expressed as a diffraction angle of 2θ, with characteristic peaks at 4.603, 9.209, 12.888, 13.920, 15.097, 18.641, 19.194, 19.700, 20.529, 22.876, 25.454, 25.664, 27.775, and 29.504.

[0033] In an optional embodiment, the E crystal form of the compound represented by formula (I) provided by the present disclosure has an X-ray powder diffraction pattern expressed as a diffraction angle 2θ, and has characteristic peaks at 4.603, 9.209, 12.888, 13.920, 15.097, 18.641, 19.194, 19.700, 20.529, 21.863, 22.876, 23.659, 24.349, 25.054, 25.454, 25.664, 27.203, 27.698, 27.775, 28.906, 29.504, 29.965, 30.747, 31.944, 34.430, 38.756, 39.263, and 42.577.

[0034] A method for preparing the E crystal form of the compound represented by formula (I), the method being selected from:

[0035] a) mixing the compound represented by formula (I) with solvent V to obtain a clear solution, and mixing the clear solution with solvent VI to obtain crystals, wherein the solvent V is selected from at least one of N,N-dimethylformamide, N,N-dimethylformamide, methanol, dimethyl sulfoxide, and dichloromethane; and the solvent VI is selected from at least one of isopropyl ether, methyl tert-butyl ether, isopropyl alcohol, dioxane, acetone, n-hexane, toluene, and acetonitrile; or

[0036] b) mixing the compound represented by formula (I) with a dichloromethane-acetonitrile solvent, dissolving it, and crystallizing it.

[0037] The present disclosure provides an F-type of a compound represented by formula (I), which has an X-ray powder diffraction pattern represented by a diffraction angle 2θ, with characteristic peaks at 4.656, 14.068, 15.183, 18.858, and 23.235.

[0038] In an optional embodiment, the F crystal form of the compound represented by formula (I) provided by the present disclosure has an X-ray powder diffraction pattern expressed as a diffraction angle 2θ, and has characteristic peaks at 2θ angles of 4.656, 9.341, 12.670, 13.443, 14.068, 14.850, 15.183, 18.858, 19.436, 20.518, 23.235, 25.591, 28.386, and 29.543.

[0039] In an optional embodiment, the F crystalline form of the compound represented by formula (I) provided by the present disclosure has an X-ray powder diffraction pattern expressed in terms of diffraction angle 2θ, with the following peaks: 4.656, 9.341, 12.670, 13.443, 14.068, 14.850, 15.183, 16.122, 18.032, 18.858, 19.436, 20.518, 22.15 There are characteristic peaks at 0, 22.571, 23.235, 24.442, 24.863, 25.591, 26.585, 27.765, 28.386, 28.934, 29.543, 30.245, 31.113, 32.116, 32.491, 36.608, 38.199, 38.620, 40.679, and 43.206.

[0040] The present disclosure provides a method for preparing the F crystal form of the compound represented by formula (I), comprising mixing the compound represented by formula (I) with ethanol-water and crystallizing.

[0041] The present disclosure provides a G crystal form of a compound represented by formula (I), which has an X-ray powder diffraction pattern represented by a diffraction angle 2θ, with characteristic peaks at 4.869, 9.735, 13.290, 14.713, and 20.020.

[0042] In an optional embodiment, the G crystal form of the compound represented by formula (I) provided by the present disclosure has an X-ray powder diffraction pattern expressed as a diffraction angle 2θ, with characteristic peaks at 4.869, 9.735, 10.863, 13.290, 14.713, 14.968, 17.852, 19.399, 20.020, 20.593, 21.796, 22.793, 24.251, 24.681, 25.510, 25.907, 26.327, 27.617, and 30.155.

[0043] In an optional embodiment, the present disclosure provides a G crystal form of the compound represented by formula (I), the X-ray powder diffraction pattern expressed in terms of diffraction angle 2θ is 4.869, 9.735, 10.863, 13.290, 14.713, 14.968, 17.852, 19.399, 20.020, 20.593, 21.796, 22.79 3. There are characteristic peaks at 24.251, 24.681, 25.510, 25.907, 26.327, 27.617, 28.051, 29.696, 30.155, 31.101, 32.405, 33.402, 35.019, 39.615, 41.021, 45.193, 46.644, and 54.898.

[0044] A method for preparing a G crystal form of a compound represented by formula (I) comprises mixing the compound represented by formula (I) with ethanol-water, dissolving the clear solution, mixing the solution with water, crystallizing the solution, and drying the solution at 40° C. for 3 days.

[0045] The present disclosure provides an H crystal form of a compound represented by formula (I), which has an X-ray powder diffraction pattern represented by a diffraction angle 2θ, with characteristic peaks at 8.608, 12.983, 13.476, 17.716, 20.144, and 23.371.

[0046] In an optional embodiment, the H crystal form of the compound represented by formula (I) provided by the present disclosure has an X-ray powder diffraction pattern expressed as a diffraction angle 2θ, with characteristic peaks at 8.608, 11.238, 12.983, 13.476, 17.716, 18.227, 19.392, 20.144, 21.435, 21.914, 22.935, 23.371, 23.814, 25.416, and 30.620.

[0047] In an optional embodiment, the H crystal form of the compound represented by formula (I) provided by the present disclosure has an X-ray powder diffraction pattern expressed in terms of diffraction angle 2θ, with the following peaks: 8.608, 11.238, 11.718, 12.983, 13.476, 14.485, 17.716, 18.227, 19.392, 20.144, 21.435, 21.914, 2 There are characteristic peaks at 2.935, 23.371, 23.814, 25.416, 26.499, 28.195, 28.887, 30.620, 31.767, 32.874, 33.970, 34.839, 35.854, 37.045, 38.478, 39.958, 41.311, 43.925, and 44.582.

[0048] A method for preparing the H crystal form of a compound represented by formula (I), comprising mixing the compound represented by formula (I) with ethanol-water and allowing crystallization to precipitate.

[0049] In certain embodiments, the method for preparing the crystalline form described in the present disclosure further comprises a filtering, washing or drying step.

[0050] The present disclosure provides a crystalline form A, a crystalline form B, a crystalline form C, a crystalline form D, a crystalline form E, a crystalline form F, a crystalline form G and a crystalline form H of a compound represented by formula (I) prepared by the above-mentioned preparation method.

[0051] In an optional embodiment, in the compound of formula (I) and its corresponding crystalline form disclosed herein, each deuterium atom (D) has an abundance of at least 20%.

[0052] In an optional embodiment, in the compound of formula (I) and its corresponding crystalline form disclosed herein, each deuterium atom (D) has an abundance of at least 50%.

[0053] In an optional embodiment, in the compound of formula (I) and its corresponding crystalline form disclosed herein, each deuterium atom (D) has an abundance of at least 90%.

[0054] In an optional embodiment, in the compound of formula (I) and its corresponding crystalline form disclosed herein, each deuterium atom (D) has an abundance of at least 98%.

[0055] The present disclosure also provides a pharmaceutical composition comprising a crystalline form of the compound represented by the aforementioned formula (I), or a crystalline form of the compound represented by the aforementioned formula (I) prepared by the aforementioned method, or a mixture thereof, and optionally a pharmaceutically acceptable carrier, diluent or excipient.

[0056] The present disclosure also provides a pharmaceutical composition prepared from the crystal form of the compound represented by the aforementioned formula (I).

[0057] The present disclosure also provides a method for preparing a pharmaceutical composition, comprising the step of mixing the crystalline form of the compound represented by the aforementioned formula (I), or the crystalline form of the compound represented by the aforementioned formula (I) prepared by the aforementioned method, or a mixture thereof with a pharmaceutically acceptable carrier, diluent or excipient.

[0058] The present disclosure also provides the use of a crystalline form of the compound represented by the aforementioned formula (I), or a crystalline form of the compound represented by the aforementioned formula (I) prepared by the aforementioned method, or a mixture thereof, or the aforementioned composition, or a composition prepared by the aforementioned method in the preparation of a medicament for treating diseases or conditions related to SHP2 regulation.

[0059] The present disclosure also provides the crystal form of the compound represented by the aforementioned formula (I), or the crystal form of the compound represented by the aforementioned formula (I) prepared by the aforementioned method, or a mixture thereof, or the aforementioned composition, or the use of the composition prepared by the aforementioned method in preparing a drug for preventing and / or treating tumors or cancer.

[0060] The present disclosure also provides a crystalline form of the compound represented by the aforementioned formula (I), or a crystalline form of the compound represented by the aforementioned formula (I) prepared by the aforementioned method, or the aforementioned composition, or use of the composition prepared by the aforementioned method in preparing a drug for preventing or treating Noonan syndrome or Leopard skin syndrome.

[0061] The present disclosure also provides a crystalline form of the compound represented by the aforementioned formula (I), or a crystalline form of the compound represented by the aforementioned formula (I) prepared by the aforementioned method, or the aforementioned composition, or the use of the composition prepared by the aforementioned method in preparing a drug for preventing or treating juvenile myelomonocytic leukemia, neuroblastoma, melanoma, acute myeloid leukemia, breast cancer, esophageal cancer, lung cancer, colon cancer, head cancer, pancreatic cancer, head and neck squamous cell carcinoma, gastric cancer, liver cancer, anaplastic large cell lymphoma and glioblastoma.

[0062] 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, which can be -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.

[0063] The "crystallization" mentioned in the present disclosure includes but is not limited to stirring crystallization, cooling crystallization, beating crystallization and volatile crystallization.

[0064] The "differential scanning calorimetry or DSC" described in this 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 the sample at a constant temperature to characterize all physical and chemical changes related to thermal effects and obtain phase change information of the sample.

[0065] The drying temperature in the present disclosure is generally 25°C-100°C, preferably 40°C-70°C, and can be dried under normal pressure or reduced pressure.

[0066] Unless otherwise indicated, when a position is specifically designated as deuterium (D), the position is understood to have an abundance of deuterium that is at least 1000 times greater than the natural abundance of deuterium, which is 0.015% (i.e., at least 10% deuterium incorporation). Examples of compounds having an abundance of deuterium greater than the natural abundance of deuterium can be at least 1000 times greater, at least 2000 times greater, at least 3000 times greater, at least 4000 times greater, at least 5000 times greater, at least 6000 times greater, or more.

[0067] The contents of WO2020259679A are incorporated herein by reference. BRIEF DESCRIPTION OF THE DRAWINGS

[0068] Figure 1 .XRPD spectrum of the amorphous form of the compound represented by formula (I).

[0069] Figure 2 .XRPD spectrum of the crystal form A of compound represented by formula (I).

[0070] Figure 3 .TGA spectrum of the crystal form A of compound represented by formula (I).

[0071] Figure 4 .DSC spectrum of crystal form A of compound represented by formula (I).

[0072] Figure 5 .XRPD comparison spectra of the crystal form A of compound represented by formula (I) before and after DVS.

[0073] Figure 6 .XRPD spectrum of the crystalline form B of compound represented by formula (I).

[0074] Figure 7 .XRPD spectrum of crystal form C of compound represented by formula (I).

[0075] Figure 8 .XRPD spectrum of the crystal form D of compound represented by formula (I).

[0076] Figure 9 .XRPD spectrum of crystal form E of compound represented by formula (I).

[0077] Figure 10 .XRPD spectrum of the crystal form F of compound represented by formula (I).

[0078] Figure 11 .XRPD spectrum of the crystalline form G of compound represented by formula (I).

[0079] Figure 12 .XRPD spectrum of H crystal form of the compound represented by formula (I). DETAILED DESCRIPTION

[0080] The present invention is further described in detail by the following examples and experimental examples. These examples and experimental examples are for illustrative purposes only and are not intended to limit the scope of the present invention.

[0081] Test conditions of the instruments used in the experiment:

[0082] The structures of the compounds were confirmed by nuclear magnetic resonance (NMR) and / or mass spectrometry (MS). NMR shifts (δ) are given in units of 10⁻⁶ (ppm). NMR measurements were performed using a Bruker AVANCE-400 NMR spectrometer. The solvents used were deuterated dimethyl sulfoxide (DMSO-d₆), deuterated chloroform (CDCl₃), and deuterated methanol (CD₃OD), with tetramethylsilane (TMS) as the internal standard.

[0083] MS was measured using a Shimadzu 2010 Mass Spectrometer or an Agilent 6110A MSD mass spectrometer.

[0084] HPLC analysis was performed using an Agilent 1260DAD high pressure liquid chromatograph (Sunfire C18 150×4.6 mm column) and a Thermo U3000 high pressure liquid chromatograph (Gimini C18 150×4.6 mm column).

[0085] HPLC determination was performed using a Shimadzu LC-20A systems, Shimadzu LC-2010HT series, or Agilent 1200 LC high pressure liquid chromatograph (Ultimate XB-C18 3.0*150 mm column or Xtimate C18 2.1*30 mm column).

[0086] Chiral HPLC analysis was performed using Chiralpak IC-3 100×4.6mm ID, 3μm, Chiralpak AD-3 150×4.6mm ID, 3μm, Chiralpak AD-3 50×4.6mm ID, 3μm, Chiralpak AS-3 150×4.6mm ID, 3μm, Chiralpak AS-3 100×4.6mm ID, 3μm, ChiralCel OD-3 150×4.6mm ID, 3μm, Chiralcel OD-3 100×4.6mm ID, 3μm, ChiralCel OJ-H 150×4.6mm ID, 5μm, and Chiralcel OJ-3 150×4.6mm ID, 3μm columns.

[0087] The thin layer chromatography silica gel plate uses Yantai Huanghai HSGF254 or Qingdao GF254 silica gel plate. The specification of the silica gel plate used in thin layer chromatography (TLC) is 0.15mm~0.2mm, and the specification used for thin layer chromatography separation and purification products is 0.4mm~0.5mm.

[0088] Column chromatography generally uses Yantai Huanghai silica gel 100-200 mesh, 200-300 mesh or 300-400 mesh silica gel as the carrier.

[0089] The chiral preparative column used was DAICEL CHIRALPAK IC (250 mm*30 mm, 10 μm) or Phenomenex-Amylose-1 (250 mm*30 mm, 5 μm).

[0090] The CombiFlash rapid preparation instrument used was Combiflash Rf150 (TELEDYNE ISCO).

[0091] Average kinase inhibition rate and IC 50 The values were determined using a NovoStar microplate reader (BMG, Germany).

[0092] 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.

[0093] Unless otherwise specified in the examples, all reactions can be carried out under an argon atmosphere or a nitrogen atmosphere.

[0094] Argon atmosphere or nitrogen atmosphere means that the reaction bottle is connected to an argon or nitrogen balloon with a capacity of about 1 L.

[0095] Hydrogen atmosphere means that the reaction bottle is connected to a hydrogen balloon with a capacity of about 1L.

[0096] The pressurized hydrogenation reaction uses a Parr 3916EKX hydrogenator and a Qinglan QL-500 hydrogen generator or an HC2-SS hydrogenator.

[0097] The hydrogenation reaction is usually carried out by evacuating the chamber and filling it with hydrogen, and the operation is repeated three times.

[0098] A CEM Discover-S 908860 microwave reactor was used for the microwave reaction.

[0099] Unless otherwise specified in the examples, the solution refers to an aqueous solution.

[0100] Unless otherwise specified in the examples, the reaction temperature is room temperature, 20°C to 30°C.

[0101] The reaction progress in the examples was monitored by thin layer chromatography (TLC). The developing solvent used in the reaction, the column chromatography eluent system used for purifying the compound, and the developing solvent system for thin layer chromatography included: A: dichloromethane / methanol system, B: n-hexane / ethyl acetate system, C: petroleum ether / ethyl acetate system, and D: petroleum ether / ethyl acetate / methanol. The volume ratio of the solvent was adjusted according to the polarity of the compound, and a small amount of alkaline or acidic reagents such as triethylamine and acetic acid could also be added for adjustment.

[0102] XRPD is X-ray powder diffraction detection: the measurement is carried out using a BRUKERD8 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: 3-48°. DSC stands for differential scanning calorimetry: measured using a METTLERTOLEDO DSC3+, a heating rate of 10°C / min, from 25 to 350°C, and a nitrogen purge rate of 50 mL / min.

[0103] TGA is thermogravimetric analysis: the detection was carried out using METTLER TOLEDO TGA2, with a heating rate of 10°C / min, the specific temperature range is referred to the corresponding spectrum, and a nitrogen purge rate of 50 mL / min.

[0104] DVS stands for dynamic moisture sorption: using Surface Measurement Systems advantage 2, starting from 50% humidity, the humidity range of investigation is 0%-95%, with a step of 10%. The judgment standard is that the mass change is less than 0.002% within 360 minutes, and the cycle is repeated twice.

[0105] The structure of metabolite 14 is as follows:

[0106]

[0107] Example 1

[0108] (S)-1′-(8-((2-amino-3-chloropyridin-4-yl)thio)imidazo[1,2-c]pyrimidin-5-yl)-5,7-dihydrospiro[cyclopenta[b]pyridine-6,4′-piperidin]-5-amine

[0109]

[0110]

[0111] first step

[0112] (3-Bromopyridin-2-yl)methanol 1b

[0113] Compound 1a (17.2 g, 79.6 mmol) was dissolved in methanol (50 mL), and sodium borohydride (15.1 g, 398 mmol) was added at 0°C. The reaction system was stirred at room temperature for 12 hours. After the reaction, saturated aqueous ammonium chloride (600 mL) was added, and the mixture was extracted with ethyl acetate (200 mL x 3). The organic phases were combined, washed with saturated sodium chloride (200 mL x 2), dried over anhydrous sodium sulfate, and concentrated under reduced pressure to obtain compound 1b (9.7 g, yield: 64.8%) as a white solid.

[0114] MS (ESI) m / z 187.8 [M+H] +

[0115] 1 H NMR: (400MHz, MeOD-d4) δ=8.52 (d, J=4.8Hz, 1H), 8.01 (dd, J=1.2, 8.0Hz, 1H), 7.26 (dd, J=4.4, 6.4Hz, 1H), 4.77 (s, 2H).

[0116] Step 2

[0117] 3-Bromo-2-(chloromethyl)pyridine 1c

[0118] Compound 1b (9.70 g, 51.6 mmol) was dissolved in dichloromethane (20 mL), and thionyl chloride (7.48 mL, 103 mmol) was added at room temperature. Stir at room temperature for 3 hours. After the reaction was complete, saturated aqueous sodium bicarbonate solution (300 mL) was added at 0°C, and the mixture was extracted with dichloromethane (80 mL x 3). The organic phases were combined, washed with saturated sodium chloride (100 mL), and dried over anhydrous sodium sulfate. Concentration under reduced pressure gave Compound 1c (10.3 g, 96.9% yield) as a pink oil.

[0119] MS (ESI) m / z 207.7 [M+H] +

[0120] 1 H NMR (400MHz, Methanol-d4) = 8.55-8.45 (m, 1H), 8.12-7.99 (m, 1H), 7.37-7.21 (m, 1H), 4.84-4.80 (m, 2H).

[0121] Step 3

[0122] 1-(tert-Butyl)4-ethyl 4-((3-bromopyridin-2-yl)methyl)piperidine-1,4-dicarboxylate 1e

[0123] Under a nitrogen atmosphere, compound 1c (9.97 g, 38.7 mmol) was dissolved in tetrahydrofuran (80 mL) and LDA (13.5 mL, 2 M tetrahydrofuran and n-hexane solution) was added dropwise at -78°C. After the addition was complete, the mixture was stirred at -78°C for 1 hour. Compound 1d (8.8 g, 35.07 mmol) was then added dropwise at -78°C and stirred at -78°C for another 9 hours. After the reaction was complete, saturated aqueous ammonium chloride (400 mL) was added, and the mixture was extracted with ethyl acetate (100 mL x 3). The organic phases were combined, washed with saturated sodium chloride solution (100 mL x 2), and dried over anhydrous sodium sulfate. The crude product was concentrated in vacuo and purified by silica gel chromatography using petroleum ether and ethyl acetate to afford compound 1e (14.8 g, 89.4% yield) as a yellow oil.

[0124] MS (ESI) m / z 429.0 [M+H] +

[0125] Step 4

[0126] 4-((3-bromopyridin-2-yl)methyl)-1-(tert-butoxycarbonyl)piperidine-4-carboxylic acid 1f

[0127] Compound 1e (14.8 g, 34.6 mmol) was dissolved in methanol (3 mL), and sodium hydroxide aqueous solution (13.8 g, 346 mmol, dissolved in 40 mL of water) was added at 0°C and stirred at 80°C for 12 hours. After the reaction was completed, the reaction solution was concentrated, and ethyl acetate (300 mL) and water (300 mL) were added thereto. Saturated sodium hydroxide aqueous solution (10 mL) was added to adjust the pH to 12, the aqueous phase was separated, and washed with ethyl acetate (80 mL × 2). 2N hydrochloric acid (25 mL) was added to the obtained aqueous phase to adjust the pH to 3, and extracted with ethyl acetate (100 mL × 3). The organic phases were combined, washed with saturated sodium chloride solution (150 mL), dried over anhydrous sodium sulfate, and concentrated under reduced pressure to obtain a white solid compound 1f (11.4 g, yield: 82.4%).

[0128] MS (ESI) m / z 344.0 [M-56+H] +

[0129] Step 5

[0130] tert-Butyl 5-carbonyl-5,7-dihydrospiro[cyclopenta[b]pyridine-6,4′-piperidine]-1′-carboxylate 1g

[0131] Under a nitrogen atmosphere, sodium hydride (60% kerosene mixture, 1.32 g, 33.1 mmol) was added to a solution of compound 1f (11.0 g, 27.6 mmol) in tetrahydrofuran (100 mL) at -15°C. The mixture was stirred at -15°C for 1 hour. The reaction solution was then cooled to -78°C, and a 2.5 M solution of n-butyllithium in n-hexane (16.5 mL, 41.3 mmol) was added dropwise, followed by stirring at -78°C for 1 hour. After completion of the reaction, saturated aqueous ammonium chloride (400 mL) was added at 0°C, and the mixture was extracted with ethyl acetate (100 mL x 3). The organic phases were combined, washed with saturated sodium chloride (100 mL x 2), and dried over anhydrous sodium sulfate. The crude product was concentrated in vacuo and purified by silica gel chromatography using dichloromethane and methanol as eluents to afford compound 1g (4.60 g, 55.2% yield) as a white solid.

[0132] MS (ESI) m / z 246.9 [M-56+H] + .

[0133] 1 H NMR (400MHz, Methanol-d4)□□=8.82 (dd, J=1.6, 4.8Hz, 1H), 8.12 (dd, J=1.6, 7.6Hz, 1H), 7.50 (dd, J=4.8, 7.6Hz, 1H), 4.08 (td, J=3.6, 13.6Hz, 2H), 3.25 (s, 2H), 3.12 (br s, 2H), 1.88-1.77 (m, 2H), 1.51 (br s, 2H), 1.49 (s, 9H).

[0134] Step 6

[0135] Tert-butyl (S)-5-((S)-tert-butylsulfonamido)-5,7-dihydrospiro[cyclopenta[b]pyridine-6,4′-piperidine]-1′-carboxylate 1i

[0136] Under a nitrogen atmosphere, tetraethyl titanate (9.4 mL, 44.6 mmol) was added to anhydrous toluene (80 mL) of compound 1g (4.50 g, 14.9 mmol) and stirred at room temperature for 10 minutes. Compound 1h (5.4 g, 44.6 mmol) was then added to the reaction solution and reacted at 120°C for 5 hours. After cooling to 0°C and adding lithium borohydride (1.58 g, 89.2 mmol), the reaction was continued for 30 minutes, then the temperature was raised to room temperature and stirred for 1 hour. After the reaction was completed, methanol (20 mL) was added dropwise at 0°C. Water (100 mL) and ethyl acetate (100 mL) were added and stirred for 5 minutes. The suspended matter was filtered off with diatomaceous earth and washed with ethyl acetate (300 mL) and water (300 mL). The organic phases were combined, washed with saturated sodium chloride (500 mL), and dried over anhydrous sodium sulfate. The crude product was concentrated under reduced pressure and purified by silica gel chromatography using petroleum ether and ethyl acetate as eluents to obtain yellow solid compound 1i (4.40 g, yield: 72.6%).

[0137] MS (ESI) m / z 408.1 [M+H] +

[0138] Step 7

[0139] (S)-N-((S)-5,7-Dihydrospiro[cyclopenta[b]pyridine-6,4′-piperidin]-5-yl)-2-methylpropane-2-sulfenamide 1j: Compound 1i (4.40 g, 10.8 mmol) was dissolved in dichloromethane (15 mL). Trifluoroacetic acid (5 mL) was added at 0°C and stirred at 0°C for 1 hour. The mixture was concentrated under reduced pressure to obtain a crude product, which was then added with 4 M aqueous sodium hydroxide until the pH reached 11. The product was extracted with chloroform and isopropanol (volume ratio 3:1) (30 mL x 3). The organic phases were combined and dried over anhydrous sodium sulfate. The mixture was concentrated under reduced pressure to obtain the product 1j (3.32 g, yield: 100%) as a yellow oil.

[0140] MS (ESI) m / z 307.9 [M+H] +

[0141] Step 8

[0142] (S)-N-((S)-1′-(8-bromoimidazo[1,2-c]pyrimidin-5-yl)-5,7-dihydrospiro[cyclopenta[b]pyridine-6,4′-piperidin]-5-yl)-2-methylpropane-2-sulfenamide 11

[0143] Under a nitrogen atmosphere, compound 1j (3.30 mg, 10.7 mmol) and compound 1k (2.50 g, 10.7 mmol) were dissolved in dimethyl sulfoxide (40 mL), and diisopropylethylamine (7.7 g, 59.8 mmol) was added. The mixture was stirred at 90°C for 2 hours. Ethyl acetate (50 mL) and water (100 mL) were added, and extraction with ethyl acetate (50 mL x 2) was performed. The organic phases were combined, washed with saturated sodium chloride solution (50 mL x 3), and dried over anhydrous sodium sulfate. The crude product was concentrated under reduced pressure and purified by silica gel chromatography using dichloromethane and methanol as eluents to obtain compound 1l (2.96 g, yield: 54.6%).

[0144] MS (ESI) m / z 503.1 [M+H] +

[0145] 1 H NMR (400MHz, METHANOL-d4)□ = 8.41 (d, J = 4.8Hz, 1H), 7.97 (s, 1H), 7.92 (d, J = 1.5Hz, 1 H), 7.81 (d, J=7.5Hz, 1H), 7.66 (d, J=1.5Hz, 1H), 7.32 (dd, J=5.0, 7.5Hz, 1H), 4.61 (br s, 2H), 3.95-3.83 (m, 2H), 3.30-3.21 (m, 2H), 2.99 (d, J=16.6Hz, 1H), 2.40 (dt, J=4.0, 12.7Hz, 1H), 2.14 (dt, J=3.6, 12.4Hz, 1H), 1.82 (br d, J=13.3Hz, 1H), 1.54 (br d, J=12.3Hz, 1H), 1.36 (s, 9H).

[0146] Step 9

[0147] (S)-N-((S)-1′-(8-((2-amino-3-chloropyridin-4-yl)thio)imidazo[1,2-c]pyrimidin-5-yl)-5,7-dihydrospiro[cyclopenta[b]pyridine-6,4′-piperidin]-5-yl)-2-methylpropane-2-sulfenamide 1n

[0148] Under a nitrogen atmosphere, compound 1l (70 mg, 0.14 mmol) and compound 1m (33 mg, 0.21 mmol, prepared using the method disclosed in patent application "WO2015107495A1") were dissolved in 1,4-dioxane (1 mL), and diisopropylethylamine (54 mg, 0.42 mmol) was added at room temperature. Tris(dibenzylideneacetone)dipalladium (13 mg, 0.014 mmol) and 2-dicyclohexylphosphine-2',6'-dimethoxybiphenyl (14 mg, 0.028 mmol) were added, and the mixture was heated and stirred at 110°C for 12 hours. After completion of the reaction, the reaction solution was filtered, the filtrate was concentrated, and the residue was purified by C-18 reverse-phase chromatography eluting with water and methanol to obtain compound 1n (45 mg, 55.1% yield) as a brown oil.

[0149] MS (ESI) m / z 583.1 [M+H] +

[0150] Step 10

[0151] (S)-1′-(8-((2-amino-3-chloropyridin-4-yl)thio)imidazo[1,2-c]pyrimidin-5-yl)-5,7-dihydrospiro[cyclopenta[b]pyridine-6,4′-piperidin]-5-amine 1

[0152] Compound 1n (25 mg, 0.035 mmol) was dissolved in 1,4-dioxane. A solution of hydrogen chloride in 1,4-dioxane (0.2 mL, 4 N) was added at 0°C and allowed to react at 2-7°C for 1 hour. After completion of the reaction, water (30 mL) was added and the mixture was extracted with ethyl acetate (15 mL x 2). The organic phases were combined, washed with saturated sodium chloride solution (20 mL), dried over anhydrous sodium sulfate, and concentrated under reduced pressure. The residue was purified by C-18 reverse-phase chromatography to yield compound 1 (3.9 mg, yield: 19.0%).

[0153] MS (ESI) m / z 479.1 [M+H] +

[0154] 1H NMR: (400MHz, MeOD-d4) δ = 8.38 (d, J = 4.8Hz, 1H), 8.06 (s, 1H), 7.90-7.84 (m, 2H), 7.57 (s, 1H), 7.50 (d, J=5.2Hz, 1H), 7.30 (dd, J=5.6, 7.6Hz, 1H), 5.90 (d, J=6.0Hz, 1H), 4.16 (s, 1H), 4.06 (br d, J=13.6Hz, 2H), 3.48-3.36(m, 2H), 3.30-3.24(m, 1H), 3.01(br d, J=16.4Hz, 1H), 2.20-2.01(m, 2H), 1.80-1.71(m, 1H), 1.61-1.53(m, 1H).

[0155] Example 2

[0156] (S)-1′-(8-((3-chloro-2-(methylamino)pyridin-4-yl)thio)-7-methylimidazo[1,2-c]pyrimidin-5-yl)-5,7-dihydrospiro[cyclopenta[b]pyridine-6,4′-piperidin]-5-amine

[0157]

[0158]

[0159] first step

[0160] (S)-N-((S)-1′-(8-bromo-7-methylimidazo[1,2-c]pyrimidin-5-yl)-5,7-dihydrospiro[cyclopenta[b]pyridine-6,4′-piperidin]-5-yl)-2-methylpropane-2-sulfenamide 2b

[0161] Compound 1j (260 mg, 0.85 mmol) and compound 2a (271 mg, 1.10 mmol) were dissolved in dimethyl sulfoxide (3 mL), and diisopropylethylamine (547 mg, 4.23 mmol) was added. The mixture was stirred at 90°C for 1 hour. Water (30 mL) was added to the reaction solution, and the mixture was extracted with ethyl acetate (30 mL × 3). The organic phases were combined, washed with saturated sodium chloride solution (50 mL × 2), dried over anhydrous sodium sulfate, and filtered. The filtrate was concentrated under reduced pressure to obtain a crude product, which was purified by silica gel chromatography using methanol and dichloromethane to obtain compound 2b (370 mg, yield: 84.5%).

[0162] MS (ESI) m / z 518.8 [M+H] +

[0163] 1H NMR (400MHz, Methanol-d4) δ=8.39 (d, J=4.8Hz, 1H), 7.82 (d, J=1.2Hz, 1H), 7.79 (d, J=8. 0Hz, 1H), 7.55 (d, J=1.6Hz, 1H), 7.30 (dd, J=4.8Hz, 7.6Hz, 1H), 3.90-3.81 (m, 2H), 3.37-3 .32(m, 1H), 3.29-3.17(m, 3H), 3.00-2.92(m, 1H), 2.57(s, 3H), 2.37(td, J=4.4Hz, 12.8Hz , 1H), 2.13 (td, J=4.4Hz, 13.2Hz, 1H), 1.83-1.75 (m, 1H), 1.56-1.49 (m, 1H), 1.34 (s, 9H).

[0164] Step 2

[0165] (S)-N-((S)-1′-(8-((3-chloro-2-(methylamino)pyridin-4-yl)thio)-7-methylimidazo[1,2-c]pyrimidin-5-yl)-5,7-dihydrospiro[cyclopenta[b]pyridine-6,4′-piperidin]-5-yl)-2-methylpropane-2-sulfenamide 2d

[0166] Compound 2b (50 mg, 0.10 mmol), compound 2c (77 mg, 0.39 mmol), and potassium phosphate (41 mg, 0.19 mmol) were dissolved in 1,4-dioxane (1 mL) and the atmosphere was replaced with nitrogen three times while stirring. Under a nitrogen atmosphere, 1,10-phenanthroline (3.5 mg, 0.02 mmol) and cuprous iodide (1.8 mg, 0.01 mmol) were quickly added. The atmosphere was replaced with nitrogen three times again, and the mixture was heated and stirred at 130°C for 10 hours. Water (50 mL) was added to the reaction solution, and the mixture was extracted with ethyl acetate (40 mL × 3). The organic phases were combined, washed with saturated sodium chloride solution (70 mL × 2), dried over anhydrous sodium sulfate, and filtered. The filtrate was concentrated under reduced pressure to obtain a crude product, which was purified by silica gel plate chromatography using dichloromethane and methanol to obtain compound 2d (36 mg, yield: 58.5%).

[0167] MS (ESI) m / z 611.1 [M+H] +

[0168] Step 3

[0169] (S)-1′-(8-((3-chloro-2-(methylamino)pyridin-4-yl)thio)-7-methylimidazo[1,2-c]pyrimidin-5-yl)-5,7-dihydrospiro[cyclopenta[b]pyridine-6,4′-piperidin]-5-amine 2

[0170] Compound 2d (36 mg, 0.059 mmol) was dissolved in dry dioxane (1 mL), and a solution of hydrogen chloride in 1,4-dioxane (1 mL, 4N) was added dropwise at 10°C. The mixture was reacted at 10°C for 15 minutes. Water (30 mL) was added to the suspended reaction solution, and the mixture was extracted with ethyl acetate (30 × 3). The aqueous phase was adjusted to pH = 8 with a saturated aqueous sodium bicarbonate solution and then extracted with chloroform (40 mL × 4). All organic phases were combined, dried over anhydrous sodium sulfate, filtered, and the filtrate was concentrated under reduced pressure to obtain a crude product. The crude product was purified by high performance liquid chromatography and lyophilized to obtain compound 2 (2.3 mg, yield: 7.7%).

[0171] MS (ESI) m / z 507.3 [M+H] +

[0172] 1H NMR (400MHz, Methanol-d4) δ=8.35 (d, J=4.4Hz, 1H), 7.85 (d, J=7.6Hz, 1H), 7.76 (d, J=1. 6Hz, 1H), 7.58 (d, J=5.6Hz, 1H), 7.48 (d, J=1.6Hz, 1H), 7.29 (dd, J=5.2Hz, 7.6Hz, 1H), 5. 75(d, J=6.0Hz, 1H), 4.12-4.00(m, 3H), 3.46-3.34(m, 2H), 3.29-3.23(m, 1H), 3.01-2.92 (m, 4H), 2.55 (s, 3H), 2.17-2.01 (m, 2H), 1.74 (d, J = 13.6Hz, 1H), 1.53 (d, J = 13.6Hz, 1H).

[0173] Example 3

[0174] (S)-1′-(8-((3-chloro-2-((methyl-d3)amino)pyridin-4-yl)thio)-7-methylimidazo[1,2-c]pyrimidin-5-yl)-5,7-dihydrospiro[cyclopenta[b]pyridine-6,4′-piperidin]-5-amine

[0175]

[0176] Intermediate 3e

[0177] Sodium 2-((methyl-d3)amino)-3-chloropyridine-4-thiolate

[0178]

[0179]

[0180] first step

[0181] 3-Chloro-4-iodo-N-(methyl-d3)pyridin-2-amine 3b

[0182] Compound 3a (3.0 g, 12 mmol) and methyl-d3-amine hydrochloride (1.2 g, 16 mmol) were dissolved in DMSO (50 mL), and DIEA (5.8 mL, 35 mmol) was added. The mixture was reacted at 70°C for 12 hours. After completion of the reaction, an ice-water mixture (50 mL) was added, filtered, and washed with ice water (50 mL x 3). The resulting solid was dried under reduced pressure to afford compound 3b (2.8 g, yield: 83%).

[0183] MS (ESI) m / z 272.0 [M+H] +

[0184] 1 H NMR: (400MHz, CDCl3) δ=7.67 (d, J=5.2Hz, 1H), 7.02 (d, J=5.2Hz, 1H), 5.14 (br s, 1H).

[0185] Step 2

[0186] Ethyl 3-((3-chloro-2-((methyl-d3)amino)pyridin-4-yl)thio)propanoate 3d

[0187] Compound 3b (2.7 g, 10 mmol) was dissolved in dichloromethane (30 mL), and ethyl 3-mercaptopropionate 3c (2.0 g, 15 mmol), tris(dibenzylideneacetone) (0.46 g, 0.50 mmol), 4,5-bis(diphenylphosphino-9,9-dimethylxanthene) (0.58 g, 0.99 mmol), and N,N-diisopropylethylamine (4.9 mL, 30 mmol) were added. The reaction was allowed to proceed at 100°C under a nitrogen atmosphere for 3 hours. After completion of the reaction, the product was filtered, and the filtrate was concentrated under reduced pressure. The resulting residue was purified by silica gel chromatography using ethyl acetate and petroleum ether as eluents to afford compound 3d (2.5 g, 90% yield).

[0188] MS (ESI) m / z 278.1 [M+H] +

[0189] 1H NMR (400MHz, CDCl3) δ=7.96 (d, J=5.6Hz, 1H), 6.45 (d, J=5.6Hz, 1H), 5.00 (br s, 1H), 4.19 (q, J=7.2Hz, 2H), 3.23 (t, J=7.6Hz, 2H), 2.72 (t, J=7.6Hz, 2H), 1.28 (t, J=7.2Hz, 3H).

[0190] Step 3

[0191] Sodium 2-((methyl-d3)amino)-3-chloropyridine-4-thiolate intermediate 3e

[0192] Compound 3d (2.4 g, 8.6 mmol) was dissolved in tetrahydrofuran (25 mL). Sodium ethoxide in ethanol (3.5 g, 10 mmol, 20% w / w) was added at 0°C and allowed to react for 1 hour at 0°C. After completion of the reaction, the reaction solution was concentrated, and a 50:1 mixture of methyl tert-butyl ether and dichloromethane (20 mL) was added. The mixture was filtered and washed with 50:1 methyl tert-butyl ether and dichloromethane (10 mL x 3). The resulting solid was dried under reduced pressure to afford intermediate 3e (2.0 g, 99% yield).

[0193] MS (ESI) m / z 178.0 [M+H] +

[0194] 1 H NMR (400MHz, DMSO_d6) δ=7.12 (d, J=5.6Hz, 1H), 6.43 (d, J=5.2Hz, 1H), 5.29 (s, 1H).

[0195] Step 4

[0196] (S)-N-((S)-1′-(8-((3-chloro-2-((methyl-d3)amino)pyridin-4-yl)thio)-7-methylimidazo[1,2-c]pyrimidin-5-yl)-5,7-dihydrospiro[cyclopenta[b]pyridine-6,4′-piperidin]-5-yl)-2-methylpropane-2-sulfenamide 3f

[0197] Under a nitrogen atmosphere, compound 2b (200 mg, 0.39 mmol) and intermediate 3e (156 mg, 0.77 mmol) were dissolved in 1,4-dioxane (5 mL). Cuprous iodide (74 mg, 0.39 mmol), N,N′-dimethylethylenediamine (34 mg, 0.39 mmol), and potassium phosphate (246 mg, 1.2 mmol) were added. The mixture was heated at 130°C under a nitrogen atmosphere for 15 hours. After completion of the reaction, aqueous ammonia (30 mL) and ethyl acetate (15 mL) were added. The aqueous phase was extracted with ethyl acetate (25 mL x 3). All organic phases were combined, washed with saturated sodium chloride aqueous solution, dried over anhydrous sodium sulfate, and concentrated. The resulting residue was purified by silica gel chromatography using dichloromethane and methanol as eluents to afford compound 3f (130 mg, 55% yield).

[0198] MS (ESI) m / z 614.3 [M+H] +

[0199] 1 H NMR (400MHz, CDCl3) δ=8.47 (d, J=4.4Hz, 1H), 7.68 (d, J=5.6Hz, 1H), 7.64 (d, J=7.2Hz, 1H), 7.48 (dd, J=1. 2, 9.2Hz, 2H), 7.17 (dd, J=4.8, 7.6Hz, 1H), 5.73 (d, J=5.6Hz, 1H), 5.28 (s, 1H), 5.03 (s, 1H), 4.64 (d, J=10 .0Hz, 1H), 4.05-3.93 (m, 2H), 3.73 (d, J=10.0Hz, 1H), 3.33-3.16 (m, 3H), 2.94 (d, J=16.4Hz, 1H), 2.54 (s, 3H), 2.52-2.44(m, 1H), 2.13(dt, J=4.0, 12.4Hz, 1H), 1.82-1.74(m, 1H), 1.52-1.44(m, 1H), 1.30(s, 9H).

[0200] Step 5

[0201] (S)-1′-(8-((3-chloro-2-((methyl-d3)amino)pyridin-4-yl)thio)-7-methylimidazo[1,2-c]pyrimidin-5-yl)-5,7-dihydrospiro[cyclopenta[b]pyridine-6,4′-piperidin]-5-amine 3

[0202] Compound 3f (130 mg, 0.21 mmol) was dissolved in dry dichloromethane (4.5 mL), and a 1,4-dioxane solution of hydrogen chloride (1.5 mL, 4N) was added dropwise at 0°C, and the mixture was reacted at 20°C for 1 hour. A 0.1 M aqueous sodium hydroxide solution (30 mL) was added to the reaction solution to adjust the pH to 14, and then extracted with dichloromethane (30 mL×2). The organic phases were combined, dried over anhydrous sodium sulfate, filtered, and the filtrate was concentrated under reduced pressure to obtain a crude product. The crude product was purified by reverse phase chromatography with 0.1% ammonia water and acetonitrile to obtain compound 3 (65 mg, yield: 41%). X-ray powder diffraction analysis showed that the solid was amorphous, and the XRPD spectrum was as shown below. Figure 1 shown.

[0203] MS (ESI) m / z 510.2 [M+H] +

[0204] 1 H NMR (400MHz, MeOD_d4) δ=8.36 (d, J=4.4Hz, 1H), 7.86 (d, J=7.6Hz, 1H), 7.76 (d, J=1.6Hz, 1H), 7.57 (d, J=5 .6Hz, 1H), 7.47 (d, J=1.6Hz, 1H), 7.29 (dd, J=4.8, 7.6Hz, 1H), 5.75 (d, J=6.0Hz, 1H), 4.11 (s, 1H), 4.05 (br d, J=13.6Hz, 2H), 3.45-3.35 (m, 2H), 3.27 (d, J=16.8Hz, 1H), 2.97 (d, J=16. 4Hz, 1H), 2.55 (s, 3H), 2.17-2.03 (m, 2H), 1.74 (brd, J=14.0Hz, 1H), 1.53 (br d, J=13.6Hz, 1H).

[0205] Example 4

[0206] (S)-1′-(8-((2-(di(methyl-d3)amino)-3-chloropyridin-4-yl)thio)-7-methylimidazo[1,2-c]pyrimidin-5-yl)-5,7-dihydrospiro[cyclopenta[b]pyridine-6,4′-piperidin]-5-amine

[0207]

[0208] Intermediate 4a

[0209] Sodium 2-(di(methyl-d3)amino)-3-chloropyridine-4-thiolate

[0210]

[0211] The synthesis steps of intermediate 4a refer to intermediate 3e, wherein dimethyl-d6-amine hydrochloride is used to replace methyl-d3-amine hydrochloride to prepare the aforementioned intermediate 4a.

[0212] MS (ESI) m / z 195.1 [M+H] +

[0213] 1 H NMR (400 MHz, DMSO_d6) δ = 7.24 (d, J = 5.2 Hz, 1H), 6.79 (d, J = 5.2 Hz, 1H).

[0214] The synthesis steps of compound 4 refer to Example 3, wherein intermediate 4a is used to replace intermediate 3e to prepare compound 4.

[0215] MS (ESI) m / z 527.2 [M+H] +

[0216] 1 H NMR (400MHz, MeOD_d4) δ=8.36 (d, J=4.4Hz, 1H), 7.86 (d, J=7.6Hz, 1H), 7.77 (d, J=1.6Hz, 1H), 7.72 (d, J=5 .2Hz, 1H), 7.47 (d, J=1.6Hz, 1H), 7.29 (dd, J=5.2, 7.6Hz, 1H), 6.06 (d, J=5.6Hz, 1H), 4.11 (s, 1H), 4.06 (br d, J=13.6Hz, 2H), 3.44-3.37 (m, 2H), 3.25 (s, 1H), 2.97 (d, J=16.8Hz, 1H), 2.56 (s, 3H), 2.14-2.03 (m, 2H), 1.75 (br d, J=13.2Hz, 1H), 1.54 (brd, J=13.6Hz, 1H).

[0217] Example 5

[0218] (S)-1′-(8-((3-chloro-2-((methyl-d2)amino)pyridin-4-yl)thio)-7-methylimidazo[1,2-c]pyrimidin-5-yl)-5,7-dihydrospiro[cyclopenta[b]pyridine-6,4′-piperidin]-5-amine

[0219]

[0220] Intermediate 5f

[0221]

[0222] first step

[0223] N,N-bis(4-methoxybenzyl)methylamine-d25b

[0224] Compound 5a (4.0 g, 16 mmol) was dissolved in methanol (50 mL), and a deuterated aqueous solution of deuterated formaldehyde (3.7 g, 23 mmol, 20% w / w) and acetic acid (0.93 g, 16 mmol) were added at room temperature. Sodium cyanoborohydride (2.9 g, 47 mmol) was then added and the mixture was allowed to react at room temperature for 15 hours. After completion of the reaction, the reaction solution was concentrated, 2 M sodium hydroxide solution was added to adjust the pH to 9-10, and the mixture was extracted with ethyl acetate (30 mL x 3). The organic phases were combined, dried over anhydrous sodium sulfate, and concentrated under reduced pressure. The resulting residue was purified by silica gel chromatography using petroleum ether and ethyl acetate as eluents to afford compound 5b (4.0 g, yield: 95%).

[0225] MS (ESI) m / z 274.3 [M+H] +

[0226] 1 H NMR.: (400MHz, CDCl3) δ = 7.28-7.25 (m, 4H), 6.89-6.84 (m, 4H), 3.88-3.74 (m, 7H), 3.45 (s, 4H).

[0227] Step 2

[0228] N-(4-Methoxybenzyl)methane-d2-amine hydrochloride 5c

[0229] Compound 5b (1.0 g, 3.7 mmol) was dissolved in methanol (20 mL), and 10% palladium on carbon (1% water content, 100 mg), 20% palladium hydroxide (100 mg), and concentrated hydrochloric acid (0.5 mL) were added. The mixture was reacted at 80°C under a 50 psi hydrogen atmosphere for 12 hours. The mixture was filtered, washed with methanol (30 mL x 3), and the filtrate was concentrated under reduced pressure to give compound 5c (0.69 g, 99% yield).

[0230] MS (ESI) m / z 153.8 [M+H] +

[0231] 1 H NMR (400MHz, DMSO-d6) δ=9.18 (br s, 2H), 7.45 (d, J=8.4Hz, 2H), 6.97 (d, J=8.4Hz, 2H), 4.01 (t, J=5.6Hz, 2H), 3.76 (s, 3H), 2.44 (brs, 1H).

[0232] Step 3

[0233] 3-Chloro-4-iodo-N-(4-methoxybenzyl)-N-(methyl-d2)pyridin-2-amine 5d

[0234] Compound 5c (638 mg, 3.4 mmol) and compound 3a (787 mg, 3.1 mmol) were dissolved in DMSO (10 mL) and DIEA (2.0 g, 15 mmol) was added. The mixture was reacted at 60°C for 15 hours. After completion of the reaction, an ice-water mixture (100 mL) was added and the mixture was extracted with ethyl acetate (30 mL x 3). The organic phases were combined, washed with saturated brine, dried over anhydrous sodium sulfate, and concentrated under reduced pressure. The resulting residue was purified by silica gel chromatography using petroleum ether and ethyl acetate as eluents to afford compound 5d (590 mg, 49% yield).

[0235] MS (ESI) m / z 391.0 [M+H] +

[0236] 1 H NMR: (400MHz, CDCl3) δ=7.76 (d, J=5.2Hz, 1H), 7.35 (d, J=5.2Hz, 1H), 7.32-7.27 (m, 2H), 6.95-6.81 (m, 2H), 4.45 (s, 2H), 3.82 (s, 3H), 2.80 (s, 1H).

[0237] Step 4

[0238] Ethyl 3-((3-chloro-2-((4-methoxybenzyl)(methyl-d2)amino)pyridin-4-yl)thio)propanoate 5e

[0239] Compound 5d (590 mg, 1.5 mmol) was dissolved in 1,4-dioxane (8 mL), and ethyl 3-mercaptopropionate 3c (304 mg, 2.3 mmol), tris(dibenzylideneacetone)dipalladium (69 mg, 0.076 mmol), 4,5-bis(diphenylphosphino-9,9-dimethylxanthene) (87 mg, 0.15 mmol), and N,N-diisopropylethylamine (586 mg, 4.5 mmol) were added. The mixture was reacted at 100°C under a nitrogen atmosphere for 5 hours. After completion of the reaction, the product was filtered, and the filtrate was concentrated under reduced pressure. The resulting residue was purified by silica gel chromatography using ethyl acetate and petroleum ether as eluents to afford compound 5e (614 mg, 94% yield).

[0240] MS (ESI) m / z 397.1 [M+H] +

[0241] 1H NMR (400MHz, CDCl3) δ=8.07 (d, J=5.6Hz, 1H), 7.34-7.29 (m, 2H), 6.93-6.83 (m, 2H), 6.71 (d, J=5.2Hz, 1H), 4.43 (s, 2H) , 4.20 (q, J=7.2Hz, 2H), 3.81 (s, 3H), 3.24 (t, J=7.6Hz, 2H), 2.80 (s, 1H), 2.75 (t, J=7.6Hz, 2H), 1.29 (t, J=7.2Hz, 3H).

[0242] Step 5

[0243] Sodium 3-chloro-2-((4-methoxybenzyl)(methyl-d2)amino)pyridine-4-thiolate intermediate 5f

[0244] Compound 5e (614 mg, 1.4 mmol) was dissolved in tetrahydrofuran (8 mL), and sodium ethoxide ethanol solution (582 mg, 1.7 mmol, 20% w / w) was added at 0°C. The mixture was reacted at 0°C for 1 hour. After completion of the reaction, the reaction solution was concentrated, and a mixed solution of methyl tert-butyl ether and dichloromethane (6 mL, v / v = 50 / 2) was added. The mixture was filtered and washed with methyl tert-butyl ether (10 mL x 3). The resulting solid was dried under reduced pressure to give intermediate 5f (445 mg, yield: 98%).

[0245] MS (ESI) m / z 297.1 [M+H] +

[0246] 1 H NMR (400MHz, Methanol_d4) δ=7.46 (d, J=5.4Hz, 1H), 7.32-7.22 (m, 2H), 7.17 (d, J=5.6Hz, 1H), 6.92-6.76 (m, 2H), 4.25 (s, 2H), 3.77 (s, 3H), 2.61 (s, 1H).

[0247] Step 6

[0248] (S)-N-((S)-1′-(8-((3-chloro-2-((4-methoxybenzyl)(methyl-d2)amino)pyridin-4-yl)thio)-7-methylimidazo[1,2-c]pyrimidin-5-yl)-5,7-dihydrospiro[cyclopenta[b]pyridine-6,4′-piperidin]-5-yl)-2-methylpropane-2-sulfenamide 5g

[0249] Under a nitrogen atmosphere, compound 2b (153 mg, 0.30 mmol) and intermediate 5f (188 mg, 0.59 mmol) were dissolved in 1,4-dioxane (5 mL). Cuprous iodide (56 mg, 0.30 mmol), N,N-dimethylethylenediamine (52 mg, 0.59 mmol), and potassium phosphate (188 mg, 0.89 mmol) were added. The mixture was heated at 130°C under a nitrogen atmosphere for 15 hours. After completion of the reaction, aqueous ammonia (20 mL) and ethyl acetate (10 mL) were added. The aqueous phase was extracted with ethyl acetate (20 mL x 3). All organic phases were combined, washed with saturated sodium chloride aqueous solution, dried over anhydrous sodium sulfate, and concentrated. The resulting residue was purified by silica gel chromatography using dichloromethane and methanol as eluents to afford compound 5g (190 mg, 45% yield).

[0250] MS (ESI) m / z 733.3 [M+H] +

[0251] Step 7

[0252] (S)-1′-(8-((3-chloro-2-((methyl-d2)amino)pyridin-4-yl)thio)-7-methylimidazo[1,2-c]pyrimidin-5-yl)-5,7-dihydrospiro[cyclopenta[b]pyridine-6,4′-piperidin]-5-amine 5

[0253] Compound 5g (160 mg, 0.22 mmol) was dissolved in TFA (3 mL) and allowed to react at room temperature for 4 hours. A solution of hydrogen chloride in 1,4-dioxane (1 mL, 4 N) was then added dropwise at 0°C and allowed to react at room temperature for 1 hour. 0.1 M aqueous sodium hydroxide solution (30 mL) was added to the reaction solution to adjust the pH to 14, and then extracted with dichloromethane (30 mL x 2). The organic phases were combined, dried over anhydrous sodium sulfate, filtered, and the filtrate was concentrated under reduced pressure to obtain a crude product. The crude product was purified by silica gel chromatography using dichloromethane and methanol as eluents to obtain compound 5 (51 mg, yield: 46%).

[0254] MS (ESI) m / z 509.2 [M+H] +

[0255] 1H NMR (400MHz, CDCl3) δ=8.45 (d, J=4.4Hz, 1H), 7.71 (d, J=5.2Hz, 1H), 7.67 (d, J=7.2Hz, 1H), 7.56 (d, J=1.2Hz, 1 H), 7.44 (d, J=1.6Hz, 1H), 7.18 (dd, J=5.2, 7.6Hz, 1H), 5.78 (d, J=5.6Hz, 1H), 5.03 (d, J=4.4Hz, 1H), 4.11 (s, 1 H), 4.04-3.93 (m, 2H), 3.33 (q, J=11.6Hz, 2H), 3.25 (d, J=16.8Hz, 1H), 3.00 (brs, 1H), 2.93 (d, J=16.8Hz, 1H), 2.58 (s, 3H), 2.11 (dt, J=4.0, 12.8Hz, 1H), 2.02 (dt, J=4.4, 12.4Hz, 1H), 1.81-1.73 (m, 1H), 1.52-1.47 (m, 1H).

[0256] Example 6

[0257] (S)-1′-(8-((3-chloro-2-((methyl- d )amino)pyridin-4-yl)thio)-7-methylimidazo[1,2-c]pyrimidin-5-yl)-5,7-dihydrospiro[cyclopenta[b]pyridine-6,4′-piperidin]-5-amine

[0258]

[0259] Intermediate 6f

[0260] N,N-bis(4-methoxybenzyl)methylamine-d6a

[0261] Compound 5a (3.0 g, 12 mmol) was dissolved in anhydrous methanol (30 mL), and aqueous formaldehyde (2.6 mL, 37% w / w) and acetic acid (0.67 mL, 12 mmol) were added at room temperature. The reaction was allowed to proceed at room temperature for 2 hours, followed by the addition of sodium borodeuteride (0.97 g, 23 mmol) at 0°C and an additional 1.5 hours at room temperature. After completion of the reaction, the reaction solution was concentrated, water (50 mL) was added, and extraction with ethyl acetate (30 mL x 3) was performed. The organic phases were combined, dried over anhydrous sodium sulfate, and concentrated under reduced pressure to afford compound 6a (3.3 g, 94% yield).

[0262] 1 H NMR: (400MHz, CDCl3) δ=7.30-7.25(m, 4H), 6.91-6.84(m, 4H), 3.81(s, 6H), 3.45(s, 4H), 2.13(s, 2H).

[0263] Step 2

[0264] Benzyl (4-methoxybenzyl)(methyl-d)carbamate 6b

[0265] Compound 6a (3.3 g, 12 mmol) was dissolved in toluene (30 mL), and benzyl chloroformate (4.1 mL, 29 mmol) was added. The mixture was reacted at 120°C under a nitrogen atmosphere for 14 hours. After completion, the reaction solution was concentrated under reduced pressure, and then washed with ethyl acetate (50 mL), water (20 mL), and saturated brine (20 mL x 2). The organic phase was dried over anhydrous sodium sulfate and concentrated under reduced pressure. The residue was purified by silica gel chromatography using petroleum ether and ethyl acetate as eluents to afford compound 6b (3.8 g crude product).

[0266] 1 H NMR (400MHz, CDCl3) δ=7.43-7.31 (m, 5H), 7.21 (d, J=7.6Hz, 1H), 7.13 (d, J=7.6Hz, 1H ), 6.91-6.82 (m, 2H), 5.20 (s, 2H), 4.45 (s, 2H), 3.82 (s, 3H), 2.86 (d, J=10.8Hz, 2H).

[0267] Step 3

[0268] N-(4-Methoxybenzyl)methane-d-amine hydrochloride 6c

[0269] Compound 6b (3.8 g, 12 mmol) was dissolved in methanol (40 mL), and 10% palladium on carbon (1.0 g) was added. The mixture was reacted at 40°C under a hydrogen atmosphere for 16 hours. The mixture was filtered, washed with methanol (80 mL), and the filtrate was concentrated under reduced pressure to give compound 6c (2 g, yield: 99%).

[0270] MS (ESI) m / z 152.9 [M+H] +

[0271] Step 4

[0272] 3-Chloro-4-iodo-N-(4-methoxybenzyl)-N-(methyl-d)pyridin-2-amine 6d

[0273] Compound 6c (2 g, 12 mmol) and compound 3a (2.3 g, 8.8 mmol) were dissolved in DMSO (4 mL) and DIEA (4.3 mL, 26 mmol) was added. The mixture was reacted at 60°C for 5 hours. After completion of the reaction, water (50 mL) was added and the mixture was extracted with ethyl acetate (30 mL x 3). The organic phases were combined, washed with saturated brine, dried over anhydrous sodium sulfate, and concentrated under reduced pressure. The resulting residue was purified by silica gel chromatography using petroleum ether and ethyl acetate as eluents to afford compound 6d (2.6 g, 76% yield).

[0274] MS (ESI) m / z 390.0 [M+H] +

[0275] 1 H NMR: (400MHz, CDCl3) δ=7.76 (d, J=4.8Hz, 1H), 7.35 (d, J=5.2Hz, 1H), 7.32-7.27 (m, 2H), 6.92-6.83 (m, 2H), 4.45 (s, 2H), 3.81 (s, 3H), 2.82 (s, 2H).

[0276] Step 5

[0277] Ethyl 3-((3-chloro-2-((4-methoxybenzyl)(methyl-d)amino)pyridin-4-yl)thio)propanoate 6e

[0278] Compound 6d (2.6 g, 6.7 mmol) was dissolved in 1,4-dioxane (30 mL), and ethyl 3-mercaptopropionate 3c (1.3 g, 10 mmol), tris(dibenzylideneacetone)dipalladium (310 mg, 0.34 mmol), 4,5-bis(diphenylphosphino)-9,9-dimethylxanthene (390 mg, 0.67 mmol), and N,N-diisopropylethylamine (3.3 mL, 20 mmol) were added. The reaction was allowed to proceed at 100°C under a nitrogen atmosphere for 6 hours. After completion of the reaction, the product was filtered, and the filtrate was concentrated under reduced pressure. The resulting residue was purified by silica gel chromatography using ethyl acetate and petroleum ether as eluents to afford compound 6e (2.3 g, 88% yield).

[0279] MS (ESI) m / z 396.1 [M+H] +

[0280] 1H NMR (400MHz, CDCl3) δ=8.07 (d, J=5.2Hz, 1H), 7.34-7.28 (m, 2H), 6.93-6.82 (m, 2H), 6.72 (d, J=5.2Hz, 1H), 4.43 (s, 2H) , 4.20 (q, J=7.2Hz, 2H), 3.81 (s, 3H), 3.24 (t, J=7.6Hz, 2H), 2.81 (s, 2H), 2.75 (t, J=7.6Hz, 2H), 1.29 (t, J=6.8Hz, 3H).

[0281] Step 6

[0282] Sodium 3-chloro-2-((4-methoxybenzyl)(methyl-d)amino)pyridine-4-thiolate intermediate 6f

[0283] Compound 6e (2.3 g, 5.9 mmol) was dissolved in tetrahydrofuran (25 mL), and sodium ethoxide ethanol solution (2.4 g, 7.1 mmol, 20% w / w) was added at 0°C. The mixture was allowed to react at 0°C for 1 hour. After completion of the reaction, the reaction solution was concentrated, and a mixed solution of methyl tert-butyl ether and dichloromethane (40 mL, v / v = 50 / 2) was added. The mixture was filtered and washed with methyl tert-butyl ether (15 mL x 3). The resulting solid was dried under reduced pressure to give intermediate 6f (1.7 g, yield: 91%).

[0284] 1 H NMR (400MHz, DMSO-d6) δ = 7.29-7.23 (m, 3H), 6.89-6.79 (m, 3H), 4.12 (s, 2H), 3.72 (s, 3H), 3.34 (s, 2H).

[0285] The synthesis steps of compound 6 refer to Example 5, wherein intermediate 6f is used to replace intermediate 5f to prepare the aforementioned compound 6.

[0286] MS (ESI) m / z 508.2 [M+H] +

[0287] 1H NMR (400MHz, CDCl3) δ=8.47 (d, J=4.4Hz, 1H), 7.68 (d, J=5.6Hz, 1H), 7.64 (d, J=7.2Hz, 1H), 7.48 (dd, J=1. 2, 9.2Hz, 2H), 7.17 (dd, J=4.8, 7.6Hz, 1H), 5.73 (d, J=5.6Hz, 1H), 5.28 (s, 1H), 5.03 (s, 1H), 4.64 (d, J=10 .0Hz, 1H), 4.05-3.93 (m, 2H), 3.73 (d, J=10.0Hz, 1H), 3.33-3.16 (m, 3H), 2.94 (d, J=16.4Hz, 1H), 2.54 (s, 3H), 2.52-2.44(m, 1H), 2.13(dt, J=4.0, 12.4Hz, 1H), 1.82-1.74(m, 1H), 1.52-1.44(m, 1H), 1.30(s, 9H).

[0288] Biological evaluation

[0289] The present disclosure is further described and explained below in conjunction with test examples, but these embodiments are not intended to limit the scope of the present disclosure.

[0290] Test Example 1: Detection of the activity of the disclosed compounds on SHP2 phosphatase

[0291] 1. Experimental materials and instruments

[0292] Instrument name Equipment Manufacturer model Constant temperature oven oscillator IMB MB-1002A Microplate Reader MD SpectraMax M5

[0293]

[0294]

[0295] 2. Experimental steps

[0296] 0.2 nM recombinantly expressed full-length SHP2 (aa 1-593), 0.5 nM activating peptide IRS1 with dual phosphorylation sites (sequence: H2N-LN(pY)IDLDLY(dPEG8)LST(pY)ASINFQK-amide) and a series of test compounds (final concentrations of 1□M, 0.3□M, 0.1□M, 0.03□M, 0.01□M, 0.003□M, 0.001□M, 0.0003□M, 0.0001□M, 0.00003□M) were added to phosphatase reaction solution (60 mM HEPES, pH 7.5 0.005% Brij-35, 75 mM NaCl, 75 mM KCl, 1 mM EDTA, 5 mM DTT) and shaken at room temperature (350 rpm) for 30 minutes. Then, DiFMUP was added to a final concentration of 30 M and reacted at room temperature for 30 minutes. The phosphatase reaction was terminated with 5 μL of a reaction stop solution (60 mM HEPES, pH 7.5, 0.2% SDS). The fluorescence values at Ex358 nm and Em455 were read on a fluorescence plate reader MD SpectraMax.

[0297] IC of the compound 50 The values were calculated using the four-parameter logit method. In the following formula, x represents the logarithm of the compound concentration; F(x) represents the effect size (the inhibition rate of cell proliferation at that concentration): F(x) = ((AD) / (1+((x / C)^B)))+D. A, B, C, and D are the four parameters. Different concentrations correspond to different inhibition rates of phosphatase activity. An inverse curve is drawn, and the IC value of the inhibitor is calculated from the curve. 50 The IC values of the compounds were calculated using Primer premier 6.0. 50 .

[0298] The in vitro activity of the disclosed compounds on SHP2 was determined by the above test, and the orally active SHP2 inhibitor SHP099 was selected as a positive drug. The structure of the compound is disclosed in the literature J.Med.Chem.2016, 59, 7773-7782. The specific compound was purchased from Shanghai Haoyuan Biopharmaceutical Technology Co., Ltd. (Medchemexpress.cn).

[0299] Measured IC 50 See Table 1 for values.

[0300] Table 1. IC values of compounds for SHP2 phosphatase 50

[0301] Example No. <![CDATA[IC 50 (nM)]]> Example No. <![CDATA[IC 50 (nM)]]> SHP099 79 1 1.7 2 2.1 3 4.5 5 4.8 6 4.7

[0302] Test Example 2: In vitro metabolic stability experiment of rat liver microsomes

[0303] The compound concentrations in the reaction system were determined by LC / MS / MS to calculate the intrinsic clearance of the test compounds and to evaluate their in vitro metabolic stability in rat liver microsomes.

[0304] Add 222.5 μL of a 1.1236 mg / mL mixture of rat liver microsomes (male Wistar Han strain, purchased from Corning, Cat. No. 452511) and 25 μL of 10 mM NADPH to the incubation plate. Vortex to mix thoroughly for 10 seconds. Incubate in a 37°C water bath for 8 minutes. Initiate the reaction by adding 2.5 μL of 100 μM test compound or positive control to the incubation plate. Vortex to mix thoroughly for 12 seconds and continue incubation in a 37°C water bath. Terminate the reaction at 0.5, 5, 10, 15, 20, and 30 minutes by transferring 20 μL of the incubation to a stop plate containing 100 μL of cold stop solution and vortex to mix thoroughly for 2 minutes. Centrifuge the stop plate at 4000 rpm for 20 minutes, then let it rest at 4°C for 30 minutes, and then centrifuge it again at 4000 rpm for 20 minutes. Transfer 40 μL of each compound supernatant to a 96-well plate and add 160 μL of purified water to dilute the sample.

[0305] The samples were quantified by ion chromatograms, and the residual rate was calculated based on the peak area of the test compound or positive control. The slope k was determined by linear regression of the natural logarithm of the residual rate against the incubation time using Microsoft Excel.

[0306] The in vitro half-life (in vitro t1 / 2) was calculated from the slope: in vitro t1 / 2 = -(0.693 / k)

[0307] The in vitro half-life was converted to intrinsic clearance (CLint, μL / min / mg protein) using the following equation:

[0308] in vitro CLint = (0.693 / t1 / 2) × (incubation volume (μL) / protein amount (mg))

[0309] The measured rat liver microsomal intrinsic clearance values are shown in Table 2.

[0310] Table 2. Rat liver microsomal intrinsic clearance

[0311]

[0312] Test Example 3: Pharmacokinetics experiment in rats

[0313] The drug concentrations in the plasma of rats at different times after oral administration of the compound of the present invention were determined by LC / MS / MS, using rats as test animals to study the pharmacokinetic behavior of the compound of the present invention in rats and evaluate its pharmacokinetic characteristics.

[0314] Experimental animals: 3 healthy 6-8 week old male SD rats per group

[0315] Drug preparation

[0316] A certain amount of drug was weighed and added with 0.5% by mass of hydropropyl methylcellulose, 0.1% by volume of Tween 80 and 99.4% by volume of water to prepare a 1 mg / mL white suspension.

[0317] Drug administration

[0318] The SD rats were fasted overnight and then gavage-administered. The dosage of the reference substance was 7.5 mg / kg, and the dosage of Example 1 was 5 mg / kg.

[0319] operate

[0320] Rats were orally administered with the compound of the present invention. 0.2 mL of blood was collected from the jugular vein at 0.25, 0.5, 1, 2, 4, 8, and 24 hours after administration. The blood was placed in a tube containing EDTA-K2 and centrifuged at 4000 rpm for 5 minutes at 4°C to separate the plasma, which was then stored at -75°C.

[0321] Determination of the test compound content in rat plasma after oral administration of different drug concentrations: 50 μL of rat plasma at each time point after administration was taken, 200 μL of acetonitrile solution of internal standard dexamethasone (50 ng / mL) was added, vortexed for 30 seconds, and centrifuged at 4°C and 4700 rpm for 15 minutes. The supernatant of the plasma sample was diluted three times with water, and 2.0 μL was taken for LC / MS / MS analysis.

[0322] Pharmacokinetic parameter results

[0323] The pharmacokinetic parameters of the compounds of the present invention in rats are shown in Table 3 below.

[0324] Table 3. Pharmacokinetic parameters of the compounds in rats

[0325]

[0326] Test Example 4: Pharmacokinetic Experiment in Cynomolgus Monkeys

[0327] Cynomolgus monkeys were used as test animals, and the plasma drug concentrations at different time points after oral administration of the compound of the present invention were determined by LC / MS / MS to study the pharmacokinetic behavior of the compound of the present invention in cynomolgus monkeys and evaluate its pharmacokinetic characteristics.

[0328] Experimental animals: 3 healthy male cynomolgus monkeys aged 2-5 years per group;

[0329] Drug preparation

[0330] Oral administration: Weigh a certain amount of drug and add 0.5% by mass of hydropropyl methylcellulose, 0.1% by volume of Tween 80 and 99.4% by volume of water to prepare a 1 mg / mL white suspension.

[0331] Drug administration

[0332] The drug was administered orally to cynomolgus monkeys after overnight fasting at a dose of 5 mg / kg.

[0333] operate

[0334] Cynomolgus monkeys were orally administered with the compound of the present invention. 0.2 mL of blood was collected from a peripheral vein at 0.25, 0.5, 1, 2, 4, 8, and 24 hours after administration. The blood was placed in a tube containing EDTA-K2 and centrifuged at 2000 rpm for 10 minutes at 2-8°C to separate the plasma, which was then stored at -75°C.

[0335] Determination of the content of the test compound in the plasma of cynomolgus monkeys after oral administration of different drug concentrations: 55 μL of cynomolgus monkey plasma at each time point after administration was taken, 200 μL of acetonitrile solution of internal standard verapamil or dexamethasone was added, vortex mixed for 30 seconds, centrifuged at 3900 rpm at 4°C for 15 minutes, and the supernatant of the plasma sample was diluted three times with water, and 15 μL was taken for LC / MS / MS analysis.

[0336] Pharmacokinetic parameter results

[0337] The pharmacokinetic parameters of the compounds of the present invention in cynomolgus monkeys are shown in Table 4 below.

[0338] Table 4. Pharmacokinetic parameters in cynomolgus monkeys

[0339]

[0340] Test Example 5: Caco-2 permeability test

[0341] The apparent permeability coefficient (P) of the drug was determined by liquid chromatography tandem mass spectrometry (LC / MS / MS) using a Caco-2 cell model. app ).

[0342] The Caco-2 cells (purchased from ATCC) were grown at a density of 7.92 × 10 5 cells / cm 2210 μL of HBSS (25 mM HEPES, pH 7.4, containing 50 μM quinidine, 30 μM benzbromarone, and 20 μM sulfasalazine) containing 10 μM of the test compound was added to the apical portion of a Transwell chamber (purchased from Corning). Immediately, 10 μL of the sample was removed and added to a 96-deep-well plate containing 90 μL of HBSS (25 mM HEPES, pH 7.4, containing 50 μM quinidine, 30 μM benzbromarone, and 20 μM sulfasalazine) as the initial drug-addition sample. 800 μL of HBSS (25 mM HEPES, pH 7.4, containing 50 μM quinidine, 30 μM benzbromarone, and 20 μM sulfasalazine) was added to the basolateral portion. The mixture was incubated at 37°C for 2 hours. At the 45-minute and 2-hour time points, 10 μL of sample was pipetted from the apical side into 90 μL of HBSS (25 mM HEPES, pH 7.4, containing 50 μM quinidine, 30 μM benzbromarone, and 20 μM sulfasalazine) in a 96-well deep-well plate. At the 45-minute and 2-hour time points, 100 μL of sample was pipetted from the basolateral side into a 96-well deep-well plate. Three volumes of pre-chilled internal standard were then added to each well. The plates were vortexed at 1000 rpm for 10 minutes and centrifuged at 4000 rpm for 20 minutes. A 100 μL sample was removed from each well, and all three samples were combined with 100 μL of purified water for LC / MS / MS analysis.

[0343] Data were calculated using Microsoft Excel, and peak areas were calculated from the chromatograms. The apparent permeability coefficient (Papp) is in cm / s and is calculated using the following formula:

[0344]

[0345] C R is the concentration of the compound to be tested at the substrate end (the superscript "120" or "45" is the sampling time, unit: minute), C D is the concentration of the compound to be tested at the top (the superscript "120" or "45" is the sampling time, unit: minutes), Area is the membrane surface area (0.33 cm 2 ), time is the total transit time (75 × 60 seconds).

[0346] The measured apparent permeability coefficient values of Caco-2 cells are shown in Table 5.

[0347] Table 5. Apparent permeability coefficients of compounds in Caco-2 cells

[0348] Example No. <![CDATA[Papp (A-B) (10 -6 ,cm / s)]]> 1 2.71 2 12.05 3 17.22

[0349] Test Example 6: CYP inhibition experiment

[0350] Pooled human liver microsomes from 150 donors (Corning, Cat. No. 452117) were used to evaluate the metabolic reactions of representative substrates of the five major human CYP isoforms (CYP1A2, CYP2C9, CYP2C19, CYP2D6, and CYP3A4 / 5). The effects of various concentrations of the test compounds on the metabolic reactions of phenacetin (CYP1A2), diclofenac sodium (CYP2C9), S-mephenytoin (CYP2C19), bufuralol hydrochloride (2D6), and midazolam (CYP3A4 / 5) were determined by liquid chromatography-tandem mass spectrometry (LC / MS / MS).

[0351] A 200 μL reaction mixture (100 mmol / L phosphate buffer, pH 7.4, containing 0.3% DMSO, 0.6% acetonitrile, and 0.1% methanol, by volume) containing 30 μM phenacetin, 10 μM diclofenac sodium, 35 μM S-mephenytoin, 5 μM bufuralol hydrochloride, 3 μM midazolam, 1 mM NADPH, test compounds (at concentrations of 0.1, 0.3, 1, 3, 10, and 30 μmol / L), positive compounds, or a blank control, was incubated with human liver microsomes (0.2 mg / mL) at 37°C for 5 minutes. Then, 200 μL of an acetonitrile solution containing 3% formic acid and 40 nM internal standard verapamil was added, and the mixture was centrifuged at 4000 rpm for 50 minutes. The mixture was cooled on ice for 20 minutes and then centrifuged at 4000 rpm for 20 minutes to precipitate the protein. A 200 μL supernatant was used for LC / MS / MS analysis.

[0352] The peak area was calculated based on the chromatogram. The residual activity ratio (%) was calculated using the following formula:

[0353] Peak area ratio = metabolite peak area / internal standard peak area

[0354] Residual activity ratio (%) = peak area ratio of the test compound group / peak area ratio of the blank group

[0355] CYP half-maximal inhibitory concentration (IC 50 ) was calculated using Excel XLfit 5.3.1.3.

[0356] The measured CYP half-maximal inhibitory concentration (IC 50 ) values are shown in Table 6.

[0357] Table 6. Half maximal inhibitory concentration (IC) of compounds on CYP 50 )

[0358]

[0359] Example 7. Preparation of Crystal Form A of the Compound Represented by Formula (I)

[0360] In the fifth step of Example 1, column chromatography using a methanol-dichloromethane solvent system was performed and rotary evaporation was performed to obtain a solid. The XRPD spectrum of the solid was as follows: Figure 2 The characteristic peak positions are shown in Table 7, which is defined as Form A; TGA spectrum ( Figure 3 ) shows that the weight loss of crystal form A is 1.33% in the range of 25-260℃, and the DSC spectrum ( Figure 4 ) shows that crystal form A has an endothermic peak with a peak value of 241.49 ° C; the X-ray powder diffraction comparison diagram before and after DVS shows that the crystal form does not change before and after DVS, see Figure 5 .

[0361] Table 7. Characteristic peaks of Form A

[0362]

[0363]

[0364] Example 8. Preparation of Crystal Form A of the Compound Represented by Formula (I)

[0365] The preparation process of Form A includes the steps of solid-liquid separation, and the crystal form is determined by X-ray powder diffraction detection.

[0366] Table 8. Preparation of Form A of the compound represented by formula (I)

[0367]

[0368]

[0369] Example 9. Preparation of Crystal Form B of the Compound Represented by Formula (I)

[0370] Weigh 50 mg of the crystalline form of compound A represented by formula (I), add 1 ml of methanol to dissolve, filter, add 5 ml of methyl tert-butyl ether, stir overnight at room temperature, filter, wash with methyl tert-butyl ether, and dry to obtain a solid. X-ray powder diffraction analysis shows that the XRPD spectrum is as follows: Figure 6 The positions of its characteristic peaks are shown in Table 9, and it is defined as Type B.

[0371] Table 9. Characteristic peaks of Form B

[0372]

[0373] Example 10. Preparation of Crystal Form C of the Compound Represented by Formula (I)

[0374] Weigh 10 mg of Form A of the compound represented by formula (I), add 0.1 ml of N-methylpyrrolidone to dissolve, add 1.0 ml of isopropanol, stir overnight at room temperature, filter to obtain a solid, and detect by X-ray powder diffraction. The XRPD spectrum is as follows: Figure 7 The positions of its characteristic peaks are shown in Table 10, which is defined as Form C; the TGA spectrum shows that Form C loses 8.83% of its weight between 25-220°C; the DSC spectrum shows that Form C has two endothermic peaks, with peak values at 197.87°C and 241.33°C, respectively.

[0375] Table 10. Characteristic peaks of crystal form C

[0376]

[0377] Example 11. Preparation of Crystal Form D of the Compound Represented by Formula (I)

[0378] Weigh 4 mg of Form A of the compound represented by formula (I), add 0.2 ml of methanol to dissolve, add 1.0 ml of isopropyl ether, stir overnight at room temperature, filter, and dry to obtain a solid. X-ray powder diffraction analysis shows that the XRPD spectrum is as follows: Figure 8 The positions of its characteristic peaks are shown in Table 11, which is defined as crystal form D; the TGA spectrum shows that crystal form D loses 2.63% of its weight between 25-215°C; the DSC spectrum shows that crystal form D has two endothermic peaks, with peak values at 195.92°C and 243.03°C, respectively.

[0379] Table 11. Characteristic peaks of Form D

[0380]

[0381] Example 12: Preparation of Crystal Form E of the Compound Represented by Formula (I)

[0382] Weigh 10 mg of Form A of the compound represented by formula (I), add 0.3 ml of dichloromethane to dissolve, add 1.0 ml of acetonitrile, stir overnight at 50°C, filter, and dry at 40°C to obtain a solid. X-ray powder diffraction analysis shows that the XRPD spectrum is as follows: Figure 9 As shown, the positions of its characteristic peaks are shown in Table 12, and it is defined as E crystal form; the TGA spectrum shows that E crystal form loses 0.33% in weight at 25-65°C and loses 6.95% in weight at 65-220°C; the DSC spectrum shows that E crystal form has two endothermic peaks, with peak values at 193.32°C and 243.68°C, respectively.

[0383] Table 12. Characteristic peaks of Form E

[0384]

[0385] Example 13. Preparation of Form F of the Compound Represented by Formula (I)

[0386] Weigh 100 mg of the E crystal form of the compound represented by formula (I), add 5 ml of 80% water-ethanol, stir overnight at room temperature, filter, wash with water, and dry at 40°C for 2 hours to obtain a solid. The XRPD spectrum of the solid is shown in the following figure: Figure 10 The positions of its characteristic peaks are shown in Table 13, which is defined as F crystal form; the TGA spectrum shows that the F crystal form loses 3.02% in weight between 25-70°C and loses 0.47% in weight between 70-260°C; the DSC spectrum shows that the F crystal form has two endothermic peaks, with peak values of 80.36°C and 243.71°C, respectively, and one exothermic peak, with a peak value of 188.21°C.

[0387] Table 13. Characteristic peaks of Form F

[0388]

[0389]

[0390] Example 14. Preparation of Form G of the Compound Represented by Formula (I)

[0391] Weigh 500 mg of the E crystal form of the compound represented by formula (I), add 12.5 ml of 20% water-ethanol to dissolve, add 37.5 ml of water, stir overnight at room temperature, filter, wash with water, and dry at 40°C for 3 days to obtain a solid. X-ray powder diffraction analysis shows that the XRPD pattern is as follows: Figure 11 The positions of its characteristic peaks are shown in Table 14, which is defined as crystal form G; the TGA spectrum shows that crystal form G loses 0.62% of its weight between 25-220°C; the DSC spectrum shows that crystal form G has one endothermic peak with a peak value of 243.37°C; and one exothermic peak with a peak value of 187.40°C.

[0392] Table 14. Characteristic peaks of G crystal form

[0393]

[0394]

[0395] Example 15. Preparation of Crystal Form H of the Compound Represented by Formula (I)

[0396] Weigh 100 mg of the E crystal form of the compound represented by formula (I), add 5 ml of 80% water-ethanol, stir overnight at room temperature, filter, wash with water, and dry at 30°C for 2 hours to obtain a solid. The XRPD spectrum of the solid is shown in FIG. Figure 12The positions of its characteristic peaks are shown in Table 15, which is defined as H crystal form; the TGA spectrum shows that the H crystal form loses 3.64% of its weight between 25-60°C; the DSC spectrum shows that the H crystal form has three endothermic peaks, with peak values of 50.81°C, 67.63°C, and 243.52°C, and one exothermic peak, with a peak value of 190.16°C.

[0397] Table 15. Characteristic peaks of H crystal form

[0398]

[0399]

[0400] Example 16: Study on the stability of influencing factors

[0401] The four crystal forms, A, F, G and H, were laid out openly and the stability of the samples was investigated under conditions of light (4500 Lux), high temperature (40°C, 60°C) and high humidity (RH75%, RH92.5%). The sampling period was 30 days.

[0402] Table 16. Stability data of factors affecting crystal form A

[0403]

[0404] Table 17. Stability data of factors affecting F crystal form

[0405]

[0406]

[0407] Table 18. Stability data of factors affecting G crystal form

[0408]

[0409] Table 19. Stability data of factors affecting H crystal form

[0410]

[0411]

[0412] Conclusion: The influencing factor experiment shows that crystal form A has good physical and chemical stability under high temperature and high humidity conditions; the chemical and physical stability of crystal forms F, G, and H under high temperature and high humidity conditions is slightly poor.

[0413] Example 17. Long-term / accelerated stability study

[0414] The stability of crystal forms A, F, G and H was investigated under nitrogen atmosphere at 25°C, 60% RH and 40°C, 75% RH, 5°C and -20°C.

[0415] Table 20. Long-term accelerated stability data of Form A

[0416]

[0417]

[0418] Table 21. Long-term accelerated stability data of Form F

[0419]

[0420] Table 22. Long-term accelerated stability data of Form G

[0421]

[0422]

[0423] Table 23. Long-term accelerated stability data of Form H

[0424]

[0425]

[0426] Conclusion: Long-term accelerated experiments show that the physical and chemical properties of crystal forms A and F are stable under long-term accelerated conditions, and crystal forms G and H undergo crystal transformation under accelerated conditions.

[0427] Example 18: Hygroscopicity Study of Crystal Forms A, F, G, and H

[0428] Using Surface Measurement Systems advantage 2, at 25°C and humidity starting from 50%, the humidity range was examined from 0% to 95% in 10% increments. The judgment criteria were that the mass change dM / dT of each gradient was less than 0.002% and TMAX was less than 360 min, and the cycle was repeated twice.

[0429] Table 24. Hygroscopicity data

[0430]

Claims

1. An X-ray powder diffraction pattern of Form A of a compound represented by formula (I) expressed in terms of a diffraction angle 2θ, There are characteristic peaks at 4.847, 9.801, 13.778, 14.770, 15.444, and 26.

077.

2. The crystal form A of the compound represented by formula (I) according to claim 1 has an X-ray powder diffraction pattern expressed in terms of a diffraction angle 2θ, with characteristic peaks at 4.847, 9.801, 13.138, 13.778, 14.770, 15.444, 18.363, 19.856, 21.092, 23.371, 26.077, and 28.

130.

3. The crystal form A of the compound represented by formula (I) according to claim 1 has an X-ray powder diffraction pattern expressed in terms of a diffraction angle 2θ, with characteristic peaks at 4.847, 9.801, 13.138, 13.778, 14.770, 15.444, 18.363, 19.856, 21.092, 22.034, 23.371, 24.460, 26.077, 28.130, 28.970, 31.894, 32.920, 33.916, and 38.

924.

4. The crystal form A of the compound represented by formula (I) according to claim 1, whose X-ray powder diffraction spectrum is shown in Figure 2.

5. A crystal form B of a compound represented by formula (I), The X-ray powder diffraction pattern expressed in diffraction angle 2θ has characteristic peaks at 4.606, 9.110, 11.423, 13.745, 16.006, and 22.

973.

6. A crystal form C of a compound represented by formula (I), The X-ray powder diffraction pattern expressed in a diffraction angle of 2θ has characteristic peaks at 8.905, 12.920, 13.428, 14.074, 18.458, and 22.

519.

7. A crystal form D of a compound represented by formula (I), The X-ray powder diffraction pattern expressed in a diffraction angle of 2θ has characteristic peaks at 4.766, 9.594, 14.089, 14.471, 18.981, 19.609, and 25.

987.

8. A crystalline form E of a compound represented by formula (I), The X-ray powder diffraction pattern expressed in diffraction angle 2θ has characteristic peaks at 4.603, 9.209, 13.920, 15.097, 19.700 and 25.

454.

9. A crystal form F of a compound represented by formula (I), The X-ray powder diffraction pattern expressed in diffraction angle 2θ has characteristic peaks at 4.656, 14.068, 15.183, 18.858, and 23.

235.

10. A crystal form G of a compound represented by formula (I), The X-ray powder diffraction pattern expressed in diffraction angle 2θ has characteristic peaks at 4.869, 9.735, 13.290, 14.713, and 20.

020.

11. A H crystal form of a compound represented by formula (I), The X-ray powder diffraction pattern expressed in a diffraction angle of 2θ degrees has characteristic peaks at 8.608, 12.983, 13.476, 17.716, 20.144, and 23.

371.

12. The crystalline form of the compound represented by formula (I) according to any one of claims 1 to 11, wherein the error range of the 2θ angle is ±0.

20.

13. The crystalline form of the compound of formula (I) according to any one of claims 1 to 11, wherein the compound of formula (I) has an abundance of at least 20% for each deuterium atom (D).

14. The crystalline form of the compound of formula (I) according to claim 12, wherein the compound of formula (I) has an abundance of at least 20% for each deuterium atom (D).

15. The crystalline form of the compound of formula (I) according to any one of claims 1 to 11, wherein the compound of formula (I) has an abundance of at least 50% for each deuterium atom (D).

16. The crystalline form of the compound of formula (I) according to claim 12, wherein the compound of formula (I) has an abundance of at least 50% for each deuterium atom (D).

17. The crystalline form of the compound of formula (I) according to any one of claims 1 to 11, wherein the compound of formula (I) has an abundance of at least 90% for each deuterium atom (D).

18. The crystalline form of the compound of formula (I) according to claim 12, wherein the compound of formula (I) has an abundance of at least 90% for each deuterium atom (D).

19. The crystalline form of the compound of formula (I) according to any one of claims 1 to 11, wherein the compound of formula (I) has an abundance of at least 98% for each deuterium atom (D).

20. The crystalline form of the compound of formula (I) according to claim 12, wherein the compound of formula (I) has an abundance of at least 98% for each deuterium atom (D).

21. A method for preparing the crystal form A of the compound of formula (I) according to any one of claims 1 to 4, which is selected from: a) mixing the compound represented by formula (I) with solvent I, dissolving it, and crystallizing it, wherein the solvent I is selected from At least one of dichloromethane and chloroform; or b) mixing the compound represented by formula (I) with solvent II to crystallize, wherein the solvent II is at least one selected from tetrahydrofuran, ethyl acetate, toluene, acetone, methanol, ethanol, acetonitrile, methyl tert-butyl ether, water, isopropyl ether, butanone, and n-hexane; or c) mixing the compound represented by formula (I) with solvent III to obtain a clear solution, mixing the clear solution with solvent IV to precipitate crystals, wherein the solvent III is selected from at least one of methanol, N,N-dimethylformamide, and N-methylpyrrolidone; and the solvent IV is selected from at least one of acetonitrile, ethyl acetate, isopropyl acetate, methyl tert-butyl ether, and isopropyl ether.

22. A pharmaceutical composition comprising the following ingredients: 1) A crystalline form of the compound of formula (I) according to any one of claims 1 to 11, or a mixture thereof and 2) Optional pharmaceutically acceptable carriers, diluents or excipients.

23. A method for preparing a pharmaceutical composition, comprising: preparing a crystalline form of a compound of formula (I) according to any one of claims 1 to 11, or a mixture thereof; 2) A step of optionally mixing with a pharmaceutically acceptable carrier, diluent or excipient.

24. Use of a crystalline form of the compound of formula (I) according to any one of claims 1 to 11 or a mixture thereof, or a composition according to claim 22, in the preparation of a medicament for treating a tumor or cancer, wherein the tumor or cancer is selected from juvenile myelomonocytic leukemia, neuroblastoma, melanoma, acute myeloid leukemia, breast cancer, esophageal cancer, lung cancer, colon cancer, pancreatic cancer, gastric cancer, liver cancer, and anaplastic large cell lymphoma.

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