A crystalline form of a triazinedione derivative and a preparation method thereof

Abstract

The present disclosure relates to a crystal form of a triazine dione derivative and a preparation method. Specifically, the present disclosure relates to different crystal forms of (S)-6-((1-(2-fluoro-5-methylphenyl)ethyl)amino)-3-(tetrahydro-2H-pyran-4-yl)-1,3,5-triazine-2,4(1H,3H)-dione and a preparation method thereof. The crystal form of (S)-6-((1-(2-fluoro-5-methylphenyl)ethyl)amino)-3-(tetrahydro-2H-pyran-4-yl)-1,3,5-triazine-2,4(1H,3H)-dione provided by the present disclosure has good stability and can be better used for clinical treatment.

Description

Technical Field

[0001] This disclosure pertains to the pharmaceutical field and relates to the crystal form of a triazine dione derivative, specifically, to the crystal form of (S)-6-((1-(2-fluoro-5-methylphenyl)ethyl)amino)-3-(tetrahydro-2H-pyran-4-yl)-1,3,5-triazine-2,4(1H,3H)-dione and its preparation method. Background Technology

[0002] Hypertrophic cardiomyopathy (HCM) is an autosomal dominant hereditary myocardial disease associated with gene mutations. Its global incidence is approximately 0.2%, and it is the leading cause of sudden death in young adults under 35 (Tuohy, CV. et al., Eur J HeartFail, 22, 2020, 228-240). Clinically, it is characterized by asymmetrical thickening of the left ventricular wall, often involving the interventricular septum, resulting in a smaller ventricular cavity, obstructed left ventricular filling, and decreased ventricular diastolic compliance. Based on the presence or absence of left ventricular outflow tract obstruction, it is classified as obstructive or non-obstructive hypertrophic cardiomyopathy. Current clinical treatments for HCM primarily involve beta-blockers and calcium channel blockers to reduce cardiac contractility and alleviate symptoms. However, these treatments only address the symptoms, not the underlying cause. In advanced stages of HCM, heart transplantation is the only option (Ramaraj, R. Cardiol Rev, 16, 2008, 172-180). Therefore, finding a treatment that targets the root cause of HCM is urgently needed.

[0003] Current research has found that 70% of HCM patients are caused by mutations in myosin genes. Multiple mutation sites are found in 5-7% of these patients. Approximately 70 pathogenic mutations have been identified, but most are family-specific, with only a few hotspots identified, such as the MYH7 R403Q and R453C mutations (Frey, N. et al., Nat RevCardiol, 9, 2011, 91-100; Sabater-Molina, M. et al., Clin Genet, 93, 2018, 3-14). Studies on the pathogenicity of gene mutations have found that MYH7 gene mutations account for approximately 30% of patients. Compared to other myosin genes, MYH7 leads to earlier disease onset and more severe myocardial hypertrophy. Myosin is a building block of the thick filaments of myofibrils and plays a crucial role in muscle movement. Its molecular shape is bean sprout-like, composed of two heavy chains and multiple light chains. Myosin heads bind to actin to form cross-bridges, significantly increasing myosin ATPase activity. This catalyzes ATP hydrolysis, generating energy to propel the cross-bridges and enable muscle contraction. Research indicates that mutations in the MYH7 gene lead to increased myosin ATPase activity, a decreased proportion of myosin super-relaxed state (SRX), and an increase in cross-bridges between myosin and actin, resulting in abnormal cardiac contractile function (Green, EM. et al., Science, 351, 2016, 617-621; Sommese, RF. et al., Proc Natl Acad Sci USA, 110, 2013, 12607-12612). Therefore, myosin is an important target for the treatment of hypertrophic cardiomyopathy.

[0004] Application No. WO2022105852 discloses and structurally characterizes a series of triazine dione derivatives, including (S)-6-((1-(2-fluoro-5-methylphenyl)ethyl)amino)-3-(tetrahydro-2H-pyran-4-yl)-1,3,5-triazine-2,4(1H,3H)-dione. Furthermore, this application also provides a biological evaluation of (S)-6-((1-(2-fluoro-5-methylphenyl)ethyl)amino)-3-(tetrahydro-2H-pyran-4-yl)-1,3,5-triazine-2,4(1H,3H)-dione, showing that the compound exhibits good inhibitory activity against myosin ATPase.

[0005] The crystal structure of a pharmaceutical active ingredient often affects its chemical stability. Different crystallization and storage conditions can lead to changes in the crystal structure of the compound, sometimes even resulting in other crystal forms. Generally, amorphous drug products lack regular crystal structures and often have other defects, such as poor product stability, fine crystals, difficulty in filtration, easy agglomeration, and poor flowability. Therefore, it is essential to improve the various properties of these products, and we need to conduct in-depth research to find crystal forms with high purity and good physicochemical stability. Summary of the Invention

[0006] This disclosure provides a myosin inhibitor, (S)-6-((1-(2-fluoro-5-methylphenyl)ethyl)amino)-3-(tetrahydro-2H-pyran-4-yl)-1,3,5-triazine-2,4(1H,3H)-dione, its crystal form, preparation method, and application.

[0007] This disclosure provides a crystalline form of (S)-6-((1-(2-fluoro-5-methylphenyl)ethyl)amino)-3-(tetrahydro-2H-pyran-4-yl)-1,3,5-triazine-2,4(1H,3H)-dione.

[0008] In some embodiments, this disclosure provides an X-ray powder diffraction pattern, expressed as a diffraction angle 2θ, of (S)-6-((1-(2-fluoro-5-methylphenyl)ethyl)amino)-3-(tetrahydro-2H-pyran-4-yl)-1,3,5-triazine-2,4(1H,3H)-dione in crystal form A, at 9.2, 10.9, 18.6, 20.1, 22.0, and 2... A characteristic peak is present at 6.9; optionally, characteristic peaks are present at 9.2, 10.9, 13.3, 16.9, 18.6, 19.2, 20.1, 22.0, 26.9, and 35.6; optionally, characteristic peaks are present at 9.2, 10.9, 13.3, 16.9, 18.6, 19.2, 20.1, 20.6, 22.0, 26.9, 28.0, and 35.6.

[0009] In some embodiments, this disclosure provides an X-ray powder diffraction pattern, expressed as a diffraction angle 2θ, of (S)-6-((1-(2-fluoro-5-methylphenyl)ethyl)amino)-3-(tetrahydro-2H-pyran-4-yl)-1,3,5-triazine-2,4(1H,3H)-dione in crystal form A, at 9.2, 10.9, 18.5, 20.1, 22.0, and 2... A characteristic peak is present at 6.9; optionally, characteristic peaks are present at 9.2, 10.9, 13.3, 16.8, 18.5, 19.1, 20.1, 22.0, 26.9, and 35.5; optionally, characteristic peaks are present at 9.2, 10.9, 13.3, 16.8, 18.5, 19.1, 20.1, 20.6, 22.0, 26.9, 27.9, and 35.5.

[0010] In some embodiments, this disclosure provides an X-ray powder diffraction pattern, expressed as a diffraction angle 2θ, for (S)-6-((1-(2-fluoro-5-methylphenyl)ethyl)amino)-3-(tetrahydro-2H-pyran-4-yl)-1,3,5-triazine-2,4(1H,3H)-dione in crystal form B, at 11.1, 15.9, 18.8, 20.4, 22.2, and 26 Å. A characteristic peak is present at 0.8; optionally, characteristic peaks are present at 6.6, 9.5, 11.1, 15.9, 18.8, 20.4, 21.0, 22.2, and 26.8; optionally, characteristic peaks are present at 6.6, 9.5, 11.1, 13.3, 14.0, 15.9, 18.8, 19.0, 19.8, 20.4, 21.0, 22.2, 26.8, and 28.0.

[0011] In some embodiments, this disclosure provides a C-crystal form of (S)-6-((1-(2-fluoro-5-methylphenyl)ethyl)amino)-3-(tetrahydro-2H-pyran-4-yl)-1,3,5-triazine-2,4(1H,3H)-dione, whose X-ray powder diffraction pattern, expressed as a diffraction angle 2θ, has characteristic peaks at 8.3, 14.4, 16.8, 17.8, 19.3, 22.3, and 23.1.

[0012] In some embodiments, this disclosure provides an X-ray powder diffraction pattern of (S)-6-((1-(2-fluoro-5-methylphenyl)ethyl)amino)-3-(tetrahydro-2H-pyran-4-yl)-1,3,5-triazine-2,4(1H,3H)-dione in its D crystal form, expressed as a diffraction angle 2θ, with characteristic peaks at 7.8, 8.7, 9.7, 13.5, 14.0, and 22.9; optionally, characteristic peaks at 7.8, 8.7, 9.7, 13.5, 14.0, 16.7, 19.6, 20.2, and 22.9; optionally, characteristic peaks at 7.8, 8.7, 9.7, 13.5, 14.0, 15.5, 16.7, 17.5, 19.6, 20.2, 22.9, and 23.9.

[0013] In some embodiments, this disclosure provides an E-crystal form of (S)-6-((1-(2-fluoro-5-methylphenyl)ethyl)amino)-3-(tetrahydro-2H-pyran-4-yl)-1,3,5-triazine-2,4(1H,3H)-dione, whose X-ray powder diffraction pattern, expressed as a diffraction angle 2θ, has characteristic peaks at 6.9, 9.3, 14.2, 17.1, 18.0, and 25.0; optionally, characteristic peaks are present at 6.9, 9.3, 13.5, 14.2, 17.1, 18.0, 22.4, 23.2, and 25.0; optionally, characteristic peaks are present at 6.9, 9.3, 13.5, 14.2, 16.1, 17.1, 18.0, 21.1, 22.4, 23.2, 23.8, and 25.0.

[0014] In an optional embodiment, the crystal form of the compound represented by formula (I) provided in this disclosure is wherein the error range of the 2θ angle is ±0.2.

[0015] This disclosure provides a method for preparing (S)-6-((1-(2-fluoro-5-methylphenyl)ethyl)amino)-3-(tetrahydro-2H-pyran-4-yl)-1,3,5-triazine-2,4(1H,3H)-dione A crystal form, selected from any of the following methods.

[0016] Method 1:

[0017] (a) The compound (S)-6-((1-(2-fluoro-5-methylphenyl)ethyl)amino)-3-(tetrahydro-2H-pyran-4-yl)-1,3,5-triazine-2,4(1H,3H)-dione is mixed with solvent (1) and dissolved by stirring or heating. Solvent (1) is selected from at least one of 7% water / ethanol, tetrahydrofuran, 10% water / isopropanol, 10% water / acetone, methanol, ethanol, and 10% water / methanol.

[0018] (b) Crystallization;

[0019] Alternatively, method two:

[0020] (a) The compound (S)-6-((1-(2-fluoro-5-methylphenyl)ethyl)amino)-3-(tetrahydro-2H-pyran-4-yl)-1,3,5-triazine-2,4(1H,3H)-dione was mixed with solvent (2) and dissolved by stirring or heating. Solvent (2) was selected from 10% water / acetone, dichloromethane.

[0021] (b) Add solvent (3) to crystallize, wherein solvent (3) is selected from at least one of water, methanol and ethanol;

[0022] Alternatively, method three:

[0023] (a) The compound (S)-6-((1-(2-fluoro-5-methylphenyl)ethyl)amino)-3-(tetrahydro-2H-pyran-4-yl)-1,3,5-triazine-2,4(1H,3H)-dione is mixed with solvent (4), wherein solvent (4) is selected from at least one of water, methanol, ethanol, isopropanol, acetone, ethyl acetate, acetonitrile, isopropyl acetate, methyl tert-butyl ether, methyl isobutyl ketone, n-heptane, cyclohexane, 1,4-dioxane, 10% water / methanol, and 7% water / ethanol.

[0024] (b) Pulping and crystallization.

[0025] This disclosure provides a method for preparing (S)-6-((1-(2-fluoro-5-methylphenyl)ethyl)amino)-3-(tetrahydro-2H-pyran-4-yl)-1,3,5-triazine-2,4(1H,3H)-dione B crystal form, selected from any of the following methods.

[0026] Method 1:

[0027] (a) The compound (S)-6-((1-(2-fluoro-5-methylphenyl)ethyl)amino)-3-(tetrahydro-2H-pyran-4-yl)-1,3,5-triazine-2,4(1H,3H)-dione was mixed with solvent (5) and dissolved by stirring or heating. Solvent (5) was selected from ethanol.

[0028] (b) Crystallization;

[0029] Alternatively, method two:

[0030] (a) The compound (S)-6-((1-(2-fluoro-5-methylphenyl)ethyl)amino)-3-(tetrahydro-2H-pyran-4-yl)-1,3,5-triazine-2,4(1H,3H)-dione was mixed with solvent (6) and dissolved by stirring or heating. Solvent (6) was selected from dichloromethane.

[0031] (b) Add solvent (7) to crystallize, wherein solvent (7) is selected from at least one of ethyl acetate, acetone, acetonitrile, and methyl isobutyl ketone.

[0032] This disclosure provides a method for preparing the (S)-6-((1-(2-fluoro-5-methylphenyl)ethyl)amino)-3-(tetrahydro-2H-pyran-4-yl)-1,3,5-triazine-2,4(1H,3H)-dione C crystal form, comprising:

[0033] (a) The compound (S)-6-((1-(2-fluoro-5-methylphenyl)ethyl)amino)-3-(tetrahydro-2H-pyran-4-yl)-1,3,5-triazine-2,4(1H,3H)-dione was mixed with solvent (9) and dissolved by stirring or heating. Solvent (9) was selected from dichloromethane.

[0034] (b) Add solvent (10) to crystallize, wherein solvent (10) is selected from isopropyl acetate.

[0035] This disclosure provides a method for preparing (S)-6-((1-(2-fluoro-5-methylphenyl)ethyl)amino)-3-(tetrahydro-2H-pyran-4-yl)-1,3,5-triazine-2,4(1H,3H)-dione D crystal form, comprising:

[0036] (a) The compound (S)-6-((1-(2-fluoro-5-methylphenyl)ethyl)amino)-3-(tetrahydro-2H-pyran-4-yl)-1,3,5-triazine-2,4(1H,3H)-dione was mixed with solvent (11), wherein solvent (11) was selected from dichloromethane.

[0037] (b) Add solvent (12) to slurry and crystallize. Solvent (12) is selected from methyl tert-butyl ether.

[0038] This disclosure provides a method for preparing (S)-6-((1-(2-fluoro-5-methylphenyl)ethyl)amino)-3-(tetrahydro-2H-pyran-4-yl)-1,3,5-triazine-2,4(1H,3H)-dione E crystal form, comprising:

[0039] (a) The compound (S)-6-((1-(2-fluoro-5-methylphenyl)ethyl)amino)-3-(tetrahydro-2H-pyran-4-yl)-1,3,5-triazine-2,4(1H,3H)-dione was mixed with solvent (13), wherein solvent (13) was selected from dichloromethane.

[0040] (b) Add solvent (14) to slurry and crystallize. Solvent (14) is selected from n-heptane.

[0041] In some embodiments, the method for preparing the crystal form described in this disclosure further includes one or more steps of filtration, washing, or drying.

[0042] This disclosure also provides a pharmaceutical composition prepared from the crystal form of the aforementioned (S)-6-((1-(2-fluoro-5-methylphenyl)ethyl)amino)-3-(tetrahydro-2H-pyran-4-yl)-1,3,5-triazine-2,4(1H,3H)-dione.

[0043] This disclosure also provides a pharmaceutical composition comprising the aforementioned crystalline form of (S)-6-((1-(2-fluoro-5-methylphenyl)ethyl)amino)-3-(tetrahydro-2H-pyran-4-yl)-1,3,5-triazine-2,4(1H,3H)-dione or a mixture thereof, or the crystalline form of (S)-6-((1-(2-fluoro-5-methylphenyl)ethyl)amino)-3-(tetrahydro-2H-pyran-4-yl)-1,3,5-triazine-2,4(1H,3H)-dione prepared by the aforementioned method, and optionally a pharmaceutically acceptable excipient.

[0044] This disclosure also provides a method for preparing a pharmaceutical composition, comprising the step of mixing the crystal form of (S)-6-((1-(2-fluoro-5-methylphenyl)ethyl)amino)-3-(tetrahydro-2H-pyran-4-yl)-1,3,5-triazine-2,4(1H,3H)-dione or a mixture thereof, or the crystal form of (S)-6-((1-(2-fluoro-5-methylphenyl)ethyl)amino)-3-(tetrahydro-2H-pyran-4-yl)-1,3,5-triazine-2,4(1H,3H)-dione prepared by the aforementioned method with a pharmaceutically acceptable excipient.

[0045] This disclosure also provides the use of (S)-6-((1-(2-fluoro-5-methylphenyl)ethyl)amino)-3-(tetrahydro-2H-pyran-4-yl)-1,3,5-triazine-2,4(1H,3H)-dione, or mixtures thereof, or crystal forms or mixtures thereof prepared by the foregoing methods, or the foregoing compositions, or compositions prepared by the foregoing methods, in the preparation of medicaments for treating diseases or conditions related to myosin regulation.

[0046] This disclosure also provides the use of (S)-6-((1-(2-fluoro-5-methylphenyl)ethyl)amino)-3-(tetrahydro-2H-pyran-4-yl)-1,3,5-triazine-2,4(1H,3H)-dione, or mixtures thereof, or the crystal forms or mixtures thereof prepared by the foregoing methods, or the foregoing compositions, or the compositions prepared by the foregoing methods, in the preparation of a medicament for treating a disease or condition selected from diastolic heart failure with preserved ejection fraction, ischemic heart disease, angina pectoris, restrictive cardiomyopathy, diastolic dysfunction, hypertrophic cardiomyopathy (HCM), non-obstructive hypertrophic cardiomyopathy (nHCM), obstructive hypertrophic cardiomyopathy (oHCM), and valves. Diseases, heart failure with normal ejection fraction (HFpEF), heart failure with median ejection fraction (HFmREF), aortic stenosis, inflammatory cardiomyopathy, Leffler endocarditis, myocardial endocardial fibrosis, infiltrative cardiomyopathy, hemochromatosis, Fabry disease, glycogen storage disease, congenital heart disease, tetralogy of Fallot, left ventricular hypertrophy, refractory angina, and Chagas disease; optionally, selected from ischemic heart disease, restrictive cardiomyopathy, hypertrophic cardiomyopathy (HCM), non-obstructive hypertrophic cardiomyopathy (nHCM), obstructive hypertrophic cardiomyopathy (oHCM), inflammatory cardiomyopathy, infiltrative cardiomyopathy, congenital heart disease, and left ventricular hypertrophy; optionally, the disease or condition is hypertrophic cardiomyopathy (HCM).

[0047] The "2θ or 2θ angle" mentioned in this disclosure refers to the diffraction angle, where θ is the Bragg angle, and the unit is ° or degree; the error range of 2θ for each characteristic peak is ±0.20 (including the case where the number has more than one decimal place after rounding), 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.

[0048] The “crystallization” or “precipitation” described in this disclosure includes, but is not limited to, stirring crystallization, pulping crystallization, cooling crystallization, and volatilization crystallization.

[0049] The “differential scanning calorimetry or DSC” described in this disclosure refers to measuring the temperature difference and heat flow difference between the sample and the reference material during the sample heating or isothermal process, in order to characterize all physical and chemical changes related to thermal effects and obtain phase transition information of the sample.

[0050] The “pharmaceuticalally acceptable excipients” described in this disclosure include, but are not limited to, any adjuvant, carrier, flow aid, sweetener, diluent, preservative, dye / coloring agent, flavoring agent, surfactant, wetting agent, dispersant, suspending agent, stabilizer, isotonic agent, or emulsifier that has been approved by the U.S. Food and Drug Administration for use in humans or livestock. Attached Figure Description

[0051] Figure 1 The amorphous XRD pattern of (S)-6-((1-(2-fluoro-5-methylphenyl)ethyl)amino)-3-(tetrahydro-2H-pyran-4-yl)-1,3,5-triazine-2,4(1H,3H)-dione is shown.

[0052] Figure 2 XRD pattern of (S)-6-((1-(2-fluoro-5-methylphenyl)ethyl)amino)-3-(tetrahydro-2H-pyran-4-yl)-1,3,5-triazine-2,4(1H,3H)-dione A crystal form.

[0053] Figure 3 XRD pattern of (S)-6-((1-(2-fluoro-5-methylphenyl)ethyl)amino)-3-(tetrahydro-2H-pyran-4-yl)-1,3,5-triazine-2,4(1H,3H)-dione B crystal form.

[0054] Figure 4 XRD pattern of (S)-6-((1-(2-fluoro-5-methylphenyl)ethyl)amino)-3-(tetrahydro-2H-pyran-4-yl)-1,3,5-triazine-2,4(1H,3H)-dione in C crystal form.

[0055] Figure 5 XRD pattern of (S)-6-((1-(2-fluoro-5-methylphenyl)ethyl)amino)-3-(tetrahydro-2H-pyran-4-yl)-1,3,5-triazine-2,4(1H,3H)-dione in D crystal form.

[0056] Figure 6 XRD pattern of (S)-6-((1-(2-fluoro-5-methylphenyl)ethyl)amino)-3-(tetrahydro-2H-pyran-4-yl)-1,3,5-triazine-2,4(1H,3H)-dione in E crystal form. Detailed Implementation

[0057] This disclosure is further described in detail through the following embodiments and experimental examples. These embodiments and experimental examples are for illustrative purposes only and are not intended to limit the scope of this disclosure.

[0058] Test conditions of the instruments used in the experiment:

[0059] The structure of the compound was determined by nuclear magnetic resonance (NMR) and / or mass spectrometry (MS). NMR shifts (δ) were expressed in 10⁻¹⁰ ohms. -6 The unit (ppm) is given. 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.

[0060] MS measurements were performed using an Agilent 1200 / 1290DAD-6110 / 6120 Quadrupole MS LC-MS system (manufacturer: Agilent, MS model: 6110 / 6120 Quadrupole MS), a Waters ACQuity UPLC-QD / SQD system (manufacturer: Waters, MS model: Waters ACQuity Qda Detector / Waters SQ Detector), or a THERMO Ultimate3000-Q Exactive system (manufacturer: THERMO, MS model: THERMO Q Exactive).

[0061] High-performance liquid chromatography (HPLC) analysis was performed using an Agilent HPLC 1200DAD, an Agilent HPLC 1200VWD, and a Waters HPLC e2695-2489 high-performance liquid chromatograph.

[0062] Chiral HPLC analysis was performed using an Agilent 1260DAD high-performance liquid chromatograph.

[0063] High performance liquid chromatography (HPLC) was performed using Waters 2767, Waters 2767-SQ Detecor2, Shimadzu LC-20AP, and Gilson-281 preparative chromatographs.

[0064] Chiral preparation was performed using a Shimadzu LC-20AP preparative chromatograph.

[0065] The CombiFlash rapid preparation system uses a CombiFlash Rf200 (TELEDYNE ISCO).

[0066] Thin-layer chromatography silica gel plates are Yantai Huanghai HSGF254 or Qingdao GF254. The silica gel plates used in thin-layer chromatography (TLC) have a diameter of 0.15 mm to 0.2 mm, and the diameter of the silica gel plates used for thin-layer chromatography separation and purification products is 0.4 mm to 0.5 mm.

[0067] Silica gel column chromatography generally uses Yantai Huanghai silica gel with a mesh size of 200-300 as the carrier.

[0068] The known starting materials disclosed herein can be synthesized using or in accordance with methods known in the art, or can be purchased from companies such as ABCR GmbH & Co. KG, Acros Organics, Aldrich Chemical Company, AccelaChemBio Inc, and Darui Chemicals.

[0069] Unless otherwise specified in the examples, the reactions can be carried out under an argon or nitrogen atmosphere.

[0070] Argon or nitrogen atmosphere refers to a reaction flask connected to an argon or nitrogen gas balloon with a volume of approximately 1L.

[0071] A hydrogen atmosphere refers to a reaction vessel connected to a hydrogen balloon with a volume of approximately 1L.

[0072] The pressurized hydrogenation reaction was performed using a Parr 3916EKX hydrogenator and a Qinglan QL-500 hydrogen generator or an HC2-SS hydrogenator.

[0073] The hydrogenation reaction is usually carried out under vacuum, filled with hydrogen gas, and repeated 3 times.

[0074] The microwave reaction was performed using a CEM Discover-S 908860 microwave reactor.

[0075] Unless otherwise specified in the examples, "solution" refers to an aqueous solution.

[0076] Unless otherwise specified in the examples, the reaction temperature is room temperature, which is 20℃~30℃.

[0077] The reaction process in the examples was monitored using thin-layer chromatography (TLC). The developing solvent used in the reaction, the eluent system used for column chromatography to purify the compounds, and the developing solvent system for TLC included: A: n-hexane / ethyl acetate system, B: dichloromethane / methanol system. The volume ratio of the solvent was adjusted according to the polarity of the compounds, and small amounts of basic or acidic reagents such as triethylamine and acetic acid could also be added for adjustment.

[0078] When the compounds in the examples contain two or more chiral centers, their relative stereochemistry is identified by NMR studies and / or X-ray diffraction. In these cases, the compounds are identified using the prefix "rel" followed by R / S nomenclature, where the R / S only provides relative stereochemical information (e.g., trans or cis) and does not indicate absolute stereochemistry.

[0079] XRPD (X-ray Powder Diffraction) was performed using a BRUKER D8 X-ray diffractometer. Specific data acquisition information included: Cu anode (40 kV, 40 mA), and monochromatic Cu-Kα rays. Scanning mode: θ / 2θ, scanning range (2θ range): 5~50° (or 3-48°).

[0080] DSC stands for Differential Scanning Calorimetry: Measurements were performed using a METTLER TOLEDO DSC 3+ differential scanning calorimeter with a heating rate of 10℃ / min. The specific temperature range was referenced from the corresponding spectra (mostly 25-300℃), and the nitrogen purging rate was 50mL / min.

[0081] TGA is thermogravimetric analysis: the test was performed using a METTLER TOLEDO TGA 2 thermogravimetric analyzer, with a heating rate of 10℃ / min, and the specific temperature range was referenced from the corresponding spectrum (mostly 25-300℃). The nitrogen purging rate was 50mL / min.

[0082] DVS stands for Dynamic Moisture Adsorption: Detection is performed using SMS DVS Advantage at 25℃, with humidity changes of 50%-95%-0%-95%-50%, in 10% increments (the final step is 5%) (the specific humidity range is subject to the corresponding spectrum; the methods listed here are the most commonly used). The judgment criterion is that dm / dt is not greater than 0.002%.

[0083] Example 1.

[0084] Preparation of (S)-6-((1-(2-fluoro-5-methylphenyl)ethyl)amino)-3-(tetrahydro-2H-pyran-4-yl)-1,3,5-triazine-2,4(1H,3H)-dione (refer to the preparation method in Example 16 of application No. WO2022105852)

[0085]

[0086] first step

[0087] 4-Isocyanoxytetrahydro-2H-pyran 1b

[0088] At 15°C, a solution of tetrahydro-2H-pyran-4-amine 1a (10.0 g, 100.0 mmol, Shanghai Shaoyuan Technology Co., Ltd.) and N,N-diisopropylethylamine (28.4 g, 220.0 mmol, Shanghai Titan Technology Co., Ltd.) in anhydrous dichloromethane (120 mL) was slowly added dropwise to a solution of bis(trichloromethyl) carbonate (11.9 g, 40.0 mmol, Shanghai Titan Technology Co., Ltd.) in anhydrous dichloromethane (120 mL). The reaction was allowed to proceed at room temperature for 2 hours. The reaction solution was concentrated under reduced pressure to obtain crude product 1b. The crude product was used directly in the next step without further purification.

[0089] Step 2

[0090] 6-(1H-pyrazol-1-yl)-3-(tetrahydro-2H-pyran-4-yl)-1,3,5-triazine-2,4(1H,3H)-dione 1d

[0091] At -10°C, 1,8-diazobisspirocyclic [5.4.0]undec-7-ene (15.2 g, 100.0 mmol, Shanghai Shaoyuan Technology Co., Ltd.) was slowly added dropwise to an anhydrous N,N-dimethylacetamide (120 mL) solution of compound 1c (10.5 g, 95.2 mmol) and crude compound 1b. The reaction was stirred at 0°C for 1 hour. Subsequently, carbonyl diimidazole (23.2 g, 142.8 mmol, Shanghai Bide Technology Co., Ltd.) and 1,8-diazobisspirocyclic [5.4.0]undec-7-ene (21.7 g, 142.8 mmol, Shanghai Shaoyuan Technology Co., Ltd.) were added at 0°C. The reaction was stirred at 60°C for 16 hours. The reaction solution was concentrated under reduced pressure, and dichloromethane was added to the resulting residue. The mixture was stirred, filtered, and the filter cake was collected and dried under reduced pressure to give the title product 1d (16.6 g, two-step yield: 63.4%). MS m / z(ESI):264.1[M+1].

[0092] Step 3

[0093] (R)-N-((S)-1-(2-fluoro-5-methylphenyl)ethyl)-2-methylpropane-2-sulfinamide 1f

[0094] (R)-N-(2-fluoro-5-methylbenzyl)-2-methylpropane-2-sulfinamide 1e (6.90 g, 28.59 mmol, prepared by the method disclosed in intermediate 3B on page 56 of patent application "WO2020092208A1") was dissolved in dichloromethane (100 mL) and purged three times with nitrogen. The reaction was cooled to -60 °C, and a 3M solution of methylmagnesium bromide in 2-methyltetrahydrofuran (19.1 mL, 57.18 mmol) was added dropwise. The reaction was stirred at room temperature for 2 hours under nitrogen protection. A saturated ammonium chloride solution (100 mL) was added, and the organic phase was separated. The aqueous phase was extracted with ethyl acetate (80 mL × 2), the organic phases were combined, washed with saturated sodium chloride solution (100 mL), dried over anhydrous sodium sulfate, filtered, concentrated under reduced pressure, and purified by high performance liquid chromatography (BostonPhlex C18 150*30 mm, 5 μm, elution system: water (10 mmol ammonium bicarbonate), acetonitrile, 20-95% acetonitrile, gradient elution for 20 min, flow rate: 30 mL / min) to give the title product 1f (5.60 g, yield: 76.1%). MS m / z (ESI): 258.0 [M+1].

[0095] Step 4

[0096] (S)-1-(2-fluoro-5-methylphenyl)ethylamine hydrochloride 1g

[0097] Compound 1f (670 mg, 2.60 mmol) was dissolved in ethanol (10 mL), cooled to 0 °C, and thionyl chloride (620 mg, 5.21 mmol) was added dropwise. The reaction was stirred at room temperature for 1 hour. The mixture was concentrated under reduced pressure to give 1 g of crude title product (493 mg, yield: 99.9%). The crude product was used directly in the next step without purification. MS m / z (ESI): 153.9 [M+1].

[0098] Step 5

[0099] (S)-6-((1-(2-fluoro-5-methylphenyl)ethyl)amino)-3-(tetrahydro-2H-pyran-4-yl)-1,3,5-triazine-2,4(1H,3H)-dione

[0100] Compound 1d (425 mg, 1.61 mmol) and compound 1g (297 mg, 1.94 mmol) were dissolved in 1,4-dioxane (10 mL) and reacted at 120 °C with stirring for 16 hours. The reaction solution was concentrated under reduced pressure and purified by high performance liquid chromatography (Boston Phlex C18 150*30 mm, 5 μm, elution system: water (10 mmol ammonium bicarbonate), acetonitrile, 20-95% acetonitrile, gradient elution for 20 min, flow rate: 30 mL / min) to give product 1 (310 mg, yield: 55.1%). MS m / z (ESI): 349.0 [M+1].

[0101] 1 H NMR(500MHz,DMSO-d6)δ10.50(brs,1H),7.28-7.05(m,4H),5.22(m,1H),4.64(m,1H),3 .90-3.86(m,2H),3.33-3.27(m,2H),2.54-2.42(m,2H),2.28(s,3H),1.45-1.39(m,5H).

[0102] Example 2. Preparation of amorphous (S)-6-((1-(2-fluoro-5-methylphenyl)ethyl)amino)-3-(tetrahydro-2H-pyran-4-yl)-1,3,5-triazine-2,4(1H,3H)-dione

[0103] The substance purified by high-performance liquid chromatography in Example 1 was lyophilized to obtain the product. X-ray powder diffraction analysis identified the product as an amorphous (S)-6-((1-(2-fluoro-5-methylphenyl)ethyl)amino)-3-(tetrahydro-2H-pyran-4-yl)-1,3,5-triazine-2,4(1H,3H)-dione. The X-ray powder diffraction pattern, expressed as a diffraction angle of 2θ, showed no characteristic peaks in the range of 3-48°. The XRD pattern is shown below. Figure 1 .

[0104] Example 3. Inhibitory effect of (S)-6-((1-(2-fluoro-5-methylphenyl)ethyl)amino)-3-(tetrahydro-2H-pyran-4-yl)-1,3,5-triazine-2,4(1H,3H)-dione on myosin ATPase activity.

[0105] The following method was used to determine the inhibitory effect of a compound on myosin ATPase activity. The experimental method is briefly described below:

[0106] 1. Experimental materials and instruments

[0107] a. Cardiac actin (Cytoskeleton, AD99)

[0108] b. Myosin Motor Protein S1 Fragment (Cytoskeleton, CS-MYS03)

[0109] c.ATP (Sigma, A7699-1G)

[0110] d. UltraPure™ 1M Tris-HCl buffer, pH 7.5 (Thermo, 15567027)

[0111] e.CytoPhos™ Phosphate Detection Biological Kit (Cytoskeleton, BK054)

[0112] f. Magnesium chloride solution (Sigma, 68475-100ML-F)

[0113] g. Potassium chloride solution (Sigma, 60142-100ML-F)

[0114] h.EGTA (Sigma, E3889-100G)

[0115] i.96 well plate (Corning, 3697)

[0116] JU-shaped bottom 96-hole plate (Corning, 3795)

[0117] k. Microplate reader (BMG, Pherastar)

[0118] l. Constant temperature incubator (Shanghai Boxun, SPX-100B-Z)

[0119] 2. Experimental Procedure

[0120] 1.61 μM of cardiac actin and 0.07 μM of myosin motor protein S1 fragment were mixed with different concentrations of small molecule compounds (initial concentration 100 μM, serially diluted 3-fold to 9 concentrations) and incubated at 37°C for 1 hour. Then, 120 μM of ATP was added and incubated at 37°C for 2 hours. Finally, 70 μL of the detection solution from the CytoPhos™ Phosphate Detection Kit was added to each well and incubated at room temperature for 10 minutes. OD values ​​at 650 nM were read using a microplate reader, and the amount of Pi was calculated based on the standard curve. Data were processed using GraphPad software, and inhibition curves were plotted based on the compound concentrations and corresponding inhibition rates. The concentration of the compound at which the inhibition rate reached 50%, i.e., the IC50, was calculated. 50 value.

[0121] Conclusion: The IC50 values ​​of (S)-6-((1-(2-fluoro-5-methylphenyl)ethyl)amino)-3-(tetrahydro-2H-pyran-4-yl)-1,3,5-triazine-2,4(1H,3H)-dione for inhibiting myosin ATPase activity were [not specified]. 50 At a concentration of 1.06 μM, it exhibits excellent inhibitory effects on myosin ATPase.

[0122] Example 4. Toxicokinetic evaluation of (S)-6-((1-(2-fluoro-5-methylphenyl)ethyl)amino)-3-(tetrahydro-2H-pyran-4-yl)-1,3,5-triazine-2,4(1H,3H)-dione in SD rats after repeated gavage administration for 14 days.

[0123] 1. Abstract

[0124] Using SD rats as test animals, the concentrations of the original drug in plasma and administration solution at different time points after gavage administration of the test compound to SD rats were determined by LC / MS / MS. The toxicokinetics of (S)-6-((1-(2-fluoro-5-methylphenyl)ethyl)amino)-3-(tetrahydro-2H-pyran-4-yl)-1,3,5-triazine-2,4(1H,3H)-dione in SD rats were investigated, and its toxicokinetics were evaluated.

[0125] 2. Experimental Design

[0126] 2.1 Test Drugs

[0127] (S)-6-((1-(2-fluoro-5-methylphenyl)ethyl)amino)-3-(tetrahydro-2H-pyran-4-yl)-1,3,5-triazine-2,4(1H,3H)-dione, compound MYK-461 Example 1 of WO2014205223A1).

[0128] 2.2 Laboratory Animals

[0129] Twenty-four SD rats, half male and half female, were divided into six groups of four rats each, with half male and half female, and were provided by Vital River Laboratory Animal Co., Ltd.

[0130] 2.3 Drug Preparation

[0131] Weigh a certain amount of (S)-6-((1-(2-fluoro-5-methylphenyl)ethyl)amino)-3-(tetrahydro-2H-pyran-4-yl)-1,3,5-triazine-2,4(1H,3H)-dione, add 15% PEG400 and 85% (10% TPGS + 1% HPMC K100LV) to prepare a pale yellow homogeneous suspension.

[0132] Weigh a certain amount of compound MYK-461 and add 0.5% MC to prepare a colorless and clear solution.

[0133] 2.4 Administration

[0134] For oral administration, the dosages of (S)-6-((1-(2-fluoro-5-methylphenyl)ethyl)amino)-3-(tetrahydro-2H-pyran-4-yl)-1,3,5-triazine-2,4(1H,3H)-dione were 5 mg / kg, 15 mg / kg, and 30 mg / kg, with an administration volume of 10 mL / kg for each. The dosages of compound MYK-461 were 0.5 mg / kg, 1.5 mg / kg, and 3 mg / kg, with an administration volume of 10 mL / kg for each.

[0135] 3. Operation

[0136] On day 1, 0.2 mL of blood was collected from the orbital cavity at 0.5 h, 1.0 h, 2.0 h, 4.0 h, 8.0 h, and 24.0 h after drug administration. On days 7 and 14, 0.2 mL of blood was collected from the orbital cavity before drug administration and at 0.5 h, 1.0 h, 2.0 h, 4.0 h, 8.0 h, and 24.0 h after drug administration. The blood samples were placed in EDTA-K2 anticoagulant tubes, centrifuged at 10,000 rpm for 1 min (4℃), and the plasma was separated within 1 hour and stored at -20℃ for analysis. The blood collection and centrifugation processes were performed under ice bath conditions. Patients ate 2 hours after drug administration.

[0137] Determination of the content of the target compound in the plasma of SD rats after gavage administration of different concentrations of the drug: 20 μL of plasma from SD rats at each time point after drug administration was taken, and 50 μL of internal standard solution ((S)-6-((1-(2-fluoro-5-methylphenyl)ethyl)amino)-3-(tetrahydro-2H-pyran-4-yl)-1,3,5-triazine-2,4(1H,3H)-dione internal standard: verapamil 100 ng / mL; internal standard of compound MYK-461: camptothecin 100 ng / mL) and 200 μL of acetonitrile were added. The mixture was vortexed for 5 minutes and centrifuged for 10 minutes (3700 rpm). 1 μL of the supernatant from the plasma sample was taken for LC / MS / MS analysis.

[0138] 4. Results of toxicokinetics

[0139] Table 1. Toxicokinetic parameters of the compounds in SD rats.

[0140]

[0141]

[0142] Conclusion: (S)-6-((1-(2-fluoro-5-methylphenyl)ethyl)amino)-3-(tetrahydro-2H-pyran-4-yl)-1,3,5-triazine-2,4(1H,3H)-dione showed no significant accumulation in SD rats after repeated gavage administration for 14 days, while compound MYK-461 showed severe accumulation in SD rats, increasing the risk of drug use. Clearly, (S)-6-((1-(2-fluoro-5-methylphenyl)ethyl)amino)-3-(tetrahydro-2H-pyran-4-yl)-1,3,5-triazine-2,4(1H,3H)-dione has a significant toxicokinetic advantage over compound MYK-461.

[0143] Example 5. Preparation of crystal form of compound A

[0144] Weigh 300 mg of (S)-6-((1-(2-fluoro-5-methylphenyl)ethyl)amino)-3-(tetrahydro-2H-pyran-4-yl)-1,3,5-triazine-2,4(1H,3H)-dione, add 6 mL of 7% water / ethanol, heat to 80 °C to dissolve, cool to room temperature, stir overnight, filter, and dry the solid under vacuum to obtain the product. X-ray powder diffraction analysis identified the product as crystal form A. The X-ray powder diffraction data are shown in Table 2, and the X-ray powder diffraction pattern is shown in... Figure 2 As shown.

[0145] The DSC spectrum shows an endothermic peak at 256.71℃. The TGA spectrum shows a weight loss of 0.59% between 30℃ and 100℃.

[0146] DVS testing showed that under normal storage conditions (i.e., room temperature, 60% RH), the sample's moisture absorption weight gain was approximately 0.12%; under accelerated testing conditions (i.e., 70% RH), the moisture absorption weight gain was approximately 0.15%; and under extreme conditions (i.e., 90% RH), the moisture absorption weight gain was approximately 0.24%. Furthermore, the crystal form did not change after DVS testing and subsequent retesting.

[0147] Table 2 Peak positions of compound A crystal form

[0148]

[0149]

[0150] Example 6. Preparation of crystal form of compound A

[0151] (S)-6-((1-(2-fluoro-5-methylphenyl)ethyl)amino)-3-(tetrahydro-2H-pyran-4-yl)-1,3,5-triazine-2,4(1H,3H)-dione was added to a solvent and stirred into a slurry. After centrifugation, the solid was dried under vacuum to obtain the product. The crystalline form was determined by X-ray powder diffraction, as shown in Table 3.

[0152] Table 3 Preparation of Crystal Form A by Pulping and Crystallization

[0153]

[0154] Example 7. Preparation of crystal form of compound A

[0155] (S)-6-((1-(2-fluoro-5-methylphenyl)ethyl)amino)-3-(tetrahydro-2H-pyran-4-yl)-1,3,5-triazine-2,4(1H,3H)-dione was added to a solvent and stirred into a slurry. After centrifugation, the solid was dried under vacuum to obtain the product. The crystalline form was determined by X-ray powder diffraction, as shown in Table 4.

[0156] Table 4. Preparation of Crystal Form A by Pulping and Crystallization

[0157]

[0158] Example 8. Preparation of crystal form of compound A

[0159] Weigh 10 mg of (S)-6-((1-(2-fluoro-5-methylphenyl)ethyl)amino)-3-(tetrahydro-2H-pyran-4-yl)-1,3,5-triazine-2,4(1H,3H)-dione, add 0.1 mL of tetrahydrofuran, stir until dissolved, and allow to evaporate and crystallize to obtain the product. X-ray powder diffraction analysis showed that the product was in crystal form A.

[0160] Example 9. Preparation of crystal form of compound A

[0161] Weigh 10 mg of (S)-6-((1-(2-fluoro-5-methylphenyl)ethyl)amino)-3-(tetrahydro-2H-pyran-4-yl)-1,3,5-triazine-2,4(1H,3H)-dione, add 1 mL of 10% water / isopropanol, stir to dissolve, and allow to evaporate and crystallize to obtain the product. X-ray powder diffraction analysis showed that the product was in crystal form A.

[0162] Example 10. Preparation of the crystal form of compound A

[0163] Weigh 10 mg of (S)-6-((1-(2-fluoro-5-methylphenyl)ethyl)amino)-3-(tetrahydro-2H-pyran-4-yl)-1,3,5-triazine-2,4(1H,3H)-dione, add 1 mL of 10% water / acetone, stir to dissolve, and allow to evaporate and crystallize to obtain the product. X-ray powder diffraction analysis showed that the product was in crystal form A.

[0164] Example 11. Preparation of the crystal form of compound A

[0165] 10 mg of (S)-6-((1-(2-fluoro-5-methylphenyl)ethyl)amino)-3-(tetrahydro-2H-pyran-4-yl)-1,3,5-triazine-2,4(1H,3H)-dione was weighed and added to 1 mL of methanol. The mixture was stirred at 50 °C until dissolved, and then cooled to crystallize, yielding the product. X-ray powder diffraction analysis showed that the product was in crystal form A.

[0166] Example 12. Preparation of crystal form of compound A

[0167] Weigh 10 mg of (S)-6-((1-(2-fluoro-5-methylphenyl)ethyl)amino)-3-(tetrahydro-2H-pyran-4-yl)-1,3,5-triazine-2,4(1H,3H)-dione, add 1 mL of ethanol, stir at 50 °C until dissolved, and cool to crystallize to obtain the product. X-ray powder diffraction analysis showed that the product was in crystal form A.

[0168] Example 13. Preparation of the crystal form of compound A

[0169] Weigh 10 mg of (S)-6-((1-(2-fluoro-5-methylphenyl)ethyl)amino)-3-(tetrahydro-2H-pyran-4-yl)-1,3,5-triazine-2,4(1H,3H)-dione, add 1 mL of 10% water / methanol, stir at 50 °C until dissolved, and cool to crystallize to obtain the product. X-ray powder diffraction analysis showed that the product was in crystal form A.

[0170] Example 14. Preparation of crystal form of compound A

[0171] Weigh 10 mg of (S)-6-((1-(2-fluoro-5-methylphenyl)ethyl)amino)-3-(tetrahydro-2H-pyran-4-yl)-1,3,5-triazine-2,4(1H,3H)-dione, add 1 mL of 7% water / ethanol, stir at 50 °C until dissolved, and cool to crystallize to obtain the product. X-ray powder diffraction analysis showed that the product was in crystal form A.

[0172] Example 15. Preparation of the crystal form of compound A

[0173] Weigh 5 mg of (S)-6-((1-(2-fluoro-5-methylphenyl)ethyl)amino)-3-(tetrahydro-2H-pyran-4-yl)-1,3,5-triazine-2,4(1H,3H)-dione, add 0.05 mL of dichloromethane and stir at room temperature until dissolved. Add 0.35 mL of methanol and stir. Slurry at room temperature for 3 days. After centrifugation, dry the solid under vacuum to obtain the product. X-ray powder diffraction analysis showed that the product is crystal form A.

[0174] Example 16. Preparation of the crystal form of compound A

[0175] Weigh 5 mg of (S)-6-((1-(2-fluoro-5-methylphenyl)ethyl)amino)-3-(tetrahydro-2H-pyran-4-yl)-1,3,5-triazine-2,4(1H,3H)-dione, add 0.05 mL of dichloromethane and stir at room temperature until dissolved. Add 0.35 mL of ethanol and stir. Slurry at room temperature for 3 days. After centrifugation, dry the solid under vacuum to obtain the product. X-ray powder diffraction analysis showed that the product was in crystal form A.

[0176] Example 17. Preparation of the crystal form of compound A

[0177] Weigh 1 g of (S)-6-((1-(2-fluoro-5-methylphenyl)ethyl)amino)-3-(tetrahydro-2H-pyran-4-yl)-1,3,5-triazine-2,4(1H,3H)-dione, add 30 mL of 10% water / acetone, stir at 50 °C until dissolved, add 30 mL of water and stir at 50 °C for 5 hours, then cool to 25 °C and stir overnight. Filter and dry the solid under vacuum to obtain the product. X-ray powder diffraction analysis showed that the product was in crystal form A.

[0178] Example 18. Preparation of the crystal form of compound B

[0179] 5 g of (S)-6-((1-(2-fluoro-5-methylphenyl)ethyl)amino)-3-(tetrahydro-2H-pyran-4-yl)-1,3,5-triazine-2,4(1H,3H)-dione was dissolved in 250 mL of anhydrous ethanol and heated to reflux until the solid dissolved completely. Heating was stopped and the mixture was allowed to stand. The mixture was filtered, the filter cake was collected, and dried to obtain the product. X-ray powder diffraction analysis identified the product as crystal form B. The X-ray powder diffraction data are shown in Table 5, and the X-ray powder diffraction pattern is shown in [Figure 5]. Figure 3 As shown.

[0180] The DSC spectrum shows an endothermic peak at 256.45℃. The TGA spectrum shows a weight loss of 1.32% for the compound between 40℃ and 190℃. DVS experimental data show that under normal storage conditions (25℃, 60% RH), the sample's moisture absorption weight gain is approximately 0.242%; under accelerated testing conditions (70% RH), the moisture absorption weight gain is approximately 0.298%; and under extreme conditions (90% RH), the moisture absorption weight gain is approximately 0.418%. XRD patterns show that the crystal form did not change before and after DVS testing.

[0181] Table 5 Peak positions of compound B crystal form

[0182]

[0183] Example 19. Preparation of the crystal form of compound B

[0184] Weigh 30 mg of (S)-6-((1-(2-fluoro-5-methylphenyl)ethyl)amino)-3-(tetrahydro-2H-pyran-4-yl)-1,3,5-triazine-2,4(1H,3H)-dione, add 0.3 mL of dichloromethane and stir at room temperature until dissolved. Add 2.1 mL of ethyl acetate and stir for 2 days at room temperature. Filter and dry the solid under vacuum to obtain the product. X-ray powder diffraction analysis showed that the product was in crystal form B.

[0185] Example 20. Preparation of the crystal form of compound B

[0186] Weigh 5 mg of (S)-6-((1-(2-fluoro-5-methylphenyl)ethyl)amino)-3-(tetrahydro-2H-pyran-4-yl)-1,3,5-triazine-2,4(1H,3H)-dione, add 0.05 mL of dichloromethane and stir at room temperature until dissolved. Add 0.35 mL of acetone and stir. Slurry at room temperature for 3 days. After centrifugation, dry the solid under vacuum to obtain the product. X-ray powder diffraction analysis showed that the product is in crystal form B.

[0187] Example 21. Preparation of the crystal form of compound B

[0188] Weigh 5 mg of (S)-6-((1-(2-fluoro-5-methylphenyl)ethyl)amino)-3-(tetrahydro-2H-pyran-4-yl)-1,3,5-triazine-2,4(1H,3H)-dione, add 0.05 mL of dichloromethane and stir at room temperature until dissolved. Add 0.35 mL of ethyl acetate and stir. Slurry at room temperature for 3 days. After centrifugation, dry the solid under vacuum to obtain the product. X-ray powder diffraction analysis showed that the product is in crystal form B.

[0189] Example 22. Preparation of the crystal form of compound B

[0190] Weigh 5 mg of (S)-6-((1-(2-fluoro-5-methylphenyl)ethyl)amino)-3-(tetrahydro-2H-pyran-4-yl)-1,3,5-triazine-2,4(1H,3H)-dione, add 0.05 mL of dichloromethane and stir at room temperature until dissolved. Add 0.35 mL of acetonitrile and stir. Slurry at room temperature for 3 days. After centrifugation, dry the solid under vacuum to obtain the product. X-ray powder diffraction analysis showed that the product is in crystal form B.

[0191] Example 23. Preparation of the crystal form of compound B

[0192] Weigh 5 mg of (S)-6-((1-(2-fluoro-5-methylphenyl)ethyl)amino)-3-(tetrahydro-2H-pyran-4-yl)-1,3,5-triazine-2,4(1H,3H)-dione, add 0.05 mL of dichloromethane and stir at room temperature until dissolved. Add 0.35 mL of methyl isobutyl ketone and stir. Slurry at room temperature for 3 days. After centrifugation, dry the solid under vacuum to obtain the product. X-ray powder diffraction analysis showed that the product is in crystal form B.

[0193] Example 24. Preparation of the C crystal form of compound

[0194] Weigh 5 mg of (S)-6-((1-(2-fluoro-5-methylphenyl)ethyl)amino)-3-(tetrahydro-2H-pyran-4-yl)-1,3,5-triazine-2,4(1H,3H)-dione, add 0.05 mL of dichloromethane and stir at room temperature until dissolved. Add 0.35 mL of isopropyl acetate and stir to crystallize. Pulp at room temperature for 3 days, centrifuge, and then dry the solid under vacuum to obtain the title product. X-ray powder diffraction analysis identified the product as C-type. The XRD pattern is shown below. Figure 4 The positions of its characteristic peaks are shown in Table 6. The DSC spectrum shows endothermic peaks at 86.45℃ and 255.56℃, and exothermic peaks at 194.44℃. The TGA spectrum shows a weight loss of 5.17% from 30 to 140℃.

[0195] Table 6. Peak positions of C crystal form of compounds

[0196]

[0197]

[0198] Example 25. Preparation of the D crystal form of compound

[0199] Weigh 5 mg of (S)-6-((1-(2-fluoro-5-methylphenyl)ethyl)amino)-3-(tetrahydro-2H-pyran-4-yl)-1,3,5-triazine-2,4(1H,3H)-dione, add 0.05 mL of dichloromethane and stir at room temperature until dissolved. Add 0.35 mL of methyl tert-butyl ether and stir to crystallize. Pulp at room temperature for 3 days, centrifuge, and then dry the solid under vacuum to obtain the product. X-ray powder diffraction analysis identified the product as crystal form D. The XRD pattern is shown below. Figure 4 The positions of its characteristic peaks are shown in Table 7. The DSC spectrum shows endothermic peaks at 96.46℃ and 256.21℃, and exothermic peaks at 160.48℃. The TGA spectrum shows a weight loss of 9.49% from 30 to 140℃.

[0200] Table 7 Peak positions of compound D crystal form

[0201]

[0202] Example 26. Preparation of the E crystal form of compound

[0203] Weigh 30 mg of (S)-6-((1-(2-fluoro-5-methylphenyl)ethyl)amino)-3-(tetrahydro-2H-pyran-4-yl)-1,3,5-triazine-2,4(1H,3H)-dione, add 0.15 mL of dichloromethane and stir at room temperature until dissolved. Add 3.15 mL of n-heptane and stir to crystallize. Pulp at room temperature for 2 days, centrifuge, and dry the solid under vacuum to obtain the product. X-ray powder diffraction analysis identified the product as crystal form E. The XRD pattern is shown below. Figure 5 The positions of its characteristic peaks are shown in Table 8. The DSC spectrum shows endothermic peaks at 104.97℃, 128.14℃, 205.60℃, 231.49℃, and 254.86℃, and exothermic peaks at 184.66℃. The TGA spectrum shows a weight loss of 0.86% from 30 to 100℃.

[0204] Table 8 Peak positions of compound E crystal form

[0205]

[0206] Experimental Example 1. Stability Study of Factors Affecting the Crystal Form of Compound A

[0207] The (S)-6-((1-(2-fluoro-5-methylphenyl)ethyl)amino)-3-(tetrahydro-2H-pyran-4-yl)-1,3,5-triazine-2,4(1H,3H)-dione A crystal form was laid flat in an open container to investigate the stability of the sample under high temperature (40℃, 60℃), light, and high humidity (RH75%, RH92.5%) conditions. The sampling period was 30 days.

[0208] Table 9 Factors affecting the stability of compound A crystal form

[0209]

[0210]

[0211] Conclusion: The influencing factor experiment shows that the free A crystal form has good physical and chemical stability under light, high temperature (40℃, 60℃) and high humidity (75%RH, 92.5%RH) conditions for 30 days.

[0212] Experimental Example 2. Long-term / accelerated stability study of compound A crystal form

[0213] The long-term accelerated stability of (S)-6-((1-(2-fluoro-5-methylphenyl)ethyl)amino)-3-(tetrahydro-2H-pyran-4-yl)-1,3,5-triazine-2,4(1H,3H)-dione A crystal form was investigated under long-term (25°C, 60% RH) and accelerated (40°C, 75% RH) conditions for 6 months.

[0214] Table 10 Long-term accelerated stability of compound A crystal form

[0215]

[0216] Conclusion: Long-term accelerated experiments show that the free A crystal form exhibits good physicochemical stability after 6 months at 25℃ and 60%RH and 40℃ and 75%RH.

[0217] Experiment Example 3. Experiment on Factors Affecting the Crystal Form of Compound B

[0218] The (S)-6-((1-(2-fluoro-5-methylphenyl)ethyl)amino)-3-(tetrahydro-2H-pyran-4-yl)-1,3,5-triazine-2,4(1H,3H)-dione B crystal form was laid flat in an open container to investigate the stability of the sample under high temperature (40℃, 60℃), light, and high humidity (RH75%, RH90%) conditions. The sampling period was 30 days.

[0219] Table 11 Factors affecting the stability of crystal form of compound B

[0220]

[0221] Conclusion: The influencing factor experiment showed that the free B crystal form had good physicochemical stability under light, high temperature (40℃, 60℃) and high humidity (75%RH, 92.5%RH) conditions for 30 days.

[0222] Experimental Example 4. Long-term / accelerated stability test of compound B crystal form

[0223] The long-term accelerated stability of (S)-6-((1-(2-fluoro-5-methylphenyl)ethyl)amino)-3-(tetrahydro-2H-pyran-4-yl)-1,3,5-triazine-2,4(1H,3H)-dione B crystal form was investigated under long-term (25℃, 60%RH) and accelerated (40℃, 75%RH) conditions for 6 months.

[0224] Table 12 Results of long-term accelerated stability experiments of compound B crystal form

[0225]

[0226] Conclusion: Long-term accelerated experiments show that the free B crystal form exhibits good physicochemical stability under conditions of 25℃ and 60%RH and 40℃ and 75%RH for 6 months.

Claims

1. An X-ray powder diffraction pattern of (S)-6-((1-(2-fluoro-5-methylphenyl)ethyl)amino)-3-(tetrahydro-2H-pyran-4-yl)-1,3,5-triazine-2,4(1H,3H)-dione in crystal form A, with characteristic peaks at 9.2, 10.9, 18.6, 20.1, 22.0 and 26.9 degrees Celsius.

2. The X-ray powder diffraction pattern of (S)-6-((1-(2-fluoro-5-methylphenyl)ethyl)amino)-3-(tetrahydro-2H-pyran-4-yl)-1,3,5-triazine-2,4(1H,3H)-dione, as described in claim 1, exhibits characteristic peaks at 9.2, 10.9, 13.3, 16.9, 18.6, 19.2, 20.1, 22.0, 26.9, and 35.6 degrees Celsius.

3. The X-ray powder diffraction pattern of (S)-6-((1-(2-fluoro-5-methylphenyl)ethyl)amino)-3-(tetrahydro-2H-pyran-4-yl)-1,3,5-triazine-2,4(1H,3H)-dione, as described in claim 1, exhibits characteristic peaks at 9.2, 10.9, 13.3, 16.9, 18.6, 19.2, 20.1, 20.6, 22.0, 26.9, 28.0, and 35.6, expressed as a diffraction angle 2θ.

4. The X-ray powder diffraction pattern of (S)-6-((1-(2-fluoro-5-methylphenyl)ethyl)amino)-3-(tetrahydro-2H-pyran-4-yl)-1,3,5-triazine-2,4(1H,3H)-dione, as described in claim 1, is shown in Figure 2.

5. An X-ray powder diffraction pattern of (S)-6-((1-(2-fluoro-5-methylphenyl)ethyl)amino)-3-(tetrahydro-2H-pyran-4-yl)-1,3,5-triazine-2,4(1H,3H)-dione in crystal form B, with characteristic peaks at 11.1, 15.9, 18.8, 20.4, 22.2 and 26.8 degrees Celsius.

6. The X-ray powder diffraction pattern of (S)-6-((1-(2-fluoro-5-methylphenyl)ethyl)amino)-3-(tetrahydro-2H-pyran-4-yl)-1,3,5-triazine-2,4(1H,3H)-dione, as described in claim 5, has characteristic peaks at 6.6, 9.5, 11.1, 15.9, 18.8, 20.4, 21.0, 22.2, 24.6, and 26.8, expressed as a diffraction angle 2θ.

7. The X-ray powder diffraction pattern of (S)-6-((1-(2-fluoro-5-methylphenyl)ethyl)amino)-3-(tetrahydro-2H-pyran-4-yl)-1,3,5-triazine-2,4(1H,3H)-dione, as described in claim 5, has characteristic peaks at 6.6, 9.5, 11.1, 13.3, 14.0, 15.9, 18.8, 19.0, 19.8, 20.4, 21.0, 22.2, 26.8, and 28.

0.

8. The X-ray powder diffraction pattern of the (S)-6-((1-(2-fluoro-5-methylphenyl)ethyl)amino)-3-(tetrahydro-2H-pyran-4-yl)-1,3,5-triazine-2,4(1H,3H)-dione, as described in claim 5, expressed in terms of diffraction angle 2θ, is shown in Figure 3.

9. The C-crystal form of (S)-6-((1-(2-fluoro-5-methylphenyl)ethyl)amino)-3-(tetrahydro-2H-pyran-4-yl)-1,3,5-triazine-2,4(1H,3H)-dione, with characteristic peaks at 8.3, 14.4, 16.8, 17.8, 19.3, 22.3 and 23.1, as expressed in diffraction angle 2θ.

10. The C-crystal form of (S)-6-((1-(2-fluoro-5-methylphenyl)ethyl)amino)-3-(tetrahydro-2H-pyran-4-yl)-1,3,5-triazine-2,4(1H,3H)-dione according to claim 9, and its X-ray powder diffraction pattern expressed in terms of diffraction angle 2θ, is shown in Figure 4.

11. An X-ray powder diffraction pattern of (S)-6-((1-(2-fluoro-5-methylphenyl)ethyl)amino)-3-(tetrahydro-2H-pyran-4-yl)-1,3,5-triazine-2,4(1H,3H)-dione in its D crystal form, expressed as a diffraction angle 2θ, with characteristic peaks at 7.8, 8.7, 9.7, 13.5, 14.0 and 22.

9.

12. The X-ray powder diffraction pattern of the (S)-6-((1-(2-fluoro-5-methylphenyl)ethyl)amino)-3-(tetrahydro-2H-pyran-4-yl)-1,3,5-triazine-2,4(1H,3H)-dione in crystal form D according to claim 11, expressed as a diffraction angle 2θ, has characteristic peaks at 7.8, 8.7, 9.7, 13.5, 14.0, 16.7, 19.6, 20.2 and 22.

9.

13. The X-ray powder diffraction pattern of the (S)-6-((1-(2-fluoro-5-methylphenyl)ethyl)amino)-3-(tetrahydro-2H-pyran-4-yl)-1,3,5-triazine-2,4(1H,3H)-dione in crystal form D according to claim 11, expressed as a diffraction angle 2θ, has characteristic peaks at 7.8, 8.7, 9.7, 13.5, 14.0, 15.5, 16.7, 17.5, 19.6, 20.2, 22.9, and 23.

9.

14. The X-ray powder diffraction pattern of the (S)-6-((1-(2-fluoro-5-methylphenyl)ethyl)amino)-3-(tetrahydro-2H-pyran-4-yl)-1,3,5-triazine-2,4(1H,3H)-dione in the D crystal form according to claim 11, expressed in terms of diffraction angle 2θ, is shown in Figure 5.

15. An X-ray powder diffraction pattern of (S)-6-((1-(2-fluoro-5-methylphenyl)ethyl)amino)-3-(tetrahydro-2H-pyran-4-yl)-1,3,5-triazine-2,4(1H,3H)-dione in its E crystal form, expressed as a diffraction angle 2θ, with characteristic peaks at 6.9, 9.3, 14.2, 17.1, 18.0 and 25.

0.

16. The X-ray powder diffraction pattern of the (S)-6-((1-(2-fluoro-5-methylphenyl)ethyl)amino)-3-(tetrahydro-2H-pyran-4-yl)-1,3,5-triazine-2,4(1H,3H)-dione according to claim 15, expressed as a diffraction angle 2θ, has characteristic peaks at 6.9, 9.3, 13.5, 14.2, 17.1, 18.0, 22.4, 23.2 and 25.

0.

17. The X-ray powder diffraction pattern of the (S)-6-((1-(2-fluoro-5-methylphenyl)ethyl)amino)-3-(tetrahydro-2H-pyran-4-yl)-1,3,5-triazine-2,4(1H,3H)-dione according to claim 15, expressed as a diffraction angle 2θ, has characteristic peaks at 6.9, 9.3, 13.5, 14.2, 16.1, 17.1, 18.0, 21.1, 22.4, 23.2, 23.8, and 25.

0.

18. The X-ray powder diffraction pattern of the (S)-6-((1-(2-fluoro-5-methylphenyl)ethyl)amino)-3-(tetrahydro-2H-pyran-4-yl)-1,3,5-triazine-2,4(1H,3H)-dione in crystal form E according to claim 15, expressed in terms of diffraction angle 2θ, is shown in Figure 6.

19. A crystalline form of (S)-6-((1-(2-fluoro-5-methylphenyl)ethyl)amino)-3-(tetrahydro-2H-pyran-4-yl)-1,3,5-triazine-2,4(1H,3H)-dione according to any one of claims 1-18, wherein, The error range of the 2θ angle is ±0.

2.

20. A method for preparing the A-type crystal form according to any one of claims 1-4, selected from any of the following methods, Method 1: (a) The compound (S)-6-((1-(2-fluoro-5-methylphenyl)ethyl)amino)-3-(tetrahydro-2H-pyran-4-yl)-1,3,5-triazine-2,4(1H,3H)-dione is mixed with solvent (1) and dissolved by stirring or heating. Solvent (1) is selected from at least one of 7% water / ethanol, tetrahydrofuran, 10% water / isopropanol, 10% water / acetone, methanol, ethanol, and 10% water / methanol. (b) Crystallization; Alternatively, method two: (a) The compound (S)-6-((1-(2-fluoro-5-methylphenyl)ethyl)amino)-3-(tetrahydro-2H-pyran-4-yl)-1,3,5-triazine-2,4(1H,3H)-dione was mixed with solvent (2) and dissolved by stirring or heating. Solvent (2) was selected from 10% water / acetone, dichloromethane, (b) Add a second solvent (3) to crystallize, wherein the solvent (3) is selected from at least one of water, methanol and ethanol; Alternatively, method three: (a) The compound (S)-6-((1-(2-fluoro-5-methylphenyl)ethyl)amino)-3-(tetrahydro-2H-pyran-4-yl)-1,3,5-triazine-2,4(1H,3H)-dione is mixed with solvent (4), wherein solvent (4) is selected from at least one of water, methanol, ethanol, isopropanol, acetone, ethyl acetate, acetonitrile, isopropyl acetate, methyl tert-butyl ether, methyl isobutyl ketone, n-heptane, cyclohexane, 1,4-dioxane, 10% water / methanol, and 7% water / ethanol. (b) Stir and beat the mixture to crystallize.

21. A pharmaceutical composition comprising the following components: i) The crystal form of (S)-6-((1-(2-fluoro-5-methylphenyl)ethyl)amino)-3-(tetrahydro-2H-pyran-4-yl)-1,3,5-triazine-2,4(1H,3H)-dione according to any one of claims 1-18; and ii) One or more pharmaceutically acceptable excipients.

22. A method for preparing a pharmaceutical composition, comprising the step of mixing the crystal form of (S)-6-((1-(2-fluoro-5-methylphenyl)ethyl)amino)-3-(tetrahydro-2H-pyran-4-yl)-1,3,5-triazine-2,4(1H,3H)-dione according to any one of claims 1-18 with a pharmaceutically acceptable excipient.

23. Use of the crystal form of (S)-6-((1-(2-fluoro-5-methylphenyl)ethyl)amino)-3-(tetrahydro-2H-pyran-4-yl)-1,3,5-triazine-2,4(1H,3H)-dione according to any one of claims 1-18, or the composition according to claim 21, in the preparation of a medicament for treating diseases or conditions related to myosin regulation.

24. The use of the (S)-6-((1-(2-fluoro-5-methylphenyl)ethyl)amino)-3-(tetrahydro-2H-pyran-4-yl)-1,3,5-triazine-2,4(1H,3H)-dione crystal form according to any one of claims 1-18, or the use of the composition according to claim 21 in the preparation of a medicament for treating a disease or condition selected from diastolic heart failure with preserved ejection fraction, heart failure with normal ejection fraction (HFpEF), heart failure with intermediate ejection fraction (HFmREF), ischemic heart disease, restrictive cardiomyopathy, and hypertrophic cardiomyopathy (HCM).

25. The use according to claim 24, wherein the hypertrophic cardiomyopathy (HCM) is selected from non-obstructive hypertrophic cardiomyopathy (nHCM) and obstructive hypertrophic cardiomyopathy (oHCM).

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