Salt of rho-associated protein kinase inhibitor, solid form of the salt, preparation method for same, and uses thereof

Abstract

To provide a salt of (6-(4-((4-(1H-pyrazol-4-yl)phenyl)amino)pyrimidin-2-yl)-1-methyl-1H-indol-2-yl)(3,3-difluoroazetidine-1-yl)methanone, which is a Rho-associated protein kinase (ROCK) inhibitor.SOLUTION: The present invention provides a salt of (6-(4-((4-(1H-pyrazol-4-yl)phenyl)amino)pyrimidin-2-yl)-1-methyl-1H-indol-2-yl)(3,3-difluoroazetidine-1-yl)methanone represented by the following formula (hereinafter referred to as "compound A"), a solid form of the salt, a method for preparing the solid form, and uses of a pharmaceutical composition comprising the solid form.SELECTED DRAWING: None

Description

Detailed Description of the Invention

[0001] [Field of the Invention] The present invention relates to a salt of (6-(4-((4-(1H-pyrazol-4-yl)phenyl)amino)pyrimidin-2-yl)-1-methyl-1H-indol-2-yl)(3,3-difluoroazetidin-1-yl)methanone (hereinafter referred to as "Compound A"), a solid form of the salt, a process for preparing the solid form, a pharmaceutical composition comprising the solid form, and the use of the solid form as a Rho-associated protein kinase (ROCK) inhibitor, preferably a selective ROCK2 inhibitor.

[0002] [Background of the invention] Rho-associated protein kinase (ROCK) is a serine / threonine kinase from the AGC kinase family, and includes two isoforms, ROCK1 and ROCK2. ROCK1 and ROCK2 are differentially expressed and regulated in specific tissues. For example, ROCK1 is ubiquitously expressed at relatively high levels, whereas ROCK2 is preferentially expressed in the heart, brain, and skeletal muscle. ROCK was the first downstream effector of Rho proteins discovered, and its biological function is achieved by phosphorylating downstream effector proteins (e.g., MLC, Lin-11, Isl-1, LIMK, ERM, MARCKS, CRMP-2, etc.). Studies have shown that various diseases (e.g., pulmonary fibrosis, cardiovascular disease, neurological diseases, and cancer) are associated with ROCK-mediated pathways. Thus, ROCK is considered an important target for the development of novel drugs.

[0003] Applicant has discovered that (6-(4-((4-(1H-pyrazol-4-yl)phenyl)amino)pyrimidin-2-yl)-1-methyl-1H-indol-2-yl)(3,3-difluoroazetidin-1-yl)methanone can be used as a potent Rho-associated protein kinase (ROCK) inhibitor (see PCT / CN2018 / 093713, which is incorporated herein by reference in its entirety), although the compound and its solid form salts have not yet been reported.

[0004] [Summary of the Invention] In one aspect, the present invention provides compound A ((6-(4-((4-(1H-pyrazol-4-yl)phenyl)amino)pyrimidin-2-yl)-1-methyl-1H-indol-2-yl)(3,3-difluoroazetidin-1-yl)methanone) shown below: [ka] of salt.

[0005] In another aspect, the present invention provides crystalline forms of salts of Compound A and solvates thereof.

[0006] The preferred crystalline forms of the present invention not only have excellent effects in the prevention or treatment of diseases mediated by Rho-associated protein kinase (ROCK), but also have other advantages. For example, the preferred crystalline forms of the present invention have excellent physical properties (including solubility, dissolution rate, light resistance, low hygroscopicity, heat resistance, high moisture resistance, flowability, etc.), and may have excellent properties in terms of bioavailability, physical and / or chemical stability, and ease of preparation. The preferred crystalline forms of the present invention have good powder properties, are more suitable and convenient for mass production and formulation, reduce irritation, improve absorption, solve metabolic rate problems, significantly reduce toxicity resulting from drug accumulation, improve safety, and effectively ensure the quality and efficacy of pharmaceutical products.

[0007] In another aspect, the present invention provides methods for preparing the crystalline forms of the present invention.

[0008] In another aspect, the present invention provides pharmaceutical compositions comprising any one or more of the crystalline forms of the present invention and one or more pharmaceutically acceptable carriers.

[0009] In another aspect, the present invention provides the use of a crystalline form of the present invention in the manufacture of a medicament as a Rho-associated protein kinase (ROCK) inhibitor, preferably a selective ROCK2 inhibitor. [Brief explanation of the drawings]

[0010] [Figure 1] 1 is an X-ray powder diffraction (XRPD) pattern of crystalline Form I of Compound A monohydrochloride monohydrate. [Figure 2] 1 is a differential scanning calorimetry (DSC) graph of crystalline Form I of Compound A monohydrochloride monohydrate. [Figure 3] 1 is a thermogravimetric analysis (TGA) graph of crystalline Form I of Compound A monohydrochloride monohydrate. [Figure 4] 1 is a scanning electron microscope image of crystalline Form I of Compound A monohydrochloride monohydrate. [Figure 5] 1 is an XRPD pattern of crystalline Form II of Compound A monohydrochloride dihydrate. [Figure 6] 1 is a DSC-TGA graph of crystalline Form II of Compound A monohydrochloride dihydrate. [Figure 7] 1 is a scanning electron microscope image of crystalline Form II of Compound A monohydrochloride dihydrate. [Figure 8] 1 is an XRPD pattern of crystalline Form III of Compound A monohydrochloride sesquihydrate. [Figure 9] 1 is a DSC-TGA pattern of crystalline Form III of Compound A monohydrochloride sesquihydrate. [Figure 10] 1 is a scanning electron microscope image of crystalline Form III of Compound A monohydrochloride sesquihydrate. [Figure 11]1 is an XRPD pattern of crystalline Form IV of Compound A monohydrochloride mono-dimethyl sulfoxide complex. [Figure 12] 1 is a DSC graph of crystalline Form IV of Compound A monohydrochloride mono-dimethyl sulfoxide complex. [Figure 13] 1 is a TGA graph of crystalline Form IV of Compound A monohydrochloride mono-dimethyl sulfoxide complex. [Figure 14] 1 is a scanning electron microscope image of crystalline Form IV of Compound A monohydrochloride mono-dimethyl sulfoxide complex. [Figure 15] 1 shows an XRPD pattern of a sample obtained in the solid stability test of Experimental Example 2. [Figure 16] 1 shows XRPD patterns of a sample before and after a DVS test in Experimental Example 4. [Figure 17] 1 shows XRPD patterns of a sample before and after a crystal form transformation test in water in Experimental Example 5. [Figure 18] 1 shows an XRPD pattern of a sample obtained in the crushing test of Experimental Example 6. [Figure 19] 1 is an XRPD spectrum of a sample obtained in the solid stability test of Experimental Example 7. [Figure 20] 1 is a TGA graph of a sample obtained in the solid stability test of Experimental Example 7.

[0011] [Detailed Description of the Invention] definition Unless otherwise defined in the context, all technical and scientific terms used herein are intended to have the same meaning as commonly understood by those skilled in the art. Reference to technology used herein is intended to refer to the technology commonly understood in the art, including the modifications of those technologies or equivalent replacements that would be obvious to those skilled in the art. Although most of the following terms are believed to be easily understood by those skilled in the art, the following definitions are nevertheless provided to better explain the present invention.

[0012] The terms "contain," "include," "comprise," "have," or "relate to," and other variations as used herein, are inclusive or open-ended and do not exclude additional, unrecited elements or method steps.

[0013] The word "about" as used herein refers to a range within an acceptable standard error of a value, such as ±0.05, ±0.1, ±0.2, ±0.3, ±1, ±2, or ±3, as would be understood by one of ordinary skill in the art.

[0014] As used herein, the term "solid form" includes all solid forms, eg, crystalline or amorphous forms, of Compound A or any solvates thereof.

[0015] As used herein, the term "amorphous" refers to any solid material that lacks order in three dimensions. In some instances, amorphous solids can be characterized by known techniques, including XRPD crystallography, solid-state nuclear magnetic resonance (ssNMR) spectroscopy, DSC, or some combination of these techniques. As shown below, amorphous solids typically give diffuse XRPD patterns composed of one or two broad peaks (i.e., peaks with a base width of about 5° 2θ or greater).

[0016] As used herein, the term "crystalline form" or "crystal" refers to any solid material that exhibits three-dimensional order, giving a characteristic XRPD pattern with well-defined peaks, in contrast to amorphous solid materials.

[0017] As used herein, the term "X-ray powder diffraction pattern (XRPD pattern)" refers to an experimentally observed diffractogram or parameters derived therefrom. XRPD patterns are typically characterized by peak positions (abscissa) and peak intensities (ordinate).

[0018] The term "2θ" as used herein refers to the peak position in degrees based on the experimental setup of an X-ray diffraction experiment, which is the common abscissa unit of a diffraction pattern. The experimental setup requires that the reflected beam be recorded at an angle 2θ (2θ) when the reflection is diffracted when the incident beam forms an angle theta (θ) with a specific lattice plane. It should be understood that references herein to specific 2θ values ​​of a particular solid form are intended to mean the 2θ values ​​(degrees) measured using the X-ray diffraction experimental conditions described herein. For example, as described herein, Cu-Kα (Kα1 (Å): 1.540598 and Kα2 (Å): 1.544426 Å) was used as the radiation source.

[0019] As used herein, "I %" refers to the percentage of peak intensity.

[0020] As used herein, the term "differential scanning calorimetry (DSC) graph" refers to a curve recorded on a differential scanning calorimeter.

[0021] As used herein, the term "thermogravimetric analysis (TGA) graph" refers to a curve recorded on a thermogravimetric analyzer.

[0022] As used herein, the term "essentially the same" in relation to X-ray diffraction peak positions means that typical peak position and intensity variations are taken into account. For example, those skilled in the art will understand that peak positions (2θ) will exhibit some variation, typically on the order of 0.1 to 0.2 degrees, depending not only on the instrument used to measure the diffraction. Furthermore, those skilled in the art will understand that relative peak intensities will exhibit variability due not only to instrument-to-instrument variability but also to crystallinity, preferred orientation, sample surface preparation, and other factors known to those skilled in the art. Similarly, as used herein, "essentially the same" in relation to DSC graphs is intended to encompass the variations associated with these analytical techniques known to those skilled in the art. For example, differential scanning calorimetry graphs typically have a variability of up to ±0.2°C for well-defined peaks and even greater variability for broad lines (e.g., up to ±1°C).

[0023] Liquid crystal nuclear magnetic resonance spectra in this application are preferably collected on a Bruker Advance 300 nuclear magnetic resonance spectrometer using DMSO-d6 as the solvent unless otherwise specified.

[0024] The polarized light microscopy data in this application are preferably collected with a polarized light microscope ECLIPSE LV100POL (Nikon, Japan).

[0025] Numeric ranges used herein (e.g., "1 to 10," "1 to 6," "2 to 10," "2 to 6," "3 to 10," "5 to 10," "3 to 6"), etc., include any point within the numerical range (e.g., 1, 2, 3, 4, 5, 6, 7, 8, 9, or 10).

[0026] The prepared salt or its crystalline form can be recovered by methods including decantation, centrifugation, evaporation, gravity filtration, suction filtration, or any other technique for recovering solids under pressure or reduced pressure. The recovered solids can optionally be dried. "Drying" in this invention is carried out under reduced pressure (preferably under vacuum) until the residual solvent content is reduced to within the limits set forth in the International Conference on Harmonization (ICH) guidelines for the Registration of Pharmaceuticals for Human Use. The residual solvent content, depending on the type of solvent, does not exceed about 5000 ppm, or preferably about 4000 ppm, or more preferably about 3000 ppm. Drying can be carried out in a tray dryer, vacuum oven, air oven, cone vacuum dryer, rotary vacuum dryer, fluidized bed dryer, spin flash dryer, flash dryer, etc. Drying can be carried out at atmospheric or reduced pressure (preferably in a vacuum) for any desired period of time (e.g., about 1, 2, 3, 5, 10, 15, 20, 24 hours, or overnight) at temperatures below about 100°C, below about 80°C, below about 60°C, below about 50°C, below about 30°C, or any other suitable temperature, as long as the quality of the salt is not compromised. Drying can be carried out for any desired time until the desired product quality is achieved. The dried product may optionally be subjected to a size reduction procedure to produce the desired particle size. Grinding or micronization may be carried out before drying the product or after drying is complete. Techniques that can be used to reduce particle size include, but are not limited to, ball mills, roller mills, and hammer mills, as well as jet mills.

[0027] As used herein, the term "anhydrous" preferably refers to a crystalline form that does not include water molecules as structural elements.

[0028] Salt of Compound A, its crystalline form and its preparation method In some embodiments, the present invention provides compound A, which is an inorganic or organic acid salt. [ka] of salt.

[0029] In a preferred embodiment, the present invention provides a salt of Compound A that is Compound A hydrochloride; Preferably, Compound A hydrochloride is Compound A monohydrochloride; Preferably, Compound A hydrochloride is crystalline form I of Compound A monohydrochloride monohydrate; Crystalline Form I has an XRPD pattern containing characteristic peaks at diffraction angles (2θ) of about 8.3±0.2°, 12.6±0.2°, and 18.5±0.2°; Preferably, an XRPD pattern comprising characteristic peaks at diffraction angles (2θ) of about 8.3±0.2°, 12.6±0.2°, 16.5±0.2°, 17.3±0.2°, 17.8±0.2°, 18.5±0.2°, 24.4±0.2°, 26.0±0.2° and 29.1±0.2°; Most preferably, the compound has an XRPD pattern comprising characteristic peaks at diffraction angles (2θ) of about 8.3±0.2°, 11.2±0.2°, 12.6±0.2°, 16.5±0.2°, 17.3±0.2°, 17.8±0.2°, 18.5±0.2°, 19.5±0.2°, 19.8±0.2°, 20.6±0.2°, 21.0±0.2°, 21.7±0.2°, 23.7±0.2°, 24.4±0.2°, 25.5±0.2°, 26.0±0.2°, 29.1±0.2°, and 33.6±0.2°.

[0030] In a more preferred embodiment, crystalline Form I has an XRPD pattern comprising peaks at the following diffraction angles (2θ): [Table 1]

[0031] In a more preferred embodiment, crystalline Form I has an XRPD pattern comprising peaks at essentially the same diffraction angles (2θ) as shown in Figure 1. In a most preferred embodiment, the XRPD pattern of crystalline Form I is essentially the same as that shown in Figure 1.

[0032] In a more preferred embodiment, crystalline Form I has a DSC graph containing endothermic peaks at about 105°C, about 146°C, and about 245°C.

[0033] In a more preferred embodiment, crystalline Form I has a DSC graph containing essentially the same characteristic peaks as shown in Figure 2. In a most preferred embodiment, crystalline Form I has a DSC graph essentially the same as shown in Figure 2.

[0034] In a more preferred embodiment, crystalline Form I has a weight loss of about 3.4% when heated to about 169°C in thermogravimetric analysis.

[0035] The DSC graph of crystalline form I has two consecutive endothermic peaks between 75 and 169°C, and the corresponding TGA graph shows a weight loss of about 3.4% before 169°C, indicating that crystalline form I begins to lose water when the temperature exceeds 75°C.

[0036] In a most preferred embodiment, crystalline Form I has a TGA graph essentially similar to that shown in FIG.

[0037] In a most preferred embodiment, crystalline Form I has a scanning electron microscope image essentially similar to that shown in FIG.

[0038] In some embodiments, the present invention provides a method for preparing crystalline Form I, comprising the steps of adding a ketone solvent (preferably a ketone having 3 to 6 carbon atoms, including, but not limited to, acetone, butanone, methyl ethyl ketone, methyl isobutyl ketone, and diethyl ketone) to Compound A, stirring under heating (preferably to about 40 to 80°C, e.g., about 50°C or about 60°C), and then adding water (the volume ratio of the ketone solvent to water is preferably about (1 to 15):1, preferably about 10:1) to completely dissolve Compound A; and then adding hydrochloric acid (the concentration of hydrochloric acid is preferably 2 to 15 mol / L, preferably 12 mol / L (i.e., concentrated hydrochloric acid), and the molar ratio of Compound A to HCl is 1:(1 to 1.3)), stirring under heating (preferably to about 40 to 80°C, e.g., about 50°C or 60°C), filtering, and optionally drying to obtain the crystalline form.

[0039] In a preferred embodiment, Compound A hydrochloride is crystalline Form II of Compound A monohydrochloride dihydrate; Crystalline Form II has an XRPD pattern containing characteristic peaks at diffraction angles (2θ) of about 9.9±0.2°, 13.2±0.2°, and 16.2±0.2°; Preferably, an XRPD pattern comprising characteristic peaks at diffraction angles (2θ) of about 9.9±0.2°, 11.0±0.2°, 13.2±0.2°, 13.5±0.2°, 13.8±0.2°, 16.2±0.2°, 19.4±0.2° and 25.2±0.2°; Most preferably, the compound has an XRPD pattern comprising characteristic peaks at diffraction angles (2θ) of about 6.5±0.2°, 9.9±0.2°, 11.0±0.2°, 11.6±0.2°, 11.8±0.2°, 13.2±0.2°, 13.5±0.2°, 13.8±0.2°, 16.2±0.2°, 16.6±0.2°, 17.1±0.2°, 18.2±0.2°, 19.4±0.2°, 20.0±0.2°, 20.3±0.2°, 20.9±0.2°, 22.6±0.2°, 24.9±0.2°, 25.2±0.2°, and 25.9±0.2°.

[0040] In a more preferred embodiment, crystalline Form II has an XRPD pattern comprising peaks at the following diffraction angles (2θ): [Table 2]

[0041] In a more preferred embodiment, crystalline Form II has an XRPD pattern comprising peaks at diffraction angles (2θ) essentially the same as those shown in Figure 5. In a most preferred embodiment, the XRPD pattern of crystalline Form II is essentially the same as that shown in Figure 5.

[0042] In a more preferred embodiment, crystalline Form II has a DSC graph comprising a broad endothermic peak between about 25-180°C and an endothermic peak between about 217-243°C.

[0043] In a more preferred embodiment, crystalline Form II has a weight loss of about 7.1% when heated to about 180°C in thermogravimetric analysis.

[0044] The DSC graph of crystalline form II has a broad endothermic peak between 25 and 180°C, and the corresponding TGA graph shows a weight loss of about 7.1% before 180°C, indicating that crystalline form II begins to lose water when the temperature exceeds 25°C.

[0045] In a most preferred embodiment, crystalline Form II has a DSC-TGA graph essentially similar to that shown in FIG.

[0046] In a most preferred embodiment, crystalline Form II has a scanning electron microscope image essentially similar to that shown in FIG.

[0047] In some embodiments, the present invention provides a method for preparing crystalline Form II of Compound A monohydrochloride dihydrate, comprising adding crystalline Form I of Compound A monohydrochloride monohydrate to an aqueous alcoholic solvent having 1 to 10 carbon atoms (the alcoholic solvent is preferably an alcohol having 1 to 6 carbon atoms, including, but not limited to, methanol, ethanol, 1-propanol (n-propanol), 2-propanol (isopropanol), 1-butanol, 2-butanol, and tert-butanol, wherein the weight-to-volume ratio (g / mL) of crystalline Form I to the alcoholic solvent is about 1:(1-100), preferably 1:50), stirring the solution at a temperature of about 10-40°C (e.g., about 20-30°C) for about 10 to 60 hours (e.g., about 50 hours), filtering, and optionally drying to obtain the crystalline form.

[0048] In a preferred embodiment, Compound A hydrochloride is Compound A monohydrochloride sesquihydrate crystalline Form III; Crystalline Form III has an XRPD pattern containing characteristic peaks at diffraction angles (2θ) of about 5.1±0.2°, 9.1±0.2°, and 10.2±0.2°; Preferably, an XRPD pattern comprising characteristic peaks at diffraction angles (2θ) of about 5.1±0.2°, 5.4±0.2°, 6.0±0.2°, 9.1±0.2°, 10.2±0.2°, 11.2±0.2° and 18.0±0.2°; Most preferably, the angle is about 5.1±0.2°, 5.4±0.2°, 6.0±0.2°, 6.3±0.2°, 7.5±0.2°, 7.8±0.2°, 8.4±0.2°, 9.1±0.2°, 10.2±0.2°, 10.7±0.2°, 11.2±0.2°, 12.2±0.2°, 12.5±0.2°, 12.9±0.2°, 13.7±0.2°, 14.0±0.2°, 15.0±0.2°, 16.0±0.2°, 17.0±0.2°, 18.0±0.2°, 19.0±0.2°, 20.0±0.2°, 21.0±0.2°, 22.0±0.2°, 23.0±0.2°, 24.0±0.2°, 25.0±0.2°, 26.0±0.2°, 27.0±0.2°, 28.0±0.2°, 29.0±0.2°, 30.0±0.2°, 31.0±0.2°, 32.0±0.2°, 33.0±0.2°, 34.0±0.2°, 35.0±0.2°, 36.0±0.2°, 37.0±0.2°, 38.0±0.2°, 39.0±0.2°, 40.0±0.2°, 41.0±0.2°, 42.0±0.2°, 43.0±0.2°, 44.0±0.2°, 45.0± It has an XRPD pattern containing characteristic peaks at diffraction angles (2θ) of 3.9±0.2°, 15.4±0.2°, 15.7±0.2°, 16.2±0.2°, 16.8±0.2°, 18.0±0.2°, 18.7±0.2°, 19.3±0.2°, 20.0±0.2°, 20.6±0.2°, 21.5±0.2° and 23.3±0.2°.

[0049] In a more preferred embodiment, crystalline Form III has an XRPD pattern comprising peaks at the following diffraction angles (2θ): [Table 3]

[0050] In a more preferred embodiment, crystalline Form III has an XRPD pattern comprising peaks at diffraction angles (2θ) essentially the same as those shown in Figure 8. In a most preferred embodiment, the XRPD pattern of crystalline Form III is essentially the same as that shown in Figure 8.

[0051] In a more preferred embodiment, crystalline Form III has a DSC graph containing a broad endothermic peak between about 25 and 134°C and endothermic peaks at about 137°C and about 251°C.

[0052] In a more preferred embodiment, crystalline Form III has a weight loss of about 4.3% when heated to about 180°C in thermogravimetric analysis.

[0053] The DSC graph of crystalline form III has a broad endothermic peak and a small endothermic peak between 25 and 180°C, and the corresponding TGA graph shows a weight loss of about 4.3% before 180°C, indicating that crystalline form III begins to lose water when the temperature exceeds 25°C.

[0054] In a most preferred embodiment, crystalline Form III has a DSC-TGA graph essentially similar to that shown in FIG.

[0055] In a most preferred embodiment, crystalline Form III has a scanning electron microscope image essentially similar to that shown in FIG.

[0056] In some embodiments, the present invention provides a method for dissolving crystalline Form I of Compound A monohydrochloride monohydrate in an aqueous alcoholic solvent having 1 to 10 carbon atoms (the alcoholic solvent is preferably an alcohol having 1 to 6 carbon atoms, including, but not limited to, methanol, ethanol, 1-propanol (n-propanol), 2-propanol (isopropanol), 1-butanol, 2-butanol, and tert-butanol), wherein the weight-to-volume ratio (g / mL) of crystalline Form I to the alcoholic solvent is about 1:( and adding an aromatic hydrocarbon solvent having 6 to 10 carbon atoms (e.g., toluene, the volume ratio of the alcoholic solvent to the aromatic hydrocarbon solvent is preferably about 1:(1-100), e.g., about 1:5) at a temperature of about 10 to 40°C (e.g., about 20 to 30°C), stirring the solution for about 1 to 6 hours (e.g., about 2 hours), filtering, and optionally drying to obtain the crystalline form.

[0057] In a preferred embodiment, Compound A hydrochloride is crystalline Form IV of Compound A monohydrochloride mono-dimethyl sulfoxide complex; Crystalline Form IV has an XRPD pattern containing characteristic peaks at diffraction angles (2θ) of about 6.6±0.2°, 8.9±0.2°, and 18.5±0.2°; Preferably, an XRPD pattern comprising characteristic peaks at diffraction angles (2θ) of about 6.6±0.2°, 8.9±0.2°, 11.6±0.2°, 14.6±0.2°, 17.7±0.2°, 18.5±0.2°, 24.7±0.2° and 25.0±0.2°; Most preferably, the angles are approximately 4.1±0.2°, 6.6±0.2°, 8.9±0.2°, 11.2±0.2°, 11.6±0.2°, 13.3±0.2°, 13.8±0.2°, 14.6±0.2°, 15.7±0.2°, 17.7±0.2°, 18.1±0.2°, 18.5±0.2°, 19.4±0.2°, 20.0±0.2°, and 21.0±0.2°. It has an XRPD pattern containing characteristic peaks at diffraction angles (2θ) of 7±0.2°, 21.4±0.2°, 22.0±0.2°, 22.6±0.2°, 23.0±0.2°, 23.8±0.2°, 24.7±0.2°, 25.0±0.2°, 26.5±0.2°, 27.9±0.2°, 29.7±0.2° and 32.8±0.2°.

[0058] In a more preferred embodiment, crystalline Form IV has an XRPD pattern comprising peaks at the following diffraction angles (2θ): [Table 4]

[0059] In a more preferred embodiment, crystalline Form IV has an XRPD pattern comprising peaks at diffraction angles (2θ) essentially the same as those shown in Figure 11. In a most preferred embodiment, the XRPD pattern of crystalline Form IV is essentially the same as that shown in Figure 11.

[0060] In a more preferred embodiment, crystalline Form IV has a DSC graph containing endothermic peaks at about 47°C, about 93°C, and about 210°C.

[0061] In a more preferred embodiment, crystalline Form IV has a DSC graph containing characteristic peaks essentially the same as that shown in Figure 12. In a most preferred embodiment, crystalline Form IV has a DSC graph essentially the same as that shown in Figure 12.

[0062] In a more preferred embodiment, crystalline Form IV has a weight loss of about 14.1% when heated to about 217°C in thermogravimetric analysis.

[0063] The DSC graph of crystalline form IV has an endothermic peak between 30 and 217°C, and the corresponding TGA graph shows a weight loss of about 14.1% before 217°C, indicating that crystalline form IV begins to lose solvent (DMSO) when the temperature exceeds 30°C.

[0064] In a most preferred embodiment, crystalline Form IV has a TGA graph essentially similar to that shown in FIG.

[0065] In a most preferred embodiment, crystalline Form IV has a scanning electron microscope image essentially similar to that shown in FIG.

[0066] In some embodiments, the present invention provides a method for preparing crystalline Form IV of Compound A monohydrochloride mono-dimethyl sulfoxide complex, comprising the steps of adding crystalline Form I of Compound A monohydrochloride monohydrate to dimethyl sulfoxide (the weight-to-volume ratio (g / mL) of crystalline Form I to dimethyl sulfoxide is about 1:(1-100), preferably about 1:50), and adding an aromatic hydrocarbon solvent having 6-10 carbon atoms (e.g., toluene, the volume ratio of dimethyl sulfoxide to aromatic hydrocarbon solvent is preferably about 1:(1-15), e.g., about 1:9) at a temperature of about 10-40°C (e.g., about 20-30°C), stirring the solution for about 1-6 hours (e.g., about 2 hours), filtering, and optionally drying to obtain the crystalline form.

[0067] Pharmaceutical compositions, methods of treatment and uses thereof In some embodiments, the present invention provides pharmaceutical compositions comprising a salt of Compound A, and any one or more of its crystalline forms I, II, III, and IV of the present invention, and one or more pharmaceutically acceptable carriers.

[0068] In some embodiments, the present invention provides the use of a salt of Compound A of the present invention and any one or more of its crystalline forms I, II, III and IV in the manufacture of a medicament as a Rho-associated protein kinase (ROCK) inhibitor, preferably a selective ROCK2 inhibitor.

[0069] In some embodiments, the present invention provides a salt of Compound A of the present invention and any one or more of its crystalline forms I, II, III and IV for use as a Rho-associated protein kinase (ROCK) inhibitor, preferably a selective ROCK2 inhibitor.

[0070] In some embodiments, the present invention provides a method for preventing or treating a disease mediated by Rho-associated protein kinase (ROCK), comprising administering to a subject, preferably a mammal, in need thereof a prophylactically or therapeutically effective amount of a salt of Compound A of the present invention and any one or more of its crystalline Forms I, II, III, and IV.

[0071] In some embodiments, diseases mediated by Rho-associated protein kinase (ROCK) include autoimmune disorders (including rheumatoid arthritis, systemic lupus erythematosus (SLE; lupus), psoriasis, Crohn's disease, atopic dermatitis, eczema, or graft-versus-host disease (GVHD)); cardiovascular disorders (including hypertension, atherosclerosis, restenosis, cardiac hypertrophy, cerebral ischemia, cerebral vasospasm, or erectile dysfunction); inflammation (including asthma, cardiovascular inflammation, ulcerative colitis, or renal inflammation); central nervous system disorders (including neuronal degeneration or spinal cord injury; the central nervous system disorder is preferably Huntington's disease, Parkinson's disease, Alzheimer's disease, amyotrophic lateral sclerosis (ALS), or multiple sclerosis); arterial thrombotic disorders (including platelet aggregation or leukocyte aggregation); fibrotic disorders (including liver fibrosis, pulmonary fibrosis, or renal fibrosis); neoplastic diseases (lymphoma, carcinoma (e.g., squamous cell carcinoma, small cell lung cancer, subcutaneous pituitary cancer, esophageal cancer, non-small cell lung cancer, lung adenocarcinoma, lung squamous cell carcinoma, peritoneal cancer, hepatocellular carcinoma, gastrointestinal cancer, pancreatic cancer, glioblastoma, cervical cancer, ovarian cancer, bladder cancer, liver cancer, breast cancer, colon cancer, colorectal cancer, endometrial or uterine cancer, salivary gland cancer, kidney cancer, prostate cancer, vulvar cancer, thyroid cancer, brain cancer, endometrial cancer, testicular cancer, bile duct cancer, gallbladder cancer, gastric cancer, melanoma or head and neck cancer), leukemia, astrocytoma, soft tissue sarcoma, sarcoma or blastoma; metabolic syndrome insulin resistance; hyperinsulinemia; type 2 diabetes; impaired glucose tolerance; osteoporosis; ocular disorders (including ocular hypertension, age-related macular degeneration (AMD), choroidal neovascularization (CNV), diabetic macular edema (DME), iris neovascularization, uveitis, glaucoma (including primary open-angle glaucoma, acute angle-closure glaucoma, pigmentary glaucoma, congenital glaucoma, normal-tension glaucoma, secondary glaucoma or neovascular glaucoma), or retinitis of prematurity (ROP)).

[0072] In some embodiments, diseases mediated by Rho-associated protein kinase (ROCK) include lupus nephritis, atherosclerosis, rheumatoid arthritis (RA), hemangiomas, angiofibromas, pulmonary fibrosis, psoriasis, corneal graft rejection, insulin-dependent diabetes mellitus, multiple sclerosis, myasthenia gravis, Crohn's disease, autoimmune nephritis, primary biliary cirrhosis, acute pancreatitis, allograft rejection, allergic inflammation, contact dermatitis, delayed hypersensitivity, inflammatory bowel disease, septic shock, osteoporosis, osteoarthritis, neuroinflammation, Ossia-Weber syndrome, restenosis, fungal infections, parasitic infections, and viral infections.

[0073] As used herein, the term "pharmaceutically acceptable carrier" in the present invention refers to a diluent, adjuvant, excipient, or vehicle with which a therapeutic agent is administered, which is suitable for contact with the tissues of human beings and animals without excessive toxicity, irritation, allergic response, or other problem or complication commensurate with a reasonable benefit / risk ratio, within the scope of sound medical judgment.

[0074] Pharmaceutically acceptable carriers that can be used in the pharmaceutical compositions of the present invention include, but are not limited to, sterile liquids such as water and oils, including those of petroleum, animal, vegetable, or synthetic origin, such as peanut oil, soybean oil, mineral oil, sesame oil, etc. Water is an exemplary carrier when the pharmaceutical composition is administered intravenously. Physiological saline and aqueous dextrose and glycerol solutions can also be used as liquid carriers, particularly for injectable solutions. Suitable pharmaceutical excipients include starch, glucose, lactose, sucrose, gelatin, maltose, chalk, silica gel, sodium stearate, glycerol monostearate, talc, sodium chloride, dried skim milk, glycerol, propylene glycol, water, ethanol, etc. The compositions can also contain minor amounts of wetting or emulsifying agents, or pH buffering agents, if desired. Oral formulations can include standard carriers such as pharmaceutical grades of mannitol, lactose, starch, magnesium stearate, sodium saccharin, cellulose, magnesium carbonate, etc. Examples of suitable pharmaceutical carriers are described, for example, in Remington's Pharmaceutical Sciences (1990).

[0075] The compositions of the invention can act systemically and / or locally and for this purpose can be administered by any suitable route, such as injection, intravenous, intraarterial, subcutaneous, intraperitoneal, intramuscular or transdermal administration, or in oral, buccal, nasal, transmucosal, topical, ophthalmic formulations or by inhalation.

[0076] For these administration routes, the compositions of the present invention can be administered in suitable dosage forms.

[0077] Dosage forms can be solid, semi-solid, liquid, or gaseous preparations, including, but not limited to, tablets, capsules, powders, granules, lozenges, hard candies, powders, sprays, creams, salves, suppositories, gels, pastes, lotions, ointments, aqueous suspensions, injections, suspensions, elixirs, and syrups.

[0078] The pharmaceutical compositions of the present invention may be manufactured by any process well known in the art, for example, by mixing, dissolving, granulating, dragee-making, comminuted, emulsifying, lyophilizing processes, and the like.

[0079] As used herein, the term "therapeutically effective amount" refers to that amount of the compound being administered which will relieve to some extent one or more of the symptoms of the disorder being treated.

[0080] Dosage regimen can be adjusted to provide optimal desired response.For example, a single bolus can be administered, or several divided doses can be administered over time, or the dose can be proportionally reduced or increased as indicated by the exigencies of the treatment situation.It should be noted that dosage value can vary according to the type and severity of the condition to be alleviated, and can include single or multiple administrations.It should further be understood that for any specific subject, specific dosage regimen should be adjusted over time according to individual need and the professional judgment of the person who manages or supervises the administration of composition.

[0081] The amount of the compound of the present invention administered will depend on the subject being treated, the severity of the disorder or condition, the rate of administration, the disposition of the compound, and the discretion of the prescribing physician. Generally, an effective dosage ranges from about 0.0001 to about 50 mg / kg body weight / day, e.g., about 0.01 to about 10 mg / kg / day, in single or divided doses. For a 70 kg human, this would amount to about 0.007 mg to about 3500 mg / day, e.g., about 0.7 mg to about 700 mg / day. In some instances, dosage levels below the lower end of the aforementioned range may be more than sufficient, provided that such higher doses are initially divided into several smaller doses for administration throughout the day; however, in other instances, even higher doses may be used without causing any adverse side effects.

[0082] The content or dosage of the compound of the present invention in the pharmaceutical composition is about 0.01 mg to about 1000 mg, suitably 0.1 to 500 mg, preferably 0.5 to 300 mg, more preferably 1 to 150 mg, particularly preferably 1 to 50 mg, for example, 1.5 mg, 2 mg, 4 mg, 10 mg, and 25 mg.

[0083] Unless otherwise indicated, the terms "treat" or "treatment" as used herein means to reverse, alleviate, inhibit the progression of, or prevent the disorder, condition, or disease to which such term applies, or one or more symptoms of such disorder, condition, or disease.

[0084] As used herein, the term "subject" includes a human or a non-human animal. Exemplary human subjects include human subjects (referred to as patients) with a disease (such as those described herein) or normal subjects. As used herein, the term "non-human animal" includes all vertebrates, such as non-mammals (e.g., birds, amphibians, reptiles), as well as mammals, such as non-human primates, livestock and / or domesticated animals (e.g., sheep, dogs, cats, cows, pigs, etc.). [Example]

[0085] The present invention will be described in more detail below with reference to examples, which are only used to illustrate the technical solutions of the present invention and are not intended to limit its scope. Those skilled in the art can make some non-essential improvements and adjustments, which still fall within the scope of the present invention.

[0086] Unless otherwise specified, all starting materials and reagents used in the following examples are commercially available or can be prepared by known methods.

[0087] The detection equipment and conditions used in the following examples are as follows: (1) X-ray powder diffraction (XRPD) (a) Instrument model: Bruker D8 advance with LynxEye detector Test conditions: the anode target material was copper, the light pipe was set at (40 KV 40 mA), the 2θ scan angle of the sample was 3°~40°, and the scan step was 0.02°. (b) Equipment model: Bruker D2 phasor Test conditions: the anode target material was copper, the light pipe was set at (30 KV 10 mA), the 2θ scan angle of the sample was 4°~50°, and the scan step was 0.02°.

[0088] (2) Differential scanning calorimetry (DSC) Instrument models: (a) TA Discovery DSC 250 (TA Instruments, USA); (b) TA Discovery DSC 25 (TA Instruments, USA) Test conditions: the heating rate was 10° C. / min and dry nitrogen was used as the purge gas.

[0089] (3) Thermogravimetric analysis (TGA) Instrument models: (a) Discovery TGA 55 (TA Instruments, USA); (b) TGA 4000 (PerkinElmer, Germany). Test conditions: automatic weighing in a heating furnace, heating rate was 10°C / min, dry nitrogen was used as purge gas.

[0090] (4) Polarized Light Microscopy (PLM) Equipment model: Polarized microscope ECLIPSE LV100POL (Nikon, Japan)

[0091] (5) Nuclear magnetic resonance ( 1 H NMR) Equipment model: Bruker Advance 400 with B-ACS 120 automated sampling system

[0092] (6) Dynamic Vapor Sorption Analysis (DVS) Device model: DVS Intrinsic (SMS, UK) Test conditions: gradient mode was used, humidity ranged from 0% to 90%, humidity increase for each gradient was 10%, and the holding time for each gradient was 1 hour.

[0093] Example 1 Preparation of (6-(4-((4-(1H-pyrazol-4-yl)phenyl)amino)pyrimidin-2-yl)-1-methyl-1H-indol-2-yl)(3,3-difluoroazetidin-1-yl)methanone (Compound A) (described in PCT / CN2018 / 093713, which is incorporated herein by reference in its entirety) [ka]

[0094] Step 1: Compound A-1 (20 g, 83.31 mmol) and ethanol (200 mL) were added to a 500 mL flask, followed by the addition of thionyl chloride (19.82 g, 166.63 mmol), and the mixture was allowed to react at 60 °C for 3 hours. Thin-layer chromatography (petroleum ether / ethyl acetate = 10:1) assay showed that the reaction was complete. The reaction solution was concentrated to give the crude product, which was dissolved in dichloromethane (500 mL), and the resulting solution was washed twice with saturated aqueous sodium bicarbonate solution (150 mL each). The organic phase was washed with saturated brine, then dried over anhydrous sodium sulfate, filtered, and concentrated to give compound A-2 (21 g, brown solid, yield: 94.01%). MS m / z (ESI): 266.1; 268.1 [MH].

[0095] Step 2: Compound A-2 (21 g, 78.33 mmol) and bis(pinacolato)diboron (26.85 g, 105.74 mmol) were dissolved in 1,4-dioxane (200 mL). Potassium acetate (23.06 g, 234.98 mmol) and Pd(dppf)Cl (3.24 g, 3.91 mmol) were added. The mixture was purged with argon three times and placed in an oil bath at 80 °C overnight. LC-MS showed the reaction was complete. The reaction solution was cooled to room temperature and concentrated under reduced pressure. The residue was purified by column chromatography (petroleum ether / ethyl acetate = 100:1 to 5:1) to give compound A-3 (17.5 g, white solid, yield: 70.89%). 1 H NMR(400 MHz,CDCl3)δ 8.92(s,1H),7.92(s,1H),7.69(d,J=8.1 Hz,1H),7.57(d,J=8.1 Hz,1H),7.22-7.18(m,1H),4.42(q,J=7.1 Hz,2H),1.42(t,J=7.1 Hz,3H),1.37(s,12H).MS m / z(ESI):316.2 [M+H].

[0096] Step 3: Compound A-3 (10.0 g, 31.8 mmol) was dissolved in tetrahydrofuran (250 mL) and, under ice bath cooling, sodium hydride (1.91 g, 47.8 mmol) was added, followed by reaction for 30 minutes. Iodomethane (13.5 g, 95.4 mmol) was slowly added to the reaction mixture, and the mixture was allowed to react at room temperature overnight. Thin-layer chromatography (petroleum ether / ethyl acetate = 5:1) showed the reaction was complete. The reaction mixture was quenched with water (100 mL) and extracted with ethyl acetate (150 mL × 2). The combined organic phase was washed successively with saturated aqueous ammonium chloride solution (200 mL × 2) and saturated brine (300 mL × 2), dried over anhydrous sodium sulfate, filtered, and concentrated. The crude product was isolated and purified by column chromatography (petroleum ether / ethyl acetate = 15:1) to give compound A-4 (6.5 g, yellow solid, yield: 62.5%). 1H NMR(400 MHz,CDCl3)δ 7.91(s,1H),7.68-7.65(m,1H),7.57(d,J=8.0 Hz,1H),7.27(s,1H),4.12(s,3H),3.91(s,3H),1.38(s,12H).

[0097] Step 4: Compound Reg-1-16 (1.00 g, 2.70 mmol) and A-4 (1.33 g, 4.04 mmol) were dissolved in ethanol / water (8:1) (120 mL), sodium carbonate (572 mg, 5.40 mmol) and Pd(PPh3)Cl2 (189 mg, 0.27 mmol) were added, the mixture was purged with argon three times, and the reaction was placed in an oil bath at 110 °C overnight. LC-MS showed the reaction was complete. The reaction was cooled to room temperature and concentrated under reduced pressure. The residue was diluted with water (30 mL), and the pH was adjusted to 1 with 6 N HCl. A large amount of solid precipitated and was filtered. The solid was washed with methanol to give compound A-5 (900 mg, yellow solid, crude product). 1 H NMR(400 MHz,DMSO-d6)δ 11.54(s,1H),8.77(s,1H),8.38(d,J=7.2 Hz,1H),8.14(s,2H),8.03(d,J=8.8 Hz,1H),7.92(d,J=8.4 Hz,1H),7.80-7.69(m,4H),7.33(s,1H),7.07(d,J=8.0 Hz,1H),4.16(s,3H).

[0098] Step 5: Compound A-5 (300 mg, 0.73 mmol) was dissolved in N,N-dimethylformamide (6 mL), and HATU (335 mg, 0.88 mmol) and DIEA (377 mg, 2.92 mmol) were added. The reaction was allowed to proceed at room temperature for 30 minutes. Compound Aa (114 mg, 0.88 mmol) was then added, and the reaction was continued at room temperature for 2 hours. LC-MS showed that the reaction was complete. The reaction solution was concentrated under reduced pressure, and the crude product was purified by high-performance liquid chromatography to give compound A (185 mg, yellow solid, yield: 52.1%). 1H NMR(400 MHz,DMSO-d6)δ 10.63(s,1H),8.52(s,1H),8.41(d,J=6.4 Hz,1H),8.09(s,2H),8.06(dd,J=8.4,1.2 Hz,1H),7.83(d,J=8.4 Hz,1H),7.77(d,J=8.0 Hz,2H),7.72(d,J=8.4 Hz,2H),7.13(s,1H),6.86(d,J=6.4 Hz,1H),4.91(s,2H),4.57(s,2H),4.05(s,3H).MS m / z(ESI):486.2 [M+H].

[0099] Example 2: Preparation of Crystalline Form I of Compound A Monohydrochloride Monohydrate Compound A (3.7 g) was added to a reaction flask, followed by acetone (250 ml). The reaction solution was heated to 50°C and stirred for 10 minutes, at which point the solid completely dissolved. Water (26.7 ml) was added, and the mixture was stirred to obtain a clear solution. Concentrated hydrochloric acid (12 mol / L) (0.641 ml) was added, causing a large amount of solid to precipitate. The mixture was stirred at 50°C for 18 hours and filtered. The solid was collected and dried under vacuum at 50°C for 8 hours to obtain a crystalline form (2.5 g, yield: 60.8%). The XRPD pattern obtained by X-ray powder diffraction analysis is shown in Figure 1. Upon DSC and TGA analysis, the DSC graph is shown in Figure 2, the TGA graph is shown in Figure 3, and the sample was examined by scanning electron microscopy; the crystalline form is shown in Figure 4. 1 H NMR(400 MHz,DMSO-d6)δ:11.19(s,1H),8.68(s,1H),8.39(d,J=6.8Hz,1H),8.11(s,2H),8.04(d,J=8Hz,1H),7.86 (d,J=8Hz,1H),7.78-7.73(m,4H),7.14(s,1H),6.99(d,J=6.4Hz,1H),4.99-4.57(brs,4H),4.07(s,3H).

[0100] Example 3: Preparation of Crystalline Form II of Compound A Monohydrochloride Dihydrate Crystalline Form I of Compound A monohydrochloride monohydrate (2.0 g) was added to methanol (100 ml, water content: 0.08%), and the mixture was stirred at 20-30°C for 48 hours. The solid was collected by filtration and dried under vacuum at 50°C for 8 hours to obtain the crystalline form (1.0 g, yield: 48.4%). The XRPD pattern obtained by X-ray powder diffraction analysis is shown in Figure 5, the DSC-TGA graph obtained by DSC and TGA analysis is shown in Figure 6, and the sample was examined by scanning electron microscope; the crystalline form is shown in Figure 7. 1 H NMR(400 MHz,DMSO-d6)δ:11.18(s,1H),8.65(s,1H),8.39(d,J=6.8Hz,1H),8.10(s,2H),8.04(d,J=8Hz,1H),7.86 (d,J=8Hz,1H),7.778-7.73(m,4H),7.14(s,1H),6.97(d,J=6.4Hz,1H),4.91-4.57(brs,4H),4.06(s,3H).

[0101] Example 4: Preparation of Crystalline Form III of Compound A Monohydrochloride Sesquihydrate Crystalline Form I of Compound A monohydrochloride monohydrate (2.0 g) was added to methanol (100 ml, water content: 0.08%), and the temperature was maintained at 20-30 °C. Toluene (560 ml) was added dropwise, resulting in precipitation of a solid. The mixture was stirred for 2 hours, filtered, and the solid was dried under vacuum at 50 °C for 8 hours to obtain the crystalline form (1.1 g, yield: 54.2%). The XRPD pattern obtained by X-ray powder diffraction analysis is shown in Figure 8. The DSC-TGA graph obtained by DSC and TGA analysis is shown in Figure 9. The sample was examined by scanning electron microscopy, and the crystalline form is shown in Figure 10. 1 H NMR(400 MHz,DMSO-d6)δ:10.98(s,1H),8.61(s,1H),8.37(d,J=6.8Hz,1H),8.10(s,2H),8.04(d,J=8Hz,1H),7.85 (d,J=8Hz,1H),7.77-7.72(m,4H),7.13(s,1H),6.94(d,J=6.4Hz,1H),4.91-4.57(brs,4H),4.05(s,3H).

[0102] Example 5: Preparation of Crystalline Form IV of Compound A Monohydrochloride Mono-Dimethyl Sulfoxide Complex Crystalline Form I of Compound A monohydrochloride monohydrate (2.0 g) was added to dimethyl sulfoxide (100 ml) and the temperature was maintained at 20-30°C. Toluene (900 ml) was added dropwise, resulting in precipitation of a solid. The mixture was stirred for 2 hours, filtered, and the solid was dried under vacuum at 50°C for 8 hours to obtain the crystalline form (0.8 g, yield: 36.0%). The XRPD pattern obtained by X-ray powder diffraction analysis is shown in Figure 11. Upon DSC and TGA analysis, the DSC graph is shown in Figure 12, the TGA graph is shown in Figure 13, and the sample was examined by scanning electron microscopy; the crystalline form is shown in Figure 14. 1 H NMR(400 MHz,DMSO-d6)δ:11.06(s,1H),8.64(s,1H),8.37(d,J=6.8Hz,1H),8.10(s,2H),8.04(d,J=8Hz,1H),7.85(d,J= 8Hz,1H),7.77-7.72(m,4H),7.13(s,1H),6.94(d,J=6.4Hz,1H),4.95-4.57(brs,4H),4.06(s,3H),2.36(s,6H).

[0103] Experimental Example Experimental Example 1: Pharmacokinetics study Compound A monohydrochloride monohydrate crystalline form I and Compound A (the vehicle for both samples was 0.5% CMC-Na) were orally administered once at a dose of 20 mpk to two groups of beagle dogs, and blood samples were taken before administration (0 hours) and at 0.25, 0.5, 1, 2, 4, 6, 8 and 24 hours after administration. The samples were processed and subjected to chromatographic analysis. The test results are shown in the table below: [Table 5]

[0104] According to the above results, the exposure of crystalline Form I of Compound A monohydrochloride monohydrate upon oral administration is significantly better than that of Compound A.

[0105] Experimental Example 2: Solid Stability Test Crystalline Form I of Compound A monohydrochloride monohydrate was subjected to powder X-ray diffraction analysis after being subjected to conditions of 40°C / 75% RH and 60°C / 90% RH for 7 days, respectively. The test results are shown in Figure 15. The results showed that crystalline Form I of Compound A monohydrochloride monohydrate remained unchanged.

[0106] Experimental Example 3: Solubility test Compound A monohydrochloride monohydrate crystalline Form I was tested for solubility in simulated gastric fluid (SGF), fed-state simulated intestinal fluid (FeSSIF), and fasted-state simulated intestinal fluid (FaSSIF). The test results are as follows: SGF: 0.0012mg / ml; FeSSIF: 0.017 mg / ml; FaSSIF: 0.00028mg / ml.

[0107] Experimental example 4: DVS test The dynamic moisture sorption / desorption curve of crystalline Form I of Compound A monohydrochloride monohydrate was determined using a DVS Intrinsic (SMS, UK) in gradient mode with a humidity range of 0% to 90% (the humidity increment was 10% for each gradient, and each gradient was maintained for 1 hour). The crystalline form of the sample before and after the DVS test was examined by XRPD. DVS showed that crystalline Form I of Compound A monohydrochloride monohydrate had a water absorption rate of 0.67% from 0 to 10% RH and 0.9% from 10 to 90% RH. The XRPD patterns of the sample before and after the DVS test are shown in Figure 16. According to Figure 16, crystalline Form I of Compound A monohydrochloride monohydrate did not change after the DVS test.

[0108] Experimental Example 5: Crystal morphology transformation test Approximately 5 mg of each of crystalline Form I, Form II, and Form III was mixed and placed in a 4 mL vial, followed by the addition of 0.8 mL of water. The suspension was stirred at room temperature for 7 days. The mixture was subjected to X-ray powder diffraction analysis on days 4 and 7, and the results are shown in Figure 17. The results showed that on day 4, the mixture transformed into a mixed crystal of crystalline Form I and Form II, and Form III disappeared. On day 7, the mixture completely transformed into crystalline Form I. This indicates that Form I is the most stable crystalline form in water at room temperature.

[0109] Experimental Example 6: Crushing test An appropriate amount of crystalline form I of Compound A monohydrochloride monohydrate was taken, crushed for 2 minutes and 5 minutes, and subjected to powder X-ray diffraction analysis. The test results are shown in Figure 18. The results show that the crystalline form did not change.

[0110] Experimental Example 7: Solid Stability Test 20.8 mg of Compound A monohydrochloride monohydrate crystalline Form I was collected and placed in a 4 mL vial. The vial was left open and dried under vacuum at 50° C. for 2 days. The sample was subjected to powder X-ray diffraction and TGA analysis, and the results are shown in FIGS. 19 and 20.

[0111] 19.3 mg of crystalline Form I of Compound A monohydrochloride monohydrate was collected and placed in a 4 mL vial. The vial was left open and placed under conditions of 25° C. / 92.5% RH for 7 days. The sample was subjected to powder X-ray diffraction analysis, and the results are shown in FIG.

[0112] The results in Figures 19 and 20 indicated that the physical stability of crystalline Form I of Compound A monohydrochloride monohydrate was good and the crystalline form did not change. The TGA weight loss of the sample after drying did not change, indicating that water molecules and drug molecules were tightly associated in crystalline Form I.

[0113] Experimental Example 8: Determination of chloride ion content in a sample by potentiometric titration 270 mg of crystalline Form I of Compound A monohydrochloride monohydrate was taken and placed in a titration beaker. 25 mL of dimethyl sulfoxide was added to dissolve the solution, followed by 15 mL of water. The solution was thoroughly stirred. Titration was performed using a Mettler T50 potentiometric titrator with a DMI141-SC silver electrode and silver nitrate titrant (0.1 mol / L). The chloride ion content in the sample was calculated based on the amount of silver nitrate titrant consumed. The chloride ion content in multiple batches of samples determined by this method ranged from 6.5% to 6.7%, whereas the theoretical chloride ion content in Compound A monohydrochloride monohydrate is 6.6%. Therefore, the measured values ​​are consistent with the theoretical value. [Table 6]

[0114] Experimental Example 9: Determining the water content of a sample by the Karl Fischer method 50 mg of crystalline Form I of Compound A monohydrochloride monohydrate was taken and added to a Karl Fischer sample bottle. The sample bottle was sealed and heated to 160°C. The water content was determined by the coulometric oven method using a Metrohm KF 831 coulometer. The water content in multiple batches of samples determined by this method ranged from 3.2% to 4.0%, while the theoretical water content of Compound A monohydrochloride monohydrate is 3.3%. Therefore, the measured values ​​are consistent with the theoretical values. [Table 7]

[0115] In addition to those described herein, various modifications of the present invention will become apparent to those skilled in the art from the foregoing description. Such modifications are intended to fall within the scope of the appended claims. Each reference, including all patents, applications, journal articles, books, and any other disclosures mentioned herein, is incorporated herein by reference in its entirety.

[0116] Claim 1. A salt of compound A, which is an inorganic or organic acid salt, preferably the hydrochloride salt. [ka]

[0117] Claim 2. Compound A hydrochloride, Preferably, the compound A hydrochloride is compound A monohydrochloride; Preferably, the Compound A hydrochloride is Compound A monohydrochloride monohydrate crystalline form I; the crystalline form I having an XRPD pattern comprising characteristic peaks at diffraction angles (2θ) of about 8.3±0.2°, 12.6±0.2°, and 18.5±0.2°; Preferably, an XRPD pattern comprising characteristic peaks at diffraction angles (2θ) of about 8.3±0.2°, 12.6±0.2°, 16.5±0.2°, 17.3±0.2°, 17.8±0.2°, 18.5±0.2°, 24.4±0.2°, 26.0±0.2° and 29.1±0.2°; Most preferably, an XRPD pattern comprising characteristic peaks at diffraction angles (2θ) of about 8.3±0.2°, 11.2±0.2°, 12.6±0.2°, 16.5±0.2°, 17.3±0.2°, 17.8±0.2°, 18.5±0.2°, 19.5±0.2°, 19.8±0.2°, 20.6±0.2°, 21.0±0.2°, 21.7±0.2°, 23.7±0.2°, 24.4±0.2°, 25.5±0.2°, 26.0±0.2°, 29.1±0.2°, and 33.6±0.2°. 2. A salt of compound A according to claim 1, having the formula:

[0118] Claim 3. Compound A hydrochloride, Preferably, the compound A hydrochloride is compound A monohydrochloride; Preferably, the Compound A hydrochloride is Compound A monohydrochloride dihydrate crystalline form II; the crystalline form II has an XRPD pattern comprising characteristic peaks at diffraction angles (2θ) of about 9.9±0.2°, 13.2±0.2°, and 16.2±0.2°; Preferably, an XRPD pattern comprising characteristic peaks at diffraction angles (2θ) of about 9.9±0.2°, 11.0±0.2°, 13.2±0.2°, 13.5±0.2°, 13.8±0.2°, 16.2±0.2°, 19.4±0.2° and 25.2±0.2°; Most preferably, an XRPD pattern comprising characteristic peaks at diffraction angles (2θ) of about 6.5±0.2°, 9.9±0.2°, 11.0±0.2°, 11.6±0.2°, 11.8±0.2°, 13.2±0.2°, 13.5±0.2°, 13.8±0.2°, 16.2±0.2°, 16.6±0.2°, 17.1±0.2°, 18.2±0.2°, 19.4±0.2°, 20.0±0.2°, 20.3±0.2°, 20.9±0.2°, 22.6±0.2°, 24.9±0.2°, 25.2±0.2°, and 25.9±0.2°. 2. A salt of compound A according to claim 1, having the formula:

[0119] Claim 4. Compound A hydrochloride, Preferably, the compound A hydrochloride is compound A monohydrochloride; Preferably, the Compound A hydrochloride is Compound A monohydrochloride sesquihydrate crystalline form III; the crystalline form III having an XRPD pattern comprising characteristic peaks at diffraction angles (2θ) of about 5.1±0.2°, 9.1±0.2°, and 10.2±0.2°; Preferably, an XRPD pattern comprising characteristic peaks at diffraction angles (2θ) of about 5.1±0.2°, 5.4±0.2°, 6.0±0.2°, 9.1±0.2°, 10.2±0.2°, 11.2±0.2° and 18.0±0.2°; Most preferably, the angle is about 5.1±0.2°, 5.4±0.2°, 6.0±0.2°, 6.3±0.2°, 7.5±0.2°, 7.8±0.2°, 8.4±0.2°, 9.1±0.2°, 10.2±0.2°, 10.7±0.2°, 11.2±0.2°, 12.2±0.2°, 12.5±0.2°, 12.9±0.2°, 13.7±0.2° XRPD pattern containing characteristic peaks at diffraction angles (2θ) of 13.9±0.2°, 15.4±0.2°, 15.7±0.2°, 16.2±0.2°, 16.8±0.2°, 18.0±0.2°, 18.7±0.2°, 19.3±0.2°, 20.0±0.2°, 20.6±0.2°, 21.5±0.2° and 23.3±0.2°. 2. A salt of compound A according to claim 1, having the formula:

[0120] Claim 5. Compound A hydrochloride, Preferably, the compound A hydrochloride is compound A monohydrochloride; Preferably, the Compound A hydrochloride is crystalline form IV of Compound A monohydrochloride mono-dimethyl sulfoxide complex; the crystalline form IV having an XRPD pattern comprising characteristic peaks at diffraction angles (2θ) of about 6.6±0.2°, 8.9±0.2°, and 18.5±0.2°; Preferably, an XRPD pattern comprising characteristic peaks at diffraction angles (2θ) of about 6.6±0.2°, 8.9±0.2°, 11.6±0.2°, 14.6±0.2°, 17.7±0.2°, 18.5±0.2°, 24.7±0.2° and 25.0±0.2°; Most preferably, the angle is about 4.1±0.2°, 6.6±0.2°, 8.9±0.2°, 11.2±0.2°, 11.6±0.2°, 13.3±0.2°, 13.8±0.2°, 14.6±0.2°, 15.7±0.2°, 17.7±0.2°, 18.1±0.2°, 18.5±0.2°, 19.4±0.2°, 20.0±0.2°, 21.0±0.2°, 22.0±0.2°, 23.0±0.2°, 24.0±0.2°, 25.0±0.2°, 26.0±0.2°, 27.0±0.2°, 28.0±0.2°, 29.0±0.2°, 30.0±0.2°, 31.0±0.2°, 32.0±0.2°, 33.0±0.2°, 34.0±0.2°, 35.0±0.2°, 36.0±0.2°, 37.0±0.2°, 38.0±0.2°, 39.0±0.2°, 40.0±0.2°, 41.0±0.2°, 42.0±0.2°, 43.0±0.2°, 44.0±0.2°, 45.0±0.2°, 46.0±0.2°, 47.0±0.2°, 48.0±0.2°, 49.0±0.2°, 50.0±0.2°, 51.0±0.2°, 52.0±0.2°, XRPD pattern containing characteristic peaks at diffraction angles (2θ) of 0.7±0.2°, 21.4±0.2°, 22.0±0.2°, 22.6±0.2°, 23.0±0.2°, 23.8±0.2°, 24.7±0.2°, 25.0±0.2°, 26.5±0.2°, 27.9±0.2°, 29.7±0.2° and 32.8±0.2° 2. A salt of compound A according to claim 1, having the formula:

[0121] Claim 6. A pharmaceutical composition comprising one or more of the salts of Compound A of claim 1, crystalline Form I of claim 2, crystalline Form II of claim 3, crystalline Form III of claim 4, and crystalline Form IV of claim 5, and one or more pharmaceutically acceptable carriers.

[0122] Claim 7. Use of any one or more of a salt of Compound A as defined in Claim 1, crystalline Form I as defined in Claim 2, crystalline Form II as defined in Claim 3, crystalline Form III as defined in Claim 4, and crystalline Form IV as defined in Claim 5 in the manufacture of a medicament as a Rho-associated protein kinase (ROCK) inhibitor, preferably a selective ROCK2 inhibitor.

[0123] Claim 8. The medicament is for treating an autoimmune disorder (including rheumatoid arthritis, systemic lupus erythematosus (SLE; lupus), psoriasis, Crohn's disease, atopic dermatitis, eczema, or graft-versus-host disease (GVHD)); a cardiovascular disorder (including hypertension, atherosclerosis, restenosis, cardiac hypertrophy, cerebral ischemia, cerebral vasospasm, or erectile dysfunction); inflammation (including asthma, cardiovascular inflammation, ulcerative colitis, or renal inflammation); a central nervous system disorder (including neuronal degeneration or spinal cord injury); The disorder is preferably Huntington's disease, Parkinson's disease, Alzheimer's disease, amyotrophic lateral sclerosis (ALS), or multiple sclerosis; arterial thrombotic disorders (including platelet aggregation or leukocyte aggregation); fibrotic disorders (including liver fibrosis, pulmonary fibrosis, or renal fibrosis); neoplastic diseases (lymphoma, carcinoma (e.g., squamous cell carcinoma, small cell lung carcinoma, pituitary carcinoma, esophageal carcinoma, non-small cell lung carcinoma, lung adenocarcinoma, lung squamous cell carcinoma, peritoneal carcinoma, hepatocellular carcinoma, gastrointestinal carcinoma, pancreatic carcinoma, glioblastoma, cervical cancer, ovarian cancer, bladder cancer, liver cancer, breast cancer, colon cancer, colorectal cancer, endometrial or uterine cancer, salivary gland cancer, kidney cancer, prostate cancer, vulvar cancer, thyroid cancer, brain cancer, endometrial cancer, testicular cancer, bile duct cancer, gallbladder cancer, stomach cancer, melanoma, or head and neck cancer), leukemia, astrocytoma, soft tissue sarcoma, sarcoma, or blastoma); metabolic syndrome; insulin resistance; hyperinsulinemia; type 2 diabetes mellitus; impaired glucose tolerance; osteoporosis; eye disorders (ocular hypertension, age-related macular degeneration) 8. The use of claim 7, for the prevention or treatment of a disease mediated by Rho-associated protein kinase (ROCK), including amyotrophic lateral sclerosis (AMD), choroidal neovascularization (CNV), diabetic macular edema (DME), iris neovascularization, uveitis, glaucoma (including primary open-angle glaucoma, acute angle-closure glaucoma, pigmentary glaucoma, congenital glaucoma, normal-tension glaucoma, secondary glaucoma or neovascular glaucoma), or retinitis of prematurity (ROP).

[0124] Claim 9. The use of claim 7, wherein the medicament is for the prevention or treatment of a disease mediated by Rho-associated protein kinase (ROCK), including lupus nephritis, atherosclerosis, rheumatoid arthritis (RA), hemangioma, angiofibroma, pulmonary fibrosis, psoriasis, corneal graft rejection, insulin-dependent diabetes mellitus, multiple sclerosis, myasthenia gravis, Crohn's disease, autoimmune nephritis, primary biliary cirrhosis, acute pancreatitis, allographic rejection, allergic inflammation, contact dermatitis, delayed-type hypersensitivity, inflammatory bowel disease, septic shock, osteoporosis, osteoarthritis, neuroinflammation, Ossia-Weber syndrome, restenosis, fungal infections, parasitic infections, and viral infections.

[0125] Claim 10. A method for preparing crystalline Form I of Compound A monohydrochloride monohydrate according to Claim 2, comprising the steps of: adding a ketone solvent (preferably a ketone having 3 to 6 carbon atoms, such as acetone, butanone, methyl ethyl ketone, methyl isobutyl ketone, and diethyl ketone) to Compound A, stirring under heating (preferably at about 40 to 80°C, for example at about 50°C or about 60°C), and then adding water (the volume ratio of the ketone solvent to water is preferably about (1 to 15):1, preferably about 10:1) to completely dissolve Compound A; and then adding hydrochloric acid (the concentration of hydrochloric acid is preferably 2 to 15 mol / L, preferably 12 mol / L, and the molar ratio of Compound A to HCl is 1:(1 to 1.3)), stirring under heating (preferably at about 40 to 80°C, for example at about 50°C or 60°C), filtering, and optionally drying to obtain the crystalline form.

Claims

[Claim 1] A salt of Compound A which is an inorganic or organic acid salt, preferably a hydrochloride salt. 【Chemistry 1】

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