Nanocrystalline formulations of ROCK2 inhibitors and methods for their preparation

Abstract

Nanocrystal formulations comprising a ROCK2 inhibitor and a stabilizer and methods for preparing the same. The present invention also relates to the use of the nanocrystal formulations in the prevention, amelioration, and / or treatment of selected diseases and conditions, particularly diseases such as idiopathic pulmonary fibrosis, fatty liver disease and / or steatohepatitis, graft-versus-host disease following hematopoietic stem cell transplantation, or viral infections.

Description

[Technical field]

[0001] The present disclosure relates to the pharmaceutical field. In particular, the present disclosure relates to nanocrystalline formulations of ROCK2 inhibitors and methods for their preparation. [Background technology]

[0002] Idiopathic pulmonary fibrosis (IPF) is a progressive respiratory disease with lung tissue fibrosis and decline and loss of lung function as the main clinical features. The median survival is 2.5-3 years. A phase II clinical trial for ROCK2-targeted drugs is being conducted in the United States for IPF (Kadmon Holdings, Inc.), and preliminary results support the safety and efficacy of ROCK2 inhibitors in the treatment of IPF.

[0003] [6-[4-[[4-(1H-pyrazol-4-yl)phenyl]amino]pyrimidin-2-yl]-1-methyl-1H-indol-2-yl](3,3-difluoroazetidin-1-yl)methanone is a new type of highly selective ROCK2 inhibitor with a new target and a new structural type, developed completely independently by Beijing Tide Pharmaceutical Co., Ltd. From the perspective of patient compliance, oral formulations are chosen for the treatment of IPF. Its high selectivity for the target greatly reduces safety risks. The company has patented this compound in the United States and has applied for compound patents in China, the European Union, Japan, South Korea, India, Canada, Australia and other countries and regions.

[0004] [6-[4-[[4-(1H-pyrazol-4-yl)phenyl]amino]pyrimidin-2-yl]-1-methyl-1H-indol-2-yl](3,3-difluoroazetidin-1-yl)methanone is a pale yellow to yellow solid powder with low solubility, insoluble in water and buffer salt solutions with pH 1.0 to pH 6.8. It has poor physical properties, high adhesion, prone to electrostatic charge, and prone to aggregation. Therefore, how to prepare a formulation of the compound and improve the dissolution of the product has become an urgent technical problem to be solved by those skilled in the art. Summary of the Invention

[0005] The objective of the present disclosure is to provide a nanocrystal formulation of ROCK2 inhibitor and its preparation method, so as to improve the dissolution of ROCK2 inhibitor. The specific technical solutions are as follows:

[0006] The present disclosure provides, first, a nanocrystal formulation comprising a ROCK2 inhibitor and a stabilizer, wherein the ROCK2 inhibitor is a compound of formula (I): [ka] [In the formula, Ring A is [ka] where the above group is attached to the pyrimidine ring through one of the two positions marked with * or ** and to the carbonyl group at the other position; R 9 and R 10 are each independently H, halogen, or C 1-6 Alkyl, C 2-6 Alkenyl, C 3-10 Carbocyclyl, 3-10 membered heterocyclyl, C 6-10 Aryl, 5-14 membered heteroaryl, C 6-12 Aralkyl, -C(=O)R 5 , and -C 1-6 alkylene-O(P=O)(OH); Each m is independently an integer of 0, 1, 2, or 3; Each n is independently an integer of 0, 1, or 2; Preferably, ring A is [ka] where the above groups are attached to the pyrimidine ring at the positions marked * and to the carbonyl group at the positions marked **; R 10 H and C 1-6 alkyl, preferably H or methyl; R is H and C 1-6 alkyl; R 1 teeth, [ka] and; R 2 H and C 1-6 alkyl; R 3 , R 4 , R 7 , and R 8 are each independently H, a halogen, or -NR 5 R 6 , -OH, C 1-6 Alkyl, and -OR 5 Selected from; The alkylene, alkyl, alkenyl, carbocyclyl, heterocyclyl, aryl, heteroaryl, and aralkyl groups each include halogen, C 1-6 Alkyl, and -OR 5 may be optionally substituted with one or more substituents independently selected from: R 5 and R 6 are independently H, C 1-6 Alkyl, C 3-10 Carbocyclyl, 3-10 membered heterocyclyl, C 6-10 Aryl, 5-14 membered heteroaryl, and C 6-12 aralkyl] or a pharma- ceutically acceptable salt, ester, stereoisomer, polymorph, solvate, N-oxide, isotopically labeled derivative, metabolite, or prodrug thereof.

[0007] The present disclosure also provides a method for preparing a nanocrystal formulation, comprising milling a ROCK2 inhibitor and a stabilizer.

[0008] Another object of the present disclosure is methods and uses of nanocrystal formulations in the prevention, alleviation, and / or treatment of idiopathic pulmonary fibrosis.

[0009] Other objects of the present disclosure will be apparent to those skilled in the art from the context and examples. [Brief description of the drawings]

[0010] In order to more clearly describe the embodiments of the present disclosure and the technical solutions of the prior art, the drawings that need to be used in the embodiments and the prior art are briefly introduced below. Obviously, the drawings in the following description only show the embodiments of the present disclosure. For those skilled in the art, other drawings can also be obtained based on these drawings without exerting creative efforts. [Figure 1] 1 and 2 show the dissolution curves of Comparative Examples 1 to 3 of the present disclosure. [Diagram 2] 1 and 2 show the dissolution curves of Comparative Examples 1 to 3 of the present disclosure. [Diagram 3] FIG. 3 shows the dissolution curves of Examples 8 to 9 of the present disclosure and a comparative example. [Figure 4] 4-6 are dissolution curves of the nanosuspension of Example 8 of the present disclosure, the nanocrystal tablets of Examples 10-15, and the comparative example, respectively. [Diagram 5] 4-6 are dissolution curves of the nanosuspension of Example 8 of the present disclosure, the nanocrystal tablets of Examples 10-15, and the comparative example, respectively. [Figure 6] 4-6 are dissolution curves of the nanosuspension of Example 8 of the present disclosure, the nanocrystal tablets of Examples 10-15, and the comparative example, respectively. [Figure 7] FIG. 7 shows the dissolution curves of the nanosuspension of Example 8, the nanocrystal tablet of Example 15, the nanocrystal capsules of Examples 24-25 of the present disclosure, and the comparative example. DETAILED DESCRIPTION OF THE PREFERRED EMBODIMENTS

[0011] definition Unless otherwise defined below, 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 technology as commonly understood in the art, including technical variations or equivalents that would be obvious to those skilled in the art. Although the following terms are believed to be fully understood by those skilled in the art, the following definitions are provided to better explain this disclosure.

[0012] The term "nanocrystals" refers to both nanocrystals and nanosuspensions, and refers to a stable colloidal dispersion system formed by nanoscale drug particles dispersed in water in the presence of a stabilizer.

[0013] As used herein, the words "include," "comprise," "have," "contain," or "involve," and other variations thereof, are inclusive or open-ended and do not exclude other, unrecited elements or method steps.

[0014] The term "alkylene," as used herein, refers to a divalent saturated hydrocarbon group, preferably having 1, 2, 3, 4, 5, or 6 carbon atoms, such as methylene, ethylene, propylene, or butylene.

[0015] The term "alkyl" as used herein is defined as a straight or branched chain saturated aliphatic hydrocarbon. In some embodiments, an alkyl group has 1 to 12 (such as 1 to 6) carbon atoms. For example, as used herein, "C 1-6 The term "alkyl" refers to a straight or branched chain group of 1 to 6 carbon atoms (e.g., methyl, ethyl, n-propyl, isopropyl, n-butyl, isobutyl, sec-butyl, tert-butyl, n-pentyl, isopentyl, neopentyl, or n-hexyl), which may be optionally substituted with one or more (e.g., 1 to 3) suitable substituents such as a halogen (in which case the group is referred to as a "haloalkyl") (e.g., CH2F, CHF2, CF3, CCl3, C2F5, C2Cl5, CH2CF3, CH2Cl, or -CH2CH2CF3, etc.). 1-4 The term "alkyl" refers to a straight or branched aliphatic hydrocarbon chain of 1 to 4 carbon atoms (i.e., methyl, ethyl, n-propyl, isopropyl, n-butyl, isobutyl, sec-butyl, or tert-butyl).

[0016] As used herein, the term "alkenyl" refers to a monovalent linear or branched chain hydrocarbon radical containing one double bond and having 2 to 6 carbon atoms ("C 2-6 Alkenyl groups include, for example, vinyl, 1-propenyl, 2-propenyl, 2-butenyl, 3-butenyl, 2-pentenyl, 3-pentenyl, 4-pentenyl, 2-hexenyl, 3-hexenyl, 4-hexenyl, 5-hexenyl, 2-methyl-2-propenyl, and 4-methyl-3-pentenyl. When the compounds of the present disclosure contain an alkenylene group, the compounds may exist in the pure E (entgegen) form, the pure Z (zusammen) form, or any mixture thereof.

[0017] As used herein, the term "alkynyl" refers to a monovalent hydrocarbon group containing one or more triple bonds and preferably having 2, 3, 4, 5, or 6 carbon atoms, such as ethynyl or propynyl.

[0018] The term "cycloalkyl" as used herein refers to a saturated monocyclic or polycyclic (including bicyclic) hydrocarbon ring (e.g., monocyclic rings such as cyclopropyl, cyclobutyl, cyclopentyl, cyclohexyl, cycloheptyl, cyclooctyl, cyclononyl, etc., or bicyclic rings containing spiro, fused, or bridged systems (e.g., bicyclo[1.1.1]pentyl, bicyclo[2.2.1]heptyl, bicyclo[3.2.1]octyl, or bicyclo[5.2.0]nonyl, decahydronaphthyl, etc.), which may be optionally substituted with one or more (e.g., 1 to 3) suitable substituents. A cycloalkyl group has 3 to 15 carbon atoms. For example, "C 3-6 The term "cycloalkyl" refers to a saturated monocyclic or polycyclic (e.g., bicyclic) hydrocarbon ring having 3 to 6 ring-forming carbon atoms (e.g., cyclopropyl, cyclobutyl, cyclopentyl, or cyclohexyl, etc.), which may be optionally substituted with one or more (e.g., 1 to 3) suitable substituents, for example, methyl-substituted cyclopropyl.

[0019] The terms "carbocyclylene", "carbocyclyl", and "carbocycle" as used herein refer to a saturated (i.e., "cycloalkylene" and "cycloalkyl") or unsaturated (i.e., having one or more double and / or triple bonds within the ring) monocyclic or polycyclic hydrocarbon ring having, for example, 3 to 10 (preferably 3 to 8, more preferably 3 to 6) ring carbons, including, but not limited to, cyclopropyl / ren / ring, cyclobutyl / ren / ring, cyclopentyl / ren / ring, cyclohexyl / ren / ring, cycloheptyl / ren / ring, cyclooctyl / ren / ring, cyclononyl / ren / ring, cyclohexenyl / ren / ring, and the like.

[0020] As used herein, the terms "heterocyclyl," "heterocyclylene," and "heterocycle" refer to a saturated (i.e., heterocycloalkyl) or partially unsaturated (i.e., having one or more double and / or triple bonds in the ring) ring group having, for example, 3 to 10 (preferably 3 to 8, more preferably 3 to 6) ring atoms, with at least one ring atom being a heteroatom selected from N, O, and S, and the remaining ring atoms being C. For example, a "3- to 10-membered heterocycle(heterocyclyl)(heterocyclylene)" is a saturated or partially unsaturated heterocycle(heterocyclyl)(heterocyclylene) having 2 to 9 (e.g., 2, 3, 4, 5, 6, 7, 8, or 9) ring carbon atoms and one or more (e.g., 1, 2, 3, or 4) heteroatoms independently selected from N, O, and S. Examples of heterocyclylenes and heterocycles (heterocyclyls) include, but are not limited to, oxiranyl / ren, aziridinyl / ren, azetidinyl / ren, oxetanyl / ren, tetrahydrofuranyl / ren, dioxolinyl / ren, pyrrolidinyl / ren, pyrrolidonyl / ren, imidazolidinyl / ren, pyrazolidinyl / ren, pyrrolinyl / ren, tetrahydropyranyl / ren, piperidinyl / ren, morpholinyl / ren, dithianyl / ren, thiomorpholinyl / ren, piperazinyl / ren, or trithianyl / ren. The group also encompasses bicyclic systems, including spiro, fused, or bridged systems, such as 8-azaspiro[4.5]decane, 3,9-diazaspiro[5.5]undecane, 2-azabicyclo[2.2.2]octane, etc. The heterocyclylene and heterocycle (heterocyclyl) may be optionally substituted with one or more (such as 1, 2, 3 or 4) suitable substituents.

[0021] As used herein, the terms "aryl / ylene" and "aromatic ring" refer to an all-carbon monocyclic or fused polycyclic aromatic group having a conjugated pi-electron system. 6-10 Aryl / Rhene" and "C 6-10The term "aromatic ring" refers to an aromatic group containing 6 to 10 carbon atoms, such as a phenylene (benzene ring) or a naphthylene (naphthalene ring). The aryl / ene and aromatic ring may have one or more (such as 1 to 3) suitable substituents (e.g., halogen, -OH, -CN, -NO2, -C, -O, -O-C ... 1-6 The alkyl group may be optionally substituted with an alkyl group.

[0022] The terms "heteroaryl / ylene" and "heteroaryl ring" as used herein refer to monocyclic, bicyclic, or tricyclic aromatic ring systems having 5, 6, 8, 9, 10, 11, 12, 13, or 14 ring atoms, in particular 1 or 2 or 3 or 4 or 5 or 6 or 9 or 10 carbon atoms, and containing at least one heteroatom (e.g., oxygen, nitrogen, or sulfur) which may be the same or different, and which in any case may be benzo-fused. In particular, the "heteroaryl / ren" or "heteroaryl ring" is selected from thienyl / ren, furyl / ren, pyrrolyl / ren, oxazolyl / ren, thiazolyl / ren, imidazolyl / ren, pyrazolyl / ren, isoxazolyl / ren, isothiazolyl / ren, oxadiazolyl / ren, triazolyl / ren, thiadiazolyl / ren, etc., and benzo derivatives thereof; or pyridyl / ren, pyridazinyl / ren, pyrimidinyl / ren, pyrazinyl / ren, triazinyl / ren, etc., and benzo derivatives thereof.

[0023] The term "aralkyl" as used herein preferably refers to an aryl or heteroaryl substituted alkyl group, where aryl, heteroaryl, and alkyl are as defined herein. In general, the aryl group may have 6-14 carbon atoms, the heteroaryl group may have 5-14 ring atoms, and the alkyl group may have 1-6 carbon atoms. Representative aralkyl groups include, but are not limited to, benzyl, phenylethyl, phenylpropyl, and phenylbutyl.

[0024] As used herein, the term "halo" or "halogen" group is defined to include F, Cl, Br, or I.

[0025] The term "nitrogen-containing heterocycle" as used herein refers to a saturated or unsaturated monocyclic or bicyclic group having 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, or 13 carbon atoms and at least one nitrogen atom in the ring, which may also optionally contain one or more (such as 1, 2, 3, or 4) ring members selected from N, O, C=O, S, S=O, and S(=O)2, which are bonded to the remainder of the molecule through the nitrogen atom in the nitrogen-containing heterocycle and any remaining ring atoms. The nitrogen-containing heterocyclic ring may be optionally benzo-fused, and is preferably bonded to the remainder of the molecule through the nitrogen atom in the nitrogen-containing heterocyclic ring and any carbon atom in the fused benzene ring.

[0026] The term "substituted" means that one or more (e.g., 1, 2, 3, or 4) hydrogens on the specified atom are replaced with a selection from the specified group, provided that the replacement does not exceed the normal valence of the specified atom in its current state and that the substitution results in a stable compound. Combinations of substituents and / or variables are permissible only if such combinations result in stable compounds.

[0027] When a substituent is described as being "optionally substituted," the substituent can be (1) unsubstituted or (2) substituted. When a carbon of a substituent is described as being optionally substituted with one or more of a list of substituents, one or more hydrogens on that carbon can be replaced (within the limits of the hydrogens present) independently and / or together with an independently selected optional substituent. When a nitrogen of a substituent is described as being optionally substituted with one or more of the listed substituents, one or more hydrogens on that nitrogen can be replaced (within the limits of the hydrogens present) with an independently selected optional substituent, respectively.

[0028] When substituents are described as being "independently selected from," each substituent is selected independently of the others. Thus, each substituent can be the same or different from the other substituents.

[0029] As used herein, the term "one or more" means one or more than one, under reasonable conditions, for example, two, three, four, five, or ten.

[0030] Unless otherwise specified, as used herein, the point of attachment of a substituent may be from any suitable position on the substituent.

[0031] When a bond to a substituent is shown to cross a bond connecting two atoms in a ring, then such substituent may be bonded to any substitutable ring-forming atom of that ring.

[0032] The present disclosure also includes all pharma- ceutically acceptable isotopically labeled compounds that are identical to the compounds of the present disclosure except that one or more atoms have been replaced by an atom having the same atomic number but a different atomic mass or mass number as found in nature. Examples of isotopes suitable for inclusion in the compounds of the present disclosure include isotopes of hydrogen (e.g., deuterium ( 2 H), tritium ( 3 H); isotopes of carbon (e.g., 11 C. 13 C, and 14 C); isotopes of chlorine (e.g. 36 Cl); isotopes of fluorine (e.g. 18 F); isotopes of iodine (e.g. 123 I and 125 I); nitrogen isotopes (e.g. 13 N and 15 N); isotopes of oxygen (e.g. 15 O. 17 O, and 18 O); isotopes of phosphorus (e.g. 32 P); and sulfur isotopes (e.g. 35Some isotopically labeled compounds of the present disclosure (e.g., compounds incorporating a radioactive isotope) may be used in drug and / or substrate tissue distribution studies (e.g., assays). 3 H) and carbon-14 (i.e. 14 C) are particularly useful for this purpose due to their ease of incorporation and detection. 11 C. 18 F, 15 O, and 13 Substitution with isotopically labeled amines (e.g., N) can be used to test substrate receptor occupancy in positron emission tomography (PET) experiments. Isotopically labeled compounds of the present disclosure can be prepared by methods analogous to those described in the accompanying schemes and / or examples and preparations, substituting the appropriate isotopically labeled reagent already utilized. Pharmaceutically acceptable solvates of the present disclosure can be prepared by methods analogous to those described in the accompanying schemes and / or examples and preparations, substituting the appropriate isotopically labeled reagent already utilized. 、 Includes solvates, which may be acetone-d6, or DMSO-d6.

[0033] The term "stereoisomer" refers to an isomer formed due to at least one asymmetric center. In compounds with one or more (e.g., 1, 2, 3, or 4) asymmetric centers, they may give rise to racemic mixtures, single enantiomers, diastereomeric mixtures, and respective diastereomers. Certain individual molecules may also exist as geometric isomers (cis / trans). Similarly, compounds of the present disclosure may exist as mixtures of two or more structurally distinct forms in rapid equilibrium (often referred to as tautomers). Representative examples of tautomers include keto-enol tautomers, phenol-ketone tautomers, nitroso-oxime tautomers, and imine-enamine tautomers. It is understood that the scope of the present disclosure encompasses all such isomers in any percentage (e.g., 60%, 65%, 70%, 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99%) or mixtures thereof.

[0034] The chemical bonds of the compounds of the present disclosure are referred to herein as [ka] The use of a solid line to draw a bond to an asymmetric carbon atom is intended to indicate that all possible stereoisomers at that carbon atom (e.g., a specific enantiomer, a racemic mixture, etc.) are included. The use of a solid or dashed wedge line to draw a bond to an asymmetric carbon atom is intended to indicate that the depicted stereoisomer exists. When present in a racemic mixture, the solid or dashed wedge line is used to define the relative stereochemistry rather than the absolute stereochemistry. Unless otherwise indicated, the compounds of the present disclosure are intended to exist in stereoisomeric forms, such as cis and trans isomers, optical isomers (e.g., R and S enantiomers), diastereomers, geometric isomers, rotamers, conformational isomers, atropisomers, and mixtures thereof. The compounds of the present disclosure may exhibit more than one type of isomerism and consist of mixtures thereof (e.g., racemic mixtures and diastereomeric pairs).

[0035] The present disclosure encompasses all possible crystalline forms or polymorphs of the compounds of the present disclosure, whether these be a single polymorph or a mixture of two or more polymorphs in any ratio.

[0036] It should also be understood that some compounds of the present disclosure may exist in free form for therapeutic use, or in appropriate cases as their pharmaceutically acceptable derivatives.In the present disclosure, pharmaceutically acceptable derivatives include, but are not limited to, pharmaceutically acceptable salts, esters, solvates, N-oxides, metabolites, or prodrugs, which can directly or indirectly provide the compounds of the present disclosure or their metabolites or residues after being administered to a patient in need of treatment.Therefore, when "compounds of the present disclosure" are mentioned herein, it is intended to include various derivative forms of the above compounds.

[0037] Pharmaceutically acceptable salts of the compounds of the present disclosure include the acid addition and base salts thereof.

[0038] Suitable acid addition salts are formed from acids which form pharma- ceutically acceptable salts. Examples include acetate, adipate, aspartate, benzoate, benzenesulfonate, bicarbonate / carbonate, bisulfate / sulfate, borate, camphorsulfonate, citrate, cyclamate, ethanedisulfonate, ethanesulfonate, formate, fumarate, glucoheptonate, gluconate, glucuronate, hexafluorophosphate, hybenzate, hydrochloride / chloride, hydrobromide / bromide, hydroiodide / iodide, isethionate, lactate, etc. Salts include, but are not limited to, malate, maleate, malonate, methanesulfonate, methylsulfate, naphthylate, 2-naphthalenesulfonate, nicotinate, nitrate, orotate, oxalate, palmitate, pamoate, phosphate / hydrogenphosphate / dihydrogenphosphate, pyroglutamate, saccharate, stearate, succinate, tannate, tartrate, toluenesulfonate, trifluoroacetate, and xinafoate.

[0039] Suitable base addition salts are formed from bases which form pharma- ceutically acceptable salts, examples of which include aluminium, arginine, benzathine penicillin, calcium, choline, diethylamine, diethanolamine, glycinate, lysine, magnesium, meglumine, ethanolamine, potassium, sodium, tromethamine and zinc salts.

[0040] For a review of suitable salts, see Stahl and Wermuth, "Handbook of Pharmaceutical Salts: Properties, Selection, and Use" (Wiley-VCH, 2002). Methods for preparing pharma- ceutically acceptable salts of the compounds of the present disclosure are known to those of skill in the art.

[0041] The term "ester" as used herein means an ester derived from a compound of each of the general formulas herein, including a physiologically hydrolyzable ester (which may be hydrolyzed under physiological conditions to release the free acid or alcohol form of the compound of the present disclosure). The compounds of the present disclosure may themselves be esters.

[0042] The compounds of the present disclosure may exist in the form of solvates, preferably hydrates, in which the compounds of the present disclosure contain a polar solvent (such as, in particular, water, methanol, or ethanol) as a structural element of the crystal lattice of the compound. The amount of polar solvent, in particular water, may be present in a stoichiometric or non-stoichiometric ratio.

[0043] Those skilled in the art will understand that not all nitrogen-containing heterocycles can form N-oxides, since nitrogen requires an available lone pair of electrons to be oxidized to oxide. Those skilled in the art will recognize nitrogen-containing heterocycles that can form N-oxides. Those skilled in the art will also recognize that tertiary amines can form N-oxides. Synthetic methods for preparing N-oxides of heterocyclic amines and tertiary amines are well known to those skilled in the art, and include the oxidation of heterocyclic amines and tertiary amines using peroxyacids such as peracetic acid and m-chloroperbenzoic acid (MCPBA), hydrogen peroxide, alkyl hydroperoxides such as tert-butyl hydroperoxide, sodium perborate, and dioxiranes such as dimethyldioxirane. These methods for preparing N-oxides have been widely described and reviewed in the literature. See, for example, TL Gilchrist, Comprehensive Organic Synthesis, vol. 7, pp 748-750; AR Katritzky and AJ Boulton, Eds., Academic Press; and GWH Cheeseman and ESG Werstiuk, Advances in Heterocyclic Chemistry, vol. 22, pp 390-392, AR Katritzky and AJ Boulton, Eds., Academic Press.

[0044] Metabolites of the compounds of the present disclosure, i.e., substances formed in vivo upon administration of the compounds of the present disclosure, are also included within the scope of the present disclosure. Such products may be produced, for example, by oxidation, reduction, hydrolysis, amidation, deamidation, esterification, enzymatic degradation, etc. of the administered compound. Thus, the present disclosure includes metabolic products of the compounds of the present disclosure, such as compounds prepared by contacting a compound of the present disclosure with a mammal for a period of time sufficient to produce a metabolic product thereof.

[0045] The present disclosure further includes within its scope prodrugs of the compounds of the present disclosure. Prodrugs are specific derivatives of the compounds of the present disclosure that may have little or no pharmacological activity themselves and can be converted, for example, by hydrolysis, to compounds of the present disclosure having the desired activity when administered in or on the body. In general, such prodrugs are functional derivatives of compounds that are easily converted in vivo to compounds with the desired therapeutic effect. Further information regarding the use of prodrugs can be found in “Pro-drugs as Novel Delivery Systems”, Volume 14, ACS Symposium Series (T. Higuchi and V. Stella). Prodrugs of the present disclosure may be prepared, for example, by replacing appropriate functional groups present in the compounds of the present disclosure with certain groups known to those skilled in the art as “pro-moieties” (e.g., as described in “Design of Prodrugs”, H. Bundgaard (Elsevier, 1985)).

[0046] The present disclosure also encompasses the compounds of the present disclosure that contain protective groups.In any process for preparing the compounds of the present disclosure, it may be necessary and / or desirable to protect sensitive or reactive groups on any related molecule, thereby forming the chemically protected form of the compounds of the present disclosure.This can be achieved by conventional protective groups, such as those described in TW Greene & PGM Wuts, Protective Groups in Organic Synthesis, John Wiley & Sons, 1991, which are incorporated herein by reference.Protective groups can be removed at a later appropriate stage by using methods known in the art.

[0047] The term "about" means within ±10%, preferably within ±5%, and more preferably within ±2% of the referred value.

[0048] The term "effective amount" refers to an amount sufficient to achieve a desired therapeutic effect under the conditions of administration resulting in the amelioration of pathological symptoms, disease progression, physiological conditions associated therewith, or the induction of resistance to the progression of said disease.

[0049] Unless otherwise specified, the term "treating" as used herein means reversing, alleviating, inhibiting the progression of the disease or condition to which such term applies, or one or more symptoms of such disease or condition, or preventing such disease or condition, or one or more symptoms of such disease or condition.

[0050] As used herein, an "individual" includes a human or a non-human animal. Examples of human individuals include human individuals (called patients) suffering from diseases such as those described herein, or normal individuals. In this disclosure, a "non-human animal" includes all vertebrates, such as non-mammals (such as birds, amphibians, reptiles, etc.) and mammals (such as non-human primates, livestock, and / or domesticated animals (such as sheep, dogs, cats, cows, pigs, etc.)).

[0051] Detailed Description In one embodiment, the present disclosure relates to a nanocrystal formulation comprising a ROCK2 inhibitor and a stabilizer.

[0052] In one embodiment, the disclosure provides a nanocrystal formulation, wherein the ROCK2 inhibitor is a compound of formula (I): [ka] [In the formula, Ring A is [ka] where the above group is attached to the pyrimidine ring through one of the two positions marked with * or ** and to the carbonyl group at the other position; R 9 and R 10are each independently H, halogen, or C 1-6 Alkyl, C 2-6 Alkenyl, C 3-10 Carbocyclyl, 3-10 membered heterocyclyl, C 6-10 Aryl, 5-14 membered heteroaryl, C 6-12 Aralkyl, -C(=O)R 5 , and -C 1-6 alkylene-O(P=O)(OH); Each m is independently an integer of 0, 1, 2, or 3; Each n is independently an integer of 0, 1, or 2; Preferably, ring A is [ka] where the above groups are attached to the pyrimidine ring at the positions marked * and to the carbonyl group at the positions marked **; R 10 H and C 1-6 alkyl, preferably H or methyl; R is H and C 1-6 alkyl; R 1 teeth, [ka] and; R 2 H and C 1-6 alkyl; R 3 , R 4 , R 7 , and R 8 are each independently H, a halogen, or -NR 5 R 6 , -OH, C 1-6 Alkyl, and -OR 5 Selected from; The alkylene, alkyl, alkenyl, carbocyclyl, heterocyclyl, aryl, heteroaryl, and aralkyl groups each include halogen, C 1-6 Alkyl, and -OR 5may be optionally substituted with one or more substituents independently selected from: R 5 and R 6 are independently H, C 1-6 Alkyl, C 3-10 Carbocyclyl, 3-10 membered heterocyclyl, C 6-10 Aryl, 5-14 membered heteroaryl, and C 6-12 aralkyl] or a pharma- ceutically acceptable salt, ester, stereoisomer, polymorph, solvate, N-oxide, isotopically labeled derivative, metabolite, or prodrug thereof.

[0053] In one embodiment, the disclosure relates to a nanocrystal formulation, wherein the ROCK2 inhibitor is a compound of formula (II): [ka] [wherein each group is as defined above] or a pharma- ceutically acceptable salt, ester, stereoisomer, polymorph, solvate, N-oxide, isotopically labeled derivative, metabolite, or prodrug thereof.

[0054] In one embodiment, the disclosure relates to a nanocrystal formulation, wherein the ROCK2 inhibitor is a compound of formula (III): [ka] [In the formula, R 10 is H or methyl, preferably methyl] or a pharma- ceutically acceptable salt, ester, stereoisomer, polymorph, solvate, N-oxide, isotopically labeled derivative, metabolite, or prodrug thereof.

[0055] In one embodiment, the disclosure relates to a nanocrystal formulation, wherein the ROCK2 inhibitor is a compound of formula (IV): [ka] or a pharma- ceutically acceptable salt (particularly the hydrochloride salt), ester, stereoisomer, polymorph, solvate, N-oxide, isotopically labeled derivative, metabolite or prodrug thereof.

[0056] The chemical name of the compound of formula (IV) is [6-[4-[[4-(1H-pyrazol-4-yl)phenyl]amino]pyrimidin-2-yl]-1-methyl-1H-indol-2-yl](3,3-difluoroazetidin-1-yl)methanone.

[0057] In the present disclosure, the compound of formula (IV) as active ingredient can be prepared by known methods or obtained through commercial routes.No matter which method is used to obtain the compound of formula (IV), it is easy for those skilled in the art to realize.Therefore, the present disclosure does not detail the obtaining of the compound of formula (IV).

[0058] Generally, stabilizers need to be added during the preparation process of nanocrystals. The main reasons are as follows: On the one hand, nanoparticulation can significantly increase the specific surface area of ​​drug particles, which also increases the free energy of the entire formulation preparation system and destabilizes the system. On the other hand, as the temperature increases due to the mechanical energy production during the grinding process, the newly formed nanoparticles will be affected by the change in free energy and re-aggregate and recrystallize. These factors may cause the reduced particle size to return to a large level, reducing the surface area for dissolution, thereby affecting bioavailability in the body. Stabilizers can prevent drug particles from re-aggregating by adsorbing onto the surface of drug particles, thereby increasing the effective surface area and significantly improving the stability of nanosuspensions.

[0059] In the present disclosure, in addition to improving the wettability of the drug and stabilizing the particle size of the drug, the stabilizer may also improve the dissolution rate of the ROCK2 inhibitor nanocrystals by improving the particle size stability of the ROCK2 inhibitor nanocrystals after solidification, drying, and redissolution. Those skilled in the art may select an appropriate stabilizer based on the description of the role of the stabilizer in the present disclosure. Stabilizers include, but are not limited to, polysorbate 20, polysorbate 40, polysorbate 60, polysorbate 65, polysorbate 80, polysorbate 85, povidone K29 / 32, polyoxyethylene fatty acid esters, poloxamer 188, poloxamer 407, hydroxypropyl cellulose (HPC), hydroxypropyl methylcellulose (HPMC 3cps), polyvinylpyrrolidone (PVP K30), poloxamers (Pluronic F68 and Pluronic F127), sodium dodecyl sulfate (SDS), sodium docusate (DSS), polyethylene glycol 15-hydroxystearate, polyoxyethylene castor oil, copovidone, and the like.

[0060] In one embodiment, the disclosure relates to a nanocrystal formulation, wherein the stabilizer is selected from one or more of polysorbates, povidone, hydroxypropyl methylcellulose, polyethylene glycol, polyvinyl alcohol, polyoxyethylene castor oil, poloxamer and sodium lauryl sulfate, lactose and mannitol.

[0061] In one embodiment, the present disclosure provides a nanocrystal formulation having a particle size D 90 is 50 to 1500 nm, preferably 50 to 1000 nm, preferably 50 to 500 nm, preferably 80 to 300 nm, and more preferably 50 nm, 100 nm, 150 nm, 200 nm, 250 nm, 300 nm, 400 nm, 500 nm, 600 nm, 700 nm, 800 nm, 900 nm, or 1000 nm.

[0062] In one embodiment, the present disclosure provides a particle size range for the ROCK2 inhibitor, preferably D 90The nanocrystal formulation has a size of 5 to 300 μm, preferably 10 to 100 μm, and more preferably 10 to 50 μm.

[0063] In one embodiment, the present disclosure provides that the weight percentage of the ROCK2 inhibitor may be 1%-55%, 4%-50%, 1%-10%, 10%-40%, 10%-35%, 20%-30%, or 30%-40%, based on the total weight of the nanocrystal formulation, and may be 1%, 2%, 3%, 4%, 4.5%, 5%, 6%, 7%, 8%, 9%, 10%, 11%, 12%, 13%, 14%, 15%, 16%, 17%, 18%, 19%, 20%, 21%, 22%, 23%, 24%, 25%, 26%, 27%, 28%, 29%, 30%, 31%, 32%, 33%, 34%, 35%, 36%, 37%, 38%, 39%, 40%, 41%, 42%, 43%, 44%, 45%, 46%, 47%, 48%, 49%, 50%, 51%, 52%, 53%, 54%, 55%, 56%, 57%, 58%, 59%, 60%, 61%, 62%, 63%, 64%, 65%, 66%, 67%, 68%, 69%, 70%, 71%, 72%, 73%, 74%, 75%, 76%, 77%, 78%, 79%, 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, 99%, 100%, 100%, 100%, 100%, 10 In some embodiments, the nanocrystal formulation may be 5%, 10%, 11%, 12%, 13%, 14%, 15%, 15%, 16%, 18%, 19%, 20%, 21%, 22%, 23%, 24%, 25%, 26%, 27%, 28%, 29%, 30%, 31%, 32%, 33%, 34%, 35%, 36%, 37%, 38%, 39%, or 40%.

[0064] In one embodiment, the disclosure relates to a nanocrystal formulation in which the weight percentage of the stabilizer, based on the total weight of the nanocrystal formulation, can be 0.1%-55%, 0.1%-30%, 0.5%-1%, 1%-10%, 10%-20%, or 20%-30%, and can also be 1%, 2%, 2.5%, 3%, 4%, 5%, 6%, 7%, 8%, 9%, 9.5%, 10%, 11%, 12%, 13%, 14%, 15%, 15%, 16%, 18%, 19%, 20%, 21%, 22%, 23%, 24%, 25%, 26%, 27%, 28%, 29%, 30%, 31%, 32%, 33%, 34%, 35%, 36%, 37%, 38%, 39%, or 40%.

[0065] In one embodiment, the present disclosure relates to nanocrystal formulations in which the weight ratio of ROCK2 inhibitor to stabilizer is 1:10-10:1, 1:9-9:1, 1:8-8:1, 1:7-7:1, 1:6-6:1, 1:5-5:1, 1:4-4:1, 1:3-3:1, 1:2-2:1, 1:1, can be 4:1-1:1 or 1:1-1:2, can be 10:1, 9:1, 8:1, 7:1, 6:1, 5:1, 4:1, 3:1, 2:1, 1:1, 1:2, 1:3, 5:4, 5:3, or 5:2.

[0066] In one embodiment, the present disclosure relates to a nanocrystal formulation further comprising an excipient.

[0067] In one embodiment, the present disclosure relates to a nanocrystal formulation, wherein the excipient is selected from one or more of a bulking agent; a wetting agent; a sweetening or flavoring agent; a surfactant; a binder; a disintegrant; a lubricant; a glidant or anti-adherent; a release modifier; a coating agent; an emulsifier; a solubilizer; and a flavoring agent.

[0068] In one embodiment, the present disclosure relates to a nanocrystal formulation, in which the excipient comprises a bulking agent. The bulking agent can improve the material properties of active ingredients, improve adhesion and electrostatic properties, thereby facilitating the subsequent shaping of the composition, such as tableting, capsule filling, etc., and can play an important role in the preparation of solid dosage forms. The bulking agent can also adjust the dissolution rate of the formulation. In the technical solution of the present disclosure, the bulking agent commonly used in the technical field can be selected from, but is not limited to, one or at least two of microcrystalline cellulose, mannitol, lactose, starch, pregelatinized starch, dextrin, calcium phosphate dihydrate, and anhydrous calcium hydrogen phosphate. In some embodiments of the present disclosure, the weight percentage of the bulking agent may be 1%-80%, more specifically, in some embodiments, the weight percentage of the bulking agent may be 20%-70%, 30%-60%, or 50-70%, such as 20%, 25%, 30%, 35%, 40%, 45%, 50%, 55%, 60%, 65%, or 70%, based on the total weight of the nanocrystal formulation. In some specific embodiments of the present disclosure, the bulking agent is selected from one or more of microcrystalline cellulose, lactose, and mannitol, preferably, the bulking agent is mannitol.

[0069] In one embodiment, the present disclosure relates to a nanocrystal formulation, in which the excipient comprises a lubricant. The lubricant facilitates various processing steps, such as ingredient mixing, tableting, etc. For example, the lubricant can make the pressure distribution during tableting uniform, make the tablet density uniform, and reduce the force required to push the tablet out of the die hole. Another possible function of the lubricant is to improve the appearance of the tablet by making the tablet surface smooth and flat. In the technical solution of the present disclosure, the lubricant commonly used in the technical field can be selected from, but not limited to, one or at least two combinations of magnesium stearate, talc powder, fine powder silica gel, sodium stearyl fumarate, glyceryl behenate, and polyethylene glycol, more preferably magnesium stearate. In some embodiments of the present disclosure, the weight percentage of the lubricant based on the total weight of the nanocrystal formulation can be 0.1%-5%, 0.1%-1.5%, or 0.5%-1%, such as 0.1%, 0.2%, 0.3%, 0.4%, 0.5%, 0.6%, 0.7%, 0.8%, 0.9%, 1.0%, 1.1%, 1.2%, 1.3%, 1.4%, or 1.5%, etc. The lubricant is selected from magnesium stearate, talc powder, finely powdered silica gel, sodium stearyl fumarate, glyceryl behenate, and polyethylene glycol.

[0070] In one embodiment, the present disclosure relates to a nanocrystal formulation, wherein the excipient comprises a disintegrant. In the technical solution of the present disclosure, the disintegrant commonly used in the art is selected from one or more of, but not limited to, croscarmellose sodium and crospovidone. In some embodiments of the present disclosure, based on the total weight of the nanocrystal formulation, the weight percentage of the disintegrant may be 0-20%, preferably 0-10%, more preferably 2-10%, such as 3%, 4%, 5%, 6%, 7%, 8%, 9%, or 10%.

[0071] In one embodiment, the present disclosure relates to a nanocrystal formulation, wherein the excipient comprises a glidant. In the technical solution of the present disclosure, the glidant commonly used in the art can be selected from, but not limited to, silicon dioxide, etc. In some embodiments of the present disclosure, based on the total weight of the nanocrystal formulation, the weight percentage of the glidant can be 0-20%, preferably 0-15%, more preferably 2-12%, such as 2%, 2.5%, 3%, 4%, 5%, 6%, 7%, 8%, 9%, 10%, or 11%.

[0072] In one embodiment, the present disclosure relates to a nanocrystal formulation further comprising a solvent. In the technical solution of the present disclosure, the solvent commonly used in the technical field can be selected from, but not limited to, water, and purified water is preferred. In some embodiments of the present disclosure, based on the total weight of the nanocrystal formulation, the weight percentage of the solvent can be 0-99%, preferably 80-99%, more preferably 85%-95%, such as 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, or 95%.

[0073] In one embodiment, the present disclosure relates to a nanocrystal formulation further comprising a bacteriostatic agent. In the technical solution of the present disclosure, the bacteriostatic agent commonly used in the technical field can be selected from one or more of, but not limited to, methylparaben, propylparaben, and the like. In some embodiments of the present disclosure, based on the total weight of the nanocrystal formulation, the weight percentage of the bacteriostatic agent can be 0-5%, preferably 0-1%, more preferably 0.01%-0.5%, such as 0.1%, 0.2%, 0.3%, 0.4%, or 0.5%.

[0074] In one embodiment, the present disclosure relates to a nanocrystal formulation selected from a suspension, tablet, capsule, granules, powder, lozenge, and pill; preferably a suspension, tablet, or capsule.

[0075] In one embodiment, the present disclosure provides a method for producing a pharmaceutical composition comprising: 1-10% ROCK2 inhibitor, preferably 1%, 2%, 3%, 4%, 4.5%, 5%, 6%, 7%, 8%, 9%, or 10% ROCK2 inhibitor, more preferably 4%, 4.5%, or 5% ROCK2 inhibitor; and 1-10% stabilizer, preferably 1%, 1.5%, 2%, 2.5%, 5%, 9%, 9.5%, or 10% stabilizer The present invention relates to a nanocrystal formulation, which is a suspension comprising:

[0076] In one embodiment, the present disclosure relates to a nanocrystal formulation, wherein the nanocrystals are a suspension comprising 11.13 g of ROCK2 inhibitor, 0.77 g of polysorbate 80, 5.00 g of povidone K29 / 32, and 233.10 g of purified water, preferably wherein the particle size of the ROCK2 inhibitor is 50-1000 nm, preferably 50-500 nm, more preferably 50-300 nm.

[0077] In one embodiment, the present disclosure relates to a nanocrystal formulation, wherein the nanocrystals are a suspension comprising 11.13 g of ROCK2 inhibitor, 0.77 g of polysorbate 80, 2.50 g of povidone K29 / 32, and 235.60 g of purified water, preferably wherein the particle size of the ROCK2 inhibitor is 50-1000 nm, preferably 50-500 nm, more preferably 50-300 nm.

[0078] In one embodiment, the present disclosure relates to a nanocrystal formulation, wherein the nanocrystals are a suspension comprising 11.13 g of ROCK2 inhibitor, 0.77 g of polysorbate 80, 2.50 g of hypromellose, and 235.60 g of purified water, preferably wherein the particle size of the ROCK2 inhibitor is 50-1000 nm, preferably 50-500 nm, more preferably 50-300 nm.

[0079] In one embodiment, the present disclosure relates to a nanocrystal formulation, wherein the nanocrystals are a suspension comprising 11.13 g of ROCK2 inhibitor, 0.77 g of polysorbate 80, 5.00 g of hypromellose, and 233.10 g of purified water, preferably wherein the particle size of the ROCK2 inhibitor is 50-1000 nm, preferably 50-500 nm, more preferably 50-300 nm.

[0080] In one embodiment, the present disclosure relates to a nanocrystal formulation, wherein the nanocrystals are a suspension comprising 250.08 g of ROCK2 inhibitor, 17.40 g of polysorbate 80, 111.60 g of povidone K29 / 32, 10.00 g of methylparaben, 1.10 g of propylparaben, and 5189.82 g of purified water, preferably wherein the particle size of the ROCK2 inhibitor is 50-1000 nm, preferably 50-500 nm, more preferably 50-300 nm.

[0081] In one embodiment, the present disclosure relates to a nanocrystal formulation, wherein the nanocrystals are a suspension comprising 55.19 g of ROCK2 inhibitor, 18.61 g of polysorbate 80, 99.25 g of polyoxyethylene castor oil, 2.23 g of methylparaben, 0.25 g of propylparaben, and 1065.09 g of purified water, preferably wherein the particle size of the ROCK2 inhibitor is 50-1000 nm, preferably 50-500 nm, more preferably 50-300 nm, and most preferably 50-150 nm.

[0082] In one embodiment, the present disclosure provides a method for producing a pharmaceutical composition comprising: a 10-30% ROCK2 inhibitor, preferably a 20-30% ROCK2 inhibitor, preferably a 20%, 22%, 25%, 28%, or 30% ROCK2 inhibitor; 1-20% stabilizer, preferably 5-20% stabilizer, more preferably 5%, 8%, 10%, 13%, 15%, 18%, or 20% stabilizer The present invention relates to a nanocrystal formulation, which is a tablet comprising:

[0083] In one embodiment, the present disclosure relates to a nanocrystal formulation, wherein the nanocrystals are a tablet comprising 11.70 g of ROCK2 inhibitor, 3.15 g of polysorbate 80, 5.26 g of lactose, 1.05 g of polyethylene glycol 6000, 16.03 g of mannitol, 4.94 g of silicon dioxide, 2.47 g of sodium lauryl sulfate, 2.96 g of microcrystalline cellulose, 2.96 g of croscarmellose sodium, and 0.32 g of magnesium stearate, preferably wherein the particle size of the ROCK2 inhibitor is 50-1000 nm, preferably 50-500 nm, more preferably 50-300 nm, and most preferably 50-150 nm.

[0084] In one embodiment, the present disclosure relates to a nanocrystal formulation, wherein the nanocrystals are a tablet comprising 32.71 g of ROCK2 inhibitor, 14.81 g of polysorbate 80, 14.71 g of lactose, 2.94 g of polyethylene glycol 6000, 43.63 g of mannitol, 16.00 g of silicon dioxide, 8.00 g of sodium lauryl sulfate, 12.80 g of microcrystalline cellulose, 12.80 g of croscarmellose sodium, and 1.60 g of sodium stearyl fumarate, preferably wherein the particle size of the ROCK2 inhibitor is 50-1000 nm, preferably 50-500 nm, more preferably 50-300 nm, and most preferably 50-150 nm.

[0085] In one embodiment, the present disclosure relates to a nanocrystal formulation, wherein the nanocrystals are a tablet comprising 3.34 g of ROCK2 inhibitor, 0.89 g of polysorbate 80, 1.80 g of lactose, 0.30 g of polyethylene glycol 6000, 0.75 g of silicon dioxide, 0.75 g of sodium lauryl sulfate, 5.82 g of microcrystalline cellulose, 1.20 g of croscarmellose sodium, and 0.15 g of sodium stearyl fumarate, preferably wherein the particle size of the ROCK2 inhibitor is 50-1000 nm, preferably 50-500 nm, more preferably 50-300 nm, and most preferably 50-150 nm.

[0086] In one embodiment, the present disclosure relates to a nanocrystal formulation, wherein the nanocrystals are a tablet comprising 3.34 g of ROCK2 inhibitor, 0.89 g of polysorbate 80, 1.80 g of lactose, 0.30 g of polyethylene glycol 6000, 0.75 g of silicon dioxide, 0.45 g of sodium lauryl sulfate, 4.50 g of microcrystalline cellulose, 1.62 g of pregelatinized starch, 1.20 g of croscarmellose sodium, and 0.15 g of sodium stearyl fumarate, preferably wherein the particle size of the ROCK2 inhibitor is 50-1000 nm, preferably 50-500 nm, more preferably 50-300 nm, and most preferably 50-150 nm.

[0087] In one embodiment, the present disclosure relates to a nanocrystal formulation, wherein the nanocrystals are a tablet comprising 11.14 g of ROCK2 inhibitor, 2.97 g of polysorbate 80, 6.01 g of lactose, 1.00 g of polyethylene glycol 6000, 1.00 g of silicon dioxide, 27.38 g of spray dried mannitol, and 0.50 g of sodium stearyl fumarate, preferably wherein the particle size of the ROCK2 inhibitor is 50-1000 nm, preferably 50-500 nm, more preferably 50-300 nm, and most preferably 50-150 nm.

[0088] In one embodiment, the present disclosure relates to a nanocrystal formulation, wherein the nanocrystals are a tablet comprising 111.20 g of ROCK2 inhibitor, 30.04 g of polysorbate 80, 59.99 g of lactose, 10.00 g of polyethylene glycol 6000, 10.00 g of silicon dioxide, 273.78 g of spray dried mannitol, and 5.00 g of sodium stearyl fumarate, preferably wherein the particle size of the ROCK2 inhibitor is 50-1000 nm, preferably 50-500 nm, more preferably 50-300 nm, and most preferably 50-150 nm.

[0089] In one embodiment, the present disclosure provides a method for producing a pharmaceutical composition comprising: a 10-30% ROCK2 inhibitor, preferably a 20-30% ROCK2 inhibitor, more preferably a 20%, 22%, 25%, 28%, or 30% ROCK2 inhibitor; 10-30% stabilizer, preferably 20-30% stabilizer, more preferably 20%, 22%, 25%, 28%, or 30% stabilizer The present invention relates to a nanocrystal formulation, which is a tablet comprising:

[0090] In one embodiment, the present disclosure relates to a nanocrystal formulation, wherein the nanocrystals are a tablet comprising 22.22 g of ROCK2 inhibitor, 20.00 g of polysorbate 80, 54.80 g of mannitol, 2.00 g of silicon dioxide, and 1.00 g of sodium stearyl fumarate, preferably wherein the particle size of the ROCK2 inhibitor is 50-1000 nm, preferably 50-500 nm, more preferably 50-300 nm, and most preferably 50-150 nm.

[0091] In one embodiment, the present disclosure relates to a nanocrystal formulation, wherein the nanocrystals are a tablet comprising 22.20 g of ROCK2 inhibitor, 10.00 g of polysorbate 80, 20.00 g of povidone K29 / 32, 44.80 g of mannitol, 2.00 g of silicon dioxide, and 1.00 g of sodium stearyl fumarate, preferably wherein the particle size of the ROCK2 inhibitor is 50-1000 nm, preferably 50-500 nm, more preferably 50-300 nm, and most preferably 50-150 nm.

[0092] In one embodiment, the present disclosure relates to a nanocrystal formulation, wherein the nanocrystals are a tablet comprising 22.20 g of ROCK2 inhibitor, 10.00 g of polysorbate 80, 20.00 g of polyethylene glycol 6000, 44.80 g of mannitol, 2.00 g of silicon dioxide, and 1.00 g of sodium stearyl fumarate, preferably wherein the particle size of the ROCK2 inhibitor is 50-1000 nm, preferably 50-500 nm, more preferably 50-300 nm, and most preferably 50-150 nm.

[0093] In one embodiment, the present disclosure relates to a nanocrystal formulation, wherein the nanocrystals are a tablet comprising 22.20 g of ROCK2 inhibitor, 10.00 g of polysorbate 80, 20.00 g of poloxamer 188, 44.80 g of mannitol, 2.00 g of silicon dioxide, and 1.00 g of sodium stearyl fumarate, preferably wherein the particle size of the ROCK2 inhibitor is 50-1000 nm, preferably 50-500 nm, more preferably 50-300 nm, and most preferably 50-150 nm.

[0094] In one embodiment, the present disclosure relates to a nanocrystal formulation, wherein the nanocrystals are a tablet comprising 22.20 g of ROCK2 inhibitor, 10.00 g of polysorbate 80, 20.00 g of polyvinyl alcohol, 44.80 g of mannitol, 2.00 g of silicon dioxide, and 1.00 g of sodium stearyl fumarate, preferably wherein the particle size of the ROCK2 inhibitor is 50-1000 nm, preferably 50-500 nm, more preferably 50-300 nm, and most preferably 50-150 nm.

[0095] In one embodiment, the present disclosure relates to a nanocrystal formulation, wherein the nanocrystals are a tablet comprising 22.20 g of ROCK2 inhibitor, 10.00 g of polysorbate 80, 16.00 g of povidone K29 / 32, 4.00 g of poloxamer 188, 44.80 g of mannitol, 2.00 g of silicon dioxide, and 1.00 g of sodium stearyl fumarate, preferably wherein the particle size of the ROCK2 inhibitor is 50-1000 nm, preferably 50-500 nm, more preferably 50-300 nm, and most preferably 50-150 nm.

[0096] In one embodiment, the present disclosure relates to a nanocrystal formulation, wherein the nanocrystals are a tablet comprising 22.20 g of ROCK2 inhibitor, 10.00 g of polysorbate 80, 4.00 g of povidone K29 / 32, 16.00 g of poloxamer 188, 44.80 g of mannitol, 2.00 g of silicon dioxide, and 1.00 g of sodium stearyl fumarate, preferably wherein the particle size of the ROCK2 inhibitor is 50-1000 nm, preferably 50-500 nm, more preferably 50-300 nm, and most preferably 50-150 nm.

[0097] In one embodiment, the present disclosure relates to a nanocrystal formulation, wherein the nanocrystals are a tablet comprising 22.20 g of ROCK2 inhibitor, 10.00 g of polysorbate 80, 10.00 g of povidone K29 / 32, 10.00 g of poloxamer 188, 44.80 g of mannitol, 2.00 g of silicon dioxide, and 1.00 g of sodium stearyl fumarate, preferably wherein the particle size of the ROCK2 inhibitor is 50-1000 nm, preferably 50-500 nm, more preferably 50-300 nm, and most preferably 50-150 nm.

[0098] In one embodiment, the present disclosure provides a method for producing a pharmaceutical composition comprising: a 10-50% ROCK2 inhibitor, preferably a 20-40% ROCK2 inhibitor, more preferably a 20%, 25%, 30%, 35%, or 40% ROCK2 inhibitor; 10-40% stabilizer, preferably 20-30% stabilizer, more preferably 20%, 22%, 25%, 28%, or 30% stabilizer The present invention relates to a nanocrystal formulation, which is a capsule comprising:

[0099] In one embodiment, the present disclosure relates to a nanocrystal formulation, wherein the nanocrystals are a capsule comprising 22.20 g of ROCK2 inhibitor, 6.00 g of polysorbate 80, 8.00 g of povidone K29 / 32, 4.00 g of poloxamer, and 20.00 g of mannitol, preferably wherein the particle size of the ROCK2 inhibitor is 50-1000 nm, preferably 50-500 nm, more preferably 50-300 nm, and most preferably 50-150 nm.

[0100] In one embodiment, the present disclosure relates to a nanocrystal formulation, wherein the nanocrystals are a capsule comprising 4.45 g of ROCK2 inhibitor, 1.20 g of polysorbate 80, 1.20 g of povidone K32 / 29, 0.80 g of poloxamer 188, and 6.80 g of mannitol, preferably wherein the particle size of the ROCK2 inhibitor is 50-1000 nm, preferably 50-500 nm, more preferably 50-300 nm, and most preferably 50-150 nm.

[0101] In one embodiment, the present disclosure relates to a nanocrystal formulation that is a nanocrystal enteric formulation selected from enteric coated tablets and enteric coated capsules.

[0102] In one embodiment, the present disclosure relates to a nanocrystalline formulation that is a nanocrystalline enteric coated tablet, wherein the enteric coating material is selected from one or more of shellac, polyvinyl alcohol acetate phthalate (PVAP), methacrylic acid copolymers, cellulose and its derivatives (cellulose acetate phthalate (CAP), cellulose acetate trimellitate (CAT), hydroxypropyl cellulose phthalate (HPMCP)), and acrylic resins (EuS100, EuL100).

[0103] In one embodiment, the present disclosure relates to a nanocrystal formulation that is a nanocrystal enteric capsule, wherein the enteric capsule is selected from a gelatin enteric capsule or a hypromellose enteric capsule, and the capsule material composition is selected from one or more of shellac, polyvinyl alcohol acetate phthalate (PVAP), methacrylic acid copolymers, cellulose and its derivatives (cellulose acetate phthalate (CAP), cellulose acetate trimellitate (CAT), hydroxypropyl cellulose phthalate (HPMCP)), and acrylic resins (EuS100, EuL100).

[0104] In one embodiment, the present disclosure provides a method for producing a pharmaceutical composition comprising: 1-20% ROCK2 inhibitor, preferably 5-15% ROCK2 inhibitor, more preferably 5%, 6%, 7%, 8%, 9%, 10%, 11%, 12%, 13%, 14%, or 15% ROCK2 inhibitor; and 1-90% stabilizer, preferably 10-40% stabilizer, more preferably 15-25% stabilizer, for example, 15%, 16%, 17%, 18%, 19%, 20%, 21%, 22%, 23%, 24%, or 25% stabilizer. The present invention relates to a nanocrystal formulation which is a nanocrystal enteric coated tablet comprising:

[0105] In one embodiment, the present disclosure relates to a nanocrystalline enteric coated tablet comprising 5.56 g of ROCK2 inhibitor, 1.50 g of polysorbate 80, 1.00 g of poloxamer, 1.50 g of povidone K29 / 32, 5.00 g of mannitol, 1.00 g of silicon dioxide, 4.00 g of crospovidone, 29.94 g of microcrystalline cellulose, 0.50 g of magnesium stearate, and 5 g of film-coated premix (enteric type), preferably, the particle size of the ROCK2 inhibitor is 50-1000 nm, preferably 50-500 nm, more preferably 50-300 nm, and most preferably 50-150 nm.

[0106] In one embodiment, the present disclosure relates to a nanocrystalline enteric coated tablet comprising 5.56 g of ROCK2 inhibitor, 1.50 g of polysorbate 80, 1.00 g of poloxamer, 1.50 g of povidone K29 / 32, 38.94 g of mannitol, 1.00 g of silicon dioxide, 0.50 g of magnesium stearate, and 5 g of film-coated premix (enteric type), preferably wherein the particle size of the ROCK2 inhibitor is 50-1000 nm, preferably 50-500 nm, more preferably 50-300 nm, and most preferably 50-150 nm.

[0107] In one embodiment, the present disclosure provides a method for producing a pharmaceutical composition comprising: 5-30% ROCK2 inhibitor, preferably 10-20% ROCK2 inhibitor, more preferably 10%, 11%, 12%, 13%, 14%, 15%, 16%, 17%, 18%, 19%, or 20% ROCK2 inhibitor; and 10-50% stabilizer, preferably 20-40% stabilizer, more preferably 30-40% stabilizer, for example 30%, 31%, 32%, 33%, 34%, 35%, 36%, 37%, 38%, 39%, or 40% stabilizer. The present invention relates to a nanocrystal formulation, which is a nanocrystal enteric capsule comprising:

[0108] In one embodiment, the present disclosure relates to a nanocrystalline formulation comprising 11.1 g of ROCK2 inhibitor, 3.0 g of polysorbate 80, 2.0 g of poloxamer, 3.0 g of povidone K29 / 32, 20.0 g of mannitol, 2.00 g of silicon dioxide, 8.0 g of crospovidone, 29.9 g of microcrystalline cellulose, 1.0 g of magnesium stearate, and an enteric capsule, preferably wherein the particle size of the ROCK2 inhibitor is 50-1000 nm, preferably 50-500 nm, more preferably 50-300 nm, and most preferably 50-150 nm.

[0109] In some embodiments of the present disclosure, in addition to a compound of Formula (I), (II), (III), or Formula (IV), or a pharma- ceutically acceptable salt or hydrate thereof, the ROCK2 inhibitor may also optionally include at least one other compound having a synergistic therapeutic effect with said compound.

[0110] In one embodiment, the present disclosure relates to a method for preparing the nanocrystal formulation, comprising milling a ROCK2 inhibitor and a stabilizer.

[0111] In one embodiment, the present disclosure provides a method for the preparation of a medicament for use in a pharmaceutical composition comprising the steps of: (a) grinding the medicament with a weight ratio of ROCK2 inhibitor to stabilizer of 1:15-15:1, 1:14-14:1, 1:13-13:1, 1:12-12:1, 1:11-11:1, 1:10-10:1, 1:9-9:1, 1:8-8:1, 1:7-7:1, 1:6-6:1, 1:5-5:1, 1:4-4:1, 1:3-3: Preferably, the weight ratio of ROCK2 inhibitor to stabilizer during grinding is 15:1 to 2:1, more preferably 15:1, 14:1, 13:1, 12:1, 11:1, 10:1, 10:3, 9:1, 8:1, 7:1, 6:1, 5:1, 4:1, 3:1, or 2:1.

[0112] In one embodiment, the present disclosure relates to a method for preparing the nanocrystal formulation, wherein the grinding media is selected from porcelain balls, glass balls, zirconia beads, steel balls, and ice beads; preferably, the grinding media is zirconia beads.

[0113] In one embodiment, the present disclosure relates to a method for preparing the nanocrystal formulation, wherein the particle size of the grinding media is 0.1-1 mm, preferably 0.1-0.5 mm, and more preferably 0.2 mm.

[0114] In one embodiment, the present disclosure relates to a method for preparing the above nanocrystal formulation, wherein the milling time is 0.1 to 6 hours, preferably 0.5 to 6 hours, preferably 4 to 6 hours, and more preferably 10 minutes, 20 minutes, 30 minutes, 40 minutes, 1 hour, 1.5 hours, 2 hours, 2.5 hours, 3 hours, 3.5 hours, 4 hours, 4.5 hours, 5 hours, 5.5 hours, or 6 hours.

[0115] In one embodiment, the present disclosure relates to a method for preparing the nanocrystal formulation, wherein the grinding speed is 1000-6000 rpm, preferably 1500 rpm-4500 rpm, and more preferably 1500 rpm, 2000 rpm, 2500 rpm, 3000 rpm, 3500 rpm, 4000 rpm, 4500 rpm, 5000 rpm, 5500 rpm, or 6000 rpm.

[0116] In one embodiment, the present disclosure relates to a method for preparing said nanocrystal formulation, wherein the loading of the grinding beads is 50%-95%, preferably 70%-90%, more preferably 70%, 80%, or 90%.

[0117] In one embodiment, the present disclosure relates to a method for preparing the nanocrystal formulation, further comprising a pre-grinding step prior to grinding of the ROCK2 inhibitor and the stabilizer.

[0118] In one embodiment, the present disclosure relates to a method for preparing the above nanocrystal formulation, wherein the pre-grinding speed is 3000-6000 rpm, preferably 3000 rpm, 3500 rpm, 4000 rpm, 4500 rpm, 5000 rpm, 5500 rpm, or 6000 rpm, and more preferably 4000 rpm; and the pre-grinding time is 1-30 minutes, preferably 2-20 minutes, and preferably 3 minutes, 4 minutes, 5 minutes, 6 minutes, 8 minutes, 10 minutes, 12 minutes, 15 minutes, 18 minutes, or 20 minutes, and more preferably 5 minutes.

[0119] In one embodiment, the present disclosure relates to a method for preparing the nanocrystal formulation, wherein stabilizers and / or excipients may be added optionally after milling.

[0120] In one embodiment, the disclosure relates to a method for preparing the nanocrystal formulation, wherein the stabilizer added after milling is selected from one or more of polysorbates, povidone, polyoxyethylene fatty acid esters, polyethylene glycol, polyvinyl alcohol, hydroxypropyl cellulose, hydroxypropyl methylcellulose, polyvinylpyrrolidone, poloxamer, sodium lauryl sulfate, sodium docusate, polyethylene glycol 15-hydroxystearate, polyoxyethylene castor oil, copovidone, lactose, and mannitol.

[0121] In one embodiment, the present disclosure relates to a method for preparing the nanocrystal formulation, wherein the stabilizer added after milling is selected from one or more of povidone K29 / 32, poloxamer 188, polyvinyl alcohol, lactose, and mannitol.

[0122] In one embodiment, the present disclosure relates to a method for preparing the above nanocrystal formulation, wherein the stabilizer added after milling is a mixture of povidone K29 / 32 and poloxamer 188, and the mixing ratio of the two is 1:10-10:1, preferably 1:9-9:1, preferably 1:8-8:1, preferably 1:7-7:1, preferably 1:6-6:1, preferably 1:5-5:1, preferably 1:4-4:1, preferably 1:3-3:1, preferably 1:2-2:1, preferably 1:1; more preferably, the mixing ratio of the two is 1:4, 4:1, or 1:1.

[0123] In one embodiment, the present disclosure relates to a method for preparing the nanocrystal formulation, comprising the steps of:

[0124] The active ingredient, a part of the stabilizer, and purified water are ground in a nano-pulverizer. After grinding, a nano-suspension is obtained, followed by adding stabilizers and / or bulking agents to obtain a nano-crystal suspension. Preferably, after grinding to obtain a nano-suspension, the mixture is solidified using spray drying or freeze drying to obtain a solid nano-crystal mixture, followed by adding bulking agents, mixing, adding lubricants, and then performing a molding process. In a specific implementation, the molding process may include steps such as granulation, optional granulation, and tableting or capsule filling. The important process in this disclosure is the nano-pulverization process. When the average particle size of the active ingredient reaches 50 nm to 1000 nm, more preferably 80 nm to 500 nm, and more preferably 80 nm to 300 nm, the grinding is completed. It should be noted that the mixing and subsequent molding process are common processes and operations in the technical field, and the present disclosure is not specifically limited herein.

[0125] In the process of preparing ROCK2 inhibitor nanocrystals, the inventors studied the effects of the grinding speed, the loading amount of grinding beads, the sample volume, and the grinding time on the particle size of the ROCK2 inhibitor. The results show that the particle size of the ROCK2 inhibitor gradually decreases with the increase of the grinding time, but the rate of decrease gradually becomes slower; as the grinding speed increases, the grinding efficiency increases; when the rotation speed is the same, the grinding efficiency increases with the loading amount of the grinding beads increasing. The sample volume has no significant effect on the grinding efficiency. The research results of the grinding parameters and particle size are shown in Table 8. Based on this, in some specific embodiments of the present disclosure, in the grinding process, 0.2 mm zirconia grinding beads are used, and the loading amount of the grinding beads is 50%-95%, preferably 70%-90%. The grinding speed is 1000 rpm-4500 rpm, preferably 1500 rpm-3500 rpm. The grinding time is 2 hours-6 hours, preferably 4 hours-5 hours.

[0126] In some embodiments of the present disclosure, when the nanocrystal formulation composition includes a lubricant, the milled nanosuspension must be spray-dried or freeze-dried to remove water, and then mixed with an optional bulking agent, either as is or after milling, followed by: A: It is mixed with all the lubricants and then the molding process is carried out; or B: It is first mixed with a portion of the lubricant, then granulated and optionally sized, then the remaining portion of the lubricant is added, followed by other forming operations (such as tableting or capsule filling).

[0127] In one embodiment, the present disclosure relates to a method for preventing, reducing, and / or treating idiopathic pulmonary fibrosis, fatty liver disease and / or steatohepatitis, graft-versus-host disease after hematopoietic stem cell transplantation, or viral infection, comprising administering to a subject a therapeutically effective amount of the nanocrystal formulation or a nanocrystal formulation prepared by the method described above, preferably the method is a method for preventing, reducing, and / or treating fatty liver disease and / or steatohepatitis, preferably the fatty liver disease is alcoholic fatty liver disease (ALFD) or non-alcoholic fatty liver disease (NALFD), the steatohepatitis is alcoholic steatohepatitis (ASH) or non-alcoholic steatohepatitis (NASH), and the hematopoietic stem cell transplantation is an allogeneic hematopoietic stem cell transplant. and the disease caused by the coronavirus is Middle East Respiratory Syndrome, Severe Acute Respiratory Syndrome, or COVID-19, preferably, the coronavirus is Severe Acute Respiratory Syndrome coronavirus 2 (SARA-CoV-2 or 2019-nCoV), and the disease caused by the coronavirus is COVID-19.

[0128] In one embodiment, the disclosure relates to a nanocrystal formulation as described above or a nanocrystal formulation prepared by the method as described above for use in the prevention, reduction, and / or treatment of idiopathic pulmonary fibrosis, fatty liver disease and / or steatohepatitis, graft-versus-host disease following hematopoietic stem cell transplantation, or viral infection, preferably, the nanocrystal formulation is used for the prevention, reduction, and / or treatment of fatty liver disease and / or steatohepatitis, preferably, the fatty liver disease is alcoholic fatty liver disease (ALFD) or non-alcoholic fatty liver disease (NALFD), the steatohepatitis is alcoholic steatohepatitis (ASH) or non-alcoholic steatohepatitis (NASH), and the hematopoietic stem cell transplant is allogeneic hematopoietic stem cell transplant; The graft-versus-host disease is acute graft-versus-host disease or chronic graft-versus-host disease, and the viral infection is a coronavirus infection, preferably, the coronavirus is selected from SARA-CoV, SARA-CoV-2, MERS-CoV, HCoV-229E, HCoV-NL63, HCoV-OC43, and HCoV-HKU1, and preferably, the disease caused by the coronavirus is Middle East Respiratory Syndrome, Severe Acute Respiratory Syndrome, or COVID-19, preferably, the coronavirus is Severe Acute Respiratory Syndrome Coronavirus 2 (SARA-CoV-2 or 2019-nCoV), and the disease caused by the coronavirus is COVID-19.

[0129] In one embodiment, the disclosure relates to the use of the nanocrystal formulation or a nanocrystal formulation prepared by the method described above in the manufacture of a medicament for preventing, reducing, and / or treating idiopathic pulmonary fibrosis, fatty liver disease and / or steatohepatitis, graft-versus-host disease after hematopoietic stem cell transplantation, or viral infection, preferably, said use is in the manufacture of a medicament for preventing, reducing, and / or treating fatty liver disease and / or steatohepatitis, preferably, said fatty liver disease is alcoholic fatty liver disease (ALFD) or non-alcoholic fatty liver disease (NALFD), said steatohepatitis is alcoholic steatohepatitis (ASH) or non-alcoholic steatohepatitis (NASH), and said hematopoietic stem cell transplantation is an allogeneic hematopoietic stem cell transplant. and wherein the graft-versus-host disease is acute graft-versus-host disease or chronic graft-versus-host disease, and the viral infection is a coronavirus infection, preferably wherein the coronavirus is selected from SARA-CoV, SARA-CoV-2, MERS-CoV, HCoV-229E, HCoV-NL63, HCoV-OC43, and HCoV-HKU1, and preferably wherein the disease caused by the coronavirus is Middle East Respiratory Syndrome, Severe Acute Respiratory Syndrome, or COVID-19, preferably wherein the coronavirus is Severe Acute Respiratory Syndrome Coronavirus 2 (SARA-CoV-2 or 2019-nCoV), and wherein the disease caused by the coronavirus is COVID-19.

[0130] In order to make the technical problems to be solved, technical solutions and beneficial effects of the present disclosure clearer, the present disclosure will be further described below with specific examples. In the following examples, unless otherwise specified, the specific conditions of the test methods are usually carried out according to conventional conditions or conditions recommended by manufacturers, and the raw materials and reagents are all commercially available or prepared using public information. EXAMPLES

[0131] The active ingredients (API) used in the following comparative examples, examples and tests are all compounds of formula (IV) above.

[0132] I. Preparation of Comparative Examples Comparative Example 1 Formulation: [Table 1]

[0133] Preparation method: The amounts of API hydrochloride, lactose, microcrystalline cellulose, and croscarmellose sodium in the formulation were weighed, mixed, and dry granulated. After granulation, magnesium stearate was added, mixed, and tableted.

[0134] Comparative Example 2 Formulation: [Table 2]

[0135] Preparation method: First, the API hydrochloride is micronized and 90 The API hydrochloride with a particle size of about 2 μm was obtained. The amounts of micronized API hydrochloride, lactose, microcrystalline cellulose, and croscarmellose sodium in the formulation were weighed, mixed, and dry granulated. After sieving, magnesium stearate was added, mixed, and compressed into tablets.

[0136] Comparative Example 3 Formulation: [Table 3]

[0137] Preparation method: The amounts of API hydrochloride, silica, pregelatinized starch 1500, and microcrystalline cellulose in the formulation, as well as half of the amounts of crospovidone and magnesium stearate, were weighed, mixed, and dry granulated. After granulation, the other half of the amounts of crospovidone and magnesium stearate were added, mixed, and tableted.

[0138] II. Preparation of Nanocrystal Formulations (Suspensions) Example 1 Formulation: [Table 4]

[0139] Preparation method: The amounts of API hydrochloride, polysorbate 80, and some purified water in the formulation were weighed out. The mixture was milled using a nano mill (0.2 mm milling beads, 70% loading) at a milling speed of 1500 rpm and a milling time of 10 minutes to obtain a nano suspension with an average particle size of 1868 nm. A solution of Povidone K29 / 32 was added to the nano suspension and the mixture was diluted to 250 g to obtain a final product with a concentration of 40 mg / mL (based on free base) and a particle size of 1875 nm.

[0140] Example 2 Referring to the formulation of Example 1, a nanosuspension with an average particle size of 1006 nm was obtained by using a milling speed of 1500 rpm and a milling time of 20 minutes. Then, a solution of Povidone K29 / 32 was added to the nanosuspension and the mixture was diluted to 250 g to obtain a final product with a concentration of 40 mg / mL (based on the free base) and a particle size of 1080 nm.

[0141] Example 3 Referring to the formulation of Example 1, a nanosuspension with an average particle size of 512 nm was obtained by using a milling speed of 1500 rpm and a milling time of 40 minutes. A solution of Povidone K29 / 32 was then added to the nanosuspension and the mixture was diluted to 250 g to obtain a final product with a concentration of 40 mg / mL (based on the free base) and a particle size of 540 nm.

[0142] Example 4 Formulation: [Table 5]

[0143] Preparation method: The amounts of API hydrochloride, polysorbate 80, and some purified water in the formulation were weighed out. The mixture was milled using a nano mill (0.2 mm milling beads, 70% loading) at a milling speed of 1500 rpm and a milling time of 6 hours to obtain a nano suspension with an average particle size of 268 nm. A solution of Povidone K29 / 32 was added to the nano suspension and the mixture was diluted to 250 g to obtain a final product with a concentration of 40 mg / mL (based on free base) and a particle size of 300 nm.

[0144] Example 5 Formulation: [Table 6]

[0145] Preparation method: The amounts of API hydrochloride, polysorbate 80, and some purified water in the formulation were weighed out. The mixture was milled using a nano mill (0.2 mm milling beads, 70% loading) at a milling speed of 1500 rpm and a milling time of 6 hours to obtain a nano suspension with an average particle size of 268 nm. A solution of Povidone K29 / 32 was added to the nano suspension and the mixture was diluted to 250 g to obtain a final product with a concentration of 40 mg / mL (based on free base) and a particle size of 302 nm.

[0146] Example 6 Formulation: [Table 7]

[0147] Preparation method: The amounts of API hydrochloride, polysorbate 80, and some purified water in the formulation were weighed out. The mixture was milled using a nano mill (0.2 mm milling beads, 70% loading) at a milling speed of 1500 rpm and a milling time of 6 hours to obtain a nano suspension with an average particle size of 268 nm. A solution of hypromellose was added to the nano suspension and the mixture was diluted to 250 g to obtain a final product with a concentration of 40 mg / mL (based on free base) and a particle size of 336 nm.

[0148] Example 7 Formulation: [Table 8]

[0149] Preparation method: The amounts of API hydrochloride, polysorbate 80, and some purified water in the formulation were weighed out. The mixture was milled using a nano mill (0.2 mm milling beads, 70% loading) at a milling speed of 1500 rpm and a milling time of 6 hours to obtain a nano suspension with an average particle size of 268 nm. A solution of hypromellose was added to the nano suspension and the mixture was diluted to 250 g to obtain a final product with a concentration of 40 mg / mL (based on free base) and a particle size of 442 nm.

[0150] Example 8 Formulation: [Table 9]

[0151] Preparation method: The amounts of API hydrochloride, polysorbate 80, and some purified water in the formulation were weighed out. The mixture was milled using a nano mill (0.2 mm milling beads, 70% loading) at a milling speed of 1500 rpm and a milling time of 4 hours to obtain a nano suspension with an average particle size of 281 nm. The previously prepared solutions of methylparaben, propylparaben, and povidone K29 / 32 were added to the nano suspension. The mixture was diluted to 5580 g to obtain a final product with a concentration of 40 mg / mL (based on free base) and a particle size of 283 nm.

[0152] Example 9 Formulation: [Table 10]

[0153] Preparation method: The amounts of API hydrochloride, polysorbate 80, and some purified water in the formulation were weighed out. The mixture was milled using a nano mill (0.2 mm milling beads, 90% loading) at a milling speed of 3000 rpm and a milling time of 4 hours to obtain a nano suspension with an average particle size of 100 nm. The previously prepared solutions of methylparaben, propylparaben, and polyoxyethylene castor oil were added to the nano suspension. The mixture was diluted to 1240.62 g to obtain a final product with a concentration of 40 mg / mL (based on the free base) and a particle size of 106 nm.

[0154] III. Preparation of Nanocrystal Formulations (Tablets) Example 10 Formulation: [Table 11]

[0155] Preparation method: The amounts of API hydrochloride, a part of polysorbate 80 (API free base: polysorbate 80 = 1:0.1), and a part of purified water in the formulation were weighed out. The mixture was milled using a nano mill (0.2 mm milling beads, 90% loading) at a milling speed of 3000 rpm and a milling time of 4 hours to obtain a nano suspension with an average particle size of 105 nm. The previously prepared solutions of polysorbate 80, polyethylene glycol 6000, and lactose were added to the nano suspension (API free base: polysorbate 80: lactose: polyethylene glycol 6000 = 1:0.3:0.5:0.1). The mixture was spray-dried at an inlet temperature of 120 °C and a spray speed of 40 rpm to obtain a spray-dried powder. Mannitol, silicon dioxide, sodium lauryl sulfate, microcrystalline cellulose, croscarmellose sodium, and magnesium stearate were added to the appropriate amount of the above spray-dried powder, mixed, and tableted as is. The tablet weight was approximately 1 g (specification: 200 mg, based on anhydrous API).

[0156] Example 11 Formulation: [Table 12]

[0157] Preparation method: The amounts of API hydrochloride, a part of polysorbate 80 (API free base: polysorbate 80 = 1:0.1), and a part of purified water in the formulation were weighed out. The mixture was milled using a nano mill (0.2 mm milling beads, 90% loading) at a milling speed of 3000 rpm and a milling time of 4 hours to obtain a nano suspension with an average particle size of 105 nm. The previously prepared solutions of polysorbate 80, polyethylene glycol 6000, a part of mannitol, and lactose were added to the nano suspension (API free base: polysorbate 80: lactose: mannitol: polyethylene glycol 6000 = 1:0.3:0.5:0.3:0.1). The mixture was spray-dried at an inlet temperature of 120 °C and a spray speed of 40 rpm to obtain a spray-dried powder. An appropriate amount of silicon dioxide was added to the above spray-dried powder, mixed, and passed through a 35 mesh sieve. Sodium lauryl sulfate, mannitol, microcrystalline cellulose, and croscarmellose sodium (inner) were then added to the powder and mixed. The mixture was wet granulated with a 10% aqueous solution of sodium lauryl sulfate, dried, and sieved. The granules were then mixed with croscarmellose sodium (outer) and sodium stearyl fumarate and compressed into tablets. Tablet weight was approximately 1 g (specification: 200 mg, based on anhydrous API).

[0158] Example 12 Formulation: [Table 13]

[0159] Preparation method: The amounts of API hydrochloride, polysorbate 80, and some purified water in the formulation were weighed out. The mixture was milled using a nano mill (0.2 mm milling beads, 90% loading) at a milling speed of 4500 rpm and milling time of 6 hours to obtain a nanocrystal suspension with a particle size of 58 nm. The previously prepared polyethylene glycol 6000 and lactose solution was added to the nanocrystal suspension, and the mixture was spray-dried. The inlet temperature was 120°C and the spray speed was 40 rpm to obtain a spray-dried powder. An appropriate amount of silicon dioxide was added to the above spray-dried powder, mixed, and passed through a 35 mesh sieve. Then microcrystalline cellulose, croscarmellose sodium (inner), and sodium stearyl fumarate (inner) were added to the powder, mixed, dry granulated, and sieved through a 24 mesh sieve. Then croscarmellose sodium (outer) and sodium stearyl fumarate (outer) were added to the granules, mixed, and tableted. The tablet weight was approximately 1 g (specification: 200 mg, based on anhydrous API).

[0160] Example 13 Formulation: [Table 14]

[0161] Preparation method: The amounts of API hydrochloride, polysorbate 80, and some purified water in the formulation were weighed out. The mixture was milled using a nano mill (0.2 mm milling beads, 90% loading) at a milling speed of 4500 rpm and milling time of 6 hours to obtain a nanocrystal suspension with a particle size of 58 nm. The previously prepared solution of polyethylene glycol 6000 and lactose was added to the nanocrystal suspension, and the mixture was spray-dried. The inlet temperature was 120°C and the spray speed was 40 rpm to obtain a spray-dried powder. An appropriate amount of silicon dioxide was added to the above spray-dried powder, mixed, and passed through a 35 mesh sieve. Then microcrystalline cellulose, pregelatinized starch, croscarmellose sodium (inner), and sodium stearyl fumarate (inner) were added to the powder, dry granulated, and sieved through a 24 mesh sieve. Then croscarmellose sodium (outer) and sodium stearyl fumarate (outer) were added to the granules, mixed, and compressed into tablets. The tablet weight was approximately 1 g (specification: 200 mg, based on anhydrous API).

[0162] Example 14 Formulation: [Table 15]

[0163] Preparation method: The amounts of API hydrochloride, polysorbate 80, and some purified water in the formulation were weighed out. The mixture was milled using a nano mill (0.2 mm milling beads, 90% loading) at a milling speed of 3000 rpm and a milling time of 6 hours to obtain a nanocrystal suspension with a particle size of 89 nm. The previously prepared polyethylene glycol 6000 and lactose solution was added to the nanocrystal suspension, and the mixture was spray-dried. The inlet temperature was 120°C and the spray speed was 40 rpm to obtain a spray-dried powder. Silicon dioxide, spray-dried mannitol, and sodium stearyl fumarate were added to the appropriate amount of the above spray-dried powder, mixed, and tableted. The tablet weight was about 1 g (specification: 200 mg, based on anhydrous API).

[0164] Example 15 Formulation: [Table 16]

[0165] Preparation method: The amounts of API hydrochloride, polysorbate 80, and some purified water in the formulation were weighed out. The mixture was milled using a nano mill (0.2 mm milling beads, 90% loading) at a milling speed of 3000 rpm and a milling time of 4 h to obtain a nanocrystal suspension with a particle size of 128 nm. The previously prepared solutions of polyethylene glycol 6000 and lactose were added to the nanocrystal suspension. The mixture was freeze-dried (the solid content of the sample solution was about 20%), i.e., pre-frozen at -40 °C for 5 h, primary dried at -5 °C for 13 h at a vacuum of 0.18 mbar, and secondary dried at 10-15 °C for 16 h at a vacuum of 0.18 mbar. After the freeze-drying was completed, the sample was milled and sized using a milling sieve. Silicon dioxide, spray-dried mannitol, and sodium stearyl fumarate (internal) were added to the sample and mixed. The mixture was dry granulated, sodium stearyl fumarate was added (external), mixed and compressed into tablets, the tablet weight was approximately 1 g (specification: 200 mg, based on anhydrous API).

[0166] Example 16 Formulation: [Table 17]

[0167] Preparation method: The amounts of API hydrochloride, a portion of polysorbate 80, and a portion of purified water in the formulation were weighed out (API free base: polysorbate 80 = 1:0.3). The mixture was milled using a nano mill (0.2 mm milling beads, 90% loading) at a milling speed of 3000 rpm and a milling time of 4.5 h to obtain a nanocrystal suspension with a particle size of 102 nm. The previously prepared polysorbate 80 and mannitol solution was then added to the nanocrystal suspension. The mixture was freeze-dried (the solid content of the sample solution was about 10%), i.e., pre-frozen at -40 °C for 5 h, primary dried at -5 °C for 13 h at a vacuum of 0.18 mbar, and secondary dried at 10-15 °C for 16 h at a vacuum of 0.18 mbar. After freeze-drying, the sample was milled and sized using a milling sieve. Silicon dioxide, spray-dried mannitol, and sodium stearyl fumarate were added to the sample, mixed, and tableted. The tablet weight was approximately 1 g (specification: 200 mg, based on anhydrous API).

[0168] Examples 17 to 23 Referring to the preparation method of Example 16, the milling formulation was API free base: polysorbate 80 = 1:0.3. After milling, the stabilizer in the table below was added to the milling liquid and dispersed evenly, and the mixture was freeze-dried. After drying, a bulking agent, lubricant, etc. were added according to the formulation in the table below to obtain Examples 17 to 23. The particle size of each Example after milling is recorded in Table 7. [Table 18]

[0169] IV. Preparation of Nanocrystal Formulations (Capsules) Example 24 Formulation: [Table 19]

[0170] Preparation method: The amounts of API hydrochloride, polysorbate 80, and purified water in the formulation were weighed out (API free base: polysorbate 80 = 1:0.3). The mixture was milled using a nano mill (0.2 mm grinding beads, 90% loading) at a milling speed of 3000 rpm and a milling time of 5 hours to obtain a nanocrystal suspension with an average particle size of 93 nm. The previously prepared solutions of povidone K29 / 32, poloxamer 188, and mannitol were then added to the nanocrystal suspension to obtain the final sample with the ratio of API free base: polysorbate 80: povidone K29 / 32: poloxamer 188: mannitol = 1:0.3:0.4:0.2:1. The samples were freeze-dried (solid content of the sample solution was approximately 10%), i.e., pre-frozen at -40°C for 5 h, primary dried at -5°C for 13 h under 0.18 mbar vacuum, and secondary dried at 10-15°C for 16 h under 0.18 mbar vacuum. After freeze-drying was completed, the samples were passed through a 40 mesh sieve and 301.2 mg of freeze-dried powder was weighed out and filled directly into capsules with a standard of 100 mg.

[0171] Example 25 Formulation: [Table 20] Preparation method: The amounts of API hydrochloride, polysorbate 80, and purified water in the formulation were weighed out (API free base: polysorbate 80 = 1:0.3). The mixture was milled using a nano mill (0.2 mm milling beads, 90% loading) at a milling speed of 3000 rpm and a milling time of 5 hours to obtain a nanocrystal suspension with an average particle size of 103 nm. The previously prepared solutions of povidone K29 / 32, poloxamer 188, and mannitol were then added to the nanocrystal suspension to obtain the final sample with API free base: polysorbate 80: povidone K29 / 32: poloxamer 188: mannitol = 1:0.3:0.3:0.2:1.7. The samples were freeze-dried (solid content of the sample solution was approximately 10%), i.e., pre-frozen at -40°C for 5 h, primary dried at -5°C for 13 h under 0.18 mbar vacuum, and secondary dried at 10-15°C for 16 h under 0.18 mbar vacuum. After freeze-drying was completed, the samples were passed through a 40 mesh sieve and 361 mg of freeze-dried powder was weighed out and filled directly into capsules with a standard of 100 mg.

[0172] V. Preparation of Enteric Coated Nanocrystal Formulations (Tablets or Capsules) Example 26 Enteric Coated Nanocrystal Tablets Formulation: [Table 21]

[0173] Preparation method: The amounts of API hydrochloride, polysorbate 80, and some purified water in the formulation were weighed out. The mixture was milled using a nano mill (0.2 mm milling beads, 90% loading) at a milling speed of 3000 rpm and a milling time of 6 h to obtain a nanocrystal suspension with a particle size of 109 nm. The previously prepared solutions of poloxamer, povidone K29 / 32, and mannitol were then added to the nanocrystal suspension. The mixture was freeze-dried (the solid content of the sample solution was about 20%), i.e., pre-frozen at -40 °C for 5 h, primary dried at -5 °C for 13 h at a vacuum of 0.18 mbar, and secondary dried at 10-15 °C for 16 h at a vacuum of 0.18 mbar. After freeze-drying, the sample was milled and sized using a milling sieve. Silicon dioxide, microcrystalline cellulose, crospovidone, and magnesium stearate were added to the sample, mixed, and tableted. The tablet weight was about 1 g (specification: 100 mg, based on anhydrous API). For enteric coating, a coating material with a solid content of 20% was prepared using a film coating premix (enteric type), and coating was performed with stirring. The inlet temperature was 45°C, and the air flow rate was 200 m 3 / min, tablet temperature was 31°C, rotation speed of the coating pan was 15-18 rpm, spray rate of the coating liquid was 5 g / min, and weight gain due to coating was 9.9%.

[0174] Example 27 Enteric Coated Nanocrystal Tablets Formulation: [Table 22]

[0175] Preparation method: The amounts of API hydrochloride, polysorbate 80, and some purified water in the formulation were weighed out. The mixture was milled using a nano mill (0.2 mm milling beads, 90% loading) at a milling speed of 3000 rpm and a milling time of 6 h to obtain a nanocrystal suspension with a particle size of 109 nm. Then the previously prepared solution of poloxamer, povidone K29 / 32, and mannitol (mannitol accounted for 15%) was added to the nanocrystal suspension. The mixture was freeze-dried (the solid content of the sample solution was about 20%), i.e., pre-frozen at -40 °C for 5 h, primary dried at -5 °C for 13 h at a vacuum of 0.18 mbar, and secondary dried at 10-15 °C for 16 h at a vacuum of 0.18 mbar. After freeze-drying, the sample was milled and sized using a milling sieve. Silicon dioxide, mannitol (33.94 g), and magnesium stearate were added to the sample, mixed, and tableted. The tablet weight was about 1 g (specification: 100 mg, based on anhydrous API). For enteric coating, a coating material with a solid content of 20% was prepared using a film coating premix (enteric type), and coating was performed with stirring. The inlet temperature was 48°C, and the air flow rate was 200 m 3 / min, tablet temperature was 36°C, rotation speed of the coating pan was 15-20 rpm, spray rate of the coating liquid was 5 g / min, and weight gain due to coating was 10.0%.

[0176] Example 28 Enteric Nanocrystal Capsules Formulation: [Table 23]

[0177] Preparation method: The amounts of API hydrochloride, polysorbate 80, and some purified water in the formulation were weighed out. The mixture was milled using a nano mill (0.2 mm milling beads, 90% loading) at a milling speed of 3000 rpm and a milling time of 6 h to obtain a nanocrystal suspension with a particle size of 109 nm. Then the previously prepared solution of poloxamer, povidone K29 / 32, and mannitol (mannitol accounted for 15%) was added to the nanocrystal suspension. The mixture was freeze-dried (the solid content of the sample solution was about 20%), i.e., pre-frozen at -40 °C for 5 h, primary dried at -5 °C for 13 h at a vacuum of 0.18 mbar, and secondary dried at 10-15 °C for 16 h at a vacuum of 0.18 mbar. After freeze-drying, the sample was milled and sized using a milling and sieving machine. Silicon dioxide, microcrystalline cellulose, mannitol (15 g), crospovidone, and magnesium stearate (0.25 g) were added to the sample, mixed, dry granulated, and sieved through a 24 mesh screen. Magnesium stearate (0.25 g) was added and mixed. The sample was hand-filled into enteric coated capsules No. 00 at a fill weight of 800 mg (specification: 100 mg, based on anhydrous API).

[0178] Drug Elution Test Test I Tablet dissolution tests of Comparative Examples 1 to 3 1.Elution method: 1) Method: Dissolution rate measurement method (Chinese Pharmacopoeia 2020 Edition, Part 4 General Provisions "0931 Dissolution and Release Rate Measurement Method" 2 (Paddle Method)); 2) Dissolution solvent: purified water, 0.3 SDS aqueous solution, 0.5% SDS aqueous solution, 0.8% SDS aqueous solution, 1.0% SDS aqueous solution, pH 2.0 hydrochloric acid solution + 1.0% SDS, pH 4.5 acetate solution + 1.0% SDS, pH 6.8 phosphate solution + 1.0% SDS, 3% Tween solution; 900 ml; 3) Rotation speed: 75 rpm; 4) Sampling time: 5min, 15min, 30min, 45min, 60min, 90min, 120min, 180min; 5) Detection method: high performance liquid chromatography, detection wavelength: 254 nm; 6) Preparation of test solutions: 5 ml of solution was taken at each time point and filtered.

[0179] Preparation of standard sample solution: Approximately 25 mg of standard sample was accurately weighed and placed in a 100 mL volumetric flask. Approximately 2 mL of DMSO was added to the volumetric flask. The standard sample was dissolved by ultrasonication. The mixture was then diluted to the mark with the corresponding solvent diluent and shaken thoroughly.

[0180] 2. Device type: [Table 24]

[0181] 3.Elution results The results of elution in each solvent for Comparative Examples 1 to 3 are shown in Table 1 and FIGS. [Table 25] Note: The dissolution results in the table are given as percentages. "ND" means not detected.

[0182] It can be seen from Table 1 and Figures 1-2 that the normal API tablet prepared in Comparative Example 3 of the present disclosure was insoluble in purified water and had significantly different dissolution rates in water containing various concentrations of sodium dodecyl sulfate (SDS) and in solvents with various pH values. The dissolution rate increased with increasing proportion of SDS. The lower the pH, the more serious the drug aggregation became and the lower the dissolution rate. Compared with Comparative Examples 1-3, the dissolution of the tablet prepared after micronizing the API was not significantly improved. Adding an appropriate amount of silicon dioxide and optimizing the formulation (Comparative Example 3) greatly improved the dissolution of the product, but the dissolution rate remained low in solvents with SDS concentration ≦0.5%.

[0183] Test II Dissolution Testing of Nanocrystal Suspensions of Examples 8 and 9 The results of dissolution in each solvent are shown in Table 2 and FIG. [Table 26] Note: Dissolution results in the table are given as percentages.

[0184] As can be seen from Table 2 and Figure 3, the dissolution rates of the nanocrystal suspension formulations prepared in Examples 8-9 of the present disclosure were significantly higher than those of normal tablets, with the dissolution rates reaching 85% or more in 15 minutes in a solvent with a low SDS concentration (3%). However, when the SDS concentration was further reduced to 2%, the samples precipitated in the latter half of the dissolution. When a 3% Tween solvent was used, the dissolution rates of the two nanocrystal suspensions of Examples 8 and 9 were significantly improved, and the samples were stable without precipitation.

[0185] Test III Dissolution test of nanocrystal tablets of Examples 10 to 15 The results of dissolution in each solvent are shown in Table 3 and FIGS. [Table 27] Note: Dissolution results in the table are given as percentages.

[0186] It can be seen from Table 3 and Figures 4 to 6 that the dissolution rates of the nanocrystal tablets prepared in Examples 10 to 15 of the present disclosure were significantly higher than those of the normal tablets. Compared to the nanocrystal suspension, the dissolution rate in the initial stage was slower, and the dissolution rate at 2 hours was similar.

[0187] Test IV Dissolution test of nanocrystal capsules of Examples 24-25 The results of dissolution in each solvent are shown in Table 4 and FIG. [Table 28]

[0188] It can be seen from Table 4 and Figure 7 that the dissolution rate of the nanocrystal capsules prepared in Examples 24-25 of the present disclosure was significantly higher than that of the normal tablets. Compared to the nanocrystal tablets, the dissolution was faster at and before 1 hour, and the dissolution rate was similar at 1 hour. Compared to the nanocrystal suspensions, the dissolution was slower at and before 15 minutes, and the dissolution rate was similar at 15 minutes.

[0189] Test V Dissolution test of nanocrystalline enteric formulations of Examples 26 to 28 In this study, compared with other examples and comparative examples, the product specification was changed from 200 mg to 100 mg, and the corresponding volume of the dissolution medium was changed from 900 ml to 500 ml.

[0190] The results of dissolution in each solvent are shown in Table 5. [Table 29]

[0191] Particle size stability test In the process of preparing nanocrystal suspension, the inventors have studied the effects of various stabilizers on the particle size stability of the product. Specific test results are shown in Tables 6 and 7.

[0192] [Table 30]

[0193] [Table 31] [Table 32] Note: "--" means not detected.

[0194] In the process of preparing nanocrystals, the inventors have studied the effects of milling speed, milling bead loading, sample volume, and milling time on the particle size of API.

Table 33

Claims

1. 1. A nanocrystal formulation comprising a ROCK2 inhibitor and a stabilizer, wherein the ROCK2 inhibitor is a compound of formula (I): 【Chemistry 1】 [In the formula, Ring A is 【Chemistry 2】 where said group is attached to the pyrimidine ring through one of the two positions marked with * or ** and to the carbonyl group at the other position; R ９ and R １０ are each independently H, a halogen, or C １－６ Alkyl, C ２－６ Alkenyl, C ３－１０ Carbocyclyl, 3- to 10-membered heterocyclyl, C ６－１０ Aryl, 5-14 membered heteroaryl, C ６－１２ Aralkyl, -C(=O)R ５ , and -C １－６ Alkylene-O(P=O)(OH) ２ Selected from: m is independently an integer of 0, 1, 2, or 3; Each n is independently an integer of 0, 1, or 2; Alternatively, ring A is 【Chemistry 3】 where the above groups are attached to the pyrimidine ring at the position marked with *, to the carbonyl group at the position marked with **, and R １０ is H and C １－６ alkyl, or is H or methyl; R is H and C １－６ alkyl; R １ teeth, 【Chemistry 4】 and R ２ is H and C １－６ alkyl; R ３ , R ４ , R ７ , and R ８ are each independently H, a halogen, or -NR ５ R ６ , -OH, C １－６ Alkyl, and -OR ５ Selected from: The alkylene, alkyl, alkenyl, carbocyclyl, heterocyclyl, aryl, heteroaryl, and aralkyl groups each include halogen, C １－６ Alkyl, and -OR ５ may be optionally substituted with one or more substituents independently selected from R ５ and R ６ are each independently H, C １－６ Alkyl, C ３－１０ Carbocyclyl, 3- to 10-membered heterocyclyl, C ６－１０ Aryl, 5- to 14-membered heteroaryl, and C ６－１２ aralkyl] or a pharma- ceutically acceptable salt, ester, stereoisomer, polymorph, solvate, N-oxide, isotopically labeled derivative, metabolite, or prodrug thereof.

2. The ROCK2 inhibitor is a compound of formula (II): 【Chemistry 5】 wherein each radical is as defined in claim 1. or a pharma- ceutically acceptable salt, ester, stereoisomer, polymorph, solvate, N-oxide, isotopically labeled derivative, metabolite, or prodrug thereof; or, The ROCK2 inhibitor is a compound of formula (III): 【Chemistry 6】 [In the formula, R １０ is H or methyl, or R １０ is methyl] or a pharma- ceutically acceptable salt, ester, stereoisomer, polymorph, solvate, N-oxide, isotopically labeled derivative, metabolite, or prodrug thereof; or, The ROCK2 inhibitor is a compound of formula (IV): 【Chemistry 7】 or a pharma- ceutically acceptable salt, ester, stereoisomer, polymorph, solvate, N-oxide, isotopically labeled derivative, metabolite, or prodrug thereof; 2. The nanocrystal formulation of claim 1.

3. The following definitions: i) the stabilizer is selected from one or more of polysorbates, povidone, polyoxyethylene fatty acid esters, polyethylene glycol, polyvinyl alcohol, hydroxypropyl cellulose, hydroxypropyl methylcellulose, polyvinylpyrrolidone, poloxamer, sodium lauryl sulfate, sodium docusate, polyethylene glycol 15-hydroxystearate, polyoxyethylene castor oil, copovidone, lactose, and mannitol; Alternatively, the stabilizing agent is selected from one or more of polysorbates, povidone, hydroxypropyl methylcellulose, polyethylene glycol 6000, polyvinyl alcohol, polyoxyethylene castor oil, poloxamer and sodium lauryl sulfate, lactose and mannitol; ii) the particle size D of the nanocrystal formulation ９０ is between 50 and 1500 nm, alternatively between 50 and 1000 nm, alternatively between 50 and 500 nm, alternatively between 80 and 300 nm, alternatively between 50 nm, 100 nm, 150 nm, 200 nm, 250 nm, 300 nm, 400 nm, 500 nm, 600 nm, 700 nm, 800 nm, 900 nm, or 1000 nm; iii) the weight percentage of said ROCK2 inhibitor is between 1% and 55%, alternatively between 4% and 50%, alternatively between 1% and 10%, alternatively between 10% and 40%, alternatively between 10% and 35%, alternatively between 20% and 30%, alternatively between 30% and 40%, alternatively between 4%, 4.5%, 5%, 10%, 15%, 20%, 25%, 30%, 35%, or 40%; iv) the weight percentage of the stabilizer is from 0.1% to 55%, alternatively from 0.1% to 30%, alternatively from 0.5% to 1%, 1% to 10%, 10% to 20%, or 20% to 30%, alternatively 1%, 2%, 5%, 10%, 15%, 20%, or 30%; v) the weight ratio of said ROCK2 inhibitor to said stabilizer is from 1:10 to 10:1, alternatively from 1:9 to 9:1, alternatively from 1:8 to 8:1, alternatively from 1:7 to 7:1, alternatively from 1:6 to 6:1, alternatively from 1:5 to 5:1, alternatively from 1:4 to 4:1, alternatively from 1:3 to 3:1, alternatively from 1:2 to 2:1, alternatively from 1:1; Alternatively, the weight ratio of said ROCK2 inhibitor to said stabilizer is 4:1 to 1:1, or 1:1 to 1:2, or 5:4, 5:3, 4:1, 3:1, 2:1, 1:1, 1:2, or 1:

3. The nanocrystal formulation of claim 1 , which satisfies one or more of the following:

4. The nanocrystal formulation of claim 1 , comprising an excipient.

5. The following definitions: i) the excipient is selected from one or more of a bulking agent; a wetting agent; a sweetening or flavoring agent; a surfactant; a binder; a disintegrant; a lubricant; a glidant or anti-adherent; a release modifier; a coating agent; an emulsifier; a solubilizer; and a flavoring; ii) the excipient comprises a bulking agent selected from one or more of microcrystalline cellulose, mannitol, lactose, starch, pregelatinized starch, dextrin, calcium phosphate dihydrate, and calcium hydrogen phosphate anhydrous; iii) the excipient comprises a bulking agent and the amount of the bulking agent is between 1% and 80%, alternatively between 20% and 70%, alternatively between 30% and 60% or between 50% and 70%; iv) the excipients include lubricants selected from one or more of magnesium stearate, talc powder, finely powdered silica gel, sodium stearyl fumarate, glyceryl behenate, and polyethylene glycol; v) the excipients include a lubricant and the amount of the lubricant is from 0.1% to 5%, alternatively from 0.1% to 1.5%, alternatively from 0.5% to 1%; vi) the excipient comprises a disintegrant selected from one or two of croscarmellose sodium and crospovidone; Alternatively, the amount of disintegrant is 0-20%, alternatively 0-10%, alternatively 2-10%; vii) the excipient comprises a glidant selected from silicon dioxide; Alternatively, the amount of glidant is 0-20%, alternatively 0-15%, alternatively 2-12%. The nanocrystal formulation of claim 4, which satisfies one or more of the following:

6. The following definitions: i) a nanocrystal formulation further comprising a solvent, Alternatively, the solvent is water or purified water; Alternatively, the amount of solvent is from 0 to 99%, alternatively from 80 to 99%, alternatively from 85% to 95%; ii) a nanocrystal formulation further comprising a bacteriostatic agent, Alternatively, the bacteriostatic agent is selected from one or two of methylparaben and propylparaben; Alternatively, the amount of the bacteriostatic agent is 0-5%, alternatively 0-1%, alternatively 0.01%-0.5%; iii) selected from suspensions, tablets, capsules, granules, powders, lozenges, and pills; Alternatively, it is a suspension, tablet, or capsule. The nanocrystal formulation of claim 1 , which satisfies one or more of the following:

7. 1-10% ROCK2 inhibitor, or 1%, 2%, 3%, 4%, 4.5%, 5%, 6%, 7%, 8%, 9%, or 10% ROCK2 inhibitor, or 4%, 4.5%, or 5% ROCK2 inhibitor; and 1-10% stabilizer, or 1%, 1.5%, 2%, 2.5%, 5%, 9%, 9.5%, or 10% stabilizer A suspension comprising: or, the nanocrystals are a suspension comprising 11.13 g of the ROCK2 inhibitor, 0.77 g of polysorbate 80, 5.00 g of povidone K29 / 32, and 233.10 g of purified water, or the particle size of the ROCK2 inhibitor is 50-1000 nm, alternatively 50-500 nm, alternatively 50-300 nm; or, the nanocrystals are a suspension comprising 11.13 g of the ROCK2 inhibitor, 0.77 g of polysorbate 80, 2.50 g of povidone K29 / 32, and 235.60 g of purified water, or the particle size of the ROCK2 inhibitor is 50-1000 nm, alternatively 50-500 nm, alternatively 50-300 nm; or, the nanocrystals are a suspension comprising 11.13 g of the ROCK2 inhibitor, 0.77 g of polysorbate 80, 2.50 g of hypromellose, and 235.60 g of purified water; or the particle size of the ROCK2 inhibitor is 50-1000 nm, alternatively 50-500 nm, alternatively 50-300 nm; or, the nanocrystals are a suspension comprising 11.13 g of the ROCK2 inhibitor, 0.77 g of polysorbate 80, 5.00 g of hypromellose, and 233.10 g of purified water; or the particle size of the ROCK2 inhibitor is 50-1000 nm, alternatively 50-500 nm, alternatively 50-300 nm; or, the nanocrystals are a suspension comprising 250.08 g of the ROCK2 inhibitor, 17.40 g of polysorbate 80, 111.60 g of povidone K29 / 32, 10.00 g of methylparaben, 1.10 g of propylparaben, and 5189.82 g of purified water, or the particle size of the ROCK2 inhibitor is 50-1000 nm, alternatively 50-500 nm, alternatively 50-300 nm; or, the nanocrystals are a suspension comprising 55.19 g of the ROCK2 inhibitor, 18.61 g of polysorbate 80, 99.25 g of polyoxyethylene castor oil, 2.23 g of methylparaben, 0.25 g of propylparaben, and 1065.09 g of purified water; or the particle size of the ROCK2 inhibitor is 50-1000 nm, alternatively 50-500 nm, alternatively 50-300 nm, alternatively 50-150 nm; 2. The nanocrystal formulation of claim 1.

8. a 10-30% ROCK2 inhibitor, alternatively a 20-30% ROCK2 inhibitor, alternatively a 20%, 22%, 25%, 28%, or 30% ROCK2 inhibitor; 1-20% stabilizer, alternatively 5-20% stabilizer, alternatively 5%, 8%, 10%, 13%, 15%, 18%, or 20% stabilizer A tablet comprising: or, the nanocrystals are a tablet comprising 11.70 g of ROCK2 inhibitor, 3.15 g of polysorbate 80, 5.26 g of lactose, 1.05 g of polyethylene glycol 6000, 16.03 g of mannitol, 4.94 g of silicon dioxide, 2.47 g of sodium lauryl sulfate, 2.96 g of microcrystalline cellulose, 2.96 g of croscarmellose sodium, and 0.32 g of magnesium stearate; or the particle size of the ROCK2 inhibitor is 50-1000 nm, alternatively 50-500 nm, alternatively 50-300 nm, alternatively 50-150 nm; or, the nanocrystals are a tablet comprising 32.71 g of the ROCK2 inhibitor, 14.81 g of polysorbate 80, 14.71 g of lactose, 2.94 g of polyethylene glycol 6000, 43.63 g of mannitol, 16.00 g of silicon dioxide, 8.00 g of sodium lauryl sulfate, 12.80 g of microcrystalline cellulose, 12.80 g of croscarmellose sodium, and 1.60 g of sodium stearyl fumarate; or the particle size of the ROCK2 inhibitor is from 50 to 1000 nm, alternatively from 50 to 500 nm, alternatively from 50 to 300 nm, alternatively from 50 to 150 nm; or, the nanocrystals are a tablet comprising 3.34 g of the ROCK2 inhibitor, 0.89 g of polysorbate 80, 1.80 g of lactose, 0.30 g of polyethylene glycol 6000, 0.75 g of silicon dioxide, 0.75 g of sodium lauryl sulfate, 5.82 g of microcrystalline cellulose, 1.20 g of croscarmellose sodium, and 0.15 g of sodium stearyl fumarate; or the particle size of the ROCK2 inhibitor is 50-1000 nm, alternatively 50-500 nm, alternatively 50-300 nm, alternatively 50-150 nm; or, the nanocrystals are a tablet comprising 3.34 g of the ROCK2 inhibitor, 0.89 g of polysorbate 80, 1.80 g of lactose, 0.30 g of polyethylene glycol 6000, 0.75 g of silicon dioxide, 0.45 g of sodium lauryl sulfate, 4.50 g of microcrystalline cellulose, 1.62 g of pregelatinized starch, 1.20 g of croscarmellose sodium, and 0.15 g of sodium stearyl fumarate; or the particle size of the ROCK2 inhibitor is from 50 to 1000 nm, alternatively from 50 to 500 nm, alternatively from 50 to 300 nm, alternatively from 50 to 150 nm; or, the nanocrystals are a tablet comprising 11.14 g of ROCK2 inhibitor, 2.97 g of polysorbate 80, 6.01 g of lactose, 1.00 g of polyethylene glycol 6000, 1.00 g of silicon dioxide, 27.38 g of spray dried mannitol, and 0.50 g of sodium stearyl fumarate; or the particle size of the ROCK2 inhibitor is 50-1000 nm, alternatively 50-500 nm, alternatively 50-300 nm, alternatively 50-150 nm; or, the nanocrystals are a tablet comprising 111.20 g of the ROCK2 inhibitor, 30.04 g of polysorbate 80, 59.99 g of lactose, 10.00 g of polyethylene glycol 6000, 10.00 g of silicon dioxide, 273.78 g of spray dried mannitol, and 5.00 g of sodium stearyl fumarate; or the particle size of the ROCK2 inhibitor is 50-1000 nm, alternatively 50-500 nm, alternatively 50-300 nm, alternatively 50-150 nm; 2. The nanocrystal formulation of claim 1.

9. a 10-30% ROCK2 inhibitor, alternatively a 20-30% ROCK2 inhibitor, alternatively a 20%, 22%, 25%, 28%, or 30% ROCK2 inhibitor; 10-30% stabilizer, alternatively 20-30% stabilizer, alternatively 20%, 22%, 25%, 28%, or 30% stabilizer A tablet comprising: or, the nanocrystals are a tablet comprising 22.22 g of the ROCK2 inhibitor, 20.00 g of polysorbate 80, 54.80 g of mannitol, 2.00 g of silicon dioxide, and 1.00 g of sodium stearyl fumarate; or the particle size of the ROCK2 inhibitor is 50-1000 nm, alternatively 50-500 nm, alternatively 50-300 nm, alternatively 50-150 nm; or, the nanocrystals are a tablet comprising 22.20 g of the ROCK2 inhibitor, 10.00 g of polysorbate 80, 20.00 g of povidone K29 / 32, 44.80 g of mannitol, 2.00 g of silicon dioxide, and 1.00 g of sodium stearyl fumarate; or the particle size of the ROCK2 inhibitor is 50-1000 nm, alternatively 50-500 nm, alternatively 50-300 nm, alternatively 50-150 nm; or, the nanocrystals are a tablet comprising 22.20 g of the ROCK2 inhibitor, 10.00 g of polysorbate 80, 20.00 g of polyethylene glycol 6000, 44.80 g of mannitol, 2.00 g of silicon dioxide, and 1.00 g of sodium stearyl fumarate; or the particle size of the ROCK2 inhibitor is 50-1000 nm, alternatively 50-500 nm, alternatively 50-300 nm, alternatively 50-150 nm; or, the nanocrystals are a tablet comprising 22.20 g of the ROCK2 inhibitor, 10.00 g of polysorbate 80, 20.00 g of poloxamer 188, 44.80 g of mannitol, 2.00 g of silicon dioxide, and 1.00 g of sodium stearyl fumarate; or the particle size of the ROCK2 inhibitor is 50-1000 nm, alternatively 50-500 nm, alternatively 50-300 nm, alternatively 50-150 nm; or, the nanocrystals are a tablet comprising 22.20 g of the ROCK2 inhibitor, 10.00 g of polysorbate 80, 20.00 g of polyvinyl alcohol, 44.80 g of mannitol, 2.00 g of silicon dioxide, and 1.00 g of sodium stearyl fumarate; or the particle size of the ROCK2 inhibitor is 50-1000 nm, alternatively 50-500 nm, alternatively 50-300 nm, alternatively 50-150 nm; or, the nanocrystals are a tablet comprising 22.20 g of ROCK2 inhibitor, 10.00 g of polysorbate 80, 16.00 g of povidone K29 / 32, 4.00 g of poloxamer 188, 44.80 g of mannitol, 2.00 g of silicon dioxide, and 1.00 g of sodium stearyl fumarate; or the particle size of the ROCK2 inhibitor is 50-1000 nm, alternatively 50-500 nm, alternatively 50-300 nm, alternatively 50-150 nm; or, the nanocrystals are a tablet comprising 22.20 g of ROCK2 inhibitor, 10.00 g of polysorbate 80, 4.00 g of povidone K29 / 32, 16.00 g of poloxamer 188, 44.80 g of mannitol, 2.00 g of silicon dioxide, and 1.00 g of sodium stearyl fumarate; or the particle size of the ROCK2 inhibitor is 50-1000 nm, alternatively 50-500 nm, alternatively 50-300 nm, alternatively 50-150 nm; or, the nanocrystals are a tablet comprising 22.20 g of the ROCK2 inhibitor, 10.00 g of polysorbate 80, 10.00 g of povidone K29 / 32, 10.00 g of poloxamer 188, 44.80 g of mannitol, 2.00 g of silicon dioxide, and 1.00 g of sodium stearyl fumarate; or the particle size of the ROCK2 inhibitor is 50-1000 nm, alternatively 50-500 nm, alternatively 50-300 nm, alternatively 50-150 nm.

2. The nanocrystal formulation of claim 1.

10. a 10-50% ROCK2 inhibitor, alternatively a 20-40% ROCK2 inhibitor, alternatively a 20%, 25%, 30%, 35%, or 40% ROCK2 inhibitor; 10-40% stabilizer, alternatively 20-30% stabilizer, alternatively 20%, 22%, 25%, 28%, or 30% stabilizer A capsule comprising: or, the nanocrystals are a capsule comprising 22.20 g of the ROCK2 inhibitor, 6.00 g of polysorbate 80, 8.00 g of povidone K29 / 32, 4.00 g of poloxamer, and 20.00 g of mannitol; or the particle size of the ROCK2 inhibitor is 50-1000 nm, alternatively 50-500 nm, alternatively 50-300 nm, alternatively 50-150 nm; or, the nanocrystals are capsules containing 4.45 g of the ROCK2 inhibitor, 1.20 g of polysorbate 80, 1.20 g of povidone K32 / 29, 0.80 g of poloxamer 188, and 6.80 g of mannitol; or the particle size of the ROCK2 inhibitor is 50-1000 nm, alternatively 50-500 nm, alternatively 50-300 nm, alternatively 50-150 nm; 2. The nanocrystal formulation of claim 1.

11. A nanocrystalline enteric formulation selected from an enteric coated tablet and an enteric coated capsule; or, Nanocrystalline enteric coated tablets, wherein the enteric coating material is selected from one or more of shellac, polyvinyl alcohol acetate phthalate (PVAP), methacrylic acid copolymers, cellulose and its derivatives (cellulose acetate phthalate (CAP), cellulose acetate trimellitate (CAT), hydroxypropyl methylcellulose phthalate (HPMCP)), and acrylic resins (EuS100, EuL100); or, A nanocrystalline enteric capsule, wherein the enteric capsule is selected from a gelatin enteric capsule or a hypromellose enteric capsule, and the composition of the capsule material is selected from one or more of shellac, polyvinyl alcohol acetate phthalate (PVAP), methacrylic acid copolymer, cellulose and its derivatives (cellulose acetate phthalate (CAP), cellulose acetate trimellitate (CAT), hydroxypropyl methylcellulose phthalate (HPMCP)), and acrylic resin (EuS100, EuL100); 2. The nanocrystal formulation of claim 1.

12. a 1-20% ROCK2 inhibitor, or a 5-15% ROCK2 inhibitor, or a 5%, 6%, 7%, 8%, 9%, 10%, 11%, 12%, 13%, 14%, or 15% ROCK2 inhibitor; and 1-90% stabilizer, alternatively 10-40% stabilizer, alternatively 15-25% stabilizer, for example 15%, 16%, 17%, 18%, 19%, 20%, 21%, 22%, 23%, 24%, or 25% stabilizer. A nanocrystalline enteric coated tablet comprising: or, a nanocrystalline enteric coated tablet comprising 5.56 g of a ROCK2 inhibitor, 1.50 g of polysorbate 80, 1.00 g of poloxamer, 1.50 g of povidone K29 / 32, 5.00 g of mannitol, 1.00 g of silicon dioxide, 4.00 g of crospovidone, 29.94 g of microcrystalline cellulose, 0.50 g of magnesium stearate, and 5 g of film-coated premix (enteric type); or wherein the particle size of said ROCK2 inhibitor is 50-1000 nm, alternatively 50-500 nm, alternatively 50-300 nm, alternatively 50-150 nm; or, a nanocrystalline enteric coated tablet comprising 5.56 g of a ROCK2 inhibitor, 1.50 g of polysorbate 80, 1.00 g of poloxamer, 1.50 g of povidone K29 / 32, 38.94 g of mannitol, 1.00 g of silicon dioxide, 0.50 g of magnesium stearate, and 5 g of film-coated premix (enteric type); or wherein the particle size of said ROCK2 inhibitor is 50-1000 nm, alternatively 50-500 nm, alternatively 50-300 nm, alternatively 50-150 nm; or, a 5-30% ROCK2 inhibitor, or a 10-20% ROCK2 inhibitor, or a 10%, 11%, 12%, 13%, 14%, 15%, 16%, 17%, 18%, 19%, or 20% ROCK2 inhibitor; and 10-50% stabilizer, alternatively 20-40% stabilizer, alternatively 30-40% stabilizer, for example 30%, 31%, 32%, 33%, 34%, 35%, 36%, 37%, 38%, 39%, or 40% stabilizer. A nanocrystalline enteric capsule comprising: or, a nanocrystalline enteric capsule comprising: 11.1 g of a ROCK2 inhibitor, 3.0 g of polysorbate 80, 2.0 g of poloxamer, 3.0 g of povidone K29 / 32, 20.0 g of mannitol, 2.00 g of silicon dioxide, 8.0 g of crospovidone, 29.9 g of microcrystalline cellulose, 1.0 g of magnesium stearate, and an enteric capsule; or wherein the particle size of the ROCK2 inhibitor is 50-1000 nm, alternatively 50-500 nm, alternatively 50-300 nm, alternatively 50-150 nm; 2. The nanocrystal formulation of claim 1.

13. 2. A method for preparing the nanocrystal formulation of claim 1, comprising milling the ROCK2 inhibitor and the stabilizer.

14. The following definitions: i) the weight ratio of said ROCK2 inhibitor to said stabilizer during grinding is 1:15-15:1, 1:14-14:1, 1:13-13:1, 1:12-12:1, 1:11-11:1, 1:10-10:1, 1:9-9:1, 1:8-8:1, 1:7-7:1, 1:6-6:1, 1:5-5:1, 1:4-4:1, 1:3-3:1, 1:2-2:1, 1:1; Alternatively, the weight ratio of said ROCK2 inhibitor to said stabilizer during grinding is from 15:1 to 2:1, alternatively 15:1, 10:1, 10:3, 5:1, 4:1, 3:1, or 2:1; ii) the grinding media is selected from porcelain balls, glass spheres, zirconia beads, steel balls, or ice beads; Alternatively, the grinding media is zirconia beads; iii) the particle size of the grinding media is from 0.1 to 1 mm, alternatively from 0.1 to 0.5 mm, alternatively 0.2 mm; iv) the milling time is from 0.1 to 6 hours, alternatively from 0.5 to 6 hours, alternatively from 4 to 6 hours, alternatively 10 minutes, 20 minutes, 30 minutes, 40 minutes, 1 hour, 1.5 hours, 2 hours, 2.5 hours, 3 hours, 3.5 hours, 4 hours, 4.5 hours, 5 hours, 5.5 hours, or 6 hours; v) the grinding speed is from 1000 to 6000 rpm, alternatively from 1500 rpm to 4500 rpm, alternatively 1500 rpm, 2000 rpm, 2500 rpm, 3000 rpm, 3500 rpm, 4000 rpm, 4500 rpm, 5000 rpm, 5500 rpm, or 6000 rpm; vi) the loading of the grinding beads is from 50% to 95%, alternatively from 70% to 90%, alternatively 70%, 80%, or 90%; The preparation method according to claim 13, wherein one or more of the following conditions are satisfied:

15. The preparation method according to claim 13, further comprising a pre-grinding step prior to grinding the ROCK2 inhibitor and the stabilizer.

16. the pre-grinding speed is 3000-6000 rpm, alternatively 3000 rpm, 3500 rpm, 4000 rpm, 4500 rpm, 5000 rpm, 5500 rpm, or 6000 rpm, alternatively 4000 rpm; The pre-grinding time is 1 to 30 minutes, alternatively 2 to 20 minutes, alternatively 3 minutes, 4 minutes, 5 minutes, 6 minutes, 8 minutes, 10 minutes, 12 minutes, 15 minutes, 18 minutes, or 20 minutes, alternatively 5 minutes; 16. The method of claim 15.

17. 14. The process according to claim 13, wherein stabilizers and / or excipients may be added optionally after grinding.

18. the stabilizer is selected from one or more of polysorbates, povidone, polyoxyethylene fatty acid esters, polyethylene glycol, polyvinyl alcohol, hydroxypropyl cellulose, hydroxypropyl methylcellulose, polyvinylpyrrolidone, poloxamer, sodium lauryl sulfate, sodium docusate, polyethylene glycol 15-hydroxystearate, polyoxyethylene castor oil, copovidone, lactose, and mannitol; Alternatively, the stabilizer is selected from povidone K29 / 32, poloxamer 188, polyvinyl alcohol, lactose, and mannitol; Alternatively, the stabilizer is a mixture of Povidone K29 / 32 and Poloxamer 188 in a ratio of 1:10 to 10:1, alternatively 1:9 to 9:1, alternatively 1:8 to 8:1, alternatively 1:7 to 7:1, alternatively 1:6 to 6:1, alternatively 1:5 to 5:1, alternatively 1:4 to 4:1, alternatively 1:3 to 3:1, alternatively 1:2 to 2:1, alternatively 1:1; alternatively 1:4, 4:1, or 1:

1.

18. The preparation method according to claim 17.

19. A pharmaceutical composition for preventing, alleviating and / or treating idiopathic pulmonary fibrosis, fatty liver disease and / or steatohepatitis, graft-versus-host disease after hematopoietic stem cell transplantation, or viral infections, comprising a nanocrystal formulation according to any one of claims 1 to 12 or a nanocrystal formulation prepared by the method according to any one of claims 13 to 18; Alternatively, the nanocrystal formulation is used for the prevention, alleviation, and / or treatment of fatty liver disease and / or steatohepatitis; Alternatively, the fatty liver disease is alcoholic fatty liver disease (ALFD) or non-alcoholic fatty liver disease (NALFD), the steatohepatitis is alcoholic steatohepatitis (ASH) or non-alcoholic steatohepatitis (NASH), the hematopoietic stem cell transplant is allogeneic hematopoietic stem cell transplant, the graft-versus-host disease is acute graft-versus-host disease or chronic graft-versus-host disease, and the viral infection is a coronavirus infection; Alternatively, the coronavirus is selected from SARA-CoV, SARA-CoV-2, MERS-CoV, HCoV-229E, HCoV-NL63, HCoV-OC43, and HCoV-HKU1; Alternatively, the disease caused by the coronavirus is Middle East Respiratory Syndrome, Severe Acute Respiratory Syndrome, or COVID-19; Alternatively, the coronavirus is severe acute respiratory syndrome coronavirus 2, and the disease caused by the coronavirus is COVID-19.

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