Rock2 inhibitors

Abstract

The present disclosure relates to inhibitors of Rho-associated protein kinase (ROCK), pharmaceutical compositions comprising said inhibitors and their use for the prevention or treatment of diseases mediated by said ROCK. In particular, the inhibitor of ROCK has selectivity for inhibition of ROCK2.

Description

Technical Field

[0001] The present disclosure relates to an inhibitor of Rho-associated protein kinase (ROCK), a pharmaceutical composition comprising the inhibitor, and the use thereof for preventing or treating a disease mediated by the ROCK. In particular, the inhibitor of ROCK is selective for inhibiting ROCK2. Background Art

[0002] Rho-associated coiled-coil kinase (ROCK) is a serine / threonine kinase from the AGC (PKA, PKG and PKC) kinase family and includes two isoforms, ROCK1 and ROCK2. These two isoforms are expressed and regulated differently in specific tissues. For example, ROCK1 is generally expressed at a relatively high level, while ROCK2 is preferentially expressed in certain tissues (including heart, brain and skeletal muscle). ROCK is a target of the small GTPase Rho and is involved in a variety of cell activities achieved by phosphorylating downstream effector proteins (MLC, LIMK, ERM, MARCKS, CRMP-2, etc.). Studies have shown that various diseases (e.g., pulmonary fibrosis, cardiovascular and cerebrovascular diseases, neurological diseases, cancer, etc.) are related to ROCK-mediated pathways. Therefore, ROCK has been considered an important target in new drug development. Summary of the invention

[0003] In one aspect, the present disclosure provides a ROCK2 inhibitor having Formula I:

[0004]

[0005] or a pharmaceutically acceptable salt thereof, wherein:

[0006] R 1 Selected from the group consisting of: H, C1-C6 alkyl, C2-C6 alkenyl, C2-C6 alkynyl, C3-C7 cycloalkyl, halo, -CN, C1-C3 perfluoroalkyl, -OR 11 、-O-(C1-C6 alkyl)-OR 11 、-(C1-C6 alkyl)-OR 11 、-NR 11 R 12 、-O-(C1-C6 alkyl)-NR 11 R 12 、-(C1-C6 alkyl)-NR 11 R 12 、-NR 11 -(C1-C6 alkyl)-NR 11 R 12 、-NR 11 -(C1-C6 alkyl)-OR 11 、-(C1-C6 alkyl)x -C(═O)R 11 、-O-(C1-C6 alkyl) x -C(═O)R 11 、-(C1-C6 alkyl) x -C(═O)OR 11 、-C(═O)-R 11 、-C(═O)OR 11 、-(C1-C6 alkyl) x -C(═O)NR 11 R 12 、-NR 11 -(C1-C6 alkyl) x -C(═O)R 11 and NR 11 -(C1-C6 alkyl) x -C(═O)OR 11 ;

[0007] R 2 Selected from the group consisting of: H, C1-C6 alkyl, C2-C6 alkenyl, C2-C6 alkynyl, C3-C7 cycloalkyl, halo, -CN, C1-C3 perfluoroalkyl, -OR 11 、-O-(C1-C6 alkyl)-OR 11 、-(C1-C6 alkyl)-OR 11 、-NR 11 R 12 、-O-(C1-C6 alkyl)-NR 11 R 12 、-(C1-C6 alkyl)-NR 11 R 12 、-NR 11 -(C1-C6 alkyl)-NR 11 R 12 、-NR 11 -(C1-C6 alkyl)-OR 11 、-(C1-C6 alkyl) x -C(═O)R 11 、-O-(C1-C6 alkyl) x -C(═O)R 11 、-(C1-C6 alkyl) x -C(═O)OR 11 、-C(═O)-R 11 、-C(═O)OR 11 、-(C1-C6 alkyl) x -C(═O)NR 11 R 12 、-NR 11 -(C1-C6 alkyl)x -C(═O)R 11 and-NR 11 -(C1-C6 alkyl) x -C(═O)OR 11 ;

[0008] Alternatively, R 1 and R 2 The fused ring may contain 0 to 2 ring heteroatoms selected from the group consisting of N, O and S, and the fused ring may be unsubstituted or substituted with 1 to 3 substituents selected from the group consisting of: C1-C6 alkyl, halo, -CN, -OH, oxo, -O-(C1-C6 alkyl), -O-(C1-C6 alkyl)-OH, -O-(C1-C6 alkyl)-O-(C1-C6 alkyl), -NR 11 R 12 、-O-(C1-C6 alkyl)-NR 11 R 12 、C1-C3 perfluoroalkyl、-NR 11 -(C1-C6 alkyl)NR 11 R 12 and-NR 11 -(C1-C6 alkyl)-OR 11 ;

[0009] X 4 is N or CH;

[0010] R 3 and R 4 Each is independently selected from the group consisting of: H, C1-C6 alkyl, C2-C6 alkenyl, C2-C6 alkynyl, C3-C7 cycloalkyl, 3 to 10 membered heterocyclyl, C6-C 10 Aryl, 5- to 14-membered heteroaryl, C 6-12 Aralkyl, -(C1-C6 alkyl)-OR 11 、-(C1-C6 alkyl)-NR 11 R 12 、-(C1-C6 alkyl) x -C(═O)R 11 、-(C1-C6 alkyl) x -C(═O)OR 11 and -(C1-C6 alkyl) x -C(═O)NR 11 R 12 ;

[0011] Alternatively, R 3 and R 4together with the nitrogen to which they are attached, provide (i) a 4- to 6-membered heterocyclic ring having 0 to 2 additional ring heteroatoms selected from N, O, and S, or (ii) a 5- to 10-membered heterobicyclic ring system having 0 to 3 additional ring heteroatoms selected from N, O, and S; wherein the heterocyclic ring or the heterobicyclic ring system is unsubstituted or substituted with 1 to 4 substituents selected from the group consisting of halo, C1-C6 alkyl, C2-C6 alkenyl, C2-C6 alkynyl, C3-C7 cycloalkyl, halo, -CN, C1-C3 perfluoroalkyl, -OR 11 , oxo, -O-(C1-C6 alkyl)-OR 11 、-(C1-C6 alkyl)-OR 11 、-NR 11 R 12 、-O-(C1-C6 alkyl)-NR 11 R 12 、-(C1

[0012] -C6 alkyl)-NR 11 R 12 、-NR 11 -(C1-C6 alkyl)-NR 11 R 12 、-NR 11 -(C1-C6 alkyl)-OR 11 、-(C1-C6 alkyl) x -C(═O)R 11 、-O-(C1-C6 alkyl) x -C(═O)R 11 、-(C1-C6 alkyl) x -C(═O)OR 11 、-C(═O)-R 11 、-C(═O)OR 11 、-(C1-C6 alkyl) x -C(═O)NR 11 R 12 、-NR 11 -(C1-C6 alkyl) x -C(═O)R 11 and-NR 11 -(C1-C6 alkyl) x -C(═O)OR 11 ;

[0013] Dashed lines represent optional double bonds;

[0014] Each R 5 independently selected from the group consisting of: H, C1-C6 alkyl, C2-C6 alkenyl, C2-C6 alkynyl, C3-C7 cycloalkyl, halo, -CN, C1-C3 perfluoroalkyl, oxo, -OR11 、-O-(C1-C6 alkyl)-OR 11 、-(C1-C6 alkyl)-OR 11 、-NR 11 R 12 、-O-(C1-C6 alkyl)-NR 11 R 12 、-(C1-C6 alkyl)-NR 11 R 12 、-NR 11 -(C1-C6 alkyl)-NR 11 R 12 、-NR 11 -(C1-C6 alkyl)-OR 11 、-(C1-C6 alkyl) x -C(═O)R 11 、-O-(C1-C6 alkyl) x -C(═O)R 11 、-(C1-C6 alkyl) x -C(═O)OR 11 、-C(═O)-R 11 、-C(═O)OR 11 、-(C1-C6 alkyl) x -C(═O)NR 11 R 12 、-NR 11 -(C1-C6 alkyl) x -C(═O)R 11 and-NR 11 -(C1-C6 alkyl) x -C(═O)OR 11 ;

[0015] n is 0 to 3;

[0016] X 1 Selected by CR 6 and N;

[0017] X 2 Selected by CHR 6 NR 7 , O and S;

[0018] X 3 Selected from the group consisting of C, CH and N;

[0019] Each R 6 independently selected from the group consisting of: H, C1-C6, alkyl, C2-C6, alkenyl, C2-C6, alkynyl, C3-C7, cycloalkyl, halo, -CN, C1-C3, perfluoroalkyl, -OR11 、-O-(C1-C6, alkyl)-OR 11 、-(C1-C6, alkyl)-OR 11 、-NR 11 R 12 、-O-(C1-C6, alkyl)-NR 11 R 12 、-(C1-C6, alkyl)-NR 11 R 12 、-NR 11 -(C1-C6, alkyl)-NR 11 R 12 、-NR 11 -(C1-C6, alkyl)-OR 11 、-(C1-C6, alkyl) x -C(═O)R 11 、-O-(C1-C6, alkyl) x -C(═O)R 11 、-(C1-C6, alkyl) x -C(═O)OR 11 、-C(═O)-R 11 、-C(═O)OR 11 、-(C1-C6, alkyl) x -C(═O)NR 11 R 12 、-NR 11 -(C1-C6, alkyl) x -C(═O)R 11 and-NR 11 -(C1-C6, alkyl) x -C(═O)OR 11 ;

[0020] Each R 7 independently selected from the group consisting of: H, C1-C6 alkyl, C2-C6 alkenyl, C2-C6 alkynyl, C3-C7 cycloalkyl, -(C1-C6 alkyl)-OR 11 、-(C1-C6 alkyl)-NR 11 R 12 、-(C1-C6 alkyl) x -C(═O)R 11 、-(C1-C6 alkyl) x -C(═O)OR 11 and -(C1-C6 alkyl) x -C(═O)NR 11 R 12 ;

[0021] Each x is independently selected from 0 and 1; and

[0022] Each R 11 and R 12 Independently selected from the group consisting of H and C1-C6 alkyl;

[0023] Or alternatively, when R 11 and R 12 When both are attached to the same nitrogen, they together form a 4- to 7-membered heterocyclic ring having 0 to 2 additional ring heteroatoms selected from the group consisting of N, O and S, and the heterocyclic ring is unsubstituted or substituted with 1 to 3 substituents selected from the group consisting of halo, C1-C6 alkyl, C2-C6 alkenyl, C2-C6 alkynyl, C3-C7 cycloalkyl, -CN, -NH2, C1-C3 perfluoroalkyl, -OH, -O-(C1-C6 alkyl) and -(C1-C6 alkyl)-OH.

[0024] In another aspect, the present disclosure provides a method for treating a disease or condition mediated by ROCK2, wherein the method comprises administering to a subject in need thereof an effective amount of a ROCK 2 inhibitor provided herein or a pharmaceutically acceptable salt thereof.

[0025] In embodiments provided by the present disclosure, the method is used to treat a disease or condition selected from the group consisting of: fibrotic diseases, inflammatory diseases, autoimmune diseases, cardiovascular disorders, central nervous system disorders, neoplastic diseases, metabolic syndrome, eye diseases, kidney diseases, lung diseases, muscular dystrophy, sickle cell disease, and viral diseases. BRIEF DESCRIPTION OF THE DRAWINGS

[0026] Figure 1 The structures of compounds according to the present disclosure and their properties are shown.

[0027] FIG. 2A to FIG. 2B The compounds of Example 1 (2A) and Example 2 (2B) are shown to inhibit IL-17 in human CD4+ T cells stimulated with Th17-biasing conditions.

[0028] Figure 3 The compounds of Example 1 and Example 2 were shown to downregulate STAT3 phosphorylation induced by Th17-biased activation in human CD4+ T cells.

[0029] Figure 4 The compounds of Example 1 and Example 2 were shown to downregulate pCofilin in human CD4+ T cells.

[0030] Figure 5 The compounds of Example 1 and Example 2 were shown to reduce profibrogenic gene expression in human lung fibroblast MRC-5 cells.

[0031] Figure 6 The compounds of Example 1 and Example 2 were shown to down-regulate type 1 collagen secretion in human lung fibroblast MRC-5 cells.

[0032] Figure 7 The compounds of Example 1 and Example 2 were shown to down-regulate adipogenesis in human adipocytes. DETAILED DESCRIPTION

[0033] The present invention will now be further described. In the following paragraphs, different aspects of the present invention are provided. Unless explicitly indicated to the contrary, each aspect presented can be combined with any other aspect. In particular, any feature indicated as being preferred or advantageous can be combined with any other one or more features indicated as being preferred or advantageous.

[0034] The compounds, compositions and methods described herein provide selective inhibitors of Rho-associated coiled-coil kinase 2 (ROCK2) for treating diseases or conditions including fibrotic diseases, inflammatory diseases, autoimmune diseases, cardiovascular disorders, central nervous system disorders, neoplastic diseases, metabolic syndrome, eye diseases, kidney diseases, lung diseases, muscular dystrophy, sickle cell disease and viral diseases.

[0035] The compound used in the methods and compositions disclosed herein is a ROCK inhibitor, particularly a ROCK2 selective inhibitor. The compound provides excellent inhibitory activity and good selectivity to ROCK (preferably ROCK2) (higher selectivity to ROCK2 compared to ROCK1). The compound may also provide one or more good physicochemical properties (e.g., solubility, physical and / or chemical stability), improved pharmacokinetic properties (e.g., improved bioavailability, appropriate half-life and duration of action) and improved safety (low toxicity and / or less side effects, wide therapeutic window). In particular, when administered orally, the ROCK2 inhibitor provided herein may show improved solubility and / or improved bioavailability.

[0036] In one aspect, the present disclosure provides a ROCK2 inhibitor having Formula I:

[0037]

[0038] or a pharmaceutically acceptable salt thereof, wherein:

[0039] R 1 Selected from the group consisting of: H, C1-C6 alkyl, C2-C6 alkenyl, C2-C6 alkynyl, C3-C7 cycloalkyl, halo, -CN, C1-C3 perfluoroalkyl, -OR 11 、-O-(C1-C6 alkyl)-OR 11、-(C1-C6 alkyl)-OR 11 、-NR 11 R 12 、-O-(C1-C6 alkyl)-NR 11 R 12 、-(C1-C6 alkyl)-NR 11 R 12 、-NR 11 -(C1-C6 alkyl)-NR 11 R 12 、-NR 11 -(C1-C6 alkyl)-OR 11 、-(C1-C6 alkyl) x -C(═O)R 11 、-O-(C1-C6 alkyl) x -C(═O)R 11 、-(C1-C6 alkyl) x -C(═O)OR 11 、-C(═O)-R 11 、-C(═O)OR 11 、-(C1-C6 alkyl) x -C(═O)NR 11 R 12 、-NR 11 -(C1-C6 alkyl) x -C(═O)R 11 and NR 11 -(C1-C6 alkyl) x -C(═O)OR 11 ;

[0040] R 2 Selected from the group consisting of: H, C1-C6 alkyl, C2-C6 alkenyl, C2-C6 alkynyl, C3-C7 cycloalkyl, halo, -CN, C1-C3 perfluoroalkyl, -OR 11 、-O-(C1-C6 alkyl)-OR 11 、-(C1-C6 alkyl)-OR 11 、-NR 11 R 12 、-O-(C1-C6 alkyl)-NR 11 R 12 、-(C1-C6 alkyl)-NR 11 R 12 、-NR 11 -(C1-C6 alkyl)-NR 11 R 12 、-NR 11 -(C1-C6 alkyl)-OR 11、-(C1-C6 alkyl) x -C(═O)R 11 、-O-(C1-C6 alkyl) x -C(═O)R 11 、-(C1-C6 alkyl) x -C(═O)OR 11 、-C(═O)-R 11 、-C(═O)OR 11 、-(C1-C6 alkyl) x -C(═O)NR 11 R 12 、-NR 11 -(C1-C6 alkyl) x -C(═O)R 11 and-NR 11 -(C1-C6 alkyl) x -C(═O)OR 11 ;

[0041] Alternatively, R 1 and R 2 The fused ring may contain 0 to 2 ring heteroatoms selected from the group consisting of N, O and S, and the fused ring may be unsubstituted or substituted with 1 to 3 substituents selected from the group consisting of: C1-C6 alkyl, halo, -CN, -OH, oxo, -O-(C1-C6 alkyl), -O-(C1-C6 alkyl)-OH, -O-(C1-C6 alkyl)-O-(C1-C6 alkyl), -NR 11 R 12 、-O-(C1-C6 alkyl)-NR 11 R 12 、C1-C3 perfluoroalkyl、-NR 11 -(C1-C6 alkyl)NR 11 R 12 and-NR 11 -(C1-C6 alkyl)-OR 11 ;

[0042] X 4 is N or CH;

[0043] R 3 and R 4 Each is independently selected from the group consisting of: H, C1-C6 alkyl, C2-C6 alkenyl, C2-C6 alkynyl, C3-C7 cycloalkyl, 3 to 10 membered heterocyclyl, C6-C 10 Aryl, 5- to 14-membered heteroaryl, C 6-12 Aralkyl, -(C1-C6 alkyl)-OR 11、-(C1-C6 alkyl)-NR 11 R 12 、-(C1-C6 alkyl) x -C(═O)R 11 、-(C1-C6 alkyl) x -C(═O)OR 11 and -(C1-C6 alkyl) x -C(═O)NR 11 R 12 ;

[0044] Alternatively, R 3 and R 4 together with the nitrogen to which they are attached, provide (i) a 4- to 6-membered heterocyclic ring having 0 to 2 additional ring heteroatoms selected from N, O, and S, or (ii) a 5- to 10-membered heterobicyclic ring system having 0 to 3 additional ring heteroatoms selected from N, O, and S; wherein the heterocyclic ring or the heterobicyclic ring system is unsubstituted or substituted with 1 to 4 substituents selected from the group consisting of halo, C1-C6 alkyl, C2-C6 alkenyl, C2-C6 alkynyl, C3-C7 cycloalkyl, halo, -CN, C1-C3 perfluoroalkyl, -OR 11 , oxo, -O-(C1-C6 alkyl)-OR 11 、-(C1-C6 alkyl)-OR 11 、-NR 11 R 12 、-O-(C1-C6 alkyl)-NR 11 R 12 、-(C1

[0045] -C6 alkyl)-NR 11 R 12 、-NR 11 -(C1-C6 alkyl)-NR 11 R 12 、-NR 11 -(C1-C6 alkyl)-OR 11 、-(C1-C6 alkyl) x -C(═O)R 11 、-O-(C1-C6 alkyl) x -C(═O)R 11 、-(C1-C6 alkyl) x -C(═O)OR 11 、-C(═O)-R 11 、-C(═O)OR 11 、-(C1-C6 alkyl) x -C(═O)NR 11 R 12 、-NR 11-(C1-C6 alkyl) x -C(═O)R 11 and-NR 11 -(C1-C6 alkyl) x -C(═O)OR 11 ;

[0046] Dashed lines represent optional double bonds;

[0047] Each R 5 independently selected from the group consisting of: H, C1-C6 alkyl, C2-C6 alkenyl, C2-C6 alkynyl, C3-C7 cycloalkyl, halo, -CN, C1-C3 perfluoroalkyl, oxo, -OR 11 、-O-(C1-C6 alkyl)-OR 11 、-(C1-C6 alkyl)-OR 11 、-NR 11 R 12 、-O-(C1-C6 alkyl)-NR 11 R 12 、-(C1-C6 alkyl)-NR 11 R 12 、-NR 11 -(C1-C6 alkyl)-NR 11 R 12 、-NR 11 -(C1-C6 alkyl)-OR 11 、-(C1-C6 alkyl) x -C(═O)R 11 、-O-(C1-C6 alkyl) x -C(═O)R 11 、-(C1-C6 alkyl) x -C(═O)OR 11 、-C(═O)-R 11 、-C(═O)OR 11 、-(C1-C6 alkyl) x -C(═O)NR 11 R 12 、-NR 11 -(C1-C6 alkyl) x -C(═O)R 11 and-NR 11 -(C1-C6 alkyl) x -C(═O)OR 11 ;

[0048] n is 0 to 3;

[0049] X 1 Selected by CR 6 and N;

[0050] X 2 Selected by CHR 6 NR 7 , O and S;

[0051] X 3 Selected from the group consisting of C, CH and N;

[0052] Each R 6 independently selected from the group consisting of: H, C1-C6, alkyl, C2-C6, alkenyl, C2-C6, alkynyl, C3-C7, cycloalkyl, halo, -CN, C1-C3, perfluoroalkyl, -OR 11 、-O-(C1-C6, alkyl)-OR 11 、-(C1-C6, alkyl)-OR 11 、-NR 11 R 12 、-O-(C1-C6, alkyl)-NR 11 R 12 、-(C1-C6, alkyl)-NR 11 R 12 、-NR 11 -(C1-C6, alkyl)-NR 11 R 12 、-NR 11 -(C1-C6, alkyl)-OR 11 、-(C1-C6, alkyl) x -C(═O)R 11 、-O-(C1-C6, alkyl) x -C(═O)R 11 、-(C1-C6, alkyl) x -C(═O)OR 11 、-C(═O)-R 11 、-C(═O)OR 11 、-(C1-C6, alkyl) x -C(═O)NR 11 R 12 、-NR 11 -(C1-C6, alkyl) x -C(═O)R 11 and-NR 11 -(C1-C6, alkyl) x -C(═O)OR 11 ;

[0053] Each R 7independently selected from the group consisting of: H, C1-C6 alkyl, C2-C6 alkenyl, C2-C6 alkynyl, C3-C7 cycloalkyl, -(C1-C6 alkyl)-OR 11 、-(C1-C6 alkyl)-NR 11 R 12 、-(C1-C6 alkyl) x -C(═O)R 11 、-(C1-C6 alkyl) x -C(═O)OR 11 and -(C1-C6 alkyl) x -C(═O)NR 11 R 12 ;

[0054] Each x is independently selected from 0 and 1; and

[0055] Each R 11 and R 12 Independently selected from the group consisting of H and C1-C6 alkyl;

[0056] Or alternatively, when R 11 and R 12 When both are attached to the same nitrogen, they together form a 4- to 7-membered heterocyclic ring having 0 to 2 additional ring heteroatoms selected from the group consisting of N, O and S, and the heterocyclic ring is unsubstituted or substituted with 1 to 3 substituents selected from the group consisting of halo, C1-C6 alkyl, C2-C6 alkenyl, C2-C6 alkynyl, C3-C7 cycloalkyl, -CN, -NH2, C1-C3 perfluoroalkyl, -OH, -O-(C1-C6 alkyl) and -(C1-C6 alkyl)-OH.

[0057] In some embodiments, the present disclosure provides a ROCK2 inhibitor having Formula II:

[0058]

[0059] or a pharmaceutically acceptable salt thereof, wherein:

[0060] R 1 Selected from the group consisting of: H, C1-C6 alkyl, C2-C6 alkenyl, C2-C6 alkynyl, C3-C7 cycloalkyl, halo, -CN, C1-C3 perfluoroalkyl, -OR 11 、-O-(C1-C6 alkyl)-OR 11 、-(C1-C6 alkyl)-OR 11 、-NR 11 R 12 、-O-(C1-C6 alkyl)-NR 11 R 12、-(C1-C6 alkyl)-NR 11 R 12 、-NR 11 -(C1-C6 alkyl)-NR 11 R 12 、-NR 11 -(C1-C6 alkyl)-OR 11 、-(C1-C6 alkyl) x -C(═O)R 11 、-O-(C1-C6 alkyl) x -C(═O)R 11 、-(C1-C6 alkyl) x -C(═O)OR 11 、-C(═O)-R 11 、-C(═O)OR 11 、-(C1-C6 alkyl) x -C(═O)NR 11 R 12 、-NR 11 -(C1-C6 alkyl) x -C(═O)R 11 and NR 11 -(C1-C6 alkyl) x -C(═O)OR 11 ;

[0061] R 2 Selected from the group consisting of: H, C1-C6 alkyl, C2-C6 alkenyl, C2-C6 alkynyl, C3-C7 cycloalkyl, halo, -CN, C1-C3 perfluoroalkyl, -OR 11 、-O-(C1-C6 alkyl)-OR 11 、-(C1-C6 alkyl)-OR 11 、-NR 11 R 12 、-O-(C1-C6 alkyl)-NR 11 R 12 、-(C1-C6 alkyl)-NR 11 R 12 、-NR 11 -(C1-C6 alkyl)-NR 11 R 12 、-NR 11 -(C1-C6 alkyl)-OR 11 、-(C1-C6 alkyl) x -C(═O)R 11 、-O-(C1-C6 alkyl) x -C(═O)R 11 、-(C1-C6 alkyl)x -C(═O)OR 11 、-C(═O)-R 11 、-C(═O)OR 11 、-(C1-C6 alkyl) x -C(═O)NR 11 R 12 、-NR 11 -(C1-C6 alkyl) x -C(═O)R 11 and-NR 11 -(C1-C6 alkyl) x -C(═O)OR 11 ;

[0062] Alternatively, R 1 and R 2 The fused ring may contain 0 to 2 ring heteroatoms selected from the group consisting of N, O and S, and the fused ring may be unsubstituted or substituted with 1 to 3 substituents selected from the group consisting of: C1-C6 alkyl, halo, -CN, -OH, oxo, -O-(C1-C6 alkyl), -O-(C1-C6 alkyl)-OH, -O-(C1-C6 alkyl)-O-(C1-C6 alkyl), -NR 11 R 12 、-O-(C1-C6 alkyl)-NR 11 R 12 、C1-C3 perfluoroalkyl、-NR 11 -(C1-C6 alkyl)NR 11 R 12 and-NR 11 -(C1-C6 alkyl)-OR 11 ;

[0063] R 3 and R 4 Each is independently selected from the group consisting of: H, C1-C6 alkyl, C2-C6 alkenyl, C2-C6 alkynyl, C3-C7 cycloalkyl, 3 to 10 membered heterocyclyl, C6-C 10 Aryl, 5- to 14-membered heteroaryl, C 6-12 Aralkyl, -(C1-C6 alkyl)-OR 11 、-(C1-C6 alkyl)-NR 11 R 12 、-(C1-C6 alkyl) x -C(═O)R 11 、-(C1-C6 alkyl) x -C(═O)OR 11 and -(C1-C6 alkyl) x-C(═O)NR 11 R 12 ;

[0064] Alternatively, R 3 and R 4 together with the nitrogen to which they are attached, provide (i) a 4- to 6-membered heterocyclic ring having 0 to 2 additional ring heteroatoms selected from N, O, and S, or (ii) a 5- to 10-membered heterobicyclic ring system having 0 to 3 additional ring heteroatoms selected from N, O, and S; wherein the heterocyclic ring or the heterobicyclic ring system is unsubstituted or substituted with 1 to 4 substituents selected from the group consisting of halo, C1-C6 alkyl, C2-C6 alkenyl, C2-C6 alkynyl, C3-C7 cycloalkyl, halo, -CN, C1-C3 perfluoroalkyl, -OR 11 , oxo, -O-(C1-C6 alkyl)-OR 11 、-(C1-C6 alkyl)-OR 11 、-NR 11 R 12 、-O-(C1-C6 alkyl)-NR 11 R 12 、-(C1

[0065] -C6 alkyl)-NR 11 R 12 、-NR 11 -(C1-C6 alkyl)-NR 11 R 12 、-NR 11 -(C1-C6 alkyl)-OR 11 、-(C1-C6 alkyl) x -C(═O)R 11 、-O-(C1-C6 alkyl) x -C(═O)R 11 、-(C1-C6 alkyl) x -C(═O)OR 11 、-C(═O)-R 11 、-C(═O)OR 11 、-(C1-C6 alkyl) x -C(═O)NR 11 R 12 、-NR 11 -(C1-C6 alkyl) x -C(═O)R 11 and-NR 11 -(C1-C6 alkyl) x -C(═O)OR 11 ;

[0066] Dashed lines represent optional double bonds;

[0067] Each R 5 independently selected from the group consisting of: H, C1-C6 alkyl, C2-C6 alkenyl, C2-C6 alkynyl, C3-C7 cycloalkyl, halo, -CN, C1-C3 perfluoroalkyl, oxo, -OR 11 、-O-(C1-C6 alkyl)-OR 11 、-(C1-C6 alkyl)-OR 11 、-NR 11 R 12 、-O-(C1-C6 alkyl)-NR 11 R 12 、-(C1-C6 alkyl)-NR 11 R 12 、-NR 11 -(C1-C6 alkyl)-NR 11 R 12 、-NR 11 -(C1-C6 alkyl)-OR 11 、-(C1-C6 alkyl) x -C(═O)R 11 、-O-(C1-C6 alkyl) x -C(═O)R 11 、-(C1-C6 alkyl) x -C(═O)OR 11 、-C(═O)-R 11 、-C(═O)OR 11 、-(C1-C6 alkyl) x -C(═O)NR 11 R 12 、-NR 11 -(C1-C6 alkyl) x -C(═O)R 11 and-NR 11 -(C1-C6 alkyl) x -C(═O)OR 11 ;

[0068] n is 0 to 3;

[0069] X 1 Selected by CR 6 and N;

[0070] X 2 Selected by CHR 6 NR 7 , O and S;

[0071] X 3 Selected from the group consisting of C, CH and N;

[0072] Each R 6 independently selected from the group consisting of: H, C1-C6, alkyl, C2-C6, alkenyl, C2-C6, alkynyl, C3-C7, cycloalkyl, halo, -CN, C1-C3, perfluoroalkyl, -OR 11 、-O-(C1-C6, alkyl)-OR 11 、-(C1-C6, alkyl)-OR 11 、-NR 11 R 12 、-O-(C1-C6, alkyl)-NR 11 R 12 、-(C1-C6, alkyl)-NR 11 R 12 、-NR 11 -(C1-C6, alkyl)-NR 11 R 12 、-NR 11 -(C1-C6, alkyl)-OR 11 、-(C1-C6, alkyl) x -C(═O)R 11 、-O-(C1-C6, alkyl) x -C(═O)R 11 、-(C1-C6, alkyl) x -C(═O)OR 11 、-C(═O)-R 11 、-C(═O)OR 11 、-(C1-C6, alkyl) x -C(═O)NR 11 R 12 、-NR 11 -(C1-C6, alkyl) x -C(═O)R 11 and-NR 11 -(C1-C6, alkyl) x -C(═O)OR 11 ;

[0073] Each R 7 independently selected from the group consisting of: H, C1-C6 alkyl, C2-C6 alkenyl, C2-C6 alkynyl, C3-C7 cycloalkyl, -(C1-C6 alkyl)-OR 11 、-(C1-C6 alkyl)-NR 11 R 12 、-(C1-C6 alkyl) x -C(═O)R 11 、-(C1-C6 alkyl) x -C(═O)OR11 and -(C1-C6 alkyl) x -C(═O)NR 11 R 12 ;

[0074] Each x is independently selected from 0 and 1; and

[0075] Each R 11 and R 12 Independently selected from the group consisting of H and C1-C6 alkyl;

[0076] Or alternatively, when R 11 and R 12 When both are attached to the same nitrogen, they together form a 4- to 7-membered heterocyclic ring having 0 to 2 additional ring heteroatoms selected from the group consisting of N, O and S, and the heterocyclic ring is unsubstituted or substituted with 1 to 3 substituents selected from the group consisting of halo, C1-C6 alkyl, C2-C6 alkenyl, C2-C6 alkynyl, C3-C7 cycloalkyl, -CN, -NH2, C1-C3 perfluoroalkyl, -OH, -O-(C1-C6 alkyl) and -(C1-C6 alkyl)-OH.

[0077] In some embodiments, the present disclosure provides a ROCK2 inhibitor having Formula IIa:

[0078]

[0079] or a pharmaceutically acceptable salt thereof, wherein:

[0080] R 3 and R 4 Each is independently selected from the group consisting of: H, C1-C6 alkyl, C2-C6 alkenyl, C2-C6 alkynyl, C3-C7 cycloalkyl, 3 to 10 membered heterocyclyl, C6-C 10 Aryl, 5- to 14-membered heteroaryl, C 6-12 Aralkyl, -(C1-C6 alkyl)-OR 11 、-(C1-C6 alkyl)-NR 11 R 12 、-(C1-C6 alkyl) x -C(═O)R 11 、-(C1-C6 alkyl) x -C(═O)OR 11 and -(C1-C6 alkyl) x -C(═O)NR 11 R 12 ;

[0081] Alternatively, R 3 and R 4together with the nitrogen to which they are attached, provide (i) a 4- to 6-membered heterocyclic ring having 0 to 2 additional ring heteroatoms selected from N, O, and S, or (ii) a 5- to 10-membered heterobicyclic ring system having 0 to 3 additional ring heteroatoms selected from N, O, and S; wherein the heterocyclic ring or the heterobicyclic ring system is unsubstituted or substituted with 1 to 4 substituents selected from the group consisting of halo, C1-C6 alkyl, C2-C6 alkenyl, C2-C6 alkynyl, C3-C7 cycloalkyl, halo, -CN, C1-C3 perfluoroalkyl, -OR 11 , oxo, -O-(C1-C6 alkyl)-OR 11 、-(C1-C6 alkyl)-OR 11 、-NR 11 R 12 、-O-(C1-C6 alkyl)-NR 11 R 12 、-(C1

[0082] -C6 alkyl)-NR 11 R 12 、-NR 11 -(C1-C6 alkyl)-NR 11 R 12 、-NR 11 -(C1-C6 alkyl)-OR 11 、-(C1-C6 alkyl) x -C(═O)R 11 、-O-(C1-C6 alkyl) x -C(═O)R 11 、-(C1-C6 alkyl) x -C(═O)OR 11 、-C(═O)-R 11 、-C(═O)OR 11 、-(C1-C6 alkyl) x -C(═O)NR 11 R 12 、-NR 11 -(C1-C6 alkyl) x -C(═O)R 11 and-NR 11 -(C1-C6 alkyl) x -C(═O)OR 11 ;

[0083] Dashed lines represent optional double bonds;

[0084] X 1 Selected by CR 6 and N;

[0085] X 2 Selected by CHR6 NR 7 , O and S;

[0086] X 3 Selected from the group consisting of C, CH and N;

[0087] Each R 6 independently selected from the group consisting of: H, C1-C6, alkyl, C2-C6, alkenyl, C2-C6, alkynyl, C3-C7, cycloalkyl, halo, -CN, C1-C3, perfluoroalkyl, -OR 11 、-O-(C1-C6, alkyl)-OR 11 、-(C1-C6, alkyl)-OR 11 、-NR 11 R 12 、-O-(C1-C6, alkyl)-NR 11 R 12 、-(C1-C6, alkyl)-NR 11 R 12 、-NR 11 -(C1-C6, alkyl)-NR 11 R 12 、-NR 11 -(C1-C6, alkyl)-OR 11 、-(C1-C6, alkyl) x -C(═O)R 11 、-O-(C1-C6, alkyl) x -C(═O)R 11 、-(C1-C6, alkyl) x -C(═O)OR 11 、-C(═O)-R 11 、-C(═O)OR 11 、-(C1-C6, alkyl) x -C(═O)NR 11 R 12 、-NR 11 -(C1-C6, alkyl) x -C(═O)R 11 and-NR 11 -(C1-C6, alkyl) x -C(═O)OR 11 ;

[0088] Each R 7 independently selected from the group consisting of: H, C1-C6 alkyl, C2-C6 alkenyl, C2-C6 alkynyl, C3-C7 cycloalkyl, -(C1-C6 alkyl)-OR 11 、-(C1-C6 alkyl)-NR11 R 12 、-(C1-C6 alkyl) x -C(═O)R 11 、-(C1-C6 alkyl) x -C(═O)OR 11 and -(C1-C6 alkyl) x -C(═O)NR 11 R 12 ;

[0089] Each x is independently selected from 0 and 1; and

[0090] Each R 11 and R 12 Independently selected from the group consisting of H and C1-C6 alkyl;

[0091] Or alternatively, when R 11 and R 12 When both are attached to the same nitrogen, they together form a 4- to 7-membered heterocyclic ring having 0 to 2 additional ring heteroatoms selected from the group consisting of N, O and S, and the heterocyclic ring is unsubstituted or substituted with 1 to 3 substituents selected from the group consisting of halo, C1-C6 alkyl, C2-C6 alkenyl, C2-C6 alkynyl, C3-C7 cycloalkyl, -CN, -NH2, C1-C3 perfluoroalkyl, -OH, -O-(C1-C6 alkyl) and -(C1-C6 alkyl)-OH.

[0092] In some embodiments, the present disclosure provides a ROCK2 inhibitor having Formula III:

[0093]

[0094] or a pharmaceutically acceptable salt thereof, wherein:

[0095] R 1 Selected from the group consisting of: H, C1-C6 alkyl, C2-C6 alkenyl, C2-C6 alkynyl, C3-C7 cycloalkyl, halo, -CN, C1-C3 perfluoroalkyl, -OR 11 、-O-(C1-C6 alkyl)-OR 11 、-(C1-C6 alkyl)-OR 11 、-NR 11 R 12 、-O-(C1-C6 alkyl)-NR 11 R 12 、-(C1-C6 alkyl)-NR 11 R 12 、-NR 11 -(C1-C6 alkyl)-NR 11 R 12、-NR 11 -(C1-C6 alkyl)-OR 11 、-(C1-C6 alkyl) x -C(═O)R 11 、-O-(C1-C6 alkyl) x -C(═O)R 11 、-(C1-C6 alkyl) x -C(═O)OR 11 、-C(═O)-R 11 、-C(═O)OR 11 、-(C1-C6 alkyl) x -C(═O)NR 11 R 12 、-NR 11 -(C1-C6 alkyl) x -C(═O)R 11 and NR 11 -(C1-C6 alkyl) x -C(═O)OR 11 ;

[0096] R 2 Selected from the group consisting of: H, C1-C6 alkyl, C2-C6 alkenyl, C2-C6 alkynyl, C3-C7 cycloalkyl, halo, -CN, C1-C3 perfluoroalkyl, -OR 11 、-O-(C1-C6 alkyl)-OR 11 、-(C1-C6 alkyl)-OR 11 、-NR 11 R 12 、-O-(C1-C6 alkyl)-NR 11 R 12 、-(C1-C6 alkyl)-NR 11 R 12 、-NR 11 -(C1-C6 alkyl)-NR 11 R 12 、-NR 11 -(C1-C6 alkyl)-OR 11 、-(C1-C6 alkyl) x -C(═O)R 11 、-O-(C1-C6 alkyl) x -C(═O)R 11 、-(C1-C6 alkyl) x -C(═O)OR 11 、-C(═O)-R 11 、-C(═O)OR 11 、-(C1-C6 alkyl) x-C(═O)NR 11 R 12 、-NR 11 -(C1-C6 alkyl) x -C(═O)R 11 and-NR 11 -(C1-C6 alkyl) x -C(═O)OR 11 ;

[0097] Alternatively, R 1 and R 2 The fused ring may contain 0 to 2 ring heteroatoms selected from the group consisting of N, O and S, and the fused ring may be unsubstituted or substituted with 1 to 3 substituents selected from the group consisting of: C1-C6 alkyl, halo, -CN, -OH, oxo, -O-(C1-C6 alkyl), -O-(C1-C6 alkyl)-OH, -O-(C1-C6 alkyl)-O-(C1-C6 alkyl), -NR 11 R 12 、-O-(C1-C6 alkyl)-NR 11 R 12 、C1-C3 perfluoroalkyl、-NR 11 -(C1-C6 alkyl)NR 11 R 12 and-NR 11 -(C1-C6 alkyl)-OR 11 ;

[0098] R 3 and R 4 Each is independently selected from the group consisting of: H, C1-C6 alkyl, C2-C6 alkenyl, C2-C6 alkynyl, C3-C7 cycloalkyl, 3 to 10 membered heterocyclyl, C6-C 10 Aryl, 5- to 14-membered heteroaryl, C 6-12 Aralkyl, -(C1-C6 alkyl)-OR 11 、-(C1-C6 alkyl)-NR 11 R 12 、-(C1-C6 alkyl) x -C(═O)R 11 、-(C1-C6 alkyl) x -C(═O)OR 11 and -(C1-C6 alkyl) x -C(═O)NR 11 R 12 ;

[0099] Alternatively, R 3 and R 4together with the nitrogen to which they are attached, provide (i) a 4- to 6-membered heterocyclic ring having 0 to 2 additional ring heteroatoms selected from N, O and S, or (ii) a 5- to 10-membered heterobicyclic ring system having 0 to 3 additional ring heteroatoms selected from N, O and S; wherein the heterocyclic ring or the heterobicyclic ring system is unsubstituted or substituted with 1 to 4 substituents selected from the group consisting of halo, C1-C6 alkyl, C2-C6 alkenyl, C2-C6 alkynyl, C3-C7 cycloalkyl, halo, -CN, C1-C3 perfluoroalkyl, -OR 11 , oxo, -O-(C1-C6 alkyl)-OR 11 、-(C1-C6 alkyl)-OR 11 、-NR 11 R 12 、-O-(C1-C6 alkyl)-NR 11 R 12 、-(C1-C6 alkyl)-NR 11 R 12 、-NR 11 -(C1-C6 alkyl)-NR 11 R 12 、-NR 11 -(C1-C6 alkyl)-OR 11 、-(C1-C6 alkyl) x -C(═O)R 11 、-O-(C1-C6 alkyl) x -C(═O)R 11 、-(C1-C6 alkyl) x -C(═O)OR 11 、-C(═O)-R 11 、-C(═O)OR 11 、-(C1-C6 alkyl) x -C(═O)NR 11 R 12 、-NR 11 -(C1-C6 alkyl) x -C(═O)R 11 and-NR 11 -(C1-C6 alkyl) x -C(═O)OR 11 ;

[0100] Dashed lines represent optional double bonds;

[0101] Each R 5 independently selected from the group consisting of: H, C1-C6 alkyl, C2-C6 alkenyl, C2-C6 alkynyl, C3-C7 cycloalkyl, halo, -CN, C1-C3 perfluoroalkyl, oxo, -OR 11、-O-(C1-C6 alkyl)-OR 11 、-(C1-C6 alkyl)-OR 11 、-NR 11 R 12 、-O-(C1-C6 alkyl)-NR 11 R 12 、-(C1-C6 alkyl)-NR 11 R 12 、-NR 11 -(C1-C6 alkyl)-NR 11 R 12 、-NR 11 -(C1-C6 alkyl)-OR 11 、-(C1-C6 alkyl) x -C(═O)R 11 、-O-(C1-C6 alkyl) x -C(═O)R 11 、-(C1-C6 alkyl) x -C(═O)OR 11 、-C(═O)-R 11 、-C(═O)OR 11 、-(C1-C6 alkyl) x -C(═O)NR 11 R 12 、-NR 11 -(C1-C6 alkyl) x -C(═O)R 11 and-NR 11 -(C1-C6 alkyl) x -C(═O)OR 11 ;

[0102] n is 0 to 3;

[0103] X 1 Selected by CR 6 and N;

[0104] X 2 Selected by CHR 6 NR 7 , O and S;

[0105] X 3 Selected from the group consisting of C, CH and N;

[0106] Each R 6 independently selected from the group consisting of: H, C1-C6, alkyl, C2-C6, alkenyl, C2-C6, alkynyl, C3-C7, cycloalkyl, halo, -CN, C1-C3, perfluoroalkyl, -OR 11、-O-(C1-C6, alkyl)-OR 11 、-(C1-C6, alkyl)-OR 11 、-NR 11 R 12 、-O-(C1-C6, alkyl)-NR 11 R 12 、-(C1-C6, alkyl)-NR 11 R 12 、-NR 11 -(C1-C6, alkyl)-NR 11 R 12 、-NR 11 -(C1-C6, alkyl)-OR 11 、-(C1-C6, alkyl) x -C(═O)R 11 、-O-(C1-C6, alkyl) x -C(═O)R 11 、-(C1-C6, alkyl) x -C(═O)OR 11 、-C(═O)-R 11 、-C(═O)OR 11 、-(C1-C6, alkyl) x -C(═O)NR 11 R 12 、-NR 11 -(C1-C6, alkyl) x -C(═O)R 11 and-NR 11 -(C1-C6, alkyl) x -C(═O)OR 11 ;

[0107] Each R 7 independently selected from the group consisting of: H, C1-C6 alkyl, C2-C6 alkenyl, C2-C6 alkynyl, C3-C7 cycloalkyl, -(C1-C6 alkyl)-OR 11 、-(C1-C6 alkyl)-NR 11 R 12 、-(C1-C6 alkyl) x -C(═O)R 11 、-(C1-C6 alkyl) x -C(═O)OR 11 and -(C1-C6 alkyl) x -C(═O)NR 11 R 12 ;

[0108] Each x is independently selected from 0 and 1; and

[0109] Each R 11 and R 12 Independently selected from the group consisting of H and C1-C6 alkyl;

[0110] Or alternatively, when R 11 and R 12 When both are attached to the same nitrogen, they together form a 4- to 7-membered heterocyclic ring having 0 to 2 additional ring heteroatoms selected from the group consisting of N, O and S, and the heterocyclic ring is unsubstituted or substituted with 1 to 3 substituents selected from the group consisting of halo, C1-C6 alkyl, C2-C6 alkenyl, C2-C6 alkynyl, C3-C7 cycloalkyl, -CN, -NH2, C1-C3 perfluoroalkyl, -OH, -O-(C1-C6 alkyl) and -(C1-C6 alkyl)-OH.

[0111] In some embodiments, the present disclosure provides a ROCK2 inhibitor having Formula IIIa:

[0112]

[0113] or a pharmaceutically acceptable salt thereof, wherein:

[0114] R 3 and R 4 Each is independently selected from the group consisting of: H, C1-C6 alkyl, C2-C6 alkenyl, C2-C6 alkynyl, C3-C7 cycloalkyl, 3 to 10 membered heterocyclyl, C6-C 10 Aryl, 5- to 14-membered heteroaryl, C 6-12 Aralkyl, -(C1-C6 alkyl)-OR 11 、-(C1-C6 alkyl)-NR 11 R 12 、-(C1-C6 alkyl) x -C(═O)R 11 、-(C1-C6 alkyl) x -C(═O)OR 11 and -(C1-C6 alkyl) x -C(═O)NR 11 R 12 ;

[0115] Alternatively, R 3 and R 4together with the nitrogen to which they are attached, provide (i) a 4- to 6-membered heterocyclic ring having 0 to 2 additional ring heteroatoms selected from N, O, and S, or (ii) a 5- to 10-membered heterobicyclic ring system having 0 to 3 additional ring heteroatoms selected from N, O, and S; wherein the heterocyclic ring or the heterobicyclic ring system is unsubstituted or substituted with 1 to 4 substituents selected from the group consisting of halo, C1-C6 alkyl, C2-C6 alkenyl, C2-C6 alkynyl, C3-C7 cycloalkyl, halo, -CN, C1-C3 perfluoroalkyl, -OR 11 , oxo, -O-(C1-C6 alkyl)-OR 11 、-(C1-C6 alkyl)-OR 11 、-NR 11 R 12 、-O-(C1-C6 alkyl)-NR 11 R 12 、-(C1

[0116] -C6 alkyl)-NR 11 R 12 、-NR 11 -(C1-C6 alkyl)-NR 11 R 12 、-NR 11 -(C1-C6 alkyl)-OR 11 、-(C1-C6 alkyl) x -C(═O)R 11 、-O-(C1-C6 alkyl) x -C(═O)R 11 、-(C1-C6 alkyl) x -C(═O)OR 11 、-C(═O)-R 11 、-C(═O)OR 11 、-(C1-C6 alkyl) x -C(═O)NR 11 R 12 、-NR 11 -(C1-C6 alkyl) x -C(═O)R 11 and-NR 11 -(C1-C6 alkyl) x -C(═O)OR 11 ;

[0117] Dashed lines represent optional double bonds;

[0118] X 1 Selected by CR 6 and N;

[0119] X 2 Selected by CHR6 NR 7 , O and S;

[0120] X 3 Selected from the group consisting of C, CH and N;

[0121] Each R 6 independently selected from the group consisting of: H, C1-C6, alkyl, C2-C6, alkenyl, C2-C6, alkynyl, C3-C7, cycloalkyl, halo, -CN, C1-C3, perfluoroalkyl, -OR 11 、-O-(C1-C6, alkyl)-OR 11 、-(C1-C6, alkyl)-OR 11 、-NR 11 R 12 、-O-(C1-C6, alkyl)-NR 11 R 12 、-(C1-C6, alkyl)-NR 11 R 12 、-NR 11 -(C1-C6, alkyl)-NR 11 R 12 、-NR 11 -(C1-C6, alkyl)-OR 11 、-(C1-C6, alkyl) x -C(═O)R 11 、-O-(C1-C6, alkyl) x -C(═O)R 11 、-(C1-C6, alkyl) x -C(═O)OR 11 、-C(═O)-R 11 、-C(═O)OR 11 、-(C1-C6, alkyl) x -C(═O)NR 11 R 12 、-NR 11 -(C1-C6, alkyl) x -C(═O)R 11 and-NR 11 -(C1-C6, alkyl) x -C(═O)OR 11 ;

[0122] Each R 7 independently selected from the group consisting of: H, C1-C6 alkyl, C2-C6 alkenyl, C2-C6 alkynyl, C3-C7 cycloalkyl, -(C1-C6 alkyl)-OR 11 、-(C1-C6 alkyl)-NR11 R 12 、-(C1-C6 alkyl) x -C(═O)R 11 、-(C1-C6 alkyl) x -C(═O)OR 11 and -(C1-C6 alkyl) x -C(═O)NR 11 R 12 ;

[0123] Each x is independently selected from 0 and 1; and

[0124] Each R 11 and R 12 Independently selected from the group consisting of H and C1-C6 alkyl;

[0125] Or alternatively, when R 11 and R 12 When both are attached to the same nitrogen, they together form a 4- to 7-membered heterocyclic ring having 0 to 2 additional ring heteroatoms selected from the group consisting of N, O and S, and the heterocyclic ring is unsubstituted or substituted with 1 to 3 substituents selected from the group consisting of halo, C1-C6 alkyl, C2-C6 alkenyl, C2-C6 alkynyl, C3-C7 cycloalkyl, -CN, -NH2, C1-C3 perfluoroalkyl, -OH, -O-(C1-C6 alkyl) and -(C1-C6 alkyl)-OH.

[0126] For each of the formulas for ROCK2 inhibitors provided herein, in the partial structure:

[0127]

[0128] The dotted circle represents one or more optional double bonds. Thus, the partial structure can be unsaturated, have a single double bond at any chemically allowed position, have two double bonds at chemically allowed positions, or be an aromatic ring system. As will be appreciated by those skilled in the art, the double bonds in such ring systems will not be in directly adjacent positions (i.e., they do not share carbon atoms). In an embodiment, the partial structure includes:

[0129]

[0130] In other embodiments, the partial structure includes:

[0131]

[0132] As used herein, the term "heteroatom" means an atom of any element other than carbon or hydrogen. Preferred heteroatoms are nitrogen, oxygen and sulfur.

[0133] The term "halogen" or "halo" means -F, -Cl, -Br or -I. Preferred halogens are -F, -Cl and -Br.

[0134] The term "hydroxy" means -OH.

[0135] As used herein, the term "oxo" refers to an oxygen atom having a double bond to another atom, particularly carbon (ie, the substituent =0).

[0136] The term "alkyl" refers to a saturated aliphatic group, including straight chain alkyl and branched chain alkyl. Thus, C1-C6 alkyl includes methyl, ethyl, n-propyl, isopropyl, n-butyl, sec-butyl, isobutyl, tert-butyl, n-pentyl, etc.

[0137] The term "cycloalkyl" refers to a saturated carbocyclic group having 3 to 7 carbons in the ring. Cycloalkyl groups include cyclopropyl, cyclobutyl, cyclopentyl and cyclohexyl.

[0138] The term "alkenyl" refers to a straight or branched hydrocarbon group having a double bond and 2 to 6 carbon atoms ("C2-C6 alkenyl"). Alkenyl includes ethenyl, 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, 4-methyl-3-pentenyl, etc. When a compound of the present disclosure contains an alkenyl group, the compound may exist as an E-form, a Z-form, or any mixture thereof.

[0139] The term "alkynyl" refers to a straight or branched hydrocarbon group having a triple bond and 2 to 6 carbon atoms ("C2-C6 alkynyl"). Alkyl groups include ethynyl, propynyl, and the like.

[0140] As used herein, the term "aryl" includes 5- and 6-membered monocyclic aromatic groups that may include zero to four heteroatoms, such as benzene, pyrene, pyrrole, furan, thiophene, imidazole, oxazole, thiazole, triazole, pyrazole, pyridine, pyrazine, pyridazine and pyrimidine, etc. Those aryl groups having heteroatoms in the ring structure may also be referred to as "aryl heterocycles", "heteroaromatics" or "heteroaryls". The term "aryl" also includes 7- to 14-membered polycyclic ring systems having two or more rings, wherein two or more carbons are shared by two adjacent rings (the rings are "fused rings"), wherein at least one of the rings is aromatic (including heteroaryls), for example, the other rings may be fused cycloalkyl, cycloalkenyl, aryl, heteroaryl and / or heterocyclic groups. Monocyclic heteroaryl groups may have 1 to 3 ring heteroatoms and fused polycyclic heteroaryls may have 1 to 5 ring heteroatoms, wherein the ring heteroatoms are selected from N, O and S.

[0141] The term "heterocyclyl" or "heterocyclic group" refers to a 3- to 10-membered ring structure, more preferably a 5- or 6-membered ring, which ring structure includes one to four heteroatoms. The heterocycle may also be polycyclic. Heterocyclic groups include, for example, thiophene, thianthrene, furan, pyran, isobenzofuran, chromene, xanthene, phenoxathiol, pyrrole, imidazole, pyrazole, isothiazole, isoxazole, pyridine, pyrazine, pyrimidine, pyridazine, indolizine, isoindole, indole, indazole, purine, quinolizine, isoquinoline, quinoline, phthalazine, naphthyridine, quinoxaline, quinazoline, cinnoline, pteridine, carbazole, carboline, phenanthridine, acridine, pyrimidine, phenanthroline, phenazine, phenarsazine, phenothiazine, furazan, phenoxazine, pyrrolidine, oxolane, thiolane, oxazole, piperidine, piperazine, morpholine, lactones, lactams such as azetidinone and pyrrolidone, sultams, sultones, and the like.

[0142] The term "aralkyl" as used herein refers to a C1-C6 alkyl group substituted with an aryl group (eg, aromatic or heteroaromatic group).

[0143] As used herein, when each expression (eg, alkyl, m, n, R, etc.) occurs more than one time in any structure, its definition is intended to be independent of its definition elsewhere in the same structure.

[0144] It is understood that "substituted," "substituted," or "substituted with" includes the implied conditions that such substitution complies with the permissible valences of the replacing atom and substituents and that the substitution results in a stable compound, e.g., which does not undergo transformation itself such as by rearrangement, cyclization, elimination, or the like.

[0145] Certain compounds provided in the present disclosure may exist in specific geometric or stereoisomeric forms. The present disclosure contemplates all such compounds falling within the scope of the present invention, including cis- and trans-isomers, R- and S-enantiomers, diastereomers, racemic mixtures thereof, and other mixtures thereof. Other asymmetric carbon atoms may be present in substituents, such as alkyl groups. All such isomers and mixtures thereof are included in the present invention.

[0146] The term "pharmaceutically acceptable salts" refers to the relatively non-toxic inorganic and organic acid addition salts of compounds disclosed herein, as well as the inorganic and organic base addition salts of compounds disclosed herein.

[0147] As described above, some embodiments of ROCK2 inhibitors may contain basic functional groups, such as amino groups, and may form pharmaceutically acceptable salts with pharmaceutically acceptable acids. These salts may be prepared in situ in the administration vehicle or in the dosage form manufacturing process, or by reacting the purified compounds of the present invention in the form of free alkali with suitable organic or inorganic acids alone and separating the salts thus formed during subsequent purification. Representative salts include hydrobromate, hydrochloride, sulfate, bisulfate, phosphate, nitrate, acetate, valerate, oleate, palmitate, stearate, laurate, benzoate, lactate, phosphate, toluenesulfonate, citrate, maleate, fumarate, succinate, tartrate, naphthoate, mesylate, glucoheptonate, lactobionate and lauryl sulfonate, etc. (see, e.g., Berge et al. (1977) "Pharmaceutical Salts", J. Pharm. Sci. 66: 1-19).

[0148] Pharmaceutically acceptable salts of the compounds of the present invention include conventional non-toxic salts or ammonium salts of the compounds, for example, from non-toxic organic or inorganic acids. For example, such conventional non-toxic salts include those derived from inorganic acids such as hydrochloric acid, hydrobromic acid, sulfuric acid, sulfamic acid, phosphoric acid, nitric acid, and salts prepared from organic acids such as acetic acid, propionic acid, succinic acid, glycolic acid, stearic acid, lactic acid, malic acid, tartaric acid, citric acid, ascorbic acid, palmitic acid, maleic acid, hydroxymaleic acid, phenylacetic acid, glutamic acid, benzoic acid, salicylic acid, p-aminobenzenesulfonic acid, 2-acetoxybenzoic acid, fumaric acid, toluenesulfonic acid, methanesulfonic acid, ethanedisulfonic acid, oxalic acid, isothionic acid, and the like.

[0149] In other cases, the compound provided in the present disclosure may contain one or more acidic functional groups, and therefore can form a pharmaceutically acceptable salt with a pharmaceutically acceptable base. These salts can be similarly prepared in situ in an administration vehicle or in a dosage form manufacturing process, or by reacting the purified compound in the form of a free acid with a suitable alkali (such as a pharmaceutically acceptable hydroxide, carbonate or bicarbonate of a metal cation), with ammonia, or with a pharmaceutically acceptable organic primary amine, secondary amine or tertiary amine alone to prepare. Representative alkali metal or alkaline earth metal salts include lithium salts, sodium salts, potassium salts, calcium salts, magnesium salts and aluminum salts, etc. Representative organic amines that can be used to form base addition salts include ethylamine, diethylamine, ethylenediamine, ethanolamine, diethanolamine, piperazine, etc. (see, for example, Berge et al., the same).

[0150] When administered orally, the ROCK2 inhibitors provided herein may show improved solubility and / or improved bioavailability. For example, the compounds of Examples 1 and 2 show kinetic solubility of 2.2 μg / ml and 2.69 μg / ml, respectively, which is significantly improved compared to Comparative Compound A, which has a kinetic solubility of <1.9 μg / ml.

[0151] Treatment

[0152] The present disclosure provides methods for preventing or treating a disease mediated by ROCK2, wherein the method comprises administering to a subject in need thereof an effective amount of a ROCK2 inhibitor as disclosed herein or a pharmaceutically acceptable salt thereof.

[0153] In some embodiments, the present disclosure provides a method for treating at least one disease or condition selected from the group consisting of fibrotic diseases, inflammatory diseases, and autoimmune diseases, the method comprising administering to a subject in need thereof a therapeutically effective amount of a compound or composition as defined herein.

[0154] In other embodiments, the disclosure provides a method for treating a cardiovascular disorder, a central nervous system disorder, a neoplastic disease or a metabolic syndrome, the method comprising administering to a subject in need thereof a therapeutically effective amount of a compound or composition as defined herein.

[0155] In some embodiments, the disease mediated by ROCK2 is an autoimmune disorder, including rheumatoid arthritis, systemic lupus erythematosus (SLE; lupus), psoriasis, psoriatic arthritis, multiple sclerosis, Crohn's disease, ulcerative colitis, atopic dermatitis, eczema, or graft-versus-host disease (GVHD; acute and chronic), idiopathic pulmonary fibrosis, and scleroderma.

[0156] Other autoimmune disorders that can be treated according to the methods provided in the present disclosure include acute disseminated encephalomyelitis (ADEM), acute necrotizing hemorrhagic leukoencephalitis, Addison's disease, alopecia areata, amyloidosis, ankylosing spondylitis, anti-GBM / anti-TBM nephritis, antiphospholipid syndrome (APS), autoimmune angioedema, autoimmune aplastic anemia, autoimmune dysautonomia, autoimmune hepatitis, autoimmune hyperlipidemia, autoimmune immunodeficiency, autoimmune inner ear disease (AIED), autoimmune myocarditis, autoimmune oophoritis, autoimmune pancreatitis, autoimmune retinopathy, autoimmune thyroid disease, autoimmune urticaria, axonal & neuronal neuropathy , Barlow's disease, Behcet's disease, bullous pemphigoid, cardiomyopathy, Castleman's disease, celiac disease, Chagas disease, chronic inflammatory demyelinating peripheral neuropathy (CIDP), chronic relapsing multifocal encephalomyelitis (CRMO), Chag-Strauss syndrome, Cogan's syndrome, coxsackievirus myocarditis, CREST disease, demyelinating neuropathy, dermatitis herpetiformis, dermatomyositis, Devic's disease (neuromyelitis optica), discoid lupus, Dressler syndrome, eosinophilic esophagitis, eosinophilic fasciitis, erythematosus sarcoidosis, Evans syndrome, fibrosing alveolitis, giant cell arteritis (temporal arteritis), giant cell myocarditis, glomerulonephritis, Goodpasture's syndrome, granulomatosis with polyangiitis (GPA), Graves' disease, Guillain-Barré syndrome Barré syndrome, Hashimoto's encephalitis, Hashimoto's thyroiditis, hemolytic anemia, glomerulonephritis, hypogammaglobulinemia, idiopathic thrombocytopenic purpura (ITP), IgA nephropathy, IgG4-related sclerosing disease, immunomodulatory lipoprotein, inclusion body myositis, interstitial cystitis, juvenile arthritis, juvenile myositis, Kawasaki syndrome, Lambert-Eaton syndrome, leukocytoclastic vasculitis, lichen planus, lichen sclerosus, woody conjunctivitis, linear IgA disease (LAD), Meniere's disease, microscopic polyangiitis, mixed connective tissue disease (MCTD), Moren's ulcer, Muhlenberg-Harris disease, myasthenia gravis, myositis, neuromyelitis optica, neutropenia, ocular cicatricial pemphigoid, optic neuritis, relapsing rheumatism disease, pediatric autoimmune neuropsychiatric disorder with streptococcal infection (PANDAS), paraneoplastic cerebellar degeneration, paroxysmal nocturnal hemoglobinuria (PNH), Parkinson-Rodriguez syndrome, Parsena-Turner syndrome, pars planitis (peripheral uveitis), pemphigus, peripheral neuropathy, perivenous encephalomyelitis, pernicious anemia, POEMS syndrome, polyarteritis nodosa, autoimmune polyglandular syndrome (types I, II, and III), polymyalgia rheumatica, polymyositis, post-myocardial infarction syndrome, postpericardiotomy syndrome, progesterone dermatitis, primary biliary cirrhosis, primary sclerosing cholangitis, psoriatic arthritis, pustular gangrene, pure red cell aplasia, Raynaud's phenomenon, reactive arthritis,Reflex sympathetic dystrophy, Reiter's syndrome, relapsing polychondritis, retroperitoneal fibrosis, sarcoidosis, Schmidt's syndrome, scleritis, Sjögren's syndrome, sperm and testicular autoimmunity, stiff-man syndrome, subacute bacterial endocarditis (SBE), Susac's syndrome, sympathetic ophthalmia, Takayasu's arteritis, temporal arteritis / giant cell arteritis, thrombocytopenic purpura (TTP), Tolman-Henrich syndrome, transverse myelitis, undifferentiated connective tissue disease (UCTD), autoimmune diabetes mellitus type 1, uveitis, vasculitis, blistering dermatosis, vitiligo, and Wegener's granulomatosis (granulomatosis with polyangiitis; GPA).

[0157] Inflammatory disorders that may be treated by the methods provided in the present disclosure include, but are not limited to, cardiovascular inflammation, lung inflammation, kidney inflammation, arteriosclerosis, and sepsis.

[0158] Fibrotic disorders that may be treated by the methods provided in the present disclosure include idiopathic pulmonary fibrosis, renal fibrosis, kidney fibrosis, ocular fibrosis, cardiac fibrosis, NASH, scleroderma, systemic sclerosis, and liver cirrhosis.

[0159] In another embodiment, the present disclosure provides a method for treating muscular dystrophy (Duchenne muscular dystrophy). In another embodiment, the present disclosure provides a method for treating myotonic dystrophy.

[0160] In other embodiments, the ROCK2 inhibitor provided herein can be used to inhibit tumor cell growth and metastasis and angiogenesis, and can be used to treat tumor diseases. Tumor diseases include any malignant growth or tumor caused by abnormal or uncontrolled cell division. Tumor diseases include lymphoma, cancer, leukemia, sarcoma and blastoma. Non-limiting examples include squamous cell carcinoma, small cell lung cancer, pituitary cancer, esophageal cancer, astrocytoma, soft tissue sarcoma, non-small cell lung cancer, lung adenocarcinoma, lung squamous cell carcinoma, peritoneal cancer, hepatocellular carcinoma, gastrointestinal cancer, pancreatic cancer, glioblastoma, cervical cancer, ovarian cancer, bladder cancer, liver cancer, hepatoma, breast cancer, colon cancer, colorectal cancer, endometrial cancer or uterine cancer, salivary gland cancer, kidney cancer, liver cancer, prostate cancer, vulvar cancer, thyroid cancer, liver tumors, brain cancer, endometrial cancer, testicular cancer, bile duct cancer, gallbladder cancer, gastric cancer, melanoma and various types of head and neck cancer.

[0161] In other embodiments, the ROCK2 inhibitors provided herein can be used to treat cardiovascular disorders, including hypertension, cardiomyopathy, cardiac remodeling, atherosclerosis, restenosis, cardiac hypertrophy, cerebral ischemia, cerebral vasospasm, and erectile dysfunction.

[0162] In other embodiments, the ROCK2 inhibitors provided herein can be used to treat pulmonary disorders, including idiopathic pulmonary fibrosis, chronic obstructive pulmonary disease, and asthma.

[0163] In other embodiments, the ROCK2 inhibitors provided herein can be used to treat central nervous system disorders, including neuronal degeneration or spinal cord injury, traumatic brain injury, cerebral cavernous malformations, Huntington's disease, Parkinson's disease, Alzheimer's disease, amyotrophic lateral sclerosis (ALS), or multiple sclerosis.

[0164] In other embodiments, the present disclosure provides methods for treating kidney disease, including polycystic kidney disease, renal fibrosis, and diabetic kidney disease.

[0165] In other embodiments, ROCK2 inhibitors provided herein can be used to treat metabolic diseases, including insulin resistance, hyperinsulinemia, type 2 diabetes, obesity, metabolic syndrome and glucose intolerance. ROCK2 inhibitors can be used to achieve weight loss and / or limit weight gain. In one embodiment, ROCK2 inhibitors are used to reduce or prevent insulin resistance or restore insulin sensitivity.

[0166] In other embodiments, the ROCK2 inhibitors provided herein can be used to treat ocular disorders, including ocular hypertension, age-related macular degeneration (AMD; wet and dry), choroidal neovascularization (CNV), choroidal tumors, diabetic macular edema (DME), iris neovascularization, uveitis, glaucoma, primary open-angle glaucoma, acute angle-closure glaucoma, pigmentary glaucoma, congenital glaucoma, normal-tension glaucoma, secondary glaucoma, neovascular glaucoma, geographic atrophy, and retinitis of prematurity (ROP).

[0167] In another embodiment, the present disclosure provides methods for treating sickle cell disease.

[0168] In other embodiments, the ROCK2 inhibitors provided herein can be used to treat (i.e., cure or reduce its severity, etc.) viral infections (particularly coronavirus infections, such as SARS-CoV-1, SARS-CoV-2, and MERS-CoV), and for treating or preventing sequelae caused by viral infections (including coronavirus infections, such as SARS-CoV-1, SARS-CoV-2, and MERS-CoV). In some embodiments, viral infection is SARS-CoV-1 infection. In some embodiments, viral infection is SARS-CoV-2 infection. In some embodiments, viral infection is MERS-CoV infection. In embodiments, sequelae include one or more of the group consisting of the following items: fatigue, dyspnea (dyspnea), cough, joint pain (osteoarthritis), myalgia, headache, chest pain, fever, palpitations, myocardial inflammation, ventricular dysfunction, stroke, abnormal lung function, fibrosis (such as pulmonary fibrosis), renal dysfunction, rash, alopecia, olfactory and / or taste dysfunction, abnormal sleep regulation, altered cognitive impairment, memory impairment, depression, anxiety, mood changes, and combinations thereof. In embodiments, sequelae include inflammation and / or fibrosis.

[0169] Pharmaceutical composition

[0170] In one aspect, the present disclosure provides a pharmaceutically acceptable composition for treating a viral disease, the pharmaceutically acceptable composition comprising a therapeutically effective amount of one or more ROCK2 inhibitors provided by the present disclosure formulated together with one or more pharmaceutically acceptable carriers. As described below, the pharmaceutical compositions of the present disclosure can be specially formulated for administration in solid or liquid form, including those suitable for: (1) oral administration, e.g., drenches (aqueous or non-aqueous solutions or suspensions), tablets (e.g., those for buccal, sublingual, and systemic absorption), boluses, powders, granules, pastes for application to the tongue; (2) parenteral administration, e.g., by subcutaneous, intramuscular, intravenous, or epidural injection, as, e.g., sterile solutions or suspensions or sustained-release formulations; (3) topical application, e.g., application to the skin as a cream, ointment, or controlled-release patch or spray; (4) intravaginal or intrarectal administration, e.g., as a suppository, pessary, cream, or foam; (5) sublingual administration; (6) ocular administration; (7) transdermal administration; or (8) nasal administration.

[0171] The phrase "pharmaceutically acceptable" is used herein to refer to those compounds, materials, compositions, and / or dosage forms which are, within the scope of sound medical judgment, suitable for use in contact with the tissues of humans and animals with toxicity, irritation, allergic response, or other problems or complications, commensurate with a reasonable benefit / risk ratio.

[0172] As used herein, the phrase "pharmaceutically acceptable carrier" means a pharmaceutically acceptable material, composition or vehicle, such as a liquid or solid filler, diluent, excipient, manufacturing aid (e.g., lubricant, talc, magnesium stearate, calcium stearate or zinc stearate, or stearic acid), solvent or solvent encapsulating material, which is involved in carrying or transporting the subject compound from one organ or part of the body to another organ or part of the body. Each carrier should be compatible with the other ingredients of the formulation and not harmful to the patient. Some examples of materials that can be used as pharmaceutically acceptable carriers include: (1) sugars such as lactose, glucose and sucrose; (2) starches such as corn starch and potato starch; (3) cellulose and its derivatives such as sodium carboxymethylcellulose, ethyl cellulose and cellulose acetate; (4) tragacanth powder; (5) malt; (6) gelatin; (7) talc; (8) excipients such as cocoa butter and suppository wax; (9) oils such as peanut oil, cottonseed oil, safflower oil, sesame oil, olive oil, corn oil and soybean oil; (10 ) glycols, such as propylene glycol; (11) polyols, such as glycerol, sorbitol, mannitol and polyethylene glycol; (12) esters, such as ethyl oleate and ethyl laurate; (13) agar; (14) buffers, such as magnesium hydroxide and aluminum hydroxide; (15) alginic acid; (16) pyrogen-free water; (17) isotonic saline; (18) Ringer's solution; (19) ethanol; (20) pH buffered solutions; (21) polyesters, polycarbonates and / or polyanhydrides; and (22) other nontoxic compatible substances used in pharmaceutical formulations.

[0173] The compounds of the present disclosure can be formulated with conventional carriers and excipients, which can be selected according to conventional practice. Tablets may contain excipients, glidants, fillers, binders, etc. Aqueous formulations are prepared in sterile form and can generally be isotonic when intended to be delivered by a method other than oral administration. All formulations may optionally contain excipients, such as those described in "Handbook of Pharmaceutical Excipients" (1986). Excipients include ascorbic acid and other antioxidants, chelating agents such as EDTA, carbohydrates such as dextran, hydroxyalkyl cellulose, hydroxyalkyl methyl cellulose, stearic acid, etc.

[0174] Although the ROCK2 inhibitors disclosed herein (referred to herein as "active ingredients") can be administered alone, it is preferred that they be provided as pharmaceutical formulations. The formulations disclosed herein for both veterinary and human use comprise at least one active ingredient as provided above, and one or more acceptable carriers for the active ingredient, and optionally other therapeutic ingredients, particularly those additional therapeutic ingredients as discussed herein.

[0175] The preparation includes preparations suitable for the administration route provided herein. The preparation can be conveniently presented in unit dosage form and can be prepared by any method well known in the pharmaceutical field. Technology and preparations are generally found in Remington's Pharmaceutical Sciences (Mack Publishing Co., Easton, Pa.). Such methods include the step of combining the active ingredient with a carrier constituting one or more auxiliary ingredients. Typically, the preparation is prepared by uniformly and closely combining the active ingredient with a liquid carrier or a finely divided solid carrier or both, and then forming the product as required.

[0176] Formulations of the present disclosure suitable for oral administration may be presented as discrete units such as capsules, cachets or tablets, each containing a predetermined amount of the active ingredient; as a powder or granules; as a solution or suspension in an aqueous or non-aqueous liquid; or as an oil-in-water liquid emulsion or a water-in-oil liquid emulsion. The active ingredient may also be administered as a bolus, electuary or paste.

[0177] Tablets can be made by compression or molding optionally with one or more auxiliary ingredients. Compressed tablets can be prepared by compressing an active ingredient in a free-flowing form (such as a powder or granule) optionally mixed with a binder, lubricant, inert diluent, preservative, surfactant or dispersant in a suitable machine. Molded tablets can be made by molding a mixture of powdered active ingredients moistened with an inert liquid diluent in a suitable machine. Tablets can be optionally coated or scored, and are optionally formulated to provide sustained release or controlled release of active ingredients therefrom.

[0178] For infections of the eye or other external tissues such as the mouth and skin, the formulation is preferably applied as a topical solution, ointment or cream containing the active ingredient. The active ingredient may be present in an amount of, for example, 0.075 to 20% w / w (including active ingredients in a range between 0.1% and 20% in increments of 0.1% w / w, such as 0.6% w / w, 0.7% w / w, etc.), preferably 0.2 to 15% w / w, most preferably 0.5 to 10% w / w. When formulated into an ointment, the active ingredient may optionally be used with a paraffin or water-miscible ointment base. Alternatively, the active ingredient may be formulated into a cream with an oil-in-water cream base.

[0179] If desired, the aqueous phase of the cream base may include, for example, at least 30% w / w of a polyol, i.e., an alcohol having two or more hydroxyl groups, such as propylene glycol, 1,3-butylene glycol, mannitol, sorbitol, glycerol, and polyethylene glycol (including PEG 400) and mixtures thereof. Topical formulations may desirably contain compounds that enhance the absorption or penetration of the active ingredient through the skin or other affected area. Examples of such skin penetration enhancers include dimethyl sulfoxide and related analogs.

[0180] The oil phase of the emulsion of the present disclosure can be composed of known ingredients in a known manner. Although this phase can contain only an emulsifier (also known as an emulgator), it ideally contains at least one emulsifier and a fat or oil or a mixture of both a fat and an oil. Preferably, a hydrophilic emulsifier is included together with a lipophilic emulsifier that acts as a stabilizer. It is also preferably comprised of oil and fat. The emulsifier, with or without a stabilizer, constitutes a so-called emulsifying wax, and the wax, together with the oil and fat, constitutes a so-called emulsifying ointment base that forms the oily dispersed phase of the ointment formulation.

[0181] Emulsifiers and emulsion stabilizers suitable for use in the formulations of the present disclosure include 60. 80, cetearyl alcohol, benzyl alcohol, myristyl alcohol, glyceryl monostearate and sodium lauryl sulfate. Other emulsifiers and emulsion stabilizers suitable for use in the formulations of the present disclosure include 80.

[0182] The selection of suitable oil or fat for preparation is based on realizing required characteristic.Emulsifiable paste should preferably be non-greasy, non-staining and washable product, and it has suitable density to avoid leakage from pipe or other containers.Straight or branched, monobasic or dibasic alkyl esters can be used, such as propylene glycol diester of diisoadipate, isocetyl stearate, coconut fatty acid, isopropyl myristate, decyl oleate, isopropyl palmitate, butyl stearate, 2-ethylhexyl palmitate or the blend of the branched esters being called as Crodamol CAP, and the last three kinds are preferred esters.Depending on required characteristic, these materials can be used alone or in combination.Alternatively, high melting point lipids such as white soft paraffin and / or liquid paraffin or other mineral oils are used.

[0183] Pharmaceutical preparations according to the present disclosure include combinations according to the present disclosure and one or more pharmaceutically acceptable carriers or excipients and optional other therapeutic agents. Pharmaceutical preparations containing active ingredients can be in any form suitable for the intended method of administration. When used for oral use, for example, tablets, lozenges, lozenges, aqueous suspensions or oil suspensions, dispersible powders or granules, emulsions, hard or soft capsules, syrups or elixirs can be prepared. Compositions intended for oral use can be prepared according to any method known in the art for making pharmaceutical compositions, and such compositions can contain one or more medicaments, including sweeteners, flavorings, coloring agents and preservatives, so as to provide a palatable preparation. Tablets containing active ingredients mixed with nontoxic pharmaceutically acceptable excipients suitable for tablet manufacturing are acceptable. These excipients can be, for example, inert diluents, such as calcium carbonate or sodium carbonate, lactose, calcium phosphate or sodium phosphate; granulating agents and disintegrants, such as corn starch or alginic acid; binding agents, such as starch, gelatin or gum arabic; and lubricants, such as magnesium stearate, stearic acid or talc. Tablets can be uncoated, or can be coated by known techniques (including microencapsulation) to delay disintegration and absorption in the gastrointestinal tract, and thereby provide sustained action over a long period of time. For example, time-delay materials, such as glyceryl monostearate or glyceryl distearate, can be used alone or with wax.

[0184] Formulations for oral use may also be presented as hard gelatin capsules in which the active ingredient is mixed with an inert solid diluent, such as starch, mannitol, calcium phosphate or kaolin; or as soft gelatin capsules in which the active ingredient is mixed with water or an oil medium, such as peanut oil, liquid paraffin or olive oil.

[0185] The aqueous suspension of the present disclosure contains an active substance mixed with an excipient suitable for making an aqueous suspension. Such excipients include suspending agents, such as sodium carboxymethylcellulose, methylcellulose, hydroxypropyl methylcellulose, sodium alginate, polyvinylpyrrolidone, tragacanth and gum arabic, and dispersants or wetting agents, such as naturally occurring phosphatides (e.g., lecithin), condensation products of alkylene oxide and fatty acids (e.g., polyoxyethylene stearate), condensation products of ethylene oxide and long-chain aliphatic alcohols (e.g., heptadecaethyleneoxycetanol), condensation products of ethylene oxide and partial esters derived from fatty acids and hexitol anhydrides (e.g., polyoxyethylene sorbitan monooleate). Aqueous suspensions may also contain one or more preservatives (such as ethyl p-hydroxybenzoate or n-propyl p-hydroxybenzoate), one or more coloring agents, one or more flavoring agents and one or more sweeteners (such as sucrose or saccharin). Other non-limiting examples of suspending agents include cyclodextrins and Captisol (=sulfobutyl ether beta-cyclodextrin; SEB-beta-CD).

[0186] Oil suspensions can be prepared by suspending the active ingredient in a vegetable oil (such as peanut oil, olive oil, sesame oil or coconut oil) or in a mineral oil (such as liquid paraffin). Oral suspensions can contain thickeners, such as beeswax, hard paraffin or cetyl alcohol. Sweeteners (such as those described above) and flavoring agents can be added to provide a palatable oral preparation. These compositions can be preserved by adding antioxidants, such as ascorbic acid.

[0187] Dispersible granules and granules of the present disclosure suitable for preparing aqueous suspensions by adding water provide active ingredients mixed with dispersants or wetting agents, suspending agents and one or more preservatives. Suitable dispersants or wetting agents and suspending agents are exemplified by those disclosed above. Additional excipients such as sweeteners, flavoring agents and coloring agents may also be present.

[0188] Pharmaceutical compositions of the present disclosure can also be in the form of oil-in-water emulsions. The oil phase can be a vegetable oil (such as olive oil or peanut oil), a mineral oil (such as liquid paraffin) or a mixture of these. Suitable emulsifiers include naturally occurring gums (such as gum arabic and gum tragacanth), naturally occurring phospholipids (such as soybean lecithin), esters or partial esters derived from fatty acids and hexitol anhydrides (such as sorbitan monooleate) and condensation products of these partial esters with ethylene oxide (such as polyoxyethylene sorbitan monooleate). Emulsions can also contain sweeteners or flavorings. Syrups and elixirs can be formulated with sweeteners such as glycerol, sorbitol or sucrose. Such preparations can also contain a demulcent, a preservative, a flavoring or a coloring agent.

[0189] The pharmaceutical composition of the present disclosure can be in the form of a sterile injectable preparation, such as a sterile injectable aqueous or oily suspension. Such suspension can be prepared according to known techniques using those suitable dispersants or wetting agents and suspending agents mentioned above. Sterile injectable preparations can also be sterile injectable solutions or suspensions in non-toxic parenteral acceptable diluents or solvents, such as solutions in 1,3-butanediol, or prepared as lyophilized powders. Acceptable vehicles and solvents that can be used include water, Ringer's solution, and isotonic sodium chloride solution. In addition, sterile non-volatile oils can be used routinely as solvents or suspension media. For this purpose, any mild non-volatile oil containing synthetic monoglycerides or diglycerides can be used. In addition, fatty acids (such as oleic acid) can also be used for the preparation of injectables. Acceptable vehicles and solvents that can be used are water, Ringer's solution, isotonic sodium chloride solution, and hypertonic sodium chloride solution.

[0190] The amount of active ingredient that can be combined with carrier material to produce single dosage form can be changed according to treatment host and specific mode of administration.For example, the timed release preparation that is intended to be used for oral administration to humans can contain about 1 to 1000mg of active substance and suitable and convenient amount of carrier material, and this carrier material can account for about 5% to about 95% (weight: weight) of total composition.Pharmaceutical composition can be prepared to provide the dosage that is easy to measure.For example, the aqueous solution that is intended to be used for intravenous infusion can include about 3 to 500μg of active ingredient / milliliter solution, so that the volume that is suitable for infusion at a rate of about 30mL / hr can be carried out.

[0191] Formulations suitable for topical administration to the eye also include eye drops in which the active ingredient is dissolved or suspended in a suitable carrier, especially an aqueous solvent for the active ingredient. The active ingredient may be present in such formulations at a concentration of 0.5% to 20%, advantageously 0.5% to 10%, and in particular about 1.5% w / w.

[0192] Formulations suitable for topical administration in the mouth include lozenges comprising the active ingredient in a flavored basis, usually sucrose and acacia or tragacanth; pastilles comprising the active ingredient in an inert basis such as gelatin and glycerin or sucrose and acacia; and mouthwashes comprising the active ingredient in a suitable liquid carrier.

[0193] Formulations for rectal administration may be presented as a suppository with a suitable base comprising for example cocoa butter or a salicylate.

[0194] Formulations suitable for intrapulmonary or nasal administration have, for example, a particle size in the range of 0.1 to 500 microns, such as 0.5, 1, 30, 35 microns, etc., which are administered by rapid inhalation through the nasal passages or by oral inhalation to reach the alveolar sacs. Suitable formulations include aqueous or oily solutions of the active ingredient. Formulations suitable for aerosol or dry powder administration can be prepared according to conventional methods and can be delivered with other therapeutic agents such as compounds.

[0195] Formulations suitable for vaginal administration may be presented as suppositories, pessaries, tampons, creams, gels, pastes, foams or spray formulations containing in addition to the active ingredient such carriers as are known in the art to be appropriate.

[0196] Formulations suitable for parenteral administration include aqueous and non-aqueous sterile injection solutions which may contain antioxidants, buffers, bacteriostats and solutes which render the formulation isotonic with the blood of the intended recipient; and aqueous and non-aqueous sterile suspensions which may include suspending agents and thickening agents.

[0197] The preparation is presented in a unit dose or multi-dose container, such as a sealed ampoule and vial, and can be stored under freeze-dried (lyophilized) conditions, requiring only the addition of a sterile liquid carrier, such as water for injection, immediately before use. Extemporaneous injection solutions and suspensions are prepared by sterile granules, granules and tablets of the aforementioned type. A preferred unit dose preparation is a preparation containing the active ingredient of a daily dose or unit daily subdose as described above or an appropriate portion thereof.

[0198] The present disclosure also provides a veterinary composition comprising at least one active ingredient as defined above and a veterinary acceptable carrier for the active ingredient.

[0199] Veterinary carriers are materials useful for the purpose of administering the composition and may be solid, liquid or gaseous materials which are otherwise inert or acceptable in the veterinary art and are compatible with the active ingredient. These veterinary compositions may be administered orally, parenterally or by any other desired route.

[0200] The compounds of the present disclosure are useful in providing controlled release pharmaceutical formulations ("controlled release formulations") containing one or more compounds of the present disclosure as active ingredients, wherein the release of the active ingredient is controlled and regulated to allow less frequent dosing or to improve the pharmacokinetic or toxicity profile of a given active ingredient.

[0201] The patient receiving the treatment is any animal in need thereof, including primates, particularly humans, and other mammals such as horses, cattle, pigs, and sheep; and poultry and pets in general.

[0202] Example

[0203] Example 1

[0204] Method A. Take the synthesis of 2-[2-(3,3-difluoroazetidine-1-carbonyl)-1-methyl-1H-indol-6-yl]-N-[4-(1H-imidazol-5-yl)phenyl]pyrimidin-4-amine as an example.

[0205]

[0206] Step 1: Synthesis of Intermediate 1: 6-bromo-2-(3,3-difluoroazetidine-1-carbonyl)-1H-indole

[0207] 6-bromoindole-2-formic acid (5.26g, 21.91mmol, 1.00 equivalent) is dissolved in anhydrous DCM. Then, oxalyl chloride (2.04ml, 24.10mmol, 1.10 equivalent) is added dropwise at 0°C in reaction mixture (25.0mL), followed by a few drops of DMF. The mixture is stirred for 1h at 0°C, then the cold bath is removed and stirring is continued for 3h at room temperature. TLC analysis shows that starting material is fully converted, and UPLC-MS analysis shows that quality corresponds to appropriate ester (with MeOH quenching sample). The reaction mixture is evaporated to dryness, co-evaporated twice with ACN. The resulting cream-colored powder was then redissolved in THF (25.0 mL) and transferred dropwise to a stirred ice-cold solution of 3,3-difluoroazetidine hydrochloride (2.84 g, 21.91 mmol, 1.00 equiv) and N,N-diisopropylethylamine (15.27 mL, 87.65 mmol, 4.00 equiv) in DCM (25.0 mL) by a syringe. After 1 hour at 0 ° C, the cold bath was removed and stirred overnight at room temperature. NaHCO3 solution was then added, and a mixture of DCM and THF was removed in vacuo. The precipitate was collected by vacuum filtration, washed with water, and dried under high vacuum for 18 h to give 6-bromo-2-(3,3-difluoroazetidine-1-carbonyl)-1H-indole (6.5 g, 90% yield). 1 HNMR (400MHz, DMSO-d6) δ11.93–11.86(m,1H),7.65–7.59(m,2H),7.21(dd,J=8.6,1.8Hz,1H),6.95(dd,J=2.2,0.9Hz,1H),4.77(m,J=170.3Hz,4H). UPLC-MS,m / z:[M+H]=314.8

[0208] Step 2: Synthesis of 6-bromo-2-(3,3-difluoroazetidine-1-carbonyl)-1-methyl-1H-indole.

[0209] To a solution of 6-bromo-2-(3,3-difluoroazetidine-1-carbonyl)-1H-indole (1.20 g, 3.81 mmol, 1.00 equiv) in anhydrous DMF (15.0 mL) was added NaH (0.12 g, 4.57 mmol, 1.20 equiv) in batches at 0 ° C. The resulting mixture was stirred for 1 h at the same temperature. Methyl iodide (0.26 mL, 4.19 mmol, 1.10 equiv) was then added dropwise. Stirring was continued at 0 ° C for 30 minutes and then stirred at room temperature overnight. TLC analysis (eluent 10% MeOH-DCM) was used to monitor the progress of the reaction. The reaction mixture was quenched with water and extracted with EtOAc. The combined organic layers were washed with brine, dried over Na2SO4, filtered and concentrated to give 6-bromo-2-(3,3-difluoroazetidine-1-carbonyl)-1-methyl-1H-indole (1.09 g, 70% yield), which was used in the next step without purification. 1 H NMR (400MHz, DMSO-d6) δ7.87–7.84(m,1H),7.59(d,J=8.5Hz,1H),7.25(dd,J=8.5,1.7Hz,1H),7.04(d,J=0.8Hz,1H),5.04–4.38(m,4H),3.93(s,3H). UPLC-MS,m / z:[M+H] + =330.75

[0210] Step 3: 2-(3,3-Difluoroazetidine-1-carbonyl)-1-methyl-6-(4,4,5,5-tetramethyl-1,3,2-dioxaborolan-2-yl)-1H-indole (Intermediate 1).

[0211] 6-Bromo-2-(3,3-difluoroazetidine-1-carbonyl)-1-methyl-1H-indole (0.74 g, 2.25 mmol, 1.00 equiv), bis(pinacolato)diboron (0.85 g, 3.37 mmol, 1.50 equiv) and potassium acetate (0.66 g, 6.74 mmol, 3.00 equiv) were dissolved in anhydrous dioxane (9.0 mL). The reaction mixture was purged with argon 4 times, and then 1,1'-bis(diphenylphosphino)-ferrocene-palladium(II) (0.05 g, 0.06 mmol, 0.03 equiv) was added. The reaction mixture was placed in a preheated oil bath at 90°C overnight. The reaction mixture was cooled to room temperature, filtered through a pad of celite, concentrated and purified using flash column chromatography (0-10% MeOH in DCM, gradient elution) to give 2-(3,3-difluoroazetidine-1-carbonyl)-1-methyl-6-(4,4,5,5-tetramethyl-1,3,2-dioxaborolan-2-yl)-1H-indole (Intermediate 1) (0.81 g, 89% yield).1 H NMR (400MHz, DMSO-d6) δ7.83 (d, J=0.9Hz, 1H), 7.63 (dd, J=8.0, 0.8Hz, 1H), 7.42 (dd, J=8. 0, 0.9Hz, 1H), 7.03 (d, J = 0.9Hz, 1H), 4.70 (d, J = 105.5Hz, 5H), 3.98 (s, 3H), 1.33 (s, 11H). UPLC-MS,m / z:[M+H] + =377.15.

[0212] Step 4: Intermediate 2: Synthesis of 2-chloro-N-[4-(1H-imidazol-5-yl)phenyl]pyrimidin-4-amine

[0213] To a solution of 4-(1H-imidazole-5-yl)aniline (0.89 g, 5.60 mmol, 1.00 equiv) in ethanol (9.0 mL) was added N,N-diisopropylethylamine (DIPEA) (1.46 mL, 8.38 mmol, 1.50 equiv) and the reaction mixture was stirred at room temperature for 15 minutes. 2,4-dichloropyrimidine (0.83 g, 5.6 mmol, 1.00 equiv) was then added and the reaction mixture was stirred for 24 h. It was filtered. The resulting solid filter cake was redissolved in boiling ethanol, filtered again, and dried under high vacuum to give 2-chloro-N-[4-(1H-imidazole-5-yl)phenyl]pyrimidine-4-amine (0.89 g, 56% yield). 1 H NMR (300MHz, DMSO-d6) δ12.90(s,1H),10.15(s,1H),8.14(d,J=5.9Hz,1H),8.03(s,2H),7.60(d,J=2.4Hz,4H),6.78(d,J=5.9Hz,1H). UPLC-MS,m / z:[M+H] + =271.95.

[0214] Step 5: 2-[2-(3,3-Difluoroazetidine-1-carbonyl)-1-methyl-1H-indol-6-yl]-N-[4-(1H-imidazol-5-yl)phenyl]pyrimidin-4-amine.

[0215] A mixture of intermediate 1 (0.20 g, 0.49 mmol, 1.40 equivalents), intermediate 2 (0.10 g, 0.35 mmol, 1.00 equivalents) and sodium carbonate (0.07 g, 0.70 mmol, 2.00 equivalents) was dissolved in a mixture of dimethoxyethane (6.0 mL) and water (2.0 mL) and degassed with argon for 15 minutes. Then, tetrakis (triphenylphosphine) palladium (0.024 g, 0.021 mmol, 0.06 equivalents) was added and the reaction mixture was stirred at 110 ° C in a microwave reactor. UPLC-MS was used to analyze and monitor the reaction process, and the raw material was completely converted after stirring for 4 hours. After cooling, the mixture was diluted with EtOAc and filtered through a diatomaceous earth pad. The filtrate was concentrated in vacuo and purified via preparative HPLC (mobile phase: H2O + 0.1% FA, ACN + 0.1% FA column C18 preparation) to provide a yellow foamy compound 2-[2-(3,3-difluoroazetidine-1-carbonyl)-1-methyl-1H-indol-6-yl]-N-[4-(1H-imidazol-5-yl)phenyl]pyrimidin-4-amine, formate salt (0.060 g, 35%). 1 H NMR (400MHz, DMSO-d6) δ9.69(s,1H),8.56(s,1H),8.41(d,J=5.8Hz,1H),8.28(d,J=7.2Hz,2H),8.21(dd,J=8.5,1.4Hz,1H),7.81(s,4H ), 7.75 (d, J = 8.5Hz, 1H), 7.71 (d, J = 1.1Hz, 1H), 7.55 (s, 1H), 7.08 (s, 1H), 6.73 (d, J = 5.8Hz, 1H), 4.73 (d, J = 111.7Hz, 4H), 4.05 (s, 3H). LCMS,m / z:[M+H] + =486.41.

[0216] Example 2

[0217] Method B - Taking the synthesis of 2-[2-(3,3-difluoroazetidine-1-carbonyl)-1-methyl-1H-indol-6-yl]-N-[4-(1H-1,2,3-triazol-4-yl)phenyl]pyrimidin-4-amine as an example.

[0218]

[0219] Step 1': Intermediate 2': Synthesis of 1-(azidomethyl)-4-methoxybenzene

[0220] A mixture of 4-methoxybenzyl chloride (2.00 g, 12.77 mmol, 1.00 equiv) and sodium azide (0.99 g, 15.32 mmol, 1.20 equiv) in anhydrous dimethyl sulfoxide (20.0 mL) was stirred at room temperature overnight. The progress of the reaction was monitored by TLC analysis (eluent: 9:1 DCM:MeOH). The mixture was partitioned between water and EtOAc. The aqueous phase was extracted with EtOAc. The combined organic layers were washed with brine, dried over Na2SO4, filtered and evaporated to dryness to give 1-(azidomethyl)-4-methoxybenzene (1.98 g, 94%) as a pale yellow liquid.

[0221] Step 2': Synthesis of 4-{1-[(4-methoxyphenyl)methyl]-1H-1,2,3-triazol-4-yl}aniline

[0222] To a mixture of 1-(azidomethyl)-4-methoxybenzene (1.98 g, 12.013 mmol, 1.0 equiv) and 4-ethynylaniline (1.40 g, 12.00 mmol, 1.00 equiv) in anhydrous dimethyl sulfoxide (29.70 mL) was added copper (II) sulfate pentahydrate (0.60 g, 2.40 mmol, 0.20 equiv), followed by sodium ascorbate (0.92 g, 4.80 mmol, 0.40 equiv), and the resulting mixture was stirred at room temperature for 2 h. Complete conversion of the starting material was observed according to UPLC-MS analysis. The mixture was poured into cold water. The precipitated solid was filtered, washed with water and dried to give 4-{1-[(4-methoxyphenyl)methyl]-1H-1,2,3-triazol-4-yl}aniline (2.5 g, 73%) as a yellow solid. 1 H NMR(300MHz,DMSO-d6)δ8.26(s,1H),7.48(d,J=8.0Hz,2H),7.37–7.23(m,2H),7 .00–6.87(m,2H),6.58(d,J=8.0Hz,2H),5.49(s,2H),5.19(s,2H),3.74(s,3H). UPLC-MS,m / z:[M+H] + =281.10.

[0223] Step 3': Synthesis of 2-chloro-N-(4-{1-[(4-methoxyphenyl)methyl]-1H-1,2,3-triazol-4-yl}phenyl)pyrimidin-4-amine

[0224] To a solution of 4-{2-[(4-methoxyphenyl)methyl]-2H-1,2,3-triazol-4-yl}aniline (2.50 g, 8.83 mmol, 1.00 equiv) in ethanol (15.0 mL) was added N,N-diisopropyl-ethylamine (DIPEA) (1.999 mL, 11.48 mmol, 1.30 equiv) and the reaction mixture was stirred at room temperature for 15 minutes. 2,4-dichloropyrimidine (1.31 g, 8.83 mmol, 1.00 equiv) was then added and the reaction mixture was stirred at 90 °C overnight. It was evaporated to dryness. The crude product was purified by FCC (0-5% MeOH in DCM) to give 2-chloro-N-(4-{1-[(4-methoxyphenyl)methyl]-1H-1,2,3-triazol-4-yl}phenyl)pyrimidin-4-amine (1.43 g, 40%) as a yellow solid. 1 H NMR (300MHz, DMSO-d6) δ10.13(s,1H),8.52(s,1H),8.18(d,J=5.9Hz,1H),7.83(d,J=8.6Hz,2H),7.66(d, J=8.3Hz,2H),7.34(d,J=8.4Hz,2H),7.01–6.91(m,2H),6.79(d,J=5.9Hz,1H),5.56(s,2H),3.74(s,3H). UPLC-MS,m / z:[M+H] - =392.10.

[0225] Step 4': 2-[2-(3,3-Difluoroazetidine-1-carbonyl)-1-methyl-1H-indol-6-yl]-N-(4-{1-[(4-methoxyphenyl)methyl]-1H-1,2,3-triazol-4-yl}phenyl)pyrimidin-4-amine.

[0226] A mixture of intermediate 1 (0.35 g, 0.42 mmol, 1.10 equivalents), intermediate 2' (0.25 g, 0.38 mmol, 1.00 equivalents) and sodium carbonate (0.12 g, 1.14 mmol, 3.00 equivalents) was dissolved in a mixture of dimethoxyethane (6.0 mL) and water (2.0 mL) and degassed with argon for 10 minutes. Then, tetrakis (triphenylphosphine) palladium (0.04 g, 0.04 mmol, 0.10 equivalents) was added and the reaction mixture was stirred at 110 ° C in a microwave reactor. UPLC-MS was used to analyze and monitor the reaction process, and the raw material was completely converted after stirring for 4 hours. After cooling, the mixture was diluted with EtOAc and filtered through a diatomaceous earth pad. The filtrate was concentrated to dryness to give a crude compound 2-[2-(3,3-difluoroazetidine-1-carbonyl)-1-methyl-1H-indol-6-yl]-N-(4-{1-[(4-methoxyphenyl)methyl]-1H-1,2,3-triazol-4-yl}phenyl)pyrimidin-4-amine (0.2 g), which was used in the next step without further purification. UPLC-MS, m / z: [M=H] + =607.10. Not yet completed 1 H NMR analysis.

[0227] Step 5': 2-[2-(3,3-Difluoroazetidine-1-carbonyl)-1-methyl-1H-indol-6-yl]-N-[4-(1H-1,2,3-triazol-4-yl)phenyl]pyrimidin-4-amine.

[0228] 2-[2-(3,3-Difluoroazetidine-1-carbonyl)-1-methyl-1H-indol-6-yl]-N-(4-{2-[(4-methoxyphenyl)methyl]-2H-1,2,3-triazol-4-yl}phenyl)pyrimidin-4-amine (0.20 g, 0.20 mmol, 1.00 equiv) was dissolved in trifluoroacetic acid (1.06 mL, 13.85 mmol, 70.00 equiv) and the reaction mixture was stirred overnight at 100° C. After this time, no partial conversion was observed. The reaction mixture was evaporated to dryness and purified using preparative HPLC (mobile phase: ACN + 0.1% FA, H2O + 0.1% FA, column C18 preparation) to give 2[2-(3,3-difluoroazetidine-1-carbonyl)-1-methyl-1H-indol-6-yl]-N-[4-(1H-1,2,3-triazol-4-yl)phenyl]pyrimidin-4-amine (0.025 g, 26%). 1H NMR (400MHz, DMSO-d6) δ9.89(s,1H),8.57(s,1H),8.44(d,J=5.9Hz,1H),8.32(d,J=16.4Hz,2H),8.21(d,J=8.6Hz ,1H),7.93(q,J=8.6Hz,4H),7.76(d,J=8.6Hz,1H),7.09(s,1H),6.78(d,J=5.8Hz,1H),4.86(s,4H),4.06(s,3H). LCMS,m / z:[M=H] + =487.18.

[0229] Example 3

[0230] Method C. The synthesis of 2-[2-(3,3-difluoroazetidine-1-carbonyl)-4H,5H,6H,7H-thieno[2,3-c]pyridin-6-yl]-N-[4-(1H-imidazol-4-yl)phenyl]pyrimidin-4-amine (Example 3) and 2-[2-(3,3-difluoroazetidine-1-carbonyl)-4H,5H,6H,7H-thieno[2,3-c]pyridin-6-yl]-N-[4-(1H-1,2,3-triazol-4-yl)phenyl]pyrimidin-4-amine (Example 3') are taken as examples.

[0231]

[0232] Step 1 and Step 2: Intermediate 1: Synthesis of 3,3-difluoro-1-{4H,5H,6H,7H-thieno[2,3-c]pyridine-2-carbonyl}azetidine

[0233] 6-[(tert-Butyloxy)carbonyl]-4H,5H,6H,7H-thieno[2,3-c]pyridine-2-carboxylic acid (0.20 g, 0.70 mmol, 1.00 equiv) was dissolved in DMF (5 mL) and DIPEA (0.50 mL, 2.83 mmol, 4.00 equiv) was then added to the reaction mixture followed by HATU (0.32 g, 0.84 mmol, 1.20 equiv). The reaction mixture was stirred at room temperature for 15 minutes and 3,3-difluoroazetidine hydrochloride (0.11 g, 0.85 mmol, 1.20 equiv) was added. The reaction mixture was stirred at room temperature overnight. The reaction mixture was quenched by the addition of water and the aqueous layer was extracted with EtOAc. The combined organic layers were washed with brine, dried over Na2SO4, filtered and concentrated to give tert-butyl 2-(3,3-difluoroazetidine-1-carbonyl)-4H,5H,6H,7H-thieno[2,3-c]pyridine-6-carboxylate, [M+ACN] +=358.90. The crude material was used in the next step without further purification. To the resulting residue was added 2M HCl in distilled water. 1-(4H,5H,6H,7H-thieno[2,3-c]pyridine-2-carbonyl}azetidine) was used directly in the next step. UPLC-MS, m / z: [M+ACN] + =299.80.

[0234] Step 3: Synthesis of 2-[2-(3,3-difluoroazetidine-1-carbonyl)-4H,5H,6H,7H-thieno[2,3-c]pyridin-6-yl]-N-[4-(1H-imidazol-5-yl)phenyl]pyrimidin-4-amine.

[0235] N, N-diisopropylethylamine (DIPEA) (0.23mL, 1.33mmol, 4.00 equivalents) was added to a solution of intermediate 1 (0.17g, 0.33mmol, 1.00 equivalents) in acetonitrile (5.0mL), and the reaction mixture was stirred at room temperature for 15 minutes. Intermediate 2 (0.10g, 0.33mmol, 1.00 equivalents) was then added and the reaction mixture was stirred at 95 ° C overnight. Low conversion of the starting material was observed (according to UPLC-MS analysis). Cs2CO3 (0.21g, 0.66mmol, 2.00 equivalents) was added to the reaction mixture, and stirring was continued for another 12hr at the same temperature. The reaction mixture was then quenched by adding water and extracted with EtOAc. The combined organic layers were dried over Na2SO4, filtered, concentrated and purified by preparative HPLC (mobile phase: ACN + 0.1% FA, H2O + 0.1% FA column C18 preparation) to provide 2-[2-(3,3-difluoroazetidine-1-carbonyl)-4H,5H,6H,7H-thieno[2,3-c]pyridin-6-yl]-N-[4-(1H-imidazol-5-yl)phenyl]pyrimidin-4-amine (0.003 g, 2%). 1 H NMR (400MHz, DMSO-d6) δ9.37 (s, 1H), 8.42 (s, 5H), 7.97 (d, J = 5.7Hz, 1H), 7.67 (dd, J = 26.6, 10.4Hz, 6 H), 7.51 (s, 1H), 7.34 (s, 1H), 6.11 (d, J = 5.7Hz, 1H), 4.98 (s, 2H), 4.04 (t, J = 5.8Hz, 3H), 2.75 (s, 3H). LCMS,m / z:[M+H] + =493.85.

[0236] Step 4: 2-[2-(3,3-Difluoroazetidine-1-carbonyl)-4H,5H,6H,7H-thieno[2,3-c]pyridin-6-yl]-N-[4-(1H-1,2,3-triazol-4-yl)phenyl]pyrimidin-4-amine.

[0237] 2-[2-(3,3-Difluoroazetidine-1-carbonyl)-4H,5H,6H,7H-thieno[2,3-c]pyridin-6-yl]-N-(4-{1-[(4-methoxyphenyl)methyl]-1H-1,2,3-triazol-4-yl}phenyl)pyrimidin-4-amine (0.20 g, 0.20 mmol, 1.00 equiv) was dissolved in trifluoroacetic acid (1.06 mL, 13.85 mmol, 70.00 equiv) and the reaction mixture was stirred overnight at 100° C. After this time, no partial conversion was observed. The reaction mixture was evaporated to dryness and purified using preparative HPLC (mobile phase: ACN + 0.1% FA, H2O + 0.1% FA, column C18 preparation) to give 22-[2-(3,3-difluoroazetidine-1-carbonyl)-4H,5H,6H,7H-thieno[2,3-c]pyridin-6-yl]-N-[4-(1H-1,2,3-triazol-4-yl)phenyl]pyrimidin-4-amine (0.025 g, 26%). 1 H NMR (400MHz, DMSO-d6) δ9.50(s,1H),8.35(s,1H),8.27(s,1H),8.00(d,J=5.7Hz,1H),7.83(d,J=8.7Hz,2H),7.75(d,J= 8.7Hz, 2H), 7.34 (s, 1H), 6.13 (d, J = 5.7Hz, 1H), 4.99 (s, 2H), 4.67 (s, 4H), 4.05 (t, J = 5.7Hz, 2H), 2.76 (d, J = 6.0Hz, 2H). Formate. LCMS, m / z: [M+H]+=495.11.

[0238] Example 4

[0239] (Method A)

[0240]

[0241] 1H NMR (300MHz, DMSO-d6) δ12.51(s,1H),9.71(s,1H),8.47(s,1H),8.40(d,J=5.9Hz,1H),8.20(dd,J=8.4,1.3Hz,1H),8.14(s,1H),7.81(d,J=2.6 Hz,5H),7.75(d,J=8.4Hz,1H),7.55(s,1H),7.09(s,1H),6.71(d,J=5.9 Hz, 1H), 5.52 (s, 2H), 4.69 (d, J = 89.6Hz, 4H), 3.18 (s, 3H), 2.86 (s, 3H). Formate. LCMS,m / z:[M+H] + =557.33.

[0242] Example 5

[0243] (Method A)

[0244]

[0245] 1 H NMR (400 MHz, DMSO-d6) δ 9.66 (s, 1H), 8.55 (s, 1H), 8.40 (d, J = 5.8 Hz, 1H), 8.19 (dd, J = 8.4, 1.4 Hz, 1H), 8.16 (s, 1H), 7.80 (s, 4H), 7.73 (d, J = 8.4 Hz, 1H), 7.70 (d, J = 1.1 Hz, 1H), 7.51 (s, 1H), 7.00 (s, 1H), 6.71 (d, J = 5.9 Hz, 1H), 4.87–4.37 (m, 6H), 2.14 (s, 6H). Two aliphatic protons CH2 overlap with H2O. Formate. LCMS, m / z: [M+H] + =543.23.

[0246] Example 6

[0247] (Method A)

[0248]

[0249] 1H NMR (400MHz, DMSO-d6) δ9.70(s,1H),8.64(s,1H),8.41(d,J=5.8Hz,1H),8.27(s,1H),8.21(dd,J=8.4,1.3Hz,1H),7.82(d,J=4.9Hz,4H ),7.75(d,J=8.4Hz,1H),7.70(d,J=1.0Hz,1H),7.49(s,1H),7.12(s,1H),6.73(d,J=5.8Hz,1H),5.02–4.41(m,10H),3.62–3.52(m,1H). Formate. LCMS,m / z:[M+H] + =542.14.

[0250] Example 7

[0251] (Method A)

[0252]

[0253] 1 H NMR (300MHz, DMSO-d6) δ9.68(s,1H),8.54(s,1H),8.40(d,J=5.9Hz,1H),8.24–8.14(m,2H),7.87–7.77(m,4H),7.74(d,J=8 .4Hz,1H),7.70(s,1H),7.53(s,1H),7.02(s,1H),6.72(d,J=5.9Hz,1H),4.31–3.96(m,4H),3.95(s,3H),3.91–3.71(m,2H). Formate. LCMS,m / z:[M+H] + =500.39.

[0254] Example 8

[0255] (Method A)

[0256]

[0257] 1H NMR(400MHz,DMSO-d6)δ9.73(s,1H),9.10(d,J=8.8Hz,1H),8.56(s,1H),8.41(d,J=5 .8Hz,1H),8.33(s,4H),8.20(dd,J=8.4,1.3Hz,1H),7.87–7.75(m,5H),7.71(d,J=1. 0Hz,1H),7.50(s,1H),7.23(s,1H),6.73(d,J=5.8Hz,1H),4.87(dt,J=15.3,7.7Hz,1 H), 4.72 (t, J = 6.9Hz, 2H), 2.60 (t, J = 6.8Hz, 2H), 2.16 (s, 6H), 1.40 (d, J = 7.0Hz, 3H). Formate (1:4). LCMS,m / z:[M+H] + =563.23.

[0258] Example 9

[0259] (Method A)

[0260]

[0261] 1 H NMR(300MHz, DMSO-d6)δ9.70(s,1H),8.99(d,J=8.9Hz,1H),8.56(s,1H),8.41(d,J=5.8Hz,1H),8.31(s,2H),8.20(dd,J=8.4,1.3Hz,1H),7.8 6–7.75(m,4H),7.70(d,J=1.1Hz,1H),7.53(s,1H),7.30(s,1H),6.73( d,J=5.9Hz,1H),4.99–4.80(m,1H),4.09(s,3H),1.40(d,J=7.1Hz,3H). Formate. LCMS,m / z:[M+H] + =506.18.

[0262] Example 10

[0263] (Method A)

[0264]

[0265] 1H NMR (400MHz, DMSO-d6) δ12.12(s,1H),9.70(s,1H),9.06(d,J=8.9Hz,1H),8.63(s,1H),8. 42(d,J=5.8Hz,1H),8.21(d,J=8.7Hz,1H),7.88–7.76(m,5H),7.71(s,1H),7.52(s,1H),7 .32(s,1H),6.73(d,J=5.9Hz,1H),5.00(dt,J=15.4,7.5Hz,2H),4.95–4.84(m,1H),4.61( dt,J=7.9,6.1Hz,2H), 4.46(t,J=6.1Hz,2H), 3.51(p,J=7.0Hz,1H), 1.40(d,J=7.0Hz,3H). LCMS,m / z:[M+H] + =562.51.

[0266] Embodiment 11

[0267] (Method A)

[0268]

[0269] 1 H NMR (400MHz, DMSO-d6) δ12.11(s,1H),9.69(s,1H),8.61(s,1H),8.41(d,J=5.8Hz, 1H),8.20(dd,J=8.4,1.3Hz,1H),7.82(s,4H),7.74(d,J=8.5Hz,1H),7.71(d,J=1.1 Hz,1H),7.52(s,1H),7.04(s,1H),6.72(d,J=5.8Hz,1H),4.81(d,J=7.4Hz,2H),4. 62 (dd, J = 7.8, 6.1 Hz, 2H), 4.42 (s, 2H), 3.98 (dd, J = 84.9, 48.3 Hz, 4H), 3.47 (s, 2H). One proton CH overlaps with H2O. LCMS, m / z: [M+H] + =556.14.

[0270] Example 12

[0271] (Method A)

[0272]

[0273] 1H NMR (400MHz, DMSO-d6) δ9.68 (s, 1H), 8.53 (s, 1H), 8.40 (d, J = 5.8Hz, 1H), 8.3 0(s,3H),8.18(dd,J=8.4,1.4Hz,1H),7.84–7.77(m,4H),7.72(d,J=8.4Hz,1H ),7.70(d,J=1.1Hz,1H),7.52(s,1H),6.90(s,1H),6.72(d,J=5.9Hz,1H),3.9 3(s,3H),3.66(t,J=6.3Hz,2H),3.55(t,J=6.6Hz,2H),1.91(d,J=8.3Hz,4H). Formate (1:3). LCMS,m / z:[M+H] + =464.39.

[0274] Example 13

[0275] (Method A)

[0276]

[0277] 1 H NMR (400MHz, DMSO-d6) δ9.70 (s, 1H), 8.57 (s, 1H), 8.42 (d, J = 5.8 Hz, 1H), 8.21 (dd, J = 8.5, 1.4 Hz, 1H), 8.14 (s, 1H), 7.87–7.73 (m, 5H), 7.55 (s, 1H), 7.06 (s, 1H), 6.73 (d, J = 5.9 Hz, 1H), 4.66 (s, 2H), 4.20 (s, 2H), 4.00 (s, 2H), 3.80 (s, 2H), 3.05 (s, 2H). Six aliphatic protons 2xCH3 overlap with DMSO-d6. LCMS, m / z: [M+H] + =557.20.

[0278] Embodiment 14

[0279] (Method A)

[0280]

[0281] 1H NMR (400MHz, DMSO-d6) δ9.75(s,1H),8.58(s,1H),8.44–8.39(m,2H),8.28(s,2H),7.91(d,J=8.3Hz,1H),7.80(q,J=8 .6Hz, 4H), 7.70 (d, J = 1.1Hz, 1H), 7.66 (d, J = 1.0Hz, 1H), 7.55 (s, 1H), 6.77 (d, J = 5.9Hz, 1H), 5.12 (s, 2H), 4.56 (s, 2H). Formate (1:2). LCMS,m / z:[M+H] + =473.29.

[0282] Embodiment 15

[0283] (Method B)

[0284]

[0285] 1 H NMR (300 MHz, DMSO-d6) δ9.82 (s, 1H), 8.56 (s, 1H), 8.44 (d, J = 5.8 Hz, 1H), 8.28 (s, 1H), 8.20 (dd, J = 8.4, 1.3 Hz, 1H), 8.14 (s, 1H), 7.97–7.87 (m, 4H), 7.74 (d, J = 8.4 Hz, 1H), 7.01 (s, 1H), 6.75 (d, J = 5.8 Hz, 1H), 4.64 (t, J = 6.0 Hz, 6H), 2.17 (s, 6H). Two aliphatic protons CH2 overlap with DMSO. Formate. LCMS, m / z: [M+H] + =544.38.

[0286] Example 16

[0287] (Method B)

[0288]

[0289] 1H NMR (400MHz, DMSO-d6) δ9.84(s,1H),8.53(s,1H),8.44(s,1H),8.40(s,3H),8.30(s,1H),8.19(dd,J=8.4,1.4Hz,1H),7.93(q,J=8 .7Hz, 4H), 7.73 (d, J = 8.4Hz, 1H), 6.91 (s, 1H), 6.76 (d, J = 5.8Hz, 1H), 3.94 (s, 3H), 3.67 (s, 2H), 3.54 (d, J = 6.9Hz, 2H), 1.90 (s, 4H). Formate (1:3). LCMS,m / z:[M+H] + =465.37.

[0290] Embodiment 17

[0291] (Method B)

[0292]

[0293] 1 H NMR (400 MHz, DMSO-d6) δ 9.84 (s, 1H), 8.55 (s, 1H), 8.44 (d, J = 5.8 Hz, 1H), 8.35 (s, 1H), 8.30 (s, 1H), 8.20 (dd, J = 8.4, 1.4 Hz, 1H), 7.93 (q, J = 8.8 Hz, 4H), 7.75 (d, J = 8.5 Hz, 1H), 7.02 (s, 1H), 6.76 (d, J = 5.8 Hz, 1H), 4.20 (s, 2H), 3.95 (s, 4H), 3.81 (s, 1H). Two aliphatic protons CH2 overlap with H2O. Formate. LCMS, m / z: [M+H] + =501.20.

[0294] Embodiment 18

[0295] (Method B)

[0296]

[0297] 1H NMR (400 MHz, DMSO-d6) δ 9.81 (s, 1H), 8.55 (s, 1H), 8.44 (d, J = 5.8 Hz, 1H), 8.26 (s, 1H), 8.19 (dd, J = 8.4, 1.3 Hz, 1H), 8.14 (s, 1H), 7.92 (q, J = 8.7 Hz, 4H), 7.74 (d, J = 8.4 Hz, 1H), 6.97 (d, J = 3.2 Hz, 1H), 6.75 (d, J = 5.8 Hz, 1H), 4.57 (s, 2H), 4.14 (s, 2H), 3.96 (s, 2H), 3.78 (s, 2H), 2.19 (s, 6H). Two aliphatic protons CH2 overlap with DMSO-d6. Formate. LCMS, m / z: [M+H] + =558.16.

[0298] Embodiment 19

[0299] (Method B)

[0300]

[0301] 1 H NMR (400MHz, DMSO-d6) δ9.91 (s, 1H), 8.58 (s, 1H), 8.46 (d, J = 5.8 Hz, 1H), 8.42 (dd, J = 8.2, 1.4 Hz, 1H), 8.39 (s, 1H), 8.32 (s, 1H), 7.96–7.88 (m, 5H), 7.67 (d, J = 1.0 Hz, 1H), 6.81 (d, J = 5.9 Hz, 1H), 5.12 (s, 2H), 4.56 (s, 2H). Formate. LCMS, m / z: [M+H] + =474.20.

[0302] Embodiment 20

[0303] (Method B)

[0304]

[0305] 1H NMR (400 MHz, DMSO-d6) δ 12.14 (s, 1H), 9.74 (s, 1H), 8.98 (d, J = 1.4 Hz, 1H), 8.46 (dd, J = 8.5, 1.5 Hz, 1H), 8.43 (d, J = 5.9 Hz, 1H), 8.08 (d, J = 8.5 Hz, 1H), 8.01 (s, 1H), 7.79 (s, 4H), 7.71 (d, J = 1.1 Hz, 1H), 7.59 (d, J = 20.2 Hz, 1H), 6.77 (d, J = 5.9 Hz, 1H), 4.85 (d, J = 199.8 Hz, 4H). The residue contains formic acid. LCMS, m / z: [M+H] + =489.10.

[0306] Embodiment 21

[0307] (Method B)

[0308]

[0309] 1 H NMR (400MHz, DMSO-d6) δ13.60(d,J=36.6Hz,1H),9.72(s,1H),8.68(d,J=34.4Hz,1H),8.50–8.33(m,2H),8.20(d,J=3.9Hz, 2H), 7.81 (s, 5H), 7.71 (d, J = 1.1Hz, 1H), 7.53 (s, 1H), 6.73 (d, J = 5.8Hz, 1H), 5.16 (d, J = 13.0Hz, 2H), 4.62 (t, J = 12.3Hz, 2H). Formate. LCMS,m / z:[M+H] + =472.88.

[0310] Embodiment 22

[0311] (Method B)

[0312]

[0313] 1 H NMR (400MHz, DMSO-d6) δ13.65(s,1H),9.85(s,1H),8.67(s,1H),8.44(d,J=5.8Hz,1H),8.37(s,2H),8.30(s,1 H), 7.93 (s, 4H), 7.81 (d, J = 12.2Hz, 1H), 6.77 (d, J = 5.8Hz, 1H), 5.18 (t, J = 12.3Hz, 2H), 4.62 (t, J = 12.2Hz, 2H). LCMS,m / z:[M+H]+ =474.25.

[0314] Embodiment 23

[0315] (Method C)

[0316]

[0317] 1 H NMR (400MHz, DMSO-d6) δ9.34 (s, 1H), 8.17 (s, 2H), 7.97 (d, J = 5.7 Hz, 1H), 7.76–7.67 (m, 3H), 7.63 (d, J = 8.4 Hz, 2H), 7.50 (s, 1H), 7.38 (s, 1H), 6.10 (d, J = 5.7 Hz, 1H), 4.96 (s, 2H), 4.12 (s, 1H), 4.04 (t, J = 5.8 Hz, 4H), 3.77 (s, 2H), 2.75 (s, 2H). Formate (1:2). LCMS, m / z: [M+H] + =520.11.

[0318] Embodiment 24

[0319] (Method C)

[0320]

[0321] 1 H NMR (400MHz, DMSO-d6) δ9.49(s,1H),8.32(s,2H),8.27(s,1H),8.00(d,J=5.7Hz,1H),7.88–7.80(m,2H),7.75(d,J=8.7Hz,2H ),7.38(s,1H),6.13(d,J=5.7Hz,1H),4.97(s,2H),4.12(s,1H),4.05(t,J=5.9Hz,4H),3.77(s,2H),2.76(s,3H),2.61(s,1H). Formate (1:2). LCMS,m / z:[M+H] + =521.09.

[0322] Embodiment 25

[0323] (Method C)

[0324]

[0325] 1H NMR (400MHz, DMSO-d6) δ12.12(s,1H),i9.34(s,1H),8.28(s,2H),7.97(d,J=5.6Hz,1H),7.73(s,2H),7.68(d,J=1.1Hz,1H),7.63(d,J=7.9 Hz,2H),7.54(s,1H),7.42(s,1H),6.10(d,J=5.7Hz,1H),4.97(s,2H),4.21(bs,1H),4.04(t,J=5.8Hz,3H),3.96–3.69(m,4H),2.76(s,2H). Formate (1:2). LCMS,m / z:[M+H] + =508.42.

[0326] Comparative Example

[0327] The structure of comparative compound A is provided below:

[0328]

[0329] It can be prepared according to the method found in WO 2019 / 001572.

[0330] Embodiment 26

[0331] ADP-Glo ​​Kinase Assay (Promega). TM The kinase assay is a luminescent ADP detection assay that allows the measurement of kinase activity based on the amount of ADP produced during the kinase reaction. The kinase reaction is performed in the presence of ATP, S6K substrate (KRRRLASLR) and ROCK kinase in the appropriate kinase reaction buffer (1x). At the end of the reaction, the ADP-Glo TM The reagent depletes unconsumed ATP. Next, a kinase detection reagent is added to convert ADP to ATP, and the newly synthesized ATP is measured using the luciferase / luciferin reaction. Luminescence is proportional to the ADP produced, and therefore proportional to the kinase activity. ADP-Glo ​​was performed in a 384-well plate as follows TM Kinase assays.

[0332] Kinase Assay Reagent Preparation: Thaw the Kinase Assay Buffer at room temperature (RT). If a precipitate is present, incubate the buffer at 37°C with constant vortexing for 15 minutes to dissolve the precipitate or remove the precipitate from the buffer by carefully aspirating the supernatant from the bottle. After both the Kinase Assay Buffer and the Kinase Assay Substrate are equilibrated to room temperature, transfer the entire volume of the Kinase Assay Buffer to the bottle containing the Kinase Assay Substrate to reconstitute the lyophilized substrate, followed by gentle mixing to obtain a homogenous solution.

[0333] Kinase reaction buffer preparation: Kinase reaction buffer was prepared freshly before each experiment. The formulation of this buffer is provided in Table 1 below:

[0334] Table 1. Preparation of Kinase Reaction Buffer (1x) for ADP-Glo ​​Kinase Assay.

[0335] Reagents: Final concentration Stock concentration Tris, pH 7.5 40mM 500mM aqueous solution DTT 0.05mM 100mM aqueous solution <![CDATA[MgCl2]]> 20mM 500mM aqueous solution BSA 0.1mg / mL 5.0mg / mL

[0336] ADP-Glo ​​Reagent Preparation. Prepare three mixtures on ice before performing the assay:

[0337] Mix 1 – contains ROCK1 or ROCK2 (1x) in kinase reaction buffer

[0338] Mix 2 – contains S6K peptide (1x) in kinase reaction buffer

[0339] Mix 3 – contains ATP (1x) in kinase reaction buffer

[0340] The final conditions for performing the ADP-Glo ​​Kinase Assay are shown in Table 2 below:

[0341] Table 2. Final conditions for the ADP-Glo ​​Kinase Assay for ROCK (concentrations in a final volume of 5 μL).

[0342] Final Concentration: ROCK1 ROCK2 Kinase 0.1ng / μL 0.1ng / μL S6K substrate 170μM 170μM ATP 10μM 40μM Time 120 minutes 120 minutes DMSO 1％ 1％

[0343] The volume ratio of the reagents in the assay is 1:1:2: Kinase reactant: ADP-Glo TM Reagents: Kinase Assay Reagents. For 384-well plates, volumes are: 5 μl Kinase Reaction, 5 μl ADP-Glo TM Reagent and 10 μl kinase assay reagent.

[0344] Generate a standard curve for the conversion of ATP to ADP. To assess the amount of ADP produced in a kinase reaction, a standard curve representing the luminescence corresponding to the conversion of ATP to ADP is prepared based on the ATP concentration used in the kinase reaction. These conversion curves represent the amount of ATP and ADP available in the reaction at a specified conversion percentage ranging from 100% conversion to 0% conversion. The standard samples used to generate the ATP-ADP conversion curves are generated by combining the appropriate volumes of ATP and ADP stock solutions.

[0345] IC of kinase inhibitors 50 Determination of value

[0346] Each plate contained a positive control (1 μM Compound A for ROCK2 and 1 μM RKI-1447 for ROCK1); a vehicle control (1% DMSO); a blank control (kinase reaction buffer (1×)); a low control (substrate and ATP, no kinase); and an autophosphorylation control (kinase and ATP, no substrate). Test and reference compounds were diluted in 100% DMSO to obtain a 10 mM stock solution. Each compound was tested in duplicate in an 8-fold serial dilution. The final concentration of DMSO in the reaction was 1% (up to 50 nL).

[0347] 5 μl / well kinase reaction buffer (1x) was dispensed into the blank control wells. 2 μl / well kinase reaction buffer (1x) was dispensed into the control wells without protein (low control) and without substrate (autophosphorylation). The plate was sealed with adhesive film and then briefly spun (1 min, 1000 rpm).

[0348] 2 μl / well kinase (2.5x) solution was dispensed into the relevant wells of the assay plate. The plate was sealed with adhesive film and then briefly spun (1 min, 1000 rpm).

[0349] 2 μl / well S6K substrate (2.5x) solution was dispensed into the relevant wells of the assay plate. The plate was sealed with adhesive film and then briefly spun (1 min, 1000 rpm).

[0350] Dispense 1 μl / well ATP (5x) solution into the relevant wells of the assay plate. The final volume of the reaction is 15 μl. Seal the plate with adhesive film and spin briefly (1 min, 1000 rpm).

[0351] The plate was incubated at 25°C with shaking at 450 rpm for 120 minutes.

[0352] After the incubation is complete, the kinase reaction is terminated, residual ATP is depleted from the kinase reaction and ADP is converted to ATP, which is measured in the luciferase / luciferin reaction. Dispense 5 μl of ADP-Glo TM The plate was sealed with adhesive film, followed by a short spin (1 min, 1000 rpm) and incubated at 25°C with a shaking speed of 450 rpm for 40 min.

[0353] Dispense 10 μl of kinase assay reagent to convert ADP to ATP and introduce luciferase and luciferin to detect ATP.The plate is sealed with adhesive film, followed by a short spin (1 min, 1000 rpm) and incubated at 25°C with a shaking speed of 450 rpm for 60 min.

[0354] After 5 minutes of dark adaptation, luminescence was measured using a PHERAstar FSX multi-mode plate reader (BMG Labtech).

[0355] IC was calculated using a 4-parameter model using GraphPad Prism 7.04 software after normalization. 50 Parameters: Log(Inhibitor) vs. Response - variable slope.

[0356]

[0357]

[0358]

[0359]

[0360] Embodiment 28

[0361] Selective ROCK2 inhibition downregulates IL-17 secretion via a pSTAT3-dependent mechanism

[0362] Rho kinase (ROCK) is a member of the serine / threonine kinase family and is often studied for its role in cell morphology, motility and shape through its effects on the cytoskeleton. Although the two isoforms of ROCK, ROCK1 and ROCK2, share more than 90% homology within their kinase domains, the functions of these proteins are not redundant and depend on the cell system in which ROCK is expressed and activated. Recent studies have shown that ROCK2, rather than ROCK1, regulates the proinflammatory IL-17 production lineage of T cells called Th17 cells via a STAT3-dependent mechanism. Dysregulated activation of Th17 cells and STAT3 phosphorylation have been implicated in the pathogenesis of inflammatory plethora and fibrotic pathologies. In addition, selective ROCK2 inhibition alters the balance between proinflammatory Th17 and immunosuppressive regulatory T cells (Treg) by increasing STAT5 phosphorylation and IL-10 secretion.

[0363] By using stimulatory antibodies against CD3 and CD28 (anti-CD3 / CD28) in combination with IL-1β and TGF-β (Th17-biased conditions), we found that novel selective ROCK2 inhibitors, such as the compounds of Example 1 and Example 2, robustly down-regulate IL-17 secretion in Th17-biased human CD4+ T cells in a dose-dependent manner with EC50 at 838 nM and 764 nM, respectively (see Figure 2A and Figure 2B In this study, peripheral blood CD4 +T cells were then stimulated with anti-CD3 / 28 mAb, IL-1β (50 ng / mL) and TGF-β (5 ng / mL) for 48 hours. Supernatants were analyzed for IL-17 by ELISA. One representative of three different experiments is shown.

[0364] The induction and expression of IL-17, as well as other proinflammatory cytokines, have been shown to depend on the activation and phosphorylation of certain transcription factors, including STAT3. + T cells resulted in a dose-dependent decrease in STAT3 phosphorylation ( Figure 3 ), which is consistent with the ability of the compounds of Example 1 and Example 2 to downregulate IL-17 secretion and pCofilin (a classical downstream ROCK target) under the same stimulation conditions ( Figure 4 In these studies, peripheral blood CD4 + T cells were then stimulated for 2 hours with anti-CD3 / 28 mAb, IL-1β (50 ng / mL) and TGF-β (5 ng / mL). Cell lysates were prepared and analyzed by Western blot. One representative of three different experiments is shown.

[0365] Embodiment 27

[0366] Selective ROCK2 inhibitor downregulates pro-fibrogenic gene expression in fibroblasts

[0367] ROCK is activated by the small GTPase Rho in response to a variety of profibrotic signals and regulates cytoskeletal dynamics, activation of different downstream intracellular targets and expression of key profibrotic genes such as fibronectin, smooth muscle actin (α-SMA) and collagen 3 (Col3A1). By using MRC-5 human fibroblasts pretreated with different doses of a selective ROCK2 inhibitor, we found that both the compounds of Example 1 and Example 2 dose-dependently downregulated TGF-β-induced fibronectin, α-SMA and Col3A1 gene expression as determined by PCR ( Figure 5 ) and Col1A protein production in the supernatant determined by ELISA ( Figure 6 ). These data further demonstrate that selective ROCK2 inhibition has a direct and robust anti-fibrotic effect.

[0368] In these studies, human lung fibroblast MRC-5 cells were obtained from the American Type Culture Collection (ATCC) and cultured in MEM supplemented with 10% fetal bovine serum. The cells were seeded in 24-well cell culture plates at 50,000 / mL / well. The next day, the cells were starved overnight in MEM containing 0.5% FBS. TGF-β1 (2.5 ng / mL) and various concentrations of the compounds of Examples 1 and 2 were applied to the cells for 48 hours. Cell mRNA was isolated and the mRNA expression of the specified genes (fibronectin, αSMA and col3A) was analyzed by RT-PCR ( Figure 5 The secreted procollagen 1a (Col1A1) in the supernatant was measured by ELISA ( Figure 6 ).

[0369] Embodiment 28

[0370] Selective ROCK2 inhibitor effectively downregulates differentiation in human adipocytes

[0371] Adipogenesis is a multi-step complex process that develops from precursor stem cells into adipocytes that synthesize and store fat. Disturbance of adipocyte function has been shown to be involved in the pathogenesis of a wide range of metabolic diseases (such as type 2 diabetes, obesity, etc.). ROCK has been considered to be involved in the regulation of adipocyte differentiation, but the isoform-specific contribution to adipogenesis and other metabolic pathways has not been well characterized. Recent data show that KD025, a selective ROCK2 inhibitor, downregulates adipocyte differentiation in both mouse 3T3-L1 preadipocytes and human adipose-derived stem cells by reducing the expression of key adipogenesis / lipidogenesis genes (such as PPARg, C / EBPa and Glut4). By using primary human adipocytes, we found that the new selective ROCK2 inhibitor Example 1 and Example 2 downregulate adipogenesis in a dose-dependent manner ( Figure 7 ), further confirming the anti-metabolic potential of ROCK2 targeting in cells. In this study, human subcutaneous adipocytes were grown to confluence and then induced to differentiate by a lipogenic cocktail (0.1 μM dexamethasone, 1 μM insulin, 200 μM indomethacin, 250 μM isobutylmethylxanthine (IBMX)) in the presence of specified concentrations of Example 1 and Example 2 for 10 days. Differentiated cells were stained with oil red, extracted with isopropanol and measured by absorbance at 492 nM.

Claims

1. A compound having formula I: or a pharmaceutically acceptable salt thereof, wherein: R 1 Selected from the group consisting of: H, C1-C6 alkyl, C2-C6 alkenyl, C2-C6 alkynyl, C3-C7 cycloalkyl, halo, -CN, C1-C3 perfluoroalkyl, -OR 11 、-O-(C1-C6 alkyl)-OR 11 、-(C1-C6 alkyl)-OR 11 、-NR 11 R 12 、-O-(C1-C6 alkyl)-NR 11 R 12 、-(C1-C6 alkyl)-NR 11 R 12 、-NR 11 -(C1-C6 alkyl)-NR 11 R 12 、-NR 11 -(C1-C6 alkyl)-OR 11 、-(C1-C6 alkyl) x -C(═O)R 11 、-O-(C1-C6 alkyl) x -C(═O)R 11 、-(C1-C6 alkyl) x -C(═O)OR 11 、-C(═O)-R 11 、-C(═O)OR 11 、-(C1-C6 alkyl) x -C(═O)NR 11 R 12 、-NR 11 -(C1-C6 alkyl) x -C(═O)R 11 and NR 11 -(C1-C6 alkyl) x -C(═O)OR 11 ; R 2 Selected from the group consisting of: H, C1-C6 alkyl, C2-C6 alkenyl, C2-C6 alkynyl, C3-C7 cycloalkyl, halo, -CN, C1-C3 perfluoroalkyl, -OR 11 、-O-(C1-C6 alkyl)-OR 11 、-(C1-C6 alkyl)-OR 11 、-NR 11 R 12 、-O-(C1-C6 alkyl)-NR 11 R 12 、-(C1-C6 alkyl)-NR 11 R 12 、-NR 11 -(C1-C6 alkyl)-NR 11 R 12 、-NR 11 -(C1-C6 alkyl)-OR 11 、-(C1-C6 alkyl) x -C(═O)R 11 、-O-(C1-C6 alkyl) x -C(═O)R 11 、-(C1-C6 alkyl) x -C(═O)OR 11 、-C(═O)-R 11 、-C(═O)OR 11 、-(C1-C6 alkyl) x -C(═O)NR 11 R 12 、-NR 11 -(C1-C6 alkyl) x -C(═O)R 11 and-NR 11 -(C1-C6 alkyl) x -C(═O)OR 11 ; Alternatively, R 1 and R 2 The fused ring may contain 0 to 2 ring heteroatoms selected from the group consisting of N, O and S, and the fused ring may be unsubstituted or substituted with 1 to 3 substituents selected from the group consisting of: C1-C6 alkyl, halo, -CN, -OH, oxo, -O-(C1-C6 alkyl), -O-(C1-C6 alkyl)-OH, -O-(C1-C6 alkyl)-O-(C1-C6 alkyl), -NR 11 R 12 、-O-(C1-C6 alkyl)-NR 11 R 12 、C1-C3 perfluoroalkyl、-NR 11 -(C1-C6 alkyl)NR 11 R 12 and-NR 11 -(C1-C6 alkyl)-OR 11 ; X 4 is N or CH; R 3 and R 4 Each is independently selected from the group consisting of: H, C1-C6 alkyl, C2-C6 alkenyl, C2-C6 alkynyl, C3-C7 cycloalkyl, 3 to 10 membered heterocyclyl, C6-C 10 Aryl, 5- to 14-membered heteroaryl, C 6-12 Aralkyl, -(C1-C6 alkyl)-OR 11 、-(C1-C6 alkyl)-NR 11 R 12 、-(C1-C6 alkyl) x -C(═O)R 11 、-(C1-C6 alkyl) x -C(═O)OR 11 and -(C1-C6 alkyl) x -C(═O)NR 11 R 12 ; Alternatively, R 3 and R 4 together with the nitrogen to which they are attached, provide (i) a 4- to 6-membered heterocyclic ring having 0 to 2 additional ring heteroatoms selected from N, O, and S, or (ii) a 5- to 10-membered heterobicyclic ring system having 0 to 3 additional ring heteroatoms selected from N, O, and S; wherein the heterocyclic ring or the heterobicyclic ring system is unsubstituted or substituted with 1 to 4 substituents selected from the group consisting of halo, C1-C6 alkyl, C2-C6 alkenyl, C2-C6 alkynyl, C3-C7 cycloalkyl, halo, -CN, C1-C3 perfluoroalkyl, -OR 11 , oxo, -O-(C1-C6 alkyl)-OR 11 、-(C1-C6 alkyl)-OR 11 、-NR 11 R 12 、-O-(C1-C6 alkyl)-NR 11 R 12 、-(C1-C6 alkyl)-NR 11 R 12 、-NR 11 -(C1-C6 alkyl)-NR 11 R 12 、-NR 11 -(C1-C6 alkyl)-OR 11 、-(C1-C6 alkyl) x -C(═O)R 11 、-O-(C1-C6 alkyl) x -C(═O)R 11 、-(C1-C6 alkyl) x -C(═O)OR 11 、-C(═O)-R 11 、-C(═O)OR 11 、-(C1-C6 alkyl) x -C(═O)NR 11 R 12 、-NR 11 -(C1-C6 alkyl) x -C(═O)R 11 and-NR 11 -(C1-C6 alkyl) x -C(═O)OR 11 ; Dashed lines represent optional double bonds; Each R 5 independently selected from the group consisting of: H, C1-C6 alkyl, C2-C6 alkenyl, C2-C6 alkynyl, C3-C7 cycloalkyl, halo, -CN, C1-C3 perfluoroalkyl, oxo, -OR 11 、-O-(C1-C6 alkyl)-OR 11 、-(C1-C6 alkyl)-OR 11 、-NR 11 R 12 、-O-(C1-C6 alkyl)-NR 11 R 12 、-(C1-C6 alkyl)-NR 11 R 12 、-NR 11 -(C1-C6 alkyl)-NR 11 R 12 、-NR 11 -(C1-C6 alkyl)-OR 11 、-(C1-C6 alkyl) x -C(═O)R 11 、-O-(C1-C6 alkyl) x -C(═O)R 11 、-(C1-C6 alkyl) x -C(═O)OR 11 、-C(═O)-R 11 、-C(═O)OR 11 、-(C1-C6 alkyl) x -C(═O)NR 11 R 12 、-NR 11 -(C1-C6 alkyl) x -C(═O)R 11 and-NR 11 -(C1-C6 alkyl) x -C(═O)OR 11 ; n is 0 to 3; X 1 Selected by CR 6 and N; X 2 Selected by CHR 6 NR 7 , O and S; X 3 Selected from the group consisting of C, CH and N; Each R 6 independently selected from the group consisting of: H, C1-C6, alkyl, C2-C6, alkenyl, C2-C6, alkynyl, C3-C7, cycloalkyl, halo, -CN, C1-C3, perfluoroalkyl, -OR 11 、-O-(C1-C6、、alkyl)-OR 11 、-(C1-C6, alkyl)-OR 11 、-NR 11 R 12 、-O-(C1-C6, alkyl)-NR 11 R 12 、-(C1-C6, alkyl)-NR 11 R 12 、-NR 11 -(C1-C6, alkyl)-NR 11 R 12 、-NR 11 -(C1-C6, alkyl)-OR 11 、-(C1-C6, alkyl) x -C(═O)R 11 、-O-(C1-C6, alkyl) x -C(═O)R 11 、-(C1-C6, alkyl) x -C(═O)OR 11 、-C(═O)-R 11 、-C(═O)OR 11 、-(C1-C6, alkyl) x -C(═O)NR 11 R 12 、-NR 11 -(C1-C6, alkyl) x -C(═O)R 11 and-NR 11 -(C1-C6, alkyl) x -C(═O)OR 11 ; Each R 7 independently selected from the group consisting of: H, C1-C6 alkyl, C2-C6 alkenyl, C2-C6 alkynyl, C3-C7 cycloalkyl, -(C1-C6 alkyl)-OR 11 、-(C1-C6 alkyl)-NR 11 R 12 、-(C1-C6 alkyl) x -C(═O)R 11 、-(C1-C6 alkyl) x -C(═O)OR 11 and -(C1-C6 alkyl) x -C(═O)NR 11 R 12 ; Each x is independently selected from 0 and 1; and Each R 11 and R 12 Independently selected from the group consisting of H and C1-C6 alkyl; Or alternatively, when R 11 and R 12 When both are attached to the same nitrogen, they together form a 4- to 7-membered heterocyclic ring having 0 to 2 additional ring heteroatoms selected from the group consisting of N, O and S, and the heterocyclic ring is unsubstituted or substituted with 1 to 3 substituents selected from the group consisting of halo, C1-C6 alkyl, C2-C6 alkenyl, C2-C6 alkynyl, C3-C7 cycloalkyl, -CN, -NH2, C1-C3 perfluoroalkyl, -OH, -O-(C1-C6 alkyl) and -(C1-C6 alkyl)-OH.

2. The compound according to claim 1, which has formula II: or a pharmaceutically acceptable salt thereof, wherein: R 1 Selected from the group consisting of: H, C1-C6 alkyl, C2-C6 alkenyl, C2-C6 alkynyl, C3-C7 cycloalkyl, halo, -CN, C1-C3 perfluoroalkyl, -OR 11 、-O-(C1-C6 alkyl)-OR 11 、-(C1-C6 alkyl)-OR 11 、-NR 11 R 12 、-O-(C1-C6 alkyl)-NR 11 R 12 、-(C1-C6 alkyl)-NR 11 R 12 、-NR 11 -(C1-C6 alkyl)-NR 11 R 12 、-NR 11 -(C1-C6 alkyl)-OR 11 、-(C1-C6 alkyl) x -C(═O)R 11 、-O-(C1-C6 alkyl) x -C(═O)R 11 、-(C1-C6 alkyl) x -C(═O)OR 11 、-C(═O)-R 11 、-C(═O)OR 11 、-(C1-C6 alkyl) x -C(═O)NR 11 R 12 、-NR 11 -(C1-C6 alkyl) x -C(═O)R 11 and NR 11 -(C1-C6 alkyl) x -C(═O)OR 11 ; R 2 Selected from the group consisting of: H, C1-C6 alkyl, C2-C6 alkenyl, C2-C6 alkynyl, C3-C7 cycloalkyl, halo, -CN, C1-C3 perfluoroalkyl, -OR 11 、-O-(C1-C6 alkyl)-OR 11 、-(C1-C6 alkyl)-OR 11 、-NR 11 R 12 、-O-(C1-C6 alkyl)-NR 11 R 12 、-(C1-C6 alkyl)-NR 11 R 12 、-NR 11 -(C1-C6 alkyl)-NR 11 R 12 、-NR 11 -(C1-C6 alkyl)-OR 11 、-(C1-C6 alkyl) x -C(═O)R 11 、-O-(C1-C6 alkyl) x -C(═O)R 11 、-(C1-C6 alkyl) x -C(═O)OR 11 、-C(═O)-R 11 、-C(═O)OR 11 、-(C1-C6 alkyl) x -C(═O)NR 11 R 12 、-NR 11 -(C1-C6 alkyl) x -C(═O)R 11 and-NR 11 -(C1-C6 alkyl) x -C(═O)OR 11 ; Alternatively, R 1 and R 2 The fused ring may contain 0 to 2 ring heteroatoms selected from the group consisting of N, O and S, and the fused ring may be unsubstituted or substituted with 1 to 3 substituents selected from the group consisting of: C1-C6 alkyl, halo, -CN, -OH, oxo, -O-(C1-C6 alkyl), -O-(C1-C6 alkyl)-OH, -O-(C1-C6 alkyl)-O-(C1-C6 alkyl), -NR 11 R 12 、-O-(C1-C6 alkyl)-NR 11 R 12 、C1-C3 perfluoroalkyl、-NR 11 -(C1-C6 alkyl)NR 11 R 12 and-NR 11 -(C1-C6 alkyl)-OR 11 ; R 3 and R 4 Each is independently selected from the group consisting of: H, C1-C6 alkyl, C2-C6 alkenyl, C2-C6 alkynyl, C3-C7 cycloalkyl, 3 to 10 membered heterocyclyl, C6-C 10 Aryl, 5- to 14-membered heteroaryl, C 6-12 Aralkyl, -(C1-C6 alkyl)-OR 11 、-(C1-C6 alkyl)-NR 11 R 12 、-(C1-C6 alkyl) x -C(═O)R 11 、-(C1-C6 alkyl) x -C(═O)OR 11 and -(C1-C6 alkyl) x -C(═O)NR 11 R 12 ; Alternatively, R 3 and R 4 together with the nitrogen to which they are attached, provide (i) a 4- to 6-membered heterocyclic ring having 0 to 2 additional ring heteroatoms selected from N, O, and S, or (ii) a 5- to 10-membered heterobicyclic ring system having 0 to 3 additional ring heteroatoms selected from N, O, and S; wherein the heterocyclic ring or the heterobicyclic ring system is unsubstituted or substituted with 1 to 4 substituents selected from the group consisting of halo, C1-C6 alkyl, C2-C6 alkenyl, C2-C6 alkynyl, C3-C7 cycloalkyl, halo, -CN, C1-C3 perfluoroalkyl, -OR 11 , oxo, -O-(C1-C6 alkyl)-OR 11 、-(C1-C6 alkyl)-OR 11 、-NR 11 R 12 、-O-(C1-C6 alkyl)-NR 11 R 12 、-(C1-C6 alkyl)-NR 11 R 12 、-NR 11 -(C1-C6 alkyl)-NR 11 R 12 、-NR 11 -(C1-C6 alkyl)-OR 11 、-(C1-C6 alkyl) x -C(═O)R 11 、-O-(C1-C6 alkyl) x -C(═O)R 11 、-(C1-C6 alkyl) x -C(═O)OR 11 、-C(═O)-R 11 、-C(═O)OR 11 、-(C1-C6 alkyl) x -C(═O)NR 11 R 12 、-NR 11 -(C1-C6 alkyl) x -C(═O)R 11 and-NR 11 -(C1-C6 alkyl) x -C(═O)OR 11 ; Dashed lines represent optional double bonds; Each R 5 independently selected from the group consisting of: H, C1-C6 alkyl, C2-C6 alkenyl, C2-C6 alkynyl, C3-C7 cycloalkyl, halo, -CN, C1-C3 perfluoroalkyl, oxo, -OR 11 、-O-(C1-C6 alkyl)-OR 11 、-(C1-C6 alkyl)-OR 11 、-NR 11 R 12 、-O-(C1-C6 alkyl)-NR 11 R 12 、-(C1-C6 alkyl)-NR 11 R 12 、-NR 11 -(C1-C6 alkyl)-NR 11 R 12 、-NR 11 -(C1-C6 alkyl)-OR 11 、-(C1-C6 alkyl) x -C(═O)R 11 、-O-(C1-C6 alkyl) x -C(═O)R 11 、-(C1-C6 alkyl) x -C(═O)OR 11 、-C(═O)-R 11 、-C(═O)OR 11 、-(C1-C6 alkyl) x -C(═O)NR 11 R 12 、-NR 11 -(C1-C6 alkyl) x -C(═O)R 11 and-NR 11 -(C1-C6 alkyl) x -C(═O)OR 11 ; n is 0 to 3; X 1 Selected by CR 6 and N; X 2 Selected by CHR 6 NR 7 , O and S; X 3 Selected from the group consisting of C, CH and N; Each R 6 independently selected from the group consisting of: H, C1-C6, alkyl, C2-C6, alkenyl, C2-C6, alkynyl, C3-C7, cycloalkyl, halo, -CN, C1-C3, perfluoroalkyl, -OR 11 、-O-(C1-C6, alkyl)-OR 11 、-(C1-C6, alkyl)-OR 11 、-NR 11 R 12 、-O-(C1-C6, alkyl)-NR 11 R 12 、-(C1-C6, alkyl)-NR 11 R 12 、-NR 11 -(C1-C6, alkyl)-NR 11 R 12 、-NR 11 -(C1-C6, alkyl)-OR 11 、-(C1-C6, alkyl) x -C(═O)R 11 、-O-(C1-C6, alkyl) x -C(═O)R 11 、-(C1-C6, alkyl) x -C(═O)OR 11 、-C(═O)-R 11 、-C(═O)OR 11 、-(C1-C6, alkyl) x -C(═O)NR 11 R 12 、-NR 11 -(C1-C6, alkyl) x -C(═O)R 11 and-NR 11 -(C1-C6, alkyl) x -C(═O)OR 11 ; Each R 7 independently selected from the group consisting of: H, C1-C6 alkyl, C2-C6 alkenyl, C2-C6 alkynyl, C3-C7 cycloalkyl, -(C1-C6 alkyl)-OR 11 、-(C1-C6 alkyl)-NR 11 R 12 、-(C1-C6 alkyl) x -C(═O)R 11 、-(C1-C6 alkyl) x -C(═O)OR 11 and -(C1-C6 alkyl) x -C(═O)NR 11 R 12 ; Each x is independently selected from 0 and 1; and Each R 11 and R 12 Independently selected from the group consisting of H and C1-C6 alkyl; Or alternatively, when R 11 and R 12 When both are attached to the same nitrogen, they together form a 4- to 7-membered heterocyclic ring having 0 to 2 additional ring heteroatoms selected from the group consisting of N, O and S, and the heterocyclic ring is unsubstituted or substituted with 1 to 3 substituents selected from the group consisting of halo, C1-C6 alkyl, C2-C6 alkenyl, C2-C6 alkynyl, C3-C7 cycloalkyl, -CN, -NH2, C1-C3 perfluoroalkyl, -OH, -O-(C1-C6 alkyl) and -(C1-C6 alkyl)-OH.

3. The compound according to claim 1, which has the formula IIa: or a pharmaceutically acceptable salt thereof, wherein: R 3 and R 4 Each is independently selected from the group consisting of: H, C1-C6 alkyl, C2-C6 alkenyl, C2-C6 alkynyl, C3-C7 cycloalkyl, 3 to 10 membered heterocyclyl, C6-C 10 Aryl, 5- to 14-membered heteroaryl, C 6-12 Aralkyl, -(C1-C6 alkyl)-OR 11 、-(C1-C6 alkyl)-NR 11 R 12 、-(C1-C6 alkyl) x -C(═O)R 11 、-(C1-C6 alkyl) x -C(═O)OR 11 and -(C1-C6 alkyl) x -C(═O)NR 11 R 12 ; Alternatively, R 3 and R 4 together with the nitrogen to which they are attached, provide (i) a 4- to 6-membered heterocyclic ring having 0 to 2 additional ring heteroatoms selected from N, O, and S, or (ii) a 5- to 10-membered heterobicyclic ring system having 0 to 3 additional ring heteroatoms selected from N, O, and S; wherein the heterocyclic ring or the heterobicyclic ring system is unsubstituted or substituted with 1 to 4 substituents selected from the group consisting of halo, C1-C6 alkyl, C2-C6 alkenyl, C2-C6 alkynyl, C3-C7 cycloalkyl, halo, -CN, C1-C3 perfluoroalkyl, -OR 11 , oxo, -O-(C1-C6 alkyl)-OR 11 、-(C1-C6 alkyl)-OR 11 、-NR 11 R 12 、-O-(C1-C6 alkyl)-NR 11 R 12 、-(C1-C6 alkyl)-NR 11 R 12 、-NR 11 -(C1-C6 alkyl)-NR 11 R 12 、-NR 11 -(C1-C6 alkyl)-OR 11 、-(C1-C6 alkyl) x -C(═O)R 11 、-O-(C1-C6 alkyl) x -C(═O)R 11 、-(C1-C6 alkyl) x -C(═O)OR 11 、-C(═O)-R 11 、-C(═O)OR 11 、-(C1-C6 alkyl) x -C(═O)NR 11 R 12 、-NR 11 -(C1-C6 alkyl) x -C(═O)R 11 and-NR 11 -(C1-C6 alkyl) x -C(═O)OR 11 ; Dashed lines represent optional double bonds; X 1 Selected by CR 6 and N; X 2 Selected by CHR 6 NR 7 , O and S; X 3 Selected from the group consisting of C, CH and N; Each R 6 independently selected from the group consisting of: H, C1-C6, alkyl, C2-C6, alkenyl, C2-C6, alkynyl, C3-C7, cycloalkyl, halo, -CN, C1-C3, perfluoroalkyl, -OR 11 、-O-(C1-C6, alkyl)-OR 11 、-(C1-C6, alkyl)-OR 11 、-NR 11 R 12 、-O-(C1-C6, alkyl)-NR 11 R 12 、-(C1-C6, alkyl)-NR 11 R 12 、-NR 11 -(C1-C6, alkyl)-NR 11 R 12 、-NR 11 -(C1-C6, alkyl)-OR 11 、-(C1-C6, alkyl) x -C(═O)R 11 、-O-(C1-C6, alkyl) x -C(═O)R 11 、-(C1-C6, alkyl) x -C(═O)OR 11 、-C(═O)-R 11 、-C(═O)OR 11 、-(C1-C6, alkyl) x -C(═O)NR 11 R 12 、-NR 11 -(C1-C6, alkyl) x -C(═O)R 11 and-NR 11 -(C1-C6, alkyl) x -C(═O)OR 11 ; Each R 7 independently selected from the group consisting of: H, C1-C6 alkyl, C2-C6 alkenyl, C2-C6 alkynyl, C3-C7 cycloalkyl, -(C1-C6 alkyl)-OR 11 、-(C1-C6 alkyl)-NR 11 R 12 、-(C1-C6 alkyl) x -C(═O)R 11 、-(C1-C6 alkyl) x -C(═O)OR 11 and -(C1-C6 alkyl) x -C(═O)NR 11 R 12 ; Each x is independently selected from 0 and 1; and Each R 11 and R 12 Independently selected from the group consisting of H and C1-C6 alkyl; Or alternatively, when R 11 and R 12 When both are attached to the same nitrogen, they together form a 4- to 7-membered heterocyclic ring having 0 to 2 additional ring heteroatoms selected from the group consisting of N, O and S, and the heterocyclic ring is unsubstituted or substituted with 1 to 3 substituents selected from the group consisting of halo, C1-C6 alkyl, C2-C6 alkenyl, C2-C6 alkynyl, C3-C7 cycloalkyl, -CN, -NH2, C1-C3 perfluoroalkyl, -OH, -O-(C1-C6 alkyl) and -(C1-C6 alkyl)-OH.

4. The compound according to claim 1, which has the formula III: or a pharmaceutically acceptable salt thereof, wherein: R 1 Selected from the group consisting of: H, C1-C6 alkyl, C2-C6 alkenyl, C2-C6 alkynyl, C3-C7 cycloalkyl, halo, -CN, C1-C3 perfluoroalkyl, -OR 11 、-O-(C1-C6 alkyl)-OR 11 、-(C1-C6 alkyl)-OR 11 、-NR 11 R 12 、-O-(C1-C6 alkyl)-NR 11 R 12 、-(C1-C6 alkyl)-NR 11 R 12 、-NR 11 -(C1-C6 alkyl)-NR 11 R 12 、-NR 11 -(C1-C6 alkyl)-OR 11 、-(C1-C6 alkyl) x -C(═O)R 11 、-O-(C1-C6 alkyl) x -C(═O)R 11 、-(C1-C6 alkyl) x -C(═O)OR 11 、-C(═O)-R 11 、-C(═O)OR 11 、-(C1-C6 alkyl) x -C(═O)NR 11 R 12 、-NR 11 -(C1-C6 alkyl) x -C(═O)R 11 and NR 11 -(C1-C6 alkyl) x -C(═O)OR 11 ; R 2 Selected from the group consisting of: H, C1-C6 alkyl, C2-C6 alkenyl, C2-C6 alkynyl, C3-C7 cycloalkyl, halo, -CN, C1-C3 perfluoroalkyl, -OR 11 、-O-(C1-C6 alkyl)-OR 11 、-(C1-C6 alkyl)-OR 11 、-NR 11 R 12 、-O-(C1-C6 alkyl)-NR 11 R 12 、-(C1-C6 alkyl)-NR 11 R 12 、-NR 11 -(C1-C6 alkyl)-NR 11 R 12 、-NR 11 -(C1-C6 alkyl)-OR 11 、-(C1-C6 alkyl) x -C(═O)R 11 、-O-(C1-C6 alkyl) x -C(═O)R 11 、-(C1-C6 alkyl) x -C(═O)OR 11 、-C(═O)-R 11 、-C(═O)OR 11 、-(C1-C6 alkyl) x -C(═O)NR 11 R 12 、-NR 11 -(C1-C6 alkyl) x -C(═O)R 11 and-NR 11 -(C1-C6 alkyl) x -C(═O)OR 11 ; Alternatively, R 1 and R 2 The fused ring may contain 0 to 2 ring heteroatoms selected from the group consisting of N, O and S, and the fused ring may be unsubstituted or substituted with 1 to 3 substituents selected from the group consisting of: C1-C6 alkyl, halo, -CN, -OH, oxo, -O-(C1-C6 alkyl), -O-(C1-C6 alkyl)-OH, -O-(C1-C6 alkyl)-O-(C1-C6 alkyl), -NR 11 R 12 、-O-(C1-C6 alkyl)-NR 11 R 12 、C1-C3 perfluoroalkyl、-NR 11 -(C1-C6 alkyl)NR 11 R 12 and-NR 11 -(C1-C6 alkyl)-OR 11 ; R 3 and R 4 Each is independently selected from the group consisting of: H, C1-C6 alkyl, C2-C6 alkenyl, C2-C6 alkynyl, C3-C7 cycloalkyl, 3 to 10 membered heterocyclyl, C6-C 10 Aryl, 5- to 14-membered heteroaryl, C 6-12 Aralkyl, -(C1-C6 alkyl)-OR 11 、-(C1-C6 alkyl)-NR 11 R 12 、-(C1-C6 alkyl) x -C(═O)R 11 、-(C1-C6 alkyl) x -C(═O)OR 11 and -(C1-C6 alkyl) x -C(═O)NR 11 R 12 ; Alternatively, R 3 and R 4 together with the nitrogen to which they are attached, provide (i) a 4- to 6-membered heterocyclic ring having 0 to 2 additional ring heteroatoms selected from N, O, and S, or (ii) a 5- to 10-membered heterobicyclic ring system having 0 to 3 additional ring heteroatoms selected from N, O, and S; wherein the heterocyclic ring or the heterobicyclic ring system is unsubstituted or substituted with 1 to 4 substituents selected from the group consisting of halo, C1-C6 alkyl, C2-C6 alkenyl, C2-C6 alkynyl, C3-C7 cycloalkyl, halo, -CN, C1-C3 perfluoroalkyl, -OR 11 , oxo, -O-(C1-C6 alkyl)-OR 11 、-(C1-C6 alkyl)-OR 11 、-NR 11 R 12 、-O-(C1-C6 alkyl)-NR 11 R 12 、-(C1-C6 alkyl)-NR 11 R 12 、-NR 11 -(C1-C6 alkyl)-NR 11 R 12 、-NR 11 -(C1-C6 alkyl)-OR 11 、-(C1-C6 alkyl) x -C(═O)R 11 、-O-(C1-C6 alkyl) x -C(═O)R 11 、-(C1-C6 alkyl) x -C(═O)OR 11 、-C(═O)-R 11 、-C(═O)OR 11 、-(C1-C6 alkyl) x -C(═O)NR 11 R 12 、-NR 11 -(C1-C6 alkyl) x -C(═O)R 11 and-NR 11 -(C1-C6 alkyl) x -C(═O)OR 11 ; Dashed lines represent optional double bonds; Each R 5 independently selected from the group consisting of: H, C1-C6 alkyl, C2-C6 alkenyl, C2-C6 alkynyl, C3-C7 cycloalkyl, halo, -CN, C1-C3 perfluoroalkyl, oxo, -OR 11 、-O-(C1-C6 alkyl)-OR 11 、-(C1-C6 alkyl)-OR 11 、-NR 11 R 12 、-O-(C1-C6 alkyl)-NR 11 R 12 、-(C1-C6 alkyl)-NR 11 R 12 、-NR 11 -(C1-C6 alkyl)-NR 11 R 12 、-NR 11 -(C1-C6 alkyl)-OR 11 、-(C1-C6 alkyl) x -C(═O)R 11 、-O-(C1-C6 alkyl) x -C(═O)R 11 、-(C1-C6 alkyl) x -C(═O)OR 11 、-C(═O)-R 11 、-C(═O)OR 11 、-(C1-C6 alkyl) x -C(═O)NR 11 R 12 、-NR 11 -(C1-C6 alkyl) x -C(═O)R 11 and-NR 11 -(C1-C6 alkyl) x -C(═O)OR 11 ; n is 0 to 3; X 1 Selected by CR 6 and N; X 2 Selected by CHR 6 NR 7 , O and S; X 3 Selected from the group consisting of C, CH and N; Each R 6 independently selected from the group consisting of: H, C1-C6, alkyl, C2-C6, alkenyl, C2-C6, alkynyl, C3-C7, cycloalkyl, halo, -CN, C1-C3, perfluoroalkyl, -OR 11 、-O-(C1-C6, alkyl)-OR 11 、-(C1-C6, alkyl)-OR 11 、-NR 11 R 12 、-O-(C1-C6, alkyl)-NR 11 R 12 、-(C1-C6, alkyl)-NR 11 R 12 、-NR 11 -(C1-C6, alkyl)-NR 11 R 12 、-NR 11 -(C1-C6, alkyl)-OR 11 、-(C1-C6, alkyl) x -C(═O)R 11 、-O-(C1-C6, alkyl) x -C(═O)R 11 、-(C1-C6, alkyl) x -C(═O)OR 11 、-C(═O)-R 11 、-C(═O)OR 11 、-(C1-C6, alkyl) x -C(═O)NR 11 R 12 、-NR 11 -(C1-C6, alkyl) x -C(═O)R 11 and-NR 11 -(C1-C6, alkyl) x -C(═O)OR 11 ; Each R 7 independently selected from the group consisting of: H, C1-C6 alkyl, C2-C6 alkenyl, C2-C6 alkynyl, C3-C7 cycloalkyl, -(C1-C6 alkyl)-OR 11 、-(C1-C6 alkyl)-NR 11 R 12 、-(C1-C6 alkyl) x -C(═O)R 11 、-(C1-C6 alkyl) x -C(═O)OR 11 and -(C1-C6 alkyl) x -C(═O)NR 11 R 12 ; Each x is independently selected from 0 and 1; and Each R 11 and R 12 Independently selected from the group consisting of H and C1-C6 alkyl; Or alternatively, when R 11 and R 12 When both are attached to the same nitrogen, they together form a 4- to 7-membered heterocyclic ring having 0 to 2 additional ring heteroatoms selected from the group consisting of N, O and S, and the heterocyclic ring is unsubstituted or substituted with 1 to 3 substituents selected from the group consisting of halo, C1-C6 alkyl, C2-C6 alkenyl, C2-C6 alkynyl, C3-C7 cycloalkyl, -CN, -NH2, C1-C3 perfluoroalkyl, -OH, -O-(C1-C6 alkyl) and -(C1-C6 alkyl)-OH.

5. The compound according to claim 1, which has the formula IIIa: or a pharmaceutically acceptable salt thereof, wherein: R 3 and R 4 Each is independently selected from the group consisting of: H, C1-C6 alkyl, C2-C6 alkenyl, C2-C6 alkynyl, C3-C7 cycloalkyl, 3 to 10 membered heterocyclyl, C6-C 10 Aryl, 5- to 14-membered heteroaryl, C 6-12 Aralkyl, -(C1-C6 alkyl)-OR 11 、-(C1-C6 alkyl)-NR 11 R 12 、-(C1-C6 alkyl) x -C(═O)R 11 、-(C1-C6 alkyl) x -C(═O)OR 11 and -(C1-C6 alkyl) x -C(═O)NR 11 R 12 ; Alternatively, R 3 and R 4 together with the nitrogen to which they are attached, provide (i) a 4- to 6-membered heterocyclic ring having 0 to 2 additional ring heteroatoms selected from N, O, and S, or (ii) a 5- to 10-membered heterobicyclic ring system having 0 to 3 additional ring heteroatoms selected from N, O, and S; wherein the heterocyclic ring or the heterobicyclic ring system is unsubstituted or substituted with 1 to 4 substituents selected from the group consisting of halo, C1-C6 alkyl, C2-C6 alkenyl, C2-C6 alkynyl, C3-C7 cycloalkyl, halo, -CN, C1-C3 perfluoroalkyl, -OR 11 , oxo, -O-(C1-C6 alkyl)-OR 11 、-(C1-C6 alkyl)-OR 11 、-NR 11 R 12 、-O-(C1-C6 alkyl)-NR 11 R 12 、-(C1-C6 alkyl)-NR 11 R 12 、-NR 11 -(C1-C6 alkyl)-NR 11 R 12 、-NR 11 -(C1-C6 alkyl)-OR 11 、-(C1-C6 alkyl) x -C(═O)R 11 、-O-(C1-C6 alkyl) x -C(═O)R 11 、-(C1-C6 alkyl) x -C(═O)OR 11 、-C(═O)-R 11 、-C(═O)OR 11 、-(C1-C6 alkyl) x -C(═O)NR 11 R 12 、-NR 11 -(C1-C6 alkyl) x -C(═O)R 11 and-NR 11 -(C1-C6 alkyl) x -C(═O)OR 11 ; Dashed lines represent optional double bonds; X 1 Selected by CR 6 and N; X 2 Selected by CHR 6 NR 7 , O and S; X 3 Selected from the group consisting of C, CH and N; Each R 6 independently selected from the group consisting of: H, C1-C6, alkyl, C2-C6, alkenyl, C2-C6, alkynyl, C3-C7, cycloalkyl, halo, -CN, C1-C3, perfluoroalkyl, -OR 11 、-O-(C1-C6, alkyl)-OR 11 、-(C1-C6, alkyl)-OR 11 、-NR 11 R 12 、-O-(C1-C6, alkyl)-NR 11 R 12 、-(C1-C6, alkyl)-NR 11 R 12 、-NR 11 -(C1-C6, alkyl)-NR 11 R 12 、-NR 11 -(C1-C6, alkyl)-OR 11 、-(C1-C6, alkyl) x -C(═O)R 11 、-O-(C1-C6, alkyl) x -C(═O)R 11 、-(C1-C6, alkyl) x -C(═O)OR 11 、-C(═O)-R 11 、-C(═O)OR 11 、-(C1-C6, alkyl) x -C(═O)NR 11 R 12 、-NR 11 -(C1-C6, alkyl) x -C(═O)R 11 and-NR 11 -(C1-C6, alkyl) x -C(═O)OR 11 ; Each R 7 independently selected from the group consisting of: H, C1-C6 alkyl, C2-C6 alkenyl, C2-C6 alkynyl, C3-C7 cycloalkyl, -(C1-C6 alkyl)-OR 11 、-(C1-C6 alkyl)-NR 11 R 12 、-(C1-C6 alkyl) x -C(═O)R 11 、-(C1-C6 alkyl) x -C(═O)OR 11 and -(C1-C6 alkyl) x -C(═O)NR 11 R 12 ; Each x is independently selected from 0 and 1; and Each R 11 and R 12 Independently selected from the group consisting of H and C1-C6 alkyl; Or alternatively, when R 11 and R 12 When both are attached to the same nitrogen, they together form a 4- to 7-membered heterocyclic ring having 0 to 2 additional ring heteroatoms selected from the group consisting of N, O and S, and the heterocyclic ring is unsubstituted or substituted with 1 to 3 substituents selected from the group consisting of halo, C1-C6 alkyl, C2-C6 alkenyl, C2-C6 alkynyl, C3-C7 cycloalkyl, -CN, -NH2, C1-C3 perfluoroalkyl, -OH, -O-(C1-C6 alkyl) and -(C1-C6 alkyl)-OH.

6. The compound according to claim 1, which is 2-[2-(3,3-difluoroazetidine-1-carbonyl)-1-methyl-1H-indol-6-yl]-N-[4-(1H-imidazol-5-yl)phenyl]pyrimidin-4-amine having the following formula: or a pharmaceutically acceptable salt thereof.

7. The compound according to claim 1, which is 2-[2-(3,3-difluoroazetidine-1-carbonyl)-1-methyl-1H-indol-6-yl]-N-[4-(1H-1,2,3-triazol-4-yl)phenyl]pyrimidin-4-amine having the following formula: or a pharmaceutically acceptable salt thereof.

8. A method for treating a disease or condition mediated by ROCK2, wherein the method comprises administering to a subject in need thereof an effective amount of a compound according to any one of claims 1 to 7 or a pharmaceutically acceptable salt thereof.

9. The method of claim 8, wherein the disease or disorder is selected from the group consisting of fibrotic diseases, inflammatory diseases, autoimmune diseases, cardiovascular disorders, central nervous system disorders, neoplastic diseases, metabolic syndrome, eye diseases, kidney diseases, lung diseases, muscular dystrophy, sickle cell disease, and viral diseases.

10. The method of claim 9, wherein the disease or condition is selected from the group consisting of a fibrotic disease, an inflammatory disease, and an autoimmune disease.

11. The method of claim 10, wherein the autoimmune disease is selected from the group consisting of rheumatoid arthritis, systemic lupus erythematosus (SLE; lupus), psoriasis, psoriatic arthritis, multiple sclerosis, Crohn's disease, ulcerative colitis, atopic dermatitis, eczema, or graft-versus-host disease (GVHD; acute and chronic), idiopathic pulmonary fibrosis, and scleroderma.

12. The method of claim 9, wherein the disease or disorder is selected from the group consisting of a cardiovascular disorder, a central nervous system disorder, a neoplastic disease, or a metabolic syndrome.

13. The method of claim 10, wherein the inflammatory disorder is selected from the group consisting of cardiovascular inflammation, lung inflammation, kidney inflammation, arteriosclerosis, and sepsis.

14. The method of claim 10, wherein the fibrotic disorder is selected from the group consisting of idiopathic pulmonary fibrosis, renal fibrosis, kidney fibrosis, ocular fibrosis, cardiac fibrosis, NASH, scleroderma, systemic sclerosis, and liver cirrhosis.

15. The method of claim 12, wherein the neoplastic disease is selected from the group consisting of ovarian cancer, breast cancer, and pancreatic cancer.

16. The method of claim 12, wherein the cardiovascular disease is selected from the group consisting of hypertension, cardiomyopathy, cardiac remodeling, atherosclerosis, restenosis, cardiac hypertrophy, cerebral ischemia, cerebral vasospasm, and erectile dysfunction.

17. The method of claim 9, wherein the pulmonary disease is selected from the group consisting of idiopathic pulmonary fibrosis, chronic obstructive pulmonary disease, and asthma.

18. The method of claim 9, wherein the central nervous system disorder is selected from the group consisting of neuronal degeneration or spinal cord injury, traumatic brain injury, cerebral cavernous malformation, Huntington's disease, Parkinson's disease, Alzheimer's disease, amyotrophic lateral sclerosis (ALS), and multiple sclerosis.

19. The method of claim 9, wherein the kidney disease is selected from the group consisting of polycystic kidney disease, renal fibrosis, and diabetic kidney disease.

20. The method of claim 9, wherein the metabolic disease is selected from the group consisting of insulin resistance, hyperinsulinemia, type 2 diabetes, obesity, metabolic syndrome, and glucose intolerance.

21. The method of claim 9, wherein the ocular disease is selected from the group consisting of ocular hypertension, age-related macular degeneration (AMD; wet and dry), choroidal neovascularization (CNV), choroidal tumors, diabetic macular edema (DME), iris neovascularization, uveitis, glaucoma, primary open-angle glaucoma, acute angle-closure glaucoma, pigmentary glaucoma, congenital glaucoma, normal-tension glaucoma, secondary glaucoma, neovascular glaucoma, geographic atrophy, and retinitis of prematurity (ROP).

22. The method of claim 9, wherein the disease is Duchenne muscular dystrophy.

23. The method of claim 9, wherein the viral infection is a coronavirus infection, such as SARS-CoV-1, SARS-CoV-2, and MERS-CoV.

Images