Prevention Or Treatment Of Diseases And Disorders Associated With Tissue Damage Patent Application (2024)

U.S. patent application number 17/632500 was filed with the patent office on 2022-09-08 for prevention or treatment of diseases and disorders associated with tissue damage.This patent application is currently assigned to Anhui Newstar Pharmaceutical Development Co., Ltd.. The applicant listed for this patent is Xiaoxiang Li. Invention is credited to Xiaoxiang Li.

PREVENTION OR TREATMENT OF DISEASES AND DISORDERS ASSOCIATED WITHTISSUE DAMAGE

Abstract

The present invention provides a method of preventing ortreating diseases and disorders associated with tissue damagesusing an MST1/2 protein kinase inhibitor. The MST1/2 protein kinaseinhibitor may be administered in its prodrug or salt forms. Abioavailability enhancing agent or an absorption enhancing agentmay be used in conjunction with the MST1/2 protein kinaseinhibitor.

Related U.S. Patent Documents

Claims

1. A method of preventing or treating a disease or disorderassociated with tissue damage using a MST1/2 protein kinaseinhibitor, said method comprising a step of: administering theMST1/2 protein kinase inhibitor to a subject having the disease ordisorder, wherein the MST1/2 protein kinase inhibitor isadministered at a dose of from about 0.1 mg/kg to about 100 mg/kgbased on the bodyweight of the subject and at a frequency of oncein a period of from 6 hours to 20 days, and the MST1/2 proteinkinase inhibitor has formulas I, II, III, IV: ##STR00114## wherein:n1 is selected from 0, 1, 2, 3 or 4; R.sub.11 is selected from: 1)C1-C6 alkyl, optionally substituted with halogen, amino, nitro,cyano; C1-C6 alkyl containing oxygen; C3-C7 cycloalkyl, which isoptionally substituted with halogen, amino, nitro, cyano; C6-C10aryl, optionally substituted by halogen, nitro, amino, hydroxy,cyano; C3-C6 alkenyl; 2) 2-N, N-dimethylaminoethyl, 2-hydroxyethyl,2-N, N-diethylaminoethyl, 2-N, N-diisopropylamino ethyl,2-morpholinyl ethyl, 2-thiomorpholinyl ethyl,2-(4-N-piperazinyl-methyl) ethyl, 3-N, N-dimethylaminopropyl, 3-N,N-diethylaminopropyl, 3-N, N-diisopropyl-aminopropyl, 3-morpholinylpropyl, 3-thiomorpholinyl propyl, 3-(4-N-methylpiperidinyl) propyl,4-N, N-dimethylamino-cyclohexyl, 4-N, N-diethylamino cyclohexyl,N-methyl-4-piperidinyl, N-ethyl-4-piperidinyl,N-isopropyl-4-piperidinyl, 1,3-dimethyl-5-pyrazolyl,1-methyl-4-pyrazolyl, 3-methyl-5-isoxazolinyl,1-(N-methyl-4-piperidinyl)-4-pyrazolyl, 1-(N-tert-butoxylformyl-4-piperidinyl)-4-pyrazolyl; 3) ##STR00115## wherein Z.sub.1,Z.sub.2, Z.sub.3, Z.sub.4, Z.sub.5 are each independently selectedfrom: (1) hydrogen, halogen, nitro, amino, hydroxy, cyano, (2)C1-C6 alkyl, C1-C6 alkoxy, C1-C6 alkyl containing oxygen, C1-C6alkyl containing fluorine, C1-C6 alkoxy containing fluorine,4-piperidinyl, N-methyl yl-4-piperidinyl, (3) N, N-dimethylamino,N, N-diethylamino, N, N-diisopropylamino, 2-N,N-dimethylaminoethylamino, 2-morpholino ethylamino, 2-ethylaminothiomorpholinyl, 2-(4-N-methylpiperazinyl) ethylamino, 3-N,N-dimethyl-aminopropyl amino, 3-N, N-diethylaminopropyl-amino, 3-N,N-diisopropylamino propylamino, 3-morpholin-propylamino,3-thiomorpholinyl propylamino, 3-(4 N-methylpiperazinyl)propylamino, N-methylpiperidinyl-4-amino,N-ethylpiperidinyl-4-amino, N-isopropyl-piperidinyl-4-amino, (4)2-N, N-dimethylaminoethoxyl, 2-N, N-diethyl-aminoethoxy, 2-N,N-diisopropyl-aminoethoxyl, 2-(N-methylpiperazinyl) ethoxyl,2-(N-acetyl-piperazinyl) ethoxyl, 2-morpholino-ethoxyl,2-thiomorpholino-ethoxyl, 2-piperidinyl ethoxyl, 3-N,N-dimethylamino-propoxyl, 3-N, N-diethylamino-propoxyl, 3-N,N-diisopropylamino propoxyl, 3-(N-methylpiperazinyl) propoxyl,3-(N-acetyl-piperazinyl) propoxyl, 3-morpholinyl-propoxyl,3-thiomorpholinyl propoxyl, 3-piperidinyl-propoxyl,2-pyridyl-methoxyl, 3-pyridyl-methoxyl, 4-pyridyl methoxyl,phenylmethoxyl, monohalogen-substituted phenylmethoxyl,hom*odihalogen-substituted phenylmethoxyl,heterodihalogen-substituted phenylmethoxyl, (5) piperidinyl, 4-N,N-dimethylamino-piperidinyl, 4-N, N-diethylamino-piperidinyl, 4-N,N-diisopropylamino piperidinyl, 4-hydroxy piperidinyl, morpholinyl,3,5-dimethyl morpholinyl, thiomorpholinyl, tetrahydropyrrolyl, 3-N,N-dimethyl-tetrahydropyrrolyl, 3-N, N-diethyl-tetrahydropyrrolyl,N-methyl-piperazinyl, N-ethyl-piperazinyl, N-isopropyl-piperazinyl,N-acetyl-piperazinyl, N-tert-butoxyl formyl piperazinyl,N-methylsulfonyl-piperazinyl, N-(2-hydroxylethyl) piperazinyl,N-(2-cyanoethyl) piperazinyl, N-(3-hydroxylpropyl) piperazinyl,N-(2-N, N-dimethylethyl) piperazinyl, N-(2-N, N-diethyl-ethyl)piperazinyl, N-(3-N, N-dimethylpropyl) piperazinyl, N-(3-N,N-diethyl-propyl) piperazinyl, 2-oxo-piperazinyl,2-oxo-piperazin-4-yl, imidazolyl, 4-imidazolyl, (6)4-(N-methylpiperazinyl) piperidinyl, 4-(N-ethyl-piperazinyl)piperidinyl, 4-(N-isopropyl-piperazinyl) piperidinyl,4-(N-acetyl-piperazinyl) piperidinyl,4-(N-t-butoxyl-formyl-piperazinyl) piperidinyl,4-(N-methylsulfonyl-piperazinyl) piperidinyl,4-(N-(2-hydroxylethyl) piperazinyl) piperidinyl,4-(N-(2-cyanoethyl) piperazinyl) piperidinyl,4-(N-(3-hydroxylpropyl) piperazinyl) piperidinyl, 4-(N-(2-N,N-dimethyl-ethyl) piperazinyl) piperidinyl, 4-(N-(2-N, N-diethylethyl) piperazinyl) piperidinyl, 4-(N-(3-N, N-dimethyl-propyl)piperazinyl) piperidinyl, 4-(N-(3-N, N-diethyl-propyl) piperazinyl)piperidinyl, 4-(tetrahydropyrrolyl) piperidinyl, 4-(3-N,N-dimethyl-tetrahydropyrrolyl) piperidinyl,N--(N-methyl-4-piperidinyl) piperazinyl, N--(N-ethyl-4-piperidinyl)piperazinyl, (7) hydroxy sulfonyl, aminosulfonyl, sulfonylmethylamino, ethylamino sulfonyl group, a sulfonyl grouppropylamino, isopropylamino-sulfonyl, aminosulfonyl cyclopropyl,cyclobutyl aminosulfonyl, cyclopentyl aminosulfonyl,piperidinyl-sulfonyl, 4-hydroxyl-piperidinyl-1-sulfonyl, 4-N,N-dimethyl-piperidinyl-1-sulfonyl, 4-N,N-diethyl-piperidinyl-1-sulfonyl, pyrrolidinyl-1-sulfonyl, 3-N,N-dimethyl-pyrrolidinyl-1-sulfonyl, 3-N,N-diethyl-pyrrolidinyl-1-sulfonyl, N-methyl-piperazinyl-sulfonyl,N-ethylpiperazinyl-1-sulfonyl, N-acetyl-piperazinyl-1-sulfonyl,N-tert-butoxylformyl-piperazinyl-1-sulfonyl, N-(2-hydroxylethyl)piperazinyl-1-sulfonyl, N-(2-cyanoethyl) piperazinyl-1-sulfonyl,N-(2-N, N-dimethyl ethyl) piperazinyl-1-sulfonyl, N-(2-N,N-diethyl-ethyl) piperazinyl-1-sulfonyl, N-(3-hydroxylpropyl)piperazinyl-1-sulfonyl, N-(3-N, N-dimethylamino-propyl)piperazinyl-1-sulfonyl, N-(3-N, N-diethylamino-propyl)piperazinyl-1-sulfonyl, morpholinyl-1-sulfonyl,3,5-dimethyl-morpholinyl-1-sulfonyl, 4-(N-methyl-1-piperazinyl)piperidinyl-1-sulfonyl, 4-(N-ethyl-1-piperazinyl)piperidinyl-1-sulfonyl, 4-(N-acetyl-1-piperazinyl)piperidinyl-sulfonyl, N--(N-methyl-4-piperidinyl)piperazinyl-1-sulfonyl, (8) amino formyl, methylamino formyl,ethylamino formyl, propylamino formyl, isopropylamino formyl,cyclopropylamino formyl, cyclobutylamino formyl, cyclopentylaminoformyl, piperidinyl-1-formyl, 4-hydroxy-piperidinyl-1-formyl, 4-N,N-dimethyl-piperidinyl-1-formyl, 4-N, N-twoethylpiperidinyl-1-formyl, tetrahydropyrrolyl-1-formyl, 3-N,N-dimethyl-tetrahydropyrrolyl-1-formyl, 3-N,N-diethyl-tetrahydropyrrolyl-1-formyl,N-methyl-piperazinyl-1-formyl, N-ethyl-piperazinyl-1-formyl,N-acetyl-piperazinyl-1-formyl,N-tert-butoxyl-formyl-piperazinyl-1-formyl, N-(2-hydroxyethyl)piperazinyl-1-formyl, N-(2-cyanoethyl) piperazinyl-1-formyl,N-(2-N, N-dimethyl-ethyl) piperazinyl-1-formyl, N-(2-N,N-diethyl-ethyl) piperazinyl-1-formyl, N-(3-hydroxypropyl)piperazinyl-1-formyl, N-(3-N, N-dimethyl-propyl)piperazinyl-1-formyl, N-(3-N, N-diethyl propyl)piperazinyl-1-formyl, morpholinyl-1-formyl,3,5-dimethyl-morpholinyl-1-formyl, 4-(N-methyl-1-piperazinyl)piperidinyl-1-formyl, 4-(N-ethyl-1-piperazinyl)piperidinyl-1-formyl, 4-(N-acetyl-1-piperazinyl)piperidinyl-1-formyl, N--(N-methyl-4-piperidinyl)piperazinyl-1-formyl, (9) hydroxyl formyl, methoxyl formyl, ethoxylformyl, propoxyl formyl, isopropoxyl formyl, n-butoxyl formyl,isobutoxy formyl, t-butoxyl formyl, (10) amino formamido,methylamino formamido, ethylamino formamido, propylamino formamido,isopropylamino formamido, cyclopropylamino formamido,cyclobutylamino formamido, cyclopentylamino formamido,piperidinyl-1-formamido, 4-hydroxy-piperidinyl-1-formamido, 4-N,N-dimethyl-piperidinyl-1-formamido, 4-N,N-diethyl-piperidinyl-1-formamido, tetrahydropyrrolyl-1-formamido,3-N, N-dimethyl-tetrahydropyrrolyl-1-formamido, 3-N,N-diethyl-tetrahydropyrrolyl-1-formamido,N-methyl-piperazinyl-1-formamido, N-ethyl-piperazinyl-1-formamido,N-acetyl-piperazinyl-1-formamido, N-tert-butoxylformyl-piperazinyl-1-formamido, N-(2-hydroxyethyl)piperazinyl-1-formamido, N-(2-cyanoethyl) piperazinyl-1-formamido,N-(2-N, N-dimethyl-ethyl) piperazinyl-1-formamido, N-(2-N,N-diethyl-ethyl) piperazinyl-1-formamido, N-(3-hydroxypropyl)piperazinyl-1-formamido, N-(3-N, N-dimethyl-propyl)piperazinyl-1-formamido, N-(3-N, N-diethyl-aminopropyl)piperazinyl-1-formamido, morpholinyl-1-formamido,3,5-dimethyl-morpholinyl-1-formamido, 4-(N-methyl-1-piperazinyl)piperidinyl-1-formamido, 4-(N-ethyl-1-piperazinyl)piperidinyl-1-formamido, 4-(N-acetyl-1-piperazinyl)piperidinyl-1-formamido, N--(N-methyl-4-piperidinyl)piperazinyl-1-formamido; or (11) amino acetamido, N-tert-butoxylformyl acetamido, N-acetylamino acetamido, acrylamido,cyclopropylamido, chloroacetamido, bromoacetamido, piperidinylacetamido, 4-hydroxy piperidinyl acetamido, 4-N,N-dimethyl-piperidinyl-acetamido, 4-N, N-diethyl-piperidinylacetamido, tetrahydropyrrolyl acetamido, 3-N,N-dimethyl-tetrahydropyrrolyl acetamido, 3-N,N-diethyl-tetrahydropyrrolyl-acetamido, N-methyl-piperazinylacetamido, N-ethyl piperazinyl-acetamido, N-acetyl-piperazinylacetamido, N-tert-butoxy formyl-piperazinyl acetamido,N-(2-hydroxyethyl) piperazinyl acetamido, N-(2-cyanoethyl)piperazinyl acetamido, N-(2-N, N-dimethylethyl) piperazinylacetamido, N-(2-N, N-diethyl-ethyl) piperazinyl acetamido,N-(3-hydroxylpropyl) piperazinyl acetamido, N-(3-N,N-dimethyl-propyl) piperazinyl acetamido, N-(3-N, N-diethyl-propyl)piperazinyl acetamido, morpholinyl acetamido,3,5-dimethyl-morpholinyl-acetamido, 4-(N-methyl-1-piperazinyl)piperidinyl acetamido, 4-(N-ethyl-1-piperazinyl) piperidinylacetamido, 4-(N-acetyl-1-piperazinyl) piperidinyl acetamido,N--(N-methyl-4-piperidinyl) piperazinyl acetamido,4-(tetrahydropyrrolyl) piperidinyl acetamido; 2-methylaminoacetamido, 2-(1-methylethyl) amino acetamido;N-benzyloxy-formyl-2-methylamino-acetamido; (12) Z.sub.2 andZ.sub.3 may form a substituted or unsubstituted oxygen-containingfive- or six-membered ring; the substituents may be selected fromthe same substituents of Z.sub.1, (13) Z.sub.2 and Z.sub.3 may forma substituted or unsubstituted nitrogen-containing five- orsix-membered ring; the substituents may be selected from the samesubstituents of Z.sub.1, 4) ##STR00116## wherein Z.sub.2, Z.sub.3,Z.sub.4, Z.sub.5 are the same as the definition 3) above; 5)##STR00117## wherein Z.sub.1, Z.sub.3, Z.sub.4, Z.sub.5 are thesame as the definition 3) above; R.sub.1 is selected from: 1) ahydrogen, halo, nitro, amino, cyano; 2) C1-C6 alkyl, optionallysubstituted by halogen, nitro, amino, cyano; --O--C1-C6 alkyl,optionally substituted by halogen, nitro, amino, cyano; a C1-C6oxygen-containing alkyl; 3) methylthio, ethylthio, isopropylthio,methylsulfinyl, ethylsulfinyl, propyl sulfinyl, methylsulfonyl,ethylsulfonyl, isopropylsulfonyl, amino sulfonyl, ethylaminosulfonyl, propylamino sulfonyl, isopropylamino-sulfonyl,cyclopropylamino sulfonyl, hydroxyl formyl, methoxyl formyl,ethoxyl formyl, propoxyl formyl, isopropoxyl formyl, n-butoxylformyl, isobutoxyl formyl, t-butoxyl formyl, amino formyl,methylamino formyl, ethylamino formyl, propylamino formyl,isopropylamino formyl, cyclopropylamino formyl, cyclobutylaminoformyl, cyclopentylamino formyl, acetamido, propionamido, n-butylamido, isobutyl amido, cyclopropyl formamido, cyclobutyl formamido,cyclopentyl formamido, methylsulfonamido, ethylsulfonamido,propylsulfonamido, isopropylsulfonamido, dimethyl phosphinyl,diethyl phosphinyl, diisopropyl phosphinyl; R.sub.31 is selectedfrom: Hydrogen; C1-C6 alkyl, optionally substituted by halogen,nitro, amino, cyano; C3-C7 cycloalkyl, which is optionallysubstituted by halogen, nitro, amino, cyano; R.sub.41 is selectedfrom: hydrogen; C1-C6 alkyl, optionally substituted by halogen,nitro, amino, cyano; C3-C7 cycloalkyl, which is optionallysubstituted by halogen, nitro, amino, cyano; R.sub.51 is selectedfrom: 1) a hydrogen, halo, nitro, amino, cyano; 2) C1-C6 alkyl,optionally substituted by halogen, nitro, amino, cyano; --O--C1-C6alkyl, optionally substituted by halogen, nitro, amino, cyano; aC1-C6 oxygen-containing alkyl; 3) methylthio, ethylthio,isopropylthio, methylsulfinyl, ethyl sulfinyl, propyl sulfinyl,methylsulfonyl, ethylsulfonyl, isopropylsulfonyl, amino sulfonyl,ethylamino sulfonyl, propylamino sulfonyl, isopropylamino-sulfonyl,cyclopropylamino sulfonyl, hydroxyl formyl, methoxyl formyl,ethoxyl formyl, propoxyl formyl, isopropoxyl formyl, n-butoxylformyl, isobutoxyl formyl, t-butoxyl formyl, amino formyl,methylamino formyl, ethylamino formyl, propylamino formyl,isopropylamino formyl, cyclopropylamino formyl, cyclobutylaminoformyl, cyclopentylamino formyl, acetamido, propionamido, n-butylamido, isobutyl amido, cyclopropyl formamido, cyclobutyl formamido,cyclopentyl formamido, methylsulfonamido, ethylsulfonamido,propylsulfonamido, isopropylsulfonamido, dimethyl phosphinyl,diethyl phosphinyl, diisopropyl phosphinyl; or a stereoisomer ofthe above compounds, a prodrug thereof, a pharmaceuticallyacceptable salt thereof, or a pharmaceutically acceptable solvatethereof.

2. The method of claim 1, wherein the disease or disorderassociated with tissue damage is a trauma to brain, a trauma tospinal cord, a trauma to peripheral nerves, a trauma to retinal ora trauma to heart.

3. The method of claim 2, wherein the trauma to brain is ischemicstroke, blunt trauma, or subarachnoid hemorrhage.

4. The method of claim 2, wherein the trauma to spinal cord isspinal cord ischemia or spinal cord blunt force trauma.

5. (canceled)

6. The method of claim 2, wherein the trauma to retinal is macularedema, diabetic retinopathy, or glaucoma.

7. The method of claim 2, wherein the trauma to heart is myocardialinfarct, or chronic heart failure.

8. The method of claim 1, wherein the disease or disorderassociated with tissue damage is an organ failure.

9. The method of claim 8, wherein the organ failure is selectedfrom diabetes mellitus type I or II, nephrosis, fatty liverdiseases, failure of gonads, failure of pancreas, failure ofkidney, failure of heart, failure of lung, failure of liver, andfailure of bowel.

10. The method of claim 1, wherein the disease or disorderassociated with tissue damage is a disease or disorder caused byexposure to a toxic agent.

11. The method of claim 10, wherein the toxic agent is selectedfrom chemotherapeutic agents, chemical agents and radiationagents.

12. The method of claim 1, wherein the disease or disorderassociated with tissue damage is an inflammatory disease.

13. The method of claim 12, wherein the inflammatory disease isselected from sepsis, inflammatory bowel diseases, Crohn's disease,ulcerative colitis, ileitis, enteritis, and acute nephritis.

14. The method of claim 1, wherein the disease or disorderassociated with tissue damage is a degenerative disease.

15. The method of claim 14, wherein the degenerative disease isselected from muscular dystrophies, myotonic dystrophy, andneurodegenerative diseases.

16. The method of claim 1, wherein the MST1/2 protein kinaseinhibitor is administered prior to onset of the disease or disorderassociated with tissue damage, during development of the disease ordisorder associated with tissue damage, and/or after the disease ordisorder associated with tissue damage has developed.

17. The method of claim 1, wherein the MST1/2 protein kinaseinhibitor is administered from one minute to about 24 hours priorto onset of the disease or disorder associated with tissuedamage.

18. The method of claim 1, wherein the MST1/2 protein kinaseinhibitor is administered at a dose of from about 1 to about 10mg/kg bodyweight.

19-20. (canceled)

21. The method of claim 1, wherein the MST1/2 protein kinaseinhibitor is administered at a frequency of once in a period offrom 8 hours to 10 days.

22-28. (canceled)

29. The method of claim 1, wherein the MST1/2 protein kinaseinhibitor is selected from: TABLE-US-00007 ##STR00118## IA-1 (TFASalt) ##STR00119## IA-2 ##STR00120## IA-3 ##STR00121## IA-4 (TFASalt) ##STR00122## IA-5 (HCl Salt) ##STR00123## IA-6 (HCl Salt)##STR00124## IA-7 (HCl Salt) ##STR00125## IB-1 (TFA Salt)##STR00126## IB-2 (TFA Salt) ##STR00127## IB-3 (TFA Salt)##STR00128## IB-4 ##STR00129## IB-5 ##STR00130## IB-6 ##STR00131##IB-7 ##STR00132## IC-1 ##STR00133## IC-2 (TFA Salt) ##STR00134##IC-3 (TFA Salt) ##STR00135## IC-4 (TFA Salt) ##STR00136## IC-5 (TFASalt) ##STR00137## IC-6 ##STR00138## IC-7 ##STR00139## IC-8##STR00140## IC-9 ##STR00141## IC-10 (HCl Salt) ##STR00142## IC-11(TFA Salt) ##STR00143## ID-1 (HCl Salt) ##STR00144## ID-2 (HClSalt) ##STR00145## ID-3 (HCl Salt) ##STR00146## IE-1 (HCl Salt)##STR00147## IE-2 (TFA Salt) ##STR00148## IE-3 (TFA Salt)##STR00149## IF-1 ##STR00150## IF-2 ##STR00151## IF-3 ##STR00152##IF-4 (TFA Salt) ##STR00153## IF-5 (TFA Salt) ##STR00154## IF-6##STR00155## IG-1 (HCl Salt) ##STR00156## IG-2 (HCl Salt)##STR00157## IG-3 ##STR00158## IH-1 ##STR00159## IH-2 (TFA Salt)##STR00160## IH-3 (TFA Salt) ##STR00161## IH-4 (TFA Salt)##STR00162## IH-5 (TFA Salt) ##STR00163## IH-6 (HCl Salt)##STR00164## IH-7 (HCl Salt) ##STR00165## IH-8 ##STR00166## IH-9##STR00167## IH-10 (HCl Salt) ##STR00168## IH-11 (HCl Salt)##STR00169## IH-12 (HCl Salt) ##STR00170## IH-13 (HCl Salt)##STR00171## IH-14 (HCl Salt) ##STR00172## II-1 (TFA Salt)##STR00173## II-2 ##STR00174## II-3 ##STR00175## II-4 ##STR00176##II-5 ##STR00177## II-6 ##STR00178## II-7 ##STR00179## II-8 (TFASalt) ##STR00180## II-9 ##STR00181## II-10 (HCl Salt) ##STR00182##II-11 (HCl Salt) ##STR00183## II-12 (HCl Salt) ##STR00184## II-13(HCl Salt) ##STR00185## II-14 (TFA Salt) ##STR00186## II-15##STR00187## II-16 ##STR00188## II-17 (TFA Salt) ##STR00189## II-18(TFA Salt) ##STR00190## II-19 (TFA Salt) ##STR00191## II-20##STR00192## II-21 ##STR00193## II-22 ##STR00194## II-23 (TFA Salt)##STR00195## II-24

##STR00196## II-25 ##STR00197## II-26 (TFA Salt) ##STR00198## II-27##STR00199## II-28 ##STR00200## I-29 ##STR00201## II-30##STR00202## III-1 (TFA Salt) ##STR00203## III-2 ##STR00204## III-3##STR00205## III-4 ##STR00206## IV-1 (TFA Salt) ##STR00207## IV-2(TFA Salt) ##STR00208## IV-3 (TFA Salt)

30. The methods of claim 1, wherein the MST1/2 protein kinaseinhibitor is selected from: ##STR00209## ##STR00210##

31-34. (canceled)

Description

TECHNICAL FIELD

[0001] The present invention relates to the field of prevention ortreatment of tissue damages, in particular, it relates to usinginhibitors of MST1/2 protein kinases for prevention or treatment ofdiseases and disorders associated with tissue damages, forpromotion of tissue or organ regeneration and repair, forprevention or treatment of diseases caused by inflammation, and forprevention or treatment of neurological disorder related diseasesand ischemic diseases.

BACKGROUND TECHNOLOGY

[0002] How organ size is controlled in multicellular organisms is afundamental question in biology. It has been proposed that themammalian target of rapamycin (mTOR) pathway and the Hippo-YAPpathway control organ size by affecting cell size and cell number,respectively (Lee et al, Annu Rev Pharmacol Toxicol (2007)47:443-467; Zhao et al, Genes Dev (2010) 24:862-874). The Hippo-YAPpathway was initially defined by genetic studies in Drosophila, inwhich mosaic mutation of the Hippo-YAP pathway genes resulted intissue overgrowth (Pan, Genes Dev (2007) 21:886-897).

[0003] Function of the Hippo-YAP pathway in organ size regulationis conserved in mammals as demonstrated by studies usinggenetically modified mouse models (Camargo et al, Curr Biol (2007)17, 2054-2060; Dong et al, Cell (2007) 130:1120-1133; Heallen etal, Science (2011) 332:458-461; Lee et al, Proc Natl Acad Sci USA(2010) 107:8248-8253; Lu et al, Proc Natl Acad Sci USA (2010)107:1437-1442; Song et al, Proc Natl Acad Sci USA (2010)107:1431-1436).

[0004] Core components of the mammalian Hippo-YAP pathway include akinase cascade of mammalian sterile 20-like kinase-1/2 (MST1/2) andLATS1/2. MST1/2, in complex with its regulatory protein Salvador(SAV1), phosphorylates and activates LATS1/2 kinases, which alsoform a complex with its regulatory protein MOB1 (Zhao et al, GenesDev (2010) 24:862-874). SAV1 forms complexes with MST1/2, whereasMOB kinase activator 1A (MOB1A) and MOB1B interact with LATS1/2.When the Hippo pathway is activated, MST1/2 activate LATS1/2 andMOB1A/1B by phosphorylation. Subsequently, LATS1/2 directlyphosphorylate YAP and TAZ. Phosphorylation inhibits YAP and TAZactivities by activating a phosphodegron that is targeted by.beta.-transducin repeat-containing protein (.beta.-TrCP), leadingto the degradation of YAP and TAZ proteins. As a result, YAP andTAZ accumulate in the nucleus and promote gene expression when theHippo pathway is not active (FIG. 1).

[0005] MST1/2 are a ubiquitously expressed serine/threonine kinase,which belongs to a mammalian sterile 20 (STE 20)--like kinasefamily consisting of PAKI, MST1, MST2, KHS, GCK, SOK1, NIK, HPK1AND SPS1. Increasing lines of evidence suggests that MST1/2 andother STE20-like family kinases play an important role in mediatingapoptosis. MST1/2 are activated by some pro-apoptotic stimuli infibroblastic and lymphocytic cell lines. MST1/2 are cleaved bycaspases and this cleavage increases kinase activities of MST1/2,which in turn activates caspase 3, thereby constituting a powerfulamplification loop of apoptotic response (Cinar et al., EMBO J.(2007) 26:4523-4534; Song and Lee, Cell Signal. (2008)20:892-906).

[0006] Conditional knockout of MST1/2 protein kinases can promoteliver regeneration (Zhou et al. Cancer Cell (2009) 16:425-438), andimmunosuppression (Mou et al. J. Exp. Med. (2012) 209:741-759).Reducing MST1/2 protein level or enzymatic activity helps reducingneuronal cell death, and thus be useful for prevention andtreatment of neurological disorders or neurodegenerative diseases,including Alzheimer's disease, multiple sclerosis, Parkinson'sdisease, stroke, etc. (Lehtinen et al. Cell (2006) 125:987-1001);and oxidizing stress-related myocardial ischemia and peripheralischemia (US 2008/0242608).

[0007] Thus, the present invention provides a method for preventingor treating tissue damage related diseases and disorder using smallmolecule inhibitors of MST1/2 protein kinases.

SUMMARY OF INVENTION

[0008] The invention provides a method of preventing or treating adisease or disorder associated with tissue damage using a MST1/2protein kinase inhibitor, said method comprising a step ofadministering the MST1/2 protein kinase inhibitor to a subjecthaving the disease or disorder, wherein the MST1/2 protein kinaseinhibitor is administered at a dose of from about 0.1 mg/kg toabout 100 mg/kg based on the bodyweight of the subject and at afrequency of once in a period of from 6 hours to 20 days, and theMST1/2 protein kinase inhibitor has a formula:

##STR00001##

or a stereoisomer of the above compounds, a prodrug thereof, apharmaceutically acceptable salt thereof, or a pharmaceuticallyacceptable solvate thereof. The definition of the substituents andsymbols are described in detail below.

[0009] In the previous embodiment, the disease or disorderassociated with tissue damage may be a trauma to brain, a trauma tospinal cord, a trauma to peripheral nerves, a trauma to retinal ora trauma to heart. In one embodiment, the trauma to brain may beischemic stroke, blunt trauma, or subarachnoid hemorrhage. Inanother embodiment, the trauma to spinal cord may be spinal cordischemia or spinal cord blunt force trauma. In yet anotherembodiment, the trauma to peripheral nerves may be sciatic nerveinjury, diabetic neuropathy, or carpal tunnel syndrome. In yetanother embodiment, the trauma to retinal may be macular edema,diabetic retinopathy, or glaucoma. In yet another embodiment, thetrauma to heart may be myocardial infarct, or chronic heartfailure.

[0010] In any one of the previous embodiments, the disease ordisorder associated with tissue damage may be an organ failure. Inone embodiment, the organ failure may be selected from diabetesmellitus type I or II, nephrosis, fatty liver diseases, failure ofgonads, failure of pancreas, failure of kidney, failure of heart,failure of lung, failure of liver, and failure of bowel.

[0011] In any one of the previous embodiments, the disease ordisorder associated with tissue damage may be a disease or disordercaused by exposure to a toxic agent. In one embodiment, the toxicagent may be selected from chemotherapeutic agents, chemical agentsand radiation agents.

[0012] In any one of the previous embodiments, the disease ordisorder associated with tissue damage may be an inflammatorydisease. In one embodiment, the inflammatory disease may beselected from sepsis, inflammatory bowel diseases, Crohn's disease,ulcerative colitis, ileitis, enteritis, and acute nephritis.

[0013] In any one of the previous embodiments, the disease ordisorder associated with tissue damage may be a degenerativedisease. In one embodiment, the degenerative disease may beselected from muscular dystrophies, myotonic dystrophy, andneurodegenerative diseases.

[0014] In any one of the previous embodiments, the MST1/2 proteinkinase inhibitor may be administered prior to onset of the diseaseor disorder associated with tissue damage, during development ofthe disease or disorder associated with tissue damage, and/or afterthe disease or disorder associated with tissue damage hasdeveloped.

[0015] In any one of the previous embodiments, the MST1/2 proteinkinase inhibitor may be administered from one minute to about 24hours, or from about 5 minutes to about 10 hours, or from about 5minutes to about 5 hours prior to onset of the disease or disorderassociated with tissue damage.

[0016] In any one of the previous embodiments, the MST1/2 proteinkinase inhibitor may be administered at a dose of from about 1 toabout 10 mg/kg bodyweight.

[0017] In any one of the previous embodiments, the MST1/2 proteinkinase inhibitor may be administered at a dose of from about 0.1 toabout 10 mg/kg bodyweight.

[0018] In any one of the previous embodiments, the MST1/2 proteinkinase inhibitor may be administered at a dose of from about 10 toabout 100 mg/kg bodyweight.

[0019] In any one of the previous embodiments, the MST1/2 proteinkinase inhibitor may be administered at a frequency of once in aperiod of from 8 hours to 10 days, from 12 hours to 7 days, or from12 hours to 3 days, or from 12 hours to 24 hours.

[0020] In any one of the previous embodiments, the MST1/2 proteinkinase inhibitor may be administered through intravenous injection,intravenous infusion, intravenous drip, subcutaneous injection,sublingual administration, or oral administration to the subject.In one embodiment, the the MST1/2 protein kinase inhibitor may beadministered by intravenous injection or intravenous infusion.

[0021] In any one of the previous embodiments, the method mayfurther comprise a step of administering an additional activeingredient that is effective in treating a degenerative disease ortissue injury. In one embodiment, the additional active ingredientand the MST1/2 protein kinase inhibitor may be administeredsimultaneously or in separate sequential administrations. Inanother embodiment, the additional active ingredient and the MST1/2protein kinase inhibitor may be administered in separate sequentialadministrations that are less than half an hour, 1 hour, 2 hours, 4hours, 6 hours, 8 hours, or 10 hours apart. In yet anotherembodiment, the additional active ingredient may be selected fromchemoprotective agents, myeloprotective agents, anti-apoptoticagents, and pro-proliferative agents.

[0022] In any one of the previous embodiments, the MST1/2 proteinkinase inhibitor may have one of formulas I, II, III, IV:

##STR00002##

[0023] wherein definition of the substituents and symbols aredescribed in detail below.

[0024] In any one of the previous embodiments, the MST1/2 proteinkinase inhibitor may be selected from the compounds in Tables1-4.

[0025] In any one of the previous embodiments, the MST1/2 proteinkinase inhibitor may be selected from:

##STR00003## ##STR00004##

[0026] In any one of the previous embodiments, the MST1/2 proteinkinase inhibitor may be administered in its prodrug form or itssalt form.

[0027] In any one of the previous embodiments, the MST1/2 proteinkinase inhibitor may be administered in a pharmaceuticalcomposition comprising a pharmaceutical excipient selected fromcarriers, diluents, fillers, buffers, bulking agents, stabilizers,and solubilizers.

[0028] In any one of the previous embodiments, the MST1/2 proteinkinase inhibitor may be administered in a pharmaceuticalcomposition comprising a solubilizing agent, an emulsifier, or asurfactant.

[0029] In any one of the previous embodiments, the MST1/2 proteinkinase inhibitor may be administered in a pharmaceuticalcomposition comprising a bioavailability enhancing agent or anabsorption enhancing agent.

BRIEF DESCRIPTION OF DRAWINGS

[0030] FIG. 1 is a schematic presentation of the Hippo pathwaywhere MST1/2 protein kinases play a pivotal role.

[0031] FIG. 2 shows inhibition of Mob1 phosphorylation by MST1/2protein kinase inhibitors at a dose dependent manner.

[0032] FIG. 3 shows the Mob1 phosphorylation inhibition curve by anMST1/2 protein kinase inhibitor.

[0033] FIGS. 4A-4D show reduction of H.sub.2O.sub.2 inducedneonatal rat cardiomyocytes (NRCM) death by the MST1/2 proteinkinase inhibitors.

[0034] FIGS. 5A-5D show reduction of isoproterenol (ISO) inducedcardiomyocytes death by the MST1/2 protein kinase inhibitors.

[0035] FIGS. 6A-6D show reduction of doxorubicin (DOX) inducedcardiomyocytes death by the MST1/2 protein kinase inhibitors.

[0036] FIG. 7A shows inhibition of apoptosis in cardiomyocytesinduced by H.sub.2O.sub.2.

[0037] FIG. 7B shows inhibition of apoptosis in cardiomyocytesinduced by ISO.

[0038] FIG. 7C shows inhibition of apoptosis in cardiomyocytesinduced by DOX.

[0039] FIG. 8 shows a study design for inhibition of heart atrophyby the MST1/2 protein kinase inhibitors in mice.

[0040] FIG. 9 shows percentages of survivals over the study periodafter treatment by Dox, optionally and additionally with the MST1/2protein kinase inhibitors.

[0041] FIG. 10 shows ratio of heart weight over bodyweight of themice after treatment by Dox, optionally and additionally with theMST1/2 protein kinase inhibitors.

[0042] FIG. 11 shows heart ejection fraction (EF %) of the miceafter treatment by Dox, optionally and additionally with the MST1/2protein kinase inhibitors.

[0043] FIG. 12 shows the rate of incorporation of EdU intocardiomyocytes of mice after treatment with the MST1/2 proteinkinase inhibitors.

[0044] FIGS. 13 and 14A show inhibition of infarct size in ischemicstroke mice after treatment with the MST1/2 protein kinaseinhibitors.

[0045] FIG. 14B shows protection of neurological functions inischemic stroke mice after treatment with the MST1/2 protein kinaseinhibitors.

[0046] FIGS. 15A-15B show inhibition of VCAM-1 expression in humanumbilical vein endothelial cells (HUVEC) as induced by TNF.alpha.using the MST1/2 protein kinase inhibitor X1.

[0047] FIGS. 16A-16B show inhibition of VCAM-1 expression in HUVECas induced by TNF.alpha. using the MST1/2 protein kinase inhibitorY1.

[0048] FIGS. 17A-17D show inhibition of expression of inflammatorymarkers in HUVEC induced by TNF.alpha. using the MST1/2 proteinkinase inhibitor X1.

[0049] FIGS. 18A-18D show inhibition of expression of inflammatorymarkers in HUVEC induced by TNF.alpha. using the MST1/2 proteinkinase inhibitor Y1.

[0050] FIGS. 19A-19B show inhibition of cell adhesion induced byTNF.alpha. using the MST1/2 protein kinase inhibitor X1.

[0051] FIGS. 20A-20B show inhibition of cell adhesion induced byTNF.alpha. using the MST1/2 protein kinase inhibitor YL.

[0052] FIGS. 21A-21D show inhibition of expression of inflammatorymarkers induced by lipopolysaccharides (LPS) using the MST1/2protein kinase inhibitor X1.

DEFINITIONS

[0053] In order to facilitate understanding of the examplesprovided herein, certain frequently occurring terms are definedherein.

[0054] In connection with a measured quantity, the term "about" asused herein refers to the normal variation in that measuredquantity that would be expected by a skilled person making themeasurement and exercising a level of care commensurate with theobjective of the measurement and the precision of the measuringequipment used. Unless otherwise indicated, "about" refers to avariation of +/-10%, +/-5%, or +/-2% of the value provided.

[0055] The term "administering" or "administration" as used hereinrefers to local and systemic administration of an inhibitor ofMST1/2 protein kinase, e.g., including enteral, parenteral,pulmonary, and topical/transdermal administration. Routes ofadministration for an inhibitor of MST1/2 as described hereininclude, e.g., oral (per os (P.O.)) administration, nasal orinhalation administration, administration as a suppository, topicalcontact, transdermal delivery (e.g., via a transdermal patch),intrathecal (IT) administration, intravenous ("i.v.")administration, intraperitoneal ("i.p.") administration,intramuscular ("im") administration, intratumoral administration,intralesional administration, or subcutaneous ("sc")administration, or the implantation of a slow-release device e.g.,a mini-osmotic pump, a depot formulation, etc., to a subject.Administration can be by any route including parenteral andtransmucosal (e.g., oral, nasal, vagin*l, rectal, or transdermal).Parenteral administration includes, e.g., intravenous,intramuscular, intra-arterial, intradermal, subcutaneous,intraperitoneal, intraventricular, ionophoretic and intracranial.Other modes of delivery include, but are not limited to, the use ofliposomal formulations, intravenous infusion, transdermal patches,etc.

[0056] The term "cancer" as used herein refers to any abnormalgrowth exhibiting malignant properties: the ability (1) to grow anddivide without respect to normal limits, (2) to invade and destroyadjacent tissues, and (3) in some instances, spread to otherlocations in the body. Cancer includes cancers or neoplasticdisorders of the central nervous system, peripheral nervous system,gastrointestinal/digestive system, genitourinary system,gynecological, head and neck, hematological/blood,musculoskeletal/soft tissue, respiratory, and breast. Furtherexamples of cancers or neoplastic disorders include, but are notlimited to, those of the brain (astrocytoma, gliobastoma, glioma),spinal cord, pituitary gland, breast (Infiltrating cancers,Pre-invasive cancers, inflammatory cancers, Paget's Disease,Metastatic and Recurrent Breast Cancer), blood (Hodgkin's Disease,Leukemia, Multiple Myeloma, Lymphoma), Lymph node cancer, Lung(Adenocarcinoma, Oat Cell lung cancer, Non-small Cell lung cancer,Small Cell lung cancer, Squamous Cell lung cancer, Mesothelioma),skin (melanoma, basal cell skin cancer, squamous cell skin cancer,Kapsosis Sarcoma), Bone Cancer (Ewings Sarcoma, Osteosarcoma,Chondrosarcoma), head and neck (laryngeal, pharyngeal (nasal cavity& sinus cavity), and esophageal cancers), oral (jaw, salivarygland, throat, thyroid, tongue, and tonsil cancers), eye,gynecological (Cervical, Endometrial, Fallopian, Ovarian, Uterine,vagin*l, and Vulvar), genitourinary (bladder, kidney, penile,prostate, testicular, and urinary cancers), adrenal (corticaladenoma, cortical carcinoma, pheochromocytoma) and gastrointestinal(appendix, bile duct (extrahepatic bile duct) colon, gallbladder,gastric, intestinal, colon, liver, pancreatic, rectal, and stomachcancers) as well as those listed below: (Fishman et al., 1985,Medicine, 2d Ed., J. B. Lippincott Co., Philadelphia): Leukemia:acute leukemia, acute lymphocytic leukemia, acute myelocyticleukemia, myeloblastic, promyelocytic, myelomonocytic, monocyticerythroleukemia, chronic leukemia, chronic myelocytic(granulocytic) leukemia, chronic lymphocytic leukemia, Polycythemiavera, Gastric carcinoma; Lymphoma (malignant and non-malignant):Hodgkin's disease, non-Hodgkin's disease, Multiple myeloma,Waldenstrom's macroglobulinemia, Heavy chain disease; Solid tumorssarcomas and carcinomas: fibrosarcoma, myxosarcoma, liposarcoma,chondrosarcoma, osteogenic sarcoma, chordoma, angiosarcoma,endotheliosarcoma, lymphangiosarcoma, lymphangioendotheliosarcoma,synovioma, mesothelioma, Ewing's tumor, leiomyosarcoma,rhabdomyosarcoma, colon carcinoma, pancreatic cancer, breastcancer, ovarian cancer, prostate cancer, squamous cell carcinoma,oral squamous cell carcinoma, hepatocellular carcinoma, basal cellcarcinoma, adenocarcinoma, sweat gland carcinoma, sebaceous glandcarcinoma, papillary carcinoma; papillary adenocarcinomas:cystadenocarcinoma, medullary carcinoma, bronchogenic carcinoma,renal cell carcinoma, hepatoma, bile duct carcinoma,choriocarcinoma, seminoma, embryonal carcinoma, Wilms' tumor,cervical cancer, cervix adenocarcinoma, uterine cancer, testiculartumor, lung carcinoma, small cell lung carcinoma, non-small celllung adenocarcinoma, bladder carcinoma, epithelial carcinoma,glioma, malignant glioma, glioblastoma, multiforme astrocyticgliomas, medulloblastoma, craniopharyngioma, ependymoma, pinealoma,hemangioblastoma, acoustic neuroma, oligodendroglioma, meningioma,melanoma, neuroblastoma, or retinoblastoma.

[0057] The term "chemical-induced toxicity" as used herein refersto injury induced by a chemical agent to a cell or tissue arisingfrom exposure to the chemical agent. Phenotypically,chemical-induced injury includes one or more of the following:structural chemical injury to a cell or tissue, inflammation,fibroproliferative tissue effects, adverse tissue remodeling,(e.g., increased neutrophil infiltration), relative to that seen ina cell or tissue not exposed to a chemical.

[0058] The term "chemotherapeutic agent" as used herein refers to achemical compound useful in the treatment of cancer. Examples ofchemotherapeutic agents include alkylating agents such as thiotepaand cyclosphosphamide (CYTOXAN.RTM.); alkyl sulfonates such asbusulfan, improsulfan and piposulfan; aziridines such as benzodopa,carboquone, meturedopa, and uredopa; ethylenimines andmethylamelamines including altretamine, triethylenemelamine,triethylenephosphoramide, triethylenethiophosphoramide andtrimethylomelamine; acetogenins (especially bullatacin andbullatacinone); delta-9-tetrahydrocannabinol (dronabinol,MARINOL.RTM.); beta-lapachone; lapachol; colchicines; betulinicacid; a camptothecin (including the synthetic analogue topotecan(HYCAMTIN.RTM.), CPT-11 (irinotecan, CAMPTOSAR.RTM.),acetylcamptothecin, scopolectin, and 9-aminocamptothecin);bryostatin; callystatin; CC-1065 (including its adozelesin,carzelesin and bizelesin synthetic analogues); podophyllotoxin;podophyllinic acid; teniposide; cryptophycins (particularlycryptophycin 1 and cryptophycin 8); dolastatin; duocarmycin(including the synthetic analogues, KW-2189 and CB1-TM1);eleutherobin; pancratistatin; a sarcodictyin; spongistatin;nitrogen mustards such as chlorambucil, chlornaphazine,chlorophosphamide, estramustine, ifosfamide, mechlorethamine,mechlorethamine oxide hydrochloride, melphalan, novembichin,phenesterine, prednimustine, trofosfamide, uracil mustard;nitrosoureas such as carmustine, chlorozotocin, fotemustine,lomustine, nimustine, and ranimnustine; antibiotics such as theenediyne antibiotics (e.g., calicheamicin, especially calicheamicingamma1I and calicheamicin omegaI1 (see, e.g., Nicolaou et al.,Angew. Chem. Intl. Ed. Engl., 33: 183-186 (1994)); CDP323, an oralalpha-4 integrin inhibitor; dynemicin, including dynemicin A; anesperamicin; as well as neocarzinostatin chromophore and relatedchromoprotein enediyne antibiotic chromophores), aclacinomysins,actinomycin, authramycin, azaserine, bleomycins, cactinomycin,carabicin, caminomycin, carzinophilin, chromomycins, dactinomycin,daunorubicin, detorubicin, 6-diazo-5-oxo-L-norleucine, doxorubicin(including ADRIAMYCIN.RTM., morpholino-doxorubicin,cyanomorpholino-doxorubicin, 2-pyrrolino-doxorubicin, doxorubicinHCl liposome injection (DOXIL.RTM.), liposomal doxorubicin TLC D-99(MYOCET.RTM.), peglylated liposomal doxorubicin (CAELYX.RTM.), anddeoxydoxorubicin), epirubicin, esorubicin, idarubicin,marcellomycin, mitomycins such as mitomycin C, mycophenolic acid,nogalamycin, olivomycins, peplomycin, porfiromycin, puromycin,quelamycin, rodorubicin, streptonigrin, streptozocin, tubercidin,ubenimex, zinostatin, zorubicin; anti-metabolites such asmethotrexate, gemcitabine (GEMZAR.RTM.), tegafur (UFTORAL.RTM.),capecitabine (XELODA.RTM.), an epothilone, and 5-fluorouracil(5-FU); folic acid analogues such as denopterin, methotrexate,pteropterin, trimetrexate; purine analogs such as fludarabine,6-mercaptopurine, thiamiprine, thioguanine; pyrimidine analogs suchas ancitabine, azacitidine, 6-azauridine, carmofur, cytarabine,dideoxyuridine, doxifluridine, enocitabine, floxuridine; androgenssuch as calusterone, dromostanolone propionate, epitiostanol,mepitiostane, testolactone; anti-adrenals such asaminoglutethimide, mitotane, trilostane; folic acid replenishersuch as frolinic acid; aceglatone; aldophosphamide glycoside;aminolevulinic acid; eniluracil; amsacrine; bestrabucil;bisantrene; edatraxate; defofamine; demecolcine; diaziquone;elformithine; elliptinium acetate; an epothilone; etoglucid;gallium nitrate; hydroxyurea; lentinan; lonidainine; maytansinoidssuch as maytansine and ansamitocins; mitoguazone; mitoxantrone;mopidanmol; nitraerine; pentostatin; phenamet; pirarubicin;losoxantrone; 2-ethylhydrazide; procarbazine; PSK.RTM.polysaccharide complex (JHS Natural Products, Eugene, Oreg.);razoxane; rhizoxin; sizofiran; spirogermanium; tenuazonic acid;triaziquone; 2,2',2'-trichlorotriethylamine; trichothecenes(especially T-2 toxin, verracurin A, roridin A and anguidine);urethan; vindesine (ELDISINE.RTM., FILDESIN.RTM.); dacarbazine;mannomustine; mitobronitol; mitolactol; pipobroman; gacytosine;arabinoside ("Ara-C"); thiotepa; taxoid, e.g., pacl*taxel(TAXOL.RTM.), albumin-engineered nanoparticle formulation ofpacl*taxel (ABRAXANE.TM.), and docetaxel (TAXOTERE.RTM.);chloranbucil; 6-thioguanine; mercaptopurine; methotrexate; platinumagents such as cisplatin, oxaliplatin (e.g., ELOXATIN.RTM.), andcarboplatin; vincas, which prevent tubulin polymerization fromforming microtubules, including vinblastine (VELBAN.RTM.),vincristine (ONCOVIN.RTM.), vindesine (ELDISINE.RTM.,FILDESIN.RTM.), and vinorelbine (NAVELBINE.RTM.); etoposide(VP-16); ifosfamide; mitoxantrone; leucovorin; novantrone;edatrexate; daunomycin; aminopterin; ibandronate; topoisomeraseinhibitor RFS 2000; difluoromethylornithine (DMF.RTM.); retinoidssuch as retinoic acid, including bexarotene (TARGRETIN.RTM.);bisphosphonates such as clodronate (for example, BONEFOS.RTM. orOSTAC.RTM.), etidronate (DIDROCAL.RTM.), NE-58095, zoledronicacid/zoledronate (ZOMETA.RTM.), alendronate (FOSAMAX.RTM.),pamidronate (AREDIA.RTM.), tiludronate (SKELID.RTM.), orrisedronate (ACTONEL.RTM.); troxacitabine (a 1,3-dioxolanenucleoside cytosine analog); antisense oligonucleotides,particularly those that inhibit expression of genes in signalingpathways implicated in aberrant cell proliferation, such as, forexample, PKC-alpha, Raf, H-Ras, and epidermal growth factorreceptor (EGF-R); vaccines such as THERATOPE.RTM. vaccine and genetherapy vaccines, for example, ALLOVECTIN.RTM. vaccine,LEUVECTIN.RTM. vaccine, and VAXID.RTM. vaccine; topoisomerase 1inhibitor (e.g., LURTOTECAN.RTM.); rmRH (e.g., ABARELIX.RTM.);BAY439006 (sorafenib; Bayer); SU-11248 (sunitinib, SUTENT.RTM.,Pfizer); perifosine, COX-2 inhibitor (e.g. celecoxib oretoricoxib), proteosome inhibitor (e.g. PS341); bortezomib(VELCADE.RTM.); CCI-779; tipifarnib (R11577); orafenib, ABT510;Bcl-2 inhibitor such as oblimersen sodium (GENASENSE.RTM.);pixantrone; EGFR inhibitors (see definition below); tyrosine kinaseinhibitors (see definition below); serine-threonine kinaseinhibitors such as rapamycin (sirolimus, RAPAMUNE.RTM.);farnesyltransferase inhibitors such as lonafarnib (SCH 6636,SARASAR.TM.); and pharmaceutically acceptable salts, acids orderivatives of any of the above; as well as combinations of two ormore of the above such as CHOP, an abbreviation for a combinedtherapy of cyclophosphamide, doxorubicin, vincristine, andprednisolone; and FOLFOX, an abbreviation for a treatment regimenwith oxaliplatin (ELOXATIN.RTM.) combined with 5-FU andleucovorin.

[0059] The term "individual" or "subject" or "patient" as usedherein refers to a mammal. Mammals include, but are not limited to,domesticated animals (e.g., cows, sheep, cats, dogs, and horses),primates (e.g., humans and non-human primates such as monkeys),rabbits, and rodents (e.g., mice and rats). In certain embodiments,the individual or subject is a human, which is commonly termed"patients".

[0060] The term "inflammatory diseases" as used herein refers tovarious diseases having an inflammatory component. Non-limitingexamples include, but are not limited to, appendicitis,blepharitis, bronchitis, bursitis, cervicitis, cholangitis,cholecystitis, chorioamnionitis, conjunctivitis, cystitis,dacryoadenitis, dermatitis, endocarditis, endometritis,epicondylitis, epididymitis, fibrositis, gastritis, gingivitis,glossitis, hidradenitis suppurativa, iritis, laryngitis, mastitis,myocarditis, myositis, nephritis, omphalitis, oophoritis, orchitis,osteitis, otitis, parotitis, pericarditis, peritonitis,pharyngitis, pleuritis, phlebitis, pneumonitis (pneumonia),prostatitis, pyelonephritis, rhinitis, salpingitis, sinusitis,stomatitis, synovitis, tonsillitis, uveitis, urethritis, vaginitis,vulvitis, asthma, systemic lupus erythematosus, myasthenia gravis,tendonitis, angiitis, chronic bronchitis, pancreatitis,osteomyelitis, arthritis (rheumatoid and psoriatic),glumeronephritis, optic neuritis, temporal arteritis, encephalitis,meningitis, traverse myelitis, dermatomyositis, polymyositis,necrotizing fasciitis, hepatitis, necrotizing entercolitis, pelvicinflammatory disease, inflammatory bowel disease (ulcerativecolitis, Crohn's disease, ileitis, and enteritis), proctitis,vasculitis, vascular stenosis, restenosis, hypotension, Type-1diabetes, Kawasaki disease, Decum's disease, chronic obstructivepulmonary disease, psoriasis, artherosclerosis, scleroderma,Sjogren's syndrome, mixed connective tissue disease, rosacea,gastric ulcers, duodenal ulcers, Alzheimer's disease, adult onsetStill's disease, acute retinal pigment epitheliitis, Tietze'ssyndrome, Bechcet's disease, white dot syndrome (acute posteriormultifocal placoid pigment epitheliopathy, serpiginous choroiditis,birdshot chorioretinopathy, multifocal choroiditis with panuveitis,diffuse subretinal fibrosis syndrome, punctuate innerchoroidopathy, multiple evanescent white dot syndrome, and diffuseunilateral subacute neuroretinitis), granuloma annulare, irritablebowel syndrome, gastroenteritis, Grave's disease, multiplesclerosis, Dupuytren's contracture, graft rejection diseases(including allograft rejection and graft-v-host disease), e.g. skingraft rejection, solid organ transplant rejection, bone marrowtransplant rejection, inflammatory dermatoses.

[0061] The phrase "pharmaceutically acceptable salt(s)" as usedherein refers to those salts of a compound of interest that aresafe and effective for topical use in mammals and that possess thedesired biological activity. Pharmaceutically acceptable saltsinclude salts of acidic or basic groups present in the specifiedcompounds. Pharmaceutically acceptable acid addition salts include,but are not limited to, hydrochloride, hydrobromide, hydroiodide,nitrate, sulfate, bisulfate, phosphate, acid phosphate,isonicotinate, acetate, lactate, salicylate, citrate, tartrate,pantothenate, bitartrate, ascorbate, succinate, maleate,gentisinate, fumarate, gluconate, glucaronate, saccharate, formate,benzoate, glutamate, methanesulfonate, ethanesulfonate,benzensulfonate, p-toluenesulfonate and pamoate (i.e.,1,1'-methylene-bis-(2-hydroxy-3-naphthoate)) salts. Certaincompounds used in the present invention can form pharmaceuticallyacceptable salts with various amino acids. Suitable base saltsinclude, but are not limited to, aluminum, calcium, lithium,magnesium, potassium, sodium, zinc, and diethanolamine salts.Illustrative salts are the ammonium, potassium, sodium, calcium,and magnesium salts. Salts derived from pharmaceutically acceptableorganic non-toxic bases include, but are not limited to, salts ofprimary, secondary, and tertiary amines, substituted aminesincluding naturally occurring substituted amines, cyclic amines andbasic ion exchange resins, such as isopropylamine, trimethylamine,diethylamine, triethylamine, tripropylamine, ethanolamine,2-dimethylaminoethanol, 2-diethylaminoethanol, dicyclohexylamine,lysine, arginine, histidine, caffeine, procaine, hydrabamine,choline, betaine, ethylenediamine, glucosamine, methylglucamine,theobromine, purines, piperazine, piperidine, N-ethylpiperidine,polyamine resins, and the like.

[0062] Illustrative organic bases are isopropylamine, diethylamine,ethanolamine, trimethylamine, dicyclohexylamine, choline, andcaffeine. See Berge et al., J. Pharm. Sci. 66:1-19 (1977),incorporated herein by reference.

[0063] The term "pharmaceutically-acceptable carrier" as usedherein refers to one or more compatible solid or liquid fillerdiluents or encapsulating substances which are suitable foradministration to a subject. The term "compatible" as used hereinmeans that the components of the composition are capable of beingcommingled with the active compound, and with each other, in amanner such that there is no interaction which would substantiallyreduce the pharmaceutical efficacy of the composition underordinary use situations. Pharmaceutically-acceptable carriers must,of course, be of sufficiently high purity and sufficiently lowtoxicity to render them suitable for administration preferably toan animal, preferably mammal being treated. Some examples ofsubstances, which can serve as pharmaceutically-acceptable carriersor components thereof, are sugars, such as lactose, glucose andsucrose; starches, such as corn starch and potato starch; celluloseand its derivatives, such as sodium carboxymethyl cellulose, ethylcellulose, and methyl cellulose; powdered tragacanth; malt;gelatin; talc; solid lubricants, such as stearic acid and magnesiumstearate; calcium sulfate; vegetable oils, such as peanut oil,cottonseed oil, sesame oil, olive oil, corn oil and oil oftheobroma; polyols such as propylene glycol, glycerine, sorbitol,mannitol. and polyethylene glycol; alginic acid; emulsifiers, suchas the TWEEN.RTM. emulsifiers; wetting agents, such sodium laurylsulfate; coloring agents; flavoring agents; tableting agents,stabilizers; antioxidants; preservatives; pyrogen-free water;isotonic saline; and phosphate buffer solutions. The choice of apharmaceutically-acceptable carrier to be used in conjunction withthe subject compound is basically determined by the way thecompound is to be administered. If the subject compound is to beinjected, the preferred pharmaceutically-acceptable carrier issterile, physiological saline, with blood-compatible suspendingagent, the pH of which has been adjusted to about 7.4. Inparticular, pharmaceutically-acceptable carriers for systemicadministration include sugars, starches, cellulose and itsderivatives, malt, gelatin, talc, calcium sulfate, vegetable oils,synthetic oils, polyols, alginic acid, phosphate buffer solutions,emulsifiers, isotonic saline, and pyrogen-free water. Preferredcarriers for parenteral administration include propylene glycol,ethyl oleate, pyrrolidone, ethanol, and sesame oil. Preferably, thepharmaceutically-acceptable carrier, in compositions for parenteraladministration, comprises at least about 90% by weight of the totalcomposition. The compositions of this invention are preferablyprovided in unit dosage form.

[0064] The term "preventing a disease or disorder" as used hereinrefers to delaying the onset, hindering the progress, hindering theappearance, protection against, inhibiting or eliminating theemergence, or reducing the incidence, of such damages, effects orsymptoms. Use of the term "prevention" is not meant to imply thatall patients in a patient population administered a preventativetherapy will never be affected by or develop symptoms in responseto the disease or disorder associated with tissue damage targetedfor prevention, but rather that the patient population will exhibita reduction in the damage, effects, or symptoms of the disease ordisorder.

[0065] The term "prodrug" as used herein refers to a compound thatmay be transformed in vivo to yield an active compound, forexample, by hydrolysis, oxidaction, or other reactions, in the gutor enzymatic conversion in blood. Common examples include, but arenot limited to, ester and amide forms of an active compound havingan active form bearing a carboxylic acid moiety. Examples ofpharmaceutically acceptable esters of the active compounds of thisinvention include, but are not limited to, alkyl esters (forexample with between about one and about six carbons) where thealkyl group is a straight or branched chain. Acceptable esters alsoinclude cycloalkyl esters and arylalkyl esters such as, but notlimited to benzyl. Examples of pharmaceutically acceptable amidesof the active compounds of this invention include, but are notlimited to, primary amides, and secondary and tertiary alkyl amides(for example with between about one and about six carbons). Amidesand esters of the active compounds of the present invention can beprepared according to conventional methods. A thorough discussionof prodrugs is provided in Bioreversible Carriers in Drug Design,ed. Edward B. Roche, American Pharmaceutical Association andPergamon Press, 1987 and Burger's Medicinal Chemistry and DrugDiscovery, (1995) 172-178, 949-982 (Manfred E. Wolff ed., 5th ed.),which are incorporated herein by reference for all purposes.

[0066] The term "prophylactically effective amount" as used hereinrefers to an amount of a compound sufficient to result in theprevention of the damage, effects or symptoms resulting from adisease or disorder associated with tissue damage. Aprophylactically effective amount can refer to the amount of thecompound sufficient to prevent the damage, effects or symptomsresulting from a disease or disorder associated with tissuedamage.

[0067] The term "radiation toxicity" or "radiation-inducedtoxicity" as used herein refers to radiation-induced injury to acell or tissue arising from exposure to a radiation agent becauseof either radiation therapy or accidental radiation exposure.Phenotypically, radiation-induced injury includes one or more ofthe following: structural radiation injury to a cell or tissue,increased neutrophil infiltration, increased collagen type IIIdeposition, and increased smooth muscle cell proliferation relativeto that seen in a cell or tissue not exposed to radiation.

[0068] The term "radiation agent" as used herein refers to anyradioactive material that may kill or injure a subject, and may beused for therapeutical purposes (e.g., radiotherapy) or as weaponsto cause bodily injuries or harm or even death to a population.Radioactive agents may include, but are not limited to .sup.137Cs,.sup.60Co, .sup.241Am, .sup.252Cf, .sup.192Ir, .sup.238Pu,.sup.90Sr, .sup.226Ra, .sup.91Sr, .sup.92Sr, .sup.95Zr, .sup.99Mo,.sup.106Ru, .sup.131Sb, .sup.132Te, .sup.139Te, .sup.140Ba,.sup.141La, .sup.144Ce, .sup.233U, .sup.235U, .sup.238U, .sup.228P,.sup.229P, .sup.230P, .sup.231P, .sup.232P, .sup.233P, .sup.234P,.sup.235P, .sup.236P, .sup.237P, .sup.238P, .sup.239P, .sup.240P,.sup.241P, .sup.242P, .sup.243P, .sup.244P, .sup.245P, .sup.246P,.sup.247P, .sup.124I, .sup.125I, .sup.127I, .sup.131I, .sup.90Y,.sup.166Ho, .sup.186Re, .sup.188Re, .sup.90Sr, .sup.226Ra,.sup.103Pd, .sup.198Au, .sup.99Tc, .sup.18F, .sup.201Th, .sup.67Ga,and .sup.111In. Exposure to the radioactive agents can result incarcinogenesis, sterilization, cataract formation, radiodermatitis,beta burns, gamma burns, loss of cells (in particular bone marrow,digestive tract cells), damage to the hematopoietic,gastrointestinal, central nervous, cardiovascular, skin, and/orreproductive systems, acute radiation syndrome, chronic radiationsyndrome, and cutaneous radiation syndrome. Acute radiationsyndrome generally results from large doses of radiation to asubject's body occurring in a short period of time. The syndromehas a predictable course starting with a feeling of nausea,vomiting, general illness and fatigue, immune system depression,loss of hair, uncontrollable bleeding (mouth, under the skin,kidneys), massive diarrhea, delirium, coma and death. Cutaneousradiation syndrome is a subset of acute radiation syndrome andrefers to radiations effects on the skin, which include, but arenot limited to, inflammation, erythema, dry or moist desquamation,hair loss, blistering, reddening, ulceration, damage to sebaceousand sweat glands, atrophy, fibrosis, decreased or increased skinpigmentation, and necrosis.

[0069] The terms "systemic administration" and "systemicallyadministered" as used herein refer to a method of administering acompound or a pharmaceutical composition to a subject so that thecompound or pharmaceutical composition is delivered to sites in thebody, including the targeted site of pharmaceutical action, via thecirculatory system. Systemic administration includes, but is notlimited to, oral, intranasal, rectal and parenteral (e.g., otherthan through the alimentary tract, such as intramuscular,intravenous, intra-arterial, transdermal and subcutaneous)administration.

[0070] The term "therapeutically effective amount" of a compound asused herein refers to a sufficient amount of the compound to treata disease or disorder, at a reasonable benefit/risk ratioapplicable to any medical treatment. It will be understood,however, that the total daily usage of the compound will be decidedby the attending physician within the scope of sound medicaljudgment. The specific therapeutically effective dose level for anyparticular patient will depend upon a variety of factors includingthe disorder being treated and the severity of the disorder;activity of the specific antibody employed; the specificcomposition employed, the age, body weight, general health, sex anddiet of the patient; the time of administration, route ofadministration, and rate of excretion of the specific antibodyemployed; the duration of the treatment; drugs used in combinationor coincidental with the specific antibody employed; and likefactors well known in the medical arts. For example, it is wellknown within the skill of the art to start doses of the compound atlevels lower than those required to achieve the desired therapeuticeffect and to gradually increase the dosage until the desiredeffect is achieved.

[0071] The terms "tissue damage" and "tissue injury" are usedherein interchangeably, which refer to any damage of a tissue thatdisrupts its physical structure resulting in the impairment of itsfunction. For example, tissue injury may be caused by any form ofchemical or physical agents, such as, drugs, environmentaltoxicants, or any other substance that contacts a subject andresults directly or indirectly, in damage to the cells of the organor tissue. Also included, is cellular damage that results fromsuccessful therapeutic treatment of a subject, such as for example,the treatment of a tumor which results in induction of apoptosis.Similarly, tissue injury might be the result of a physical agentsuch as, for example, exposure to an environmental condition suchas a hypoxic condition or air or water pollution. Alternatively,the physical agent might be a physical trauma event, whetherself-imposed or not, such as for example, exercise, smoking, bluntforce trauma, stroke, etc.

[0072] The term "tissue protective activity" or "tissue protection"as used herein refers to the effect of inhibiting or delayingdamage or death of a cell, tissue, or organ. Unless otherwisenoted, the "delay" in damage or death of a cell, tissue or organ isevaluated relative to a control condition in the absence of acompound of the invention. Tissue protective activity is specificto tissue, cells, and/or organs expressing a tissue protectivereceptor complex (i.e., a responsive tissue cell, and/or organ,respectively), such as, but not limited to, the tissues of thecentral nervous system. In specific embodiments, the responsivecells are not erythrocyte progenitor cells.

[0073] The term "toxic agent" as used herein refers to a chemicalagent or a radiation agent disclosed herein. The term "chemicalagent" as used herein refers a chemical substance that isadministered to a subject for therapeutical purposes or used aschemical weapon to cause severe injuries or harm to the subject.Therapeutical chemical agent includes many pharmaceutical compoundsthat may also cause toxicity to the subject (i.e., side effects).The chemical agents used as weapons can be classified by theirmethod of action such as: blood agents, blister agents, nerveagents, pulmonary agents, and incapacitating agents.

[0074] The term "treatment," "treat," or "treating" as used hereinrefers to clinical intervention in an attempt to alter the naturalcourse of the individual being treated, and can be performed eitherfor prophylaxis or during the course of clinical pathology. Forpurposes of this invention, beneficial or desired clinical resultsinclude, but are not limited to, alleviation of symptoms;diminishment of extent of disorder or disease; stabilized (i.e. notworsening) state of disorder or disease; delay or slowing ofdisorder, or disease progression; amelioration of the disorder ordisease state, remission (whether partial or total), whetherdetectable or undetectable; or enhancement or improvement of thedisorder or disease. Treatment includes eliciting a cellularresponse that is clinically significant, without excessive levelsof side effects. Treatment also includes prolonging survival ascompared to expected survival if not receiving treatment.

DETAILED DESCRIPTION OF PREFERRED EMBODIMENTS

[0075] The description that follows includes systems, methods,techniques, instruction sequences, and computing machine programproducts that embody illustrative embodiments of the presentinvention. In the following description, for purposes ofexplanation, numerous specific details are set forth in order toprovide an understanding of various embodiments, it will beevident, however, to those skilled in the art that embodiments ofthe inventive subject matter may be practiced without thesespecific details. In general, well-known instruction instances,protocols, structures, and techniques have not been shown indetail.

[0076] Additionally, the terminology used herein is for the purposeof description and not for limitation. Furthermore, althoughcertain methods are described with reference to steps that arepresented herein in a certain order, in many instances, these stepscan be performed in any order as may be appreciated by one skilledin the art; the novel method is therefore not limited to theparticular arrangement of steps disclosed herein. It is to beunderstood that each component, compound, substituent, or parameterdisclosed herein is to be interpreted as being disclosed for usealone or in combination with one or more of each and every othercomponent, compound, substituent, or parameter disclosedherein.

[0077] It must be noted that as used herein and in the appendedclaims, the singular forms "a", "an", and "the" include pluralreferences unless the context clearly dictates otherwise.Furthermore, the terms "a" (or "an"), "one or more", and "at leastone" can be used interchangeably herein. The terms "comprising","including", "having" and "constructed from" can also be usedinterchangeably.

[0078] Unless otherwise indicated, all numbers expressingquantities of ingredients, properties such as molecular weight,percent, ratio, reaction conditions, and so forth used in thespecification and claims are to be understood as being modified inall instances by the term "about," whether or not the term "about"is present. Accordingly, unless indicated to the contrary, thenumerical parameters set forth in the specification and claims areapproximations that may vary depending upon the desired propertiessought to be obtained by the present disclosure.

[0079] The present invention provides a method for preventing ortreating a disease or disorder associated with tissue damages usingan inhibitor of MST1/2 protein kinases, its prodrugs, or apharmaceutical composition comprising the inhibitor of MST1/2protein kinases.

[0080] The inhibitors of MST1/2 protein kinases, their prodrugs andpharmaceutical compositions, are described previously in PCTpublication WO 2017/148406 A1, which are incorporated herein byreference in its entirety. Briefly, the inhibotrs of MST1/2 proteinkinases may be represented by the following general formula:

##STR00005##

wherein R.sub.1 is selected: 1) C1-C6 alkyl, optionally substitutedby halogen, nitro, cyano; C1-C6 alkyl group containing oxygen;C3-C7 cycloalkyl, which is optionally substituted by halogen,nitro, cyano; --O--C1-C6 alkyl, optionally substituted by halogen,nitro, amino, cyano; --O-C3-C7 cycloalkyl, which is optionallysubstituted with halogen, nitro, amino, cyano; C6-C10 aryl, whichis optionally substituted by halogen, nitro, amino, cyano;--O-C6-C10 aryl, which is optionally substituted by halogen, nitro,amino, cyano; C2-C6 alkenyl group; 2) 3-N,N-dimethylamino-propenyl, 3-pyrrolidin-propenyl; 3) amino,cyclopropylamino, cyclobutylamino, cyclopentylamino,cyclohexylamino, N, N-dimethylamino, N, N-diethylamino, N,N-diisopropylamino, 2-N, N-dimethyl-ehthylamino,2-hydroxyethylamino, 2-morpholinyl-ethylamino, 2-thiomorpholinylethylamino, 2-(4-N-methyl piperazinyl) ethylamino, 3-N,N-dimethyl-aminopropyl amino, 3-N, N-diethyl aminopropyl amino,3-N, N-diisopropyl-aminopropyl amino, amino-3-hydroxylpropyl,3-morpholinyl-propylamino, 3-thiomorpholinyl propylamino,3-(4-N-methylpiperazinyl) propylamino, N-methylpiperidinyl-4-amino,N-ethylpiperidiny-4-amino, N-isopropyl-piperidinyl-4-amino,N-acetyl-piperidinyl-4-amino; 4) hydroxyl,2-N,N-dimethylaminoethoxyl, 2-N, N-diethyl-aminoethoxyl, 2-N,N-diisopropyl-aminoethoxyl, 2-(N-methylpiperazinyl) ethoxyl,2-(N-acetyl-piperazinyl) ethoxyl, 2-morpholinyl-ethoxyl,2-thiomorpholinyl ethoxyl, 2-piperidinyl-ethoxyl, 3-N,N-dimethylamino-propoxyl, 3-N, N-diethylamino-propoxyl, 3-N,N-diisopropylamino propoxyl, 3-(N-methylpiperazinyl) propoxyl,3-(N-acetyl-piperazinyl) propoxyl, 3-morpholinyl-propoxyl,3-thiomorpholinyl propoxyl, 3-piperidinyl-propoxyl,2-pyridyl-methoxyl, 3-pyridyl-methoxyl, 4-pyridyl methoxyl,phenylmethoxyl, monohalogen-substituted phenylmethoxyl,hom*odihalogen-substituted phenylmethoxyl,heterodihalogen-substituted phenylmethoxyl; 5) selected from thegroup of five- or six-membered heterocyclic rings comprising one ora more of N, S and O heteroatoms, said five- or six-memberedheterocyclic rings are optionally substituted with C1-C6 alkyl,C1-C6 alkoxy, hydroxy, amino, C1-C6 acyl, cyano, or heterocyclicgroup, including but not limited to: piperidinyl, 4-N,N-dimethylamino-piperidinyl, 4-N, N-diethylamino-piperidinyl, 4-N,N-diisopropylamino piperidinyl, 4-hydroxypiperidinyl,4-(N-methylpiperazinyl) piperidinyl, 4-(N-ethyl-piperazinyl)piperidinyl, 4-(N-isopropyl-piperazinyl) piperidinyl,4-(N-acetyl-piperazinyl) piperidinyl, 4-(N-tert-butoxylformyl-piperazinyl) piperidinyl, 4-(N-methylsulfonyl-piperazinyl)piperidinyl, 4-(N-(2-hydroxylethyl) piperazinyl) piperidinyl,4-(N-(2-cyanoethyl) piperazinyl) piperidinyl,4-(N-(3-hydroxylpropyl) piperazinyl) piperidinyl, 4-(N-(2-N,N-dimethyl-ethyl) piperazinyl) piperidinyl, 4-(N-(2-N,N-diethyl-ethyl) piperazinyl) piperidinyl, 4-(N-(3-N,N-dimethyl-propyl) piperazinyl) piperidinyl,4-(N-(3-N,N-diethyl-propyl) piperazinyl) piperidinyl,4-(pyrrolidinyl) piperidinyl, 4-(3-N, N-dimethyl-pyrrolidinyl)piperidinyl; N-methyl-piperazinyl, N-ethyl-piperazinyl,N-isopropyl-piperazinyl, N-acetyl-piperazinyl, N-tert formylpiperazinyl, N-methylsulfonyl-piperazinyl piperazinyl,N-(2-hydroxylethyl) piperazinyl, N-(2-cyanoethyl) piperazinyl,N-(3-hydroxylpropyl) piperazinyl, N-(2-N, N-dimethylethyl)piperazinyl, N-(2-N, N-diethyl-ethyl) piperazinyl, N-(3-N,N-dimethyl-propyl) piperazinyl, N-(3-N, N-diethyl-propyl)piperazinyl, 2-oxo-piperazin-4-yl, N--(N-methyl-4-piperidinyl)piperazinyl, N--(N-ethyl-4-piperidinyl) piperazinyl,N--(N-acetyl-4-piperidinyl) piperazinyl; morpholinyl, 3,5-dimethylmorpholinyl, thiomorpholinyl, tetrahydropyrrolyl, 3-N,N-dimethyl-tetrahydropyrrolyl, 3-N, N-diethyl-tetrahydropyrrolyl;R.sub.2 is selected from: 1) a hydrogen, halo, nitro, amino, cyano;2) C1-C6 alkyl, optionally substituted by halogen, nitro, amino,cyano; --O-C1-C6 alkyl, optionally substituted by halogen, nitro,amino, cyano; a C1-C6 oxygen-containing alkyl; 3) methylthio,ethylthio, isopropylthio, methylsulfinyl, ethyl sulfinyl, propylsulfinyl, methylsulfonyl, ethylsulfonyl, isopropylsulfonyl, aminosulfonyl, ethylamino sulfonyl, propylamino sulfonyl, isopropylaminosulfonyl, cyclopropylamino sulfonyl, hydroxyl formyl, methoxylformyl, ethoxyl formyl, propoxyl formyl, isopropoxyl formyl,n-butoxyl formyl, isobutoxyl formyl, t-butoxyl formyl, aminoformyl, methylamino formyl, ethylamino formyl, propylamino formyl,isopropylamino formyl, cyclopropylamino formyl, cyclobutylaminoformyl, cyclopentylamino formyl, acetamido, propionamido, n-butylamido, isobutyl amido, cyclopropyl formamido, cyclobutyl formamido,cyclopentyl formamido, methylsulfonamino, ethylsulfonamino,propylsulfonamido, isopropylsulfonamido, dimethyl phosphinyl,diethyl phosphinyl, diisopropyl phosphinyl; R.sub.3 is selectedfrom: hydrogen; C1-C6 alkyl, optionally substituted by halogen,nitro, amino, cyano; C3-C7 cycloalkyl, which is optionallysubstituted with halogen, nitro, amino, cyano; alternatively,R.sub.3 may form a five-member ring with its connected N atom and Catom in the S1 ring; R.sub.4 is selected from: hydrogen; C1-C6alkyl, optionally substituted by halogen, nitro, amino, cyano;C3-C7 cycloalkyl, which is optionally substituted with halogen,nitro, amino, cyano; R.sub.5 is selected from: 1) a hydrogen, halo,nitro, amino, cyano; 2) C1-C6 alkyl, optionally substituted byhalogen, nitro, amino, cyano; --O-C1-C6 alkyl, optionallysubstituted by halogen, nitro, amino, cyano; a C1-C6oxygen-containing alkyl; 3) methylthio, ethylthio, isopropylthio,methylsulfinyl, ethyl sulfinyl, propyl sulfinyl, methylsulfonyl,ethylsulfonyl, isopropylsulfonyl, amino sulfonyl, ethylaminosulfonyl, propylamino sulfonyl, isopropylamino sulfonyl,cyclopropylamino sulfonyl, hydroxyl formyl, methoxyl formyl,ethoxyl formyl, propoxyl formyl, isopropoxyl formyl, n-butoxylformyl, isobutoxyl formyl, t-butoxyl formyl, amino formyl,methylamino formyl, ethylamino formyl, propylamino formyl,isopropylamino formyl, cyclopropylamino formyl, cyclobutylaminoformyl, cyclopentylamino formyl, acetamido, propionamido, n-butylamido, isobutyl amido, cyclopropyl formamido, cyclobutyl formamido,cyclopentyl formamido, methylsulfonamido, ethylsulfonamido,propylsulfonamido, isopropylsulfonamido, dimethyl phosphinyl,diethyl phosphinyl, diisopropyl phosphinyl; X=O, NH or a directbond; Y=S, C, P, N, OH, NH.sub.2 or CH.sub.2; m=0, 1 or 2; n=0, 1,2, 3 or 4;

##STR00006##

is aryl or heteroaryl group fused with a seven memberedtwo-nitrogen heterocyclic ring, such as benzene ring, a thiophenering, a furan ring, a pyridine ring, an oxazole ring, or thiazolylring fused with a seven membered two nitrogen heterocyclic ringgroup;

##STR00007##

is aryl or heteroaryl, such as a benzene ring or a pyrazole ring;or a stereoisomer of the above compounds, a prodrug thereof, apharmaceutically acceptable salt thereof, or a pharmaceuticallyacceptable solvate thereof.

[0081] A second aspect, the inhibitors of MST1/2 protein kinaseshave the following formulas I, II, III, IV:

##STR00008##

wherein: n1 is selected from 0, 1, 2, 3 or 4; R.sub.11 is selectedfrom: 1) C1-C6 alkyl, optionally substituted with halogen, amino,nitro, cyano; C1-C6 alkyl containing oxygen; C3-C7 cycloalkyl,which is optionally substituted with halogen, amino, nitro, cyano;C6-C10 aryl, optionally substituted by halogen, nitro, amino,hydroxy, cyano; C3-C6 alkenyl; 2) 2-N, N-dimethylaminoethyl,2-hydroxyethyl, 2-N, N-diethylaminoethyl, 2-N, N-diisopropylaminoethyl, 2-morpholinyl ethyl, 2-thiomorpholinyl ethyl,2-(4-N-piperazinyl-methyl) ethyl, 3-N, N-dimethylaminopropyl, 3-N,N-diethylaminopropyl, 3-N, N-diisopropyl-aminopropyl, 3-morpholinylpropyl, 3-thiomorpholinyl propyl, 3-(4-N-methylpiperidinyl) propyl,4-N, N-dimethylamino-cyclohexyl, 4-N, N-diethylamino cyclohexyl,N-methyl-4-piperidinyl, N-ethyl-4-piperidinyl,N-isopropyl-4-piperidinyl, 1,3-dimethyl-5-pyrazolyl,1-methyl-4-pyrazolyl, 3-methyl-5-isoxazolinyl,1-(N-methyl-4-piperidinyl)-4-pyrazolyl, 1-(N-tert-butoxylformyl-4-piperidinyl)-4-pyrazolyl; 3)

##STR00009##

wherein Z.sub.1, Z.sub.2, Z.sub.3, Z.sub.4, Z.sub.5 are eachindependently selected from: (1) hydrogen, halogen, nitro, amino,hydroxy, cyano, (2) C1-C6 alkyl, C1-C6 alkoxy, C1-C6 alkylcontaining oxygen, C1-C6 alkyl containing fluorine, C1-C6 alkoxycontaining fluorine, 4-piperidinyl, N-methyl yl-4-piperidinyl, (3)N, N-dimethylamino, N, N-diethylamino, N, N-diisopropylamino, 2-N,N-dimethylaminoethylamino, 2-morpholino ethylamino, 2-ethylaminothiomorpholinyl, 2-(4-N-methylpiperazinyl) ethylamino, 3-N,N-dimethyl-aminopropyl amino, 3-N, N-diethylaminopropyl-amino, 3-N,N-diisopropylamino propylamino, 3-morpholin-propylamino,3-thiomorpholinyl propylamino, 3-(4 N-methylpiperazinyl)propylamino, N-methylpiperidinyl-4-amino,N-ethylpiperidinyl-4-amino, N-isopropyl-piperidinyl-4-amino, (4)2-N, N-dimethylaminoethoxyl, 2-N, N-diethyl-aminoethoxy, 2-N,N-diisopropyl-aminoethoxyl, 2-(N-methylpiperazinyl) ethoxyl,2-(N-acetyl-piperazinyl) ethoxyl, 2-morpholino-ethoxyl,2-thiomorpholino-ethoxyl, 2-piperidinyl ethoxyl, 3-N,N-dimethylamino-propoxyl, 3-N, N-diethylamino-propoxyl, 3-N,N-diisopropylamino propoxyl, 3-(N-methylpiperazinyl) propoxyl,3-(N-acetyl-piperazinyl) propoxyl, 3-morpholinyl-propoxyl,3-thiomorpholinyl propoxyl, 3-piperidinyl-propoxyl,2-pyridyl-methoxyl, 3-pyridyl-methoxyl, 4-pyridyl methoxyl,phenylmethoxyl, monohalogen-substituted phenylmethoxyl,hom*odihalogen-substituted phenylmethoxyl,heterodihalogen-substituted phenylmethoxyl, (5) piperidinyl, 4-N,N-dimethylamino-piperidinyl, 4-N, N-diethylamino-piperidinyl, 4-N,N-diisopropylamino piperidinyl, 4-hydroxy piperidinyl, morpholinyl,3,5-dimethyl morpholinyl, thiomorpholinyl, tetrahydropyrrolyl, 3-N,N-dimethyl-tetrahydropyrrolyl, 3-N, N-diethyl-tetrahydropyrrolyl,N-methyl-piperazinyl, N-ethyl-piperazinyl, N-isopropyl-piperazinyl,N-acetyl-piperazinyl, N-tert-butoxyl formyl piperazinyl,N-methylsulfonyl-piperazinyl, N-(2-hydroxylethyl) piperazinyl,N-(2-cyanoethyl) piperazinyl, N-(3-hydroxylpropyl) piperazinyl,N-(2-N, N-dimethylethyl) piperazinyl, N-(2-N, N-diethyl-ethyl)piperazinyl, N-(3-N, N-dimethylpropyl) piperazinyl, N-(3-N,N-diethyl-propyl) piperazinyl, 2-oxo-piperazinyl,2-oxo-piperazin-4-yl, imidazolyl, 4-imidazolyl, (6)4-(N-methylpiperazinyl) piperidinyl, 4-(N-ethyl-piperazinyl)piperidinyl, 4-(N-isopropyl-piperazinyl) piperidinyl,4-(N-acetyl-piperazinyl) piperidinyl,4-(N-t-butoxyl-formyl-piperazinyl) piperidinyl,4-(N-methylsulfonyl-piperazinyl) piperidinyl,4-(N-(2-hydroxylethyl) piperazinyl) piperidinyl,4-(N-(2-cyanoethyl) piperazinyl) piperidinyl,4-(N-(3-hydroxylpropyl) piperazinyl) piperidinyl, 4-(N-(2-N,N-dimethyl-ethyl) piperazinyl) piperidinyl, 4-(N-(2-N, N-diethylethyl) piperazinyl) piperidinyl, 4-(N-(3-N, N-dimethyl-propyl)piperazinyl) piperidinyl, 4-(N-(3-N, N-diethyl-propyl) piperazinyl)piperidinyl, 4-(tetrahydropyrrolyl) piperidinyl, 4-(3-N,N-dimethyl-tetrahydropyrrolyl) piperidinyl,N--(N-methyl-4-piperidinyl) piperazinyl, N--(N-ethyl-4-piperidinyl)piperazinyl, (7) hydroxy sulfonyl, aminosulfonyl, sulfonylmethylamino, ethylamino sulfonyl group, a sulfonyl grouppropylamino, isopropylamino-sulfonyl, aminosulfonyl cyclopropyl,cyclobutyl aminosulfonyl, cyclopentyl aminosulfonyl,piperidinyl-sulfonyl, 4-hydroxyl-piperidinyl-1-sulfonyl, 4-N,N-dimethyl-piperidinyl-1-sulfonyl, 4-N,N-diethyl-piperidinyl-1-sulfonyl, pyrrolidinyl-1-sulfonyl, 3-N,N-dimethyl-pyrrolidinyl-1-sulfonyl, 3-N,N-diethyl-pyrrolidinyl-1-sulfonyl, N-methyl-piperazinyl-sulfonyl,N-ethylpiperazinyl-1-sulfonyl, N-acetyl-piperazinyl-1-sulfonyl,N-tert-butoxylformyl-piperazinyl-1-sulfonyl, N-(2-hydroxylethyl)piperazinyl-1-sulfonyl, N-(2-cyanoethyl) piperazinyl-1-sulfonyl,N-(2-N, N-dimethyl ethyl) piperazinyl-1-sulfonyl, N-(2-N,N-diethyl-ethyl) piperazinyl-1-sulfonyl, N-(3-hydroxylpropyl)piperazinyl-1-sulfonyl, N-(3-N, N-dimethylamino-propyl)piperazinyl-1-sulfonyl, N-(3-N, N-diethylamino-propyl)piperazinyl-1-sulfonyl, morpholinyl-1-sulfonyl,3,5-dimethyl-morpholinyl-1-sulfonyl, 4-(N-methyl-1-piperazinyl)piperidinyl-1-sulfonyl, 4-(N-ethyl-1-piperazinyl)piperidinyl-1-sulfonyl, 4-(N-acetyl-1-piperazinyl)piperidinyl-sulfonyl, N--(N-methyl-4-piperidinyl)piperazinyl-1-sulfonyl, (8) amino formyl, methylamino formyl,ethylamino formyl, propylamino formyl, isopropylamino formyl,cyclopropylamino formyl, cyclobutylamino formyl, cyclopentylaminoformyl, piperidinyl-1-formyl, 4-hydroxy-piperidinyl-1-formyl, 4-N,N-dimethyl-piperidinyl-1-formyl, 4-N, N-twoethylpiperidinyl-1-formyl, tetrahydropyrrolyl-1-formyl, 3-N,N-dimethyl-tetrahydropyrrolyl-1-formyl, 3-N,N-diethyl-tetrahydropyrrolyl-1-formyl,N-methyl-piperazinyl-1-formyl, N-ethyl-piperazinyl-1-formyl,N-acetyl-piperazinyl-1-formyl,N-tert-butoxyl-formyl-piperazinyl-1-formyl, N-(2-hydroxyethyl)piperazinyl-1-formyl, N-(2-cyanoethyl) piperazinyl-1-formyl,N-(2-N, N-dimethyl-ethyl) piperazinyl-1-formyl, N-(2-N,N-diethyl-ethyl) piperazinyl-1-formyl, N-(3-hydroxypropyl)piperazinyl-1-formyl, N-(3-N, N-dimethyl-propyl)piperazinyl-1-formyl, N-(3-N, N-diethyl propyl)piperazinyl-1-formyl, morpholinyl-1-formyl,3,5-dimethyl-morpholinyl-1-formyl, 4-(N-methyl-1-piperazinyl)piperidinyl-1-formyl, 4-(N-ethyl-1-piperazinyl)piperidinyl-1-formyl, 4-(N-acetyl-1-piperazinyl)piperidinyl-1-formyl, N--(N-methyl-4-piperidinyl)piperazinyl-1-formyl, (9) hydroxyl formyl, methoxyl formyl, ethoxylformyl, propoxyl formyl, isopropoxyl formyl, n-butoxyl formyl,isobutoxy formyl, t-butoxyl formyl, (10) amino formamido,methylamino formamido, ethylamino formamido, propylamino formamido,isopropylamino formamido, cyclopropylamino formamido,cyclobutylamino formamido, cyclopentylamino formamido,piperidinyl-1-formamido, 4-hydroxy-piperidinyl-1-formamido, 4-N,N-dimethyl-piperidinyl-1-formamido, 4-N,N-diethyl-piperidinyl-1-formamido, tetrahydropyrrolyl-1-formamido,3-N, N-dimethyl-tetrahydropyrrolyl-1-formamido, 3-N,N-diethyl-tetrahydropyrrolyl-1-formamido,N-methyl-piperazinyl-1-formamido, N-ethyl-piperazinyl-1-formamido,N-acetyl-piperazinyl-1-formamido, N-tert-butoxylformyl-piperazinyl-1-formamido, N-(2-hydroxyethyl)piperazinyl-1-formamido, N-(2-cyanoethyl) piperazinyl-1-formamido,N-(2-N, N-dimethyl-ethyl) piperazinyl-1-formamido, N-(2-N,N-diethyl-ethyl) piperazinyl-1-formamido, N-(3-hydroxypropyl)piperazinyl-1-formamido, N-(3-N, N-dimethyl-propyl)piperazinyl-1-formamido, N-(3-N, N-diethyl-aminopropyl)piperazinyl-1-formamido, morpholinyl-1-formamido,3,5-dimethyl-morpholinyl-1-formamido, 4-(N-methyl-1-piperazinyl)piperidinyl-1-formamido, 4-(N-ethyl-1-piperazinyl)piperidinyl-1-formamido, 4-(N-acetyl-1-piperazinyl)piperidinyl-1-formamido, N--(N-methyl-4-piperidinyl)piperazinyl-1-formamido; or (11) amino acetamido, N-tert-butoxylformyl acetamido, N-acetylamino acetamido, acrylamido,cyclopropylamido, chloroacetamido, bromoacetamido, piperidinylacetamido, 4-hydroxy piperidinyl acetamido, 4-N,N-dimethyl-piperidinyl-acetamido, 4-N, N-diethyl-piperidinylacetamido, tetrahydropyrrolyl acetamido, 3-N,N-dimethyl-tetrahydropyrrolyl acetamido, 3-N,N-diethyl-tetrahydropyrrolyl-acetamido, N-methyl-piperazinylacetamido, N-ethyl piperazinyl-acetamido, N-acetyl-piperazinylacetamido, N-tert-butoxy formyl-piperazinyl acetamido,N-(2-hydroxyethyl) piperazinyl acetamido, N-(2-cyanoethyl)piperazinyl acetamido, N-(2-N, N-dimethylethyl) piperazinylacetamido, N-(2-N, N-diethyl-ethyl) piperazinyl acetamido,N-(3-hydroxylpropyl) piperazinyl acetamido, N-(3-N,N-dimethyl-propyl) piperazinyl acetamido, N-(3-N, N-diethyl-propyl)piperazinyl acetamido, morpholinyl acetamido,3,5-dimethyl-morpholinyl-acetamido, 4-(N-methyl-1-piperazinyl)piperidinyl acetamido, 4-(N-ethyl-1-piperazinyl) piperidinylacetamido, 4-(N-acetyl-1-piperazinyl) piperidinyl acetamido,N--(N-methyl-4-piperidinyl) piperazinyl acetamido,4-(tetrahydropyrrolyl) piperidinyl acetamido; 2-methylaminoacetamido, 2-(1-methylethyl) amino acetamido;N-benzyloxy-formyl-2-methylamino-acetamido; (12) Z.sub.2 andZ.sub.3 may form a substituted or unsubstituted oxygen-containingfive- or six-membered ring; the substituents may be selected fromthe same substituents of Z.sub.1, (13) Z.sub.2 and Z.sub.3 may forma substituted or unsubstituted nitrogen-containing five- orsix-membered ring; the substituents may be selected from the samesubstituents of Z.sub.1, 4)

##STR00010##

wherein Z.sub.2, Z.sub.3, Z.sub.4, Z.sub.5 are the same as thedefinition 3) above; 5)

##STR00011##

wherein Z.sub.1, Z.sub.3, Z.sub.4, Z.sub.5 are the same as thedefinition 3) above; R.sub.21 is selected from: 1) a hydrogen,halo, nitro, amino, cyano; 2) C1-C6 alkyl, optionally substitutedby halogen, nitro, amino, cyano; --O-C1-C6 alkyl, optionallysubstituted by halogen, nitro, amino, cyano; a C1-C6oxygen-containing alkyl; 3) methylthio, ethylthio, isopropylthio,methylsulfinyl, ethylsulfinyl, propyl sulfinyl, methylsulfonyl,ethylsulfonyl, isopropylsulfonyl, amino sulfonyl, ethylaminosulfonyl, propylamino sulfonyl, isopropylamino-sulfonyl,cyclopropylamino sulfonyl, hydroxyl formyl, methoxyl formyl,ethoxyl formyl, propoxyl formyl, isopropoxyl formyl, n-butoxylformyl, isobutoxyl formyl, t-butoxyl formyl, amino formyl,methylamino formyl, ethylamino formyl, propylamino formyl,isopropylamino formyl, cyclopropylamino formyl, cyclobutylaminoformyl, cyclopentylamino formyl, acetamido, propionamido, n-butylamido, isobutyl amido, cyclopropyl formamido, cyclobutyl formamido,cyclopentyl formamido, methylsulfonamido, ethylsulfonamido,propylsulfonamido, isopropylsulfonamido, dimethyl phosphinyl,diethyl phosphinyl, diisopropyl phosphinyl; R.sub.31 is selectedfrom: Hydrogen; C1-C6 alkyl, optionally substituted by halogen,nitro, amino, cyano; C3-C7 cycloalkyl, which is optionallysubstituted by halogen, nitro, amino, cyano; R.sub.11 is selectedfrom: hydrogen; C1-C6 alkyl, optionally substituted by halogen,nitro, amino, cyano; C3-C7 cycloalkyl, which is optionallysubstituted by halogen, nitro, amino, cyano; R.sub.51 is selectedfrom: 1) a hydrogen, halo, nitro, amino, cyano; 2) C1-C6 alkyl,optionally substituted by halogen, nitro, amino, cyano; --O-C1-C6alkyl, optionally substituted by halogen, nitro, amino, cyano; aC1-C6 oxygen-containing alkyl; 3) methylthio, ethylthio,isopropylthio, methylsulfinyl, ethyl sulfinyl, propyl sulfinyl,methylsulfonyl, ethylsulfonyl, isopropylsulfonyl, amino sulfonyl,ethylamino sulfonyl, propylamino sulfonyl, isopropylamino-sulfonyl,cyclopropylamino sulfonyl, hydroxyl formyl, methoxyl formyl,ethoxyl formyl, propoxyl formyl, isopropoxyl formyl, n-butoxylformyl, isobutoxyl formyl, t-butoxyl formyl, amino formyl,methylamino formyl, ethylamino formyl, propylamino formyl,isopropylamino formyl, cyclopropylamino formyl, cyclobutylaminoformyl, cyclopentylamino formyl, acetamido, propionamido, n-butylamido, isobutyl amido, cyclopropyl formamido, cyclobutyl formamido,cyclopentyl formamido, methylsulfonamido, ethylsulfonamido,propylsulfonamido, isopropylsulfonamido, dimethyl phosphinyl,diethyl phosphinyl, diisopropyl phosphinyl; or a stereoisomer ofthe above compounds, a prodrug thereof, a pharmaceuticallyacceptable salt thereof, or a pharmaceutically acceptable solvatethereof.

[0082] A third aspect, the inhibitors of MST1/2 protein kinases arerepresented by the following structural formula:

##STR00012##

wherein m2 is selected from 0, 1, 2, 3 or 4; n2 is selected from 0,1, 2, 3 or 4; R.sub.12 is selected from: 1) selected from the groupof five membered or six-membered heterocyclic rings comprising oneor more N, O and S heteroatoms, the five membered or six-memberedheterocyclic rings are optionally substituted with C1-C6 alkyl,C1-C6 alkoxy, hydroxy, amino, alkylamino, dialkylamino, C1-C6 acyl,cyano, optionally substituted C1-C6 alkyl, --O-C1-C6 alkyl,hydroxy, hydroxy C1-C6 alkyl, C1-C6 acyl, alkylamino, dialkylaminosubstituted heterocyclic group, including but not limited to: 4-N,N-dimethylamino-piperidinyl, 4-N, N-diethylamino-piperidinyl, 4-N,N-diisopropylamino-piperidinyl, 4-hydroxy-piperidinyl,4-(N-methylpiperazinyl) piperidinyl, 4-(N-ethyl-piperazinyl)piperidinyl, 4-(N-isopropyl-piperazinyl) piperidinyl,4-(N-acetyl-piperazinyl) piperidinyl, 4-(N-tert-butoxylformyl-piperazinyl) piperidinyl, 4-(N-methylsulfonyl-piperazinyl)piperidinyl, 4-(N-(2-hydroxylethyl) piperazinyl) piperidinyl,4-(N-(2-cyanoethyl) piperazinyl) piperidinyl,4-(N-(3-hydroxylpropyl) piperazinyl) piperidinyl, 4-(N-(2-N,N-dimethyl-ethyl) piperazinyl) piperidinyl, 4-(N-(2-N,N-diethyl-ethyl) piperazinyl) piperidinyl, 4-(N-(3-N,N-dimethyl-propyl) piperazinyl) piperidinyl, 4-(N-(3-N,N-diethyl-propyl) piperazinyl) piperidinyl, 4-(tetrahydropyrrolyl)piperidinyl, 4-(3-N, N-dimethyl-tetrahydropyrrolyl) piperidinyl; 2)amino, cyclopropylamino, cyclobutylamino, cyclopentylamino,cyclohexylamino, N, N-dimethylamino, N, N-diethylamino, N,N-diisopropylamino, 2-N, N-dimethylaminoethylamino,2-hydroxyethylamino, 2-morpholinyl-ethylamino,2-(4-N-methylpiperazinyl) ethylamino, 3-N,N-dimethylamino-propylamino, 3-N, N-diethylamino-propylamino, 3-N,N-diisopropylamino-propylamino, 3-hydroxy-propylamino,3-morpholinyl-propylamino, 3-(4-N-methylpiperazinyl) propylamino,N-methylpiperidinyl-4-amino, N-ethylpiperidinyl-4-amino,N-isopropyl piperidinyl-4-amino, N-acetyl piperidinyl-4-amino;N-methyl-piperazinyl, N-ethyl-piperazinyl, N-isopropyl-piperazinyl,N-acetyl piperazinyl, N-tert-butoxyl formyl-piperazinyl,N-methylsulfonyl-piperazinyl, N-(2-hydroxylethyl) piperazinyl,N-(2-cyanoethyl) piperazinyl, N-(3-hydroxylpropyl) piperazinyl,N-(2-N, N-dimethylethyl) piperazinyl, N-(2-N, N-diethyl aminoethyl)piperazinyl, N-(3-N, N-dimethyl-propyl) piperazinyl, N-(3-N,N-diethyl-propyl) piperazinyl, 2-oxo-piperazin-4-yl,N--(N-Methyl-4-piperidinyl) piperazinyl, N--(N-ethyl-4-piperidinyl)piperazinyl, N--(N-acetyl-4-piperidinyl) piperazinyl; morpholinyl,3,5-dimethyl morpholinyl, thiomorpholinyl, tetrahydropyrrolyl, 3-N,N-dimethyl-tetrahydropyrrolyl, 3-N, N-diethyl-tetrahydropyrrolyl;3) C1-C6 alkyl, optionally substituted by halogen, nitro, cyano; 4)C3-C7 cycloalkyl, which is optionally substituted by halogen,nitro, cyano; 5) --O--C1-C6 alkyl, optionally substituted byhalogen, nitro, amino, cyano; 6) --O--C3-C7 cycloalkyl, which isoptionally substituted by halogen, nitro, amino, cyano; 7) C6-C10aryl, which is optionally substituted by halogen, nitro, amino,cyano; --O-C6-C10 aryl group, which is optionally substituted byhalogen, nitro, amino, cyano; 8) C2-C6 alkenyl; 9) hydroxyl, 2-N,N-dimethylaminoethoxyl, 2-N, N-diethyl-aminoethoxyl, 2-N,N-diisopropyl-aminoethoxyl, 2-(N-methylpiperazine-yl) ethoxyl,2-(N-acetyl-piperazinyl) ethoxyl, 2-morpholinyl-ethoxyl,2-thiomorpholinyl ethoxyl, 2-piperidinyl-ethoxyl, 3-N,N-dimethylamino-propoxyl, 3-N, N-diethylamino-propoxyl, 3-N,N-diisopropylamino propoxyl, 3-(N-methylpiperazinyl) propoxyl,3-(N-acetyl-piperazinyl) propoxyl, 3-morpholinyl-propoxyl,3-thiomorpholinyl propoxyl, 3-piperidinyl-propoxyl,2-pyridyl-methoxyl, 3-pyridyl-methoxyl, 4-pyridyl methoxyl,phenylmethoxyl, monohalogen-substituted phenylmethoxyl,hom*odihalogen substituted phenyl methoxyl,heterodihalogen-substituted phenylmethoxyl; R.sub.22 is selectedfrom: 1) a hydrogen, halo, nitro, amino, cyano; 2) C1-C6 alkyl,optionally substituted by halogen, nitro, amino, cyano; --O-C1-C6alkyl, optionally substituted by halogen, nitro, amino, cyano;C1-C6 oxygen-containing alkyl; 3) methylthio, ethylthio,isopropylthio, methylsulfinyl, ethyl sulfinyl, propyl sulfinyl,methylsulfonyl, ethylsulfonyl, isopropylsulfonyl, amino sulfonyl,ethylamino sulfonyl, propylamino sulfonyl, isopropylamino-sulfonyl,cyclopropylamino sulfonyl, hydroxyl formyl, methoxyl formyl,ethoxyl formyl, propoxyl formyl, isopropoxyl formyl, n-butoxylformyl, isobutoxyl formyl, t-butoxyl formyl, amino formyl,methylamino formyl, ethylamino formyl, propylamino formyl,isopropylamino formyl, cyclopropylamino formyl, cyclobutylaminoformyl, cyclopentylamino formyl, acetamido, propionamido, n-butylamido, isobutyl amido, cyclopropyl formamido, cyclobutyl formamido,cyclopentyl formamido, methylsulfonamiso, ethylsulfonamiso,propylsulfonamiso, isopropylsulfonamiso, dimethyl phosphinyl,diethyl phosphinyl, diisopropyl phosphinyl; R.sub.32 is selectedfrom: hydrogen; C1-C6 alkyl, optionally substituted by halogen,nitro, amino, cyano; C3-C7 cycloalkyl, which is optionallysubstituted with halogen, nitro, amino, cyano; R.sub.42 is selectedfrom: hydrogen; C1-C6 alkyl, optionally substituted by halogen,nitro, amino, cyano; C3-C7 cycloalkyl, which is optionallysubstituted with halogen, nitro, amino, cyano; R.sub.52 is selectedfrom: 1) a hydrogen, halo, nitro, amino, cyano; 2) C1-C6 alkyl,optionally substituted by halogen, nitro, amino, cyano; --O-C1-C6alkyl, optionally substituted by halogen, nitro, amino, cyano;C1-C6 oxygen-containing alkyl; 3) methylthio, ethylthio,isopropylthio, methylsulfinyl, ethyl sulfinyl, propyl sulfinyl,methylsulfonyl, ethylsulfonyl, isopropylsulfonyl, amino sulfonyl,ethylamino sulfonyl, propylamino sulfonyl, isopropylamino-sulfonyl,cyclopropyl aminosulfonyl, hydroxyl formyl, methoxyl formyl,ethoxyl formyl, propoxyl formyl, isopropoxyl formyl, n-butoxylformyl, isobutoxyl formyl, t-butoxyl formyl, amino formyl,methylamino formyl, ethylamino formyl, propylamino formyl,isopropylamino formyl, cyclopropylamino formyl, cyclobutylaminoformyl, cyclopentylamino formyl, acetamido, propionamido, n-butylamido, isobutyl amido, cyclopropyl formamido, cyclobutyl formamido,cyclopentyl formamido, methylsulfonamido, ethylsulfonamido,propylsulfonamido, isopropylsulfonamido, dimethyl phosphinyl,diethyl phosphinyl, diisopropyl phosphinyl; Or a stereoisomer ofthe above compounds, a prodrug thereof, a pharmaceuticallyacceptable salt thereof, or a pharmaceutically acceptable solvatethereof.

[0083] A fourth aspect, the inhibitors of MST1/2 protein kinasesare represented by the following structural formula:

##STR00013##

wherein n3 is selected from 0, 1, 2, 3 or 4; R.sub.23 is--S--O.sub.2X, wherein X is selected from: hydroxy; C1-C6 alkyl,optionally substituted by halogen, nitro, amino, cyano; C3-C7cycloalkyl, which is optionally substituted with halogen, nitro,amino, cyano; R.sub.13 is selected from: hydrogen; C1-C6 alkyl,optionally substituted by halogen, nitro, amino, cyano; C3-C7cycloalkyl, which is optionally substituted with halogen, nitro,amino, cyano; R.sub.33 is selected from: 1) a hydrogen, halo,nitro, amino, cyano; 2) C1-C6 alkyl, optionally substituted byhalogen, nitro, amino, cyano; --O--C1-C6 alkyl, optionallysubstituted by halogen, nitro, amino, cyano; a C1-C6oxygen-containing alkyl; 3) methylthio, ethylthio, isopropylthio,methylsulfinyl, ethyl sulfinyl, propyl sulfinyl, methylsulfonyl,ethylsulfonyl, isopropylsulfonyl, amino sulfonyl, ethylaminosulfonyl, propylamino sulfonyl, isopropylamino-sulfonyl,cyclopropylamino sulfonyl, hydroxyl formyl, methoxyl formyl,ethoxyl formyl, propoxyl formyl, isopropoxyl formyl, n-butoxylformyl, isobutoxyl formyl, t-butoxyl formyl, amino formyl,methylamino formyl, ethylamino formyl, propylamino formyl,isopropylamino formyl, cyclopropyl aminoformyl, cyclobutylaminoformyl, cyclopentylaminoformyl, acetamido, propionamido,n-butyl amido, isobutyl amido, cyclopropyl formamido, cyclobutylformamido, cyclopentyl formamido, methylsulfonamido,ethylsulfonamido, propylsulfonamido, isopropylsulfonamido, dimethylphosphinyl, diethyl phosphinyl, diisopropyl phosphinyl; R.sub.53 isselected from: hydrogen; C1-C6 alkyl, optionally substituted byhalogen, nitro, amino, cyano; C3-C7 cycloalkyl, which is optionallysubstituted with halogen, nitro, amino, cyano; R.sub.43 is selectedfrom: hydrogen; C1-C6 alkyl, optionally substituted by halogen,nitro, amino, cyano; C3-C7 cycloalkyl, which is optionallysubstituted with halogen, nitro, amino, cyano; Or a stereoisomer ofthe above compounds, a prodrug thereof, a pharmaceuticallyacceptable salt thereof, or a pharmaceutically acceptable solvatethereof.

[0084] MST1/2 protein kinase inhibitors having Formula I areexemplified by the compounds in Table 1.

TABLE-US-00001 TABLE 1 MST1/2 protein kinase inhibitors havingFormula I ##STR00014## IA-1 (TFA Salt) ##STR00015## IA-2##STR00016## IA-3 ##STR00017## IA-4 (TFA Salt) ##STR00018## IA-5(HCl Salt) ##STR00019## IA-6 (HCl Salt) ##STR00020## IA-7 (HClSalt) ##STR00021## IB-1 (TFA Salt) ##STR00022## IB-2 (TFA Salt)##STR00023## IB-3 (TFA Salt) ##STR00024## IB-4 ##STR00025## IB-5##STR00026## IB-6 ##STR00027## IB-7 ##STR00028## IC-1 ##STR00029##IC-2 (TFA Salt) ##STR00030## IC-3 (TFA Salt) ##STR00031## IC-4 (TFASalt) ##STR00032## IC-5 (TFA Salt) ##STR00033## IC-6 ##STR00034##IC-7 ##STR00035## IC-8 ##STR00036## IC-9 ##STR00037## IC-10 (HClSalt) ##STR00038## IC-11 (TFA Salt) ##STR00039## ID-1 (HCl Salt)##STR00040## ID-2 (HCl Salt) ##STR00041## ID-3 (HCl Salt)##STR00042## IE-1 (HCl Salt) ##STR00043## IE-2 (TFA Salt)##STR00044## IE-3 (TFA Salt) ##STR00045## IF-1 ##STR00046## IF-2##STR00047## IF-3 ##STR00048## IF-4 (TFA Salt) ##STR00049## IF-5(TFA Salt) ##STR00050## IF-6 ##STR00051## IG-1 (HCl Salt)##STR00052## IG-2 (HCl Salt) ##STR00053## IG-3 ##STR00054## IH-1##STR00055## IH-2 (TFA Salt) ##STR00056## IH-3 (TFA Salt)##STR00057## IH-4 (TFA Salt) ##STR00058## IH-5 (TFA Salt)##STR00059## IH-6 (HCl Salt) ##STR00060## IH-7 (HCl Salt)##STR00061## IH-8 ##STR00062## IH-9 ##STR00063## IH-10 (HCl Salt)##STR00064## IH-11 (HCl Salt) ##STR00065## IH-12 (HCl Salt)##STR00066## IH-13 (HCl Salt) ##STR00067## IH-14 (HCl Salt)

[0085] A fifth aspect, the inhibitors of MST1/2 protein kinases arerepresented by the following structural formula:

##STR00068##

wherein n4 is selected from 0, 1 or 2; R.sub.14 is selected from:1) selected from the group of five-membered heterocyclic orsix-membered heterocyclic rings comprising one or more N, O and Sheteroatoms, the five-membered heterocyclic or six-memberedheterocyclic rings are optionally substituted with C1-C6 alkyl,C1-C6 alkoxy, hydroxy, amino, alkylamino, dialkylamino, C1-C6 acyl,cyano, optionally substituted C1-C6 alkyl, --O-C1-C6 alkyl,hydroxyl, hydroxyl C1-C6 alkyl, C1-C6 acyl, alkylamino,dialkylamino substituted heterocyclic group, including but notlimited to: 4-N, N-dimethylamino-piperidinyl, 4-N,N-diethylamino-piperidinyl, 4-N, N-diisopropylamino-piperidinyl,4-hydroxyl-piperidinyl, 4-(N-methylpiperazinyl) piperidinyl,4-(N-ethyl-piperazinyl) piperidinyl, 4-(N-isopropyl-piperazinyl)piperidinyl, 4-(N-acetyl-piperazinyl) piperidinyl,4-(N-tert-butoxyl formyl-piperazinyl) piperidinyl,4-(N-methylsulfonyl-piperazinyl) piperidinyl,4-(N-(2-hydroxylethyl) piperazinyl) piperidinyl,4-(N-(2-cyanoethyl) piperazinyl) piperidinyl, 4-(N-(3-hydroxylpropyl) piperazinyl) piperidinyl, 4-(N-(2-N, N-dimethyl-3 ethyl)piperazinyl) piperidinyl, 4-(N-(2-N, N-diethyl-ethyl) piperazinyl)piperidinyl, 4-(N-(3-N, N-dimethyl-propyl) piperazinyl)piperidinyl, 4-(N-(3-N, N-diethyl-propyl) piperazinyl) piperidinyl,4-(tertahydropyrrolyl) piperidinyl, 4-(3-N,N-dimethyl-tetrahydropyrrolyl) piperidinyl; 2) amino,cyclopropylamino, cyclobutylamino, cyclopentylamino,cyclohexylamino, N, N-dimethylamino, N, N-diethylamino, N,N-diisopropylamino, 2-N, N-dimethylamino ethylamino,2-hydroxylethylamino, 2-morpholinyl-ethylamino,2-(4-N-methylpiperazinyl) ethylamino, 3-N, N-dimethyl-aminopropylamino, 3-N, N-diethylamino propylamino, 3-N,N-diisopropylamino-propylamino, 3-hydroxypropyl,3-morpholinyl-propylamino, 3-(4-N-methylpiperazinyl) propylamino,N-methyl-piperidinyl-4-amino, N-ethylpiperidinyl-4-amino,N-isopropyl piperidinyl-4-amino, N-acetylpiperidinyl-4-amino;N-methyl-piperazinyl, N-ethyl-piperazinyl, N-isopropyl-piperazinyl,N-acetyl piperazinyl, N-tert-butoxyl formyl-piperazinyl,N-methylsulfonyl-piperazinyl, N-(2-hydroxylethyl) piperazinyl,N-(2-cyanoethyl) piperazinyl, N-(3-hydroxylpropyl) piperazinyl,N-(2-N, N-dimethylethyl) piperazinyl, N-(2-N, N-diethyl aminoethyl)piperazinyl, N-(3-N, N-dimethyl-propyl) piperazinyl, N-(3-N,N-diethyl-propyl) piperazinyl, 2-oxo-piperazin-4-yl,N--(N-methyl-4-piperidinyl) piperazinyl, N--(N-ethyl-4-piperidinyl)piperazinyl, N--(N-acetyl-4-piperidinyl) piperazinyl; morpholinyl,3,5-dimethyl morpholinyl, thiomorpholinyl, tetrahydropyrrolyl, 3-N,N-dimethyl-tetrahydropyrrolyl, 3-N, N-diethyl-tetrahydropyrrolyl;3) C1-C6 alkyl, optionally substituted by halogen, nitro, cyano;[0086] 4) C3-C7 cycloalkyl, which is optionally substituted byhalogen, nitro, cyano; 5) --O--C1-C6 alkyl, optionally substitutedby halogen, nitro, amino, cyano; [0087] 6) --O--C3-C7 cycloalkyl,which is optionally substituted by halogen, nitro, amino, cyano; 7)C6-C10 aryl, which is optionally substituted by halogen, nitro,amino, cyano; --O-C6-C10 aryl, which is optionally substituted byhalogen, nitro, amino, cyano; [0088] 8) C2-C6 alkenyl; [0089] 9)hydroxyl, 2-N, N-dimethylaminoethoxyl, 2-N, N-diethyl-aminoethoxyl,2-N, N-diisopropyl-aminoethoxyl, 2-(N-methylpiperazinyl) ethoxyl,2-(N-acetyl-piperazinyl) ethoxyl, 2-morpholinyl-ethoxyl,2-thiomorpholinyl ethoxyl, 2-piperidinyl-ethoxyl, 3-N,N-dimethylamino-propoxyl, 3-N, N-diethylamino-propoxyl, 3-N,N-diisopropylamino propoxyl, 3-(N-methylpiperazinyl) propoxyl,3-(N-acetyl-piperazinyl) propoxyl, 3-morpholinyl-propoxyl,3-thiomorpholinyl propoxyl, 3-piperidinyl-propoxyl,2-pyridyl-methoxyl, 3-pyridyl-methoxyl, 4-pyridyl methoxyl,phenylmethoxyl, monhalogen-substituted phenylmethoxyl,hom*odihalogen substituted phenylmethoxyl, heterodihalo-substitutedphenylmethoxyl; R.sub.24 is selected from: 1) a hydrogen, halo,nitro, amino, cyano; 2) C1-C6 alkyl, optionally substituted byhalogen, nitro, amino, cyano; --O-C1-C6 alkyl, optionallysubstituted by halogen, nitro, amino, cyano; C1-C6oxygen-containing alkyl; 3) methylthio, ethylthio, isopropylthio,methylsulfinyl, ethyl sulfinyl, propyl sulfinyl, methylsulfonyl,ethylsulfonyl, isopropylsulfonyl, amino sulfonyl, ethylaminosulfonyl, propylamino sulfonyl, isopropylamino sulfonyl,cyclopropylamino sulfonyl, hydroxyl formyl, methoxyl formyl,ethoxyl formyl, propoxyl formyl, isopropoxyl formyl, n-butoxylformyl, isobutoxyl formyl, t-butoxyl formyl, amino formyl,methylamino formyl, ethylamino formyl, propylamino formyl,isopropylamino formyl, cyclopropylamino formyl, cyclobutylaminoformyl, cyclopentylamino formyl, acetamido, propionamido,n-butylamido, isobutyl amido, cyclopropyl formamido, cyclobutylformamido, cyclopentyl formamido, methylsulfonamido,ethylsulfonamido, propylsulfonamido, isopropylsulfonamido, dimethylphosphinyl, diethyl phosphinyl, diisopropyl phosphinyl; R.sub.34 isselected from: hydrogen; C1-C6 alkyl, optionally substituted byhalogen, nitro, amino, cyano; C3-C7 cycloalkyl, which is optionallysubstituted by halogen, nitro, amino, cyano; R.sub.4 is selectedfrom: hydrogen; C1-C6 alkyl, optionally substituted by halogen,nitro, amino, cyano; C3-C7 cycloalkyl, which is optionallysubstituted by halogen, nitro, amino, cyano; Or a stereoisomer ofthe above compounds, a prodrug thereof, a pharmaceuticallyacceptable salt thereof, or a pharmaceutically acceptable solvatethereof.

[0090] A sixth aspect, the inhibitors of MST1/2 protein kinases arerepresented by the following structural formula:

##STR00069##

wherein m5 is selected from 0, 1, 2, 3 or 4; n5 is selected from 0,1 or 2; R.sub.15, R.sub.25 are independently selected from: 1)hydrogen; C1-C6 alkyl, optionally substituted by halogen, nitro,amino, cyano; C3-C7 cycloalkyl, which is optionally substituted byhalogen, nitro, amino, cyano; a C1-C6 oxygen-containing alkyl; 2)--SO.sub.2C1-C6 alkyl, optionally substituted by halogen, nitro,amino, cyano; --SO.sub.2C2-C6 alkenyl, which is optionallysubstituted with halogen, nitro, amino, cyano substituted;--COC1-C6 alkyl, optionally substituted by halogen, nitro, amino,cyano; --COC2-C6 alkenyl, which is optionally substituted byhalogen, nitro, amino, cyano; Or, together with R.sub.15 andR.sub.25 and the N atom to which they are attached forming ahexaheterocyclic ring that contains one or more heteroatomsselected from N, O and S, said hexahetrerocyclic ring is optionallysubstituted with C1-C6 alkyl, hydroxyl, or amino group; R.sub.35 isselected from: 1) a hydrogen, halo, nitro, amino, cyano; 2) C1-C6alkyl, optionally substituted by halogen, nitro, amino, cyano;--O--C1-C6 alkyl, optionally substituted by halogen, nitro, amino,cyano; a C1-C6 oxygen-containing alkyl; 3) methylthio, ethylthio,isopropylthio, methylsulfinyl, ethyl sulfinyl, propyl sulfinyl,methylsulfonyl, ethylsulfonyl, isopropylsulfonyl, amino sulfonyl,ethylamino sulfonyl, propylamino sulfonyl, isopropylamino-sulfonyl,cyclopropylamino sulfonyl, hydroxyl formyl, methoxyl formyl,ethoxyl formyl, propoxyl formyl, isopropoxyl formyl, nbutoxylformyl, isobutoxyl formyl, t-butoxyl formyl, amino formyl,methylamino formyl, ethylamino formyl, propylamino formyl,isopropylamino formyl, cyclopropylamino formyl, cyclobutylaminoformyl, cyclopentylamino formyl, acetamido, propionamido, n-butylamido, isobutyl amido, cyclopropyl formamido, cyclobutyl formamido,cyclopentyl formamido, methylsulfonamido, ethylsulfonamido,propylsulfonamido, isopropylsulfonamido, dimethyl phosphinyl,diethyl phosphinyl, diisopropyl phosphinyl; R.sub.45 is selectedfrom:

[0091] hydrogen; C1-C6 alkyl, optionally substituted by halogen,nitro, amino, cyano; C3-C7 cycloalkyl, which is optionallysubstituted by halogen, nitro, amino, cyano;

R.sub.55 is selected from: hydrogen; C1-C6 alkyl, optionallysubstituted by halogen, nitro, amino, cyano; C3-C7 cycloalkyl,which is optionally substituted by halogen, nitro, amino, cyano;R.sub.65 is selected from: 1) a hydrogen, halo, nitro, amino,cyano; 2) C1-C6 alkyl, optionally substituted by halogen, nitro,amino, cyano; --O-C1-C6 alkyl, optionally substituted by halogen,nitro, amino, cyano; a C1-C6 oxygen-containing alkyl; 3)methylthio, ethylthio, isopropylthio, methylsulfinyl, ethylsulfinyl, propyl sulfinyl, methaylsulfonyl, ethylsulfonyl,isopropylsulfonyl, amino sulfonyl, ethylamino sulfonyl, propylaminosulfonyl, isopropylamino-sulfonyl, cyclopropylamino sulfonyl,hydroxyl formyl, methoxyl formyl, ethoxyl formyl, propoxyl formyl,isopropoxyl formyl, n-butoxyl formyl, isobutoxyl formyl, t-butoxylformyl, amino formyl, methylamino formyl, ethylamino formyl,propylamino formyl, isopropylamino formyl, cyclopropylamino formyl,cyclobutylamino formyl, cyclopentylamino formyl, acetamido,propionamido, n-butyl amido, isobutyl amido, cyclopropyl formamido,cyclobutyl foramido, cyclopentyl foramido, methylsulfonamido,ethylsulfonamido, propylsulfonamido, isopropylsulfonamido, dimethylphosphinyl, diethyl phosphinyl, diisopropyl phosphinyl; Or astereoisomer of the above compounds, a prodrug thereof, apharmaceutically acceptable salt thereof, or a pharmaceuticallyacceptable solvate thereof.

[0092] A seventh aspect, the inhibitors of MST1/2 protein kinasesare represented by the following structural formula:

##STR00070##

n6 is selected from 0, 1 or 2; R.sub.16 is selected from: 1)hydrogen; C1-C6 alkyl, optionally substituted by halogen, nitro,amino, cyano; C3-C7 cycloalkyl, which is optionally substituted byhalogen, nitro, amino, cyano; a C1-C6 oxygen-containing alkyl; 2)--SO.sub.2C1-C6 alkyl, optionally substituted by halogen, nitro,amino, cyano; --SO.sub.2C2-C6 alkenyl, which is optionallysubstituted with halogen, nitro, amino, cyano substituted;--COC1-C6 alkyl, optionally substituted by halogen, nitro, amino,cyano; --COC2-C6 alkenyl, which is optionally substituted byhalogen, nitro, amino, cyano; 3) piperidinyl, 4-N,N-dimethylamino-piperidinyl, 4-N, N-diethylamino-piperidinyl, 4-N,N-diisopropylamino-piperidinyl, 4-hydroxyl piperidinyl,4-(N-methylpiperazinyl) piperidinyl, 4-(N-ethyl-piperazinyl)piperidinyl, 4-(N-isopropyl-piperazinyl) piperidine,4-(N-acetyl-piperazinyl) piperidinyl, 4-(N-tert-butoxylformyl-piperazinyl) piperidinyl, 4-(N-methylsulfonyl-piperazinyl)piperidinyl, 4-(N-(2-hydroxylethyl) piperazinyl) piperidinyl,4-(N-(2-cyanoethyl) piperazinyl) piperidinyl,4-(N-(3-hydroxylpropyl) piperazinyl) piperidinyl, 4-(N-(2-N,N-dimethyl-ethyl) piperazinyl) piperidinyl, 4-(N-(2-N, N-diethylethyl) piperazinyl) piperidinyl, 4-(N-(3-N, N-dimethyl-propyl)piperazinyl) piperidinyl, 4-(N-(3-N, N-diethyl-propyl) piperazinyl)piperidinyl, 4-(tetrahydropyrrolyl) piperidinyl, 4-(3-N,N-dimethyl-tetrahydropyrrolyl) piperidinyl; R.sub.26 is selectedfrom: 1) a hydrogen, halo, nitro, amino, cyano; 2) C1-C6 alkyl,optionally substituted by halogen, nitro, amino, cyano; --O-C1-C6alkyl, optionally substituted by halogen, nitro, amino, cyano; aC1-C6 oxygen-containing alkyl; 3) methylthio, ethylthio,isopropylthio, methylsulfinyl, ethyl sulfinyl, propyl sulfinyl,methylesulfonyl, ethylsulfonyl, isopropylsulfonyl, amino sulfonyl,ethylamino sulfonyl, propylamino sulfonyl, isopropylamino-sulfonyl,cyclopropylamino sulfonyl, hydroxyl formyl, methoxyl formyl,ethoxyl formyl, propoxyl formyl, isopropoxyl formyl, n-butoxylformyl, isobutoxyl formyl, t-butoxyl formyl, amino formyl,methylamino formyl, ethylamino formyl, propylamino formyl,isopropylamino formyl, cyclopropylamino formyl, cyclobutylaminoformyl, cyclopentylamino formyl, acetamido, propionamido, n-butylamido, isobutyl amido, cyclopropyl formamido, cyclobutyl formamido,cyclopentyl formamido, methylsulfonamido, ethylsulfonamido,propylsulfonamido, isopropylsulfonamido, dimethyl phosphinyl,diethyl phosphinyl, diisopropyl phosphinyl; R.sub.36 is selectedfrom: hydrogen; C1-C6 alkyl, optionally substituted by halogen,nitro, amino, cyano; C3-C7 cycloalkyl, which is optionallysubstituted with halogen, nitro, amino, cyano; R.sub.46 is selectedfrom: hydrogen; C1-C6 alkyl, optionally substituted by halogen,nitro, amino, cyano; C3-C7 cycloalkyl, which is optionallysubstituted with halogen, nitro, amino, cyano; Or a stereoisomer ofthe above compounds, a prodrug thereof, a pharmaceuticallyacceptable salt thereof, or a pharmaceutically acceptable solvatethereof.

[0093] An eighth aspect, the inhibitors of MST1/2 protein kinasesare represented by the following structural formula:

##STR00071##

wherein n7 is selected from 0, 1 or 2; R.sub.17 is selected from:1) hydrogen; C1-C6 alkyl, optionally substituted by halogen, nitro,amino, cyano; C3-C7 cycloalkyl, which is optionally substituted byhalogen, nitro, amino, cyano; a C1-C6 oxygen-containing alkyl; 2)--SO.sub.2C1-C6 alkyl, optionally substituted by halogen, nitro,amino, cyano; --SO.sub.2C2-C6 alkenyl, which is optionallysubstituted with halogen, nitro, amino, cyano substituted;--COC1-C6 alkyl, optionally substituted by halogen, nitro, amino,cyano; --COC2-C6 alkenyl, which is optionally substituted byhalogen, nitro, amino, cyano; 3) piperidinyl, 4-N,N-dimethylamino-piperidinyl, 4-N, N-diethylamino-piperidinyl, 4-N,N-diisopropylamino-piperidinyl, 4-hydroxyl piperidinyl,4-(N-methylpiperazinyl) piperidinyl, 4-(N-ethyl-piperazinyl)piperidinyl, 4-(N-isopropyl-piperazinyl) piperidinyl,4-(N-acetyl-piperazinyl) piperidinyl, 4-(N-tert-butoxylformyl-piperazinyl) piperidinyl, 4-(N-methylsulfonyl-piperazinyl)piperidinyl, 4-(N-(2-hydroxylethyl) piperazinyl) piperidinyl,4-(N-(2-cyanoethyl) piperazinyl) piperidinyl,4-(N-(3-hydroxylpropyl) piperazinyl) piperidinyl, 4-(N-(2-N,N-dimethyl-ethyl) piperazinyl) piperidinyl, 4-(N-(2-N, N-diethylethyl) piperazinyl) piperidinyl, 4-(N-(3-N, N-dimethyl-propyl)piperazinyl) piperidinyl, 4-(N-(3-N, N-diethyl-propyl) piperazinyl)piperidinyl, 4-(tetrahydropyrrolyl) piperidinyl, 4-(3-N,N-dimethyl-tetrahydropyrrolyl) piperidinyl; R.sub.27 is selectedfrom: 1) a hydrogen, halo, nitro, amino, cyano; 2) C1-C6 alkyl,optionally substituted by halogen, nitro, amino, cyano; --O-C1-C6alkyl, optionally substituted by halogen, nitro, amino, cyano; aC1-C6 oxygen-containing alkyl; 3) methylthio, ethylthio,isopropylthio, methylsulfinyl, ethyl sulfinyl, propyl sulfinyl,methylsulfonyl, ethylsulfonyl, isopropylsulfonyl, amino sulfonyl,ethylamino sulfonyl, propylamino sulfonyl, isopropylamino-sulfonyl,cyclopropylamino sulfonyl, hydroxyl formyl, methoxyl formyl,ethoxyl formyl, propoxyl formyl, isopropoxyl formyl, n-butoxylformyl, isobutoxyl formyl, t-butoxyl formyl, amino formyl,methylamino formyl, ethylamino formyl, propylamino formyl,isopropylamino formyl, cyclopropylamino formyl, cyclobutylaminoformyl, cyclopentylamino formyl, acetamido, propionamido, n-butylamido, isobutyl amido, cyclopropyl formamido, cyclobutyl formamido,cyclopentyl formamido, methylsulfonamido, ethylsulfonamido,propylsulfonamido, isopropylsulfonamido, dimethyl phosphinyl,diethyl phosphinyl, diisopropyl phosphinyl; R.sub.37 is selectedfrom: Hydrogen; C1-C6 alkyl, optionally substituted by halogen,nitro, amino, cyano; C3-C7 cycloalkyl, which is optionallysubstituted by halogen, nitro, amino, cyano; R.sub.47 is selectedfrom: Hydrogen; C1-C6 alkyl, optionally substituted by halogen,nitro, amino, cyano; C3-C7 cycloalkyl, which is optionallysubstituted by halogen, nitro, amino, cyano; Or a stereoisomer ofthe above compounds, a prodrug thereof, a pharmaceuticallyacceptable salt thereof, or a pharmaceutically acceptable solvatethereof.

[0094] The inhibotors of MST1/2 protein kinases having the formulaII are exemplified by the compounds in Table 2.

TABLE-US-00002 TABLE 2 MST1/2 protein kinase inhibitors havingFormula II ##STR00072## II-1 (TFA Salt) ##STR00073## II-2##STR00074## II-3 ##STR00075## II-4 ##STR00076## II-5 ##STR00077##II-6 ##STR00078## II-7 ##STR00079## II-8 (TFA Salt) ##STR00080##II-9 ##STR00081## II-10 (HCl Salt) ##STR00082## II-11 (HCl Salt)##STR00083## II-12 (HCl Salt) ##STR00084## II-13 (HCl Salt)##STR00085## II-14 (TFA Salt) ##STR00086## II-15 ##STR00087## II-16##STR00088## II-17 (TFA Salt) ##STR00089## II-18 (TFA Salt)##STR00090## II-19 (TFA Salt) ##STR00091## II-20 ##STR00092## II-21##STR00093## II-22 ##STR00094## II-23 (TFA Salt) ##STR00095## II-24##STR00096## II-25 ##STR00097## II-26 (TFA Salt) ##STR00098## II-27##STR00099## II-28 ##STR00100## II-29 ##STR00101## II-30

[0095] A ninth aspect, the inhibitors of MST1/2 protein kinases arerepresented by the following structural formula:

##STR00102##

wherein n8 is selected from 0, 1 or 2; R.sub.18 is selected from:1) selected from the group of five-membered heterocyclic orsix-membered heterocyclic rings comprising one or more heteroatomsselected from N, O and S, the five-membered heterocyclic orsix-membered heterocyclic rings are optionally substituted withC1-C6 alkyl, C1-C6 alkoxy, hydroxy, amino, C1-C6 acyl, a cyano, asubstituted heterocyclic group, including but not limited to: 4-N,N-dimethylamino-piperidinyl, 4-N, N-diethylamino-piperidinyl, 4-N,N-diisopropylamino-piperidinyl, 4-hydroxyl-piperidinyl,4-(N-methylpiperazinyl) piperidinyl, 4-(N-ethyl-piperazinyl)piperidinyl, 4-(N-isopropyl-piperazinyl) piperidinyl,4-(N-acetyl-piperazinyl) piperidinyl, 4-(N-tert-butoxylformyl-piperazinyl) piperidinyl, 4-(N-methylsulfonyl-piperazinyl)piperidinyl, 4-(N-(2-hydroxyethyl) piperazinyl) piperidinyl,4-(N-(2-cyanoethyl) piperazinyl) piperidinyl, 4-(N-(3-hydroxylpropyl) piperazinyl) piperidinyl, 4-(N-(2-N, N-dimethyl-ethyl)piperazinyl) piperidinyl, 4-(N-(2-N, N-diethyl-ethyl) piperazinyl)piperidinyl, 4-(N-(3-N, N-dimethyl-propyl) piperazinyl)piperidinyl, 4-(N-(3-N, N-diethyl-propyl) piperazinyl) piperidinyl,4-(tetrahydropyrrolyl) piperidinyl, 4-(3-N,N-dimethyl-tetrahydropyrrolyl) piperidinyl; 2) amino,cyclopropylamino, cyclobutylamino, cyclopentylamino,cyclohexylamino, N, N-dimethylamino, N, N-diethylamino, N,N-diisopropylamino, 2-N, N-dimethylethylamino, 2-hydroxyethylamino,2-morpholinyl-ethylamino, 2-(4-N-methylpiperazinyl) ethylamino,3-N, N-dimethyl-aminopropyl amino, 3-N, N-diethylamino propylamino,3-N, N-diisopropylpropylamino, 3-hydroxypropyl amino,3-morpholinyl-propylamino, 3-(4-N-methylpiperazinyl) propylamino,N-methylpiperidiny-4-amino, N-ethylpiperidinyl-4-amino, N-isopropylpiperidinyl-4-amino, N-acetylpiperidinyl-4-amino;N-methyl-piperazinyl, N-ethyl-piperazinyl, N-isopropyl-piperazinyl,N-acetyl piperazinyl, N-tert-butoxyl formyl-piperazinyl,N-methylsulfonyl-piperazinyl, N-(2-hydroxylethyl) piperazinyl,N-(2-cyanoethyl) piperazinyl, N-(3-hydroxylpropyl) piperazinyl,N-(2-N, N-dimethylethyl) piperazinyl, N-(2-N, N-diethyl aminoethyl)piperazinyl, N-(3-N, N-dimethyl-propyl) piperazinyl, N-(3-N,N-diethyl-propyl) piperazinyl, 2-oxo-piperazin-4-yl,N--(N-methyl-4-piperidinyl) piperazinyl, N--(N-ethyl-4-piperidinyl)piperazinyl, N--(N-acetyl-4-piperidinyl) piperazinyl; morpholinyl,3,5-dimethyl morpholinyl, thiomorpholinyl, tetrahydropyrrolyl, 3-N,N-dimethyl-tetrahydropyrrolyl, 3-N, N-diethyl-tetrahydropyrrolyl;3) C1-C6 alkyl, optionally substituted by halogen, nitro, cyano; 4)C3-C7 cycloalkyl, which is optionally substituted by halogen,nitro, cyano; 5) --O-C1-C6 alkyl, optionally substituted byhalogen, nitro, amino, cyano; 6) --O-C3-C7 cycloalkyl, which isoptionally substituted by halogen, nitro, amino, cyano; 7) C6-C10aryl, which is optionally substituted by halogen, nitro, amino,cyano; --O-C6-C10 aryl, which is optionally substituted by halogen,nitro, amino, cyano; 8) C2-C6 alkenyl; 9) hydroxyl, 2-N,N-dimethylaminoethoxyl, 2-N, N-diethyl-aminoethoxyl, 2-N,N-diisopropyl-aminoethoxyl, 2-(N-methylpiperazine-yl) ethoxyl,2-(N-acetyl-piperazinyl) ethoxyl, 2-morpholinyl-ethoxyl,2-thiomorpholinyl ethoxyl, 2-piperidinyl-ethoxyl, 3-N,N-dimethylamino-propoxyl, 3-N, N-diethylamino-propoxyl, 3-N,N-diisopropylamino propoxyl, 3-(N-methylpiperazinyl) propoxyl,3-(N-acetyl-piperazinyl) propoxyl, 3-morpholinyl-propoxyl,3-thiomorpholinyl propoxyl, 3-piperidinyl-propoxyl,2-pyridyl-methoxyl, 3-pyridyl-methoxyl, 4-pyridyl methoxyl,phenylmethoxyl, monohalogen-substituted phenylmethoxyl,hom*odihalogen-substituted phenylmethoxyl,heterodihalogen-substituted phenylmethoxyl; R.sub.28 is selectedfrom: 1) a hydrogen, halo, nitro, amino, cyano; 2) C1-C6 alkyl,optionally substituted by halogen, nitro, amino, cyano; --O-C1-C6alkyl, optionally substituted by halogen, nitro, amino, cyano; aC1-C6 oxygen-containing alkyl; 3) methylthio, ethylthio,isopropylthio, methylsulfinyl, ethyl sulfinyl, propyl sulfinyl,methylsulfonyl, ethylsulfonyl, isopropylsulfonyl, amino sulfonyl,ethylamino sulfonyl, propylamino sulfonyl, isopropylamino sulfonyl,cyclopropylamino sulfonyl, hydroxyl formyl, methoxyl formyl,ethoxyl formyl, propoxyl formyl, isopropoxyl formyl, n-butoxylformyl, isobutoxyl formyl, t-butoxyl formyl, amino formyl,methylamino formyl, ethylamino formyl, propylamino formyl,isopropylamino formyl, cyclopropylamino formyl, cyclobutylaminoformyl, cyclopentylamino formyl, acetamido, propionamido, n-butylamido, isobutyl amido, cyclopropyl formamido, cyclobutyl formamido,cyclopentyl formamido, methylsulfonamido, ethylsulfonamido,propylsulfonamido, isopropylsulfonamido, dimethyl phosphinyl,diethyl phosphinyl, diisopropyl phosphinyl; R.sub.38 is selectedfrom: hydrogen; C1-C6 alkyl, optionally substituted by halogen,nitro, amino, cyano; C3-C7 cycloalkyl, which is optionallysubstituted with halogen, nitro, amino, cyano; R.sub.48 is selectedfrom: hydrogen; C1-C6 alkyl, optionally substituted by halogen,nitro, amino, cyano; C3-C7 cycloalkyl, which is optionallysubstituted with halogen, nitro, amino, cyano; Or a stereoisomer ofthe above compounds, a prodrug thereof, a pharmaceuticallyacceptable salt thereof, or a pharmaceutically acceptable solvatethereof.

[0096] Inhibitors of MST1/2 protein kinases having the formula IIIare exemplified by the compounds in Table 3.

TABLE-US-00003 TABLE 3 MST1/2 protein kinase inhibitors havingFormula III ##STR00103## III-1 (TFA Salt) ##STR00104## III-2##STR00105## III-3 ##STR00106## III-4

[0097] A tenth aspect, the inhibitors of MST1/2 protein kinases arerepresented by the following structural formula:

##STR00107##

wherein n9 is selected from 0, 1, 2 or 3; R.sub.19 is selectedfrom: 1) hydrogen; C1-C6 alkyl, optionally substituted by halogen,nitro, amino, cyano; C3-C7 cycloalkyl, which is optionallysubstituted by halogen, nitro, amino, cyano; a C1-C6oxygen-containing alkyl; 2) --SO.sub.2C1-C6 alkyl, optionallysubstituted by halogen, nitro, amino, cyano; --SO.sub.2C2-C6alkenyl, which is optionally substituted with halogen, nitro,amino, cyano substituted; --COC1-C6 alkyl, optionally substitutedby halogen, nitro, amino, cyano; --COC2-C6 alkenyl, which isoptionally substituted by halogen, nitro, amino, cyano; R.sub.29 isselected from: 1) a hydrogen, halo, nitro, amino, cyano; 2) C1-C6alkyl, optionally substituted by halogen, nitro, amino, cyano;--O-C1-C6 alkyl, optionally substituted by halogen, nitro, amino,cyano; a C1-C6 oxygen-containing alkyl; 3) methylthio, ethylthio,isopropylthio, methylsulfinyl, ethyl sulfinyl, propyl sulfinyl,methylsulfonyl, ethylsulfonyl, isopropylsulfonyl, amino sulfonyl,ethylamino sulfonyl, propylamino sulfonyl, isopropylamino sulfonyl,cyclopropylamino sulfonyl, hydroxyl formyl, methoxyl formyl,ethoxyl formyl, propoxyl formyl, isopropoxyl formyl, n-butoxylformyl, isobutoxyl formyl, t-butoxyl formyl, amino formyl,methylaminoformyl, ethylamino formyl, propylamino formyl,isopropylamino formyl, cyclopropylamino formyl, cyclobutylaminoformyl, cyclopentylamino formyl, acetamido, propionamido, n-butylamido, isobutyl amido, cyclopropyl formamido, cyclobutyl formamido,cyclopentyl formamido, methylsulfonamido, ethylsulfonamido,propylsulfonamido, isopropylsulfonamido, dimethyl phosphinyl,diethyl phosphinyl, diisopropyl phosphinyl; R.sub.49 is selectedfrom: hydrogen; C1-C6 alkyl, optionally substituted by halogen,nitro, amino, cyano; C3-C7 cycloalkyl, which is optionallysubstituted with halogen, nitro, amino, cyano; Or a stereoisomer ofthe above compounds, a prodrug thereof, a pharmaceuticallyacceptable salt thereof, or a pharmaceutically acceptable solvatethereof. In some embodiments, R.sub.19 is selected from hydrogen;C1-C6 alkyl; C3-C7 cycloalkyl; --SO.sub.2C1-C6 alkyl;--SO.sub.2C2-C6 alkenyl group; --COC1-C6 alkyl, which is optionallysubstituted by halogen, nitro, amino, cyano; --COC2-C6 alkenyl,which is optionally substituted by halogen, nitro, amino,cyano.

[0098] An eleventh aspect, the inhibitors of MST1/2 protein kinasesare represented by the following structural formula:

##STR00108##

wherein n.sub.0 is selected from 0, 1, 2 or 3; R.sub.10 is selectedfrom: 1) selected from the group of five-membered heterocyclic orsix-membered heterocyclic rings comprising one or more heteroatomsselected from N, O and S, the five-membered heterocyclic orsix-membered heterocyclic rings are optionally substituted withC1-C6 alkyl, C1-C6 alkoxy, hydroxy, amino, alkylamino,dialkylamino, C1-C6 acyl, cyano, optionally substituted C1-C6alkyl, --O-C1-C6 alkyl, hydroxy, hydroxy C1-C6 alkyl, C1-C6 acyl,alkylamino, dialkylamino substituted heterocyclic group, includingbut not limited to: 4-N, N-dimethylamino-piperidinyl, 4-N,N-diethylamino-piperidinyl, 4-N, N-diisopropylamino-piperidinyl,4-hydroxyl-piperidinyl, 4-(N-methylpiperazinyl) piperidinyl,4-(N-ethyl-piperazinyl) piperidinyl, 4-(N-isopropyl-piperazinyl)piperidinyl, 4-(N-acetyl-piperazinyl) piperidinyl,4-(N-tert-butoxyl formyl-piperazinyl) piperidinyl,4-(N-methylsulfonyl-piperazinyl) piperidinyl,4-(N-(2-hydroxylethyl) piperazinyl) piperidinyl,4-(N-(2-cyanoethyl) piperazinyl) piperidinyl, 4-(N-(3-hydroxylpropyl) piperazinyl) piperidinyl, 4-(N-(2-N, N-dimethyl-ethyl)piperazinyl) piperidinyl, 4-(N-(2-N, N-diethyl-ethyl) piperazinyl)piperidinyl, 4-(N-(3-N, N-dimethyl-propyl) piperazinyl)piperidinyl, 4-(N-(3-N, N-diethyl-propyl) piperazinyl) piperidinyl,4-(tetrahydropyrrolyl) piperidinyl, 4-(3-N,N-dimethyl-tetrahydropyrrolyl) piperidinyl; 2) amino,cyclopropylamino, cyclobutylamino, cyclopentylamino,cyclohexylamino, N, N-dimethylamino, N, N-diethylamino, N,N-diisopropylamino, 2-N, N-dimethylaminoethyl,2-hydroxylethylamino, 2-morpholinyl-ethylamino,2-(4-N-methylpiperazinyl) ethylamino, 3-N, N-dimethylaminopropylamino, 3-N, N-diethylaminopropyl amino, 3-N,N-diisopropylamino-propyl amino, 3-hydroxylpropyl amino,3-morpholinyl-propylamino, 3-(4-N-methylpiperazinyl) propylamino,N-methyl-piperidinyl-4-amino, N-ethylpiperidinyl-4-amino,N-isopropyl piperidinyl-4-amino, N-acetylpiperidinyl-4-amino;N-methyl-piperazinyl, N-ethyl-piperazinyl, N-isopropyl-piperazinyl,N-acetyl piperazinyl, N-tert-butoxyl formyl-piperazinyl,N-methylsulfonyl-piperazinyl, N-(2-hydroxylethyl) piperazinyl,N-(2-cyanoethyl) piperazinyl, N-(3-hydroxylpropyl) piperazinyl,N-(2-N, N-dimethylethyl) piperazinyl, N-(2-N, N-diethyl aminoethyl)piperazinyl, N-(3-N, N-dimethyl-propyl) piperazinyl, N-(3-N,N-diethyl-propyl) piperazinyl, 2-oxo-piperazin-4-yl,N--(N-methyl-4-piperidinyl) piperazinyl, N--(N-ethyl-4-piperidinyl)piperazinyl, N--(N-acetyl-4-piperidinyl) piperazinyl; morpholinyl,3,5-dimethyl morpholinyl, thiomorpholinyl, tetrahydropyrrolyl, 3-N,N-dimethyl-tetrahydropyrrolyl, 3-N, N-diethyl-tetrahydropyrrolyl;3) C1-C6 alkyl, optionally substituted by halogen, nitro, cyano; 4)C3-C7 cycloalkyl, which is optionally substituted by halogen,nitro, cyano; 5) --O-C1-C6 alkyl, optionally substituted byhalogen, nitro, amino, cyano; 6) --O-C3-C7 cycloalkyl, which isoptionally substituted by halogen, nitro, amino, cyano; 7) C6-C10aryl group, which is optionally substituted by halogen, nitro,amino, cyano; --O-C6-C10 aryl, which is optionally substituted byhalogen, nitro, amino, cyano; 8) C2-C6 alkenyl group; 9) hydroxyl,2-N, N-dimethylaminoethoxyl, 2-N, N-diethyl-aminoethoxyl, 2-N,N-diisopropyl-aminoethoxyl, 2-(N-methylpiperazinyl) ethoxyl,2-(N-acetyl-piperazinyl) ethoxyl, 2-morpholinyl-ethoxyl,2-thiomorpholinyl ethoxyl, 2-piperidinyl-ethoxyl, 3-N,N-dimethylamino-propoxyl, 3-N, N-diethylamino-propoxyl, 3-N,N-diisopropylamino propoxyl, 3-(N-methylpiperazinyl) propoxyl,3-(N-acetyl-piperazinyl) propoxyl, 3-morpholinyl-propoxyl,3-thiomorpholinyl propoxyl, 3-piperidinyl-propoxyl, 2-pyridylmethoxyl, 3-pyridyl-methoxyl, 4-pyridyl methoxyl, phenylmethoxyl,monohalogen-substituted phenylmethoxyl, hom*odihalogen-substitutedphenyl methoxyl, heterodihalo-substituted phenylmethoxyl; R.sub.20is selected from: 1) a hydrogen, halo, nitro, amino, cyano; 2)C1-C6 alkyl, optionally substituted by halogen, nitro, amino,cyano; --O-C1-C6 alkyl, optionally substituted by halogen, nitro,amino, cyano; a C1-C6 oxygen-containing alkyl; 3) methylthio,ethylthio, isopropylthio, methylsulfinyl, ethyl sulfinyl, propylsulfinyl, methylsulfonyl, ethylsulfonyl, isopropylsulfonyl, aminosulfonyl, ethylamino sulfonyl, propylamino sulfonyl,isopropylamino-sulfonyl, cyclopropylamino sulfonyl, hydroxylformyl, methoxyl formyl, ethoxyl formyl, propoxyl formyl,isopropoxyl formyl, n-butoxyl formyl, isobutoxyl formyl, t-butoxylformyl, amino formyl, methylamino formyl, ethylamino formyl,propylamino formyl, isopropylamino formyl, cyclopropylamino formyl,cyclobutylamino formyl, cyclopentylamino formyl, acetamido,propionamido, n-butyl amido, isobutyl amido, cyclopropyl formamido,cyclobutyl formamido, cyclopentyl formamido, methylsulfonamido,ethylsulfonamido, propylsulfonamido, isopropylsulfonamido, dimethylphosphinyl, diethyl phosphinyl, diisopropyl phosphinyl; R.sub.40 isselected from: hydrogen; C1-C6 alkyl, optionally substituted byhalogen, nitro, amino, cyano; C3-C7 cycloalkyl, which is optionallysubstituted with halogen, nitro, amino, cyano; Or a stereoisomer ofthe above compounds, a prodrug thereof, a pharmaceuticallyacceptable salt thereof, or a pharmaceutically acceptable solvatethereof.

[0099] Inhibitors of MST1/2 protein kinases having the formula IVare exemplified by the compounds in Table 4.

TABLE-US-00004 TABLE 4 MST1/2 protein kinase inhibitors havingFormula IV ##STR00109## IV-1 (TFA Salt) ##STR00110## IV-2 (TFASalt) ##STR00111## IV-3 (TFA Salt)

[0100] Some of the inhibitors of the MST1/2 protein kinases haveactivities in inhibiting MST1 and MST2 as shown in Table 5.

TABLE-US-00005 TABLE 5 IC50 of the MST1/2 protein kinase inhibitorsNo MST1 IC.sub.50 (.mu.M) MST2 IC.sub.50 (.mu.M) IA-2 2.38 14.80IA-4 2.51 2.74 IA-5 24.43 23.30 IA-6 >30.00 >30.00 IB-1>30.00 >30.00 1B-2 >30.00 >30.00 1B-3 >30.00>30.00 IC-1 0.07 0.11 IC-2 2.32 7.80 IC-3 0.67 0.52 IC-4>30.00 >30.00 IC-5 1.91 1.80 IC-7 0.50 0.09 IC-8 13.50 0.20IC-9 0.18 2.33 IC-10 0.24 0.08 ID-1 0.04 0.11 ID-2 0.08 0.39 ID-38.65 >30.00 IE-1 0.83 2.45 IE-2 15.76 >30.00 IE-3 2.72 8.21IF-2 0.63 3.86 IF-3 2.44 15.69 IF-4 6.46 >30.00 IF-5 >30.00>30.00 IF-6 9.76 >30.00 IG-1 1.55 2.70 IG-2 5.80 14.64 IG-3>30.00 >30.00 IH-1 5.01 4.81 IH-2 0.89 1.68 IH-4 0.15 0.19IH-5 2.40 2.73 IH-6 >30.00 >30.00 IH-3 >30.00 >30.00IH-7 3.12 2.49 IH-8 2.17 3.36 IH-9 5.67 11.63 IH-10 5.93 >30.00IH-11 0.23 0.44 IH-12 1.66 1.94 IH-13 7.82 9.47 IH-14 >30.0022.53 II-1 0.14 0.04 II-2 2.71 >30.00 II-3 0.85 1.72 II-4 0.170.22 II-5 4.64 8.89 II-6 0.26 0.02 II-7 1.10 0.15 II-8 0.08 1.03II-10 1.02 1.97 II-11 >30.00 >30.00 II-12 0.57 0.22 II-130.33 0.26 II-14 2.16 2.75 II-15 0.65 1.99 II-16 >30.00 >30.00II-17 0.61 2.51 II-18 3.34 13.00 II-19 3.03 12.59 II-20 0.11 2.32II-21 0.25 3.10 II-22 >30.00 >30.00 II-23 0.77 4.76 II-240.22 2.25 II-25 0.58 2.24 II-27 3.44 >30.00 II-26 0.82 0.41II-28 0.54 0.36 II-30 0.81 0.44 IV-1 6.07 3.97 IV-2 >30.00>30.00 IV-3 0.43 1.61

[0101] The inhibitors of MST1/2 protein kinases also encompass thepharmaceutically acceptable salts of the compounds described above,including inorganic or organic acid salts, wherein the inorganicsalt is a hydrochloride, hydrobromide, hydroiodide, nitrate,bicarbonate, and salts of carbonates, sulfates or phosphates, theorganic acid salt is a formate, acetate, propionate, benzoate,maleate, fumarate, succinate, tartrate, citrate, ascorbate,alpha-ketoglutarate, alpha-glycerophosphate, alkyl sulfonate oraryl sulfonate. Preferably, the alkyl sulfonate is methylsulfonateor ethylsulfonate; and aryl sulfonate is benzylsulfonate orp-toluenylsulfonate.

[0102] In another aspect, the inhibitors of Mst1/2 protein kinasesalso encompass prodrugs of the compounds described above.

[0103] The inhibitors of Mst1/2 protein kinases may be formulatedinto a pharmaceutical composition, which also comprises one or moresuitable pharmaceutical excipients, such as, carriers, diluents,fillers, buffers, bulking agents, stabilizers, solubilizers, andthe like. The pharmaceutical composition may be in hard or softshell capsules, swallowable tablets, buccal tablets, pills,troches, elixirs, suspensions, syrups, wafers, and the likeformulations. For instance, tablets and pills may be coated withgelatin, wax, shellac or sugar. In addition, the pharmaceuticalcomposition may be in an injection or infusion formulation ofsolution or dispersion suitable for sterile injectable or infusibleformulation of the instant (optionally encapsulated in liposomes)in sterile aqueous solutions or dispersions or sterile powders.

[0104] The pharmaceutical composition may be in unit dosage form,which unit dosage form is a physically discrete unit containing aunit dose, suitable for administration to humans and other mammals.The unit dosage form can be a capsule or tablet, or a lot ofcapsules or tablets. The pharmaceutical composition may also havenew dosage forms such as liposomes, microspheres and nanospheres,such as using fine particle dispersion comprising polymericmicelles (polymeric micelles), nanoemulsion (nanoemulsion),submicron emulsion (submicroemuls micro capsule (microcapsule),microspheres (microsphere), liposomes (liposomes) and lipidvesicles (niosomes) (also known as non-ionic surfactant vesicles)in the manufacture of a medicament and the like.

[0105] More examples of dosage forms of the pharmaceuticalcompositions include tablets; caplets; capsules, such as softelastic gelatin capsules; cachets; troches; lozenges; dispersions;suppositories; ointments; cataplasms (poultices); pastes; powders;dressings; creams; plasters; solutions; patches; aerosols (e.g.,nasal sprays or inhalers); gels; liquid dosage forms suitable fororal or mucosal administration to a patient, including suspensions(e.g., aqueous or non-aqueous liquid suspensions, oil-in-wateremulsions, or a water-in-oil liquid emulsions), solutions, andelixirs; liquid dosage forms suitable for parenteral administrationto a patient; and sterile solids (e.g., crystalline or amorphoussolids) that can be reconstituted to provide liquid dosage formssuitable for parenteral administration to a patient. SeePharmaceutical Dosage Forms: Parenteral Medications, vol. 1, 2nded., Avis et al., Eds., Mercel Dekker, New York, N. Y. 1992.

[0106] In some embodiments, poorly soluble inhibitors of MST1/2protein kinases may be incorporated into liquid dosage forms (anddosage forms suitable for reconstitution) with the aid ofsolubilizing agents, emulsifiers and surfactants such as, but notlimited to, cyclodextrins (e.g., .alpha.-cyclodextrin,.beta.-cyclodextrin, Captisol.RTM., and Encapsin.RTM. (see, e.g.,Davis and Brewster, Nat. Rev. Drug Disc. 3:1023-1034 (2004)),Labrasol.RTM., Labrafil.RTM., Labrafac.RTM., cremafor, andnon-aqueous solvents, such as, but not limited to, ethyl alcohol,isopropyl alcohol, ethyl carbonate, ethyl acetate, benzyl alcohol,benzyl benzoate, propylene glycol, 1,3-butylene glycol, dimethylformamide, dimethyl sulfoxide (DMSO), biocompatible oils (e.g.,cottonseed, groundnut, corn, germ, olive, castor, and sesame oils),glycerol, tetrahydrofurfuryl alcohol, polyethylene glycols, fattyacid esters of sorbitan, and mixtures thereof (e.g.,DMSO:cornoil).

[0107] The pharmaceutical composition may be in parenteral dosageforms that can be administered to patients by various routesincluding intramuscular, intravenous (including bolus injection),subcutaneous, intraperitoneal, subdermal, intradermal,intraarticular, intrathecal, sublingual, oral and the like.Examples of parenteral dosage forms include solutions ready forinjection, dry products ready to be dissolved or suspended in apharmaceutically acceptable vehicle for injection, suspensionsready for injection, and emulsions. Suitable vehicles that can beused to provide parenteral dosage forms of the invention are wellknown to those skilled in the art. Examples include: Water forInjection USP; aqueous vehicles such as Sodium Chloride Injection,Ringer's Injection, Dextrose Injection, Dextrose and SodiumChloride Injection, and Lactated Ringer's Injection; water-misciblevehicles such as ethyl alcohol, polyethylene glycol, andpolypropylene glycol; and non-aqueous vehicles such as corn oil,cottonseed oil, peanut oil, sesame oil, ethyl oleate, isopropylmyristate, and benzyl benzoate.

[0108] The pharmaceutical composition may be formulated to enhancebioavailability of the inhibitors of MST1/2 protein kinases. Anincrease in bioavailability of the MST1/2 protein kinase inhibitorsmay be achieved by complexation of the inhibitors with one or morebioavailability or absorption enhancing agents or inbioavailability or absorption enhancing formulations. Suchbioavailability or absorption enhancing agents include, but are notlimited to, various surfactants such as various triglycerides, suchas from butter oil, monoglycerides, such as of stearic acid andvegetable oils, esters thereof, esters of fatty acids, propyleneglycol esters, the polysorbates, sodium lauryl sulfate, sorbitanesters, sodium sulfosuccinate, among other compounds. By alteringthe surfactant properties of the inhibitors, it is possible to, forexample, allow the inhibitors to have greater intestinal contactover a longer period of time that increases uptake and reduces sideeffects.

[0109] Further examples of such bioavailability or absorptionenhancing agents include carrier molecules such as cyclodextrin andderivatives thereof, well known in the art for their potential ascomplexation agents capable of altering the physicochemicalattributes of the inhibitors. For example, cyclodextrins maystabilize (both thermally and oxidatively), reduce the volatilityof, and alter the solubility of, the inhibitors with which they arecomplexed. Cyclodextrins are cyclic molecules composed ofglucopyranose ring units that form toroidal structures. Theinterior of the cyclodextrin molecule is hydrophobic and theexterior is hydrophilic, making the cyclodextrin moleculewater-soluble. The degree of solubility can be altered throughsubstitution of the hydroxyl groups on the exterior of thecyclodextrin. Similarly, the hydrophobicity of the interior can bealtered through substitution, though generally the hydrophobicnature of the interior allows accommodation of relativelyhydrophobic guests within the cavity. Examples of cyclodextrinderivatives include sulfobutylcyclodextrin, maltosylcyclodextrin,hydroxypropylcyclodextrin, and salts thereof.

[0110] Accommodation of one molecule within another is known ascomplexation and the resulting product is referred to as aninclusion complex. Complexation of the inhibitors of MST1/2 proteinkinases with a carrier molecule such as cyclodextrin to form aninclusion complex may thereby reduce the dose of the inhibitors ofMST1/2 protein kinases needed for therapeutic efficacy by enhancingthe bioavailability of the administered inhibitors of MST1/2protein kinases.

[0111] The pharmaceutical composition may also be in amicroemulsion to enhance bioavailability of the inhibitors ofMST1/2 protein kinases. A microemulsion is a fluid and stablehom*ogeneous solution composed of four major constituents,respectively, a hydrophilic phase, a lipophilic phase, at least onesurfactant (SA) and at least one cosurfactant (CoSA). A surfactantis a chemical compound possessing two groups, the first is polar orionic, which has a great affinity for water, the second contains alonger or shorter aliphatic chain and is hydrophobic. Thesechemical compounds having marked hydrophilic character are intendedto cause the formation of micelles in aqueous or oily solution.Examples of suitable surfactants include mono-, di- andtriglycerides and polyethylene glycol (PEG) mono- and diesters. Acosurfactant, also sometimes known as "co-surface-active agent", isa chemical compound having hydrophobic character, intended to causethe mutual solubilization of the aqueous and oily phases in amicroemulsion. Examples of suitable co-surfactants include ethyldiglycol, lauric esters of propylene glycol, oleic esters ofpolyglycerol, and related compounds.

[0112] The pharmaceutical composition may be formulated withvarious polymers to enhance bioavailability of the inhibitors ofMST1/2 protein kinases by increasing adhesion to mucosal surfaces,by decreasing the rate of degradation by hydrolysis or enzymaticdegradation of the inhibitors, and by increasing the surface areaof the inhibitors relative to the size of the particle. Suitablepolymers can be natural or synthetic, and can be biodegradable ornon-biodegradable. Representative natural polymers include proteinssuch as zein, modified zein, casein, gelatin, gluten, serumalbumin, and collagen, polysaccharides such as cellulose, dextrans,and polyhyaluronic acid. Synthetic polymers are generally preferreddue to the better characterization of degradation and releaseprofiles. Representative synthetic polymers includepolyphosphazenes, polyvinyl alcohols), polyamides, polycarbonates,polyacrylates, polyalkylenes, polyacrylamides, polyalkyleneglycols, polyalkylene oxides, polyalkylene terephthalates,polyvinyl ethers, polyvinyl esters, polyvinyl halides,polyvinylpyrrolidone, polyglycolides, polysiloxanes, polyurethanesand copolymers thereof. Examples of suitable polyacrylates includepory(methyl-methacrylate), poly(ethyl methacrylate), poly(butylmethacrylate), poly(isobutyl methacrylate), poly(hexylmethacrylate), poly(isodecyl methacrylate), poly(laurylmethacrylate), poly(phenyl methacrylate), poly(methyl acrylate),poly(isopropyl acrylate), poly(isobutyl acrylate) andpoly(octadecyl acrylate). Synthetically modified natural polymersinclude cellulose derivatives such as alkyl celluloses,hydroxyalkyl celluloses, cellulose ethers, cellulose esters, andnitrocelluloses. Examples of suitable cellulose derivatives includemethyl cellulose, ethyl cellulose, hydroxypropyl cellulose,hydroxypropyl methyl cellulose, hydroxybutyl methyl cellulose,cellulose acetate, cellulose propionate, cellulose acetatebutyrate, cellulose acetate phthalate, carboxymethyl cellulose,cellulose triacetate and cellulose sulfate sodium salt.

[0113] The polymers described above can be separately characterizedas biodegradable, non-biodegradable, and bioadhesive polymers, asdiscussed in more detail below. Representative synthetic degradablepolymers include polyhydroxy acids such as polylactides,polyglycolides and copolymers thereof, poly(ethyleneterephthalate), poly(butic acid), poly(valeric acid),poly(lactide-co-caprolactone), polyanhydrides, polyorthoesters andblends and copolymers thereof. Representative natural biodegradablepolymers include polysaccharides such as alginate, dextran,cellulose, collagen, and chemical derivatives thereof(substitutions, additions of chemical groups, for example, alkyl,alkylene, hydroxylations, oxidations, and other modificationsroutinely made by those skilled in the art), and proteins such asalbumin, zein and copolymers and blends thereof, alone or incombination with synthetic polymers. In general, these materialsdegrade either by enzymatic hydrolysis or exposure to water invivo, by surface or bulk erosion. Examples of non-biodegradablepolymers include ethylene vinyl acetate, poly(meth)acrylic acid,polyamides, polyethylene, polypropylene, polystyrene, polyvinylchloride, polyvinylphenol, and copolymers and mixtures thereof.Hydrophilic polymers and hydrogels tend to have bioadhesiveproperties. Hydrophilic polymers that contain carboxylic groups(e.g., poly[acrylic acid]) tend to exhibit the best bioadhesiveproperties. Polymers with the highest concentrations of carboxylicgroups are preferred when bioadhesiveness on soft tissues isdesired. Various cellulose derivatives, such as sodium alginate,carboxymethylcellulose, hydroxymethylcellulose and methylcellulosealso have bioadhesive properties. Some of these bioadhesivematerials are water-soluble, while others are hydrogels. Polymerssuch as hydroxypropylmethylcellulose acetate succinate (HPMCAS),cellulose acetate trimellitate (CAT), cellulose acetate phthalate(CAP), hydroxypropylcellulose acetate phthalate (HPCAP),hydroxypropylmethylcellulose acetate phthalate (HPMCAP), andmethylcellulose acetate phthalate (MCAP) may be utilized to enhancethe bioavailability of hypoglycemic agent and/or copper antagonistwith which they are complexed. Rapidly bioerodible polymers such aspoly(lactide-co-glycolide), polyanhydrides, and polyorthoesters,whose carboxylic groups are exposed on the external surface astheir smooth surface erodes, can also be used for bioadhesivehypoglycemic agent/copper chelator delivery systems. In addition,polymers containing labile bonds, such as polyanhydrides andpolyesters, are well known for their hydrolytic reactivity. Theirhydrolytic degradation rates can generally be altered by simplechanges in the polymer backbone. Upon degradation, these materialsalso expose carboxylic groups on their external surface, andaccordingly, these can also be used for bioadhesive deliverysystems for the inhibitors of MST1/2 protein kinases.

[0114] Other agents that may enhance bioavailability or absorptionof the MST1/2 inhibitors can act by facilitating or inhibitingtransport across the intestinal mucosa. For example, blood flow inthe stomach and intestine is a factor in determining intestinaldrug absorption and drug bioavailability, so that agents thatincrease blood flow, such as vasodilators, may increase the rate ofabsorption of orally administered inhibitors of MST1/2 proteinkinases by increasing the blood flow to the gastrointestinal tract.Vasodilators have been used in combination with other drugs. Forexample, in EPO Publication EP0106335, the use of a coronaryvasodilator, diltiazem, is reported to increase oralbioavailability of drugs which have an absolute bioavailability ofnot more than 20%, such as adrenergic beta-blocking agents (e.g.,propranolol), catecholamines (e.g., dopamine), and benzodiazepinederivatives (e.g., diazepam).

[0115] The inhibitors of MST1/2 protein kinases, optionally theirsalts, prodrugs, or in a pharmaceutical composition as describedherein, are administered to a subject in a prophylacticallyeffective amount to prevent, or in a therapeutically effectiveamount to treat, a disease or disorder associated with tissuedamages. The inhibitors of MST1/2 protein kinases of the presentinvention promote both cell protection by inhibition of cellapoptosis and tissue regeneration by stimulating cellproliferation. See FIG. 1.

[0116] Specifically, MST1/2 protein kinases activation induces cellapoptosis through multiple mechanisms: (1) phosphorylating Beclin1to inhibit autophagy but promote apoptosis (Nature Medicine,19:1367-1368, 2013); (2) translocating its C terminus into thenucleus to phosphorylate H2B to induce apoptosis (Nature Medicine,20:385-397, 2014); (3) inhibiting YAP/TAZ/TEAD1-4 signal thatnormally induces antiapoptotic gene expression. Therefore,inhibiting MST1/2 protein kinases leads to the inhibition of cellapoptosis.

[0117] In addition, YAP/TAZ/TEAD1-4 is inhibited when MST1/2protein kinases are activated. Inhibiting MST1/2 protein kinaseswill activate YAP/TAZ/TEAD1-4, which normally leads to cellproliferation. As such, inhibiting MST1/2 protein kinases promotescell proliferation and tissue regeneration.

[0118] Further, the inhibition of MST1/2 protein kinases can alsoreduce tissue inflammation via decreasing endothelial expression ofVCAM and other inflammatory cytokines.

[0119] Accordingly, the present invention uses the inhibitors ofMST1/2 protein kinases to prevent or treat a disease or disorderassociated with tissue damages by reducing cell apoptosis, inducingtissue regeneration, and/or inhibiting inflammation. As such, theinhibitors of MST1/2 protein kinases may act on multiple mechanismsto treat these diseases and disorders. There are at least fivetypes of diseases and disorders associated with tissue damages.

[0120] The first type of diseases and disorders have tissue damagesresulted from trauma to the brain (ischemic stroke, blunt trauma,subarachnoid hemorrhage), spinal cord (ischemia, blunt forcetrauma), peripheral nerves (sciatic nerve injury, diabeticneuropathy, carpal tunnel syndrome), retinal (macular edema,diabetic retinopathy, glaucoma), and heart (myocardial infarct,chronic heart failure). In a preferred embodiment, the first typeof diseases and disorders are ischemic stroke and acute myocardialinfarct (hear attack). Stroke includes both focal and globalischemia, as well as transient cerebral ischemic attacks and othercerebral vascular problems accompanied by cerebral ischemia.

[0121] The second type of diseases and disorders have tissuedamages resulted from an organ failure. For example, diabetesmellitus type I or II, nephrosis, fatty liver diseases, failure ofgonads, pancreas, kidney, heart, lung, liver and bowel. In apreferred embodiment, the second type of diseases and disorders arediabetes mellitus type I or II, fatty liver diseases, and heartfailure.

[0122] The third type of diseases and disorders have tissue damagesresulted from exposure to a toxic agent, such chemotherapeuticagents, chemical agents, radiation agents. The toxic agent cancause tissue damages to many organs, for example, organ failure.For example, liver may be damaged by consumption of alcohol andheart may be damaged (cardic injury) by chemotherapeutic agents.Nephrotoxicity may be induced by exposure to a contrast imagingagent. Further, exposure to radiation causes extensive tissuedamages to many organs. Many pharmaceutical used to treat otherdiseases may have side effects of causing tissue damages in someorgans, such as liver toxicity and renal toxicity. In a preferredembodiment, the third type of diseases and disorders are cardiacinjury induced by a chemotherapeutic agent (e.g., doxorubicin),acute liver damage induced by alcohol, and nephrotoxicity inducedby a contrast imaging agent.

[0123] The fourth type of diseases and disorders have tissuedamages resulted from inflammatory diseases, including sepsis,inflammatory bowel diseases, Crohn's disease, ulcerative colitis,ileitis, and enteritis, acute nephritis, and other inflammatorydiseases (either acute or chronic). In a preferred embodiment, thefourth type of diseases and disorders are sepsis, inflammatorybowel disease, Crohn's disease, ulcerative colitis, and acutenephritis.

[0124] The fifth type of diseases and disorders have tissue damagesresulted from degenerative diseases including muscular dystrophies,myotonic dystrophy, neurodegenerative diseases. Some examples ofneurodegenerative diseases include age-related loss of cognitivefunction and senile dementia, chronic seizure disorders,Alzheimer's disease, Parkinson's disease, dementia, memory loss,amyotrophic lateral sclerosis, multiple sclerosis, tuberoussclerosis, Wilson's disease, cerebral and progressive supranuclearpalsy, Guam disease, Lewy body dementia, prion diseases, such asspongiform encephalopathies, e.g., Creutzfeldt-Jakob disease,Huntington's disease, Freidrich's ataxia and other ataxias.Degenerative diseases also include diseases caused by excessivebone loss or cartilage or matrix degradation such as: osteoporosis,glucocorticoid induced osteoporosis, Paget's disease, abnormallyincreased bone turnover, periodontal disease, gingivitis, toothloss, bone fractures, arthritis, rheumatoid arthritis,osteoarthritis, periprosthetic osteolysis, osteogenesis imperfecta,or metastatic bone disease.

[0125] The inhibitors of MST1/2 protein kinases or pharmaceuticalcompositions of the present invention may be effectivelyadministered at a plurality of times associated with tissue damage,including prior to actual damage, during various stages of damagedevelopment, and/or after damage has occurred. Specifically, theinhibitors of MST1/2 protein kinases or pharmaceutical compositionscan be used prophylactically to protect a subject against thedevelopment of tissue damage, such as from one minute to about 24hours, or from about five minutes to about 10 hours, or from aboutfive minutes to about five hours prior to onset of the tissuedamage. The inhibitors of MST1/2 protein kinases or pharmaceuticalcompositions can also be used after the development of tissuedamage for a short period of time, such as from one minute to about24 hours, or from about five minutes to about 10 hours, or fromabout five minutes to about five hours.

[0126] The inhibitors of MST1/2 protein kinases or pharmaceuticalcompositions can also be used after the development of tissuedamage for an extended period of time, such as days to weeks,months, or even years to prevent continuing primary or secondaryinjury. Finally, the inhibitors of MST1/2 protein kinases orpharmaceutical compositions can be used after the tissue damage hasbeen alleviated (or there is reason to believe it has beenimproved), in an attempt to prevent the onset of a similar tissuedamage in the future.

[0127] To treat a disease or disorder associated with tissuedamages, the inhibitors of MST1/2 protein kinases may beadministered through intravenous administration (by injection orinfusion or drip), subcutaneous injection, sublingualadministration, or oral administration to a subject at a dose inthe range of from about 1 to about 10 mg/kg bodyweight. In someembodiments, the dose may be from at least about 2, 3, 4, 5, 6, 7,8, and 9 mg/kg bodyweight up to about 10 mg/kg bodyweight. In someembodiments, the dose may be from about 1 mg/kg bodyweight up toabout 2, 3, 4, 5, 6, 7, 8, and 9 mg/kg bodyweight. In someembodiments, the dose may be from any one of about 2, 3, 4, 5, 6,7, 8, and 9 mg/kg bodyweight to any one of about 2, 3, 4, 5, 6, 7,8, and 9 mg/kg bodyweight.

[0128] In some embodiments, the dosage of the inhibitors of MST1/2protein kinases may be in the range of from about 0.1 to about 100mg/kg bodyweight. In some embodiments, the dosage of the inhibitorsof MST1/2 protein kinases may be in the range of from about 0.1 toabout 10 mg/kg bodyweight. In some embodiments, the dosage of theinhibitors of MST1/2 protein kinases may be in the range of fromabout 1 to about 100 mg/kg bodyweight. In some embodiments, thedosage of the inhibitors of MST1/2 protein kinases may be in therange of from about 10 to about s100 mg/kg bodyweight. In someembodiments, the dosage of the inhibitors of MST1/2 protein kinasesmay be in the range of from any one of about 10, 20, 30, 40, 50,60, 70, 80, 90 mg/kg bodyweight to any one of about 20, 30, 40, 50,60, 70, 80, 90, 100 mg/kg bodyweight.

[0129] In the embodiment where the inhibitors of MST1/2 proteinkinases are administered by intravenous infusion or intravenousinjection or intravenous drip, the entire dose of the inhibitorsmay be administered for a period of from about 0.3 hour to about 12hours, or from about 0.5 hour to about 10 hours, or from about 1hour to about 8 hours, or from 1 hour to about 6 hours, or fromabout 1 hour to about 4 hours.

[0130] In some embodiments, the inhibitors of MST1/2 proteinkinases may be administered before a tissue injury is expected tooccur, such as at the time when angina occurs in ischemic heartdisease patients, or before a cancer patient receiving cardiotoxicchemotherapy agents. In some embodiments, the inhibitors of MST1/2protein kinases may be administered at the time with injuryinducers when e a tissue injury is expected to occur, such as acancer patient receiving cardiotoxic chemotherapy agents. In someother embodiments, the inhibitors of MST1/2 protein kinases may beadministered after tissue injury, such as after stroke or acuteliver injury, or after inflammatory diseases have developed.

[0131] The inhibitors of MST1/2 protein kinases may be administeredone time or twice before the tissue injury is expected to occur.After the tissue injury has occurred, the inhibitors of MST1/2protein kinases may be administered four times per day, three timesper day, twice per day, or once every 1, 2, 3, 4, 5, 6, 7, 8, 9,10, 11, 12, 13, 14, 15, 16, 17, 18, 19, or 20 day(s), or at otherschedules, such as once every a few days.

[0132] In some embodiments, the inhibitors of MST1/2 proteinkinases are administered intravenously (by injection or infusion ordrip) or subcutaneously to a subject having or potentiallydeveloping tissue injury due to heart attack, cardiac injuryinduced by chemotherapy, stroke, acute liver damage induced byalcohols, acute nephritis, or nephrotoxicity induced by contrastimaging agent.

[0133] In some embodiments, the inhibitors of MST1/2 proteinkinases are administered sublingually or orally to a subject havingor potentially developing tissue injury due to heart failure, fattyliver diseases, sepsis, inflammatory diseases, and diabetes.

[0134] In some embodiments, the inhibitors of MST1/2 proteinkinases may be co-administered (either simultaneously or inseparate sequential administrations) with one or more furtheractive ingredients known to treat degenerative diseases/tissueinjuries. Preferably, if the administration is not simultaneous,the inhibitors of MST1/2 protein kinases and the further activeingredients are administered in a close time proximity to eachother, within half an hour, or 1 hour, or 2 hours, or 4 hours, or 6hours, or 8 hours, or 10 hours. Furthermore, the inhibitors ofMST1/2 protein kinases and the further active ingredients may beadministered in the same dosage form or different dosage forms,e.g. one compound may be administered intravenously and anothercompound may be administered orally.

[0135] Examples of further active ingredients for use incombination with the inhibitors of MST1/2 protein kinases includebut are not limited to: chemoprotective or myeloprotective agentssuch as G-CSF, BB10010 (Clemons et al., Breast Cancer Res.Treatment, 57, 127, 1999), amifostine (Ethyol) (Fetscher et al.,Current Opinion in Hemat., 7, 255-60, 2000), SCF, IL-11, MCP-4,IL-1-beta, AcSDKP (Gaudron et al., Stem Cells, 17,100-6, 1999),TNF-a, TGF-b, MIP-1a (Egger et al., Bone Marrow Transpl., 22(Suppl. 2), 34-35, 1998), and other molecules identified as havinganti-apoptotic, survival or proliferative properties.

[0136] Additional examples of further active ingredients for use incombination with the inhibitors of MST1/2 protein kinases includebut are not limited to: stem cell, megakaryocyte, neutrophilmobilizers such as chemotherapeutic agents (i.e., cytoxan,etoposide, cisplatin, Ballestrero A. et al., Oncology, 59:7-13,2000), chemokines, IL-8, Gro-beta (King, A. G. et al. J. Immun.,164:3774-82, 2000), receptor agonist or antagonist antibodies,small molecule cytokine or receptor agonists or antagonists, SCF,Flt3 ligand, adhesion molecule inhibitors or antibodies such as:anti-VLA-4 (Kikuta T. et al., Exp. Hemat., 28:311-7, 2000) oranti-CD44 (Vermeulen M. et al., Blood, 92:894-900, 1998),cytokine/chemokine/interleukin or receptor agonist or antagonistantibodies, MCP-4 (Berkhout T A., et al., J. Biol. Chem.,272:16404-16413, 1997; Uguccioni M. et al., J. Exp. Med.,183:2379-2384, 1996).

[0137] In some embodiments, further active ingredients for use incombination with the inhibitors of MST1/2 protein kinases alsoinclude, but are not limited to, carbamates (pyridostigmine,physostigmine, aminostigmine, neostigmine, synostigmine,Epastigmine, Mobam, decarbofuran), anticholingerics(trihexyphenidyle, benactyzine, Biperidene, Scopolamine, aprophen,atropine, hyoscin, adiphenine, Caramiphen, pentmethonium,Mecamylamine, Trihexyphenidyle) PANPAL, aminophenols (eseroline),organophosphates (TEPP, Paraxon, Ethyl-4-nitrophenylphosphate),tacrine, 7-MEO-TA, huperzine A, Cholinesterases (BuChE, AChE,triesterase, paraoxonase), oximes/reactivators (HI-6, PAM,Obidoxime, Trimedoxime, Methoxime, Hlo-7, BI-6, K048, K033,pralidoxime chloride (2-PAM Cl), P2S, TMB4, 2-PAMI), Suramine,Benzodiazepines, tubocurine, Memantine, Procyclidine, Nimodipin,Clonidine, pralidoxime, diazepam, enkephalins, phenylmethylsulfonylfluoride, natrium bicarbonate, vitamin E analogs(.alpha.-tocopherol succinate, .gamma.-tocotrienol), superoxidedismutase/catalase mimic (EUK189), selenium, benzyl styryl sulfone,truncated flagellin, statins, genistein, galantamine, hypothermia,5-androstenediol, CpG-oligodeoxynucleotides, antimicrobials, stemcell transplants, amifostine, Tempol, isoflavones, benzylsulfoneanalogs, GM-CSF, G-CSF, potassium iodide, aluminum hydroxide,Prussian blue, chelating agents (diethylenetriaminepentaacetate(Ca-DTPA), zinc diethylenetriaminepentaacetate (Zn-DTPA)),keratinocyte growth factor, intestinal peptide hormones, betaglucan, octreotide, pentoxifylline, angiotensin converting enzymeinhibitors, angiotensin II receptor blockers, methemoglobin formers(amyl nitrite, sodium nitrite), sodium thiosulfate, cobaltcompounds (hydroxycobalamin (Vitamin B12a), toxoids, antitoxins,vaccines, passive antibodies, chemotherapeutic agents including,but not limited to, methotrexate, taxol, mercaptopurine,thioguanine, hydroxyurea, cytarabine, cyclophosphamide, ifosfamide,nitrosoureas, cisplatin, carboplatin, mitomycin, dacarbazine,procarbizine, etoposides, campathecins, bleomycin, doxorubicin,idarubicin, daunorubicin, dactinomycin, plicamycin, mitoxantrone,asparaginase, vinblastine, vincristine, vinorelbine, pacl*taxel,and docetaxel; Radiation: .alpha.-radiation; Alkylating agents;Nitrogen mustards: cyclophosphamide, Ifosfamide trofosfamide,Chlorambucil; Nitrosoureas: carmustine (BCNU), Lomustine (CCNU),Alkylsulphonates busulfan, Treosulfan; Triazenes: Dacarbazine;Platinum containing compounds: Cisplatin carboplatin, PlantAlkaloids; Vinca alkaloids: vincristine, Vinblastine, Vindesine,Vinorelbine; Taxoids: pacl*taxel, Docetaxol; DNA TopoisomeraseInhibitors Epipodophyllins: etoposide, Teniposide, Topotecan,9-aminocamptothecin irinotecan (Campto.RTM.), crisnatol;Mytomycins: Mytomycin C, Mytomycin C; Anti-metabolites,Anti-folates: DHFR inhibitors: methotrexate, Trimetrexate; IMPdehydrogenase Inhibitors: mycophenolic acid, Tiazofurin, RibavirinEICAR; Ribonuclotide reductase inhibitors: hydroxyurea;deferoxamine; Pyrimidine analogs: Uracil analogs, 5-Fluorouracil,Floxuridine, Doxifluridine, Ratitrexed; Cytosine analogs:cytarabine (ara C) Cytosine arabinoside fludarabine; Purineanalogs: mercaptopurine, Thioguanine; Hormonal therapies; Receptorantagonists: Anti-estrogens, Tamoxifen, Raloxifene megestrol; LHRHagonists: goserelin, Leuprolide acetate; Anti-androgens: flutamide,bicalutamide; Retinoids/Deltoids Vitamin D3 analogs: EB 1089, CB1093, KH 1060; Photodyamic therapies: vertoporfin (BPD-MA),Phthalocyanine photosensitizer, Pc4 Demethoxy-hypocrellin A(2BA-2-DMHA) Cytokines: Interferon-.alpha., Interferon-.gamma.,Tumor necrosis factor; Isoprenylation inhibitors: Lovastatin;Dopaminergic neurotoxins: 1-methyl-4-phenylpyridinium ion; Cellcycle inhibitors: staurosporine; Actinomycins: Actinomycin D,Dactinomycin; Bleomycins: bleomycin A2, Bleomycin B2, Peplomycin;Anthracyclines: daunorubicin, Doxorubicin (adriamycin), Idarubicin,Epirubicin, Pirarubicin, Zorubicin, Mitoxantrone; MDR inhibitors:verapamil; Ca.sup.2+ ATPase inhibitors: thapsigargin; TNF-.alpha.inhibitors/thalidomide angiogenesis inhibitors3-(3,4-dimethoxy-phenyl)-3-(1-oxo-1,3-dihydro-isoindol-2-yl)-propionamide (SelCIDs.TM.) ImiDs.TM.,Revlimid.TM., Actimid.TM.. In another aspect of the presentinvention, a pharmaceutical composition according to the presentinvention may include an MST1/2 inhibitor in a formulation with atleast one small molecule that exhibits tissue protectivefunctionality. Suitable small molecules include, but are notlimited to, steroids (e.g., lazaroids and glucocorticoids),antioxidants (e.g., coenzyme Q.sub.10, alpha lipoic acid, andNADH), anticatabolic enzymes (e.g., glutathione peroxidase,superoxide dimutase, catalase, synthetic catalytic scavengers, aswell as mimetics), indole derivatives (e.g., indoleamines,carbazoles, and carbolines), nitric acid neutralizing agents,adenosine/adenosine agonists, phytochemicals (flavanoids), herbalextracts (ginko biloba and turmeric), vitamins (vitamins A, E, andC), oxidase electron acceptor inhibitors (e.g., xanthine oxidaseelectron inhibitors), minerals (e.g., copper, zinc, and magnesium),non-steriodal anti-inflammatory drugs (e.g., aspirin, naproxen, andibuprofen), and combinations thereof. Additionally agentsincluding, but not limited to, anti-inflammatory agents (e.g.,corticosteroids, prednisone and hydrocortisone), glucocorticoids,steroids, non-steriodal anti-inflammatory drugs (e.g., aspirin,ibuprofen, diclofenac, and COX-2 inhibitors), beta-agonists,anticholinergic agents and methyl xanthines), immunomodulatoryagents (e.g., small organic molecules, T cell receptor modulators,cytokine receptor modulators, T-cell depleting agents, cytokineantagonists, monokine antagonists, lymphocyte inhibitors, oranti-cancer agents), gold injections, sulphasalazine,penicillamine, anti-angiogenic agents (e.g., angiostatin),TNF-.alpha. antagonists (e.g., anti-TNF.alpha. antibodies), andendostatin), dapsone, psoralens (e.g., methoxalen and trioxsalen),anti-malarial agents (e.g., hydroxychloroquine), anti-viral agents,anti-histamines and antibiotics (e.g., erythromycin andpenicillin).

[0138] In some embodiments, the inhibitors of MST1/2 proteinkinases may be administered in conjunction with methods oftreatment such as chemotherapy, radiation therapy (x-ray radiation,high-energy megavoltage (radiation of greater that 1 MeV energy),electron beam, orthovoltage x-ray radiation, gamma-ray emittingradioisotopes (radioactive isotopes of radium, cobalt and otherelements)), hyperbaric chambers, heart bypass machine, angioplasty,hypothermia, surgery, angioplasty, etc. to to achieve additive,synergistic or offsetting (to counteract side effects of thetherapeutic method) benefits of the effects of the inhibitors ofMST1/2 protein kinases. As an example, in a specific embodiment,the inhibitors of MST1/2 protein kinases can be administered to apatient that has undergone chemotherapy or radiation therapy. Inanother specific embodiment, a chemotherapeutic agent or radiationtherapy is administered prior or subsequent to administration ofthe inhibitors of MST1/2 protein kinases, preferably at least anhour, two hours, three hours, five hours, 12 hours, a day, twodays, three days, or a week. The inhibitors of MST1/2 proteinkinases may be administered as a supplement to chemotherapy orradiation therapy where the chemotherapy or the radiation therapyhas proven or may prove too toxic, e.g., results in unacceptable orunbearable side effects, for the patient being treated. Theinhibitors of MST1/2 protein kinases may be administered,simultaneously with or following treatment with chemotherapy orradiation in an effort to prevent or ameliorate the toxic sideeffects of the treatment method.

Exemplary Diseases and Disorders

[0139] In verifying applications of the inhibitors of MST1/2protein kinases for these exemplary diseases and disorders, sixexemplary compounds were used:

##STR00112## ##STR00113##

[0140] The activity of these compounds in preventingphosphorylation of Mob1 is an important event in preventing cellapoptosis and inducing tissue/organ regeneration (FIG. 1). Thecompounds show dose-dependent inhibition of phosphorylation of Mob1(FIG. 2). The IC50 in inhibiting Mob1 phosphorylation through MST1and MST2 is in the nano-molar range. For example, one of thesecompounds has IC50 at 71.0.+-.15 nM for MST1 and 38.5.+-.6.2 nM forMST2 (FIG. 3). Thus, these six compounds have excellent activity ininhibiting MST1/2 and suppression of the Hippo pathway, which leadto expression of the target genes, including anti-apoptosis genes,cell division related genes and genes involved in inhibition ofcell differentiation.

A. Cardio Protection

[0141] The cardiomyocytes were isolated from neonatal rats andcultured using the following protocol: [0142] 1. Pre-sterilizeforceps, scissors, and beaker with a stir bar. Also preparesterilized dH.sub.2O ahead of time. [0143] 2. Coating 12 well ofPlate by SureCoat solution and leave at 37.degree. C. till use.[0144] 3. Prepare wash buffer: Prepare 2 of 100 mm dishes with1.times. cold PBS and 1 of 50 ml of tube with 1.times. cold PBS.Keep them cold. [0145] 4. Prepare D2: 28 ml of dH.sub.2O+20 ml ofCellutron D2+2 ml of EC; filter to sterilize. This is to be kept atroom temperature. [0146] 5. Prepare 10 ml of FBS (5 ml is enoughfor one isolation). [0147] 6. Sterilize neonates with 70% EtOH;Collect hearts into one dish with cold PBS, and remove atria, andtransfer hearts into the 2nd dish with cold PBS. Wash multipletimes with the 50 ml cold PBS (this can prevent contamination lateron). Transfer the hearts into enzymes. [0148] 7. Digestion is doneat 37.degree. C. in a small beaker with a stirrer bar(pre-sterilized) and stirrer at 150 rpm). Each digestion is 9 min/5ml (total 10 digestions), followed by pipetting to break downcells. The cell is to be pelleted at 1000 g for 1 min. Then cellsare collected into the tube of FBS until the end. [0149] 8. Pelletcells down and remove FBS, re-suspend in DMEM 11995+10% FBS+P/S.Use 50 ml for 10 neonatal rat hearts. [0150] 9. Filter through acell strainer is to remove the clumps. [0151] 10. Count cells withTrypan blue before pre-plating to get an idea how many live cellsthere would be, to get an idea how many plates should be coated(the actual cell # will be smaller, only .about.2/3 afterpre-plating at 37.degree. C. for 1.about.1.5 hrs). [0152] 11.Pre-plate in uncoated tissue culture dish (3 of 100 mm dishes/per10 neonatal rat hearts) in 370 C CO.sub.2 incubator for 1.about.1.5hrs. [0153] 12. Move unattached cells to sterile 50-mL tubes. Countlive cell density. [0154] 13. Plating: Adjust cells with culturemedium (DMEM+10% FBS+P/S)+100 .mu.M BrDU (To block fibroblasts fromgrowing up) to 10.sup.6 cell/mL, and plate cells in 6-well cellculture plates with 2.times.10.sup.6 cells/well. [0155] 14. Nextday, wash with DMEM with PPennecilin/Streptomycin without FBS toremove dead cells (around 2.about.3 times; check cell undermicroscope). [0156] 15. For cell live/death assay, add DMEM with10% FBS and 1% P/S or 1% FBS and 1% P/S, and several concentrationsof treatments according to experimental design. [0157] 16. For EdUassay use DMEM with 1% FBS and 1% P/S. Add 0.5 .mu.M EdU per wellexcept adding different concentration of compounds. [0158] 17.Medium and treatment need to be changed every day to preventcontamination if cells need culture more than one day. For EdUassay, change medium with same compounds and EdU and culture foranother 24 hr.

[0159] The isolated rat cardiomyocytes (neonatal rat cardiomyocytesor NRCM) were subjected to a live/died cell assay. Specifically,the isolated rat cardiomyocytes were incubated with differentconcentration of compounds (X1, X2, Y1 or Y2) at 37.degree. C. and5% CO.sub.2 for 1 hr. Chemical agents that are toxic to the cellswere then added to the incubated cells, including H.sub.2O.sub.2,isoproterenol (ISO), and doxorubicin (DOX). The cardiomyocytes withthe compounds and the chemical agents were further incubated at 370C and 5% CO.sub.2O/N for 24 hrs. The cardiomyocytes were stainedwith 0.4% Trypan Blue at room temperature for 15 min to estimatethe percentage of dead cells which were stained blue by Trypan blue(cell death rate). Images of the stained cells were taken for 3-5random fields per well for analysis of the death rate.

[0160] The MST1/2 inhibitors were found to significantly reducedeath rate of cardiomyocytes induced by 20 .mu.MH.sub.2O.sub.2(FIGS. 4A-4D), by 100 .mu.M ISO (FIGS. 5A-5D) or by0.2 .mu.M DOX (FIGS. 6A-6D). These experiments indicated that theinhibitors of MST1/2 protein kinases are effective in preventing ortreating injuries to cardiomyocytes induced by these toxicagents.

[0161] The apoptosis in cardiomyocytes induced by the toxic agents(H.sub.2O.sub.2, ISO and DOX) was measured by the apoptosisdetection assay using the DeadEnd.TM. Fluorometric TUNEL System bythe following protocol: [0162] 1. Cardiomyocytes were fixed byadding 4% PFA in PBS for 1 hr. at RT after incubated 24 hours.[0163] 2. Wash twice by 1.times. PBS, 5 minutes each time. [0164]3. Drain, and dry the wells completely. Rehydrate with 1.times.PBS.[0165] 4. Permeabilize cells by 0.2% Triton.RTM. X-100 in PBS for 5minutes. [0166] 5. Wash cells twice by 1.times. PBS, 5 minutes eachtime. [0167] 6. Equilibrate: Add 100 .mu.l Equilibration Buffer.Equilibrate at room temperature for 5-10 minutes. [0168] 7. Label:Add 50 .mu.l of TdT reaction mix to the cells on an area no largerthan 5 square centimeters. Do not allow cells to dry completely.Incubate plate for 60 minutes at 37.degree. C. in a humidifiedincubator; avoid exposure to light from this step forward. [0169]8. Stop Reaction by adding 2.times.SSC to well for 15 minutes.[0170] 9. Wash cells three times by 1.times. PBS, 5 minutes eachtime. [0171] 10. Stained nuclei by DAPI. [0172] 11. Analyze: Detectlocalized green fluorescence of apoptotic cells by fluorescencemicroscopy. Stained nuclei by DAPI will be blue. [0173] 12.Randomly take images of 5-10 field per well and save image foranalysis.

[0174] The apoptosis of the cardiomyocytes was determined bypercentages of Tunnel positive stain. As shown in FIGS. 7A-7C, thetoxic agents (H.sub.2O.sub.2, ISO and DOX) induced apoptosis in thecardiomyocytes. If the cardiomyocytes were pretreated with theinhibitors of MST1/2 protein kinases, the apoptosis wassignificantly reduced (FIGS. 7A-7C).

[0175] In vivo cardio protection in mice by the inhibitors ofMST1/2 protein kinases was verified by measuring heart function andatrophy induced by DOX (FIG. 8). Two inhibitors X1 (drug 1) and Y1(drug 2) were administered to the mice daily by i.p. during theentire study period. After administration of the inhibitors for sixdays, DOX was administered every two days by i.p. for four times.Blood samples were collected periodically from the mice byultra-sound. EdU was administered to the mice during the final fourdays of the study (FIG. 8).

[0176] The survival of the mice was summarized in FIG. 9, whichshowed that the inhibitors of MST1/2 protein kinases can delaydeath of mice induced by DOX. Further, the mice were disected atthe death or at the end of the study and the weight of the heartswas measured. The ratio of heart weight and body weight of the mice(mg/g) was presented in FIG. 10, which showed that the inhibitorsof MST1/2 protein kinases can reduce or prevent heart weight lossinduced by DOX. This indicated that the inhibitors of MST1/2protein kinases can reduce death and heart atrophy induced bychemotherapeutic agents such as DOX.

[0177] The heart function measured as EF % (ejection fraction) wasimpaired by DOX over the four-week study period. The inhibitors ofMST1/2 protein kinases X1 and Y1 reduced the EF % decrease that wasinduced by DOX by week four (FIG. 11).

[0178] Finally, the cardiomyocytes from the rat hearts wereincubated with EdU and DOX, optionally in combination with theinhibitors of MST1/2 protein kinases. The cardiomyocytes wereexamined for the rate of EdU incorporation into the cardiomyocytes,which indicated heart regeneration (cardiomyocyte divisions). Therate of EdU incorporation was measured by The Click-iT.RTM. PlusEdU Imaging Assay, using the following protocol: [0179] 1. Fromcell isolation and culture of step 16, cells were fixed by adding4% PFA in PBS for 1 hr. at room temperature after incubated 48hours. [0180] 2. Wash twice by 1.times. PBS, 5 minutes each time.[0181] 3. Drain, and dry the wells completely, then circle withimmunoEdge pen (to minimize the volume of reagents needed), let itdry. Rehydrate with 1.times.PBS. [0182] 4. Permeabilize cells by0.2% Triton.RTM. X-100 in PBS for 5 minutes. [0183] 5. Wash cellstwice by 1.times. PBS, 5 minutes+each time. [0184] 6. Blocking: 3%BSA/1.times.PBS, RT 30 min-1h. [0185] 7. Cool down centrifuge to 4C; spin primary antibody anti-.alpha. sarcomeric actin (MA121597,mouse IgM) at 10,000 g for 5 min at 4 C; dilute 1:100 in 3%BSA/1.times.PBS and add to cover the cells; incubate at 4.degree.C. O/N. [0186] 8. Wash cells twice by 1.times. PBS, 5 minutes eachtime from overnight incubated with primary antibody. Then continueEdU staining. (No light from here all the way down (fluorescentstaining). [0187] 9. Prepare 1.times.Click-iT EdU reaction buffer(component D) working solution by diluting component D 1:10 withdH.sub.2O. Add 2 ml dH.sub.2O to component F to result in 10.times.stock solution of the Click-iT EdU buffer additive (stored at-20.degree. C. in aliquots) (if it turns brown, it's bad; stillgood if light brown). Right before use, prepare 1.times. Click-iTEdU buffer additive by diluting the 10.times. solution 1:10 indH.sub.2O (prepare fresh and use right away). [0188] 10. PrepareClick-iT Plus reaction co*cktail as in the table below in the orderas listed (and use within 15 min):

TABLE-US-00006 [0188] Reaction components* 1 1X Click-iT .RTM.reaction buffer (prepared in step 9) 440 .mu.L Copperprotectant(Component E) 10 .mu.L Alexa Fluor .RTM. picolyl azide(Component B) 1.2 .mu.L Reaction buffer additive (prepared in step9) 50 .mu.L Total volume 500 .mu.L

[0189] Component B is fluorescent azides reacting with incorporatedEdU in newly synthesized DNA. [0190] 11. Add Click-iT reactionco*cktail to cover the cells; Incubate for 30 min at RT (no light).[0191] 12. Wash clean with 1.times.PBS. [0192] 13. Apply secondaryAnti-Mouse IgM antibody in 3% BSA/1.times.PBS for 1h at RT (A21043Invitrogen, 568 goat anti-mouse IgM; 1:1000)--add DAPI directlywithout washing, and incubate for another 10 min. Thorough washwith 1.times.PBS. [0193] 14. Check stain quality and take image byrandomly choosing 5-10 field per well and save the images foroffline analysis.

[0194] The rate of EdU incorporation into the cardiomyocytes waspresented in FIG. 12, which showed that significantly more EdU wasincorporated into cardiomyocytes when the inhibitors of MST1/2protein kinases were present.

[0195] Accordingly, these series of experiments verified that theinhibitors of MST1/2 protein kinases of the present invention areeffective in preventing or treating heart atrophy induced by toxicagents such as chemotherapeutic agent DOX.

B. Ischemic Stroke

[0196] To verify the protective effect of the inhibitors of MST1/2protein kinases on the brain injury after ischemic stroke, X1 wasadministered to mouse cerebral ischemia model induced by transientmiddle cerebral artery occlusion (MCAO). Briefly, 8-12 week-oldC57B1/6 mice (both male and female mice) were intraperitonealinjection with vehicle (DMSO) or 2 mg/kg X1 twice at 24 hours and 3hours prior to MCAO procedure respectively. Mice were thenanesthetized with 2.0% isoflurane, with their body temperaturemaintained at 37.degree. C. with a heating pad. Following a midlinecervical skin incision, the proximal right common carotid arteryand the right external carotid artery were ligated. A 6-0 siliconrubber-coated monofilament (6023910PK10; Doccol, Sharon, Mass.) wasinserted via the right internal carotid artery to occlude theorigin of the right middle cerebral artery. The suture was left inplace for 60 minutes to cause ischemia and then removed to allowfor reperfusion.

[0197] Real-time cerebral blood flow was monitored using a laserspeckle contrast imager (PeriCam PSI HR System, Perimed, Sweden) toconfirm occlusion of the MCAO and reperfusion after removal of thefilament. After 23 hours of reperfusion, mice were then sacrificedand their brains were harvested and sectioned into 1 mm sections.Brain sections were stained with 2% 2,3,5-Triphenyltetrazoliumchloride (T8877; Sigma-Aldrich, St. Louis, Mo.) and scanned (HPScanjet G4010) to allow quantification of infarcted and uninjuredbrain tissue. Statistical analysis of the data was performed usingPrism 6 (GraphPad). Student's t test (mean.+-.standard error of themean) was used to determine statistical significance. P<0.05 wasregarded as statistically significant. As shown in FIGS. 13 and14A, X1 significantly attenuated the infarct size in the cerebralischemia model mice.

[0198] In addition, neurological function of the MCAO-inducedcerebral ischemia model mice was evaluated after treatment with theMST1/2 protein kinases inhibitor X1. Neurological function wasevaluated using a 0-4-point neurological score: 0=no neurologicaldysfunction; 1=failure to extend left forelimb fully when lifted bytail: 2=circling to the contralateral side; 3=falling to the left;4=no spontaneous walk or in a comatose state, or barrel rolling.All scores were performed while being blinded to the study groups.The MST1/2 protein kinases inhibitor X1 demonstrated the capabilityof reducing the damages to neurological functions in the cerebralischemia model mice, in comparison with vehicle treatment (FIG.14B).

[0199] Accordingly, the inhibitors of MST1/2 protein kinases of thepresent invention are effective in preventing or treating ischemicstroke by reducing the infarct size and preserving neurologicalfunctions.

C. Inflammatory Diseases

[0200] The applications of the inhibitors of MST1/2 protein kinasesin preventing or treating inflammatory diseases were verified byinhibiting expression of cell adhesion molecules and/orinflammatory cytokines.

[0201] Arterial recruitment of inflammatory cells from thecirculation and their transendothelial migration are key events inthe early phase of cardiovascular and inflammatory diseases such asatherosclerosis, restenosis, heart failure and ischemic stroke. Inresponse to several inflammatory stimuli, such as tumor necrosisfactor (TNF)-.alpha. and interleukin-1.beta. (IL)-1.beta.,endothelial cells (ECs) undergo inflammatory activation, resultingin an increased surface expression of cell adhesion molecules, suchas intercellular adhesion molecule (ICAM)-1, vascular cell adhesionmolecule (VCAM)-1, and E-selectin, which contributes to recruitmentof inflammatory cells to arterial wall and their transmigrationacross the arterial wall. Genetic deficiencies of adhesionmolecules in mice are associated with decreased atherosclerosis.Therefore, modulation of the expression of adhesion molecules onECs is an important indicator for the prevention and treatment ofinflammatory diseases such as atherosclerosis.

[0202] The effect of inhibitors of MST1/2 protein kinases (X1 andY1) on the expression of cell adhesion molecules in vascularendothelial cells was measured. First, human umbilical vascular ECs(HUVECs) were pretreated with 1 .mu.M MST1/2 inhibitors (X1 and Y1)for 1 hour and then stimulated with TNF-.alpha. (20 ng/mL) andIL-1.beta. (10 ng/mL) for 12 hours. The expression of VCAM-1 andICAM-1 was detected by western blot. The expression of celladhesion molecules was reduced by the inhibitors of MST1/2 proteinkinases. For example, as shown in FIG. 15A, X1 at the concentrationof 1 .mu.M substantially inhibited VCAM-1 expression, while theexpression of ICAM-1 was barely affected. In addition, X1 inhibitsVCAM-1 expression in a dose dependent manner with an IC50 of 1.29.mu.M (FIG. 15B). Likewise, Y1 had a similar inhibitor effect onthe expression of VCAM-1 in HUVECs, with an IC50 of 0.49 .mu.M(FIGS. 16A-16B). Furthermore, both compounds X1 and Y1 markedlyinhibited the mRNA levels of inflammatory cytokines in HUVEC, suchas MCP-1 and IL-6, in response to TNF.alpha. stimulation, asdetermined by qPCR (FIGS. 17A-17D and 18A-18D).

[0203] To substantiate the functional significance of inhibitors ofMST1/2 protein kinases in activating endothelial cells, themonocyte adhesion to the activated endothelial cells was furtherexamined. As shown in FIGS. 19A-19B and 20A-20B, the adhesion ofTHP-1 cells to TNF.alpha.-activated ECs was markedly inhibited byboth X1 and Y1, indicating that inhibitors of MST1/2 proteinkinases exerted potent anti-inflammatory effects in ECs.

[0204] Cell adhesion was assessed as the follows: THP1 cells werelabeled with calcein-AM (Invitrogen) according to the instructionsof the manufacturer. After the HUVECs were stimulated and washed,2.5.times.105 calcein-labeled THP1 cells were added to each welland allowed to interact with MST1/2 inhibitors for 60 minutes at370.degree. C. Unbound cells were removed by gently washing withcomplete medium, and the number of attached cells was counted on aninverted fluorescent microscope.

[0205] The therapeutic capacity of inhibitors of MST1/2 proteinkinases for inflammatory diseases was also demonstrated by a murineseptic model by injecting mice with LPS (2.5 mg/kg, i.p.). In thisregard, the mice were pretreated with X1 twice at 24 hrs and 3 hrs,respectively, prior to the injection of lipopolysaccharides (LPS).6 hrs after injection of LPS to induce the sepsis model, theexpression of inflammatory cytokines in the mouse lung wasdetermined by real-time PCR. As shown in FIGS. 21A-21D, X1 markedlysuppressed the mRNA levels of ICAM-1, VCAM-1, MCP-1 and IL-6,indicating that X1 has an in vivo anti-inflammatory effect.

[0206] Accordingly, the inhibitors of MST1/2 protein kinases of thepresent invention are effective in preventing or treatinginflammatory diseases, such as sepsis.

Other Diseases and Disorders

[0207] In addition to these exemplary diseases and disordersdescribed above, utility of the inhibitors of MST1/2 proteinkinases in preventing or treating other diseases and disorders maybe determined or confirmed by various assays. For example, thetissue protective activity of the MST1/2 inhibitors may beconfirmed using various assays known in the art and disclosedwithin U.S. Patent Publication Nos: US 2009/0136519A1, US2004/0122216A1, and US 2009/0221482A1. Additionally, one ofordinary skill in the art will recognize that the inhibitors ofMST1/2 protein kinases' ability to prevent, mitigate or treat adisease or disorder associated with tissue damage or damage,effects or symptoms resulting therefrom may be confirmed throughvarious assays both in vitro and in vivo, although in certainembodiments in vivo assays may be preferred.

D. Tissue Protective Assays and Models

[0208] The inhibitors of MST1/2 protein kinases of the presentinvention exhibit tissue protective properties, i.e.anti-apoptotic, neuritogenic, neuroprotective, anti-cachectic,anti-inflammatory etc. These inhibitors may be tested for tissueprotective activity, e.g., protecting cells, tissues or organs.Protective activities may be further tested using in vitro and invivo assays. In vitro tests that are indicative of tissueprotective activity include, for example, cell proliferationassays, cell differentiation assays, or detecting the presence ofproteins or nucleic acids upregulated by tissue protective receptorcomplex, e.g. tissue protective cytokine receptor complex,activity, e.g., nucleolin, neuroglobin, cytoglobin, or frataxin.Neuroglobin, for example, may be involved in facilitating thetransport or the short-term storage of oxygen. Therefore, oxygentransport or storage assays may be used as an assay to identify orscreen for compounds which modulate tissue protective activity.

[0209] Neuroglobin is expressed in cells and tissues of the centralnervous system in response to hypoxia or ischemia and may provideprotection from injury (Sun et al. PNAS, 98:15306-15311, 2001;Schmid et al., J. Biol. Chem., 276:1932-1935, 2003, each of whichis incorporated by reference herein in its entirety). Cytoglobinmay play a similar role in protection of tissues or organs, but isexpressed in a variety of tissues at varying levels (Pesce et al.,EMBO, 3:1146-1151, 2002, which is incorporated by reference hereinin its entirety). In one embodiment of the invention, the levels ofan upregulated protein in a cell may be measured before and aftercontacting the MST1/2 inhibitors to a cell. In certain embodiments,the presence of an upregulated protein associated with tissueprotective activity in a cell, may be used to confirm the tissueprotective activities of the MST1/2 inhibitors.

[0210] Nucleolin may protect cells from certain damage. It playsnumerous roles in cells including modulation of transcriptionprocesses, sequence specific RNA-binding protein, cytokinesis,nucleogensis, signal transduction, apoptosis induced by T-cells,chromatin remodelling, or replication. It can also function as acell surface receptor DNA/RNA helicase, DNA-dependent ATPase,protein shuttle, transcription factor component, or transcriptionalrepressor (Srivastava and Pollard, FASEB J., 13:1911-1922, 1999;and Ginisty et al., J. Cell Sci., 112:761-772, 1999, each of whichis incorporated by reference herein in its entirety).

[0211] Frataxin is a protein involved with mitochondrial ironmetabolism and has previously been shown to be stronglyup-regulated by EPO both in vivo and in vitro (Sturm et al., Eur JClin Invest 35: 711, 2005, which is incorporated by referenceherein in its entirety).

[0212] Expression of a protein may be detected by detecting mRNAlevels corresponding to the protein in a cell. The mRNA can behybridized to a probe that specifically binds a nucleic acidencoding the upregulated protein. Hybridization may consist of, forexample, Northern blot, Southern blot, array hybridization,affinity chromatography, or in situ hybridization. The mRNAexpression levels can be quantitated with new generations ofRNA-seq techniques.

[0213] Tissue protective activity of the inhibitors of MST1/2protein kinases can also be detected using in vitro neuroprotectionassays. For example, primary neuronal cultures may be prepared fromnew born rat hippocampi by trypsinization, and cultured as by anymethod known in the art and/or described herein e.g. in MEM-IIgrowth medium (Invitrogen), 20 mM D-glucose, 2 mM L-glutamine, 10%Nu-serum (bovine; Becton Dickinson, Franklin Lakes, N.J.), 2% B27supplement (Invitrogen), 26.2 mM NaHCO.sub.3, 100 U/ml penicillin,and 1 mg/ml streptavidin (see, e.g., Leist et al., Science305:239-242, 2004, hereby incorporated by reference in itsentirety). One day after seeding, 1 .mu.Mcytosinearabino-furanoside is added. 13 day old cultures are thenpreincubated with increasing doses of the MST1/2 inhibitors ofinterest (3-3000 .mu.M) for 24 h. On day 14, the medium is removedand the cultures challenged with 300 .mu.M NMDA in PBS at roomtemperature (RT). After 5 min, pre-conditioned medium is returnedto the cultures which are then returned to the incubator for 24 h.The cells are fixed in paraformaldehyde, stained by Hoechst 33342(Molecular Probes, Eugene, Oreg.) and condensed apoptotic nucleimay be counted. NGF (50 ng/ml) and MK801 (1 .mu.M) are included aspositive controls.

[0214] Animal model systems can be used to demonstrate the tissueprotective activity of the inhibitors of MST1/2 protein kinases todemonstrate their safety and efficacy for different types of tissuedamage, disease, condition, or syndrome of interest. Animal modelsfor various diseases and disorders are known in the art. Forexample, protection against the onset of acute experimentalallergic encephalomyelitis in Lewis rats, restoration or protectionfrom diminished cognitive function in mice after receiving braintrauma, cerebral ischemia ("stroke") or seizures stimulated byexcitotoxins (Brines et al., PNAS, 97:10295-10672, 2000, which isincorporated by reference herein in its entirety), protection frominduced retinal ischemia (Rosenbaum et al., Vis. Res. 37:3443-51,1997, which is incorporated by reference herein in its entirety),protection from injury to the sciatic nerve, and protection fromischemia-reperfusion injury to the heart (in vitro cardiomyocytestudies and in vivo ischemia-reperfusion injury, see, e.g.,Calvillo et al., PNAS 100:4802-4806, 2003, and Fiordaliso et al.,PNAS 102:2046-2051, 2005, each of which is hereby incorporated byreference in its entirety). Animal models related to spinal cordinjury, ischemic stroke, peripheral nerve damage, wounds, or damageto the heart, eyes, kidneys, etc. are also known in the art. Suchassays are described in further detail in Grasso et al. Med SciMonit 10: BR1-3, 2004, PCT publication Nos. WO 2002/053580 and WO2007/019545, each of which is incorporated by reference herein inits entirety.

E. Assays for Specific Indications

[0215] Specific indications for the inhibitors of MST1/2 proteinkinases may be verified by various assays. Two exemplaryindications are provided for illustrative purposes. One ispreventing or treating tissue damages induced by a toxic agent.Another is preventing or treating tissue damages associated withinflammation.

[0216] A variety of assays known in the art may be used todetermine the inhibitors of MST1/2 protein kinases' ability toprevent, treat, ameliorate, or manage damage, effects or symptomsresulting from exposure to a toxic agent. In general, this isaccomplished by selecting an appropriate cell line, subjecting thatcell to a toxic agent of interest and treating a portion of thecells with an inhibitor of MST1/2 protein kinases and determiningthe cells survival or response in the presence of the toxic agentand the inhibitor of MST1/2 protein kinases of interest. If thecell exhibits improved survival or a reduction of damage, effectsor symptoms in the presence of the inhibitor of MST1/2 proteinkinases, the inhibitor of MST1/2 protein kinases can be consideredto be a possible therapeutic for toxic exposure. Further one ofordinary skill in the art will recognize that the inhibitors ofMST1/2 protein kinases' protective ability can be evaluated bytreating the cells with the inhibitors of MST1/2 protein kinasesprior to the toxic agent challenge.

[0217] Exemplary cell lines for testing protective ability againsttissue damages induced by a chemical agent include: a) skin celllines such as J-774 (mouse macrophage derived cell line), CHO-K1(strain of epithelial cell line derived from Chinese hamster ovarycells), and HeLa (human cervical carcinoma) (Sawyer, et al.,Eplasty, 8:e25, 2008); b) corneal cell lines for vesicant agents(Amir, et al., Proceedings of the U.S. Army Medical DefenseBioscience Review, Aberdeen Proving Ground, MD (2004)); c)macrophages (Amir, et al., J Appl Toxicol, 20 Suppl 1:S51-8, 2000);d) upper respiratory tract cell lines (Andrew and Lindsay, Hum ExpToxicol 17(7):387-95, 1998; Calvet et al., Hum Exp Toxicol18(2):77-81, 1999; Langford, et al., Hum Exp Toxicol 15(8):619-24,1996); e) skin models (Blaha et al., J Appl Toxicol 20 Suppl1:S101-8, 2000) Exemplary cell lines for testing protective abilityagainst tissue damages induced by a radiation agent include: a)endothelial cells (Abderrahmani, et al., Radioprotection 2008, vol43, no. 5, 2008), b) neuroimmune cells (afferent nerves, entericsensory nerves, mast cells) (Wang, et al., British Journal ofRadiology, 80:S41-S48, 2007), c) blood or lymphocyte cultures(Lloyd, et al., Phys Med Biol 18(3):421-31, 1973; Lloyd, et al.,Mutat. Res. 179(2):197-208, 1987; Blakely et al., Stem Cells 13(Suppl 1):223-30, 1995; Gotoh et al., Int. J. Radiation. Biol.81(1):33-40, 2005).

[0218] Further, suitable in vivo assays are known in the art forevaluating the effect of tissue protection after toxic agentexposure. Animal models using rats, mice, guinea pigs, rabbits,pigs, sheep, ferrets, dogs and non-human primates are contemplatedas well as transgenic animals that are particularly susceptible toa toxic agent (CD46 mice). In particular, exemplary animal modelsfor testing protective ability against tissue damages induced bychemical agents include: (1) Reid, Sulfur mustard induced skinburns in weanling swine evaluated clinically andhistopathologically, Journal of applied toxicology,20(S1):5153-5160, 2001; (2) Isidore, et al., A dorsal model forcutaneous vesicant injury 2-chloroethyl ethyl sulfide using c57b1/6mice, Cutaneous and ocular toxicology, 26(3):265-276, 2007; (3)Kassa, et al., The Choice: HI-6, pradoxime or Obidoxime againstNerve Agents?, www.asanite.com/ASANews-97/Antidot-Choice.html, (4)Shih, et al., Organophosphorus nerve agents-induced seizures andefficacy of atropine sulfate as anticonvulsant treatment,Pharmacol-Biochem-Behav. 64(1):147-53, 1999, (5) Luo, et al.,Comparison of oxime reactivation and aging of the nerveagent-inhibited monkey and human acetylcholinesterases,Chemico-Biological Interactions, 175(1-3):261-266, 2008.

[0219] Exemplary animal models for testing protective abilityagainst tissue damages induced by radiation agents include: (1)Blakely et al., In Vitro and Animal Models of Partial-Body DoseExposure: Use of Cytogenic and Molecular Biomarkers for Assessmentof Inhom*ogeneous Dose Exposures and Radiation Injury, PB-Rad-Injury2008 Workshop, May 5-6, 2008 AFRRI, Bethesda, Md.; (2) Augustine,et al., Meeting Report: Animal Models of Radiation Injury,Protection and Therapy, Radiation Research 164:100-109, 2005; (3)Houchen, et al. Prosurvival and antiapoptotic effects of PGE.sub.2in radiation injury are mediated by EP.sub.2 receptor in intestine,Am J Physiol Gastrointest Liver Physiol, 284:G490-G498, 2003; (4)Chen, Animal Models for Acquired Bone Marrow Failure Syndromes,Clinical Medicine & Research 3(2):102-108, 2005.

[0220] Additionally, various in vitro models of inflammation may beused to evaluate inhibitors of MST1/2 protein kinases' ability toprotect or treat the damage, symptoms, or effects of inflammationon the body. Initially, the ability of the inhibitors of MST1/2protein kinases to modulate an inflammatory mediator (e.g., ICAMand VCAM) can be confirmed by measuring the levels of theinflammatory mediator in an inflammatory assay after treatment withthe inhibitors of MST1/2 protein kinases, including but not limitedto, ELISA, cytometric bead array analysis, high-sensitivity andimmunonephelometric assays. For example, to determine if theinhibitors of MST1/2 protein kinases modulate either TNF.alpha. orIL-1, a murine model of LPS-mediated cytokine production would beperformed. Some mice in the murine model would be pretreated withthe inhibitors of MST1/2 protein kinases and then challenged withLPS while others would be saline treated. Blood would then becollected and the TNF.alpha. and IL-1 levels in the blood could bedetermined by an ELISA kit (OPT-EIA mouse TNF.alpha. and IL-1 ELISAkits from BD Biosciences). If the TNF.alpha. levels in the treatedanimals are lower than the TNF.alpha. levels in the saline treatedanimals then the inhibitors of MST1/2 protein kinases could beconsidered to modulate TNF.alpha.. Preferably, the inhibitors ofMST1/2 protein kinases would be tested for its ability to modulatemore than one inflammatory mediator, and more preferably it wouldbe a mediator other than or in addition to TNF.alpha., and mostpreferably it would be histamine. Similarly, the inhibitors ofMST1/2 protein kinases may be tested in additional in vitro assaysincluding, but not limited to, those disclosed in Lopata, CurrentAllergy & Clinical Immunology, 19:18-20, 2006, (histamine andtryptase assays), and Arulmozhi et al., Indian Journal ofPharmacology, 37:96-102, 2005, (5-lipoxygenase (5-LO),cyclo-oxygenase (COX), Leukotrine B4 (LTB4) and nitric oxidesynthase (NOS)).

[0221] Further, in vivo assays of inflammation may be useful inevaluating the inhibitors of MST1/2 protein kinases' utility forpreventing or treating inflammation. In vivo assays include, butnot limited to, murine EAE models, those utilizing transgenic micesuch as MDBiosciences DSS IBD murine model of severe colitis, theMDBioscience TNBS IBD murine model of inflammatory bowel disease,models involving IL-1 knockout mice disclosed within U.S. Pat. No.6,437,216, or models of transgenic mice involving TNF.alpha. asdisclosed within Probert et al., PNAS, 92:11294-11298, 1995,Kontoyiannis et al., Immunity 10:387-398, 1999, Keffer et al., EMBOJ. 10(13):4025-31, 1991, or models using chemical or syntheticchallenges to induce the inflammation such as models of asthma andchronic obstructive pulmonary disease disclosed in JPET307:373-385, 2003, adjuvant arthritis models as disclosed in EP 1777 234; murine LPS shock models, murine LPS lung models, acute pawinflammation models, or histidine challenge wheal formation modelas known in the art.

[0222] Further, the efficacy of the inhibitors of MST1/2 proteinkinases in humans may be verified by using well-known clinicalstudies such as the skin prick test and bronchoprovocation testdisclosed in Ravensberg et al., Clinical and Experimental Allergy,37:100-107, 2007; asthma studies as disclosed in Diamant et al.,Respiratory Medicine, 102:332-338, 2008, or nasal allergenchallenge as disclosed in Boot et al. Allergy, 62:378-384,2007.

[0223] In the present application, the invention has been describedwith reference to specific embodiments thereof. It will, however,be evident that various modifications and changes may be madethereto without departing from the broader spirit and scope of theinvention. The specification and drawings are, accordingly, to beregarded in an illustrative rather than a restrictive sense. Allcitations (e.g., scientific journal publications, patents, andother reference material) mentioned herein are hereby incorporatedherein by reference to the same extent as if each individualcitation was specifically and individually indicated to beincorporated by reference.

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References

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