Methods and compositions related to modulators of annexin and cartilage homeostasis

Abstract

The present invention provides a method of treating a subject with arthritis or an arthritic disease or preventing arthritis or arthritic disease in a subject, comprising administering to the subject a therapeutically effective amount of an agent that attenuates annexin function. Also provided are various methods of screening for agents and genes that increase collagen synthesis, decrease collagen degradation, or reduce or delay apoptosis, and methods of using the identified agents or nucleic acids for attaining or maintaining cartilage homeostasis, promoting cartilage repair, increasing collagen synthesis, decreasing collagen degradation, or reducing or delaying apoptosis. The invention also provides methods of decreasing mineralization of vesicles derived from chondrocytes and of promoting endochondral bone growth using agents that attenuates annexin function. The present invention also provides a composition comprising isolated chondrocytes, an agent that attenuates annexin function, and collagen fragments or other means for challenging chondrocytes, including for example, collagen fragments, collagen peptides, or immune mediators.

What is claimed is:  
     
         1 . A method of treating a subject with arthritis or arthritic disease or preventing arthritis or arthritic disease in a subject, comprising administering to the subject a therapeutically effective amount of an agent that attenuates annexin function.  
     
     
         2 . The method of  claim 1 , wherein the attenuated annexin function is a function of an annexin that binds collagen.  
     
     
         3 . The method of  claim 1 , wherein the annexin binds type II collagen.  
     
     
         4 . The method of  claim 3 , wherein the annexin that binds type II collagen is annexin V or annexin X.  
     
     
         5 . The method of  claim 1 , wherein the treatment or prevention is effected by increasing collagen synthesis or decreasing collagen degradation.  
     
     
         6 . The method of  claim 1 , wherein the agent has the structure I:  
       
         
           
           
               
               
           
         
         wherein y is from 1 to 4, wherein each R 1  is, independently, hydrogen, a branched or straight chain alkyl group, an alkenyl group, an alkynyl group, a branched or straight chain alkoxy group, an aryl group, an aralkyl group, a cycloalkyl group, an ester group, a substituted or unsubstituted amino group, a cyano group, an amide group, a nitro group, a hydroxy group, a halo group, a thio group, or a trihalomethyl group;  
         R 2  and R 6  are, independently, hydrogen, hydroxy, or branched or straight chain alkyl;  
         R 3  is hydrogen, a branched or straight chain alkyl group, or a substituted or unsubstituted aryl group;  
         R 4  is hydrogen, a branched or straight chain alkyl group, an acyl group, a cycloalkyl group, oxygen, or a group having the structure II  
         
           
             
             
                 
                 
             
           
         
         wherein W is carbon or nitrogen; Z is oxygen or H 2 ; n is 1 or 2; and R 7  is a branched or straight chain alkyl group, a branched or straight chain alkoxy group, an aryl group, an aralkyl group, a cycloalkyl group, or a heteroaryl group;  
         R 5  is A-R 10  or R 10 , wherein A is a C 1-4  branched or straight chain alkyl group, a hydroxyalkyl group, an acyl group, an amino group, an amide group, an ester group, a keto group, a substituted or unsubstituted aryl group, a substituted or unsubstituted heteroaryl group, a sulfonamide group, or a combination thereof; or  
         R 5  and R 6  are collectively ═C(H)R 10 ;  
         wherein R 10  is substituted or unsubstituted aryl, or substituted or unsubstituted heteroaryl;  
         V is hydrogen; an aryl group, a heteroaryl group, an alkoxy group, or an alkenyloxy group;  
         X is oxygen, sulfur, hydrogen, an aryl group, a heteroaryl group, an alkoxy group, an alkenyloxy group, or NR 8 , wherein R 8  is hydrogen, a branched or straight chain alkyl group, a substituted or unsubstituted aryl group, or a substituted or unsubstituted heteroaryl group; or  
         Y is carbon, oxygen, sulfur, a sulfone group, a sulfoxide group, or NR 9 , wherein R 9  is hydrogen, a branched or straight chain alkyl group, an alkenyl group, an alkynyl group, a cycloalkyl group, an ester group, an amino group, an amide group, a cyano group, or a trihalomethyl group;  
         wherein when bond a is a double bond, then R 3  is present and R 2  is not present; or when bond a is a single bond, then R 2  and R 3  are present;  
         wherein when bond c is a double bond, then R 5  is present and R 6  is not present; or when bond c is a single bond, then R 5  and R 6  are present;  
         wherein bonds a and c are not simultaneously double bonds;  
         wherein when bonds b, d, and e are present, then R 3 -E-R 4  is a substituted or unsubstituted alkylene group, or a substituted or unsubstituted alkylene group containing at least one heteroatom;  
         wherein when bond f is a double bond, then bond i and V are not present; or when bond f is a single bond, then bond i is a single bond and V is present;  
         wherein when bond f is a single bond or a double bond, then bonds g and h are not present; or when bond f is a single bond or a double bond, and bonds g and h are present, then F is a substituted or unsubstituted alkylene group, or a substituted or unsubstituted alkylene group containing at least one heteroatom;  
         and a pharmaceutically acceptable salt thereof.  
       
     
     
         7 . The method of  claim 6 , wherein Y is NR 9 , wherein R 9  is a branched or straight chain alkyl group.  
     
     
         8 . The method of  claim 6 , wherein Y is carbon.  
     
     
         9 . The method of  claim 7 , wherein bond f is a double bond; bonds g and h are not present; and X is oxygen.  
     
     
         10 . The method of  claim 7 , wherein R 5  is a group having the structure III 
 wherein R 13 -R 15  are, independently, hydrogen, a branched or straight chain                          alkyl group, an acyl group, a cycloalkyl group, or an aryl group; and    x is from 1 to 4, wherein each R 16  is, independently, hydrogen, a branched or straight chain alkyl group, an alkenyl group, an alkynyl group, a branched or straight chain alkoxy group, an aryl group, an aralkyl group, a cycloalkyl group, an ester group, a substituted or unsubstituted amino group, a cyano group, an amide group, a nitro group, a hydroxy group, a halo group, a thio group, or a trihalomethyl group.    
     
     
         11 . The method of  claim 7 , wherein R 5  has the structure IV  
       
         
           
           
               
               
           
         
       
     
     
         12 . The method of  claim 7 , wherein bond a is a double bond and bond c is a single bond.  
     
     
         13 . The method of  claim 7 , wherein y is 4 and each R 1  is hydrogen, a branched or straight chain alkyl group, an alkenyl group, an alkynyl group, a branched or straight chain alkoxy group, an aryl group, an aralkyl group, a cycloalkyl group, an ester group, a substituted or unsubstituted amino group, a cyano group, an amide group, a nitro group, a hydroxy group, a halo group, a thio group, or a trihalomethyl group.  
     
     
         14 . The method of  claim 7 , wherein R 3  comprises a substituted or unsubstituted phenyl group.  
     
     
         15 . The method of  claim 7 , wherein R 4  is a branched or straight chain alkyl group or an acyl group.  
     
     
         16 . The method of  claim 6 , wherein bonds a and f are double bonds; bond c is a single bond; bonds g and h are not present; X is oxygen; Y is NR 9 ; y is 4; each R 1  is hydrogen; R 3  comprises a substituted or unsubstituted phenyl group; R 4  is a branched or straight chain alkyl group or an acyl group; and R 5  has the structure III  
       
         
           
           
               
               
           
         
         wherein R 13 -R 15  are, independently, hydrogen, a branched or straight chain alkyl group, an acyl group, a cycloalkyl group, or an aryl group;  
         x is from 1 to 4, wherein each R 16  is, independently, hydrogen, a branched or straight chain alkyl group, an alkenyl group, an alkynyl group, a branched or straight chain alkoxy group, an aryl group, an aralkyl group, a cycloalkyl group, an ester group, a substituted or unsubstituted amino group, a cyano group, an amide group, a nitro group, a hydroxy group, a halo group, a thio group, or a trihalomethyl group.  
       
     
     
         17 . The method of  claim 1 , wherein the agent is 3-(R,S)-(L-tryptophanyl)-1,3-dihydro-1-methyl-5-phenyl-2H-1,4-benzodiazepine-2-one.  
     
     
         18 . The method of  claim 6 , wherein Y is sulfur.  
     
     
         19 . The method of  claim 18 , wherein V and X are hydrogen.  
     
     
         20 . The method of  claim 19 , wherein bonds g and h are not present, and bond f is a single bond.  
     
     
         21 . The method of  claim 18 , wherein R 4  has the structure II.  
     
     
         22 . The method of  claim 21 , wherein in structure II, W is nitrogen; Z is oxygen; n is 2, and R 7  is CH 2 Ph.  
     
     
         23 . The method of  claim 18 , wherein bonds a and c are single bonds, and bonds d and e are not present.  
     
     
         24 . The method of  claim 18 , wherein R 1  is branched or straight chain alkoxy.  
     
     
         25 . The method of  claim 6 , wherein bonds a, c, and f are single bonds; bonds d, e, g and h are not present; Y is sulfur; R 1  is branched or straight chain alkoxy; and R 4  has the structure II.  
     
     
         26 . The method of  claim 1 , wherein the agent is 4-(3-(1-(4-benzyl)piperidinyl)propionyl)-7-methoxy-2,3,4,5-tetrahydro-1,4-benzothiazepine.  
     
     
         27 . The method of  claim 1 , wherein the agent is not 1,3-dihydro-1-methyl-5-phenyl-2H-1,4-benzodiazepine-2-one.  
     
     
         28 . The method of  claim 1 , wherein the agent comprises a benzodiazepine compound, a benzothiazepine compound, or a combination thereof.