Methods for preventing or reducing ischemia/reperfusion induced myocardial cell death

Abstract

Provided herein are methods relating to inhibition of PUMA function for preventing or reducing myocardial cell death. A method for preventing or reducing ischemia/reperfusion induced myocardial cell death in a human is provided as are methods for identifying pharmaceuticals which interfere with PUMA function. Also herein provided is a method for reducing stem cell death in stem cell explants wherein the explants are intended to restore cardiac efficiency following cell death in response to coronary infarct.

1 . A method for preventing or reducing ischemia/reperfusion induced myocardial cell death in a human, the method comprising administering to a human a composition effective to inhibit PUMA function in cardiomyocytes in the human.  
     
     
         2 . The method of  claim 1  wherein the composition is administered during or subsequent to an ischemic attack.  
     
     
         3 . The method of  claim 1  wherein the composition is administered to an individual at risk for an ischemic attack.  
     
     
         4 . The method of  claim 1  wherein the composition downregulates PUMA expression.  
     
     
         5 . The method of  claim 1  wherein the composition inhibits binding of PUMA to Bcl-2.  
     
     
         6 . The method of  claim 1  wherein the composition inhibits binding of PUMA to Bcl-xL.  
     
     
         7 . The method of  claim 1  wherein the composition inhibits the mitochondrial translocation of Bax.  
     
     
         8 . A method for identifying a drug which inhibits PUMA expression, the method comprising: 
 a) providing PUMA expressing cells in culture;    b) providing a candidate drug to be tested for its ability to inhibit PUMA expression;    c) contacting the PUMA-expressing cells in culture with the candidate drug;    d) assaying the PUMA-expressing cells in culture which have been contacted with the candidate drug for PUMA expression;    e) relating PUMA expression levels in the cells which have been contacted with the candidate drug to PUMA expression levels in matched cells which have not been contacted with the candidate drug; and    f) identifying a drug based on its ability to inhibit PUMA expression to a significant degree as related in step e).    
     
     
         9 . A method for identifying a drug which substantially inhibits the binding of PUMA to Bcl-2, the method comprising: 
 a) providing a PUMA protein;    b) providing a Bcl-2 protein;    c) providing a candidate drug to be tested for its ability to interfere with the binding of a) to b);    d) incubating a), b) and c) under conditions appropriate for the binding of a) to b);    e) determining the extent of binding of a) to b); and    f) comparing the extent of binding determined in step e) to the extent of binding in an otherwise identical incubation in which no candidate drug is included, a substantial difference between the former and the latter indicating that the candidate drug is effective in substantially inhibiting the binding of PUMA to Bcl-2.    
     
     
         10 . A method for identifying a drug which substantially inhibits the binding of PUMA to Bcl-xL, the method comprising: 
 a) providing a PUMA protein;    b) providing a Bcl-xL protein;    c) providing a candidate drug to be tested for its ability to interfere with the binding of a) to b);    d) incubating a), b) and c) under conditions appropriate for the binding of a) to b);    e) determining the extent of binding of a) to b); and    f) comparing the extent of binding determined in step e) to the extent of binding in an otherwise identical incubation in which no candidate drug is included, a substantial difference between the former and the latter indicating that the candidate drug is effective in substantially inhibiting the binding of PUMA to Bcl-xL.    
     
     
         11 . The method of claims  9  or  10  wherein the method is a biochemical method.  
     
     
         12 . The method of claims  9  or  10  wherein the method is carried out in cell culture.  
     
     
         13 . A method for reducing stem cell death in stem cell explants intended to restore cardiac efficiency following cell death in response to coronary infarct, the method comprising contacting the stem cells with a composition effective to inhibit PUMA function prior to introduction.  
     
     
         14 . The method of  claim 13  wherein the composition downregulates PUMA expression.  
     
     
         15 . The method of  claim 14  wherein the composition is a PUMA antisense RNA.  
     
     
         16 . The method of  claim 15  wherein the stem cells express the PUMA antisense RNA.  
     
     
         17 . The method of  claim 13  wherein the composition is a peptide which binds to PUMA protein.  
     
     
         18 . The method of  claim 13  wherein the composition inhibits binding of PUMA to Bcl-2.  
     
     
         19 . The method of  claim 13  wherein the composition inhibits binding of PUMA to Bcl-xL.  
     
     
         20 . The method of  claim 13  wherein the composition inhibits the mitochondrial translocation of Bax.