USE OF TUMOR NECROSIS FACTOR-alpha RECEPTOR p75 FOR TREATMENT OF ISCHEMIA-INDUCED NEOVASCULARIZATION

Abstract

Improvements on the basic method used for BEAMing increase sensitivity and increase the signal-to-noise ratio. The improvements have permitted the determination of intrinsic error rates of various DNA polymerases and have permitted the detection of rare and subtle mutations in DNA isolated from plasma of cancer patients.

1 . A method of treating, reducing the severity of, or preventing ischemia in a subject having or at risk of developing ischemia, the method comprising,
 a) contacting a cell of the subject with a nucleic acid molecule encoding a p75/TNFR2 polypeptide or a fragment thereof; and   b) expressing the p75/TNFR2 polypeptide in the cell, wherein the method treats or prevents ischemia in the subject.   
     
     
         2 . A method of enhancing angiogenesis in a tissue before, during, or after an ischemic event, the method comprising
 a) contacting a cell with a nucleic acid molecule encoding a p75/TNFR2 polypeptide or a fragment thereof; and   b) expressing the p75/TNFR2 polypeptide in the cell, wherein the method enhances angiogenesis in the tissue.   
     
     
         3 . The method of  claim 1 ,
 further comprising administering the cell to the subject, wherein the method enhances angiogenesis.   
     
     
         4 . The method of  claim 1 , wherein
 the method reduces apoptosis in the subject.   
     
     
         5 . The method of  claim 1 , wherein the method enhances the local release of angiogenic growth factors and cytokines in the tissue. 
     
     
         6 . The method of  claim 1 , further comprising the step of administering to the subject an angiogenic factor selected from the group consisting of: vascular endothelial growth factor (VEGF), hepatocyte growth factor (HGF), basic fibroblast growth factor (bFGF), angiopoietin 1, angiopoietin 2 and monocyte chemotactic protein-1 (MCP-1). 
     
     
         7 . The method of  claim 1 , further comprising the step of administering to the subject an endothelial cell mitogen selected from the group consisting of acidic and basic fibroblast growth factors, vascular endothelial growth factor, epidermal growth factor, transforming growth factor a and 13, platelet-derived endothelial growth factor, platelet-derived growth factor, tumor necrosis factor a, hepatocyte growth factor, insulin like growth factor, erythropoietin, colony stimulating factor, macrophage-CSF, granulocyte/macrophage CSF and nitric oxide synthase. 
     
     
         8 - 10 . (canceled) 
     
     
         11 . The method of  claim 3 , wherein the cell is delivered directly to an ischemic tissue or is delivered systemically. 
     
     
         12 . (canceled) 
     
     
         13 . The method of  claim 1 , wherein the nucleic acid molecule is present in a vector. 
     
     
         14 - 15 . (canceled) 
     
     
         16 . A method of enhancing p75/TNFR2 expression in a cell, the method comprising
 a) contacting a cell with a nucleic acid molecule encoding a p75/TNFR2 polypeptide or a fragment thereof; and   b) expressing the p75/TNFR2 polypeptide in the cell.   
     
     
         17 - 21 . (canceled) 
     
     
         22 . The method of claim  20 , wherein the cell is an endothelial progenitor cell or bone marrow derived cell. 
     
     
         23 . An expression vector comprising a nucleic acid molecule encoding a mammalian p75/TNFR2 polypeptide or a fragment thereof operably linked to a promoter sufficient to direct expression of the p75 TNFR2 receptor polypeptide in a cell. 
     
     
         24 - 26 . (canceled) 
     
     
         27 . The vector of claim  26 , wherein the vector is selected from the group consisting of adenoviral vectors, adeno-associated viral vectors, retroviral vectors, lentiviral vectors, alphaviral vectors, and herpes virus vectors. 
     
     
         28 - 30 . (canceled) 
     
     
         31 . A host cell comprising the vector of  claim 23 . 
     
     
         32 - 34 . (canceled) 
     
     
         35 . The host cell of  claim 31 , wherein the cell is an endothelial progenitor cell or a bone marrow-derived cell. 
     
     
         36 . (canceled) 
     
     
         37 . The host cell of  claim 31 , wherein the cell is in vitro or in vivo. 
     
     
         38 . A pharmaceutical composition comprising an effective amount of an expression vector encoding a human p75/TNFR2 polypeptide or a fragment thereof in a pharmaceutically acceptable excipient, wherein the p75/TNFR2 polypeptide is operably linked to a promoter sufficient to drive expression of the p75/TNFR2 polypeptide in a mammalian cell. 
     
     
         39 . (canceled) 
     
     
         40 . The pharmaceutical composition of  claim 38 , wherein the promoter is sufficient to drive expression in an endothelial progenitor cell or a bone marrow-derived cell. 
     
     
         41 . A kit for the treatment or prevention of ischemia, the kit comprising an effective amount of an expression vector encoding a human p75/TNFR2 polypeptide or a fragment thereof in a pharmaceutically acceptable excipient, wherein the p75/TNFR2 polypeptide is operably linked to a promoter sufficient to drive expression of the p75/TNFR2 polypeptide in a mammalian cell. 
     
     
         42 . (canceled) 
     
     
         43 . A method of monitoring a subject being treated for ischemia, the method comprising
 a) administering a treatment that enhances the expression of a p75/TNFR2 polypeptide in a cell of the subject; and   b) measuring apoptosis or an increase in angiogenesis in a tissue of the subject relative to a reference, wherein a decrease in apoptosis or an increase in angiogenesis indicates a reduced severity of ischemia in the subject.   
     
     
         44 - 50 . (canceled) 
     
     
         51 . A method for identifying a candidate compound a candidate compound useful for the treatment of ischemia, the method comprising the steps of:
 (a) contacting a cell expressing p75/TNFR2 polypeptide with a candidate compound; and   (b) detecting an increase in the level or the biological activity of a p75/TNFR2 polypeptide or nucleic acid molecule in the cell relative to a reference, wherein an increase in the level or the biological activity of p75/TNFR2 polypeptide identifies a candidate compound useful for the treatment of ischemia.   
     
     
         52 . (canceled)