Rgd-binding compounds and methods of use

Abstract

Provided herein are compositions and methods for preventing and treating diseases and risk factors associated with metabolic syndrome by targeting the RGD-binding site of selected intra- and extracellular proteins. Exemplary compositions include RGD-polyphenol conjugates via an ester linkage; polyphenol polymer conjugated to RGD analogs or mimetics; and RGD polymer conjugates linked to polyphenol.

What is claimed is: 
     
         1 . A method for identifying genes encoding the amino acid sequence Arg-Gly-Asp that are regulated in opposing directions by insulin and at least one RGD-binding compound, the method comprising:
 (a) selectively stimulating expression of an unknown gene product that encodes the amino acid sequence Arg-Gly-Asp;   (b) isolating said gene product from control, resveratrol- and insulin-stimulated cells alone and in combination;   (c) pooling said gene products; and   (c) analyzing the pools of gene product to identify genes regulated in opposing directions by insulin and at least one RGD-binding compound.   
     
     
         2 . The method of  claim 1 , wherein said analyzing step includes performing a gene expression microarray. 
     
     
         3 . A method of screening for indicators of insulin sensitivity that encode the amino acid sequence Arg-Gly-Asp, the method comprising:
 (a) providing an affinity column containing at least one compound that binds to the amino acid sequence Arg-Gly-Asp;   (b) introducing serum collected from a subject exposed to nutritional manipulations, or from a subject afflicted with a condition associated with metabolic syndrome to said column, said serum containing at least one indicator of insulin sensitivity; and   (c) collecting at least one indicator that binds to the compounds in the affinity column, wherein an indicator that binds to the compounds in the affinity column encode the amino acid sequence Arg-Gly-Asp and represents an indicator of insulin sensitivity.   
     
     
         4 . The method of  claim 3 , wherein said indicators are extra-cellular secretory proteins. 
     
     
         5 . A method of screening a test compound for modulating insulin sensitivity, the method comprising:
 (a) providing a cell culture comprising a first cell line overexpressing a recombinant expression construct containing at least one intra-cellular protein encoding the amino acid sequence Arg-Gly-Asp, and a second cell line overexpressing a recombinant expression construct containing at least one mutant derivative of the protein expressed in said first cell line;   (b) adding the test compound to the cell culture;   (c) assaying the cell culture to determine whether the test compound is taken up by the first and second cell lines;   (d) assaying the first and second cell lines to determine whether the test compound binds the protein expressed by the first and second cell line, wherein binding of the test compound to the protein expressed by the first cell line, but not the protein expressed by the second cell line, indicates that the test compound modulates insulin activity.   
     
     
         6 . The method of  claim 5 , wherein said mutant derivative encodes the amino acid sequence Arg-Gly-Glu. 
     
     
         7 . The method of  claim 5 , wherein the first cell line in step (a) alternatively contains an extra-cellular protein encoding the amino acid sequence Arg-Gly-Asp. 
     
     
         8 . The method of  claim 5 , wherein the first and second cell line contains preadipocytes. 
     
     
         9 . The method of  claim 5 , wherein the test compound modulates insulin activity by preventing insulin stimulated fat cell differentiation or insulin stimulated fat accumulation. 
     
     
         10 . The method of  claim 5 , wherein the test compound modulates insulin activity by potentiating insulin stimulated fat cell differentiation or insulin stimulated fat accumulation. 
     
     
         11 . The method of  claim 5 , wherein the first cell line in step (a) alternatively comprises a reporter gene driven by the promoter of a gene product encoding the amino acid sequence Arg-Gly-Asp, and wherein the second cell line in step (a) alternatively comprises a reporter gene driven by the promoter of a gene product encoding the amino acid sequence Arg-Gly-Glu. 
     
     
         12 . The method of  claim 5 , further comprising the step of adding insulin to the cell culture prior to performing step (b). 
     
     
         13 . The method of  claim 5 , wherein the assaying step of step (d) includes determining whether the test compound binds the promoter expressed by the first and second cell line, wherein binding of the test compound to the promoter expressed by the first cell line, but not the promoter expressed by the second cell line, indicates that the test compound modulates insulin activity. 
     
     
         14 . The method of  claim 5 , wherein the method further comprises the step of assaying libraries for a hit.