Assay systems for leptin-enhancing agents

Abstract

The present invention relates to methods for identifying therapeutic agents that enhance the effect of leptin, an adipocyte-derived cytokine that regulates food intake and body weight. The invention further provides for use of agents identified using this assay system to enhance the interaction between leptin and its receptor, OB-R, thereby boosting leptin's weight-reducing effects in obese individuals.

We claim:  
     
         1 . A method of screening for a molecule capable of competing with OB-R for binding to SHP2-SH2 comprising: 
 a) contacting a phosphopeptide comprising the Y986 domain of OB-R under conditions in which the Y986-containing phosphopeptide is capable of binding to a peptide comprising the SH2 domain of SHP2;    b) determining the amount of the Y986-containing domain that binds to the SHP2-SH2 containing peptide;    c) contacting a known amount of the Y986 containing phosphopeptide to the SHP2-SH2 containing peptide, in the presence of a sample suspected of containing the molecule capable of competing with the Y986 containing phosphopeptide, under the same conditions used in b);    d) determining the amount of the Y986-containing phosphopeptide that binds to the SHP2-SH2 containing peptide;    e) comparing the amount from (b) with the amount from (d), wherein a lesser amount in (d) indicates the presence of a molecule capable of competing with the Y986 containing phosphopeptide for binding to the SHP2-SH2 domain    
     
     
         2 . The method of  claim 1 , wherein the Y986 containing phosphopeptide or the SHP2-SH2 containing peptide is bound to a solid support.  
     
     
         3 . The method of  claim 2 , wherein the Y986 containing phosphopeptide or the SHP2-SH2 containing peptide is detectably labeled.  
     
     
         4 . The method of  claim 1  wherein the Y986 containing phosphopeptide is the cytoplasmic domain of OB-R.  
     
     
         5 . The method of  claim 1  wherein the Y986 containing phosphopeptide is a dimer.  
     
     
         6 . An assay system comprising 
 a) an SH2 domain of an SH2 containing phosphatase;    b) a peptide containing the tyrosine phosphorylated SH2 binding domain of a cytokine receptor;    c) a means for detecting the interaction between said SH2 domain and said SH2 binding domain.    
     
     
         7 . The assay system of  claim 6  wherein said SH2 binding domain containing peptide is the cytoplasmic domain of a cytokine receptor that has a downstream signaling pathway that involves an SH2-containing phosphatase.  
     
     
         8 . The assays system of  claim 7  wherein said SH2-containing phosphatase is SHP1or SHP2.  
     
     
         9 . The assays system of  claim 7  wherein said cytokine receptor is selected from the group consisting of OB-R and erythropoietin.  
     
     
         10 . An agent identified according to the method of  claim 1 .  
     
     
         11 . An agent that enhances the therapeutic effect of leptin response comprising an agent according to claim  10 .