Function homology screening

Abstract

A method of screening biologically active agent based on the analysis of complex biological responses in culture. Methods for selecting cells and culture conditions for such screens are provided, as well as the identification of an optimized set of discrete parameters to be measured, and the use of biomap analysis for rapid identification and characterization of drug candidates, genetic sequences acting pathways, and the like. A feature of the invention is simultaneous screening of a large number of cellular pathways, and the rapid identification of compounds that cause cellular responses.

1 - 32 . (canceled)  
     
     
         33 . A method for characterization of a biologically active agent according to its mechanism of action, the method comprising: 
 contacting said agent with cells in at least one cell culture assay combination comprising at least two factors acting on said cells;    recording changes in at least three different cellular parameter readouts as a result of introduction of said agent;    deriving a biomap from said changes in parameter readouts, wherein said biomap comprises data normalized to be a ratio of test to control data on the same cell type under control conditions in the absence of said biologically active agent, and said parameters are optimized so that the set of data in the biomap is sufficiently informative that it can discriminate the mechanism of action of said agent;    analyzing said biomap by a multiparameter pattern recognition algorithm to quantify relatedness of said biomap to reference biomaps that include predetermined agents that target specific pathways, wherein the presence or absence of relatedness to said reference biomaps provides a characterization of said agent mechanism of action.    
     
     
         34 . The method of  claim 33 , wherein said analyzing step comprises: 
 classifying said biomap for a single biologically active agent for relatedness to a plurality of biomaps in a database.    
     
     
         35 . The method of  claim 33 , wherein said biologically active agent is an organic compound.  
     
     
         36 . The method of  claim 33 , wherein the cells are primary human endothelial cells, and the at least two factors are selected from IL1β, TNFα, and IFNγ.  
     
     
         37 . The method of  36 , wherein the parameters are selected from E-selectin, VCAM-1, ICAM-1, MCP-1, Mig, IL-8, HLA-DR, uPAR, Tissue Factor and SRB.  
     
     
         38 . The method of  claim 33 , wherein the cells are primary human endothelial cells, and the at least two factors are IL4 and histamine.  
     
     
         39 . The method of  38 , wherein the parameters are selected from VEGFRII, P-selectin, VCAM-1, uPAR, Eotaxin-3, MCP-1 and SRB.  
     
     
         40 . The method of  claim 33 , wherein the cells are primary human fibroblasts and primary human keratinocytes, and the at least two factors are selected from IL1β, TNFα, IFNγ and TGFβ.  
     
     
         41 . The method of  40 , wherein the parameters are selected from MCP-1, ICAM-1, Collagen I, IP-10, Mig, IL1α, M-CSF, PAI-1, TGFβ1, and uPAR.  
     
     
         42 . The method of  claim 33 , wherein the cells are primary human endothelial cells and human smooth muscle cells, and the at least two factors are selected from IL1β, TNFα, and IFNγ.  
     
     
         43 . The method of  42 , wherein the parameters are selected from endothelin-1, E-selectin, MCP-1, VCAM-1, thrombomodulin, tissue factor, uPAR, IL-8, MIG, HGLA-DR, LDL-r, m-CSF and SRB.  
     
     
         44 . The method of  claim 33 , wherein the cells are human smooth muscle cells, and the at least two factors are selected from IL-1β, TNFα and IFNγ.  
     
     
         45 . The method of  44 , wherein the parameters are selected from MCP-1, VCAM-1, thrombomodulin, tissue factor, uPAR, IL-8, MIG, HLA-DR, LDL-r, m-CSF and SRB.  
     
     
         46 . The method of  claim 33 , wherein the cells are human primary endothelial cells and Th2 T cells, and the at least two factors are selected from TCR stimulation, IL-12, and IL-2.  
     
     
         47 . The method of  46 , wherein the parameters are selected from MCP-1, eotaxin 3, VCAM-1, CD38, CD40, E-selectin, uPAR, IL-8, CD69, HGLA-DR, MIG and SRB.  
     
     
         48 . The method of  claim 33 , wherein the cells are primary human endothelial cells and the at least two factors are TNFα and IL-4.  
     
     
         49 . The method of  claim 33 , wherein the cells are primary human endothelial cells and the at least two factors are TNFα and IL-4.  
     
     
         50 . The method of  claim 33 , wherein the cells are fibroblasts and the at least two factors are selected from IL1, TNFα, IFNγ, EGF, FGF and PDGF.