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 - 16 . (canceled)  
     
     
         17 . A method for characterization of a biologically active agent, the method comprising: 
 contacting said agent with human primary 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 known agents that target specific pathways, wherein the presence or absence of relatedness to said reference biomaps provides a characterization of said agent.    
     
     
         18 . The method according to  claim 17 , wherein one of said assay combinations simulates an in vivo physiological state.  
     
     
         19 . The method according to  claim 17 , wherein one of said cell cultures is a control cell culture selected from the group consisting of: a basal cell culture in the absence of said factors; a basal cell culture in the absence of said factors and in the presence of an agent of known physiological activity; said cell culture in the presence of said factors, or said cell culture in the presence of said factors and a compound of known physiological activity.  
     
     
         20 . The method according to  claim 18 , wherein said physiological state environment comprises said biologically active agent and a control assay combination is free of said agent.  
     
     
         21 . The method according to  claim 17 , wherein said cell culture comprises a plurality of different cell types associated with said physiological state.  
     
     
         22 . The method according to  claim 17 , wherein said cells are primary cells.  
     
     
         23 . The method according to  claim 17 , wherein said cells are selected from the group consisting of endothelial cells, leukocytes, neoplastic cells and epithelial cells.  
     
     
         24 . The method according to  claim 17 , wherein said cells are genomically modified cells.  
     
     
         25 . The method according to  claim 17 , wherein said biologically active agent is an organic compound.  
     
     
         26 . The method according to  claim 17 , wherein the cells are primary endothelial cells, and said at least two factors are selected from VEGF, FGF, EGF, TNF-α, IL-4, IL-13, histamine, IL-8, angiotensin-II, thrombin, IFN-γ, PDGF, oxidized LDL and IL-1.  
     
     
         27 . The method according to  claim 17 , wherein said cells are T lymphocytes and said at least two factors are selected from  Staphylococcal  enterotoxin B, anti-CD28, anti-CD3, anti-CD49d, IL-12, IL-1, II-2, IL-4, IL-6, IL-7, IL-13, IL-15, IL-18, and TGFβ.  
     
     
         28 . A method for preparing a biomap dataset for a plurality of pathways associated with a plurality of physiological states of interest of cells in cell culture, said method comprising: 
 contacting a biologically active agent with human primary 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.    
     
     
         29 . The method according to  claim 28 , further comprising the step of compiling a plurality of said biomaps in a database.  
     
     
         30 . A screening system comprising: 
 at least two cell culture assay combinations comprising cells and at least two factors affecting at least four pathways for inducing said physiological state of interest on said cells, wherein at least one of said assay combinations comprises said biologically active agent; assay reagents for measuring at least two parameters associated with said pathways; and a data processor for analyzing the data from said biological culture in relation to at least one control biological culture of known activity.    
     
     
         31 . The system according to  claim 30 , wherein said data processor comprises a plurality of biomap datasets in a database, wherein said biomap datasets are prepared according to the method set forth in  claim 17.