Method of epitope discovery

Abstract

A method of epitope discovery comprising the step of selecting an epitope from a population of peptide fragments of an antigen associated with a target cell, wherein the fragments have a known or predicted affinity for a major histocompatibility complex class I receptor peptide binding cleft, wherein the epitope selected corresponds to a proteasome cleavage product of the target cell.

1 . A method for selecting and/or producing a T cell epitope, comprising contacting a peptide or polypeptide to a proteasome to permit processing of the peptide or polypeptide by the proteasome to determine the c-terminus of said T cell epitope.  
     
     
         2 . The method of  claim 1 , wherein said proteasome is an immune proteasome.  
     
     
         3 . The method of  claim 1 , wherein said proteasome is a housekeeping proteasome.  
     
     
         4 . The method of  claim 1 , wherein said proteasome is a proteasome exposed to interferon-gamma.  
     
     
         5 . The method of  claim 1 , wherein said peptide or polypeptide is synthetic.  
     
     
         6 . The method of  claim 1 , wherein said peptide or polypeptide is a fragment of an antigen.  
     
     
         7 . The method of  claim 1 , wherein said peptide or polypeptide is an epitope cluster.  
     
     
         8 . The method of  claim 1 , further comprising identifying additional characteristics of said T cell epitope.  
     
     
         9 . The method of  claim 8 , wherein said characteristics comprises at least one characteristic selected from the group consisting of MHC binding affinity, identity of a c-terminus, N-terminus, and proteolytic cleavage sites.  
     
     
         10 . A T cell epitope obtainable by a method selected from the group consisting of the method of  claim 1 ,  2 ,  3 ,  4 ,  5 ,  6 ,  7 ,  8  and  9 .  
     
     
         11 . A nucleic acid encoding at least one T-cell epitope according to  claim 10 .  
     
     
         12 . The nucleic acid of  claim 11 , comprising at least two sequences, each encoding a T cell epitope, wherein said at least two sequences are separated by at least one proteolytic cleavage site.  
     
     
         13 . A vector comprising a nucleic acid selected from the group consisting of the nucleic acid of  claim 11  and  claim 12 .  
     
     
         14 . A composition comprising a vector of  claim 13 .  
     
     
         15 . A composition comprising a T cell epitope of  claim 10 .  
     
     
         16 . A polypeptide obtainable by expressing a nucleic acid of  claim 11  or  claim 12 .  
     
     
         17 . A composition comprising the polypeptide of  claim 16 .  
     
     
         18 . A composition comprising an adjuvant and a composition selected from the group consisting of the composition of  claim 14 ,  15  and  17  the polypeptide of  claim 16 .  
     
     
         19 . A method for selecting and/or producing a T cell epitope, comprising subjecting a precursor peptide or polypeptide to the action of a 20S proteasome, in particular an IFN-.gamma. or other immune-related proteasome or a functional equivalent thereof to determine the location of the c-terminus of said T cell epitope.  
     
     
         20 . A method for selecting and/or producing according to  claim 1  wherein said precursor peptide or polypeptide is tested for other T cell epitope characteristics.  
     
     
         21 . A method according to  claim 20 , wherein said further testing comprises testing for the right anchor residues, the proper cleavage sites, the proper pathway signals, and/or the stability of the MHC-T cell epitope complex.  
     
     
         22 . A T cell epitope obtainable by a method according to any one of claims  19 - 21 .  
     
     
         23 . A nucleic acid encoding at least one T cell epitope according to  claim 22 .  
     
     
         24 . A nucleic acid according to  claim 23  comprising at least two sequences encoding a T cell epitope, separated by at least one proteolytic cleavage site.  
     
     
         25 . A gene delivery vehicle comprising a nucleic acid according to  claim 22  or  23 .  
     
     
         26 . A pharmaceutical composition comprising a gene delivery vehicle according to  claim 7 .  
     
     
         27 . A pharmaceutical composition comprising at least one epitope according to  claim 22 .  
     
     
         28 . A proteinaceous molecule obtainable by expression of a nucleic acid according to  claim 23  or  24 .  
     
     
         29 . A pharmaceutical composition comprising a proteinaceous molecule according to  claim 28 .  
     
     
         30 . A pharmaceutical composition according to  claim 26 ,  27  or  29  further comprising an adjuvant.  
     
     
         31 . A pharmaceutical composition according to  claim 26 ,  27 ,  29  or  30  comprising an antigen presenting moiety.  
     
     
         32 . A pharmaceutical composition according to  claim 31  wherein said moiety comprises a major histocompatibility molecule.  
     
     
         33 . A pharmaceutical composition according to  claim 32  wherein said major histocompatibility molecule is present on an antigen presenting cell, such as a dendritic cell.  
     
     
         34 . A method of epitope discovery comprising the step of selecting an epitope from a population of peptide fragments of an antigen associated with a target cell, wherein the fragments have a known or predicted affinity for a major histocompatibility complex class I receptor peptide binding cleft, wherein the epitope selected corresponds to a proteasome cleavage product of the target cell.