MHC Multimers, Methods for Their Generation, Labeling and Use

Abstract

The present invention describes novel methods to generate MHC multimers and methods to improve existing and new MHC multimers. The invention also describes improved methods for the use of MHC multimers in analysis of T-cells in samples including diagnostic and prognostic methods. Furthermore, the use of MHC multimers in therapy are described, e.g. anti-tumour and anti-virus therapy, including isolation of antigen specific T-cells capable of inactivation or elimination of undesirable targeT-cells or isolation of specific T-cells capable of regulation of other immune cells.

1 . An MHC multimer comprising (a-b-P) n , wherein n>1,
 wherein a and b together form a functional MHC protein capable of binding peptide P, when P is present,   wherein (a-b-P) is an MHC-peptide complex formed when the peptide P binds to the functional MHC protein,   wherein each MHC-peptide complex is associated with one or more multimerization domains, and   wherein one or more detectable labelling molecules selected from the group consisting of DNA and RNA are attached to the MHC-peptide complex or to the one or more multimerization domains.   
     
     
         2 . The MHC multimer according to  claim 1 , wherein said one or more detectable labelling molecules uniquely designate the MHC multimer. 
     
     
         3 . The MHC multimer according to  claim 1 , wherein the one or more labelling molecules are covalently attached to the MHC multimer. 
     
     
         4 . The MHC multimer according to  claim 1 , wherein the one or more labelling molecules are non-covalently attached to the MHC multimer. 
     
     
         5 . The MHC multimer according to  claim 1 , wherein the one or more labelling molecules are attached directly to the MHC multimer. 
     
     
         6 . The MHC multimer according to  claim 5 , wherein the one or more labelling molecules are attached directly to the MHC complex via the heavy chain, β2m, or peptide. 
     
     
         7 . The MHC multimer according to  claim 5 , wherein the one or more labelling molecules are attached directly to the one or more multimerization domains. 
     
     
         8 . The MHC multimer according to  claim 1 , wherein the one or more labelling molecules are conjugated to a linker. 
     
     
         9 . The MHC multimer according to  claim 8 , wherein the linker attaches the detectable labelling molecule to the one or more multimerization domains. 
     
     
         10 . The MHC multimer according to  claim 1 , wherein one or more of the n MHC-peptide complexes are covalently linked to the one or more multimerization domains. 
     
     
         11 . The MHC multimer according to  claim 1 , wherein the one or more multimerization domains comprise a polymer, to which is optionally attached one or more scaffolds. 
     
     
         12 . The MHC multimer according to  claim 11 , wherein the polymer comprises a polysaccharide. 
     
     
         13 . The MHC multimer according to  claim 12 , wherein the polysaccharide is one or more dextran moieties. 
     
     
         14 . The MHC multimer according to  claim 1 , wherein the MHC complexes comprise chemically modified MHC complexes. 
     
     
         15 . The MHC multimer according to  claim 14 , wherein the chemically modified MHC complexes comprise one or more types of cross-linkers. 
     
     
         16 . The MHC multimer according to  claim 15 , wherein the cross-linkers are one or more chemical cross linkers. 
     
     
         17 . A composition comprising a plurality of MHC multimers according to  claim 1 . 
     
     
         18 . A method for generating the MHC multimer according to  claim 1 , said method comprising the steps of:
 providing one or more MHC-peptide complexes;   providing one or more multimerization domains; and   reacting said one or more MHC-peptide complexes and said one or more multimerization domains, thereby generating the MHC multimer according to  claim 1 .