Method for the detection, preparation and depletion of cd4+ t lymphocytes

Abstract

The present invention relates an in vitro method for detecting class II restricted CD4+ T cells in a sample. Herein a sample is contacted with an isolated complex of an MHC class II molecule and a peptide. This peptide comprises an MHC class II restricted T cell epitope of an antigenic protein and immediately adjacent thereof, or separated by a linker of at most 7 amino acids a sequence with a [CST]-xx-C or C-xx-[CST] motif. CD4+ T cells are detected by measuring the binding of the complex with cells in the sample, wherein the binding of the complex to a cell is indicative for the presence of CD4+ T cells in the sample. The present invention further relates to an isolated complex of an MHC Class II molecule and a peptide comprising an MHC class II restricted T cell epitope of an antigenic protein and immediately adjacent thereof, or separated by a linker of at most 7 amino acids a sequence with a [CST]-xx-C or C-xx-[CST] motif.

Description

FIELD OF INVENTION[0001]The present invention relates to synthetic peptides encompassing class I l-restricted major histocompatibility complex (MHC) T cell epitopes containing a thioredox motif within flanking residues for the detection, preparation or depletion of CD4+ T lymphocytes in or from body fluids or culture medium.BACKGROUND OF THE INVENTION[0002]Lymphocytes play a central role in the elaboration of immune responses against foreign antigens and in the control of diseases. Such recruitment and activation of lymphocytes can be beneficial as in responses against infectious agents, or detrimental, such as exemplified by auto-immune diseases, responses to allergens and in graft rejection. There are therefore a large number of circumstances in which it would be advantageous to detect, enumerate, purify or deplete such lymphocytes. Current methods to achieve these goals are, however, unsatisfactory.[0003]Lymphocytes are divided in several lineages and subsets according to the pre...

AI Technical Summary

Benefits of technology

[0049]The WO2008017517 patent application describes the use of oxidoreductase motifs inserted into the flanking residues of class II-restricted epitopes for the therapy of a number of diseases. It is demonstrated that the oxidoreductase motif stabilizes the synapse formed between the MHC-peptide complex and cognate TCR, as reported in Carlier et al., PloS ONE 7(10): e45366 1-14. The increased synapse formation described therein is supported by experiments with purified APC and CD4+ T cells, which show an increase in number and duration of dimer formation between APC and CD4+ T cells, which results in an effect on various functional properties of CD4+ T cells. An effect on the binding affinity between APC and CD4+ is not discussed in the prior art.

[0050]We made the unexpected observation that addition of an oxidoreductase motif within flanking residues of class II epitopes, increases the binding affinity of MHC-epitope complexes with CD4+ T cells, also when the MCH molecules are used as isolated complexes. This allows using peptides with a T cell epitope sequence and a redox motif in the detection and preparation of antigen-specific CD4+ T cells and for all subsequent manipulation of such cells.

Problems solved by technology

Current methods to achieve these goals are, however, unsatisfactory.

TCR low affinity results in inefficient detection, in particular in complex body fluids.

However, although both the specificity and sensitivity of these multimers have been significantly increased, these detection tools still remain relatively inefficient under circumstances of low T cell frequency or when TCR affinity is a limiting factor, such as for T cells in naïve configuration.

The contrast between the increasing need for methods to detect CD4+ T cells in areas as diverse as auto-immune diseases, cancer or evaluation of responses towards vaccination, to cite just a few, and the poor efficiency of available methods makes it an urgent need to improve such methods