Identification of an immunogenic epitope from RASGRP2 with specificity for HLA-DR15 in multiple sclerosis

Identifying autoantigens in multiple sclerosis (MS) has been challenging. If successful, this could facilitate development of autoantigen-specific tolerogenic therapies, by exposing antigen presenting cells (APC) to tolerogenic stimuli with autoantigens specific to the human leukocyte antigen (HLA) haplotype of patients. RAS guanyl releasing protein 2 (RASGRP2), which is expressed by lymphocytes and striatal neurons, is a putative autoantigen in HLA-DRB1*15:01 (DR15)-positive individuals. We aimed to identify antigenic RASGRP2 epitope(s) that could be used to develop tolerogenic therapies. RASGRP2-derived peptides of varying DR15 binding affinities were exposed to peripheral blood mononuclear cells (PBMCs). Immunoeptidomic techniques demonstrated strong and moderate binding affinity RASGRP2 peptides could be presented by HLA-DR on PBMCs from DR15-homozygous or DR15-negative patients. Moderate affinity peptides produced the greatest increase in CD80 expression and pro-inflammatory cytokine (IFN-γ, IL-17, IL-22) secretion by PBMCs, particularly in DR15-positive patients. One RASGRP2 peptide eliciting the highest pro-inflammatory responses was used to generate HLA-DR15 tetramers. CD4+ T-cells specific for this peptide were four-fold higher in frequency in DR15-positive patients versus controls, more pro-inflammatory in phenotype and demonstrated increased peptide-stimulated proliferation. In conclusion, we identified a novel immunogenic RASGRP2 peptide with relative selectivity for HLA-DR15-positive people with MS, which could form the basis for autoantigen-specific tolerogenic therapy.