The heterogeneous nuclear ribonucleoprotein A2 (hnRNP-A2) has been described as an important autoantigen in rheumatoid arthritis (RA) since it is targeted by autoantibodies, autoreactive T cells, and is aberrantly expressed in synovial cells in patients. To identify hnRNP-A2-specific T-cell epitopes possibly associated with pathogenicity, we used an innovative approach. We first scanned 280 overlapping hnRNP-A2 peptides for binding to the RA-associated class II molecules HLA-DR4 and HLA-DR1, leading to a comprehensive selection of binders. The selected peptides were tested in IFN-gamma-specific ELISPOT assay: PBMC from 18% of RA patients showed a significant IFN-gamma response to hnRNP-A2 peptides, 15% to the overlapping sequences 117-133 and/or 120-133, whereas PBMC from healthy individuals tested negative. We measured proliferative responses to these two peptides in another cohort of patients with RA or osteoarthritis: positive responses were found in 28% of RA, but also in 11% of osteoarthritis patients and these responses could be blocked by anti-MHC class II Ab. Remarkably, the presence of 117/120-133-specific T cells was significantly associated with active disease in RA patients, and bone erosion appeared to be more common in T-cell positive patients. These data suggest involvement of hnRNP-A2 specific cellular autoimmune responses in RA pathogenesis.

Abstract Human cytomegalovirus (HCMV) is contained by T-lymphocyte responses focused towards the major tegument protein pp65. To systematically identify T-cell epitopes, we applied the following strategy: 441 overlapping 15mer peptides spanning the entire HCMV pp65 antigen in 1-aa steps were screened in enzyme-linked immunospot (ELIspot) assays for interferon gamma (IFN-γ) secretion by peripheral blood mononuclear cells (PBMCs) from nine healthy HCMV-seropositive subjects expressing human leukocyte antigen (HLA)-A2. This analysis confirmed a number of previously known epitopes and revealed several new ones. A total of 26 epitopes were identified, including 14 HLA-A2, four HLA-B7, -B35, -B12 and -B44 restricted class I epitopes, six class II epitopes, and two epitopes of unknown restriction. Three novel HLA-A2 epitopes were confirmed using T2-cells, and one peptide for which only binding data had been published so far was verified. Two novel class II epitopes were confirmed by intracellular cytokine staining. Responses were usually oligoclonal against up to seven HLA-A2 epitopes, albeit with a few dominating epitopes. Clusters of overlapping epitopes (hot-spots) were identified. These and the newly identified T-cell epitopes may be of great value for epitope-based immunotherapeutic approaches, including peptide vaccines.