Citrullinated and native GRP78 peptides are targeted by circulating naïve CD8+ T cells in type 1 diabetic and healthy donors

Introduction. CD8+ T-cell-mediated autoimmune β-cell destruction occurs in T1D. Such destruction may be favored by an increased β-cell vulnerability through the presentation of neo-epitopes more prone to escape tolerance. In this respect, post-translationally modified target peptides have been described, including citrullinated glucose-regulated protein 78 (GRP78) peptides in the NOD mouse and in the islet infiltrates of T1D patients. Here we report about the recognition of HLA-A2-restricted native and citrullinated GRP78 peptides by circulating CD8+ T cells.

Methods. GRP78 native (R form) and citrullinated (X form) candidate epitopes were identified in silico and confirmed to bind HLA-A2 in vitro. Peptide-reactive CD8+ T cells were identified by combinatorial HLA-A2 multimer staining ex vivo on PBMCs from T1D and healthy donors, and the relative recognition of the R vs. X form assessed.

Results. All the GRP78 candidate epitopes tested were recognized by circulating CD8+ T cells. The latter were predominantly naïve and fell in the expected frequency range reported for known islet epitopes, which was similar in T1D and healthy donors for all peptides. Exceptions were noted for GRP78_435-443R and, to a lesser extent, GRP78_360-368R, which were preferentially recognized in T1D patients and displayed a higher proportion of antigen-experienced CD8+ T cells. CD8+ T-cell recognition exhibited a variable preference for the R or X GRP78 peptide forms: 9-18R, 261-269X, 298-307R, 360-368R, 435-443R, 502-511X. With few exceptions (261-269, 435-443), this preference was rather exclusive, and very few CD8+ T cells recognized both R and X forms. In conclusion, GRP78 is a novel islet antigen targeted by circulating CD8+ T cells, whose recognition is not invariably biased toward citrullinated forms.