Tolerogenic multi-epitope DNA vaccination in autoimmune diabetes

BACKGROUND: Antigen-specific tolerance has not yet been achieved in Type 1 diabetes (T1D) patients, who harbor diabetogenic T cells reacting against multiple beta-cell antigens. Plasmid DNA (pDNA)-encoding such antigens have been evaluated with the goal to achieve tolerance, and have the advantages of easy production at low cost, and prolonged expression of the antigens. Thus far, the use of a single antigen produced in native form may limit the fraction of diabetogenic T cells that can be engaged, yet there is numerous evidence of epitope spreading and post-transcriptional peptide modifications of multiple antigens. We investigated whether delivery of major epitopes from multiple beta-cell antigens confers any potential benefit over delivery of a single or multiple proteins as source of antigens.

METHODS: We designed two epitope-expressing constructs, both expressing the same major epitopes from several beta-cell antigens, including native epitopes and mimotopes. We used these DNAs to immunize NOD mice. By MHC-tetramer staining and TCR-transgenic adoptive cell transfer, we studied the antigen-specific immune responses and DNA therapeutic efficacy.

RESULTS: DNA delivered intramuscularly or intradermally was successfully expressed for at least 1 week at the site of a single injection. Both non-hematopoietic and dendritic cells (DCs) expressed the pDNA at the site of injection but only DCs were found in the draining lymph nodes (LNs). Antigen-specific CD4+ and CD8+ T cells specific to at least 4 different beta-cell antigens expressed were engaged after DNA vaccination in draining and pancreatic LNs, where they accumulated with minimal clonal expansion. The T cell response was dominated by anergy, with some regulatory T cells induced. Finally, vaccination with multi-epitope encoding DNA significantly delayed the onset of T1D in NOD mice, unlike vaccination with single antigen-encoding pDNA. Furthermore, mice receiving combinations of several antigen-encoding plasmids were protected to a greater extent than those receiving proinsulin-encoding pDNA only.