Although both intracellular and secreted constructs were found to elicit antigen-specific T cell responses, only the secreted form of Endotope mediated disease protection in NOD mice, and was therefore the only form used in subsequent experiments. The authors sought to determine whether route of inoculation (i.e., intramuscular versus intradermal) could affect the platform’s efficacy, and also compared this with Proinsulin pDNA treatment. With weekly injections, both vaccines reduced disease incidence in mice when administered intramuscularly, though not significantly. In contrast, type 1 diabetes incidence was significantly reduced through both treatments when administered intradermally. Additionally, the authors determined that longer, continuous DNA delivery is required to achieve stable, long-term protection against type 1 diabetes.
Since the authors discovered that antigens were present as late as 14 days following treatment, they next investigated whether weekly treatments were too frequent and treated NOD mice via both inoculation routes every other week. Interestingly, Endotope provided only partial and non-significant protection against type 1 diabetes, suggesting that antigen presentation beyond one week may not contribute to disease protection.
Proinsulin pDNA vaccination is particularly effective at preventing type 1 diabetes onset at the dysglycemic stage, and so the authors examined whether Endotope has the same capability. Following six weeks of twice-weekly treatment, 78% of control mice developed diabetes compared to 50% of Endotope-treated and 22% of Proinsulin-treated mice. However, Endotope-treated mice who had not developed diabetes were more protected in the long term than Proinsulin-treated mice. The authors also determined that the Proinsulin vaccine is more effective at preventing the milder form of disease, whereas the Endotope vaccine is modestly effective at preventing both mild and aggressive forms. Additionally, while both vaccines significantly reduced insulitis after 10 weeks of treatment, Endotope-treated mice exhibited significantly more islets without immune cell infiltration.