Role of Peptide Modification and Multivalent Delivery on the Efficacy and Safety of Peptide-based Immunotherapy in Type 1 Diabetes

Peptide-based immunotherapy offers a more targeted approach to reestablish tolerance in several human autoimmune diseases, but issues of bioavailability, enzymatic degradation and anaphylaxis may limit their clinical application.  We previously showed that soluble antigen arrays (SAgAs), made of multiple antigenic peptides grafted onto a hyaluronic acid polymer, efficiently deliver p79 and 2.5HIP peptides and protect NOD mice against autoimmune diabetes. However, some mice developed anaphylaxis following repeated dosing. Thus, we further investigated two SAgA variants and their corresponding modified peptides for their effect on both disease prevention and anaphylaxis in NOD mice. To that end, two different N-terminal peptide modifications were used for grafting the peptides to make SAgAs: aminooxy (AO) for hydrolysable linkers and homopropargyl (HP) for non-hydrolysable linkers. AO-functionalized peptides and corresponding SAgAs led to significantly accelerated anaphylaxis compared to HP-functionalized peptides and SAgAs. Moreover, anaphylaxis occurred significantly faster when using free peptides than with SAgAs. Anaphylaxis was dose-dependent and mainly mediated by IgG1 against p79. Importantly, both SAgA variants efficiently prevented the development of spontaneous diabetes while free peptides failed to provide any significant protection at equivalent peptide doses. The protection appears to be mediated by a combination of induced regulatory T cells, exhaustion and/or deletion of peptide-specific T cells, depending on the time of analysis and forms of SAgA. In sum, we provide evidence that SAgAs are more effective than free peptides to induce tolerance and block autoimmune diabetes, and those made with HP-modified peptides are best to minimize risks of anaphylaxis.