EXTH-97. Enhancing T cell responses to glioma-specific vaccines through optimized altered peptide ligands

Abstract Gliomas are diffusely infiltrating and incurable primary brain tumors. While glioma immunotherapy presents promising strategies, clinical implementation faces significant challenges, particularly in achieving durable responses. We have previously conducted phase 1 clinical trials demonstrating safety, immunogenicity, and biological activity of mutation-specific long peptide vaccines, such as vaccines targeting the glioma driver mutations IDH1R132H and H3K27M. Immunogenic epitopes from these driver mutations are presented on MHC class II molecules across various allelotypes to stimulate mutation-specific CD4 T cell responses in patients. As the magnitude of immune responses was variable and transient in some patients, we rationalized the synthetic optimization of the H3K27M long peptide to improve immunogenicity. Here, we aimed at enhancing peptide-MHC class II interactions, specifically targeting the HLA-DRB1 P1 pocket, known for its preference for large hydrophobic residues that are critical for peptide anchoring. Using a late-stage-functionalization method, we generated a library of potential MHC ligands with a variety of synthetic substituents as anchor residues for our proof-of-concept peptide, derived from the influenza A matrix 1 (M1) antigen. Peptide-MHC stability assays confirmed that large, nonpolar substituents increase MHC binding stability by up to 30%, while highly polar modifications drastically reduce stability. Immunization studies in A2DR1 humanized mice using the M1 altered peptide ligand (APL) with modification X demonstrated enhanced antigen-specific T cell response compared to the wildtype peptide. Importantly, the APL also showed improved cross-reactive T cell responses. These findings were validated using the clinically relevant 27mer H3K27M peptide, which showed comparable enhancement in immune responses, confirming the broader applicability of our modification strategy. Next steps will include evaluating the therapeutic efficacy of the optimized H3K27M APL in an orthotopic glioma model, with emphasis on tumor regression and survival outcomes. Our approach to optimizing these peptides shows promise for enhancing anti-tumor peptide vaccination, which could lead to durable responses.