Mapping the Modification Landscape of MHC-I Epitopes: A Framework for Immunogenic Peptidomimetic Antigen Design

ABSTRACT Peptide-based cancer vaccines offer a promising strategy to target tumor-specific neoantigens. This approach is increasingly critical as post-translationally modified peptides, driven by altered tumor metabolism, emerge as a unique class of neoantigens. Because these chemically distinct epitopes cannot be genetically encoded by mRNA or viral platforms, synthetic peptide vaccines are poised to be the primary route to target these types of neoantigens. Yet, their clinical translation is restricted by poor metabolic stability, limited intracellular permeability, and structural requirements for MHC-I binding and T cell receptor (TCR) recognition. Although peptidomimetic modifications have been widely explored to improve pharmacokinetics, their impact on antigen presentation and immune recognition remains poorly understood. Here, we undertook a comprehensive evaluation of peptidomimetics geared at MHC-I neoantigens and generated a diverse library of systematically modified peptides that incorporate backbone N -methylation, peptoid substitution, and stereochemical inversion. Integrated assays revealed a highly position-dependent tolerance to peptidomimetic modifications, while subsequent combinatorial designs demonstrated non-additive effects on the balance between immunogenicity and pharmacokinetics. Collectively, these findings establish design principles and provide a framework for balancing immune recognition with enhanced stability and permeability in peptidomimetic antigen design.