Endoplasmic Reticulum-Targeted Phototherapy Remodels the Tumor Immunopeptidome to Enhance Immunogenic Cell Death and Adaptive Anti-Tumor Immunity

<b>Background:</b> Endoplasmic reticulum (ER)-targeted phototherapy has emerged as a promising approach to amplify ER stress, induce immunogenic cell death (ICD), and enhance anti-tumor immunity. However, its impact on the antigenicity of dying tumor cells remains poorly understood. <b>Methods:</b> Laser activation of the ER-targeted photosensitizer ER-Cy-<i>po</i>NO₂ was performed to investigate its effects on tumor cell antigenicity. Transcriptomic analysis was carried out to assess gene expression changes. Immunopeptidomics profiling was used to identify high-affinity major histocompatibility complex class I (MHC-I) ligands. In vitro functional studies were conducted to evaluate dendritic cell maturation and T lymphocyte activation, while in vivo experiments were performed by combining the identified peptide with poly IC to evaluate anti-tumor immunity. <b>Results:</b> Laser activation of ER-Cy-<i>po</i>NO₂ significantly remodeled the antigenic landscape of 4T-1 tumor cells, enhancing their immunogenicity. Transcriptomic analysis revealed upregulation of antigen processing and presentation pathways. Immunopeptidomics profiling identified multiple high-affinity MHC-I ligands, with IF4G3_986-994 (QGPKTIEQI) showing exceptional immunogenicity. In vitro, IF4G3_986-994 promoted dendritic cell maturation and enhanced T lymphocytes activation. In vivo, the combination of IF4G3_986-994 with poly IC elicited robust anti-tumor immunity, characterized by increased CD8⁺ T lymphocytes infiltration, reduced regulatory T cells (Tregs) in the tumor microenvironment, elevated systemic Interferon-gamma (IFN-γ) levels, and significant tumor growth inhibition without systemic toxicity. <b>Conclusions:</b> These findings establish a mechanistic link between ER stress-driven ICD, immunopeptidome remodeling, and adaptive immune activation, highlighting the potential of ER-targeted phototherapy as a platform for identifying immunogenic peptides and advancing peptide-based cancer vaccines.