Abstract 3751: A pan-cancer intracellular tumor antigen atlas

T-cell-based immunotherapies have transformed the treatment, offering cures to subsets of cancer patients. The rate-limiting step in developing safe and effective new immunotherapies is the identification of antigens that are tumor-specific, highly abundant, recurrent, and homogeneously expressed. Current approaches have revealed few molecules that fulfill these criteria, restricting the patient populations that benefit from immunotherapies. HLA molecules present a snapshot of the human proteome on the cell surface, rendering intracellular antigens accessible for various cancer immunotherapies. While many publicly available immunopeptidome dataset exist, exhaustively selecting such HLA antigens requires not only canonical proteome, but genomics-guided search space to include all molecular alterations - a challenge that hasn’t been tackled until now. We addressed this challenge by performing a comprehensive pan-cancer analysis encompassing 9,270 RNA-Seq and 1,520 immunopeptidome across 21 tumor types. Our approach provides an exhaustive search for antigens arising from both canonical and 15 non-canonical alternations known to result from tumor-associated molecular events, such as alternative splicing and transposable elements. Our pan-cancer analysis uncovered numerous novel tumor antigens across tumor histologies that are suitable for immunotherapeutic approaches including PC-CARs, TCRs and vaccines. We validated several tumor-specific antigens using targeted mass spectrometry, peptide-HLA refolding and ribosome profiling. For example, we demonstrate that antigens derived from alternatively spliced peptides can be preferentially presented on the cell surface, highlighting a skipped exon event in PMEL that we suggest will provide a superior target to the canonical PMEL peptide targeted in the clinic. Notably, we identified widespread presentation of ORF2 peptides originating from the L1 element, which have remained undetectable by conventional methods, suggesting that ORF2 may encode a short-lived protein prone to proteasomal degradation. We also observed that tumor-resident pathogens may invade colon and ovarian cancer cells, providing new potential avenues for therapeutic targeting. We highlight antigens derived from cryptic open reading frames located in the untranslated regions of neuroblastoma, expanding the repertoire of potential targets in immuno-cold cancers. We have made this atlas available to the scientific community through a web portal and released our search tool hosted on the Cancer Genome Cloud (CGC), enabling identification of 16 classes of tumor-specific antigens from any in-house dataset. We expect that the pan-cancer atlas and this toolkit will significantly accelerate discovery of actionable targets for cancer immunotherapy and reveal new insights in antigen presentation and tumor biology. Citation Format: Guangyuan Li, Omar U. Guzmán-Bringas, Aman Sharma, Xinya Liu, Darren Taylor, Rachel Yamin, Maxence Dellacherie, Rowan Beck, Tariq Khoyratty, Zelia Worman, Mark Yarmarkovich. A pan-cancer intracellular tumor antigen atlas [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2025; Part 1 (Regular Abstracts); 2025 Apr 25-30; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2025;85(8_Suppl_1):Abstract nr 3751.