Abstract

Pancreatic ductal adenocarcinoma (PDAC) is refractory to immunotherapy due to its immunologically cold microenvironment and the scarcity of mutation-derived neoantigens. Here, we introduce NeoAPP, a computational tool designed to systematically decode neoantigens arising from tumor-specific transcripts (TSTs) generated by transcriptional dysregulation. By integrating multi-cohort transcriptomic data from 413 PDAC samples, NeoAPP identified a median of 351 neoantigens per sample, originating from 56 neoantigen-encoding TSTs (neoTSTs), exceeding mutation-derived neoantigens in both abundance and patient coverage. Mechanistic analyses show that noncanonical splicing junctions and transposable element activation drive neoantigen generation, while alternative promoter usage regulated by FOXA2 constitutes a potential major source of neoTSTs. In addition, a subset of neoTSTs was detectable in plasma-derived extracellular vesicles and was associated with patient survival, suggesting their potential as minimally invasive biomarkers. Vaccination with neoTSTs induced CD8 + T cell responses in HLA-A*02:01 and HLA-A*11:01 transgenic mice and suppressed tumor growth in syngeneic PDAC models. Collectively, this work establishes TST-derived neoantigens as a dominant and therapeutically actionable antigen reservoir in PDAC, advancing a transcriptome-guided framework for neoantigen discovery with potential to overcome immune resistance in low-mutation cancers.