Long-range alternative splicing contributes to neoantigen specificity in glioblastoma

Recent advances in neoantigen research have accelerated the development of immunotherapies for cancers, such as glioblastoma (GBM). Neoantigens resulting from genomic mutations and dysregulated alternative splicing have been studied in GBM. However, these studies have primarily focused on annotated alternatively-spliced transcripts, leaving non-annotated transcripts largely unexplored. Circular ribonucleic acids (circRNAs), abnormally regulated in tumors, are correlated with the presence of non-annotated linear transcripts with exon skipping events. But the extent to which these linear transcripts truly exist and their functions in cancer immunotherapies remain unknown. Here, we found the ubiquitous co-occurrence of circRNA biogenesis and alternative splicing across various tumor types, resulting in large amounts of long-range alternatively-spliced transcripts (LRs). By comparing tumor and healthy tissues, we identified tumor-specific LRs more abundant in GBM than in normal tissues and other tumor types. This may be attributable to the upregulation of the protein quaking in GBM, which is reported to promote circRNA biogenesis. In total, we identified 1057 specific and recurrent LRs in GBM. Through in silico translation prediction and MS-based immunopeptidome analysis, 16 major histocompatibility complex class I-associated peptides were identified as potential immunotherapy targets in GBM. This study revealed long-range alternatively-spliced transcripts specifically upregulated in GBM may serve as recurrent, immunogenic tumor-specific antigens.