Long-read isoform sequencing reveals survival-associated splicing in breast cancer

Tumors display widespread transcriptome alterations, but the full repertoire of isoform-level alternative splicing in cancer is not known. We developed a long-read RNA sequencing and analytical platform that identifies and annotates full-length isoforms, and infers tumor-specific splicing events. Application of this platform to breast cancer samples vastly expands the known isoform landscape of breast cancer, identifying thousands of previously unannotated isoforms of which ~30% impact protein coding exons and are predicted to alter protein localization and function, including of the breast cancer-associated genes ESR1 and ERBB2 . We performed extensive cross-validation with -omics data sets to support transcription and translation of novel isoforms. We identified 3,059 breast tumor-specific splicing events, including 35 that are significantly associated with patient survival. Together, our results demonstrate the complexity, cancer subtype-specificity, and clinical relevance of novel isoforms in breast cancer that are only annotatable by LR-seq, and provide a rich resource of immuno-oncology therapeutic targets. ### Competing Interest Statement D.F.T.V., A.D.M. and J.B. are named inventors in the patent application PCT/US2019/039150 (pending) filed by The Jackson Laboratory in June 26, 2019. This patent application covers the method used for discovery of tumor-specific AS events described in Fig. 4. A.D.M. owns shares of Pacific Biosciences. J.B. serves on the Board of Directors (BOD) for Neovacs, is a BOD member and stock holder for Ascend Biopharmaceuticals, Scientific Advisory Board (SAB) member for Cue Biopharma, and stock holder for Sanofi. The remaining authors declare that they have no competing interests.