Abstract 6155: Pediatric rhabdomyosarcomas recapitulate the transcriptomic and epigenomic development of skeletal myogenesis

Abstract

Abstract Rhabdomyosarcoma (RMS), is a pediatric tumor that histologically mimics developing skeletal muscle. Importantly, RMS tumors feature heterogeneous expression of the skeletal muscle transcription factor myogenin (MYOG). However, the degree by which RMS tumors model the development of normal skeletal muscle remains unclear. Here, we have utilized droplet-based single-cell RNA-sequencing (scRNA-seq) of 158,070 cells or nuclei isolated from 20 patient RMS tumors (14 embryonal RMS [ERMS], 6 alveolar RMS [ARMS]). Surprisingly, we found that cells from ERMS tumors recapitulate the entire developmental hierarchy of muscle development, with a subset of cells expressing the presomitic mesoderm homeobox gene, MEOX2. In contrast, cells from ARMS tumors were more narrowly restricted to expression of the myoblast- and myocyte-specific transcription factors MYF5 and MYOG. We extended this observation using a developmental indexing approach that leverages single-nuclear transcriptomic data from the Mouse Organogenesis Cell Atlas. By identifying latent cellular states within developing non-malignant embryonal muscle and projecting those latent states onto malignant cells, our developmental index estimates the equivalent myogenic developmental state of each individual malignant cell. ERMS tumors consistently displayed a broad diversity of development indices, even in relapsed tumors; alveolar RMS tumors, however, were restricted to late myoblast and myocyte developmental states. We applied our developmental index to an additional 310,937 cells isolated from 19 orthotopic patient-derived xenografts (O-PDXs), which preserved the developmental diversity of their originating tumor. Next, we used lentiviral barcoding with a high-complexity library to demonstrate that developmental states are interrelated rather than isolated subclones. Additionally, we transduced O-PDX cell suspensions with RFP-encoding retrovirus to lineage trace proliferative cells to show that ERMS tumors transition from immature to mature myogenic states in a unidirectional manner. Finally, we utilized single-cell assay of transposase-accessible chromatin with sequencing (scATAC-seq) in combination with scRNA-seq to show that core regulatory circuit superenhancer accessibility is heterogeneously specific to developmental state within both ERMS and ARMS. Taken together, our findings reveal that pediatric RMS consist of transformed cells that recapitulate the transcriptomic and epigenomic diversity of embryonic muscle. Citation Format: Anand G. Patel, Xiang Chen, Michael Clay, Brittney Gordon, Jim Houston, Asa Karlstrom, Jackie Norrie, Elizabeth Stewart, Michael Dyer. Pediatric rhabdomyosarcomas recapitulate the transcriptomic and epigenomic development of skeletal myogenesis [abstract]. In: Proceedings of the Annual Meeting of the American Association for Cancer Research 2020; 2020 Apr 27-28 and Jun 22-24. Philadelphia (PA): AACR; Cancer Res 2020;80(16 Suppl):Abstract nr 6155.