Abstract B10: Rhabdomyosarcoma requires MYC family genomic events to pathogenically subvert core-regulatory circuitry
Authors: Durbin Adam D. , Kugener Guillaume , Zimmerman Mark W. , Yan Chuan , Dharia Neekesh V. , Frank Elizabeth S. , Chen Xiang , Ross Ken N. , Paolella Brenton , Krill-Burger Michael , Root David E. , Boehm Jesse S. , Vazquez Francisca , Hong Andrew L. , Tsherniak Aviad , Langenau David M. , Hahn William C. , Golub Todd R. , Abraham Brian J. , Young Richard A. , Look A. Thomas , Stegmaier Kimberly 
Published: August 2020

Abstract
Rhabdomyosarcoma (RMS) is the most common soft-tissue sarcoma of childhood. Despite multimodality therapy and trials of molecularly targeted agents, limited improvements in overall survival have been realized for patients with high-risk disease. Thus, we aimed to determine the landscape of tumor-specific gene dependencies that underlie tumorigenesis in RMS and therefore provide a valuable group of targets for the development of novel therapeutics. Using unbiased genome-scale CRISPR-Cas9 approaches, we identified a set of RMS-specific tumor dependencies involved in tumor cell growth and survival. RMS dependencies were enriched for nucleic acid binding proteins, including transcription factors (TFs). We then used genome-wide chromatin-immunoprecipitation coupled to high-throughput sequencing analysis to demonstrate that a small number of essential TFs—MYCN, MYOD1, TCF12, SOX8, ZEB2, ZNF217, and SIX1—are members of the transcriptional core regulatory circuitry (CRC) that maintains the malignant cell state of RMS. Both c-MYC and MYCN were associated with gene and enhancer copy number increases in cell lines and primary tumors and represented strong dependencies in the RMS cell lines screened. c-MYC and MYCN function to similarly invade and regulate the CRC in respectively dependent cells. To disable the CRC, we tested A485, an inhibitor of the histone acetyltransferase enzymes involved in the establishment of super-enhancer elements that are associated with high level expression of the CRC factors. A485 caused a reversible and rapid loss of CRC factor and c-MYC and/or MYCN expression, and prolonged treatment resulted in cell cycle arrest, differentiation, and apoptosis in vitro and in vivo. This phenotype is rescued by exogenous re-expression of either c-MYC or MYCN in a manner insensitive to A485, indicating a mechanism by which these genes subvert a myogenic CRC to produce an oncogenic fate. This study defines a common set of critical dependency genes in RMS and identifies key genomic events surrounding the c-MYC and MYCN loci that lead to elevated expression and tumorigenesis.
Citation Format: Adam D. Durbin, Guillaume Kugener, Mark W. Zimmerman, Chuan Yan, Neekesh V. Dharia, Elizabeth S. Frank, Xiang Chen, Ken N. Ross, Brenton Paolella, Michael Krill-Burger, David E. Root, Jesse S. Boehm, Francisca Vazquez, Andrew L. Hong, Aviad Tsherniak, David M. Langenau, William C. Hahn, Todd R. Golub, Brian J. Abraham, Richard A. Young, A. Thomas Look, Kimberly Stegmaier. Rhabdomyosarcoma requires MYC family genomic events to pathogenically subvert core-regulatory circuitry [abstract]. In: Proceedings of the AACR Special Conference on the Advances in Pediatric Cancer Research; 2019 Sep 17-20; Montreal, QC, Canada. Philadelphia (PA): AACR; Cancer Res 2020;80(14 Suppl):Abstract nr B10.