Abstract 95: The landscape of extrachromosomal circular DNA in medulloblastoma subgroups
Authors: Chapman Owen S. , Wang Shanqing , Luebeck Jens , Garancher Alexandra , Larson Jon D. , Lange Joshua , Crawford John , Pomeroy Scott L. , Mischel Paul , Fraenkel Ernest , Wechsler-Reya Robert J. , Bafna Vineet , Mesirov Jill P. , Chavez Lukas 
Published: July 2021

Abstract
Extrachromosomal circular DNA (ecDNA) has been shown to be an important driver of particularly aggressive human cancers. However, it remains unknown to what extent and by which molecular mechanisms ecDNA promotes tumor development and progression in the different molecular subgroups of medulloblastoma (MB), the most common malignant pediatric brain tumor. To answer these questions, we have assembled a multi-institutional retrospective cohort of 472 MB patient samples with available whole genome sequencing (WGS) data, drawing from three non-overlapping public cancer genomic data repositories and covering all four MB subgroups (i.e. WNT, SHH, Group 3 and Group 4). Using genomic cloud computing platforms, local computing resources and recent computational methods for the detection and reconstruction of ecDNA, we find ecDNA in 66 patients (14%) and observe that the presence of ecDNA is associated with significantly poorer outcomes. By subgroup, ecDNA was found in 0/13 WNT (0%), 18/69 SHH (26%), 12/76 G3 (16%) and 14/123 G4 (11%) patients. Affected genomic loci harbor up to hundredfold amplification of oncogenes including MYC, MYCN, and TERT; as well as amplifications of novel putative driver genes involved in chromatin remodeling, DNA repair, and kinase signaling pathways. To investigate the frequency and properties of ecDNA in model systems of MB tumors, we further analyzed WGS data from 24 patient-derived xenografts (PDX) and four cell lines. ecDNA was substantially more frequent in these patient-derived models (17 of 29, 59%) than in our patient cohort. To elucidate the functional regulatory landscapes of ecDNAs in MB, we generated transcriptional (RNA-seq), accessible chromatin (ATAC-seq), and chromatin interaction (Hi-C) profiles from 6 MB tumor samples. In each case, we identify regulatory interactions that cross fusion breakpoints on the ecDNA, representing potential “enhancer rewiring” events that are likely to contribute to transcriptional activation of co-amplified oncogenes. To test this hypothesis, we are currently conducting in-vitro CRISPRi screens targeting regulatory regions on the ecDNA with the aim of testing whether enhancer-rewiring increases the transcription of co-amplified oncogenes and promotes proliferation and cell line growth. In summary, our study analyzes the frequency, diversity and functional relevance of regulatory enhancers co-amplified with oncogenes on ecDNA across MB subgroups and provides strong scientific justification for continued mechanistic studies of ecDNA in MB tumors and tumor models with the potential to uncover new therapeutic approaches.
Citation Format: Owen S. Chapman, Shanqing Wang, Jens Luebeck, Alexandra Garancher, Jon D. Larson, Joshua Lange, John Crawford, Scott L. Pomeroy, Paul Mischel, Ernest Fraenkel, Robert J. Wechsler-Reya, Vineet Bafna, Jill P. Mesirov, Lukas Chavez. The landscape of extrachromosomal circular DNA in medulloblastoma subgroups [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2021; 2021 Apr 10-15 and May 17-21. Philadelphia (PA): AACR; Cancer Res 2021;81(13_Suppl):Abstract nr 95.