Abstract 5710: Identification of convergent gene repression mechanisms through integrative genomic and DNA methylation analysis in TRACERx
Authors: Gimeno-Valiente Francisco , Castignani Carla , Larose-Cadieux Elizabeth , Chen Kezhong , Mensah Nana , Chervova Olga , Watkins Thomas , Dhami Pawan , Vaikkinen Heli , Feber Andrew , Consortium TRACERx , Demeulemeester Jonas , Tanic Miljana , Beck Stephan , Van Loo Peter , Swanton Charles , Kanu Nnennaya 
Published: June 2022

Abstract
Introduction: Lung TRACERx is a prominent study employing multi-region and longitudinal multi-omics sequencing to unravel the evolutionary trajectories of lung cancer. Aberrant DNA methylation patterns have been widely described in nearly all human cancers, yet their interplay with DNA mutations in lung cancer is not well understood. Incorporating the contribution of epigenetic modifications to cancer evolution trajectories within TRACERx could improve our understanding of the intricate relationship between genetic and epigenetic changes in non-small cell lung cancer (NSCLC) evolution.
Methods: Multi-region sampling from 38 TRACERx patients including 112 tumor regions and 37 matched normal adjacent tissue samples was performed. Reduced representation bisulfite sequencing (RRBS) was performed to assess DNA methylation and the CAMDAC (Larose-Cadieux et al, 2020) was applied to estimate purified tumor methylation rates and correct for copy number changes. Whole exome sequencing and somatic copy number alterations (SCNAs) were inferred using the ASCAT tool (Van Loo et al, 2010) and Methsig (Pan et al, 2021) was performed to discover new methylation driver genes.
Results: Using multi-region sequencing, we identified ubiquitous hypermethylation of 29 known driver genes in both lung adenocarcinoma (LUAD) and squamous cell lung cancer (LUSC), together with an additional 9 and 27 genes exclusive to LUSC and LUAD, respectively. We also identified 13 and 7 driver genes non-ubiquitously hypermethylated exclusively in LUSC and LUAD, respectively. Using a differential methylation based approach, we describe a method to determine the extent of intra-tumor methylation heterogeneity akin to established ITH scores based on genomics data. In addition, we report the identification of novel subtype-specific methylation driver genes enriched in HOX family members which are related to cancer progression. Through integration of DNA methylation and genomic sequencing data, we identify parallel mechanisms contributing towards ubiquitous tumor suppressor gene alterations. At the patient level, multiple driver genes such as NSD1, GATA3 and MGA were subject to repression by both copy number loss and DNA hypermethylation. Finally, we describe dosage-compensation of genes such as the Notch ligands JAG2 and DLK1 that are proximal to amplified oncogenes and hypermethylated during tumor evolution.
Conclusion: We describe the contribution of DNA methylation and genomic alterations to altering the landscape of NSCLC. Leveraging DNA methylation, we can determine the extent of convergent repression mechanisms in different regions of the same tumor, assess DNA methylation heterogeneity, and discover DNA methylation-based driver genes in NSCLC.
Citation Format: Francisco Gimeno-Valiente, Carla Castignani, Elizabeth Larose-Cadieux, Kezhong Chen, Nana Mensah, Olga Chervova, Thomas Watkins, Pawan Dhami, Heli Vaikkinen, Andrew Feber, TRACERx Consortium, Jonas Demeulemeester, Miljana Tanic, Stephan Beck, Peter Van Loo, Charles Swanton, Nnennaya Kanu. Identification of convergent gene repression mechanisms through integrative genomic and DNA methylation analysis in TRACERx [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 5710.