Insights into Neuroblastoma Initiation and Disease Progression Through integrative Genomics and Epigenomics
In this dissertation, we use integrative genomics to shed new insights into the molecular lesions and mechanisms that drive neuroblastoma. In Part 1, we use imputation and epigenetic profiling in order to identify the causal germline SNP that drives differential susceptibility to neuroblastoma at the LMO1 oncogene locus. In Part 2, we use whole genome sequencing and Bayesian statistical modeling to understand the clonal evolution that occurs between diagnosis and relapse. Part 1: Neuroblastoma is a pediatric malignancy that typically arises in early childhood, and is derived from the developing sympathetic nervous system. A previous genome-wide association study identified common polymorphisms at the LMO1 gene locus that are highly associated with neuroblastoma susceptibility and oncogenic addiction to LMO1 in the tumor cells. Here we investigate the causal DNA variant at this locus. We show that SNP rs2168101 G>T is the most highly associated variant and resides in a super-enhancer defined by extensive acetylation of histone H3 lysine 27 within the first intron of LMO1. The ancestral G allele that is associated with tumor formation resides in a conserved GATA transcription factor binding motif. We show that the newly evolved protective TATA allele ablates GATA3 binding and enhancer activity, and is associated with decreased total and allele-specific LMO1 expression in neuroblastoma primary tumors. These findings indicate that a recently evolved polymorphism within a super-enhancer element in the first intron of LMO1 influences neuroblastoma susceptibility through differential GATA transcription factor binding and direct modulation of LMO1 expression in cis. Part 2: The majority of high-risk neuroblastomas initially respond to chemotherapy, but over half of patients will experience therapy-resistant relapses which are nearly always fatal. The molecular defects driving relapse and drug resistance are unknown. We performed whole genome sequencing of 23 paired diagnostic and relapsed neuroblastomas, and corresponding normal lymphocyte DNAs, to define genetic alterations associated with relapse. Unbiased pathway analysis of the somatic mutations detected in the relapse tissues identified a strong enrichment in genes associated with RAS-MAPK signaling (18 of 23 patients). These RAS-MAPK mutations were clonally enriched at relapse and exist within clonal or major subclonal tumor populations. Similar MAPK pathway mutations were detected in 11 of 18 human neuroblastoma-derived cell lines, and these lesions are predicted to be sensitive to small molecule inhibition of MEK in vitro and in vivo. In this study of 23 neuroblastoma cases, MAPK pathway mutations were highly enriched in the relapsed genomes, providing a potential biomarker for new therapeutic approaches to chemotherapy refractory disease. Collectively, these studies provide important insights into the genetic and epigenetic factors driving neuroblastoma, and suggest new opportunities for pathway-targeted therapies.
Sharon J. Diskin