Abstract 5249: Identification of genome-wide alternative splicing events in high-risk MYCN amplified neuroblastomas

Abstract

Abstract Introduction: High-risk neuroblastoma (H-NB) is an aggressive tumor of the sympathetic nervous system for which there are few predictors of therapeutic response. We sought to determine the association of global alternative splicing (AS) gene expression patterns with NB aggressiveness and MYCN gene status. Methods: As part of the NCI-sponsored Therapeutically Applicable Research to Generate Effective Treatments (TARGET) initiative, we profiled 190 primary human NBs using Affymetrix Human Exon arrays. Tumors were obtained from children who participated in Children's Oncology Group trials with informed consent. The high-risk cohort included tumors with (H-NBA n=48) and without MYCN amplification (H-NBN n=111), a key molecular determinant of clinical aggressiveness. Twenty-four samples from low-risk NB lacking MYCN amplification (L-NB) were also profiled as a comparison group. Robust multichip analysis (RMA) was used for summarization of probe selection regions (PSR) and transcript regions. Finding isoforms using RMA (FIRMA) was used to generate PSR isoform scores. After an unsupervised pre-filtering step based on coefficient of variation, ANOVA analyses were conducted comparing H-NBA or H-NBN against the L-NB group for differential transcript and isoform expression. Last, gene set enrichment analyses (GSEA) comparing high-risk to low-risk NBs were conducted using unfiltered transcript data and the Molecular Signature Database's (MsigDB) canonical gene sets. Results: 1914 out of the 22,011 core transcripts passed our strict prefiltering criteria that required a high coefficient of variation. There was a larger number of genes with significant differential expression between H-NBA vs. L-NB than H-NBN vs. L-NB (23.5% versus 4.6%, P<0.001). There was also significantly larger number of PSRs exhibiting differential FIRMA scores between H-NBA vs. L-NB compared to H-NBN vs. L-NB (14.6% versus 2.3% of PSRs; P<0.001). NTRK1, a gene with known alternative splicing pattern among neuroblastomas, was identified in our data to have lower expression in both H-NBA and H-NBN vs. the L-NB subgroup but showed significant evidence of alternative splicing only in the H-NBA group. GSEA analyses also identified more differentially expressed pathways in H-NBA vs. L-NB than in H-NBN versus L-NB neuroblastomas (66 versus 31 pathways; P<0.001). Pathways related to cell cycle and axon guidance were among the top differentially expressed pathways in the H-NBA vs. L-NB comparison. Conclusion: Significant differences in gene expression and pathway expression are identified among H-NBA compared to H-NBN tumors. In addition, we show for the first time a high degree of alternative splicing events in the H-NBA tumors. These findings could aid in understanding the regulatory networks in the two subgroups of high-risk neuroblastomas and suggest possible therapeutic targets for further validation. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 5249.