Abstract 2850: Noncoding regulatory mutations as driving event for the oncogenic core regulatory circuitries of neuroblastoma
Authors: Mario Capasso, Vincenzo Aievola, Vito Alessandro Lasorsa, Annalaura Montella, Ferdinando Bonfiglio, Marianna Avitabile, Teresa Maiorino, Matilde Tirelli, Giuseppe D'Alterio, Matthias Fischer, Frank Westermann, Achille Iolascon
Journal: Cancer Research (2024)

Abstract Background and Aim: Neuroblastoma (NB) is the most aggressive solid tumor of infancy and arises from neural-crest-derived progenitor cells. It is composed by cell types reflecting developmental stages of the adrenergic lineage: mesenchymal (MES) and adrenergic (ADRN) cell types. These cell identities result from the activity of NB specific Core Regulatory Circuitries (CRCs), transcription factor (TF) sets that co-occupy regulatory regions together impacting the expression of their own genes and those of lineage-specific ones governing cell state transitions and cellular identities. Our hypothesis is that noncoding somatic single nucleotide variants (SNVs) located in NB CRCs TF binding sites (TFBSs) underlie NB CRCs activity, affecting cellular differentiation and promoting tumor onset. We aim to investigate such pattern of regulatory elements and identify putative driver SNVs, exploring their role and characterizing their action mechanism. Methods: Transcriptionally active TFBSs (aTFBSs) bound by ADRN CRC TFs (GATA3, HAND2, ISL1, TBX2, MYCN, PHOX2B, ASCL1 and LMO1) were identified integrating 27 ChIP-seq and 11 ATAC-seq experiments in 6 NB ADRN cell lines (SKNBE2C, Kelly, NGP, COGN415, LAN5 and NB1643). In particular, aTFBSs were defined as ChIP-seq peaks overlapping ATAC-seq signals in the same cell line. SNVs from 317 NB whole genomes were mapped on aTFBSs and the enrichment of mutations in core regions respect to flanking ones was tested using Fisher test. Observed mutation rate in aTFBSs was compared with background mutation rate due to local sequence context. SNVs in mutated aTFBSs were selected and their impact on TF binding affinity was evaluated with FABIAN-variant and motifbreakR tools. aTFBSs target-genes were identified through promoter capture HiC (CHiC) in ADRN NB cells (SKNBE2C and SHSY5Y). Results: We found a significant (FDR &amp;lt; 0.05) mutation enrichment in aTFBSs of ISL1, PHOX2B, TBX2, GATA3 and MYCN, suggesting a common somatic mutation burden for a subset of ADRN CRC TFBSs in ADRN NB cells. Starting from 839 SNVs falling in significantly mutated aTFBSs, we selected 269 mutations impacting the binding of ADRN CRC TFs. This mutations subset mapped in aTFBSs interacting with genes that significantly enriched pathways involved in neuronal development and migration (FDR &amp;lt; 0.05), thus suggesting their possible role in affecting cell identity definition. As an example, chr6:137753475:G&amp;gt;T mapped in aTFBSs interacting with the promoter of OLIG3, which takes part in neuronal differentiation. Interestingly, chr6:137753475:G&amp;gt;T was predicted to break the binding to ISL1 (FABIAN score: -0.24), a key regulator of ADRN CRC. Conclusions: These results demonstrate that somatic noncoding SNVs can act synergistically to promote NB onset, affecting CRCs activity and, in turn, the normal cell identity definition. Citation Format: Mario Capasso, Vincenzo Aievola, Vito Alessandro Lasorsa, Annalaura Montella, Ferdinando Bonfiglio, Marianna Avitabile, Teresa Maiorino, Matilde Tirelli, Giuseppe D'Alterio, Matthias Fischer, Frank Westermann, Achille Iolascon. Noncoding regulatory mutations as driving event for the oncogenic core regulatory circuitries of neuroblastoma [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 2850.