Author Description

Nearly half of patients with neuroblastoma present with high-risk disease, which often has poor outcome. The proto-oncogene MYCN is amplified in half of all high-risk neuroblastoma. Patients with high-risk disease inevitably relapse, and show frequent mutational activation of MAPK signaling. High-risk disease remains a clinical enigma, and the lack of actionable targets has prevented sufficient advances in improving survival. To better understand the genetic requirements of high-risk neuroblastoma, we generated a human induced pluripotent stem cell (iPSC) model of the disease. Human iPSC models are superior to genetically engineered mouse models or cell line models at representing the chromosomal landscape and telomere biology of human neuroblastoma. Human iPSCs were differentiated toward trunk neural crest, the cell of origin for neuroblastoma. We optimized protocols to generate this progenitor cell. We transduced candidate and established drivers of neuroblastoma including MYCN and hyperactive mutant ALK (F1174L) and orthotopically implanted the resulting trunk neural crest cells (NCCs) into the renal capsules of immunocompromised mice. The combination of ALK (F1174L) and MYCN significantly accelerated tumor formation. Tumors expressed markers typically found in neuroblastoma on histological and RT-PCR analyses, while lacking markers of other tumors. We found gene expression changes specifically enriched in MYCN/ALK vs MYCN alone-driven tumors, including increased integrin and PI3 kinase signaling. We also introduced CRISPRi/a gene expression modulation into the iPSC system to detangle TERT and MYCN biology that could regulate neuroblastoma tumorigenesis. We further found that MYCN P44L mutation significantly decreases tumor latency compared to WT MYCN misexpression, while CMYC misexpression results in tumors that progress with similar rapidity to MYCN. ChIP-seq and RNA-seq studies in MYCN/CMYC/P44L mutants clarify the differential transcriptional landscapes in va