Human iPSC modeling recapitulates in vivo sympathoadrenal development and reveals an aberrant developmental subpopulation in familial neuroblastoma [Bulk RNA-seq]

Pediatric malignancies, including neuroblastoma, are best understood as disorders of development. Neuroblastoma explicitly represents a failure of sympathoadrenal development. Yet, its molecular pathogenesis remains elusive. The application of an in vitro model of human sympathoadrenal development would allow for studying the normal developmental trajectory. Such a model could also interrogate the early steps toward neuroblastoma transformation utilizing genetic manipulation. However, in vitro models have thus far been unable to generate the cells of interest reliably. We developed and characterized a human in vitro pluripotent stem cell-based model via sequential single-cell RNA sequencing throughout sympathoadrenal development. We demonstrate the power of our model to study early events of the development of human neuroblastoma. We do so by identifying the differences between normal and patient-specific induced pluripotent stem cells, derived from a child with familial neuroblastoma, harboring a germline mutation in the Anaplastic Lymphoma Kinase (ALK) gene. Induced pluripotent stem cells (iPSC) were differentiated along the sympathoadrenergic precursor (SAP) development during a 40 day long differentiation process. We performed bulk RNA sequencing on selected time-points (D0/Start, D3, D7, D11, D16, D21, D23, D25, D27, D29, D31, D35, D40) covering the entire differentiation process. The experiments were repeated four times, twice with a human induced pluripotent stem cell line (J2) and twice with a human embryonic stem cell line (H9) for a total of four biological replicates.