Merlin-deficient iPSCs show altered pluripotency and constitute a potential in vitro model for NF2-related schwannomas

NF2-related schwannomatosis is an autosomal dominant syndrome that predisposes to the development of benign tumors of the nervous system. Schwannomas, particularly bilateral vestibular schwannomas (VS), are the most characteristic features of the disease. These tumors are caused by the bi-allelic inactivation of the NF2 gene in a cell of the Schwann cell lineage. Our current understanding of the molecular pathogenesis of the NF2 gene, as well as the development of new effective therapies is hampered by the absence of human non-perishable cell-based bearing distinct NF2 pathogenic variants. With this aim, we generated and characterized three isogenic paired induced pluripotent stem cell (iPSC) lines with single or bi-allelic inactivation of NF2 by combining the direct reprogramming of VS cells with the use of CRISPR/Cas9 editing. Our results show a critical function of NF2 for the maintenance of a stable pluripotent state. However, we were able to nudge them towards the Neural Crest-Schwann Cell (NC-SC) axis by applying a 3D Schwann cell differentiation protocol. NF2(+/-) and NF2(-/-) spheroids homogeneously expressed classical markers of the NC-SC lineage. In addition, NF2(-/-) SC-like spheroids showed dysregulation of multiple signaling pathways already described for merlin-deficient SC, and altered in human schwannomas. Therefore, NF2(+/-) and NF2(-/-) SC-like spheroids can represent a bona fide human in vitro cellular model to study the role of NF2 pathogenesis.