Spinal Muscular Atrophy Phenotype Is Ameliorated Either By SMN Increase Or Modulation Of Secondary Cell Death Events With RNA Therapy (S56.006)

OBJECTIVE: The aim of the study was to develop therapeutic strategies based on RNA modulation for Spinal Muscular Atrophy (SMA) using a stem cell model.BACKGROUND: Spinal muscular atrophy (SMA) is a genetic disorder caused by mutations of the survival motor neuron gene (SMN1) leading to the degeneration of motor neurons, and clinically causing paralysis and death. No effective treatment is available so far, but antisense therapy to increase the SMN level is a promising strategy.DESIGN/METHODS: We generated induced pluripotent stem cell (iPSC) lines derived from human skin fibroblasts with lentiviral constructs and with a non-viral non integrating method based on the expression of reprogramming factors with episomal vectors. We differentiated iPSCs using a protocol to promote motor neuronal phenotype. The phenotype of these cells was analyzed by morphological, functional, gene expression, and protein analysis. RNA strategy based on antisense morpholino, ExSpeU1 and siRNA aiming at increasing SMN level or inhibiting Fas activation were tested.RESULTS: We show here that SMA iPSC-motor neurons recapitulate the disease phenotype with a significantly fewer and smaller motor neurons at later time periods in culture compared to wild-type subject iPSC lines. These features were ameliorated in SMA motor neurons treated with antisense morpholino or ExSpeU1 that rescue SMN level. During motor neuron development, SMA lines showed an increase in Fas ligand-mediated apoptosis and increased caspase-8 activation. Importantly, this could be mitigated by Fas silencing with siRNA.CONCLUSIONS: Our data confirm the utility of SMA iPSCs as in vitro disease model, suggesting that RNA therapy can be a possible therapeutic strategy for SMA trough SMN up-regulation and modulation of disease pathways that can be achieved with different therapeutic tools.