Targeted-Deletion of a Tiny Sequence via Prime Editing to Restore SMN Expression.

Spinal muscular atrophy (SMA) is a devastating autosomal recessive motor neuron disease associated with mutations in the survival motor neuron 1 (SMN1) gene, the leading genetic cause of infant mortality. A nearly identical copy gene (SMN2) is retained in almost all patients with SMA. However, SMN2 fails to prevent disease development because of its alternative splicing, leading to a lack of exon 7 in the majority of SMN2 transcripts and yielding an unstable truncated protein. Several splicing regulatory elements, including intronic splicing silencer-N1 (ISS-N1) of SMN2 have been described. In this study, targeted-deletion of ISS-N1 was achieved using prime editing (PE) in SMA patient-specific induced pluripotent stem cells (SMA-iPSCs) with a high efficiency of 7/24. FL-SMN expression was restored in the targeted-deletion iPS clones and their derived motor neurons (iMNs). Notably, the apoptosis of the iMNs, caused by the loss of SMN protein that leads to the hyperactivity of endoplasmic reticulum (ER) stress, was alleviated in targeted-deletion iPSCs derived-iMNs. Thus, this is the first study to demonstrate that the targeted-deletion of ISS-N1 via PE for restoring FL-SMN expression holds therapeutic promise for SMA.