Genome editing in patient iPSC efficiently corrects the most prevalent USH2A mutations and reveals intriguing mutant mRNA expression profiles

Abstract Inherited retinal dystrophies (IRDs) are characterized by progressive photoreceptor degeneration and vision loss. Usher syndrome (USH) is a syndromic IRD characterized by retinitis pigmentosa (RP) and hearing loss. USH is clinically and genetically heterogeneous, and the most prevalent causative gene is USH2A. USH2A mutations also account for a large number of isolated autosomal recessive RP (arRP) cases. This high prevalence is due to two recurrent USH2A mutations, c.2276G>T and c.2299delG. Due to the large size of the USH2A cDNA, gene augmentation therapy is inaccessible. However, CRISPR/Cas9-mediated genome editing is a viable alternative. We used eSpCas9 to successfully achieve seamless correction of the two most prevalent USH2A mutations in induced pluripotent stem cells (iPSCs) of patients with USH or arRP. Our results highlight features that promote high target efficacy and specificity of eSpCas9. Consistently, we did not identify any off-target mutagenesis in the corrected iPSC, which also retained pluripotency and genetic stability. Furthermore, analysis of USH2A expression unexpectedly identified aberrant mRNA levels associated with the c.2276G>T and c.2299delG mutations that were reverted following correction. Taken together, our efficient CRISPR/Cas9-mediated strategy for USH2A mutation correction brings hope for a potential treatment for USH and arRP patients.