Allele-specific gene-editing approach for vision loss restoration in -associated retinitis pigmentosa.

Mutant is the most frequent genetic cause of autosomal dominant retinitis pigmentosa (adRP). Here, we developed an allele-specific gene-editing therapeutic drug to selectively target the human T17M mutant allele while leaving the wild-type allele intact for the first time. We identified a Cas9 (SaCas9) guide RNA that was highly active and specific to the human T17M allele. experiments using HEK293T cells and patient-specific induced pluripotent stem cells (iPSCs) demonstrated active nuclease activity and high specificity. Subretinal delivery of a single adeno-associated virus serotype 2/8 packaging SaCas9 and single guide RNA (sgRNA) to the retinas of the humanized mice showed that this therapeutic drug targeted the mutant allele selectively, thereby downregulating the mutant mRNA expression. Administration of this therapeutic drug resulted in a long-term (up to 11 months after treatment) improvement of retinal function and preservation of photoreceptors in the heterozygous mutant humanized mice. Our study demonstrated a dose-dependent therapeutic effect . Unwanted off-target effects were not observed at the whole-genome sequencing level. Our study provides strong support for the further development of this effective therapeutic drug to treat -T17M-associated adRP, also offers a generalizable framework for developing gene-editing medicine. Furthermore, our success in restoring the vision loss in the suffering humanized mice verifies the feasibility of allele-specific CRISPR/Cas9-based medicines for other autosomal dominant inherited retinal dystrophies.