%0 Journal Article
%T In vivo genome editing via CRISPR/Cas9-mediated homology-independent targeted integration for Bietti crystalline corneoretinal dystrophy treatment.
%A Meng X
%A Jia R
%A Zhao X
%A Zhang F
%A Chen S
%A Yu S
%A Liu X
%A Dou H
%A Feng X
%A Zhang J
%A Wang N
%A Xu B
%A Yang L
%J Nat Commun
%V 15
%N 1
%D 2024 May 6
%M 38710738
%F 17.694
%R 10.1038/s41467-024-48092-9
%X Bietti crystalline corneoretinal dystrophy (BCD) is an autosomal recessive chorioretinal degenerative disease without approved therapeutic drugs. It is caused by mutations in CYP4V2 gene, and about 80% of BCD patients carry mutations in exon 7 to 11. Here, we apply CRISPR/Cas9 mediated homology-independent targeted integration (HITI)-based gene editing therapy in HEK293T cells, BCD patient derived iPSCs, and humanized Cyp4v3 mouse model (h-Cyp4v3mut/mut) using two rAAV2/8 vectors via sub-retinal administration. We find that sgRNA-guided Cas9 generates double-strand cleavage on intron 6 of the CYP4V2 gene, and the HITI donor inserts the carried sequence, part of intron 6, exon 7-11, and a stop codon into the DNA break, achieving precise integration, effective transcription and translation both in vitro and in vivo. HITI-based editing restores the viability of iPSC-RPE cells from BCD patient, improves the morphology, number and metabolism of RPE and photoreceptors in h-Cyp4v3mut/mut mice. These results suggest that HITI-based editing could be a promising therapeutic strategy for those BCD patients carrying mutations in exon 7 to 11, and one injection will achieve lifelong effectiveness.