In vivo genome editing via CRISPR/Cas9-mediated homology-independent targeted integration for Bietti crystalline corneoretinal dystrophy treatment
Autor: | Xiang Meng, Ruixuan Jia, Xinping Zhao, Fan Zhang, Shaohong Chen, Shicheng Yu, Xiaozhen Liu, Hongliang Dou, Xuefeng Feng, Jinlu Zhang, Ni Wang, Boling Xu, Liping Yang |
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Jazyk: | angličtina |
Rok vydání: | 2024 |
Předmět: | |
Zdroj: | Nature Communications, Vol 15, Iss 1, Pp 1-18 (2024) |
Druh dokumentu: | article |
ISSN: | 2041-1723 86539132 |
DOI: | 10.1038/s41467-024-48092-9 |
Popis: | Abstract 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-Cyp4v3 mut/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-Cyp4v3 mut/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. |
Databáze: | Directory of Open Access Journals |
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