[HTML][HTML] Repair of the TGFBI gene in human corneal keratocytes derived from a granular corneal dystrophy patient via CRISPR/Cas9-induced homology-directed …

Y Taketani, K Kitamoto, T Sakisaka, M Kimakura… - Scientific reports, 2017 - nature.com
Y Taketani, K Kitamoto, T Sakisaka, M Kimakura, T Toyono, S Yamagami, S Amano…
Scientific reports, 2017nature.com
Granular corneal dystrophy (GCD) is an autosomal dominant hereditary disease in which
multiple discrete and irregularly shaped granular opacities are deposited in the corneal
stroma. GCD is caused by a point mutation in the transforming growth factor-β-induced
(TGFBI) gene, located on chromosome 5q31. Here, we report the first successful application
of CRISPR-Cas9-mediated genome editing for the correction of a TGFBI mutation in GCD
patient-derived primary corneal keratocytes via homology-directed repair (HDR). To correct …
Abstract
Granular corneal dystrophy (GCD) is an autosomal dominant hereditary disease in which multiple discrete and irregularly shaped granular opacities are deposited in the corneal stroma. GCD is caused by a point mutation in the transforming growth factor-β-induced (TGFBI) gene, located on chromosome 5q31. Here, we report the first successful application of CRISPR-Cas9-mediated genome editing for the correction of a TGFBI mutation in GCD patient-derived primary corneal keratocytes via homology-directed repair (HDR). To correct genetic defects in GCD patient cells, we designed a disease-specific guide RNA (gRNA) targeting the R124H mutation of TGFBI, which causes GCD type 2 (GCD2). An R124H mutation in primary human corneal keratocytes derived from a GCD2 patient was corrected by delivering a CRISPR plasmid expressing Cas9/gRNA and a single-stranded oligodeoxynucleotide HDR donor template in vitro. The gene correction efficiency was 20.6% in heterozygous cells and 41.3% in homozygous cells. No off-target effects were detected. These results reveal a new therapeutic strategy for GCD2; this method may also be applicable to other heredity corneal diseases.
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