Gene Therapy of Dominant CRX-Leber Congenital Amaurosis using Patient Stem Cell-Derived Retinal Organoids.

Mutations in the photoreceptor transcription factor gene cone-rod homeobox (CRX) lead to distinct retinopathy phenotypes, including early-onset vision impairment in dominant Leber congenital amaurosis (LCA). Using induced pluripotent stem cells (iPSCs) from a patient with CRX-I138fs48 mutation, we established an in vitro model of CRX-LCA in retinal organoids that showed defective photoreceptor maturation by histology and gene profiling, with diminished expression of visual opsins. Adeno-associated virus (AAV)-mediated CRX gene augmentation therapy partially restored photoreceptor phenotype and expression of phototransduction-related genes as determined by single-cell RNA-sequencing. Retinal organoids derived from iPSCs of a second dominant CRX-LCA patient carrying K88N mutation revealed the loss of opsin expression as a common phenotype, which was alleviated by AAV-mediated augmentation of CRX. Our studies provide a proof-of-concept for developing gene therapy of dominant CRX-LCA and other CRX retinopathies.