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Cells from an X-linked RP (XLRP) patient’s skin punch have gene defect for blindness corrected with gene editing tools

Researchers based at the University of Iowa and Columbia University, New York have provided the first demonstration of successful gene correction in human induced pluripotent stem cells (iPSCs) associated with inherited retinal degeneration. The results of the study, published in the journal Science Reports (DOI: 10.1038/srep19969), show that skin tissue taken from a patient with a retinal degeneration can be induced into pluripotent stem cells from which retinal cells can be generated following correction of the primary genetic mutation. The approach represents a step change from current gene therapy strategies for similar retinal degenerations where whole genes are delivered to treat a disorder. Instead of replacing up to thousands of normal DNA bases, gene-editing represents a highly specific approach directed to correcting a single base mutation to bring about therapeutic repair.


The US-based research team used tissue from two brothers with a diagnosis of RPGR-associated XLRP in which significant vision loss had been documented. Sequencing of their RPGR gene revealed a novel RPGR mutation (c.3070G > T, pGlu1024X) within the ORF15 exon of the gene, a region in which >60% of all XLRP mutations are found. A skin punch biopsy from which patient fibroblasts were cultured in the lab were transformed into patient-specific iPSCs and these cells were observed to differentiate into all three germ layers and express markers of pluripotency including Oct-4, Sox-2, SSEA-4, TRA-1-60 and alkaline phosphatase expression. From this stage of the cell development trajectory the cells could be induced into retinal cells however, to generate a population of retinal cells that were corrected for the RPGR gene mutation the researchers first infected the iPSCs with the gene-editing guide RNA / CRISPR cassette. Sequencing of such transfected cells showed that 13% of reads contained the precise correction of the mutation. The next stage for this technology will be to re-introduce the corrected iPSCs to patients and from there assess the therapeutic benefit in terms of cell viability and visual acuity.


A further rationale for the gene-editing-iPSC approach, according to the researchers, is the persistence of a number of challenges with current gene therapy strategies. For example, the researchers highlighted that while a number of clinical trials for RPE-associated Leber congenital amaurosis (LCA) had shown early signs of safety and benefit, three-year follow-up data indicated that photoreceptor sensitivity may diminish over time and objective measures of visual function may not be as clinically meaningful as had been hoped for. In addition, gene therapy in animal models with comparable doses may produce better outcomes when subsequently compared with human clinical results and this may be due to interspecies genetic differences and other factors. If gene replacement is analogous to replacing a whole book to correct a spelling mistake, gene-editing dispenses with replacing the whole book and instead focuses solely on correcting the actual “spelling error” in the affected gene.