Induced Pluripotent Stem Cells and Genome-Editing Tools in Determining Gene Function and Therapy for Inherited Retinal Disorders

Benati, Daniela; Leung, Amy; Perdigão, Pedro; Toulis, Vasileios; Spuy, Jacqueline van der; Recchia, Alessandra

doi:10.3390/ijms232315276

Cited by 2 publications

(2 citation statements)

References 122 publications

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“…The low efficiency of HDR mediated KI in hPSCs has also been improved by a number of methodologies [ 10 , 66 , 67 ]. As a result, Cas9 has become a standardized approach for gene perturbation or correction in hPSCs as evidenced by numerous review articles [ 68 – 70 ]. However, the recently developed pencil like-editing tools (i.e.…”

Section: Applications Of “Pencil” In Hpscsmentioning

confidence: 99%

Gene editing with ‘pencil’ rather than ‘scissors’ in human pluripotent stem cells

Park

Lee

et al. 2023

Stem Cell Res Ther

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Owing to the advances in genome editing technologies, research on human pluripotent stem cells (hPSCs) have recently undergone breakthroughs that enable precise alteration of desired nucleotide bases in hPSCs for the creation of isogenic disease models or for autologous ex vivo cell therapy. As pathogenic variants largely consist of point mutations, precise substitution of mutated bases in hPSCs allows researchers study disease mechanisms with “disease-in-a-dish” and provide functionally repaired cells to patients for cell therapy. To this end, in addition to utilizing the conventional homologous directed repair system in the knock-in strategy based on endonuclease activity of Cas9 (i.e., ‘scissors’ like gene editing), diverse toolkits for editing the desirable bases (i.e., ‘pencils’ like gene editing) that avoid the accidental insertion and deletion (indel) mutations as well as large harmful deletions have been developed. In this review, we summarize the recent progress in genome editing methodologies and employment of hPSCs for future translational applications.

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Section: Applications Of “Pencil” In Hpscsmentioning

confidence: 99%

Gene editing with ‘pencil’ rather than ‘scissors’ in human pluripotent stem cells

Park

Lee

et al. 2023

Stem Cell Res Ther

View full text Add to dashboard Cite

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“…Patient-derived iPSC are valuable tools for in vitro disease modelling (disease-in-a-dish) and drug screening for retinal diseases [ 40 ]. In addition, iPSC with corrected mutations using CRISPR/Cas9 editing can be used to study gene function or generate cell therapy products as an alternative treatment to replace the degenerated retina in advanced disease [ 41 – 44 ]. At present, only two PROM1 -related iPSC lines, both associated with the same mutation c.619G > T (p.E207X), have been reported [ 45 ].…”

Section: Introductionmentioning

confidence: 99%

Clinical exome analysis and targeted gene repair of the c.1354dupT variant in iPSC lines from patients with PROM1-related retinopathies exhibiting diverse phenotypes

Puertas-Neyra,

Coco-Martin,

Hernandez-Rodriguez

et al. 2024

Stem Cell Res Ther

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Background Inherited retinal dystrophies (IRD) are one of the main causes of incurable blindness worldwide. IRD are caused by mutations in genes that encode essential proteins for the retina, leading to photoreceptor degeneration and loss of visual function. IRD generates an enormous global financial burden due to the lack of understanding of a significant part of its pathophysiology, molecular diagnosis, and the near absence of non-palliative treatment options. Patient-derived induced pluripotent stem cells (iPSC) for IRD seem to be an excellent option for addressing these questions, serving as exceptional tools for in-depth studies of IRD pathophysiology and testing new therapeutic approaches. Methods From a cohort of 8 patients with PROM1-related IRD, we identified 3 patients carrying the same variant (c.1354dupT) but expressing three different IRD phenotypes: Cone and rod dystrophy (CORD), Retinitis pigmentosa (RP), and Stargardt disease type 4 (STGD4). These three target patients, along with one healthy relative from each, underwent comprehensive ophthalmic examinations and their genetic panel study was expanded through clinical exome sequencing (CES). Subsequently, non-integrative patient-derived iPSC were generated and fully characterized. Correction of the c.1354dupT mutation was performed using CRISPR/Cas9, and the genetic restoration of the PROM1 gene was confirmed through flow cytometry and western blotting in the patient-derived iPSC lines. Results CES revealed that 2 target patients with the c.1354dupT mutation presented monoallelic variants in genes associated with the complement system or photoreceptor differentiation and peroxisome biogenesis disorders, respectively. The pluripotency and functionality of the patient-derived iPSC lines were confirmed, and the correction of the target mutation fully restored the capability of encoding Prominin-1 (CD133) in the genetically repaired patient-derived iPSC lines. Conclusions The c.1354dupT mutation in the PROM1 gene is associated to three distinct AR phenotypes of IRD. This pleotropic effect might be related to the influence of monoallelic variants in other genes associated with retinal dystrophies. However, further evidence needs to be provided. Future experiments should include gene-edited patient-derived iPSC due to its potential as disease modelling tools to elucidate this matter in question. Graphical Abstract

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Induced Pluripotent Stem Cells and Genome-Editing Tools in Determining Gene Function and Therapy for Inherited Retinal Disorders

Cited by 2 publications

References 122 publications

Gene editing with ‘pencil’ rather than ‘scissors’ in human pluripotent stem cells

Gene editing with ‘pencil’ rather than ‘scissors’ in human pluripotent stem cells

Clinical exome analysis and targeted gene repair of the c.1354dupT variant in iPSC lines from patients with PROM1-related retinopathies exhibiting diverse phenotypes

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