Gene correction of the CLN3 c.175G>A variant in patient‐derived induced pluripotent stem cells prevents pathological changes in retinal organoids

Molecular Genetics & Genomic Medicine

Jennifer A. Thompson¹  Terri L. McLaren²  Dan Zhang²  Samuel McLenachan²  Zhiqin Huang²  Tina M. Lamey²  John N. De Roach²  Xiao Zhang²  Fred K. Chen²  Shang‐Chih Chen³  Luke Jennings³ 

[1] Australian Inherited Retinal Disease Registry and DNA Bank Department of Medical Technology and Physics Sir Charles Gairdner Hospital Perth WA Australia;Centre for Ophthalmology and Visual Science The University of Western Australia Perth WA Australia;Ocular Tissue Engineering Laboratory Lions Eye Institute Perth WA Australia;

关键词: Batten disease;    CLN3;    induced pluripotent stem cells;    retinal organoid;    retinopathy;   

DOI  :  10.1002/mgg3.1601

来源: DOAJ

【 摘 要 】

Abstract Background Mutations in CLN3 cause Batten disease, however non‐syndromic CLN3 disease, characterized by retinal‐specific degeneration, has been also described. Here, we characterized an induced pluripotent stem cell (iPSC)‐derived disease model derived from a patient with non‐syndromic CLN3‐associated retinopathy. Methods Patient‐iPSC, carrying the 1 kb‐deletion and c.175G>A variants in CLN3, coisogenic iPSC, in which the c.175G>A variant was corrected, and control iPSC were differentiated into neural retinal organoids (NRO) and cardiomyocytes. CLN3 transcripts were analyzed by Sanger sequencing. Gene expression was characterized by qPCR and western blotting. NRO were characterized by immunostaining and electron microscopy. Results Novel CLN3 transcripts were detected in adult human retina and control‐NRO. The major transcript detected in patient‐NRO displayed skipping of exons 2 and 4–9. Accumulation of subunit‐C of mitochondrial ATPase (SCMAS) protein was demonstrated in patient‐derived cells. Photoreceptor progenitor cells in patient‐NRO displayed accumulation of peroxisomes and vacuolization of inner segments. Correction of the c.175G>A variant restored CLN3 mRNA and protein expression and prevented SCMAS and inner segment vacuolization. Conclusion Our results demonstrate the expression of novel CLN3 transcripts in human retinal tissues. The c.175G>A variant alters splicing of the CLN3 pre‐mRNA, leading to features consistent with CLN3 deficiency, which were prevented by gene correction.

【 授权许可】

Unknown