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Open AccessArticle

Antisense Oligonucleotide-Based Downregulation of the G56R Pathogenic Variant Causing NR2E3-Associated Autosomal Dominant Retinitis Pigmentosa

1
Center for Medical Genetics Ghent, Ghent University and Ghent University Hospital, Corneel Heymanslaan 10, 9000 Ghent, Belgium
2
Cancer Research Institute Ghent (CRIG), Ghent University, 9000 Ghent, Belgium
3
Bioinformatics Institute Ghent (BIG), Ghent University, 9000 Ghent, Belgium
*
Author to whom correspondence should be addressed.
These authors contributed equally to this work.
Genes 2019, 10(5), 363; https://doi.org/10.3390/genes10050363
Received: 1 April 2019 / Revised: 28 April 2019 / Accepted: 6 May 2019 / Published: 10 May 2019
(This article belongs to the Special Issue Molecular Therapies for Inherited Retinal Diseases)
The recurrent missense variant in Nuclear Receptor Subfamily 2 Group E Member 3 (NR2E3), c.166G>A, p.(Gly56Arg) or G56R, underlies 1%–2% of cases with autosomal dominant retinitis pigmentosa (adRP), a frequent, genetically heterogeneous inherited retinal disease (IRD). The mutant NR2E3 protein has a presumed dominant negative effect (DNE) by competition for dimer formation with Cone-Rod Homeobox (CRX) but with abolishment of DNA binding, acting as a repressor in trans. Both the frequency and DNE of G56R make it an interesting target for allele-specific knock-down of the mutant allele using antisense oligonucleotides (AONs), an emerging therapeutic strategy for IRD. Here, we designed gapmer AONs with or without a locked nucleic acid modification at the site of the mutation, which were analyzed for potential off-target effects. Next, we overexpressed wild type (WT) or mutant NR2E3 in RPE-1 cells, followed by AON treatment. Transcript and protein levels of WT and mutant NR2E3 were detected by reverse transcription quantitative polymerase chain reaction (RT-qPCR) and Western blot respectively. All AONs showed a general knock-down of mutant and WT NR2E3 on RNA and protein level, showing the accessibility of the region for AON-induced knockdown. Further modifications are needed however to increase allele-specificity. In conclusion, we propose the first proof-of-concept for AON-mediated silencing of a single nucleotide variation with a dominant negative effect as a therapeutic approach for NR2E3-associated adRP. View Full-Text
Keywords: retinitis pigmentosa; autosomal dominant; NR2E3; G56R; putative dominant negative effect; gapmer antisense oligonucleotides; allele-specific knockdown retinitis pigmentosa; autosomal dominant; NR2E3; G56R; putative dominant negative effect; gapmer antisense oligonucleotides; allele-specific knockdown
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MDPI and ACS Style

Naessens, S.; Ruysschaert, L.; Lefever, S.; Coppieters, F.; De Baere, E. Antisense Oligonucleotide-Based Downregulation of the G56R Pathogenic Variant Causing NR2E3-Associated Autosomal Dominant Retinitis Pigmentosa. Genes 2019, 10, 363.

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