Antisense oligonucleotide therapy corrects splicing in the common Stargardt disease type 1-causing variant ABCA4 c.5461-10T>C

Melita Kaltak; Petra de Bruijn; Davide Piccolo; Sang-Eun Lee; Kalyan Dulla; Thomas Hoogenboezem; Wouter Beumer; Andrew R Webster; Rob W J Collin; Michael E Cheetham; Gerard Platenburg; Jim Swildens

doi:10.1016/j.omtn.2023.02.020

Antisense oligonucleotide therapy corrects splicing in the common Stargardt disease type 1-causing variant ABCA4 c.5461-10T>C

Mol Ther Nucleic Acids. 2023 Feb 18:31:674-688. doi: 10.1016/j.omtn.2023.02.020. eCollection 2023 Mar 14.

Authors

Melita Kaltak^{1

2

3}, Petra de Bruijn¹, Davide Piccolo⁴, Sang-Eun Lee⁴, Kalyan Dulla¹, Thomas Hoogenboezem¹, Wouter Beumer¹, Andrew R Webster^{4

5}, Rob W J Collin^{2

3}, Michael E Cheetham⁴, Gerard Platenburg¹, Jim Swildens¹

Affiliations

¹ ProQR Therapeutics, Zernikedreef 9, 2333 CK Leiden, the Netherlands.
² Department of Human Genetics, Radboud University Medical Center, Geert Grooteplein-Zuid 10, 6525 GA Nijmegen, the Netherlands.
³ Academic Alliance Genetics, Radboud University Medical Center, Geert Grooteplein-Zuid 10, 6525 GA Nijmegen, and Maastricht University Medical Center+, P. Debyelaan 25, 6229 HX Maastricht, the Netherlands.
⁴ UCL, Institute of Ophthalmology, 11-43 Bath Street, EC1V 9EL London, UK.
⁵ Moorfields Eye Hospital, 162 City Road, EC1V 2PD London, UK.

Abstract

Stargardt disease type 1 (STGD1) is the most common hereditary form of maculopathy and remains untreatable. STGD1 is caused by biallelic variants in the ABCA4 gene, which encodes the ATP-binding cassette (type 4) protein (ABCA4) that clears toxic byproducts of the visual cycle. The c.5461-10T>C p.[Thr1821Aspfs∗6,Thr1821Valfs∗13] variant is the most common severe disease-associated variant, and leads to exon skipping and out-of-frame ABCA4 transcripts that prevent translation of functional ABCA4 protein. Homozygous individuals typically display early onset STGD1 and are legally blind by early adulthood. Here, we applied antisense oligonucleotides (AONs) to promote exon inclusion and restore wild-type RNA splicing of ABCA4 c.5461-10T>C. The effect of AONs was first investigated in vitro using an ABCA4 midigene model. Subsequently, the best performing AONs were administered to homozygous c.5461-10T>C 3D human retinal organoids. Isoform-specific digital polymerase chain reaction revealed a significant increase in correctly spliced transcripts after treatment with the lead AON, QR-1011, up to 53% correct transcripts at a 3 μM dose. Furthermore, western blot and immunohistochemistry analyses identified restoration of ABCA4 protein after treatment. Collectively, we identified QR-1011 as a potent splice-correcting AON and a possible therapeutic intervention for patients harboring the severe ABCA4 c.5461-10T>C variant.

Keywords: ABCA4; MT: Oligonucleotides: Therapies and Applications; RNA therapy; Stargardt disease; antisense oligonucleotides; retinal organoids; splicing correction.

Grants and funding

205041/Z/16/Z/WT_/Wellcome Trust/United Kingdom