Antisense Oligonucleotide-Based Rescue of Aberrant Splicing Defects Caused by 15 Pathogenic Variants in ABCA4

Abstract

The discovery of novel intronic variants in the ABCA4 locus has contributed significantly to solving the missing heritability in Stargardt disease (STGD1). The increasing number of variants affecting pre-mRNA splicing makes ABCA4 a suitable candidate for antisense oligonucleotide (AON)-based splicing modulation therapies. In this study, AON-based splicing modulation was assessed for 15 recently described intronic variants (three near-exon and 12 deep-intronic variants). In total, 26 AONs were designed and tested in vitro using a midigene-based splice system. Overall, partial or complete splicing correction was observed for two variants causing exon elongation and all variants causing pseudoexon inclusion. Together, our results confirm the high potential of AONs for the development of future RNA therapies to correct splicing defects causing STGD1.

RNA-based therapeutics for neurological diseases

“…There is extensive pre-clinical and clinical research on the use of ssAONs to correct splicing defects in inherited retinal dystrophies [ 35–37 ]. For example, for Stargardt disease, ssAONs to correct intronic mutations which cause exon elongation or pseudoexons in the ATP-binding cassette transporter type 4 subfamily A, ABCA4 , transcript show promise when tested in various cell models including induced pluripotent stem cell (iPSCs)-derived photoreceptor precursor cells [ 38 , 39 ]. The ssAON, sepofarsen, is under clinical development as a treatment for Leber congenital amaurosis [ 40 ] and an exon skipping ssAON is being investigated for the treatment of Usher syndrome type 2 [ 36 , 41 ].…”

Antisense oligonucleotides targeting exon 11 are able to partially rescue the Neurofibromatosis Type 2 phenotype in vitro

“…It has only been possible through CRISPR/Cas technology by the induction of single nucleotide substitutions [46,47]. Allele-specific approaches have been successfully tested for intronic variants through splice modulation ASOs that mask the erroneous signalling and given their intronic location and distance to splicing regulatory regions, proper protein synthesis can be recovered [48][49][50][51][52].…”

Therapy Approaches for Stargardt Disease

“…Patients with the deep intronic mutation c.2991 + 1655A > G in CEP290, which causes the inherited retinal dystrophy Leber congenital amaurosis, have so far received an intravitreal injection of sepofarsen (ProQR Therapeutics), a 17-mer 2 -O-methylmodified phosphonothioate RNA AON, followed by three monthly top-up injections. Importantly, no serious adverse events have been reported with encouraging signs of improvements to visual acuity at 3 months post treatment [91]. More recently, it was revealed that one patient achieved a sustained response to just a single dose of sepofarsen lasting up to 15 months after treatment [92].…”