Rescue of Aberrant Splicing Caused by a Novel Complex Deep-intronic ABCA4 Allele

Area of Science:

• Genetics and Ophthalmology
• Molecular Biology and Genetic Medicine

Background:

• Stargardt disease (STGD1) is an inherited retinal disorder causing progressive central vision loss, linked to pathogenic variants in the ABCA4 gene.
• Disease onset and severity in STGD1 can vary significantly, influenced by the specific genetic mutations involved.

Purpose of the Study:

• To investigate a complex deep-intronic allele in the ABCA4 gene identified in a patient with STGD1.
• To characterize the splicing alterations caused by this allele and explore potential therapeutic strategies.

Main Methods:

• Whole exome sequencing (WES), whole gene sequencing, and whole genome sequencing (WGS) were employed for genetic analysis.
• Minigene assays coupled with nanopore sequencing were used to analyze splicing patterns and identify pseudoexons.
• Antisense oligonucleotides (AONs) were designed and tested for their ability to correct aberrant splicing.

Main Results:

• A novel complex deep-intronic ABCA4 allele was identified in a patient with STGD1.
• This allele significantly increased the abundance of transcripts containing pseudoexons from ABCA4 intron 11.
• Two AONs were effective in restoring normal splicing, with one achieving reference levels.

Conclusions:

• Minigene assays and nanopore sequencing are powerful tools for uncovering low-abundance transcripts and splicing abnormalities.
• The identified complex ABCA4 allele contributes to STGD1 pathogenesis through aberrant splicing.
• AONs show promise as a potential therapeutic approach for correcting splicing defects in STGD1.