Abstract
There is a major need for therapeutics that treat disease caused by premature termination codons (PTCs). Splice-switching antisense oligonucleotides (ASOs) can be directed to block splicing and cause exon skipping, a process that can be used to effectively remove PTCs from an mRNA. This ASO-induced exon skipping can restore protein coding potential when the exons on either side of the skipped exon are in the same reading frame, or symmetrical. We demonstrate the potential of this approach as a therapeutic using the cystic fibrosis (CF) transmembrane regulator (CFTR) gene, which has CF-associated, PTC-causing variants in all 27 of its exons. We functionally screened all CFTR isoforms that can be generated by deletion of symmetrical exons and identify four that are functionally responsive to CFTR modulators. We identified ASOs that induce skipping of these exons and show that they recover CFTR function in airway cells derived from individuals with CFTR PTC variants. This study demonstrates that systematic functional analysis of in-frame exon-deleted protein isoforms can successfully identify targets for ASO-based splice-switching therapies, a therapeutic concept that can be broadly applied to any multi-exon protein-coding gene disrupted by PTCs.