A scalable platform for exon-skipping antisense oligonucleotide therapy development for inborn genetic diseases

Abstract

Antisense oligonucleotides (ASOs) are a versatile therapeutic modality for inborn genetic diseases. ASOs can induce skipping of "dispensable" exons containing disease-causing variants to rescue protein amount and function, but this approach has been studied for only a small number of genes. We developed a high-throughput in silico tool for assessing exon dispensability and designing exon-skipping ASO sequences. Parameters were optimized using known dispensable and in-frame indispensable exons. Across 72,644 exons of 5,057 disease genes, we identified thousands of new targets for exon-skipping ASOs (3.4% of exons with most stringent filters, 24.5% with less stringent filters) that collectively include 0.97%-15.6% of disease-causing variants in large-scale databases. To facilitate recognition of DNA variants potentially amenable to exon skipping as a therapeutic strategy, we established the HAWK-EYE database as a repository of exon dispensability predictions and corresponding in silico-optimized ASO sequences, available as an open-access web application (https://hawk-eye.research.sickkids.ca/). To illustrate translational utility, we experimentally validated a subset of the in silico-optimized ASO sequences that were generated for all Dispensable exons in the HAWK-EYE database, and showed that skipping a Dispensable exon in SOX5 preserves protein function using in vivo and in vitro assays. This scalable platform approach to exon-skipping ASOs will accelerate identification and testing of amenable genetic variants.