• PE
• ASO
• poison exons
• antisense oligonucleotides
• genome-wide

Poison exons (PEs) are naturally occurring alternative exons that contain a premature termination codon. More than 20% of isoforms in the human transcriptome contain PEs, which direct the carrier transcripts to nonsense-mediated decay (NMD). Splice-switching antisense oligonucleotides (ASOs) are short, chemically modified RNAs that can block the inclusion of naturally occurring PEs by modulating RNA splicing, thereby up-regulating productive mRNA and protein levels. Recently, such strategy has been deployed in ASO development to treat diseases caused by mutations in haploinsufficient genes (Lim, 2020). However, our knowledge of where PEs reside in the genome is still limited, as it is challenging to detect such NMD-sensitive transcripts with low abundance in total RNA samples. Here, we identify thousands of novel PEs and generate a genome-wide atlas of gene targets for therapeutic ASO development for Mendelian diseases. To capture previously undetectable PE-containing transcripts, we use a variety of strategies to inhibit NMD in patient-derived fibroblasts, neurons, astrocytes, and retinal pigmented epithelium cells, followed by deep stranded RNA sequencing. We propose a gene annotation-agnostic pipeline for genome-wide, tissue-specific identification of PEs. So far, our pipeline has discovered >4,000 novel PEs and confirmed >4000 known PEs, while missing <100 high-confidence PEs from previous studies. We experimentally validate 30 PEs using RT-PCR, 22 out of which in haploinsufficient genes or oncogenes. We are developing splice-switching ASOs for PEs within genes relevant to Central Nervous System monogenic diseases. In the pilot exploration for therapeutic targets in MAPK8IP3-Related Neurodevelopmental Disorder, we identify 8 ASOs targeting a PE that can up-regulate MAPK8IP3 expression in fibroblasts. Our study delineates a rich set of interventional targets for therapeutic ASO development for several Mendelian disorders.