Abstract
Purpose :
Stoke Therapeutics is developing medicines to treat diseases by leveraging antisense oligonucleotides (ASOs) to increase gene expression. Stoke’s approach, TANGO (Targeted Augmentation of Nuclear Gene Output) exploits naturally-occurring non-productive splicing events identified using a proprietary bioinformatics pipeline to increase target protein levels via splicing modulation.Diseases of the eye are well suited to TANGO due to ASO capacity to reach target cells in the back of the eye. We are exploring a non-productive splicing event in CD274 to increase expression of PD-L1 to treat autoimmune diseases, such as uveitis. In addition to TANGO, ASOs can also be used to switch gene isoforms. For example, we can switch from the membrane-bound isoform of vascular endothelial growth factor (mbVEGF), which promotes pathologic neovascularization in age related macular degeneration, to the soluble isoform (sVEGF), which inhibits neovascularization. Finally, we evaluated the durability of uniformly modified 2’-O-methoxyethyl (MOE) ASO activity in the mouse retina using an ASO targeting Cep290.
Methods :
RNA-seq data were evaluated with proprietary transcriptome-independent approaches to identify annotated and novel non-productive splicing events. Cells were treated with various ASOs, and RNA/protein was extracted to verify RNA splicing effects and changes in protein expression. ASO targeting Cep290 was intravitreally injected in mice, and retinas were collected to evaluate ASO impact on splicing over time.
Results :
Stoke has identified >100,000 non-productive splicing events amenable to our TANGO technology in >50% of genes. We developed ASOs targeting CD274 (PD-L1) that increase its mRNA and protein levels. We also demonstrate the ability to switch VEGF isoforms, functionally decreasing VEGF signaling. Finally, using a MOE-ASO targeting Cep290, we show that ASOs have activity in the mouse retina for >9 months following a single intravitreal injection.
Conclusions :
Stoke’s technology offers an ASO-mediated strategy to increase target gene expression to treat a wide variety of diseases caused by haplo-insufficiency of a critical gene or by insufficient expression of specific pathway genes. We have demonstrated feasibility of this approach for treating ocular diseases. As over 50% of genes possess non-productive splicing events, the potential target opportunity is significant and largely untapped.
This abstract was presented at the 2019 ARVO Annual Meeting, held in Vancouver, Canada, April 28 - May 2, 2019.