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Christoph Jüschke, Sebastian Swirski, Thomas Klopstock klopstock, Bernd Wissinger, John Per Neidhardt; A novel treatment of OPA1 splice defects by U1 snRNA adaptions in a DOA mouse model and patient-derived cells. Invest. Ophthalmol. Vis. Sci. 2020;61(7):1916.
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© ARVO (1962-2015); The Authors (2016-present)
Dominant optic atrophy (DOA) is amongst the most common forms of inherited optic neuropathies with a prevalence of about 1:30,000 worldwide. DOA is mainly caused by mutations in the optic atrophy 1 (OPA1) gene with haploinsufficiency being the main mechanism of the disease. Almost 30% of the cases are due to splice defects. Our goal was to correct OPA1 splice defects using engineered splice factors (U1 snRNAs) in both, a mouse model of DOA and in primary patient-derived fibroblasts.
Both, the studied patients and the mouse model carried the homologous point mutation c.1065+5G>A in the human OPA1 or murine Opa1 gene. The consequence of the mutation was highly similar in human and mice and caused skipping of exon 10. Patient-derived and control fibroblast cell lines were cultured and transduced with lentiviral particles expressing engineered splice factors targeting the spliceosome towards different locations within exon 10 and intron 10 of OPA1. For the mouse model, the engineered splice factor U1 snRNA was applied via sub-retinal injection of AAVs. Reduction of mis-splicing was quantified by semi-quantitative RT-PCR.
We developed a novel treatment of OPA1 splice defects using sequence-adapted splice factors. We tested a series of U1 snRNAs binding to different locations in OPA 1 exon 10 and intron 10 in patient-derived fibroblast cell lines. With the best U1-constructs we were able to achieve a reduction of the level of exon skipped transcripts to about 25% of total OPA1 transcripts. Sub-retinal injections of AAV carrying fully mutation-adapted U1-snRNAs yielded in a robust increase of correctly spliced OPA1 transcripts in the eyes of OPA1+/- mutant mice compared to mock- or non-injected littermate controls.
As a proof-of-concept study, our data indicate for the first time the feasibility of splice site correction as a treatment option for DOA. Increasing the amount of correctly spliced OPA1 transcript might suffice to overcome the haploinsufficiency. Investigations will be performed to address the question of potential side effects. Our findings further support that engineered splice factors offer a novel treatment option for blinding disease.
This is a 2020 ARVO Annual Meeting abstract.
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