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Katharine J Liang, Kenton T Woodard, R. Jude Samulski; AAV-mediated activation of the Nrf2 pathway in the Nrf2-deficient mouse. Invest. Ophthalmol. Vis. Sci. 2014;55(13):1738.
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© ARVO (1962-2015); The Authors (2016-present)
Retinal degeneration is due at least in part to redox deregulation. Under conditions of oxidative stress, the Nrf2 transcription factor activates the antioxidant response element, driving expression of endogenous antioxidant genes. The Nrf2-deficient mouse exhibits an increased susceptibility to exogenous oxidative stress, but limited overt pathology. Because the Nrf2-/- mouse exhibits an age-related retinal degeneration of incomplete penetrance, we applied novel AAV vectors to dampen oxidative stress in the light-stressed Nrf2-deficient mouse.
In order to increase uniformity of disease progression and mimic environmental factors contributing to retinal degeneration, we exposed Nrf2-deficient mice to a light-induced retinal degeneration protocol. An AAV2.5-CBh-Nrf2 vector was engineered using a chimeric capsid vector and promoter selected for retinal transduction via intravitreal injection and delivered prior to light damage exposure. Functional efficacy of AAV-Nrf2 was verified using in vitro luciferase reporter and fluorescent ROS detection assays. Downstream gene activation was assessed by western blot. Therapeutic efficacy was assessed using ERG, fundus imaging, and OCT.
AAV2.5 combines the improved transduction properties of AAV1 with reduced antigenic cross-reactivity against antibodies directed at both parental serotypes while keeping the receptor binding properties of AAV2. AAV-Nrf2 successfully mediates Nrf2 expression, activates downstream genes, and suppresses reactive oxygen species in vitro. In vivo, intravitreally delivered AAV-Nrf2 was sufficient to activate downstream gene expression up for up to six months. Light-induced degeneration in Nrf2-/- mice and AAV-mediated rescue is being characterized by ERG, fundus imaging, and OCT.
AAV vector development combined with gene replacement offers a unique opportunity to optimize and test both efficient transgene delivery as well as potent therapeutic potential. Our research provides a targeted molecular approach to drive antioxidant gene expression, and offers a promising alternative to currently limited therapeutics for multifactorial diseases involving ROS deregulation.
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