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Chen Matsevich, Deniz Dalkara, Alexey Obolensky, Melissa Desrosiers, Ayala Ejzenberg, Dror Sharon, Eyal Banin, Avigail Beryozkin; Subretinal and Intravitreal Delivery of the Photoreceptor-Specific AAV2-7m8-hGRK1-GFP Viral Vector in Mice. Invest. Ophthalmol. Vis. Sci. 2019;60(9):2924. doi: https://doi.org/.
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© ARVO (1962-2015); The Authors (2016-present)
FAM161A mutations are the most common cause of autosomal recessive retinitis pigmentosa (RP) in Israel. In preparation for testing gene augmentation therapy in a FAM161A mouse model of RP and later-on in humans, our purpose was to evaluate the transduction efficacy and safety of the viral vector AAV2-7m8-hGRK1-GFP following subretinal (SR) versus intravitreal (IV) injection in mice.
2µl of an AAV2-7m8 viral vector containing 2.8x10E10 viral particles carrying the reporter GFP gene under control of the photoreceptor-specific promoter hGRK1 were delivered subretinally or intravitreally into the eyes of naïve mice (n=19) that are the background for the FAM161A model. Each animal was treated unilaterally with the vector while the fellow eye received a similar injection of BSS and served as control. At 1, 2, 3, 4.5, 6 and 7 months following injection, GFP expression was monitored in-vivo using the Micron III system equipped with the proper filters. Retinal structure was studied by histological analysis (H&E staining) following enucleation at 7 months post-injection. Efficacy and specificity of transduction in photoreceptors was confirmed by anti-GFP immunohistochemistry (IHC).
Following both SR and IV injections, in-vivo imaging showed increase of the GFP signal over the initial 2-3 months, which then stabilized. The pattern of GFP expression differed between the two administration routes: Following SR delivery (n=10), GFP expression was relatively strong but limited sharply to the area that corresponds to the SR bleb. Following IV delivery (n=9), GFP expression was more disperse and mainly detected near the optic nerve, along the major retinal blood vessels and along the far retinal periphery. H&E showed that retinal structure was well-preserved in all eyes, confirming safety of both routes of injection. IHC showed a high level of GFP expression in the outer nuclear (photoreceptors) layer.
The vector may be safely delivered into the SR space or vitreous providing specific transfection of the photoreceptors in naïve mice that serve as the background for the FAM161A model. We are planning to use a similar vector carrying the normal FAM161A gene for gene therapy of FAM161A knockout mice in an attempt to slow/halt progression of disease. Results of these experiments may serve as an important step towards future application of gene therapy in FAM161A-RP patients.
This abstract was presented at the 2019 ARVO Annual Meeting, held in Vancouver, Canada, April 28 - May 2, 2019.
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