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Agostina Puppo, Giulia Cesi, Elena Marrocco, Alberto Auricchio; Retinal transduction profiles by high capacity viral vectors. Invest. Ophthalmol. Vis. Sci. 2014;55(13):3326.
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© ARVO (1962-2015); The Authors (2016-present)
Retinal gene therapy with adeno-associated viral (AAV) vectors is safe and effective in humans. However, the limited cargo capacity of AAV prevents their use for therapy of those inherited retinopathies (IRs) due to mutations in large (>5 kb) genes. Viral vectors derived from Adenovirus (Ad), Lentivirus (LV) and Herpesvirus (HV) can package large transgenic DNA sequences but do not target efficiently retinal photoreceptor (PRs) where the majority of genes responsible for IRs are expressed. Here we aimed at the identification of Ad, LV and HV vectors with high PRs tropism and transduction efficiency.
We have evaluated the murine PR transduction efficacy of vectors derived from 16 different Ad serotypes, 7 LV pseudotypes, and from one bovine HV. All vectors expressed either EGFP or lacZ under the control of the ubiquitous CMV promoter. After subretinal delivery, fluorescent microscopy and X-gal staining of retinal cryosections was used to compare expression efficiency and cell type specificity. Vectors that showed potential for enhanced PR transduction were additionally tested using the PR specific Rho promoter.
The retinal pigment epithelium (RPE) was transduced with almost all the vectors tested. LV-GP64 outperformed the most studied LV-VSVG in transducing PRs although transduction was confined to a restricted area. More extensive PR transduction than with LV was found with Ad1-, Ad2- and Ad5/F35++ -Rho-EGFP, although none significantly outperformed the canonical Ad5.
The high-capacity vectors tested have overall similar PR transduction ability than the canonical Ad5 or LV-VSVG and none matched the extent of PR transduction of AAV2/8, a highly effective AAV serotype for PR targeting. The larger size of the high-capacity particles we have investigated compared to AAV2/8 might cause a limited diffusion in the subretinal space where hindrance may be exerted by anatomical barriers as the inter-photoreceptor matrix or the outer limiting membrane. The research leading to these results has received funding from the European Research Council under the European Union's Seventh Framework Programme (FP/2007-2013) / ERC Grant Agreement n.  and from Fondazione Telethon Grant n. TGM11MT1.
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