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GS Yang, J Lindbloom, Z Yan, J Engelhardt, M Schmidt, R Kotin, B Davidson; Viral mediated transduction of murine retina with AAV2 and AAV5 . Invest. Ophthalmol. Vis. Sci. 2002;43(13):4619.
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© ARVO (1962-2015); The Authors (2016-present)
Purpose: Inherited retinal degeneration such as retinitis pigmentosa (RP) causes blindness. Approaches to treat RP may include gene replacement to photoreceptor cells, a method which requires identification of an efficient vector with appropriate cell specificity and minimal toxicity. Methods: In this study we quantified the cellular specificity and efficiency of rAAV5 vs. rAAV2 in mice after subretinal delivery. Viruses, all containing expression cassettes with reporters driven by the CMV promoter, were injected for either intra or inter-eye comparisons and gene transfer was evaluated using unbiased stereology (Bioquant software). Results: Our data showed that rAAV2 and AAV5 can transduce RPE and photoreceptor cells. AAV5.nlsGFP.Long vs. AAV2.dsRed.Long (107 particles per eye) showed mean transgene expression in photoreceptor cells for AAV5 and AAV2 to be 63,000 and 0.0 at 5 weeks (p<0.0001), 200,000 and 63,000 at 15 weeks (p=0.016), and 79,000 and 6,300 at 31 weeks. AAV2/5.GFP.Long (AA2 genome in AAV5 capsid) vs. AAV2.GFP.Long (109 particles per eye) showed mean transgene expression in photoreceptor cells for AAV2/5 and AAV2 to be 140,000 and 15.0 at 5 weeks (p=0.0002) and 400,000 and 1,000 at 15 weeks (p=0.02). AAV5.GFP.Short vs. AAV2.GFP.Short (109 particles per eye) showed mean transgene expression in photoreceptor cells for AAV5 and AAV2 to be 250,000 and 50,000 at 5 weeks (p=0.04), and 320,000 and 580,000 at 15 weeks (p=0.024). AAV2.GFP.Short transduced more photoreceptor cells than AAV2.GFP.Long at both 5 (p=0.007) and 15 weeks (p<0.0001). We harvested Hirt DNA of retina 14 weeks after subretinal delivery and showed that AAV2.GFP.Long vs. AAV2/5.GFP.Long had 16,000 and 450,000 genome copies (p=0.0003) and AAV2.GFP.Short vs. AAV2.GFP.Long had 790 and 16,000 genome copies (p<0.0001). Conclusion: Our results showed that the kinetics of AAV5 expression was faster and that, with full length AAV genomes, more photoreceptor cells were transgene positive than AAV2.
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