Abstract
Purpose :
As improvements are made in engineered and recombinant AAV serotypes that are potentially more potent than naturally occurring serotypes, there is an urgent need to explore an effective and direct method for testing the transduction efficiency and tropism of these AAV serotypes in human retinal tissue. Human retinal explants serve as a robust, informative model to evaluate transduction efficiency and tropism in human cells, while the efficacy of non-human primate (NHP) retinal explants needs to be directly compared to human tissue to determine its use as a predictive model. Herein, this study examines the transduction efficiency and tropism of ten different AAV serotypes in the human retinal and non-human primate (NHP) retinal explants ex vivo.
Methods :
AAV-3b, AAV-6.2, AAV-6TM, AAV-9rh, AAV-32.33, AAV-PHP.B, AAV-PHP.eB, AAV-rh10, AAV-rh39, AAV-rh43 were tested. AAV2/2 and AAV2/8 served as positive controls. All vectors driving expression of GFP under control of the CB6 promoter packaged by the University of Massachusetts Chan Medical School Vector Core. Three independent experiments were performed, each using a different human donor and NHP eye. Retinal explant cultures were maintained for 7 days post transduction. Immunohistochemistry was performed for GFP and the total number of GFP+ cells was calculated using ImageJ manually. Retinal cell tropism for each AAV serotype was evaluated qualitatively by recording whether cells of the inner nuclear layer (INL) or outer nuclear layer (ONL) that were GFP+ overlapped with markers of different retinal cell types, 7G6, PNA, GS, rhodopsin, and arvalbumin.
Results :
AAV-32.33 and AAV-rh10 have the high transducing efficiency for photoreceptor cells and Muller cells, compared with AAV2/2 and AAV2/8 in NHP and human retinal explants. Furthermore, the retinal cell tropism in NHP retinal explants for all serotypes was consistent with retinal cell tropism in human retinal explants.
Conclusions :
Our study shows that AAV-32.33 and AAV-rh10 could be useful candidates for intraocular gene delivery to photoreceptor and Muller cells. These results show that human retina explants serve as an effective and direct method for predicting transduction efficiency and tropism of new AAV serotypes in human retina.
This abstract was presented at the 2022 ARVO Annual Meeting, held in Denver, CO, May 1-4, 2022, and virtually.