Assessment of transduction efficiency and tropism of different adeno-associated virus (AAV) serotypes based on route of delivery in rats

Abstract

Purpose : Adeno-associated virus (AAV)-mediated gene replacement has been shown to restore vision in patients with RPE65-associated Leber congenital amaurosis, paving the way for ocular gene therapies. Our group recently showed that transduction efficiency and tropism differ between various AAV subtypes when tested in organotypic human retinal explants ex vivo. However, little is known about serotype-specific differences in tropic behavior based on the route of delivery in vivo. This study assesses the retinal transduction efficiency and tropism of different AAV serotypes based on different routes of ocular delivery in rats.

Methods : Seven different serotypes (1, 2, 4, 5, 6, 8, 9) of recombinant AAV expressing eGFP under control of the cytomegalovirus promoter were packaged and titer matched. For each AAV serotype, 10 µl of viral vector (1x10¹² vg/ml) were delivered via intravitreal, subretinal, or suprachoroidal injection into 2 month old Sprague Dawley rats (3 eyes per serotype per route of delivery). Two weeks after injection, the eyes were examined in vivo using a rodent-specific fluorescent fundus camera (Micron III, Phoenix Laboratories) to evaluate the intensity of GFP signal and the distribution relative to the injection sites. The animals were then sacrificed, and the eyes enucleated and processed for immunohistochemistry. Tropism of the various AAV serotypes for the different retinal layers was then compared for each route of delivery.

Results : Retinal transduction efficiency and tropism varied greatly depending on the AAV subtype and route of delivery. AAV2 serotype 2 demonstrated low transduction efficiency across all 3 routes of delivery, whereas serotype 6 demonstrated high transduction efficiency, with the largest areas of GFP expression noted via suprachoroidal injection. Several serotypes, including 1, 4, and 5, demonstrated limited or no transduction via intravitreal delivery compared to subretinal or suprachoroidal delivery.

Conclusions : AAV vectors show differences in retinal transduction efficiency and tropism depending on serotype and route of delivery. These differences may be important for the strategic selection of AAV serotype and route of delivery for gene-augmentation therapies in human subjects.

This is an abstract that was submitted for the 2018 ARVO Annual Meeting, held in Honolulu, Hawaii, April 29 - May 3, 2018.