Photoreceptor-targeted gene delivery using intravitreally administered AAV vectors in dogs

Abstract

Delivery of therapeutic transgenes to retinal photoreceptors using adeno-associated virus (AAV) vectors has traditionally required subretinal injection. Recently, photoreceptor transduction efficiency following intravitreal injection (IVT) has improved in rodent models through use of capsid-mutant AAV vectors; but remains limited in large animal models. Thickness of the inner limiting membrane (ILM) in large animals is thought to impair retinal penetration by AAV. Our study compared two newly developed AAV vectors containing multiple capsid amino acid substitutions following IVT in dogs. The ability of two promoter constructs to restrict reporter transgene expression to photoreceptors was also evaluated. AAV vectors containing the interphotoreceptor-binding protein (IRBP) promoter drove expression exclusively in rod and cone photoreceptors, with transduction efficiencies of ~ 4% of cones and 2% of rods. Notably, in the central region containing the cone-rich visual streak, 15.6% of cones were transduced. Significant regional variation existed, with lower transduction efficiencies in the temporal regions of all eyes. This variation did not correlate with ILM thickness. Vectors carrying a cone-specific promoter failed to transduce a quantifiable percentage of cone photoreceptors. The newly developed AAV vectors containing the IRBP promoter were capable of producing photoreceptor-specific transgene expression following IVT in the dog.

Noninvasive gene delivery to foveal cones for vision restoration

“…We selected 3 previously described promoters in view of their utility in driving gene expression in cones (4,18,19,42,43) and tested them for specificity and strength of cone transduction side by side. All promoters tested in vivo in mouse retinas led to transgene expression in the photoreceptor layer when delivered subretinally.…”

Impact of Heparan Sulfate Binding on Transduction of Retina by Recombinant Adeno-Associated Virus Vectors

“…In previous studies, it was shown that rAAV2(quadY-F) and rAAV2(quadY-FϩT-V) vectors promoted significant transduction of photoreceptors from Ivt injection in both small-and large-animal models (30,38,52). Of the AAV2-based variants to be tested, rAAV2(tripleY-FϩT-V) has the strongest liver transduction following intravascular delivery and exhibits maximal nuclear translocation (35).…”

Insight into the mechanisms of enhanced retinal transduction by the engineered AAV2 capsid variant ‐7m8

“…All rights reserved 12 of efficient retinal transduction from the vitreous have been designed only recently, using rational design (Boyd et al, 2015;Boye et al, 2016;Kay et al, 2013;Petrs-silva et al, 2011) or in vivo directed evolution (Cronin et al, 2014;Dalkara et al, 2013). In the latter study, we used directed evolution of random peptide libraries displayed on AAV2 capsid for selection of capsid variants that overcome the natural AAV transduction barriers of retinal tissue from the vitreous.…”