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

Abstract

Recently, we described a modified AAV2 vector-AAV2-7m8-having a capsid-displayed peptide insertion of 10 amino acids with enhanced retinal transduction properties. The insertion of the peptide referred to as 7m8 is responsible for high-level gene delivery into deep layers of the retina when virus is delivered into the eye's vitreous. Here, we further characterize AAV2-7m8 mediated gene delivery to neural tissue and investigate the mechanisms by which the inserted peptide provides better transduction away from the injection site. First, in order to understand if the peptide exerts its effect on its own or in conjunction with the neighboring amino acids, we inserted the 7m8 peptide at equivalent positions on three other AAV capsids, AAV5, AAV8, and AAV9, and evaluated its effect on their infectivity. Intravitreal delivery of these peptide insertion vectors revealed that only AAV9 benefited from 7m8 insertion in the context of the retina. We then investigated AAV2-7m8 and AAV9-7m8 properties in the brain, to better evaluate the spread and efficacy of viral transduction in view of the peptide insertion. While 7m8 insertion led to higher intensity gene expression, the spread of gene expression remained unchanged compared to the parental serotypes. Our results indicate that the 7m8 peptide insertion acts by increasing efficacy of cellular entry, with little effect on the spread of viral particles in neural tissue. The effects of peptide insertion are capsid and tissue dependent, highlighting the importance of the microenvironment in gene delivery using AAV. Biotechnol. Bioeng. 2016;113: 2712-2724. © 2016 Wiley Periodicals, Inc.

Noninvasive gene delivery to foveal cones for vision restoration

“…In order to find vector-promoter combinations suitable for strong and specific cone targeting away from the injection site, we compared several AAVs after intravitreal and subretinal delivery in mouse retinas. To enable efficient cone photoreceptor targeting, we used an engineered AAV variant called AAV2-7m8, which has been shown to target photoreceptors efficiently via both administration routes (16,17). Specific targeting of cone cells has never been attempted using vitreally administered AAV.…”

“…Based on this, we reasoned that an enhanced AAV9 variant might afford efficient transduction of foveal cones from a distal bleb. We previously described a variant called AAV9-7m8, which provides a 30-fold increase in infectivity over AAV9 (17). To promote foveal cone gene delivery through a distal subretinal injection site, we used an AAV9-7m8 variant.…”

“…Intravitreal injections are surgically simple ways to deliver therapeutics without retinal detachment. Gene therapy vectors can target the outer retina via intravitreal injections in rodents without damage to the photoreceptors (16,17). However, safe and efficient gene delivery to primate cones via intravitreal injection had not been achieved so far, likely due to the substantial dilution of the vector in the vitreous and resulting loss of efficacy.…”

“…16) or rational design (AAV9-7m8; ref. 17), can achieve therapeutic objectives where parental serotypes fail to provide sufficient gene delivery. Indeed, AAV2 and AAV9 cannot perform efficient noninvasive foveal targeting (30, 31) while 7m8 modified vectors bridge this gap.…”

Heparan Sulfate Binding Promotes Accumulation of Intravitreally Delivered Adeno-associated Viral Vectors at the Retina for Enhanced Transduction but Weakly Influences Tropism

“…We predict that rAAV5 capsid should have enhanced transduction with the HSPG-binding mutation. In addition, other laboratories have tested rAAV5 and rAAV8 using a CAG promoter and have shown expression (47). This serves as a reason to reevaluate all rAAV capsid transduction following intravitreal delivery by FISH.…”