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

Abstract

Adeno-associated viruses (AAVsA deno-associated viruses (AAVs) are nonpathogenic, singlestranded packaging DNA dependoparvoviruses within the family Parvoviridae. As the genus name implies, wild-type (WT) AAV is dependent on helper viruses, such as those in the Adenoviridae and Herpesviridae families, for replication. For its safety and the availability of a wide variety of naturally occurring serotypes displaying various tissue tropisms, recombinant AAV (rAAV) has found wide utility as both a gene therapy vector and biotechnological tool (1). Capsid crystal structures also have been determined for many of the AAV serotypes (2-9), leading to a revolution in identifying the structural determinants of receptor attachment, tropism, and transduction efficiency and, by extension, the ability to modify the capsid to generate variants with desired transduction profiles.One of the major clinical applications of AAV is as a gene therapy vector for the treatment of blindness. The eye is a particularly well-suited organ for gene therapy due to its small size, compartmentalization, and immune-privileged status (10). Clinical trials for RPE65-Leber congenital amaurosis (LCA2) demonstrate the ability to deliver a therapeutic transgene to the retinal pigment epithelium (RPE), thereby restoring retinal function and visually evoked behavior to patients (11-13). However, the majority of inherited retinal diseases are caused by defects in photoreceptors, highlighting the need to identify serotypes capable of transducing this cell type. We and others have shown that various subretinally delivered AAV serotypes are capable of efficient transduction of photoreceptors in nonhuman primates (NHP) (14-19). However, subretinal injection of AAV2 under the fovea of some LCA2 patients led to central retinal thinning and loss of visual acuity (20). Similar decreases in retinal thickness were ob-

Noninvasive gene delivery to foveal cones for vision restoration

“…Using the same optogenetic cone reactivation strategy, we showed that this approach also affords robust light responses mediated by Jaws but in a higher percentage of cones compared with a intravitreal route. AAV9-7m8's behavior is similar to a previously described AAV2-derived mutant that exhibits enhanced lateral spread after subretinal injections in mouse retina (44). However, in the macaque retina, we believe that the transduction beyond the bleb with AAV9-7m8 is correlated with its increased infectivity compared with its parental serotype (17) rather than with altered binding to its primary receptor.…”

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

“…Based on our data, we suggest that any capsid that has an HS-binding motif would be capable of accumulating at the ILM. The recent study published by Boye et al used HSPGbinding rAAV5 and rAAV8 but was unsuccessful in enhancing their GFP fluorescence, indicating this HS-binding enhancement is not universal to all capsids (46). We believe this to be related to their shortened CBA promoter and that use of a different promoter may reveal expression.…”

“…Other attempts to enhance transduction used capsids modified to avoid proteasomal degradation (50), thereby enhancing intracellular trafficking to help increase the transgene expression of intravitreally delivered capsids. However, a recent publication suggests that these vectors may not benefit from proteasome avoidance (46). These capsid changes work downstream of ILM binding once the vector has penetrated the retina.…”

Highly Efficient Delivery of Adeno-Associated Viral Vectors to the Primate Retina

“…Interaction of the capsid with heparan sulfate proteoglycan (HSPG), as shown by binding affinity of the capsid to a heparin column, is a strict requirement for intravitreally mediated transduction when the ILM is intact. 64 Interestingly, these two capsid mutants retain the ability to bind heparin, but with lower affinity than wild-type AAV2. 20,64 It has been hypothesized that the improved transduction is partially due to the fact that these variants interact with the ILM via HSPG but do not become sequestered there and thus can traffic further into the retina.…”

“…64 Interestingly, these two capsid mutants retain the ability to bind heparin, but with lower affinity than wild-type AAV2. 20,64 It has been hypothesized that the improved transduction is partially due to the fact that these variants interact with the ILM via HSPG but do not become sequestered there and thus can traffic further into the retina. These two capsids have already been shown to have tropism for Mü ller glia and ON BCs and, therefore, they will also be interesting candidates for testing via this route.…”