Postsurgical Assessment and Long-term Safety of Recombinant Adeno-Associated Virus–Mediated Gene Transfer Into the Retinas of Dogs and Primates

Meur, Guylène Le

doi:10.1001/archopht.123.4.500

Cited by 36 publications

(26 citation statements)

References 36 publications

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“…We found that retinal adhesion differs between individual RPE65-deficient animals and this affected the size of the retinal detachment created during the subretinal injection. This variation, which has not been observed in the retina of normal dogs or primates, 22,23,27 is likely to be associated with RPE65…”

Section: Discussionmentioning

confidence: 68%

“…In previous studies involving subretinal injection of rAAV vectors in normal dogs and macaques, such abnormalities were never observed. 22,23,27 Fortunately, the detected retinal abnormalities seen in all animals treated under a year of age did not prevent rescue of retinal function. All the treated dogs, except A6 that was treated at a much older age, showed retinal and visual functions.…”

Section: Discussionmentioning

confidence: 94%

“…In all animals, only one eye was treated, with the contralateral eye serving as an internal control for ophthalmic examination and vision testing. Based on our previous experience with subretinal injection in dogs and primates, 22,23 our initial aim was to inject 400-500 ml of vector suspension into each treated retina. We observed that in contrast to normal retinas, the retinal detachment, or bleb, created during subretinal injections in RPE65 À/À dogs was not reproducible in all animals, and was likely due to individual variation in retinal adhesion.…”

Section: Resultsmentioning

confidence: 99%

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Restoration of vision in RPE65-deficient Briard dogs using an AAV serotype 4 vector that specifically targets the retinal pigmented epithelium

Meur

Stieger

Smith³

et al. 2006

Gene Ther

182

150

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Previous studies have tested gene replacement therapy in RPE65-deficient dogs using recombinant adeno-associated virus 2/2 (rAAV2/2), -2/1 or -2/5 mediated delivery of the RPE65 gene. They all documented restoration of dark-and light-adapted electroretinography responses and improved psychophysical outcomes. Use of a specific RPE65 promoter and a rAAV vector that targets transgene expression specifically to the RPE may, however, provide a safer setting for the long-term therapeutic expression of RPE65. Subretinal injection of rAAV2 pseudotyped with serotype 4 (rAAV2/4) specifically targets the RPE. The purpose of our study was to evaluate a rAAV2/4 vector carrying a human RPE65cDNA driven by a human RPE65 promoter, for the ability to restore vision in RPE65 À/À purebred Briard dogs and to assess the safety of gene transfer with respect to retinal morphology and function. rAAV2/4 and rAAV2/2 vectors containing similar human RPE65 promoter and cDNA cassettes were generated and administered subretinally in eight affected dogs, ages 8-30 months (n ¼ 6 with rAAV2/4, n ¼ 2 with rAAV2/2). Although fluorescein angiography and optical coherence tomography examinations displayed retinal abnormalities in treated retinas, electrophysiological analysis demonstrated that restoration of rod and cone photoreceptor function started as soon as 15 days post-injection, reaching maximal function at 3 months postinjection, and remaining stable thereafter in all animals treated at 8-11 months of age. As assessed by the ability of these animals to avoid obstacles in both dim and normal light, functional vision was restored in the treated eye, whereas the untreated contralateral eye served as an internal control. The dog treated at a later age (30 months) did not recover retinal function or vision, suggesting that there might be a therapeutic window for the successful treatment of RPE65 À/À dogs by gene replacement therapy.

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Section: Discussionmentioning

confidence: 68%

Section: Discussionmentioning

confidence: 94%

Section: Resultsmentioning

confidence: 99%

See 1 more Smart Citation

Restoration of vision in RPE65-deficient Briard dogs using an AAV serotype 4 vector that specifically targets the retinal pigmented epithelium

Meur

Stieger

Smith³

et al. 2006

Gene Ther

182

150

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“…Large animal studies to date, including primate safety studies, have mainly treated the area centralis or central retinal region (14,(41)(42)(43), so this region is a likely target for subretinal gene therapy. Thus, our finding of early foveal cone loss has strong clinical relevance.…”

Section: Discussionmentioning

confidence: 99%

Human cone photoreceptor dependence on RPE65 isomerase

Jacobson

Alemán

Cideciyan

et al. 2007

Proc. Natl. Acad. Sci. U.S.A.

140

144

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The visual (retinoid) cycle, the enzymatic pathway that regenerates chromophore after light absorption, is located primarily in the retinal pigment epithelium (RPE) and is essential for rod photoreceptor survival. Whether this pathway also is essential for cone photoreceptor survival is unknown, and there are no data from man or monkey to address this question. The visual cycle is naturally disrupted in humans with Leber congenital amaurosis (LCA), which is caused by mutations in RPE65, the gene that encodes the retinoid isomerase. We investigated such patients over a wide age range (3-52 years) for effects on the cone-rich human fovea. In vivo microscopy of the fovea showed that, even at the youngest ages, patients with RPE65-LCA exhibited cone photoreceptor loss. This loss was incomplete, however, and residual cone photoreceptor structure and function persisted for decades. Basic questions about localization of RPE65 and isomerase activity in the primate eye were addressed by examining normal macaque. RPE65 was definitively localized by immunocytochemistry to the central RPE and, by immunoblotting, appeared to concentrate in the central retina. The central retinal RPE layer also showed a 4-fold higher retinoid isomerase activity than more peripheral RPE. Early cone photoreceptor losses in RPE65-LCA suggest that robust RPE65-based visual chromophore production is important for cones; the residual retained cone structure and function support the speculation that alternative pathways are critical for cone photoreceptor survival.optical coherence tomography ͉ retinal degeneration ͉ retinoid cycle ͉ Leber congenital amaurosis ͉ gene therapy V ertebrate retinas have two types of photoreceptors: rods for the perception of dim light and cones for the perception of bright light. Light absorbed by visual pigments of photoreceptors triggers a series of enzymatic reactions known as phototransduction (1). The light-capturing part of the visual pigment, the chromophore, must then be regenerated. The pathway for regeneration of rhodopsin (the rod visual pigment) is known as the visual (retinoid) cycle (2, 3) and involves both rod cells and the adjacent retinal pigment epithelium (RPE). All-trans-retinal is reisomerized to 11-cis-retinal by a reaction in the RPE that requires the retinal isomerase, retinal pigment epithelium-specific 65-kDa protein (RPE65), and lecithinretinol acyltransferase (LRAT) activities. Functional visual pigment is then reformed by combining the 11-cis-chromophore with the photoreceptor opsin apoprotein (4). Molecular knowledge of the visual cycle has increased as the genes encoding the enzymes of this pathway have been elucidated (2, 5, 6).Decades ago, cone pigment regeneration was proposed to be RPE cell-independent (7, 8). More recently, cone-dominant ground squirrel and chicken retinas have been shown to exhibit retinoid isomerase activity that is distinct from RPE65 (5, 9, 10). Murine double knockout experiments, however, provide evidence that RPE65 and the RPE are essential for cone function (11...

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“…The advantages of AAV as a gene transfer vector include its lack of pathogenicity and long-lasting gene expression (Grimm and Kay, 2003). AAV has been used extensively to transduce cells in the retina (Auricchio et al, 2001;Bennett, 2003;Lotery et al, 2003;Acland et al, 2005;Dinculescu et al, 2005;Le Meur et al, 2005;Gorbatyuk et al, 2007). Its tropism in the retina differs from that of LV, raising the possibility of analyzing the effects of target gene knockdown in distinct cell populations.…”

Section: Knockdown In Vivo Using Aav Vectorsmentioning

confidence: 99%

Rapid and Stable Knockdown of an Endogenous Gene in Retinal Pigment Epithelium

et al. 2007

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The selective silencing of target genes in specific cell types by RNA interference (RNAi) represents a powerful approach both to gene therapy of dominantly active mutant alleles, and to the investigation of normal gene function in animal models in vivo. We established a simple and versatile in vitro method for screening the efficacy of DNA-based short hairpin RNAs (shRNAs), and identified a highly effective shRNA targeting basic fibroblast growth factor (bFGF), a gene thought to play important roles in endogenous neuroprotective responses in the rat retina. We used two viral vectors, based on lentivirus and adeno-associated virus (AAV), to deliver shRNAs and silence bFGF in retinal pigment epithelial cells in vivo. The AAV experiments made use of a "stabilized double-stranded" version of these vectors with rapid onset of gene expression. In the rat retinal pigment epithelium, shRNAs delivered by either vector reduced bFGF immunoreactivity to undetectable levels in transduced cells, whereas a nonfunctional control construct incorporating a two-base pair mutation had no measurable effect on bFGF expression. Silencing commenced within a few days after injection of virus and remained stable throughout the period of observation, as long as 60 days. Viral delivery of RNAi constructs offers a powerful and versatile approach for both gene therapy and the analysis of fundamental questions in retinal biology. 871

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Postsurgical Assessment and Long-term Safety of Recombinant Adeno-Associated Virus–Mediated Gene Transfer Into the Retinas of Dogs and Primates

Cited by 36 publications

References 36 publications

Restoration of vision in RPE65-deficient Briard dogs using an AAV serotype 4 vector that specifically targets the retinal pigmented epithelium

Restoration of vision in RPE65-deficient Briard dogs using an AAV serotype 4 vector that specifically targets the retinal pigmented epithelium

Human cone photoreceptor dependence on RPE65 isomerase

Rapid and Stable Knockdown of an Endogenous Gene in Retinal Pigment Epithelium

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