Dosage Thresholds for AAV2 and AAV8 Photoreceptor Gene Therapy in Monkey

Abstract

Gene therapy is emerging as a therapeutic modality for treating disorders of the retina. Photoreceptor cells are the primary cell type affected in many inherited diseases of retinal degeneration. Successfully treating these diseases with gene therapy requires the identification of efficient and safe targeting vectors that can transduce photoreceptor cells. One serotype of adeno-associated virus, AAV2, has been used successfully in clinical trials to treat a form of congenital blindness that requires transduction of the supporting cells of the retina in the retinal pigment epithelium (RPE). Here, we determined the dose required to achieve targeting of AAV2 and AAV8 vectors to photoreceptors in nonhuman primates. Transgene expression in animals injected subretinally with various doses of AAV2 or AAV8 vectors carrying a green fluorescent protein transgene was correlated with surgical, clinical, and immunological observations. Both AAV2 and AAV8 demonstrated efficient transduction of RPE, but AAV8 was markedly better at targeting photoreceptor cells. These preclinical results provide guidance for optimal vector and dose selection in future human gene therapy trials to treat retinal diseases caused by loss of photoreceptors.

A New Promoter Allows Optogenetic Vision Restoration with Enhanced Sensitivity in Macaque Retina

“…37 One primate was injected intravitreally with two vector doses (one eye with 1 Â 10 11 and the other with 5 Â 10 11 vg), with CatCh expressed under the SNCG in fusion with GFP to monitor gene expression in vivo (NHP1). As GFP can be immunogenic, 24 we used CatCh without a fluorescent tag for the remaining nine primates. 18 macaque eyes were injected intravitreally with either 1 Â 10 11 (n = 5 eyes) or 5 Â 10 11 (n = 5 eyes) or 1 Â 10 12 AAV2 particles encoding CatCh under the SNCG or CMV promoter (n = 4 eyes) ( Table 1).…”

“…First, optogenetically equipped cells require very high-level opsin expression, as there is no amplification cascade behind microbial opsins. 19 Although adeno-associated viruses (AAVs) have a favorable safety profile in clinical gene therapy, [20][21][22][23] their safety is dose dependent, 24,25 limiting the injected dose. This makes our ability to obtain functional-level microbial opsin expression with a safe viral dose uncertain.…”

“…AAV2 was chosen for gene delivery due to its efficacy in targeting RGCs and to the large body of knowledge using this serotype in non-human primate (NHP) studies 24,28,29 and in human clinical trials. [20][21][22][23]30 To obtain functional responses at lower light intensities, we optimized several parameters.…”

Leber Congenital Amaurosis Caused by Mutations in GUCY2D

“…Keeping in mind that the primary target in LCA1 is the central, cone-rich retina, focus must be placed on each serotype's ability to transduce cone photoreceptors. Only partial cone transduction was achieved following subretinal delivery of elevated doses of AAV8-CMV-GFP (10 11 vg delivered) in nonhuman primates (Vandenberghe et al 2011(Vandenberghe et al , 2013. At a lower dose (10 10 vg delivered), this serotype failed to transduce foveal, parafoveal, or perifoveal cones (Vandenberghe et al 2011).…”

Gene Therapy of Inherited Retinopathies: A Long and Successful Road from Viral Vectors to Patients

“…Over the past years, AAV vectors derived from naturally-occurring serotypes (Auricchio et al, 2001;Gao et al, 2002Gao et al, , 2005 or with rationallymodified (Zhong et al, 2008b) or in vitro evolved capsids (Perabo et al, 2008) have been generated, significantly expanding the potential of AAVs for the treatment of IRs. The most relevant findings have been: (i) the identification of AAV2/8 as the most efficient naturally-occurring serotype for PR transduction in mice (Allocca et al, 2007;Auricchio, 2011), pigs (Mussolino et al, 2011a), and nonhuman primates (NHPs) (Vandenberghe et al, 2011); (ii) the availability of AAV2/7, 2/9, and 2/5 as alternatives to AAV2/8 for PR transduction (Auricchio et al, 2001;Lotery et al, 2003;Allocca et al, 2007;Mussolino et al, 2011a); and (iii) the generation of tyrosine-mutated AAV capsids (Zhong et al, 2008b) that avoid targeting of AAVs to the proteasome, thus increasing the vector load that reaches the nucleus, resulting in increased transgene expression (Zhong et al, 2008a). In particular: (i) the single and triple mutants from AAV2/2 (Y444F and Y444,500,730F) and 2/8-Y733F determine stronger and more widespread transduction than their wild type (wt) counterparts when delivered SR in mice (Petrs-Silva et al, 2009); and (ii) quadruple and pentuple tyrosine-mutant AAV2/2 vectors are capable of transducing murine PRs and RPE when injected IV (Petrs-Silva et al, 2011).…”