title: The human rhodopsin kinase promoter in an AAV5 vector confers rod- and cone-specific expression in the primate retina.
abstract: Adeno-associated virus (AAV) has proven an effective gene delivery vehicle for the treatment of retinal disease. Ongoing clinical trials using a serotype 2 AAV vector to express RPE65 in the retinal pigment epithelium have proven safe and effective. While many proof-of-concept studies in animal models of retinal disease have suggested that gene transfer to the neural retina will also be effective, a photoreceptor-targeting AAV vector has yet to be used in the clinic, principally because a vector that efficiently but exclusively targets all primate photoreceptors has yet to be demonstrated. Here, we evaluate a serotype 5 AAV vector containing the human rhodopsin kinase (hGRK1) promoter for its ability to target transgene expression to rod and cone photoreceptors when delivered subretinally in a nonhuman primate (NHP). In vivo fluorescent fundus imaging confirmed that AAV5-hGRK1-mediated green fluorescent protein (GFP) expression was restricted to the injection blebs of treated eyes. Optical coherence tomography (OCT) revealed a lack of gross pathology after injection. Neutralizing antibodies against AAV5 were undetectable in post-injection serum samples from subjects receiving uncomplicated subretinal injections (i.e., no hemorrhage). Immunohistochemistry of retinal sections confirmed hGRK1 was active in, and specific for, both rods and cones of NHP retina. Biodistribution studies revealed minimal spread of vector genomes to peripheral tissues. These results suggest that AAV5-hGRK1 is a safe and effective AAV serotype/promoter combination for targeting therapeutic transgene expression protein to rods and cones in a clinical setting.
authors: Department of Ophthalmology, University of Florida College of Medicine, Gainesville, FL 32610, USA. shannon.boye@eye.ufl.edu
authors: Shannon E
authors: SE
authors: Boye
authors: John J
authors: JJ
authors: Alexander
authors: Sanford L
authors: SL
authors: Boye
authors: Clark D
authors: CD
authors: Witherspoon
authors: Kristen J
authors: KJ
authors: Sandefer
authors: Thomas J
authors: TJ
authors: Conlon
authors: Kirsten
authors: K
authors: Erger
authors: Jingfen
authors: J
authors: Sun
authors: Renee
authors: R
authors: Ryals
authors: Vince A
authors: VA
authors: Chiodo
authors: Mark E
authors: ME
authors: Clark
authors: Christopher A
authors: CA
authors: Girkin
authors: William W
authors: WW
authors: Hauswirth
authors: Paul D
authors: PD
authors: Gamlin
mesh_terms: D000818:Animals; D057134:Antibodies, Neutralizing / Q000276:immunology; D001483:Base Sequence; D000229:Dependovirus / Q000235:genetics*; D005654:Fundus Oculi; D051606:G-Protein-Coupled Receptor Kinase 1 / Q000235:genetics*; D015870:Gene Expression; D015316:Genetic Therapy; D005822:Genetic Vectors / Q000235:genetics; D049452:Green Fluorescent Proteins / Q000378:metabolism; D006801:Humans; D007150:Immunohistochemistry; D008251:Macaca / Q000378:metabolism*; D008297:Male; D051379:Mice; D008810:Mice, Inbred C57BL; D009928:Organ Specificity; D011401:Promoter Regions, Genetic / Q000235:genetics*; D012160:Retina / Q000378:metabolism*; D017949:Retinal Cone Photoreceptor Cells / Q000378:metabolism*; D017948:Retinal Rod Photoreceptor Cells / Q000378:metabolism*; D014018:Tissue Distribution; D041623:Tomography, Optical Coherence
chemical_list: D057134:Antibodies, Neutralizing; D049452:Green Fluorescent Proteins; D051606:G-Protein-Coupled Receptor Kinase 1; C496764:GRK1 protein, human