Purchase this article with an account.
Joshua T. Bartoe, Freya Mowat, Astra Dinculescu, Sanford Boye, William Hauswirth, Simon Petersen-Jones; AAV Vectors With Engineered Capsid Mutations Efficiently Transduce Outer Retina With Intravitreal Delivery And Result In Rapid Gene Expression With Subretinal Delivery In Dogs. Invest. Ophthalmol. Vis. Sci. 2011;52(14):1420.
Download citation file:
© ARVO (1962-2015); The Authors (2016-present)
To compare transduction onset time and cellular tropism of GFP-expressing AAV vectors with Try-to-Phe capsid mutations compared with self-complimentary AAV vectors following either intravitreal or subretinal injection in canine eyes.
12 normal beagle dogs (24 eyes) were used. All AAV genomes included eGFP driven by the chicken beta actin promoter. Vectors were tittered and diluted to 5x1011 vg/ml. 4 right eyes (OD) received intravitreal AAV serotype 2/2 with 4 Tyr-to-Phe capsid mutations (mutAAV2); 4 contralateral left eyes (OS) received intravitreal AAV serotype 2/2 with a self-complimentary genome (scAAV2). Following 3-port pars plana vitrectomy, 4 OD received subretinal mutAAV2; 4 OS received self-complimentary AAV serotype 2/5 (scAAV5). Following vitrectomy, the remaining 4 OD received subretinal AAV serotype 2/8 with 1 Tyr-to-Phe capsid mutation (mutAAV8); 4 OS received scAAV5. Ophthalmic examination and color and fluorescent fundus photography were performed regularly. Fundic images were identically computer enhanced to detect low-level fluorescence. Eyes were fixed following euthanasia at 28 days post-injection. Retinal cryosections were evaluated for GFP. Contralateral eyes were compared statistically via paired t-test.
Onset of GFP expression was detected in all dogs with intravitreal injection of mutAAV2 by 9 days and scAAV2 by 8 days; fluorescence was similar for both vectors throughout the study. GFP expression was detected in all dogs with subretinal injection of mutAAV2 and mutAAV8 by 3 days and scAAV5 by 5 days. However, fluroescence was greater for mutAAV8 compared to both mutAAV2 and scAAV5. Histology showed no difference in transduction of inner retinal cells between intravitreal mutAAV2 and scAAV2; however, mutAAV2 more efficiently transduced the outer retina and RPE. All subretinal injected vectors efficiently transduced outer retina and RPE.
AAV 2/2 vectors with engineered capsid mutations are efficient in transduction of outer retina following intravitreal delivery. AAV 2/2 and 2/8 vectors with engineered capsid mutations produce more rapid and robust gene expression following subretinal delivery. AAV vectors with engineered capsid mutations are likely to be a critical resource in gene replacement therapy for rapidly progressive heritable retinal degenerations.
This PDF is available to Subscribers Only