Long Term Rescue Following AAV-Mediated Cone Targeting Gene Therapy to Cpfl5 Mice, a Model for Human Achromatopsia 2 With Cnga3 Mutation

Abstract

Purpose: : Mutations in gene encoding the alpha-subunit of the cone cyclic nucleotide-gated (CNGA3) channels cause cone function loss in mammals. We test if cone targeted CNGA3 gene delivery can restore the cone system function in cpfl5 (Cone Photoreceptor Function Loss 5) mice, a natural model of human Achromatopsia 2 with CNGA3 mutation.

Methods: : At postnatal day 14, 1 µl of AAV5-PR2.1-CNGA3 vector (1 x 10¹³ genome containing viral particles/ml) was injected subretinally into one eye of 20 cpfl5 mice. The untreated contralateral eye was used as control. Dark- and light-adapted ERGs were recorded periodically starting from 3 weeks after injections. 10 months after injection, visual function was assessed with routine ERGs and 10 Hz flicker ERGs. Visual acuity and contrast sensitivity were also tested. Then both treated and control eyes were harvested for histochemical studies.

Results: : In treated eyes, restored light-adapted ERG waveforms were observed at 3 weeks after injections. The restored light-adapted ERGs remained stable for at least 10 months following treatment; the amplitudes were about 50% of those of the normal uninjected C57BL/6J mice. In the contralateral untreated eyes, the cone-driven ERGs were not recordable. The flicker ERGs following 8 hours dark adaption indicated that the treatment significantly improved the cone-driven responses. Behavioral tests showed nearly normal cone-driven visual acuity and contrast sensitivity in the treated eyes of the cpfl5 mice, but not in the untreated eyes. In the treated cpfl5 retinas, immunohistochemistry showed CNGA3 staining in the inner and outer segments of the cones. However, no CNGA3 expression was observed in the untreated eye from the same mouse.

Conclusions: : AAV mediated gene therapy corrects CNGA3 deficiency in a naturally occurring mouse model of human Achromatopsia 2. The genetic intervention restores and maintains the cone function as demonstrated by ERG and behavioral tests for at least 10 months.

Keywords: gene transfer/gene therapy • retinal degenerations: cell biology • color vision