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W. W. Hauswirth, B. Chang, S. L. Boye, J. Alexanda, J.-J. Pang; Gene Therapy Prevents Early Cone Degeneration in a Natural Animal Model of RRP65 Leber Congenital Amaurosis: The rd12 Mouse. Invest. Ophthalmol. Vis. Sci. 2007;48(13):1688.
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© ARVO (1962-2015); The Authors (2016-present)
To test whether AAV-mediated gene delivery can prevent cone degeneration in a rodent model with a recessive Rpe65 mutation, the rd12 mouse.
1 µl (~1010 vector genome particles) of self-complimentary AAV vector containing a small CBA promoter driving expression of human RPE65 (scAAV5-smCBA-hRPE65) was injected subretinally into one eye of rd12 mice at postnatal day 14 (P14). Three weeks later at P35, the contra lateral eye was injected with the same vector. Electroretinograms (ERGs) were recorded regularly. Five months after the second injection, both eyes were enucleated and retinal whole mounts were prepared for peanut agglutinin (PNA) staining. For comparison, PNA staining was also done on uninjected eyes of rd12 mice at 2 weeks, 5 weeks and 5 months.
In rd12 eyes treated with scAAV5-smCBA-hRPE65 at P14, restored dark-adapted (rod) and light-adapted (cone) ERGs were detected as early as 4 days after treatment. These ERGs became stable in amplitude around 2 weeks post-treatment and remained so for an additional 5 months, the longest time assessed following treatment. Restored rod and cone b-wave amplitudes were 60-70% of those from isogenic normal mice. Restoration of rod ERGs were also observed soon after P35 treatment of the second eye. By 5 months after this treatment, rod ERGs remained stable and approximately equivalent in both the P14 and P35 treated eyes. However, there was a distinct difference between cone function in eyes treated at P14 vs. P35. A 60% of normal cone b-wave amplitude remained in the P14 treated eye as apposed to no recordable cone ERG in the partner eye treated at P35. To help understand this finding, retinal whole mount PNA staining of untreated and treated rd12 eyes was carried out to determine the number of cones remaining over time. In untreated eyes, cone number was relatively normal at P14, while at 5 weeks and 5 months cones were absent except in some regions of the dorsal and temporal quadrants. In eyes treated at P14, cones remained relatively normal throughout the whole retina. In contrast, P35 treated eyes showed a significant loss of cones, again with the exception of the dorsal and temporal quadrants, a pattern that resembled an untreated rd12 eye.
AAV mediated gene therapy can prevent early cone degeneration as seen in this naturally occurring mouse lacking RPE65 function, but only if therapy precedes early cone cell death. This work also shows that a self-complimentary AAV5 vector can restore substantial visual function within 4 days of treatment.
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