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Diego S. Fajardo, Hilda Petrs-Silva, William W. Hauswirth, Alfred S. Lewin; Supplementation Of Rds In rds/+ Mice Model For Gene Therapy Using Hirbp Promoter In An Aav Vector. Invest. Ophthalmol. Vis. Sci. 2011;52(14):481.
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© ARVO (1962-2015); The Authors (2016-present)
A variety of retinal diseases are associated with mutations in the photoreceptor specific Rds/Peripherin transmenbrane glycoprotein. Rds plays an important role in the morphogenesis and maintenance of the of the disk rim structure of both rods and cones. Consequently, developing a gene therapy for rds/peripherin-associated disease must target both rod and cone photoreceptor cells. Mutations in the RDS gene are haploinsufficient (requiring a full complement of Rds protein). For this reason, we tested the efficacy of a gene supplementation approach of RDS specifically targeting photoreceptors by utilizing the human Interphotoreceptor retinoid-binding protein (hlRBP) promoter. We expect the AAV delivered RDS to partially rescue the rds retinas, resulting in a slower course of photoreceptor loss.
The RDS gene was cloned in recombinant Adeno associated virus of serotype 5 (rAAV5) vectors under the control of the 1.3-kbp human IRBP promoter segment or a 487 bp mouse rod opsin (MOPs) promoter. Virus particles were delivered in only one eye by subretinal injections (1µl at 1 x 1013 vg/ml) in postnatal day 15 mice. Injected mice were monitored on a monthly basis by full-field scotopic electroretinography (ERG). Rds/+/- mice were used to mimic the human disease with one wild type and one mutant allele.
An initial test of the hIRBP promoter driving the humanized green fluorescent protein (hGFP) in rAAV5 showed strong expression of hGFP in cones and rods photoreceptors. Scotopic ERG of injected eyes showed an initial reduction in ERG a- and b-waves amplitude within a month after injections with both the hIRBP and MOPs driven RDS compared to untreated eyes. However, at two months post injection, a slight increase (not statistically significant) in scotopic ERG amplitudes of treated eyes was detected compare to untreated eyes. This slight was maintained for up to five months post injection.
Our results suggest that a gene supplementation approach to RDS haploinsufficency may not be enough to completely rescue the Rds/+/- retina. We are currently testing a suppression and replacement approach in which levels of endogenous wild-type and mutant mRNA RDS are reduced by siRNA and a siRNA-resistant version of the mRNA (hardened RDS) is delivery simultaneously.
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