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X. Cai, S. M. Conley, M. J. Cooper, S. J. Fliesler, M. I. Naash; Targeting the RPE: A Novel Application for Compacted-DNA Nanoparticles. Invest. Ophthalmol. Vis. Sci. 2008;49(13):4782.
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Previously we have shown that compacted DNA nanoparticles containing either a reporter gene or a therapeutic gene can successfully transfect both the adult and developing retina. In the present study we evaluated the ability of nanoparticles to specifically deliver genes to the developing retinal pigment epithelium (RPE).
Vectors containing the cDNA for either EGFP or the human RPE-specific retinoid isomerohydrolase, RPE65, and either the CMV or the RPE-specific vitelliform macular dystrophy (VMD2) promoter were constructed and compacted as acetate-CK30PEG10k nanoparticles. Nanoparticles (with naked DNA and saline as controls) were delivered by subretinal injection at postnatal day 5 to one eye of wild-type or RPE65-/- mice (with the contralateral uninjected eye serving as a control). Gene expression levels were analyzed by quantitative RT-PCR at 2, 7, and 30 days post- injection (PI), while expression patterns were analyzed by fluorescence microscopy and immunohistochemistry.
After injection at postnatal day 5, gene expression from all four nanoparticles peaked at PI-2 with levels from CMV-RPE65 injected eyes up to 10-fold higher than in the uninjected eyes. Expression levels at PI-7 were lower than at PI-2, but still significantly higher than in controls. Injection of all nanoparticles (including those with the RPE-specific promoter) resulted in detectable levels of the delivered gene’s product in nascent photoreceptor outer segments at PI-2 and PI-7, as well as strong RPE layer labeling detectable at later timepoints (PI-30).
Successful transfer of both reporter and therapeutic genes to the RPE has been achieved using compacted DNA nanoparticles, and was accompanied by high levels of gene expression for at least 30 days post-injection. Compacted DNA nanoparticles provide a promising gene therapy candidate for treating RPE-associated disorders,e.g., Leber congenital amaurosis type 2 (LCA2), congenital stationary night blindness (CSNB), and autosomal recessive retinitis pigmentosa (AR-RP).
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