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Da Sun, Rebecca Schur, Avery Sears, Songqi Gao, Rui Xin, Akiko Maeda, Krzysztof Palczewski, Zheng-Rong Lu; Efficient Non-viral Gene Therapy for Stargardt’s Disease with pH-Sensitive Multifunctional Lipid ECO Plasmid DNA Nanoparticles. Invest. Ophthalmol. Vis. Sci. 2019;60(9):2915.
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© ARVO (1962-2015); The Authors (2016-present)
Monogenic retinal dystrophies can be treated with gene replacement therapy (GRT) due to easy localized delivery into retina. Here, we developed a nanoparticle-based GRT using pH-sensitive multifunctional lipid ECO and therapeutic ABCA4 plasmids to treat Stargardt’s disease (STGD), which requires delivery of large ABCA4 gene.
Therapeutic plasmids expressing ABCA4 induced by rod photoreceptor-specific RHO promoter and non-specific CMV promoter were designed. Subretinal treatments were performed using ECO/pRHO-ABCA4 and ECO/pCMV-ABCA4 nanoparticles to abca4-/- mice (STGD model). Treatment efficacy was evaluated by analysis of A2E accumulation in the RPE after 6 months. ABCA4 expression was evaluated by qRT-PCR and IHC at 7 days and 8 months after a single injection.
GRT using nanoparticles reduced A2E accumulation at least for 6 months. Demonstrated by HPLC (Figure 1A) and quantitative A2E levels (Figure 1B) of treated abca4-/- mice, an averaged 35% reduction in A2E levels was observed for ECO/pRHO-ABCA4 and a 15% reduction for ECO/CMV-ABCA4. ABCA4 mRNA (Figure 1C) were measured by PCR in mice treated with both nanoparticles. Expression driven by RHO promoter was 10-fold higher in treated eye than control at 8 months, but the expression driven by CMV was slightly but not significantly higher than control. IHC staining for ABCA4 protein (Figure 1D) revealed significant protein expression for ECO/pRHO-ABCA4 treated group and no significant difference between ECO/pCMV-ABCA4 group and control. The expression driven by RHO demonstrated excellent tissue specificity in the retinal outer segments.
Successful GRT of STGD using ECO/pABCA4 nanoparticles was achieved with significant tissue specific expression of ABCA4 up to 8 months, and slowdown of A2E accumulation and STGD progression up to 6 months. The non-viral ECO/plasmid nanoparticles are a promising GRT strategy for a broad range of visual dystrophies.
This abstract was presented at the 2019 ARVO Annual Meeting, held in Vancouver, Canada, April 28 - May 2, 2019.
Figure 1. GRT using ECO/pRHO-ABCA4 and ECO/pCMV-ABCA4 nanoparticles in abca4-/- mice. HPLC analysis of A2E in abca4-/- mice 6 months after treatments with (A) chromatograms showing retention time of A2E and (B) A2E levels from all treated mice. (C) ABCA4 mRNA levels and (D) expression in the retina of abca4-/- mice after 8 months. (Control: contralateral eye injected with PBS. **p < 0.05. Green: ABCA4.)
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