Enhancement of rAAV2-Mediated Transgene Expression in Retina Cells In Vitro and In Vivo by Coadministration of Low-Dose Chemotherapeutic Drugs

PURPOSE. Recombinant adeno-associated viral vector serotype 2 (rAAV2) has been used with success to deliver retina-targeted gene therapeutics in retinal degeneration. However, one of the major limitations of this approach is the vector's low transduction efficiency. This study is designed to increase AAV2 transduction efficiency in vitro and in vivo. METHODS. Green fluorescence protein (GFP) or luciferase reporter gene-carried rAAV2 vectors were applied to cultured human RPE cells (ARPE-19) or animal eyes with or without chemotherapeutic agents. GFP transduction efficiency was evaluated by image, flow cytometry analysis, and Western blot. The ciliary neurotrophic factor (rAAV2-CNTF)-carried AAV2 vector was coinjected to subretinal space with or without chemotherapeutic agent. The therapeutic efficacy was evaluated by counting numbers of remaining photoreceptors in retina sections of treated or untreated eyes. RESULTS. Coadministration of 0.1 mu g/mL doxorubicin (DXR), 0.14 mu g/mL cytarabine (Ara-C), 1 mu g/mL etoposide (VP-16), or 20 mu g/mL cisplatin (DDP) significantly increased rAAV2-mediated GFP and/or luciferase expression in cultured hRPE cells without any detectable toxicity. Pretreatment with DXR for 24 h prior to infection was most effective in enhancing rAAV2 transgene expression in hRPE cells. In addition, subretinal coinjection of rAAV2-CMV-ciliary neurotrophic factor (rAAV2-CNTF) and DXR into the eyes of rats with inherited retinal degeneration resulted in an approximately 2-fold increase in photoreceptor layer thickness and cellular density of the outer nuclear layer (ONL) compared to rAAV2-CNTF alone, reflecting a pronounced protection effect mediated by the enhanced expression of CNTF. CONCLUSIONS. The method described here to improve rAAV2-based gene delivery is simple and feasible without any detectable toxicity. This strategy might be therapeutically exploited in the gene therapy of degenerative retinal diseases. (Invest Ophthalmol Vis Sci. 2012;53:2675-2684) DOI:10.1167/iovs.11-8856