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Gabriele Thumann, Martina Kropp, Nina Harmening, ; Transposon-Based, Targeted Ex Vivo Gene Therapy to Treat Age-related Macular Degeneration. Invest. Ophthalmol. Vis. Sci. 2015;56(7 ):1820.
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The current treatment for neovascular AMD of repeated life-long intravitreal injections of anti-VEGF antibodies may have significant side-effects. The TargetAMD project, funded within the EU 7th framework program, comprises 13 partners from 3 universities, 2 hospitals, 3 research institutes and 5 industrial partners from 8 countries, who together will develop procedures, reagents, devices, and technologies to successfully complete 2 phase Ib/IIa clinical trials for the treatment of exudative AMD. In the approach, autologous retinal (clinical trial 1) and iris pigment epithelial cells (clinical trial 2) will be isolated and transfected with the PEDF gene using the non-viral Sleeping Beauty (SB100X) transposon system; then they will be transplanted to the subretinal space of patients within one surgical session.
Using the SB100X transposon system, we have electroporated with the PEDF gene 10,000 pigment epithelial cells, a number that can be obtained from a peripheral retina or iris biopsy. Transfection efficiency and safety was optimized by the use of pFAR4 miniplasmids. The electroporation protocol is adapted by modifying an electroporation device, already approved for the use in clinical trials, to efficiently transfect low cell numbers. The electroporation buffer was optimized to obtain high transfection efficiencies and high cell viability. In vivo experiments are performed to evaluate the safety and the benefit of the transfected and transplanted cells.
By using the pFAR4 miniplasmids and the modified electroporation device, primary IPE and RPE cells could be efficiently and stably transfected with PEDF and the fluorescent Venus protein (20-55% efficiency). With the new developed electroporation buffer, transfection efficiency and viability was comparable or better than obtained with commercially available buffers. Transplantation of PEDF-transfected cells subretinally showed feasibility of the procedure and significant reduction of choroidal neovascularization (CNV) in a rat model.
During the last two years, the consortium has made significant and essential progress towards the goal of performing phase Ib/IIa clinical trials to treat neovascular AMD with a gene therapeutic approach. Here we have shown that the approach is not only feasible, but it is beneficial as shown by the reduced neovascularization in a laser-induced CNV model.
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