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R.J. Marano, N. Wimmer, P.S. Kearns, B.G. Thomas, I. Toth, A.S. Wilson, M. Brankov, P.E. Rakoczy; Use of Synthetic Dendrimer Peptide’s to Mediate the Delivery of a Sense Oligonucleotide . Invest. Ophthalmol. Vis. Sci. 2003;44(13):1078.
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Purpose: To determine if lipid-lysine dendrimers are a viable option for the delivery of oligonucleotides for use in gene therapy. Methods: D407 cells were transfected with nine different dendrimers complexed with an oligonucleotide (ODN-1) proven to possess an anti vascular endothelial growth factor (VEGF) effect. The efficacy of the dendrimers to deliver ODN-1 to the target site was determined by calculating the levels of VEGF protein and mRNA expression under hypoxic conditions at 24 and 48 hours post transfection using ELISA and RT-PCR respectively, and comparing this to results obtained using a commercially available transfecting agent. The two most effective dendrimer complexes were subsequently injected into the vitreous of rat eyes and later laser photocoagulated to induce choroidal neovascularisation (CNV). The extent of CNV was determined using fluorescein angiography. Results: In vitro data indicated that all of the dendrimer / ODN-1 complexes resulted in a 40% to 60% decrease in the production of both VEGF protein and mRNA in the first 24 hour period. However, after 48 hours, several of the dendrimers were unable to maintain a reduction in the expression of VEGF indicating poor DNA protection qualities. Both the transfecting and protective ability seemed to be related to the length and number of lipidic amino acids (Laa's) associated with each dendrimer. It was found that dendrimer 4, which possessed two C14 Laa's and eight free amino groups, achieved the second highest transfection efficacy of 89% and in addition maintained the greatest reduction in VEGF expression for the 24 and 48 hour time periods (48% - 50% respectively). In vivo, eyes that were treated with dendrimer 4 showed a 70% lower rate of CNV compared to that of eyes treated with dendrimer minus the oligonucleotide for up to 3 months post injection / lasering. Conclusion: We have shown that synthetic lipophilic charged dendrimers can be used for gene delivery both in vivo and in vitro, resulting in a therapeutic outcome and will be a valuable tool in gene therapy.
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