Abstract
Two synthetic peptides comprising an RGD moiety for integrin binding and a polylysine moiety for DNA binding were tested for transfection efficiency under a variety of different conditions. Binding of target cells to the peptide was shown to be strongly dependent on cyclisation of the peptides via cysteine residues. Low (10 μ M) concentrations of chloroquine, added to assist endocytic exit, unexpectedly reduced transfection efficiency in two of the cell lines tested, COS-7 and ECV304. However, transfection efficiency increased at higher chloroquine concentrations and exceeded that in the absence of chloroquine in the case of the COS-7 and A375M cell lines. With the ECV304 cell line, optimum transfection occurred in the absence of chloroquine. Transfection efficiency of the peptides was greatest at peptide:DNA ratios of 4:1 (w/w), which were calculated to generate complexes containing approximately 5000 peptide molecules per plasmid. This represented approximately a 6:1 ratio of positive to negative charges. Peptide 5 was shown to have a higher transfection efficiency under most conditions, possibly because of more efficient stabilisation of cyclisation by two cysteine–cysteine bonds.
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Hart, S., Collins, L., Gustafsson, K. et al. Integrin-mediated transfection with peptides containing arginine-glycine-aspartic acid domains. Gene Ther 4, 1225–1230 (1997). https://doi.org/10.1038/sj.gt.3300513
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DOI: https://doi.org/10.1038/sj.gt.3300513
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