Cryoconserved shielded and EGF receptor targeted DNA polyplexes: cellular mechanisms.
Recently, cryoconservable polyethylene glycol (PEG)-shielded and epidermal growth factor receptor (EGFR)-targeted polyplexes (EGF+ polyplexes) were engineered in our laboratory for tumor-directed transfer and expression of DNA. Here, we further analyzed specificity and kinetics of EGFR-mediated cellular uptake of these polyplexes. Similar to our previous results, EGF+ polyplexes significantly enhanced the transfection efficiency as compared to polyplexes without EGF (EGF- polyplexes) in HUH-7 hepatoma cells and Renca-EGFR renal carcinoma cells. EGF+ polyplexes rapidly associated with the cells within 30 min of exposure, and binding of EGF+ polyplexes to the cells after 4 h was significantly higher than that of EGF- polyplexes. In the presence of free EGF, both cell association and transfection efficiency of EGF+ polyplexes were markedly reduced indicating that these effects were primarily mediated via ligand receptor interaction. Fluorescence microscopy revealed that the cell-associated EGF+ polyplexes aggregated to micrometer sized clusters, resembling typical clustering of receptors upon ligand binding. In conclusion, EGFR-targeting enhances transfection efficiency due to accelerated and increased cell association followed by aggregation of the bound EGF+ polyplexes.
von Gersdorff
Katharina
K
Pharmaceutical Biology-Biotechnology, Department of Pharmacy, Ludwig-Maximilians-Universität, Munich, Germany.
Ogris
Manfred
M
Wagner
Ernst
E
D002478:Cells, Cultured; D004247:DNA / Q000378:metabolism*; D004815:Epidermal Growth Factor / Q000378:metabolism; D066246:ErbB Receptors / Q000378:metabolism*; D018014:Gene Transfer Techniques; D006801:Humans; D014162:Transfection
D004815:Epidermal Growth Factor; D004247:DNA; D066246:ErbB Receptors