ENHANCED CELLULAR UPTAKE OF OLIGONUCLEOTIDES BY EGF RECEPTOR-MEDIATEDENDOCYTOSIS IN A549 CELLS

Purpose. The goal of this study was to investigate the feasibility of utilizing epidermal growth factor (EGF) receptor-mediated endocytosis to enhance cellular uptake and targeting of oligonucleotides in epithe lial cancer cells. To overcome the problem of endosomal entrappment as sociated with receptor-mediated delivery, we also examined the effects of two fusogenic peptides, polymyxin B and influenza HA2 peptide, for their capability to promote cytoplasmic delivery of oligonucleotides. Methods. A molecular conjugate consisting of EGF and poly-L-lysine (P L) was synthesized and complexed with 5' fluorescently-labeled oligonu cleotide. Cellular uptake of the complex in presence or absence of the fusogenic peptides was monitored fluorometrically. Microscopic studie s were performed to visualize the intracellular distribution of the ol igonucleotide. Results. Cells treated with the complex exhibited intra cellular fluorescence intensity significantly enhanced over free oligo nucleotide-treated controls. The uptake of the complex was shown to oc cur via the EGF receptor-mediated pathway. Fluorescence microscopic st udies revealed cellular internalization of the complex, however, the c omplex appeared to be accumulated in endocytic vesicles. Exposure of t he cells to complex in presence of HA2 peptide and polymyxin B resulte d in a more diffused intracellular fluorescence pattern and a correspo nding increase in fluorescence intensity. These results are consistent with the known fusion and destabilizing activities of the peptides. C onclusions. Since EGF receptors are overexpressed in many cancer cell types, the EGF-PL conjugate may potentially be used as an effective an d selective delivery system to enhance uptake of oligonucleotides into cancer cells.