Development of transferrin-polycation/DNA based vectors for gene delivery to melanoma cells.
We describe the comparison of non-viral polycation transfection reagents, adenovirus-enhanced transferrinfection (AVET), polyethylenimine (PEI800) and transferrin-conjugated PEI800 (Tf-PEI800) in their ability to transfect murine and primary human melanoma cell lines. Expression of a reporter gene, cell surface marker and secreted protein (interleukin-2) was assessed for each vector system. Testing for luciferase reporter gene expression in murine and primary human cell lines, AVET and Tf-PEI800, both showed high levels of expression and comparable activity. Furthermore, when the melanoma cell line B16F10 was transfected with a cell surface marker up to approximately 97% of the cells expressed the protein on the cell surface. Assessing the levels of secreted IL-2 in murine cell lines, AVET/IL-2, Tf-PEI800/IL-2 and PEI800/IL-2 all expressed high levels of the cytokine (up to 20 microg IL-2/10(6) cells/24 h). In primary human melanoma cell lines, AVET/IL-2 transfected cells secreted more IL-2 than cells transfected with either Tf-PEI800/IL-2 or PEI800/IL-2. In murine melanoma cell culture experiments, positively charged PEI800/DNA and Tf-PEI800/DNA complexes gave similar transfection efficiencies. However, when subcutaneous tumors in mice were injected with the luciferase reporter gene complexed with either Tf-PEI800 or AVET, higher transfection activity was measured in the tumors as compared to ligand free PEI800/DNA complexes.
Wightman
L
L
Boehringer Ingelheim Austria, Vienna. lionel.wightman@vie.boehringer-ingelheim.com
Patzelt
E
E
Wagner
E
E
Kircheis
R
R
D000256:Adenoviridae / Q000737:chemistry; D000818:Animals; D002412:Cations / Q000737:chemistry*; D002460:Cell Line; D016207:Cytokines / Q000378:metabolism; D004247:DNA / Q000378:metabolism*; D016503:Drug Delivery Systems / Q000379:methods*; D017930:Genes, Reporter / Q000235:genetics; D005822:Genetic Vectors / Q000235:genetics*; D006801:Humans; D007376:Interleukin-2 / Q000378:metabolism; D008156:Luciferases / Q000235:genetics; D008545:Melanoma / Q000378:metabolism*; D008565:Membrane Proteins / Q000378:metabolism; D051379:Mice; D011094:Polyethyleneimine / Q000737:chemistry; D014162:Transfection / Q000235:genetics; D014168:Transferrin / Q000235:genetics*
D002412:Cations; D016207:Cytokines; D007376:Interleukin-2; D008565:Membrane Proteins; D014168:Transferrin; D011094:Polyethyleneimine; D004247:DNA; D008156:Luciferases