title: Folate copolymer-mediated transfection of cultured cells.
abstract: Poly(ethylene glycol) of various sizes was used as a molecular spacer to separate the cell-targeting ligand, folate, from the surface of poly-L-lysine. The resulting ternary macromolecule (pLys-PEG-folate) was investigated in various formulations for its ability to transfect reporter plasmids into receptor-bearing HeLa and IGROV cell lines. Formulations were optimized with respect to DNA content, +/- charge ratio, and the size and amount of PEG substitution off the pLys backbone. Transfection activity was highest 48 h after sample introduction, and PEG 3400 was determined to be the most favorable spacer size tested. pLys-PEG-folate:DNA transfection was also found to be both concentration dependent and saturable; plus, it was blocked by the addition of excess-free folate, indicative of a specific mechanism of uptake. Transfection activity was virtually identical for complexes formed in 10% serum-supplemented media, deionized water, or Hepes buffer. And, cell viability remained greater than 85% at the highest concentrations of pLys-PEG-folate:DNA complexes tested (4.8 microg/mL pLys 331 000; 12 microg/mL DNA). Taken together, these observations provide evidence that pLys-PEG-folate:DNA complexes are taken up specifically by the folate endocytosis pathway, and that the intramolecular spatial distance of the ligand from the pLys backbone dramatically influences transfection.
authors.affiliation: Department of Oligomer Development, GlaxoWellcome Research Institute, 5 Moore Drive, Research Triangle Park, North Carolina 27709, USA. Leamon@Endocyte.com
authors.forename: C P
authors.initials: CP
authors.lastname: Leamon
authors.forename: D
authors.initials: D
authors.lastname: Weigl
authors.forename: R W
authors.initials: RW
authors.lastname: Hendren
mesh_terms: D002412:Cations; D002460:Cell Line; D002470:Cell Survival; D004247:DNA / Q000008:administration & dosage* / Q000737:chemistry; D004705:Endocytosis; D005492:Folic Acid / Q000737:chemistry* / Q000378:metabolism; D006367:HeLa Cells; D006651:Histocytochemistry; D006801:Humans; D007700:Kinetics; D008081:Liposomes; D010957:Plasmids / Q000235:genetics; D011092:Polyethylene Glycols / Q000737:chemistry*; D011107:Polylysine / Q000737:chemistry*; D013329:Structure-Activity Relationship; D014162:Transfection; D001616:beta-Galactosidase / Q000032:analysis / Q000235:genetics
chemical_list: D002412:Cations; D008081:Liposomes; D011107:Polylysine; D011092:Polyethylene Glycols; D004247:DNA; D005492:Folic Acid; D001616:beta-Galactosidase