Receptor-targeted transfection using stable maleimido-transferrin/thio-poly-L-lysine conjugates.
Cell surface ligand-receptor systems provide favorable routes for DNA transfection due to target cell specificity, transfer efficiency, and low toxicity. Using the transferrin receptor system as a model, an approach to transfection is developed here within which involves the complexing of DNA to stable maleimido-transferrin/thio-poly-L-lysine conjugates. These studies establish the importance of precise stoichiometry for activity of ligand:poly-L-lysine conjugates, as well as a chemistry for their controlled conjugation. Also considered quantitatively are effects of the following related parameters on the efficiency of receptor-mediated transfection: lysine polymer length, conjugate concentration, DNA:conjugate ratio, and treatment of target cells with chloroquine and desferrioxamine. Compared directly to standard procedures (electroporation, modified DEAE-dextran, lipofection, and modified Ca2PO4 protocols), transfection via this transferrin receptor-mediated system was > or = 10-fold more efficient, and essentially nontoxic to erythroleukemic F-MEL and J2E cells. Following transfection these cells retained the physiological capacity to undergo induced differentiation in response to dimethyl sulfoxide (F-MEL cells), or to erythropoietin (J2E cells), the natural hormonal regulator of erythropoiesis. Thus, an optimized approach to transferrin receptor-mediated transfection is developed which should be widely applicable for alternate cells and ligand-receptor systems both in vitro and in vivo.
Taxman
D J
DJ
Department of Molecular and Cell Biology, Pennsylvania State University, University Park 16802.
Lee
E S
ES
Wojchowski
D M
DM
D000818:Animals; D002454:Cell Differentiation / Q000187:drug effects; D004247:DNA / Q000737:chemistry / Q000235:genetics*; D004355:Drug Stability; D004705:Endocytosis; D004921:Erythropoietin / Q000494:pharmacology; D005622:Friend murine leukemia virus; D004915:Leukemia, Erythroblastic, Acute / Q000235:genetics / Q000382:microbiology / Q000473:pathology; D008301:Maleimides / Q000737:chemistry / Q000378:metabolism; D051379:Mice; D011107:Polylysine / Q000031:analogs & derivatives / Q000737:chemistry; D011990:Receptors, Transferrin / Q000235:genetics / Q000378:metabolism / Q000502:physiology*; D014162:Transfection / Q000379:methods*; D014168:Transferrin / Q000031:analogs & derivatives* / Q000737:chemistry / Q000378:metabolism*; D014407:Tumor Cells, Cultured
D008301:Maleimides; D011990:Receptors, Transferrin; D014168:Transferrin; C072481:transferrin-polylysine conjugate; D004921:Erythropoietin; D011107:Polylysine; D004247:DNA