title: Transferrin-associated lipoplexes as gene delivery systems: relevance of mode of preparation and biophysical properties.
abstract: The successful application of gene therapy depends highly on understanding the properties of gene carriers and their correlation with the ability to mediate transfection. An important parameter that has been described to improve transfection mediated by cationic liposomes involves association of ligands to cationic liposome-DNA complexes (lipoplexes). In this study, ternary complexes composed of 1,2-dioleoyl-3-(trimethylammonium) propane:cholesterol, plasmid DNA and transferrin (Tf, selected as a paradigm of a ligand) were prepared under various conditions, namely, in medium with different ionic strengths (HEPES-buffered saline [HBS] or dextrose), at different lipid/DNA (+/-) charge ratios and using different modes for component addition. We investigated the effect of these formulation parameters on transfection (in the absence and presence of serum), size of the complexes, degree of DNA protection and extent of their association with cells (in terms of both lipid and DNA). Our results show that all the tested parameters influenced to some extent the size of the complexes and their capacity to protect the carried genetic material, as well as the levels of cell association and transfection. The best transfection profile was observed for ternary complexes (Tf-complexes) prepared in high ionic strength solution (HBS), at charge ratios close to neutrality and according to the following order of component addition: cationic liposomes-Tf-DNA. Interestingly, in contrast to what was found for dextrose-Tf-complexes, transfection mediated by HBS-Tf-complexes in the presence of serum was highly enhanced.
authors.affiliation: Center for Neuroscience and Cell Biology, University of Coimbra, Coimbra 3000, Portugal.
authors.forename: Nuno
authors.initials: N
authors.lastname: Penacho
authors.forename: Ana
authors.initials: A
authors.lastname: Filipe
authors.forename: Sérgio
authors.initials: S
authors.lastname: Simões
authors.forename: Maria C
authors.initials: MC
authors.lastname: Pedroso de Lima
mesh_terms: D000818:Animals; D019556:COS Cells; D002522:Chlorocebus aethiops; D002784:Cholesterol / Q000737:chemistry*; D004247:DNA / Q000737:chemistry*; D005229:Fatty Acids, Monounsaturated / Q000737:chemistry*; D005947:Glucose / Q000737:chemistry; D006801:Humans; D008081:Liposomes / Q000737:chemistry*; D009994:Osmolar Concentration; D000644:Quaternary Ammonium Compounds / Q000737:chemistry*; D014162:Transfection / Q000379:methods*; D014168:Transferrin / Q000737:chemistry*
chemical_list: D005229:Fatty Acids, Monounsaturated; D008081:Liposomes; D000644:Quaternary Ammonium Compounds; D014168:Transferrin; D004247:DNA; D002784:Cholesterol; D005947:Glucose; C070046:1,2-dioleoyloxy-3-(trimethylammonium)propane