Lipid mixing between lipoplexes and plasma lipoproteins is a major barrier for intravenous transfection mediated by cationic lipids.
It has been previously shown that transfection activity of cationic liposome/DNA lipoplexes delivered systemically is drastically inhibited by lipoproteins (Tandia, B. M., Vandenbranden, M., Wattiez, R., Lakhdar, Z., Ruysschaert, J. M., and Elouahabi, A. (2003) Mol Ther. 8, 264-273). In this work, we have compared the binding/uptake and transfection activities of DOTAP (N-[1-(2,3-dioleoyloxy)propyl]-N,N,N-trimethylammonium chloride) and diC14-amidine (3-tetradecylamino-N-tert-butyl-N'-tetra-decylpropionamidine)-containing lipoplexes in the presence or absence of purified low density lipoproteins and high density lipoprotein. Binding/uptake of both lipoplexes by the mouse lung endothelial cell line was inhibited to a similar extent in the presence of lipoproteins. In contrast, transfection activity of diC14-amidine-containing lipoplexes was almost completely inhibited (approximately by 95%), whereas approximately 40% transfection activity of DOTAP-containing lipoplexes was preserved in the presence of lipoproteins. Interestingly, the ability of lipoproteins to inhibit the transfection efficiency of lipoplexes was well correlated with their ability to undergo lipid mixing with the cationic lipid bilayer as revealed by fluorescence resonance energy transfer assay. Incubation of lipoplexes with increased doses of lipoproteins resulted in enhanced lipid mixing and reduced transfection activity of the lipoplexes in mouse lung endothelial cells. The role of lipid mixing in transfection was further demonstrated using lipid-mixing inhibitor, lyso-phosphatidylcholine, or activator (dioleoylphosphatidylethanolamine). Incorporation of Lyso-PC into diC14-amidine-containing lipoplexes completely abolished their capacity to undergo lipid mixing with lipoproteins and allowed them to reach a high transfection efficiency in the presence of lipoproteins. On the other hand, the incorporation of dioleoylphosphatidylethanolamine into DOTAP/DNA lipoplex activated lipid mixing with the lipoproteins and was shown to be detrimental toward the transfection activity of these lipoplexes. Taken together, these results indicate that fusion of lipoplexes with lipoproteins is a limiting factor for in vivo transfection.
Tandia
Bouna-Moussa
BM
Center of Structural Biology and Bioinformatics, Laboratory of Structure and Function of Biological Membranes, Université Libre de Bruxelles, Campus Plaine CP 206/2, B-1050 Brussels, Belgium.
Lonez
Caroline
C
Vandenbranden
Michel
M
Ruysschaert
Jean-Marie
JM
Elouahabi
Abdelatif
A
D000818:Animals; D002152:Calorimetry, Differential Scanning; D002412:Cations; D004247:DNA / Q000378:metabolism; D003902:Detergents / Q000494:pharmacology; D004305:Dose-Response Relationship, Drug; D042783:Endothelial Cells / Q000378:metabolism; D005229:Fatty Acids, Monounsaturated / Q000737:chemistry; D031541:Fluorescence Resonance Energy Transfer; D018014:Gene Transfer Techniques; D008051:Lipid Bilayers / Q000737:chemistry; D050356:Lipid Metabolism; D008055:Lipids / Q000737:chemistry; D008074:Lipoproteins / Q000097:blood* / Q000737:chemistry*; D008075:Lipoproteins, HDL / Q000378:metabolism; D008081:Liposomes / Q000378:metabolism; D008168:Lung / Q000166:cytology; D008244:Lysophosphatidylcholines / Q000378:metabolism; D051379:Mice; D010714:Phosphatidylethanolamines / Q000378:metabolism; D010957:Plasmids / Q000378:metabolism; D011485:Protein Binding; D000644:Quaternary Ammonium Compounds / Q000737:chemistry; D013696:Temperature; D013997:Time Factors; D014162:Transfection; D014357:Trypsin / Q000494:pharmacology
D002412:Cations; D003902:Detergents; D005229:Fatty Acids, Monounsaturated; D008051:Lipid Bilayers; D008055:Lipids; D008074:Lipoproteins; D008075:Lipoproteins, HDL; D008081:Liposomes; D008244:Lysophosphatidylcholines; D010714:Phosphatidylethanolamines; D000644:Quaternary Ammonium Compounds; C094877:dioleoyl phosphatidylethanolamine; D004247:DNA; D014357:Trypsin; C070046:1,2-dioleoyloxy-3-(trimethylammonium)propane