Membrane perturbation and the mechanism of lipid-mediated transfer of DNA into cells

Mixtures of cationic lipids and unsaturated phosphatidylethanolamine are us ed extensively for the intracellular delivery of plasmids and antisense oli godeoxynucleotides (ODN) in vitro. However, the mechanism by which cytoplas mic delivery of these large molecules is achieved remains unclear. The comm on hypothesis is that phosphatidylethanolamine promotes fusion of lipid/DNA particles with endosomal membranes, but this is inconsistent with several reports that have failed to correlate the fusogenic activity of a wide vari ety of lipid/DNA particles, measured by lipid mixing techniques, with their transfection activity. To address this issue further we have conducted a d etailed analysis of the lipid mixing and DNA transfer activity of two, phys ically similar but functionally different, lipid/DNA particles composed of equimolar dioleyldimethylammonium chloride (DODAC) and dioleoylphosphatidyl ethanolamine (DOPE) or dioleoylphosphatidylcholine (DOPC). In combination w ith DODAC both phospholipids form almost identical lipid/DNA particles, the y are endocytosed by cells to the same extent and each undergoes equivalent lipid mixing with cell membranes after uptake. Despite this, DNA transfer is 10- to 100-fold more extensive for lipid/DNA particles containing DOPE. We conclude that lipid mixing between lipid-based delivery systems and endo somal membranes must occur for DNA transfer to occur. However, the potency of different lipid/DNA particles correlates better with the ability of the exogenous lipid to disrupt membrane integrity. (C) 2000 Elsevier Science B. V. All rights reserved.