Optimization of cationic liposome/DNA complexes for in vivo applications:: Correlation between transfection activity and morphology

The morphology and transfection activity of cationic liposome-DNA complexes (CLDC) have been investigated to establish structure-function relationships relevant for high transfection activities under both in vivo and in vitro conditions. The effect of the helper Lipid (Cholesterol [Chol] versus DOPE), the influence of lipid components providing steric stabilization such as PEG-PE, as well as pre-condensation of plasmid DNA by spermidine were studied on both parameters of CLDC, transfection activity and morphology. The in vitro studies were performed an SK-BR-3 cells in the presence of serum-containing media and the in vivo studies were carried out in mice following i.v, injection. The morphology of the CLDC was monitored by freeze-fracture electron microscopy after mixing dth tell growth medium or with mouse serum respectively. Complexes containing DOPE rather than Chol as helper lipid precipitate in the present of cell medium as well as serum and convert into hexagonal lipid (H-n) phase. Despite their high transfection activity in vitro, such complexes show wry little transfection activity in vivo. Substitution of DOPE with Chol, and the addition of PEG-PE are stabilizing CLDC with respect to time and scram. Such complexes show high expression of a marker gene in mouse lungs, but relatively low expression in SK-BR-3 cells. Additionally, some of the complexes containing pre-condensed DNA look like "map pins" showing heads of the size of liposomes and short, stiff and tapering tails. Their in vivo transfection activity is highest. These comparisons raveal a fundamental difference between in vitro and in vivo activity of CLDC: High in vitro transfection activity seems to be associated with hexagonal lipid precipitates whereas high in vivo activity seems to be related with small, stabilized complexes, which additionally exhibit some "map-pin" structures.