Equilibrium and Kinetic Studies of the Binding of DNA to Cationic Liposomes

The objective of this work was to study the formation of DNA-cationic vesicle complexes (lipoplexes). These complexes are interesting not only from a fundamental point of view but also as promising candidates for gene therapy. The approach taken was to study first the cationic vesicles by themselves. This will allow us to better understand the "complexation mechanism" upon the addition of DNA. The effects of vesicle preparation method, concentration and counterion of the amphiphiles, and additives on vesicle formation and physical stability were examined. Addition of DNA to cationic vesicles (or vice versa) leads to the spontaneous formation of complexes. The structure of these DNA-cationic vesicles in dilute solutions used in this study is the same as in more concentrated samples (precipitate). However the lipoplexes in solution are not stable against aggregation and precipitation occurred within days. Calorimetric results show that the interaction between DNA and cationic vesicles is entropically driven and that the interaction between DNA and cationic vesicles can be correlated with the lipid composition of the cationic lipid bilayer. Measurements made in other polyelectrolyte-cationic vesicle systems, either by changing the lipids or the polyelectrolyte suggest that the observed energetics is a general feature occurring in the formation of complexes made by double-chain lipids and polyelectrolytes. The transient properties upon combining vesicles with DNA to form lipoplexes were addressed using time-resolved techniques: turbidity, fluorescence, small-angle neutron scattering. The formation of the lipoplexes occurred in 3 steps. The first step was the coverage of one DNA strand with one single vesicle. This interaction is strong enough to rupture the vesicle upon contact of the cationic vesicle with a DNA molecule. This intermediate with locally cylindrical structure appears within seconds. Association of previously ruptured vesicles leads to continue aggregation and growth of DNA-cationic vesicle complexes into a multilamellar structure. The knowledge of the time-dependent structural changes may contribute to future developments and biological applications of DNA-cationic vesicle complexes