Structural Stability against Disintegration by Anionic Lipids Rationalizes the Efficiency of Cationic Liposome/DNA Complexes

Abstract

Reported here is the correlation between the transfection efficiency of cationic liposome/DNA complexes (lipoplexes) and the structural evolution that they undergo when interacting with anionic membrane lipids. Multicomponent lipoplexes, incorporating from three to six lipid species simultaneously, presented a much higher transfection efficiency than binary lipoplexes, which are more commonly used for gene-delivery purposes. The discovery that a high transfection efficiency can be achieved by employing multicomponent complexes at a lower-than-ever-before membrane charge density of lipoplexes was of primary significance. Synchrotron small-angle X-ray diffraction (SAXD) experiments showed that anionic liposomes made of dioleoylphosphatidylglycerol (DOPG) disintegrated the lamellar phase of lipoplexes. DNA unbinding was measured by electrophoresis on agarose gels. Most importantly, structural changes induced by anionic lipids strictly depended on the lipid composition of lipoplexes. We found evidence of the existence of three different regimes of stability related to the interaction between complexes and anionic membranes. Both unstable (with low membrane charge density, sigmaM) and highly stable lipoplexes (with high sigmaM) exhibited low transfection efficiency whereas highly efficient multicomponent lipoplexes exhibited an "optimal stability". This intermediate regime reflects a compromise between two opposing constraints: protection of DNA in the cytosol and endosomal escape. Here we advance the concept that structural stability, upon interaction with cellular anionic lipids, is a key factor governing the transfection efficiency of lipoplexes. Possible molecular mechanisms underlying experimental observations are also discussed.

Factors Determining the Superior Performance of Lipid/DNA/Protammine Nanoparticles over Lipoplexes

“…Insight into parameters that determine stabilization and destabilization of these complexes, which are intricately related to their efficiency in gene delivery, requires an understanding of factors that govern amphiphileÀDNA interaction and subsequent complex formation. Upon complex formation, reentrant condensation and charge inversion 20,21 occurred in both systems (Figure 3). Both charge inversion and charge and size saturation of LPD complexes were found to occur at R V values smaller than those observed when lipoplexes were used.…”

The interdigitated gel phase in mixtures of cationic and zwitterionic phospholipids

“…Cationic lipids have attracted a great amount of interest for their potential as DNA transfecting lipoplexes [10,29,30], transfection reagents for RNA [31,32], co-adjuvants for vaccines [33,34] and as facilitators for drug delivery [35]. Ethyl-PC lipids are of particular interest due to their relatively low toxicity, which is likely a result of their similarity to cellular lipids and because they can be metabolized [28,36].…”

Lipid rafts-mediated endocytosis and physiology-based cell membrane traffic models of doxorubicin liposomes

“…The unsaturated DOPE lipid promotes the fusion between liposomes and cell membrane, while the saturated DPPC lipid protects liposomes from disintegration by cellular lipids [22]. We hypothesized that if charge density caused by DOTAP or DDAB of liposome is a more important factor then substituting the neutral lipids (DPPC or DOPE) would not affect cellular uptake of Dox loaded liposomes, whereas if fusogenic properties are more important the DOPE formulations would be superior to the DPPC formulations.…”

“…DDAB and DOTAP contribute positive surface charge of liposomes. DOPE promotes the fusion of liposomes and cell/endosome membranes, while DPPC promotes more stable laminar structures [18,21,22]. PEG-DSPE was used in some formulations to achieve surface pegylation.…”