Stabilization and regulated fusion of liposomes containing a cationic lipid using amphipathic polyethyleneglycol derivatives

Current fusogenic liposomal delivery systems have Limited applicability in vivo due to poor stability in the blood and rapid clearance from the circulation. This is particularly true for liposomes composed of dioleoylphosphatidylethanolamine (DOPE) and cationic lipids, currently being developed for systemic delivery of gene-based drugs. This paper describes a potential strategy to overcome these problems, involving the incorporation of exchangeable amphipathic polyethyleneglycol (PEG) derivatives to transiently stabilize fusogenic liposomes while in the circulation, but where the PEG coating dissipates to reveal fusogenic character at later times after arrival at target sites. It is shown here that large unilamellar vesicles (LUVs) containing DOPE and the cationic lipid, N,N-dioleoyl-N,N-dimethylammonium chloride (DODAC) can be stabilized against serum-induced aggregation and fusion by inclusion of at least 2 mol% of PEG coupled to phosphatidylethanolamine (PEG-PE) or ceramide (PEG-Cer). However, low in vitro recovery of fusogenic activity was obtained for the PEG-PE-containing system, presumably due to electrostatic interactions between the negatively charged PEG-PE and the cationic lipid which prevent PEG-PE dissociation from the LUV. Improved recovery of fusogenic activity was achieved for LUVs stabilized by the neutral PEG-Cer derivatives, with shorter chain ceramides exhibiting more rapid recovery rates. Biodistribution studies showed that DOPE/DODAC (85:15, mol/mol) LUVs were rapidly cleared from the circulation, whereas inclusion of 10 mol% PEG-Cer(C-20) resulted in significantly prolonged circulation time. Inclusion of shorter ceramide chain lengths resulted in decreased circulation times, consistent with increased exchangeability. These findings demonstrate the feasibility of developing a cationic liposome that is stable in the circulation, but retains its ability to fuse with membranes. This work represents the first step toward the rationale design of fusogenic cationic liposomes for the systemic delivery of gene-based drugs to target tissues, such as tumors.