Intermembrane transfer of polyethylene glycol-modified phosphatidylethanolamine as a means to reveal surface-associated binding ligands on liposomes

In order to explore the use of exchangeable poly(ethylene glycol) (PEG)-mod ified diacylphosphatidylethailolamines (PE) to temporarily shield binding l igands attached to the surface of liposomes, a model reaction based on inhi bition and subsequent recovery of biotinylated liposome binding to streptav idin immobilized on superparamagnetic iron oxide particles (SA magnetic par ticles) was developed. PEG-lipid incorporation into biotinylated liposomes decreased liposome binding to SA magnetic particles in a non-linear fashion , where as little as 0.1 mol% PEG-PE resulted in a 20% decrease in binding. Using an assay based on inhibition of binding, PEG(2000)-PE transfer from donor liposomes to biotinylated acceptor liposomes could be measured. The i nfluence of temperature and acyl chain composition on the transfer of PEG-d iacyl PEs from donor liposomes to acceptor liposomes, consisting of 1,2-dio leoyl-sn-glycero-3-phosphocholine. cholesterol and N-(6-biotinoyl)amino)hex anoyl)-1,2-distearoyl-sn-glycero-3-phosphoethanolamine (54.9:45:0.1 mole ra tio), was measured. Donor liposomes were prepared using 1,2-distearoyl-sn-g lycero-3-phosphocholine (50 mol%), cholesterol (45 mol%) and 5 mol%, of eit her PEG-derivatized 1,2-dimyristoyl-sn-glycero-3-phosphoethanolamine (DMPE- PEG(2000)), 1,2-dipalmitoyl-sn-glycero-3-phosphoethanolamine (DPPE-PEG(2000 ), or 1,2-distearoyl-sn-glycero-3-phosphoethanolamine (DSPE-PEG(2000). Tran sfer of DSPE-PEG(2000) to the donor liposomes was not detected under the co nditions employed. In contrast, DMPE-PEG(2000) was transferred efficiently even at 4 degreesC. Using an acceptor to donor liposome ratio of 1:4. the t ime required for DMPE-PEG(2000) to become evenly distributed between the tw o liposome populations (T-EQ) at 4 degreesC and 37 degreesC was approx. 2 a nd < 0.5 h, respectively. An increase in acyl chain length from C14:0 to C1 6:0 of the PEG-lipid resulted in a significant reduction in the rate of tra nsfer as measured by this assay. The transfer of PEG-lipid out of biotinyla ted liposomes was also studied in mice following intravenous administration . The relative rates of transfer for the various PEG-lipids were found to b e comparable under in vivo and in vitro conditions. These results suggest t hat it is possible to design targeted liposomes with the targeting ligand p rotected while in the circulation through the use of PEG-lipids that are se lected on the basis of exchange.