FACTORS INFLUENCING THE RETENTION AND CHEMICAL-STABILITY OF POLY(ETHYLENE GLYCOL)-LIPID CONJUGATES INCORPORATED INTO LARGE UNILAMELLAR VESICLES

Poly(ethylene glycol)(PEG)-lipid anchor conjugates can prolong the cir culation lifetimes of liposomes following intravenous injection. In th is work we investigate the influence of the lipid anchor and the natur e of the chemical link between the PEG and lipid moieties on circulati on life time. It is shown th at incorporation of N-(monomethoxypoly(et hylene inyl)-1-palmitoyl-2-oleoylphosphatidylethanolamide (MePEG(2000) -S-POPE) into large unilamellar vesicles (LUVs) composed of distearoyl phosphatidylcholine (DSPC) and cholesterol (DSPC/cholesterol/MePEG(200 0)-S-POPE, 50:45:5, mol/mol) results in only small increases in the ci rculation lifetimes as observed in mice. This is shown to be due to ra pid removal of the hydrophilic coating in vivo, which likely arises fr om exchange of the entire PEG-lipid conjugate from the liposomal membr ane, although chemical breakdown of the PEG-lipid conjugate is also po ssible. The chemical stability of four different linkages was tested, including succinate, carbamate and amide linkages between MePEG deriva tives and the amino head group of PE, as well as a direct link to the phosphate head group of phosphatidic acid (PA). The succinate linkage was found to be the most labile. The anchoring capability of DSPE as c ompared to POPE in PEG-PE conjugates was also examined. It is shown th at incorporation of MePEG(2000)-S-DSPE conjugates into DSPC/cholestero l LUVs results in little loss of the PEG coating in vivo, long circula tion lifetimes and reduced chemical breakdown of the PEG-lipid conjuga te. This work establishes that DSPE is a considerably more effective a nchor for PEG(2000) than POPE and that the chemical stability of PEG-P E conjugates is sensitive to the nature of the linkage and exchangeabi lity of the PEG-PE complex. We suggest that retention of the PEG coati ng is of paramount importance for prolonged circulation lifetimes.