Immunogenicity and pharmacokinetic attributes of poly(ethylene glycol)-grafted immunoliposomes.

Immunoliposomes composed of hydrogenated soy phosphatidylcholine, cholesterol, methoxypoly(ethylene glycol)-distearoyl phosphatidylethanolamine (mPEG-DSPE), and hydrazide-PEG-DSPE (mole ratio, 57:38:3.3:1.7) linked to periodate-oxidized chimerized mouse IgG (C225, anti-human epidermal growth factor receptor) were prepared by an optimized aggregation-free procedure. The antigen-binding activity of the immunoliposomes was well preserved. When injected intravenously into naive rats, the immunoliposomes (approximately 18 IgG per 100 nm liposome) exhibited long circulation times (MRT = 8.5 h, Cl = 0.2 ml/h). Subsequent injections of the immunoliposomes into the same animals resulted in rapid clearance (MRT < or = 0.7 h, Cl > or = 7 ml/h), which was accompanied by a significant increase in anti-C225 specific titers. Upon repeated injection or coinjection with the parent liposomes free C225 consistently exhibited prolonged circulation without any increase in C225-specific antisera, but was cleared quickly when administered into animals that had been pretreated with the immunoliposomes. Screening of the immunoliposome induced antisera against human polyclonal IgG and C225-derived Fab' fragment revealed that the immune response was specifically triggered by the constant human region of C225. These results demonstrate that the preparations of PEG-grafted immunoliposomes are more immunogenic than the free IgG component, which is of profound importance to the antibody-mediated liposomal drug delivery effort.

Peptide attachment to extremities of liposomal surface grafted PEG chains: preparation of the long-circulating form of laminin pentapeptide, YIGSR.

Poly(ethylene glycol) (PEG)-grafted liposomes offer new opportunities as long-circulating platforms presenting biologically relevant ligands. In pursuit of this goal, liposomal conjugates of YIGSR were prepared by mild periodate oxidation of TYIGSR-NH2 and incubation of the product with hydrazide-PEG-(distearoylphosphatidyl)ethanolamine-containing liposomes. The peptide-carrying liposomes, with up to 500 YIGSR residues per vesicle, despite exhibiting faster blood clearance rates than the parent liposomes in rats, remained in circulation for extended periods of time. Mean residence times for the parent liposomal formulation and conjugated preparations containing 200 and 500 YIGSR residues per vesicle were 28, 25, and 23 h, respectively. The results have important implications for systemic delivery of peptides and for their use as targeting moieties for PEG-grafted liposomes.

New amphipatic polymer-lipid conjugates forming long-circulating reticuloendothelial system-evading liposomes.

Lipid-conjugates of two amphipatic polymers, poly(2-methyl-2-oxazoline) (PMOZ) and poly(2-ethyl-2-oxazoline) (PEOZ) (degree of polymerization approximately 50) were synthesized by linking glutarate esters of the polymers to distearoylphosphatidylethanolamine (DSPE) or alternatively by termination of the polymerization process with DSPE. Surface-modified liposomes (90 +/- 5 nm) prepared from either conjugate (5 mol % of total lipid) were injected into rats and followed by blood level and tissue distribution measurements. Both polymers PEOZ and PMOZ were found to convey long circulation and low hepatosplenic uptake to liposomes to the same extent as polyethylene glycol (PEG), the best known material for this purpose. This is the first demonstration of protection from rapid recognition and clearance conveyed by alternative polymers, which is equal to the effect of PEG.