Targeting Liposomes Through Immunoglobulin Superfamily Domains: P0 Protein as a Model

Abstract

The objective of this study was to evaluate the potential of P0 protein, a cell adhesion molecule from peripheral nerve myelin, as a targeting ligand for liposomes. To evaluate binding characteristics and identify possible binding domains, cell-interaction studies were carried out with P0 protein reconstituted into liposomes (P0 liposomes) under various conditions. P0 liposomes with intact P0 protein were tested after endoglycosidase F treatment (cleaves the carbohydrate moiety) or trypsin digestion (removes the hydrophobic portion, residues 1-79, leaving the carbohydrate portion intact). The cellular uptake was quantitated using radioactive lipids in the liposome bilayer and a liposome-entrapped water soluble compound (inulin). The presence of intact P0 protein in the liposome bilayer increased the rate of interaction of liposomes 3-4 times with M21 melanoma and HTB-11 neuroblastoma cells (cells of neuroectodermal origin), two times with Caki-1 renal carcinoma cells, and marginally with 3T3 fibroblasts (mesodermal origin). This binding was inhibited by anti-chick P0 antibodies and Fab fragments. A control transmembrane glycoprotein, glycophorin A., when reconstituted in liposomes had no effect on the binding of liposomes with M21 cells. The results indicate that P0 protein plays a specific role in the binding of liposomes to cells. Removal of the N-asparagine linked carbohydrate from the P0 protein in the liposomes resulted in an increase of their association with M21 cells five times that of control liposomes (no protein) and two times that of the non-endoglycosidase F-treated P0 liposomes. To further characterize the binding domains of P0 reconstituted into liposomes, competition studies were carried out in the presence of synthetic P0-peptides. The competition studies indicated that both the extracellular (residues 90-96) and intracellular (residues 201-207) domain of P0 protein may be involved in the interaction with cell membranes. The results suggest that P0 protein is capable of mediating specific heterophilic interactions with various cell lines and targeting to cells of neuroectodermal origin may be achieved.