Receptor-mediated disposition of polypeptides: Kinetic analysis of the transport of epidermal growth factor as a model peptide using in vitro isolated perfused organs and in vivo system

Abstract

The concept of “transport receptor” is now well established and receptor-mediated endocytosis (RME) is known as a general mechanism in the uptake of biologically important peptides and proteins. Using epidermal growth factor (EGF) as a model polypeptide, we have developed in vivo and in vitro methods to define the kinetic characteristics of RME. Kinetic analysis of the tissue distribution of EGF in rats performed in vivo clarified the following points. (1) The uptake of EGF by the liver, kidney and some other tissues showed clear saturation, which may represent RME. (2) The hepatic extraction ratio of EGF at a low dose was 0.8 and its uptake was limited mostly by the hepatic plasma flow rate. (3) In the intact animal, the bulk (ca. 50%) of the removal of EGF from the circulation was accounted for by hepatic clearance, followed by kidney clearance (ca. 15%). We then analyzed the hepatic and renal handling of EGF by isolated perfused organs, and the kinetic parameters have been obtained. The analysis clarified that, in the overall receptormediated processing of EGF by rat liver, the ligand-receptor association process is rapid (mean time < 1 min), the degradation process is much slower (mean time 4.5 h) and the receptor internalization process is intermediate (mean time 4.5 min). An internalization rate constant comparable with that in the liver was obtained also in the kidney. The extent of the receptor downregulation and the recovery rate of the available receptors were determined in several organs after the bulk of the receptors had been down-regulated by injecting excess EGF in vivo. The mean time required for the recovery showed inter-organ differences and the value (ca. 30 min) in the liver was much shorter than those (2–5 h) in other organs. Such recovery was considered at least in the liver to reflect the processes of recycling of internalized EGF receptors to the cell surface. Finally, time profiles of EGF plasma concentrations and their surface receptors in the liver were simulated with a physiological pharmacokinetic model.