Asialoglycoprotein/asialoglycoprotein receptor (AGP-AGPr) interaction is an important mechanism for the uptake of FK506 by hepatocytes

Abstract

Hepatic tissue concentrations of FK506 have been correlated with early acute rejection following liver transplantation. Asialoglycoproteins (AGP) reputedly bind FK506 in blood. AGP are removed from the circulation by the liver via the AGP receptor (AGPr), which resides on hepatocytes. This study was undertaken to determine if the AGP-AGPr mechanism enhances the delivery of FK506 to hepatocytes. Human orosomucoid (OM) was used as a representative AGP. asialoOM (aOM) was prepared by desialation of OM. Fresh rat hepatocytes were isolated by collagenase digestion. Tritium labeled FK506 (FK) was used to identify and quantitate FK506. Quantitation of FK in serum and culture media was by direct counting. FK in animal tissues used a method developed in our laboratory for the purpose. AGPr on resting hepatocytes was demonstrated by flow cytometry using FITC-orosomucoid and FITC-BSA controls. AGPr were enhanced by 2 g glucose/L. Two serum FK-binding fractions, 44 kD and 15 kD, were identified by gel filtration. Exogenous OM avidly bound FK and displaced FK activity from the 15 kD fraction. Serum (1%) and the 44 kD fraction enhanced the uptake of FK by hepatocytes, while serum depleted of OM-aOM by affinity chromatography was only 72.5% as effective as control serum; aOM enhanced the uptake of FK by hepatocytes to a degree similar to that of control serum but OM did not significantly affect the uptake of FK. Cold FK506 blocked the uptake and was dose dependent; cold CsA had no effect. Affinity extraction of OM from serum to which FK had previously been added removed 28.4% of FK activity. Following i.v. infusion, the kidney had the highest and liver the lowest tissue concentration of FK at 1 hr and 3 hr. In contrast, after oral administration the liver had the highest concentrations of the other tissues tested. The AGP-AGPr mechanism plays a significant role in the delivery of FK506 to hepatocytes and is likely responsible for the differences in bioavailability observed after oral and i.v. administration. Factors governing the AGP-AGPr mechanism are germane to understanding both the efficacy and toxicity of FK506 and the development optimal therapeutic strategies.