Purpose: The purpose of this study was to assess whether LLC-PK1 renal epithelial cells could serve as an in vitro model for studying the renal tubular reabsorption of protein drugs.
Methods: The association of 111In-labeled model protein drugs, bovine serum albumin (BSA), superoxide dismutase (SOD), soybean trypsin inhibitor (STI), and [Asu1.7]-eel calcitonin (Asu-ECT), with the monolayers of LLC-PK1 renal epithelial cells was characterized under various conditions.
Results: The cellular association of these proteins was temperature-dependent and varied according to the protein. Saturation kinetics were observed for STI association, with the apparent Km and Vmax values determined to be 66.3 micrograms/ml and 250 ng/mg protein/min, respectively. The association of STI decreased with increases in medium pH from 5.4 to 8.4 and was inhibited significantly by 2,4-dinitrophenol, sodium azide, cytochalasin B, and colchicine, suggesting that the cellular association involved endocytosis. Mutual inhibition was observed in competitive binding experiments with the four protein drugs, suggesting that they shared a common binding site on the luminal membrane of LLC-PK1 cells. Taken together, these findings show that a variety of protein drugs bind to LLC-PK1 cells in a non-specific manner and possibly undergo endocytosis, a phenomenon that is similar to in vivo proximal tubular reabsorption.
Conclusions: LLC-PK1 renal epithelial cells would be a suitable model system for the study of the renal proximal tubular reabsorption of protein drugs.