Comparison of Methods for Estimating Unbound Intracellular-to-Medium Concentration Ratios in Rat and Human Hepatocytes Using Statins

摘要

It is essential to estimate concentrations of unbound drugs inside the hepatocytes to predict hepatic clearance, efficacy, and toxicity of the drugs. The present study was undertaken to compare predictability of the unbound hepatocyte-to-medium concentration ratios (K-p,K-uu) by two methods based on the steady-state cell-to-mediumtotal concentration ratios at 37 degrees C and on ice (K-p,K- uu,K- ss) and based on their initial uptake rates (K-p,K-uu,K-Vo). Poorly metabolized statins were used as test drugs because of their concentrative uptake via organic anion-transporting polypeptides. K-p,K- uu,K- ss values of these statins provided less interexperimental variation than the K-p,K-uu,K-V0 values, because only data at longer time are required for K-p,K- uu,K- ss. K-p,K- uu,K- V0 values for pitavastatin, rosuvastatin, and pravastatin were 1.2-to 5.1-fold K-p,K- uu,K- ss in rat hepatocytes; K-p,K- uu,K- V0 values in human hepatocytes also tended to be larger than corresponding K-p,K- uu,K- ss. To explain these discrepancies, theoretical values of (Kp, uu, ss) and K-p,K- uu,K- V0 were compared with true K-p,K- uu (K-p,K- uu, true), considering the insidenegative membrane potential and ionization of the drugs in hepatocytes and medium. Membrane potentials were approximately 230 mV in human hepatocytes at 37 degrees C and almost abolished on ice. Theoretical equations considering the membrane potentials indicate that K-p,K- uu,K- ss values for the statins are 0.85-to 1.2-fold K-p,K- uu, true, whereas K-p,K- uu,K- V0 values are 2.2-to 3.1-fold K-p,K-uu, true, depending on the ratio of the passive permeability of the ionized to nonionized forms. In conclusion, K-p,K-uu, ss values of anions are similar to K-p,K- uu, true when the inside-negative membrane potential is considered. This suggests that K-p,K- uu,K- ss is preferable for estimating the concentration of unbound drugs inside the hepatocytes.