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Saturable Uptake of Lipophilic Amine Drugs into Isolated Hepatocytes: Mechanisms and Consequences for Quantitative Clearance Prediction

Centre for Applied Pharmacokinetic Research, School of Pharmacy and Pharmaceutical Sciences, University of Manchester, United Kingdom

The hepatic uptake of quinine, fluvoxamine, and fluoxetine (0.1–10 µM) was investigated with freshly isolated rat hepatocytes. The cell-to-medium concentration ratios (K_p) were concentration-dependent: the mean maximum K_p values (at 0.1 µM) were 150 (quinine), 500 (fluvoxamine), and 2000 (fluoxetine). There was also a large capacity site that was not saturable over the concentration range used (possibly partition into the phospholipid component of membranes); representing this site, the mean minimum K_p values (at 10 µM) were 30 (quinine), 200 (fluvoxamine), and 500 (fluoxetine). To eliminate concomitant metabolism, cells were pretreated with the irreversible P450 inhibitor, aminobenzotriazole. The saturable uptake was substantially eliminated after exposure to carbonyl cyanide 4-(trifluoromethoxy)phenylhydrazone (ATP inhibitor). The difference between the maximum and minimum K_p for these three amine drugs, as well as for dextromethorphan, propranolol, and imipramine, was within a limited range of 3-fold, indicating a common magnitude of saturable uptake. Basic, permeable drugs are expected to be sequestered into lysosomes, which actively maintain their low internal pH (5) using ATP, and this process is predictable from the combined effects of pH-driven ion accumulation and unsaturable binding representing partition into membranes. The resultant predicted maximum K_p correlated strongly with the observed maximum K_p. Thus, at low substrate concentrations, the fraction of drug unbound in the hepatocyte incubation (critical for assessing drug clearance and drug-drug interaction potential) may be dependent upon saturable as well as unsaturable binding, and for lipophilic, basic drugs, this can be readily estimated assuming a common degree of uptake into lysosomes.

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