Physiologically Based Pharmacokinetic Modeling of Transporter-Mediated Hepatic Clearance and Liver Partitioning of OATP and OCT Substrates in Cynomolgus Monkeys.

In the present investigations, we evaluate in vitro hepatocyte uptake and partitioning for the prediction of in vivo clearance and liver partitioning. Monkeys were intravenously co-dosed with rosuvastatin and bosentan, substrates of the organic anion transporting polypeptides (OATPs), and metformin, a substrate of organic cation transporter 1 (OCT1). Serial plasma and liver samples were collected over time. Liver and plasma unbound fraction was determined using equilibrium dialysis. In vivo unbound partitioning (Kpu,u) for rosuvastatin, bosentan, and metformin, calculated from total concentrations in the liver and plasma, were 243, 553, and 15, respectively. A physiologically based pharmacokinetic monkey model that incorporates active and passive hepatic uptake was developed to fit plasma and liver concentrations. In addition, a two-compartment model was used to fit in vitro hepatic uptake curves in suspended monkey hepatocyte to determine active uptake, passive diffusion, and intracellular unbound fraction parameters. At steady-state in the model, in vitro Kpu,u was determined. The results demonstrated that in vitro values under-predicted in vivo active uptake for rosuvastatin, bosentan, and metformin by 6.7-, 28-, and 1.5-fold, respectively, while passive diffusion was over-predicted. In vivo Kpu,u values were under-predicted from in vitro data by 30-, 79-, and 3-fold. In conclusion, active uptake and liver partitioning in monkeys for OATP substrates were greatly under-predicted from in vitro hepatocyte uptake, while OCT-mediated uptake and partitioning scaled reasonably well from in vitro, demonstrating substrate- and transporter-dependent scaling factors. The combination of in vitro experimental and modeling approaches proved useful for assessing prediction of in vivo intracellular partitioning.

Rosuvastatin Liver Partitioning in Cynomolgus Monkeys: Measurement In Vivo and Prediction Using In Vitro Monkey Hepatocyte Uptake.

Unbound plasma concentrations may not reflect those in target tissues, and there is a need for methods to predict tissue partitioning. Here, we investigate the unbound liver partitioning (Kpu,u) of rosuvastatin, a substrate of hepatic organic anion transporting peptides, in cynomolgus monkeys and compare it with that determined using hepatocytes in vitro. Rosuvastatin (3 mg/kg) was administered orally to monkeys and plasma and liver (by ultrasound-guided biopsy) collected over time. Uptake into monkey hepatocytes was evaluated up to steady state. Binding in monkey plasma, liver, and hepatocytes was determined using equilibrium dialysis. Mean in vivo Kpu,u was 118 after correcting total liver partitioning by plasma and liver binding. In vitro uptake data were analyzed by compartmental modeling to determine active uptake clearance, passive diffusion, the intracellular unbound fraction, and Kpu,u. In vitro Kpu,u underpredicted that in vivo, resulting in the need for an empirical in vitro to in vivo scaling factor of 10. Adjusting model parameters using hypothetical scaling factors for transporter expression and surface area or assuming no effect of protein binding on active transport increased partitioning values by 1.1-, 6-, and 9-fold, respectively. In conclusion, in vivo rosuvastatin unbound liver partitioning in monkeys was underpredicted using hepatocytes in vitro. Modeling approaches that allow integrating corrections from passive diffusion or protein binding on active uptake could improve the estimation of in vivo intracellular partitioning of this organic anion transporting peptide substrate. A similar assessment of other active hepatic transport mechanisms could confirm and determine the extent to which limited accumulation in isolated hepatocytes needs to be considered in drug development.

Serologic evaluation of clinical and subclinical secondary hepatic amyloidosis in rhesus macaques (Macaca mulatta).

Secondary hepatic amyloidosis in nonhuman primates carries a grave prognosis once animals become clinically ill. The purpose of this study was to establish serologic parameters that potentially could be used to identify rhesus macaques undergoing subclinical development of secondary hepatic amyloidosis. A retrospective analysis was completed by using serum biochemical profiles from 26 histologically diagnosed amyloidotic macaques evaluated at 2 stages of disease, clinical and subclinical (3 to 32 mo prior to clinical signs of disease). Standard serum biochemistry values for cases were compared with institutional age- and gender-specific references ranges by construction of 95% confidence intervals for the difference between means. In addition, 19 histologically diagnosed amyloidotic macaques and 19 age-matched controls were assayed for changes in various parameters by using routinely banked, frozen (-80 degrees C) sera available from clinical and subclinical time points. Clinically amyloidotic animals displayed increased levels of alkaline phosphatase, aspartate aminotransferase, lactate dehydrogenase, gamma glutamyltranspeptidase, and macrophage colony-stimulating factor and significantly decreased quantities of albumin and total cholesterol. Subclinical amyloidotic animals displayed increased levels of alkaline phosphatase, aspartate aminotransferase, lactate dehydrogenase, and serum amyloid A and decreased concentrations of albumin and total cholesterol. The serologic parameters studied indicate a temporal relationship of these factors not previously described, show a clear pattern of disease progression, and could be useful in subclinical disease detection.