Experimental and Mathematical Analysis of Hepatic Uptake

Introduction: To investigate the nonlinear kinetics of in vitro hepatic uptake the OATP substrate, Pitivastatin, was used as a probe. Experiments were conducted using freshly isolated rat hepatocytes, utilising the ‘oil spin’ methodology described by Hassen et al [1]. Briefly, freshly isolated rat hepatocytes were incubated with Pitvastatin (5–300μM). At 10 s, 30 s, 50 s and 70 s aliquots were spun through a silicone oil layer to separate the hepatocytes from the media. [Pitivastatin]hepatocyte was detemined using LCMSMS. Results: Uptake to rat hepatocytes was saturable and progressed according to Michaelis-Menten kinetics. The Km and Vmax of Pitvastatin were 2050 pmols/min/10 cells and 33 μM respectively, which was in in good agreement with other literature reports [2]. Mathematical Modelling: A nonlinear pharmacokinetic model has been derived to characterise the uptake process. A structural identifiablity analysis was performed on the model to establish that all unknown parameters could be identified from the experimental observations available. The model was then subsequently used for parameter estimation and model validation using the data collected. Sensitivity analysis and model robustness analyses were also performed. Once fully validated the model has the potential to perform robust, predictive simulations to ascertain optimal levels of uptake and the effects of the use of appropriate inhibitors. This work was supported by AZ & TG was supported by AZ and the MRC Capacity Building initiative.