Transport-Metabolism Interplay of Atazanavir in Rat Hepatocytes

Abstract

The aim of this study was to explore the mechanisms governing the intra-to extracellular unbound concentration ratio (Kp u,u ) for the HIV protease inhibitor atazanavir (ATV) in rat hepatocytes. We had previously proposed a new method to determine Kp u,u by using the unbound Km values from metabolism studies with suspended rat hepatocytes and rat liver microsomes. Following that method, we determined that the value of ATV Kp u,u was 0.32, indicating that ATV hepatocellular clearance is uptake ratelimited. This hypothesis was supported by the linear correlation between Kp u,u and active uptake clearance (P = 0.04; R 2 =0.82) in the presence of increasing concentrations of the uptake transport inhibitor losartan. Moreover, in contrast to an expected increase of Kp u,u upon inhibition of ATV metabolism, a decrease of Kp u,u was observed, suggesting an increased impact of sinusoidal efflux. In summary, involvement of active uptake transport does not guarantee high intracellular accumulation; however, it has a key role in regulating intracellular drug concentrations and drug metabolism. These findings will help improve future in vitro-to-in vivo extrapolations and likewise physiologically based pharmacokinetic models.

Identification of novel inhibitors of rat Mrp3

“…Hepatocytes in suspension are known to be across the sinusoidal membrane (Bow et al, 2008;De Bruyn et al, 2011;Keemink et al, 2015;Nicolaï, 2015). The big advantage of using suspended hepatocytes over e.g.…”

Quantitative determination of colistin A/B and colistin methanesulfonate in biological samples using hydrophilic interaction chromatography tandem mass spectrometry

“…The oil‐spin method 17 was used for the hepatocyte uptake sample preparation with small modifications in rat and human hepatocytes. After the incubation, triplicate 200 μL aliquots were rapidly pipetted on top of an oil layer (82:18 silicon oil/mineral oil, 1.051 g/mL) above NaCl solution (8%, w/v) in 1.5‐mL test tubes.…”

PBPK Modeling as a Tool for Predicting and Understanding Intestinal Metabolism of Uridine 5′-Diphospho-glucuronosyltransferase Substrates

“…Atazanavir is a hepatic uptake transporter substrate [ 80 ], and the PBPK modeling for atazanavir did not explicitly incorporate the role of transporters for atazanavir. Importantly, Nicolai et al [ 82 ] studied the interplay of transporters and metabolism for atazanavir in rats in vitro, and found that involvement of active uptake transport did not cause high intracellular levels in this case. For atazanavir, they found that the ratio of unbound intracellular to extracellular concentration in hepatocytes was approximately 0.3, and likewise, the unbound Michaelis–Menten constant, K m,u , was >three-fold higher in hepatocytes than microsomes.…”