Evaluation of Hepatic Uptake of OATP1B Substrates by Short Term-Cultured Plated Human Hepatocytes: Comparison With Isolated Suspended Hepatocytes

Abstract

Hepatic uptake clearance has been measured in suspended human hepatocytes (SHH). Plated human hepatocytes (PHH) after short-term culturing are increasingly employed to study hepatic transport driven mainly by its higher throughput. To know pros/cons of both systems, the hepatic uptake clearances of several organic anion transporting polypeptide 1B substrates were compared between PHH and SHH by determining the initial uptake velocities or through dynamic model-based analyses. For cerivastatin, pitavastatin and rosuvastatin, initial uptake clearances (PS inf) obtained using PHH were comparable to those using SHH, while cell-to-medium concentration (C/M) ratios were 2.7-to 5.4-fold higher. For pravastatin and dehydropravastatin, hydrophilic compounds with low uptake/cellular binding, their PS inf and C/M ratio in PHH were 1.8-to 3.2-fold lower than those in SHH. These hydrophilic substrates are more prone to wash-off during the uptake study using PHH, which may explain the apparently lower uptake than SHH. The C/M ratios obtained using PHH were stable over an extended time, making PHH suitable to estimate the C/M ratios and hepatocyte-to-medium unbound concentration ratios (K p,uu). In conclusion, PHH is useful in evaluating hepatic uptake/efflux clearances and K p,uu of OATP1B substrates in a high-throughput manner, however, a caution is warranted for hydrophilic drugs with low uptake/cellular binding.

Advances in Animal Models and Cutting-Edge Research in Alternatives: Proceedings of the Third International Conference on 3Rs Research and Progress, Vishakhapatnam, 2022

“…Thus, 999 Elite and MetMax Cryopreserved Human Hepatocytes can be routinely used for the evaluation of human-specific drug properties during drug development. 40–45…”

“…Thus, 999 Elite and MetMax Cryopreserved Human Hepatocytes can be routinely used for the evaluation of human-specific drug properties during drug development. [40][41][42][43][44][45] 3-D bioprinted liver sinusoidal model: Focusing on the metabolic zonation Dr Falguni Pati (Associate Professor, Department of Biomedical Engineering, IIT Hyderabad, India) described the development and design of a 3-D printed mini bioreactor that mimics the arrangement of hepatocytes and the sinusoidal network in vivo. He touched on the fact that the liver is the largest organ in the human body, performing a wide range of metabolic functions, and that it is the primary organ involved in drug metabolism.…”

The Next Frontier in ADME Science: Predicting Transporter-Based Drug Disposition, Tissue Concentrations and Drug-Drug Interactions in Humans

“…Kumar, Yin, et al, 2020;Miyauchi et al, 2018;Sachar et al, 2020;Watanabe et al, 2011). Both in-house (unpublished) and published data suggest that the number of washes can have impact on the estimate of drug uptake (Yoshikado et al, 2021). Also, to reduce non-specific binding of the drug, some add bovine serum albumin to the washing buffer (Niessen et al, 2009).…”

“…Instead, the transport assay is terminated by rapidly separating the cells from the incubation medium by passaging them through an oil layer. However, this method does not necessarily reduce non-specific binding of highly lipophilic drugs as any drug adsorbed to the cell surface could also passage into the oil layer (Yoshikado et al, 2021). To improve solubility of lipophilic drugs in the uptake buffer, organic solvents are often used (e.g., dimethyl sulfoxide).…”

Including active excretion in physiologically based kinetic models for new approach methodologies

“…For biliary excretion (Chapter 2), the in vitro kinetic model used in the present thesis consisted of freshly isolated primary hepatocytes either cultivated in suspension or in monolayer (Brock and Vore 1984;Brouwer et al 1987;Ishizuka et al 1998;Kouzuki et al 1999). This model is well established and widely used to obtain especially metabolism data but is also frequently applied to obtain transport data (Louisse et al 2020;Yoshikado et al 2021). Advantages of using this primary hepatocyte model is that it is close to the real liver, and still contains all the elements of a hepatocyte including the expression and functioning of the transporters.…”