Polarized location of SLC and ABC drug transporters in monolayer-cultured human hepatocytes

Vée, Marc Le; Jouan, Elodie; Noel, G. R.; Stieger, Bruno; Fardel, Olivier

doi:10.1016/j.tiv.2015.03.019

Cited by 25 publications

(12 citation statements)

References 53 publications

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“…As shown in Figure 1 a, SLC transporters were very poorly expressed in HuH-7 cells comparatively to hepatocytes. Indeed, HuH-7 cells failed to express the sinusoidal uptake transporters NTCP, OATP1B1 ( SLCO1B1 ), OAT2 ( SLC22A7 ) and OCT1, which, by contrast, were highly expressed in human hepatocytes, in agreement with previous data [ 45 ]. OATP1A2 ( SLCO1A2 ) mRNAs were also not detected in HuH-7 cells, knowing, however, that they were similarly not present in human hepatocytes ( Figure 1 a).…”

Section: Resultssupporting

confidence: 91%

Drug Transporter Expression and Activity in Human Hepatoma HuH-7 Cells

et al. 2016

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Human hepatoma cells may represent a valuable alternative to the use of human hepatocytes for studying hepatic drug transporters, which is now a regulatory issue during drug development. In the present work, we have characterized hepatic drug transporter expression, activity and regulation in human hepatoma HuH-7 cells, in order to determine the potential relevance of these cells for drug transport assays. HuH-7 cells displayed notable multidrug resistance-associated protein (MRP) activity, presumed to reflect expression of various hepatic MRPs, including MRP2. By contrast, they failed to display functional activities of the uptake transporters sodium taurocholate co-transporting polypeptide (NTCP), organic anion-transporting polypeptides (OATPs) and organic cation transporter 1 (OCT1), and of the canalicular transporters P-glycoprotein and breast cancer resistance protein (BCRP). Concomitantly, mRNA expressions of various sinusoidal and canalicular hepatic drug transporters were not detected (NTCP, OATP1B1, organic anion transporter 2 (OAT2), OCT1 and bile salt export pump) or were found to be lower (OATP1B3, OATP2B1, multidrug and toxin extrusion protein 1, BCRP and MRP3) in hepatoma HuH-7 cells than those found in human hepatocytes, whereas other transporters such as OAT7, MRP4 and MRP5 were up-regulated. HuH-7 cells additionally exhibited farnesoid X receptor (FXR)- and nuclear factor erythroid 2-related factor 2 (Nrf2)-related up-regulation of some transporters. Such data indicate that HuH-7 cells, although expressing rather poorly some main hepatic drug transporters, may be useful for investigating interactions of drugs with MRPs, notably MRP2, and for studying FXR- or Nrf2-mediated gene regulation.

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Section: Resultssupporting

confidence: 91%

Drug Transporter Expression and Activity in Human Hepatoma HuH-7 Cells

et al. 2016

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“…Relative quantification of the steady-state target mRNA levels was calculated after normalization of the total amount of cDNA tested to the 18S RNA endogenous reference using the 2 (−Δ C t) method. Data were finally expressed in arbitrary units relative to 18S RNA content as described previously [ 23 ].…”

Section: Methodsmentioning

confidence: 99%

Evaluation of P-Glycoprotein Inhibitory Potential Using a Rhodamine 123 Accumulation Assay

et al. 2016

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In vitro evaluation of P-glycoprotein (P-gp) inhibitory potential is now a regulatory issue during drug development, in order to predict clinical inhibition of P-gp and subsequent drug–drug interactions. Assays for this purpose, commonly based on P-gp-expressing cell lines and digoxin as a reference P-gp substrate probe, unfortunately exhibit high variability, raising thus the question of developing alternative or complementary tests for measuring inhibition of P-gp activity. In this context, the present study was designed to investigate the use of the fluorescent dye rhodamine 123 as a reference P-gp substrate probe for characterizing P-gp inhibitory potential of 16 structurally-unrelated drugs known to interact with P-gp. 14/16 of these P-gp inhibitors were found to increase rhodamine 123 accumulation in P-gp-overexpressing MCF7R cells, thus allowing the determination of their P-gp inhibitory potential, i.e., their half maximal inhibitor concentration (IC50) value towards P-gp-mediated transport of the dye. These IC50 values were in the range of variability of previously reported IC50 for P-gp and can be used for the prediction of clinical P-gp inhibition according to Food and Drug Administration (FDA) criteria, with notable sensitivity (80%). Therefore, the data demonstrated the feasibility of the use of rhodamine 123 for evaluating the P-gp inhibitory potential of drugs.

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“…However, human primary hepatocytes remain stable with time in culture, with a polarized status. Thus, monolayer-cultured human hepatocytes are also a valuable tool for the study of hepatic transporters since, contrary to that referred for rodent monolayer primary-cultured hepatocytes, the de-differentiation process is not expected to occur [ 170 ]. In fact, Schaefer and colleagues (2012) quantified sinusoidal and canalicular hepatic transporters by liquid chromatography/mass spectrometry (LC/MS) using both monolayer- and sandwich-cultured human primary hepatocytes and no major differences between them were found [ 171 ].…”

Section: Study Models For Abc Transportersmentioning

confidence: 99%

Cellular Models and In Vitro Assays for the Screening of modulators of P-gp, MRP1 and BCRP

et al. 2017

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Adenosine triphosphate (ATP)-binding cassette (ABC) transporters are highly expressed in tumor cells, as well as in organs involved in absorption and secretion processes, mediating the ATP-dependent efflux of compounds, both endogenous substances and xenobiotics, including drugs. Their expression and activity levels are modulated by the presence of inhibitors, inducers and/or activators. In vitro, ex vivo and in vivo studies with both known and newly synthesized P-glycoprotein (P-gp) inducers and/or activators have shown the usefulness of these transport mechanisms in reducing the systemic exposure and specific tissue access of potentially harmful compounds. This article focuses on the main ABC transporters involved in multidrug resistance [P-gp, multidrug resistance-associated protein 1 (MRP1) and breast cancer resistance protein (BCRP)] expressed in tissues of toxicological relevance, such as the blood-brain barrier, cardiovascular system, liver, kidney and intestine. Moreover, it provides a review of the available cellular models, in vitro and ex vivo assays for the screening and selection of safe and specific inducers and activators of these membrane transporters. The available cellular models and in vitro assays have been proposed as high throughput and low-cost alternatives to excessive animal testing, allowing the evaluation of a large number of compounds.

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Polarized location of SLC and ABC drug transporters in monolayer-cultured human hepatocytes

Cited by 25 publications

References 53 publications

Drug Transporter Expression and Activity in Human Hepatoma HuH-7 Cells

Drug Transporter Expression and Activity in Human Hepatoma HuH-7 Cells

Evaluation of P-Glycoprotein Inhibitory Potential Using a Rhodamine 123 Accumulation Assay

Cellular Models and In Vitro Assays for the Screening of modulators of P-gp, MRP1 and BCRP

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