Hepatic Drug Transporters Research Articles

Integrating Pharmacogenetics for Assessment of Herb-Drug Interactions: Has the Time Come?

It is increasingly common for patients to use western medications concurrently with Traditional Chinese Medicine or other herbal medicines. Indeed, it is estimated that one-third of people in developed countries are using alternative therapies including herbal medicines. Metabolism of drugs/herbs takes place to a large extent in the liver and small intestine via various drug-metabolizing enzymes. Therefore, the concurrent use of western and herbal medicines has potential for drug-herb interactions at these biological locations. There are several hepatic metabolic enzymes and drug transporters that have been documented to be involved in the herb-drug interactions. These include the cytochrome P450 (CYP450) enzymes that contribute to metabolism of various drugs, food constituents, and herbs, and the ATPbinding cassette (ABC) transporters, particularly P-glycoprotein (P-gp), and solute carrier (SLC) transporters. Pharmacogenetic variance in CYP450s and drug transporters in humans and animals may lead to pharmacokinetic and pharmacodynamics changes of target substrates which could be drugs and/or herbal medicines. This paper presents a synthesis of the emerging evidence on the role of pharmacogenetics in the mechanism of the salient herb-drug interactions, and why the time has come to consider integrating this new strand of postgenomics knowledge to clinical and public health practice. Keywords: CYP 450, drug transporter, genetic polymorphism, herb-drug interaction, pharmacogenetics.

Evaluation of Organic Anion-Transporting Polypeptide 1B1 and CYP3A4 Activities in Primary Human Hepatocytes and HepaRG Cells Cultured in a Dynamic Three-Dimensional Bioreactor System

The long-term stability of liver cell functions is a major challenge when studying hepatic drug transport, metabolism, and toxicity in vitro. The aim of the present study was to investigate organic anion-transporting polypeptide (OATP) 1B1 and CYP3A4 activities in fresh primary human hepatocytes and differentiated cryopreserved HepaRG cells when cultured in a three-dimensional (3D) bioreactor system. OATP1B1 activity was determined by loss from media experiments of [(3)H]estradiol-17β-D-glucuronide and atorvastatin acid (ATA) for up to 7 days in culture. ATA metabolite formation was determined at days 3 to 4 to evaluate CYP3A4 activity. Overall, the results showed that freshly isolated human hepatocytes inoculated in the bioreactor retained OATP1B1 activity for at least 7 days, whereas in HepaRG cells no OATP1B1 activity was observed beyond day 2. The activity data were in agreement with immunohistochemical stainings, which showed that OATP1B1 protein expression was preserved for at least 9 days in fresh human hepatocytes, whereas OATP1B1 was expressed markedly lower in HepaRG cells after 9 days in culture. Fresh human hepatocytes and HepaRG cells exhibited similar CYP3A4 activity in bioreactor culture, and immunohistochemical stainings supported these findings. Activity and mRNA expression of OATP1B1 and CYP3A4 in primary human hepatocytes compared with HepaRG cells in fresh suspensions were in agreement with data obtained in bioreactor culture. In conclusion, freshly isolated human hepatocytes cultured in a 3D bioreactor system preserve both OATP1B1 and CYP3A4 activities, allowing long-term in vitro studies on drug disposition and toxicity.

Drug disposition and clinical practice in neonates: Cross talk between developmental physiology and pharmacology

Drug dosing in infants should be based on their physiological characteristics and the pharmacokinetic and -dynamic profile of the compound. Since maturational physiological changes are most prominent in infancy, variability is the key feature of clinical pharmacology in infancy: developmental physiology drives developmental pharmacology. This is illustrated by the link between renal physiology and renal drug clearance and between hepatic physiology and hepatic drug elimination for some specific compounds. However, the maturational profiles of the individual elimination processes differ substantially at the enzyme and transporter level. This implies that it is important to integrate all ontogeny-related knowledge of the different elimination routes to predict compound specific, phenotypic in vivo observations of infancy. In addition to the introduction of already available in vivo observations to validate mechanistic (estimated to in vivo observations) or physiology based pharmacokinetic (PBPK, developmental physiology related estimated to in vivo observations) models, a simultaneous use of both approaches (mechanistic and PBPK) to search for discrepancies between both approaches may also unveil 'missing' links in maturational physiology or clinical pharmacology (e.g. ontogeny renal or hepatic drug transporters): developmental pharmacology drives developmental pharmacology.

Looking for synergy or additivity between 188Re and sorafenib on hepatoma cell lines.

247 Background: In case of non resectable HCC, radioembolization and sorafenib (S) are therapeutic options respectively for intermediate and advanced stages. In some other cancers, there is an increase of efficacy when external beam radiotherapy is done concomitantly with systemic chemotherapy or targeted therapies. So we wondered if there could be a synergistic or an additive activity when S is combined with a radionuclide. Methods: Hepatoma cell lines N1S1 (murine HCC), HepG2 (human hepatoblastoma) and HepaRG (human HCC) were treated with increasing concentrations of rhenium-188 (188Re) or S. On each cell line, we have studied the cellular toxicities of S and 188Re using Tetrazolium dye test, extra-cellular medium LDH level and morphologic analysis. This was done for different dosage of S and 188Re. We measured the lethal concentration killing 25% of cells (LC25) with the results of the Tetrazolium dye test. Secondly, we looked for synergy or additivity on cellular toxicity of these two compounds according to cell lines by combined treatment. Synergy or additivity was estimated with the combination index (CI) method (synergy if CI lower than 1, additivity if CI = 1, antagonism if CI upper to 1) based on the Tetrazolium dye test’s results. Results: Monotherapy dose-dependent toxicities were observed for all three cell lines with 188Re and for the N1S1 and HepG2 cell lines only with S. Combined treatment with 188Re and S showed synergy on HepaRG and N1S1 cell lines and additivity on the HepG2 cell line. Conclusions: The additive, and even synergistic, interest of a combined treatment with 188Re and S is demonstrated in vitro (for the first time to our knowledge) on hepatoma cell lines. This results, in particular for the HepaRG cell line (human HCC), could be explained by the down-regulation of the hepatic drug transporters which are responsible for the Sorafenib efflux in case of simultaneous DNA damages due to a radionuclide exposition. This promising approach now needs to be confirmed in vivo. [Table: see text]

Pregnane X receptor regulates drug metabolism and transport in the vasculature and protects from oxidative stress

AimsCirculating endogenous, dietary, and foreign chemicals can contribute to vascular dysfunction. The mechanism by which the vasculature protects itself from these chemicals is unknown. This study investigates whether the pregnane X receptor (PXR), the major transcriptional regulator of hepatic drug metabolism and transport that responds to such xenobiotics, mediates vascular protection by co-ordinating a defence gene programme in the vasculature.Methods and resultsPXR was detected in primary human and rat aortic endothelial and smooth muscle cells (SMC) and blood vessels including the human and rat aorta. Metabolic PXR target genes cytochrome P450 3A, 2B, 2C, and glutathione S-transferase mRNA and activity were induced by PXR ligands in rodent and human vascular cells and absent in the aortas from PXR-null mice stimulated in vivo or in rat aortic SMC expressing dominant-negative PXR. Activation of aortic PXR by classical agonists had several protective effects: increased xenobiotic metabolism demonstrated by bioactivation of the pro-drug clopidogrel, which reduced adenosine diphosphate-induced platelet aggregation; increased expression of multidrug resistance protein 1, mediating chemical efflux from the vasculature; and protection from reactive oxygen species-mediated cell death.ConclusionPXR co-ordinately up-regulates drug metabolism, transport, and antioxidant genes to protect the vasculature from endogenous and exogenous insults, thus representing a novel gatekeeper for vascular defence.

Sodium fluorescein is a probe substrate for hepatic drug transport mediated by OATP1B1 and OATP1B3

The aim of this study was to characterize the in vitro hepatic uptake kinetics of sodium fluorescein (NaFluo) and identify the transporters involved. NaFluo exhibited saturable uptake kinetics in suspended rat and human hepatocytes as reflected by Km values of 22.5 and 14.1 µM, and Vmax values of 98.3 and 5.8 pmol/(million cells • min), respectively. Coincubation with known inhibitors (e.g. rifampicin) of organic anion transporting polypeptide (OATP/Oatp; SLCO gene family) significantly decreased NaFluo uptake in hepatocytes. In contrast, neither inhibitors/substrates of the organic cation transporter or organic anion transporter family nor depletion of extracellular sodium resulted in significant inhibition of NaFluo uptake. To explore the contribution of individual uptake transporters, NaFluo uptake was determined in Chinese hamster ovary cells transfected with OATP1B1, OATP1B3, and OATP2B1. Transporter‐mediated uptake of NaFluo was observed in OATP1B1‐ and OATP1B3‐transfected cells (Km = 4.2 and 10.9 µM; Vmax = 30.9 and 135 [pmol/(mg protein • min)], respectively). NaFluo can be used as a probe substrate to study Oatp/OATP1B‐mediated drug interactions in fluorescence‐based in vitro transport assays of rat and human liver. Labeling of drugs or bile salts with a fluorescein moiety can be expected to result in fluorescent conjugates with substantially altered hepatic uptake characteristics as compared with the unconjugated compounds. © 2011 Wiley‐Liss, Inc. and the American Pharmacists Association J Pharm Sci 100:5018–5030, 2011

Effects of Chronic Renal Failure on Brain Drug Transporters in Rats

Studies demonstrated that chronic renal failure (CRF) affects the expression and activity of intestinal, hepatic, and renal drug transporters. Such drug transporters are expressed in brain cells and at the blood-brain barrier (BBB), where they limit the entry and distribution of drugs in the brain. Perturbations in brain drug transporter equilibrium by CRF could lead to central drug toxicity. This study evaluates how CRF affects BBB drug transporters using a 5/6 nephrectomized rat model. Protein and mRNA expression of influx transporters [organic anion-transporting polypeptide (Oatp), organic anion transporter (Oat)] and efflux transporters [P-glycoprotein (P-gp), multidrug resistance-related protein (Mrp), breast cancer resistance protein (Bcrp)] were measured in CRF and control rat brain. Intracerebral accumulation of radiolabeled benzylpenicillin, digoxin, doxorubicin, and verapamil was used to evaluate BBB drug permeability. Protein expression of the transporters was evaluated in rat brain endothelial cells (RBECs) and astrocytes incubated with control and CRF rat serum. We demonstrated significant decreases (30-50%) in protein and mRNA levels of Bcrp, Mrp2 to -4, Oat3, Oatp2 and -3, and P-gp in CRF rat brain biopsies, as well as in astrocytes and RBECs incubated with CRF serum. These decreases did not correlate with in vivo changes because BBB permeability of benzylpenicillin was decreased by 30% in CRF rats, whereas digoxin, doxorubicin, and verapamil permeabilities were unchanged. It thus seems that even with decreased drug transporters, BBB integrity and function is conserved in CRF.

Transporter-Mediated Drug–Drug Interactions

Drug-drug interactions are a serious clinical issue. An important mechanism underlying drug-drug interactions is induction or inhibition of drug transporters that mediate the cellular uptake and efflux of xenobiotics. Especially drug transporters of the small intestine, liver and kidney are major determinants of the pharmacokinetic profile of drugs. Transporter-mediated drug-drug interactions in these three organs can considerably influence the pharmacokinetics and clinical effects of drugs. In this article, we focus on probe drugs lacking significant metabolism to highlight mechanisms of interactions of selected intestinal, hepatic and renal drug transporters (e.g., organic anion transporting polypeptide [OATP] 1A2, OATP2B1, OATP1B1, OATP1B3, P-gp, organic anion transporter [OAT] 1, OAT3, breast cancer resistance protein [BCRP], organic cation transporter [OCT] 2 and multidrug and toxin extrusion protein [MATE] 1). Genotype-dependent drug-drug interactions are also discussed.

Automated Applications of Sandwich-Cultured Hepatocytes in the Evaluation of Hepatic Drug Transport

Predictions of the absorption, distribution, metabolism, excretion, and toxicity of compounds in pharmaceutical development are essential aspects of the drug discovery process. B-CLEAR is an in vitro system that uses sandwich-cultured hepatocytes to evaluate and predict in vivo hepatobiliary disposition (hepatic uptake, biliary excretion, and biliary clearance), transporter-based hepatic drug-drug interactions, and potential drug-induced hepatotoxicity. Automation of predictive technologies is an advantageous and preferred format in drug discovery. In this study, manual and automated studies are investigated and equivalence is demonstrated. In addition, automated applications using model probe substrates and inhibitors to assess the cholestatic potential of drugs and evaluate hepatic drug transport are examined. The successful automation of this technology provides a more reproducible and less labor-intensive approach, reducing potential operator error in complex studies and facilitating technology transfer.

Regulation of drug transporter mRNA expression by interferon-γ in primary human hepatocytes

Interferon (IFN)-γ is known to downregulate expression of drug detoxifying proteins such as cytochromes P-450 (CYPs) in human hepatocytes. The present study was designed to determine whether IFN-γ may also impair expression of influx and efflux drug transporters, which constitute important determinants of the liver detoxification pathway. Exposure of primary human hepatocytes to 10 ng/mL IFN-γ was found to downregulate mRNA levels of sinusoidal influx transporters such as sodium-taurocholate cotransporting polypeptide, organic anion transporting polypeptide (OATP) 2B1, OATP1B1, and OATP1B3. IFN-γ concomitantly reduced mRNA expression of drug efflux pumps such as multidrug resistance gene 1, multidrug resistance protein (MRP) 2, MRP3, breast cancer resistance protein and bile salt export pump. Such IFN-γ-mediated repression of major hepatic drug transporters may contribute to impaired liver clearance of drugs administrated to patients suffering from inflammation or viral infections associated with increased secretion of IFN-γ.

Impact of Polyinosinic/Polycytidylic Acid on Placental and Hepatobiliary Drug Transporters in Pregnant Rats

Although inflammation is known to impose changes in the expression and activity of drug transporters, little is known about the impact of inflammatory stimuli on these transporters during pregnancy. Our objective was to study the effect of viral-induced inflammation on key maternal hepatic and placental drug transporters and their endogenous substrates. Acute inflammation was induced in pregnant Sprague-Dawley rats (gestation day 17-18, n = 5-6/group) by single intraperitoneal doses of polyinosinic/polycytidylic acid [poly(I:C)] (2.5 or 5.0 mg/kg) with saline as a control. Tissues were harvested 24 h later. Expression of transporters was measured via real-time polymerase chain reaction and Western blotting. Maternal plasma levels of cytokines, bile acids, and bilirubin and fetal levels of bile acids were examined. Plasma concentrations of interferon-γ, tumor necrosis factor-α, and interleukin-6 were significantly induced in poly(I:C)-treated rats, compared with controls (p < 0.001). Significant down-regulation of placental Abcb1a/b, Abcc1, Abcc3, Abcg2, Slco1a4, and Slco4a1 mRNA and of hepatic Abcc2, Abcg2, Slco1a4, Slc10a1, and Cyp3a2 mRNA was observed in poly(I:C)-treated rats. Hepatic Abcb1b and Abcc3 mRNA levels were significantly induced. Hepatic protein levels of P-glycoprotein, multidrug resistance-associated protein 2, and breast cancer resistance protein were significantly down-regulated relative to those for controls (p < 0.05). Total bile acids in maternal plasma were significantly increased at the higher dose of poly(I:C). In summary, the poly(I:C) model of viral infection imposes significant changes in the expression of key drug transporters in placental and hepatic tissues of pregnant rats. Because many clinically important endogenous and exogenous compounds are substrates of these transporters, inflammation-mediated changes in transporter expression could affect their maternal disposition and fetal exposure.

Upregulation of multidrug resistance-associated protein 4 in the early stage of vascular invasion of hepatocellular carcinoma.

4146 Background: In human liver, many drug transporters play important physiological and pharmacological roles. The expression of hepatic drug transporters is altered in various pathophysiologic co...

Sulindac and Its Metabolites Inhibit Multiple Transport Proteins in Rat and Human Hepatocytes

Sulindac is a commonly used nonsteroidal anti-inflammatory drug. This study tested the hypothesis that sulindac-mediated drug-drug interactions and/or hepatotoxicity may be caused, in part, by inhibition of proteins responsible for the hepatic transport of drugs and/or bile acids by sulindac and/or sulindac metabolites [sulindac sulfone (S-sulfone) and sulindac sulfide (S-sulfide)]. The uptake and excretion of model substrates, [(3)H]taurocholate (TC), [(3)H]estradiol 17-beta-glucuronide (E217G), and nitrofurantoin (NF), were investigated in rat and human suspended and sandwich-cultured hepatocytes (SCH). In suspended rat hepatocytes, S-sulfone and S-sulfide inhibited Na(+)-dependent TC initial uptake (IC(50) of 24.9 +/- 6.4 and 12.5 +/- 1.8 microM, respectively) and Na(+)-independent E217G initial uptake (IC(50) of 12.1 +/- 1.6 and 6.3 +/- 0.3 microM, respectively). In rat SCH, sulindac metabolites (100 microM) decreased the in vitro biliary clearance (Cl(biliary)) of TC, E217G, and NF by 38 to 83%, 81 to 97%, and 33 to 57%, respectively; S-sulfone and S-sulfide also decreased the TC and NF biliary excretion index by 39 to 55%. In suspended human hepatocytes, S-sulfone and S-sulfide inhibited Na(+)-dependent TC initial uptake (IC(50) of 42.2 and 3.1 microM, respectively); S-sulfide also inhibited the TC Cl(biliary) in human SCH. Sulindac/metabolites markedly inhibited hepatic uptake and biliary excretion of E217G by 51 to 100% in human SCH. In conclusion, sulindac and metabolites are potent inhibitors of the uptake and biliary clearance of bile acids in rat and human hepatocytes and also inhibit substrates of rat breast cancer resistance protein, rat and human organic anion-transporting polypeptides, and human multidrug resistance-associated protein 2. Inhibition of multiple hepatic transport proteins by sulindac/metabolites may play an important role in clinically significant sulindac-mediated drug-drug interactions and/or liver injury.

Screening of OATP1B1/3 and OCT1 inhibitors in cryopreserved hepatocytes in suspension

Drug–drug interactions involving hepatic drug transporters may have clinical consequences and jeopardize development of promising drug candidates. Organic anion transporting polypeptides (OATP/Oatp) and the organic cation transporters (OCT/Oct) are among the most important transporters involved in xenobiotic uptake in the liver. In the present study, 179 molecules have been tested as inhibitors of the uptake of estradiol-17βD-glucuronide (E 217βG), substrate of OATP1B1/3 (rOatp), or 1-methyl-4-phenylpyridinium (MPP+), substrate of OCT1 (rOct1), into suspended cryopreserved hepatocytes from humans and rats. Uptake was assessed in 96-well plates by measuring intracellular accumulation of radioactive substrate in hepatocytes in presence or absence of inhibitor. In rat hepatocytes 140 compounds were identified as inhibitors (inhibition at 20 μM ≥ 30%) of E 217βG uptake and 77 compounds inhibitors of MPP+ uptake. The most potent inhibitors of rOatp and rOct1 were dantrolene sodium ( K i = 2 ± 9 μM) and bepridil ( K i = 14 ± 2 μM), respectively. In human hepatocytes, the most potent inhibitors of E 217βG and MPP+ uptake were capsazepine ( K i = 14 ± 5 μM) and cyproheptadine ( K i = 19 ± 3 μM), respectively. Structure–activity relationship (SAR) analysis of all tested compounds suggested that lipophilicity, polarity, p K a and the number of hydrogen bond donors and acceptors play a role in their interaction with the transporters investigated. The method used here is a simple tool to screen large number of compounds as inhibitors of the uptake of substrates into suspended hepatocytes.

Impact of Hyperlipidemia on Plasma Protein Binding and Hepatic Drug Transporter and Metabolic Enzyme Regulation in a Rat Model of Gestational Diabetes

It is currently unknown whether gestational diabetes mellitus (GDM), a prevalent obstetrical complication, compounds the changes in drug disposition that occur naturally in pregnancy. Hyperlipidemia occurs in GDM. Using a rat model of GDM, we determined whether excess lipids compete with drugs for plasma protein binding. Because lipids activate nuclear receptors that regulate drug transporters and metabolic enzymes, we used proteome analysis to determine whether hyperlipidemia indirectly leads to the dysregulation of these proteins in the liver. GDM was induced on gestational day 6 (GD6) via streptozotocin injection. Controls received either vehicle alone or streptozotocin with subsequent insulin treatment. Liver and plasma were collected on GD20. Glyburide and saquinavir protein binding was determined by ultrafiltration, and an established solvent method was used for plasma delipidation. Proteomics analysis was performed by using isobaric tags for relative and absolute quantitation methodology with membrane-enriched hepatic protein samples. Relative to controls, GDM rat plasma contained more cholesterol and triglycerides. Plasma protein binding of glyburide and saquinavir was decreased in GDM. Delipidation normalized protein binding in GDM plasma. Proteins linked to lipid metabolism were strongly affected in the GDM proteomics data set, with prohyperlipidemic and antihyperlipidemic changes observed, and formed networks that implicated several nuclear receptors. Up-regulation of drug transporters and metabolic enzymes was observed (e.g., multidrug resistance 1/2, CYP2A1, CYP2B9, and CYP2D3). In this study, GDM-induced hyperlipidemia decreased protein binding and was associated with drug transporter and metabolic enzyme up-regulation in the liver. Both of these findings could change drug disposition in affected pregnancies, compounding changes associated with pregnancy itself.

Transport mechanism and substrate specificity of human organic anion transporter 2 (hOat2 [SLC22A7

Human organic anion transporter 2 (hOat2[SLC22A7]) is highly expressed in the human liver. Although localization, gene expression, substrate specificity and transport mechanisms of other human Oat isoforms such as human Oat1 (hOat1), human Oat3 (hOat3) and human Oat4 (hOat4) have been elucidated, information concerning human Oat2 (hOat2) is less defined. The objective of this study was to provide further information on the transport mechanism and substrate specificity of hOat2. When expressed in Xenopus laevis oocytes, the transport of organic compounds mediated by hOat2 was not affected by the replacement of extracellular sodium with lithium, choline and mannitol. The uptake of estrone sulfate (ES) in hOat2-expressing oocytes was significantly trans-stimulated by preloading the oocytes with fumarate and succinate, but not glutarate. Moreover, we observed that hOat2 mediates the transport of bumetanide, ES, glutarate, dehydroepiandrosterone sulfate, allopurinol, prostaglandin E2, 5-fluorouracil, paclitaxel and L-ascorbic acid. These compounds are identified for the first time as hOat2 substrates. A wide range of structurally unrelated organic compounds inhibited the hOat2-mediated uptake of tetracycline, except for sulfobromophthalein. All of these findings indicate that hOat2 is a sodium-independent multi-specific organic anion/dimethyldicarboxylate exchanger. Our present findings thus provide further insights into the role of hOat2 in hepatic drug transport.

HIV protease inhibitors are substrates for OATP1A2, OATP1B1 and OATP1B3 and lopinavir plasma concentrations are influenced by SLCO1B1 polymorphisms

OATP1B1 and OATP1B3 are major hepatic drug transporters whilst OATP1A2 is mainly located in the brain but is also located in liver and several other organs. These transporters affect the distribution and clearance of many endobiotics and xenobiotics and have been reported to have functional single nucleotide polymorphisms (SNPs). We have assessed the substrate specificities of these transporters for a panel of antiretrovirals and investigated the effects of SNPs within these transporters on the pharmacokinetics of lopinavir. SLCO1A2, SLCO1B1 and SLCO1B3 were cloned, verified and used to generate cRNA for use in the Xenopuslaevis oocyte transport system. Using the oocyte system, antiretrovirals were tested for their substrate specificities. Plasma samples (n=349) from the Liverpool therapeutic drug monitoring registry were genotyped for SNPs in SLCO1A2, SLCO1B1 and SLCO1B3 and associations between SNPs and lopinavir plasma concentrations were analysed. Antiretroviral protease inhibitors, but not non-nucleoside reverse transcriptase inhibitors, are substrates for OATP1A2, OATP1B1 and OATP1B3. Furthermore, ritonavir was not an inhibitor of OATP1B1. The 521T>C polymorphism in SLCO1B1 was significantly associated with higher lopinavir plasma concentrations. No associations were observed with functional variants of SLCO1A2 and SLCO1B3. These data add to our understanding of the factors that contribute to variability in plasma concentrations of protease inhibitors. Further studies are now required to confirm the association of SLCO1B1 521T>C with lopinavir plasma concentrations and to assess the influence of other polymorphisms in the SLCO family.

Hepatitis C Virus-related Cirrhosis is a Major Determinant of the Expression Levels of Hepatic Drug Transporters

Hepatic drug transporters are responsible for both hepatic uptake and the biliary excretion of drugs. Expression changes in hepatic drug transporter genes have been observed in various pathophysiological conditions. However, it has not been comprehensively investigated what factors substantially influence the mRNA levels of hepatic drug transporters. In this study, we quantified the mRNA expression of 17 drug transporters using noncancerous liver tissue samples and carried out stepwise multiple regression analysis to identify the factors affecting their expression from 18 clinical variables. For 17 drug transporters, the mRNA level of organic anion transporting polypeptide (OATP) 2B1 was highest, followed by that of organic cation transporter 1, organic anion transporter 2, OATP1B1, OATP1B3, multidrug resistance-associated protein (MRP) 6, and MRP3. Stepwise multiple regression analysis demonstrated MRP4 mRNA level to be predicted with the greatest accuracy among 17 drug transporters. Of clinical variables entered into the prediction model for MRP4, hepatitis C virus (HCV) infection and liver cirrhosis were crucial factors affecting MRP4 mRNA and protein levels. Furthermore, HCV-related cirrhosis influenced the mRNA levels of 8 drug transporters besides MRP4. These findings indicate that HCV-related cirrhosis is a crucial factor affecting the expression of hepatic drug transporters, especially MRP4.

Human Skeletal Muscle Drug Transporters Determine Local Exposure and Toxicity of Statins

The 3-hydroxy-3-methylglutaryl coenzyme A reductase inhibitors, or statins, are important drugs used in the treatment and prevention of cardiovascular disease. Although statins are well tolerated, many patients develop myopathy manifesting as muscle aches and pain. Rhabdomyolysis is a rare but severe toxicity of statins. Interindividual differences in the activities of hepatic membrane drug transporters and metabolic enzymes are known to influence statin plasma pharmacokinetics and risk for myopathy. Interestingly, little is known regarding the molecular determinants of statin distribution into skeletal muscle and its relevance to toxicity. We sought to identify statin transporters in human skeletal muscle and determine their impact on statin toxicity in vitro. We demonstrate that the uptake transporter OATP2B1 (human organic anion transporting polypeptide 2B1) and the efflux transporters, multidrug resistance-associated protein (MRP)1, MRP4, and MRP5 are expressed on the sarcolemmal membrane of human skeletal muscle fibers and that atorvastatin and rosuvastatin are substrates of these transporters when assessed using a heterologous expression system. In an in vitro model of differentiated, primary human skeletal muscle myoblast cells, we demonstrate basal membrane expression and drug efflux activity of MRP1, which contributes to reducing intracellular statin accumulation. Furthermore, we show that expression of human OATP2B1 in human skeletal muscle myoblast cells by adenoviral vectors increases intracellular accumulation and toxicity of statins and such effects were abrogated when cells overexpressed MRP1. These results identify key membrane transporters as modulators of skeletal muscle statin exposure and toxicity.

Regulation of human hepatic drug transporter expression by pro-inflammatory cytokines

Background: Sinuosidal and canalicular hepatic drug transporters, involved in drug uptake in the liver and drug secretion in the bile, respectively, play a major role in liver drug clearance. Inflammation is well-known to impair expression of these transporters in rodents; data about this topic have been more recently reported in human hepatocytes. Objective: The present review is designed to summarise the effects of pro-inflammatory cytokines such as IL-1β, TNF-α and IL-6 toward human hepatic drug and bile salt transporters. Methods: Recent studies aimed at analyzing transporter expression and activity in cytokines-exposed primary human hepatocytes and well-differentiated human hepatoma cells are resumed and discussed. Results/conclusion: Exposure to IL-1β, TNF-α or IL-6 markedly alters expression profile of human hepatic transporters. Bile salt transporters as well as sinusoidal solute carrier transporters are usually repressed, whereas ATP-binding cassette drug efflux pumps remain unchanged or are either downregulated or upregulated. These changes are observed at mRNA levels, but also, for some of the transporters, at protein and activity levels. They are likely to contribute to alterations of drug pharmacokinetics, impairment of bile salt secretion and cholestasis caused by inflammation in humans.