Drug transporters are critically important for the absorption, distribution, metabolism, and excretion (ADME) of many drugs and endogenous compounds. Therefore, disruption of these pathways by ...inhibition, induction, genetic polymorphisms, or disease can have profound effects on overall physiology, drug pharmacokinetics, drug efficacy, and toxicity. This white paper provides a review of changes in transporter function associated with acute and chronic disease states, describes regulatory pathways affecting transporter expression, and identifies opportunities to advance the field.
There is an increasing interest in transporter induction (i.e., decreased systemic drug exposure due to increased efflux‐limited absorption or transporter‐mediated clearance) as a mechanism of ...drug–drug interactions (DDIs), although evidence of clinical relevance is still evolving. Intestinal P‐glycoprotein (P‐gp) and hepatic organic anion transporting polypeptides 1B (OATP1B) can be important determinants of drug absorption and disposition, as well as targets for DDIs. Current data indicate that intestinal P‐gp protein levels can be induced up to threefold to fourfold in humans primarily with pregnane X receptor (PXR) activators, and that this induction can decrease the systemic exposure of drugs with P‐gp efflux‐limited absorption (e.g., ≤ 67% decrease in the exposure of total dabigatran following rifampin multiple oral dosing). Evaluation of the clinical relevance of P‐gp induction as a DDI mechanism must consider the induction potential of the perpetrator drug for P‐gp and attenuation of exposure of the victim drug in the context of its therapeutic window. Practical drug development recommendations are provided herein. Reports are contradictory on OATP1B induction by PXR activators in human hepatocytes and liver biopsies. Some clinical investigations demonstrated that rifampin pretreatment decreased exposure of OATP1B substrates, while other studies found no differences, and the potential involvement of other mechanisms in these observed DDIs cannot be definitively ruled out. Thus, further studies are needed to understand hepatic OATP1B induction and potential involvement of other mechanisms contributing to reduced exposure of OATP1B substrates. This review critically summarizes the state‐of‐the‐art on intestinal P‐gp and hepatic OATP1B induction, and highlights implications for drug development.
This white paper examines recent progress, applications, and challenges in predicting unbound and total tissue and intra/subcellular drug concentrations using in vitro and preclinical models, imaging ...techniques, and physiologically based pharmacokinetic (PBPK) modeling. Published examples, regulatory submissions, and case studies illustrate the application of different types of data in drug development to support modeling and decision making for compounds with transporter‐mediated disposition, and likely disconnects between tissue and systemic drug exposure. The goals of this article are to illustrate current best practices and outline practical strategies for selecting appropriate in vitro and in vivo experimental methods to estimate or predict tissue and plasma concentrations, and to use these data in the application of PBPK modeling for human pharmacokinetic (PK), efficacy, and safety assessment in drug development.
Drug-induced liver injury (DILI) is the leading cause of acute liver failure and a major concern in drug development. Altered bile acid homeostasis via inhibition of the bile salt export pump (BSEP) ...is one mechanism of DILI. Dasatinib, pazopanib, and sorafenib are tyrosine kinase inhibitors (TKIs) that competitively inhibit BSEP and increase serum biomarkers for hepatotoxicity in ∼25-50% of patients. However, the mechanism(s) of hepatotoxicity beyond competitive inhibition of BSEP are poorly understood. This study examined mechanisms of TKI-mediated hepatotoxicity associated with altered bile acid homeostasis. Dasatinib, pazopanib, and sorafenib showed bile acid-dependent toxicity at clinically relevant concentrations, based on the C-DILI assay using sandwich-cultured human hepatocytes (SCHH). Among several bile acid-relevant genes, cytochrome P450 (CYP) 7A1 mRNA was specifically upregulated by 6.2- to 7.8-fold (dasatinib) and 5.7- to 9.3-fold (pazopanib), compared with control, within 8 hours. This was consistent with increased total bile acid concentrations in culture medium up to 2.3-fold, and in SCHH up to 1.4-fold, compared with control, within 24 hours. Additionally, protein abundance of sodium taurocholate co-transporting polypeptide (NTCP) was increased up to 2.0-fold by these three TKIs. The increase in NTCP protein abundance correlated with increased function; dasatinib and pazopanib increased hepatocyte uptake clearance (CL
) of taurocholic acid, a probe bile acid substrate, up to 1.4-fold. In conclusion, upregulation of
and NTCP in SCHH constitute novel mechanisms of TKI-associated hepatotoxicity. SIGNIFICANCE STATEMENT: Understanding the mechanisms of hepatotoxicity associated with tyrosine kinase inhibitors (TKIs) is fundamental to development of effective and safe intervention therapies for various cancers. Data generated in sandwich-cultured human hepatocytes, an
model of drug-induced hepatotoxicity, revealed that TKIs upregulate bile acid synthesis and alter bile acid uptake and excretion. These findings provide novel insights into additional mechanisms of bile acid-mediated drug-induced liver injury, an adverse effect that limits the use and effectiveness of TKI treatment in some cancer patients.
Autosomal dominant polycystic kidney disease (ADPKD) is a common form of inherited polycystic kidney disease (PKD) and is a leading cause of kidney failure. Fluid-filled cysts develop in the kidneys ...of patients with ADPKD, and cysts often form in their liver and other organs. Previous data have shown that bile acids are increased in the liver of polycystic kidney (PCK) rats, a rodent model of PKD; these changes may be associated with alterations in liver transporter expression and function. However, the impact of PKD on hepatic transporters has not been characterized. Therefore, this preclinical study was designed to investigate hepatic transporter expression and function in PCK compared with wild-type (WT) Sprague-Dawley rats. Transporter gene expression was measured by quantitative polymerase chain reaction, and protein levels were quantified by Western blot and liquid chromatography-tandem mass spectroscopy (LC-MS/MS)-based proteomic analysis in rat livers. Transporter function was assessed in isolated perfused livers (IPLs), and biliary and hepatic total glutathione content was measured. Protein expression of Mrp2 and Oatp1a4 was decreased 3.0-fold and 2.9-fold, respectively, in PCK rat livers based on Western blot analysis. Proteomic analysis confirmed a decrease in Mrp2 and a decrease in Oatp1a1 expression (PCK/WT ratios, 0.368 ± 0.098 and 0.563 ± 0.038, respectively; mean ± S.D.). The biliary excretion of 5(6)-carboxy-2',7'-dichlorofluorescein, a substrate of Oatp1a1, Mrp2, and Mrp3, was decreased 28-fold in PCK compared with WT rat IPLs. Total glutathione was significantly reduced in the bile of PCK rats. Differences in hepatic transporter expression and function may contribute to altered disposition of Mrp2 and Oatp substrates in PKD.
Candida infections are a leading cause of infectious disease-related death in children supported by extracorporeal membrane oxygenation (ECMO). The ECMO circuit can alter drug pharmacokinetics (PK); ...thus, standard fluconazole dosing may result in suboptimal drug exposures. The objective of our study was to determine the PK of fluconazole in children on ECMO. Forty children with 367 PK samples were included in the analysis. The PK data were analyzed using nonlinear mixed-effect modeling (NONMEM). A one-compartment model best described the data. Weight was included in the base model for clearance (CL) and volume of distribution (V). The final model included the effect of serum creatinine (SCR) level on CL and the effect of ECMO on V as follows: CL (in liters per hour) = 0.019 × weight × (SCR/0.4)(-0.29) × exp(ηCL) and V (in liters) = 0.93 × weight × 1.4(ECMO) × exp(ηV). The fluconazole V was increased in children supported by ECMO. Consequently, children on ECMO require a higher fluconazole loading dose for prophylaxis (12 mg/kg of body weight) and treatment (35 mg/kg) paired with standard maintenance doses to achieve exposures similar to those of children not on ECMO.
The effect of membrane transporters on drug disposition, efficacy and safety is now well recognized. Since the initial publication from the International Transporter Consortium, significant progress ...has been made in understanding the roles and functions of transporters, as well as in the development of tools and models to assess and predict transporter-mediated activity, toxicity and drug-drug interactions (DDIs). Notable advances include an increased understanding of the effects of intrinsic and extrinsic factors on transporter activity, the application of physiologically based pharmacokinetic modelling in predicting transporter-mediated drug disposition, the identification of endogenous biomarkers to assess transporter-mediated DDIs and the determination of the cryogenic electron microscopy structures of SLC and ABC transporters. This article provides an overview of these key developments, highlighting unanswered questions, regulatory considerations and future directions.
Per- and polyfluoroalkyl substances (PFAS) have become internationally recognized over the past three decades as persistent organic pollutants used in the production of various consumer and ...industrial goods. Research efforts continue to gauge the risk that historically used, and newly produced, PFAS may cause to human health. Numerous studies report toxic effects of PFAS on the human liver as well as increased serum cholesterol levels in adults. A major concern with PFAS, also dubbed "forever chemicals", is that they accumulate in liver and kidney and persist in serum. The mechanisms responsible for their disposition and excretion in humans are poorly understood. A better understanding of the interaction of PFAS with liver transporters, as it pertains to the disposition of PFAS and other xenobiotics, could provide mechanistic insight into human health effects and guide efforts toward risk assessment of compounds in development. This review summarizes the current state of the literature on the emerging relationships (e.g., substrates, inhibitors, modulators of gene expression) between PFAS and specific hepatic transporters. The adaptive and toxicological responses of hepatocytes to PFAS that reveal linkages to pathologies and epidemiological findings are highlighted. The evidence suggests that our understanding of the molecular landscape of PFAS must improve to determine their impact on the expression and function of hepatocyte transporters that play a key role in PFAS or other xenobiotic disposition. From here, we can assess what role these changes may have in documented human health outcomes.
The bile salt export pump (BSEP) plays an important role in bile acid excretion. Impaired BSEP function may result in liver injury. Bile acids also undergo basolateral efflux, but the relative ...contributions of biliary (CLBile) versus basolateral efflux (CLBL) clearance to hepatocellular bile acid excretion have not been determined. In the present study, taurocholic acid (TCA; a model bile acid) disposition was characterized in human and rat sandwich-cultured hepatocytes (SCH) combined with pharmacokinetic modeling. In human SCH, biliary excretion of TCA predominated (CLBile = 0.14 ± 0.04 ml/min per g liver; CLBL = 0.042 ± 0.019 ml/min per g liver), whereas CLBile and CLBL contributed approximately equally to TCA hepatocellular excretion in rat SCH (CLBile = 0.34 ± 0.07 ml/min per g liver; CLBL = 0.26 ± 0.07 ml/min per g liver). Troglitazone decreased TCA uptake, CLBile, and CLBL; membrane vesicle assays revealed for the first time that the major metabolite, troglitazone sulfate, was a noncompetitive inhibitor of multidrug resistance-associated protein 4, a basolateral bile acid efflux transporter. Simulations revealed that decreased CLBile led to a greater increase in hepatic TCA exposure in human than in rat SCH. A decrease in both excretory pathways (CLBile and CLBL) exponentially increased hepatic TCA in both species, suggesting that 1) drugs that inhibit both pathways may have a greater risk for hepatotoxicity, and 2) impaired function of an alternate excretory pathway may predispose patients to hepatotoxicity when drugs that inhibit one pathway are administered. Simulations confirmed the protective role of uptake inhibition, suggesting that a drug's inhibitory effects on bile acid uptake also should be considered when evaluating hepatotoxic potential. Overall, the current study precisely characterized basolateral efflux of TCA, revealed species differences in hepatocellular TCA efflux pathways, and provided insights about altered hepatic bile acid exposure when multiple transport pathways are impaired.
The sodium taurocholate cotransporting polypeptide (NTCP; gene name
) is the primary hepatic basolateral uptake transporter for conjugated bile acids and the entry receptor for the hepatitis B and D ...virus (HBV/HDV). Regulation of human NTCP remains a knowledge gap due to significant species differences in substrate and inhibitor selectivity and plasma membrane expression. In the present study, various kinase inhibitors were screened for inhibition of NTCP function and taurocholate (TCA) uptake using NTCP-transfected HuH-7 cells. This study identified everolimus, an mTOR inhibitor and macrocyclic immunosuppressive drug, as an NTCP inhibitor with modest potency (IC
= 6.7-8.0 µM). Further investigation in differentiated HuH-7 cells expressing NTCP and NTCP-overexpressing Flp-In T-REx 293 cells revealed that the mechanism of action of everolimus on NTCP is direct inhibition and mTOR-independent. Structural analogs of everolimus inhibited NTCP-mediated TCA uptake, however, functional analogs did not affect NTCP-mediated TCA transport, providing further evidence for direct inhibition. This work contributes to the growing body of literature suggesting that NTCP-mediated bile acid uptake may be inhibited by macrocyclic peptides, which may be further exploited to develop novel medications against HBV/HDV.
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