Effect of changes in metabolic enzymes and transporters on drug metabolism in the context of liver disease: Impact on pharmacokinetics and drug-drug interactions.

Changes in the pharmacokinetic and resulting pharmacodynamic properties of drugs are common in many chronic liver diseases, leading to adverse effects, drug interactions and increased risk of over- or underdosing of medications. Structural and functional hepatic impairment can have major effects on drug metabolism and transport. This review summarizes research on the functional changes in phase I and II metabolic enzymes and in transport proteins in patients with metabolic diseases such as type 2 diabetes, metabolic dysfunction-associated steatotic liver disease, metabolic dysfunction-associated steatohepatitis and cirrhosis, providing a clinical perspective on how these changes affect drug uptake and metabolism. Generally, a decrease in expression and/or activity of many enzymes of the cytochrome P450 family (e.g. CYP2E1 and CYP3A4), and of influx and efflux transporters (e.g. organic anion-transporting polypeptide [OATP]1B1, OATP2B1, OAT2 and bile salt export pump), has been recently documented in patients with liver disease. Decreased enzyme levels often correlate with increased severity of chronic liver disease. In subjects with hepatic impairment, there is potential for strong alterations of drug pharmacokinetics due to reduced absorption, increased volume of distribution, metabolism and extraction. Due to the altered pharmacokinetics, specific drug-drug interactions are also a potential issue to consider in patients with liver disease. Given the huge burden of liver disease in western societies, there is a need to improve awareness among all healthcare professionals and patients with liver disease to ensure appropriate drug prescriptions.

Honey Mushroom, Armillaria mellea (Agaricomycetes) and Its Fermentation Products Target Regulation of OAT1/OAT3 Proteins to Reduce Hyperuricemia in Mice.

BACKGROUND: Disorders of purine metabolism are the main cause of hyperuricemia. Current drugs for the treatment of hyperuricemia usually cause a degree of cardiovascular damage. METHODS: This study aimed to investigate the therapeutic effects of Armillaria mellea fruiting body (AFB), Armillaria rhizomorph (AR) and Armillaria mellea fermentation product (after rhizomorphs removal) (AFP) on hyperuricemic mice. The hyperuricemia mouse model was established by oral administration of potassium oxonate 0.9 g⋅kg-1 and hypoxanthine 0.5 g⋅kg-1 for two weeks. Starting from the third week, the intragastric administration of the intervention drug group was as follows: Allopurinol 0.013 g⋅kg-1, AFB (3.9 and 7.8 g⋅kg-1), AR (3.9 and 7.8 g⋅kg-1), AFP (1.95 and 3.9 g⋅kg-1) once daily for 14 days. RESULTS: Results showed that AFB, AR, and AFP reduced the contents of serum uric acid, serum creatinine, and blood urea nitrogen in hyperuricemic mice and the mechanism of action might be through up-regulation of the expression levels of organic anion transporter 1/organic anion transporter 3 proteins in kidney tissue. AR and AFP both exhibited better uric acid-lowering effects than AFB, which may be due to the higher purine content of AFB. CONCLUSIONS: Armillaria mellea and its fermentation products can treat hyperuricemia by up-regulating OAT1 protein and OAT3 protein, reducing uric acid content in mice.

Organic anion transporting polypeptide 3a1 is a novel influx pump for Perfluorooctane sulfonate in Sertoli cells and contributes to its reproductive toxicity.

Persistent organic pollutant perfluorooctane sulfonate (PFOS) is strongly associated with male reproductive disorders, but the related mechanisms are still not fully understood. In this study, we used in vivo and in vitro models to explore the role of organic anion transporting polypeptide 3a1 (Oatp3a1) on PFOS-induced male reproductive injury. Thirty male C57BL/6 (B6) mice were orally given PFOS (0-10 mg/kg/bw) for 28 days. Body weight, organ index, sperm count, histology, and blood-testis barrier (BTB) integrity were evaluated. Primary Sertoli cells were used to describe the related molecular mechanisms of male reproductive injury caused by PFOS. Our results showed that PFOS induced a decrease in sperm count, morphological damage to testicular Sertoli cells, and disruption of BTB. In the in vitro model, exposure to PFOS significantly increased Oatp3a1 mRNA and protein levels and decreased miR-23a-3p expression in Sertoli cells, accompanied by reduced trans-epithelial electrical resistance (TEER) value. By performing the 14C-PFOS uptake experiment, we showed that 14C-PFOS uptake in HEK293-Oatp3a1 cells was apparently higher than in HEK293-MOCK cells. Meanwhile, treating Sertoli cells with Oatp3a1 siRNA significantly decreased Oatp3a1 expression and rescued PFOS-induced decreases in TEER value. As such, the present study highlights that Oatp3a1 may play an important role in the toxic effect of PFOS on Sertoli cells, advancing our understanding of molecular mechanisms for PFOS-induced male reproductive disorders.

[Development of Prediction Methods for Drug-Drug Interactions Based on the Construction of Physiologically Based Pharmacokinetic Models for Hepatic OATP Substrate Drugs and Endogenous Substrates].

Quantitative prediction of the potential for drug-drug interaction (DDI) is essential to guarantee the safety and efficacy of drugs. DDI screening, modeling, and prediction is standard practice in the pharmaceutical industry. This review describes our work on (1) the establishment of a standard framework for determining physiologically based pharmacokinetic (PBPK) model structures and parameters useful for quantitatively analyzing DDIs via hepatic organic anion transporting polypeptides (OATPs). By analyzing clinically observed DDIs involving several statins as substrates, and cyclosporin A and rifampicin as inhibitors, similar in vivo inhibition constants for OATPs by each inhibitor were obtained, regardless of the substrate. (2) We took a PBPK modeling-based approach to define rate-determining processes in hepatic elimination of several OATPs and CYP3A dual substrates using our clinical DDI data with specific inhibitors for OATPs and CYP3A. Essential in vivo parameters (the passive diffusion/active transport ratio in the uptake, and the fraction of intrinsic clearance in the total drug elimination from the hepatocytes) dominating the rate-determining process in hepatic elimination were estimated quantitatively. (3) Finally, using our clinical DDI data with rifampicin, we established a PBPK model for coproporphyrin I (CP-I), which is expected to act as an endogenous substrate (biomarker) supporting the prediction of DDI involving hepatic OATPs. Our PBPK modeling-based approach with several in vitro experiments using CP-I and OATP probe substrates (statins) demonstrated the usefulness of the translation of the effect of an OATP inhibitor on CP-I pharmacokinetics into that on OATP probe substrates in drug discovery and development.

Pulmonary inflammation caused by cigarette smoke combined with lipopolysaccharide up-regulated OATP2B1 in rat lung tissue and pulmonary epithelial cells.

Organic anion transport polypeptide 2B1 (OATP2B1), as an uptake transporter, is involved in the transport of many related substrate drugs and endogenous substances in the lungs. A large amount of data shows that cigarette smoke plays an important role in the occurrence and development of lung diseases such as chronic obstructive pulmonary disease (COPD), asthma and bronchitis. However, the effect of cigarette smoke combined with lipopolysaccharide-induced pulmonary inflammation on the expression of OATP2B1 is not clear. In this study, we used cigarette smoke combined with lipopolysaccharide to establish a lung inflammation model in vivo and in vitro to explore the effect of inflammation on the expression of OATP2B1. Our study found that cigarette smoke combined with lipopolysaccharide-induced pulmonary inflammation upregulated the mRNA and protein expression of OATP2B1 and related inflammatory factors, and the expression level of related proteins was higher with the aggravation of inflammation. The experimental results of animals in vivo were consistent with those of cells in vitro. In summary, these findings provide a model and basis for a follow-up study of the mechanism of OATP2B1 in pulmonary inflammation.

The System Profile of Renal Drug Transporters in Tubulointerstitial Fibrosis Model and Consequent Effect on Pharmacokinetics.

With the widespread clinical use of drug combinations, the incidence of drug-drug interactions (DDI) has significantly increased, accompanied by a variety of adverse reactions. Drug transporters play an important role in the development of DDI by affecting the elimination process of drugs in vivo, especially in the pathological state. Tubulointerstitial fibrosis (TIF) is an inevitable pathway in the progression of chronic kidney disease (CKD) to end-stage renal disease. Here, the dynamic expression changes of eleven drug transporters in TIF kidney have been systematically investigated. Among them, the mRNA expressions of Oat1, Oat2, Oct1, Oct2, Oatp4C1 and Mate1 were down-regulated, while Oat3, Mrp2, Mrp4, Mdr1-α, Bcrp were up-regulated. Pearson correlation analysis was used to analyze the correlation between transporters and Creatinine (Cr), OCT2 and MATE1 showed a strong negative correlation with Cr. In contrast, Mdr1-α exhibited a strong positive correlation with Cr. In addition, the pharmacokinetics of cimetidine, ganciclovir, and digoxin, which were the classical substrates for OCT2, MATE1 and P-glycoprotein (P-gp), respectively, have been studied. These results reveal that changes in serum creatinine can indicate changes in drug transporters in the kidney, and thus affect the pharmacokinetics of its substrates, providing useful information for clinical use.

Biological effects and activity optimization of small-molecule, drug-like synthetic anion transporters.

The balance of normal anion concentrations in cells provides basis for maintaining cellular morphology and function. Disrupting the homeostasis of cellular anions and lysosomal pH, in particular with high selectivity for cancer cells over normal cells may serve as a promising approach for the treatment of cancers. Small-molecule organic compounds with transmembrane anion transport activity, namely synthetic anion transporters are able to destroy the homeostasis of cellular anions, in particular chloride anions to trigger cell death and thus may be developed as a new class of anti-tumor drugs. This paper reviews the latest advance in the investigation into the in vitro anion transport, promising anti-tumor activity and probable mechanism of biological action of synthetic anion transporters. The strategies for optimizing the biological activity of synthetic anion transporters and improving the selectivity for cancer cells over normal cells are also discussed.

Synthetic Anion Transporters as Endoplasmic Reticulum (ER) Stress Inducers.

Cl--ion transporters (2a-2h) were synthesized based on the binding motifs of prodigiosin. Transporter 2e clearly displays Cl--ion transportation activity across both model and live cell membranes. Furthermore, 2e can disrupt Ca2+ homeostasis and increase the intracellular concentration of Ca2+ in the DLD-1 cell. This disruption can lead to Caspase-dependent apoptosis supported by CHOP expression (a marker of ER stress) and the appearance of the cleaved forms of Caspase 3 and PARP.

Treatment with proteasome inhibitor bortezomib decreases organic anion transporting polypeptide (OATP) 1B3-mediated transport in a substrate-dependent manner.

OATP1B1 and OATP1B3 mediate hepatic uptake of many drugs (e.g., statins) and can mediate transporter-mediated drug-drug-interactions (DDIs). Bortezomib is the first-in-class proteasome inhibitor drug approved by the U. S. Food and Drug Administration for the treatment of multiple myeloma. The potential of bortezomib to cause OATP-mediated DDIs has not been assessed. The current study investigated the involvement of the ubiquitin-proteasome system (UPS) in OATP1B1 and OATP1B3 degradation and determined the effects of proteasome inhibitors on OATP1B1- and OATP1B3-mediated transport. Co-immunoprecipitation of FLAG-OATP1B1/1B3 and HA-ubiquitin was observed in human embryonic kidney (HEK) 293 cells co-transfected with FLAG-tagged OATP1B1/OATP1B3 and hemagglutinin (HA)-tagged ubiquitin, suggesting that OATP1B1 and OATP1B3 can be ubiquitin-modified. Although blocking proteasome activity by bortezomib treatment (50 nM, 7 h) increased the endogenous ubiquitin-conjugated FLAG-OATP1B1 and FLAG-OATP1B3 in HEK293-FLAG-OATP1B1 and-OATP1B3 cells, such treatment did not affect the total protein levels of OATP1B1 and OATP1B3, suggesting that the UPS plays a minor role in degradation of OATP1B1 and OATP1B3 under current constitutive conditions. Pretreatment with bortezomib (50-250 nM, 2-7 h) significantly decreased transport of [3H]CCK-8, a specific OATP1B3 substrate, in HEK293-OATP1B3 and human sandwich-cultured hepatocytes (SCH). However, bortezomib pretreatment had negligible effects on the transport of [3H]E217ßG and [3H]pitavastatin, dual substrates of OATP1B1 and OATP1B3, in HEK293-OATP1B1/1B3 cells and/or human SCH. Compared with vehicle control treatment, bortezomib pretreatment significantly decreased the maximal transport velocity (Vmax) of OATP1B3-mediated transport of CCK-8 (92.25 ± 14.2 vs. 133.95 ± 15.5 pmol/mg protein/min) without affecting the affinity constant (Km) values. Treatment with other proteasome inhibitors MG132, epoxomicin, and carfilzomib also significantly decreased OATP1B3-mediated [3H]CCK-8 transport. In summary, the current studies for the first time report ubiquitination of OATP1B1 and OATP1B3 and the apparent substrate-dependent inhibitory effect of bortezomib on OATP1B3-mediated transport. The data suggest that bortezomib has a low risk of causing OATP-mediated DDIs.

High impact of Oatp1a/1b transporters on in vivo disposition of the hydrophobic anticancer drug paclitaxel.

PURPOSE: Organic anion-transporting polypeptides (OATP) mediate the cellular uptake of a broad range of drugs. The hydrophobic anticancer drug, paclitaxel (PTX), was recently identified as a substrate for OATP1B3 in vitro. We investigated the role of Oatp1a/1b transporters in the pharmacokinetics of PTX in vivo, as well as their impact at different dose levels of PTX and methotrexate (MTX). EXPERIMENTAL DESIGN: Recently generated Slco1a/1b(-/-) (lacking all Oatp1a/1b transporters) and wild-type mice were intravenously dosed with 2, 10, or 50 mg/kg of PTX, or with 10, 50, or 500 mg/kg of MTX, and plasma and tissue drug concentrations were measured. RESULTS: In spite of its hydrophobicity, PTX systemic exposure (at 10 mg/kg) was increased by greater than 2-fold in Slco1a/1b(-/-) mice compared with wild-type, whereas PTX liver uptake was reduced by about 2-fold. Oatp1a/1b transporters displayed a high impact on PTX and MTX pharmacokinetics over a broad dose range. For MTX, even at 500 mg/kg, saturation of Oatp1a/1b was not observed, with a 3.4-fold increase in plasma and 30-fold decrease in liver levels in Slco1a/1b(-/-) mice compared with wild-type. Although beginning saturation of Oatp1a/1b was observed at the highest dose of PTX, plasma levels in Slco1a/1b(-/-) mice were still 1.7-fold increased and liver levels 1.5-fold decreased compared with wild-type. CONCLUSION: Oatp1a/1b transporters play a pronounced role in determining plasma levels and tissue distribution of MTX and PTX, thus affecting even highly hydrophobic drugs. Variation in OATP1A/1B transporter activity, due to genetic variation, inhibition, and/or tumor expression might affect toxicity and therapeutic efficacy of these anticancer drugs.

Interactions of human organic anion transporters with aristolochic acids.

Aristolochic acids (AAs), contained in Chinese herbal preparations, have been considered to induce nephropathy. In order to elucidate the molecular mechanisms of AA-induced nephrotoxicity, we have elucidated the interaction of human organic anion transporters (hOATs) with AAs using their stable cell lines. AA-I and AA-II inhibited organic anion uptake by hOAT1, hOAT3, and hOAT4 in dose-dependent manners. Treatment of hOAT3 with AA-I resulted in a significant reduction in viability compared with that of mock, which was rescued by the organic anion transport inhibitor probenecid. In conclusion, hOAT3-mediated AA-I uptake may be associated with the induction of nephrotoxicity.

Influence of SLCO1B1, 1B3, 2B1 and ABCC2 genetic polymorphisms on mycophenolic acid pharmacokinetics in Japanese renal transplant recipients.

OBJECTIVE: We investigated the association between mycophenolic acid (MPA) pharmacokinetics and organic anion-transporting polypeptide (OATP/SLCO)1B1, 1B3, 2B1 and multidrug resistance-association protein 2 (MRP2/ABCC2) genetic polymorphisms and diarrhea. METHODS: Eighty-seven renal allograft recipients were given repeated doses of mycophenolate mofetil every 12 h at a designated time (09:00 and 21:00). The pharmacokinetics of MPA were analyzed on day 28 posttransplantation. RESULTS: The dose-adjusted area under the cuve (AUC)(6-12) of MPA, an estimate of enterohepatic recirculation, was greater in SLCO1B3 T334G GG (or G699A AA) carriers than in TT carriers (or G699A GG) (40 vs. 25 ng h/mL per milligram, respectively, P = 0.0497). None of the polymorphism of SLCO1B1, SLCO2B1, or ABCC2 C-24T were associated with MPA pharmacokinetics or diarrhea. However, the oral clearance of MPA in recipients having both the SLCO1B3 T334G GG genotype and the ABCC2 C-24T T allele was significantly lower than in patients having both the SLCO1B3 T334G TT and ABCC2 C-24T CC genotypes (0.15 vs. 0.18 L/h per kilogram, respectively, P = 0.0010). CONCLUSIONS: MPA excretion into bile in patients with SLCO1B3 T334G GG (or G699A AA) was higher than in those with T334G TT (or G699A GG), probably resulting in a higher AUC(6-12) value of MPA. MPA uptake into hepatocytes and excretion into bile at first pass may be greater in SLCO1B3 T334G GG carriers than in TT carriers. In addition, the ABCC2 C-24T polymorphism also seems to be associated with enhanced enterohepatic circulation of MPA. The SLCO1B3 and ABCC2 transporters rather than uridine diphosphate-glucuronosyltransferase (UGT) may partly affect interindividual variety in plasma MPA concentration.

Expression of rat Multidrug Resistance Protein 2 (Mrp2) in male and female rats during normal and pregnenolone-16alpha-carbonitrile (PCN)-induced postnatal ontogeny.

The normal maturation of biliary organic anion excretion in newborn rats can be enhanced by microsomal enzyme-inducing chemical treatment, yet the mechanism for this phenomenon is not known. Multidrug Resistance Protein 2 (Mrp2) is a biliary efflux transporter that is inducible by select microsomal enzyme-inducing chemicals. Thus, the aims of this study were to compare the normal and pregnenolone-16alpha-carbonitrile (PCN)-induced postnatal ontogeny of Mrp2 in male and female rats. Mrp2 protein increased in an age-dependent manner in both sexes between 0 and 90 days of age. At birth, Mrp2 protein in both male and female rats was the same, approximately 70% of adult levels. Mrp2 protein in both sexes reached maximal expression levels that were higher than adult levels (male: days 25-40; female: day 45), then decreased to adult levels, at which age Mrp2 protein expression in male and female rats was the same. Second, male and female rats of various ages were treated with PCN (75 mg/kg, ip) or corn oil for 4 days, after which livers were removed and analyzed for Mrp2 protein and mRNA expression. PCN accelerated the expression of Mrp2 protein in male and female rats as early as 10 days of age, whereas, PCN did not affect male and female Mrp2 mRNA ontogeny. These data suggest that PCN increased Mrp2 protein by a sex-independent posttranscriptional mechanism.