依诺沙星对大鼠肝药酶的抑制作用

在肝微粒体水平上，采用依诺沙里４００ｍｇ／ｋｇ，灌胃ｑｄ×７ｄ能显著减少大鼠肝微粒体细胞色素Ｐ４５０。含量，经依诺沙星处理的大鼠肝微粒体中氨基比林－Ｎ－脱甲基酶、ＮＡＤＰＨ－细胞色素Ｃ还原酶、７－乙氧基香豆素－Ｏ－脱乙基酶、苯并芘羟化酶、戊巴比妥侧链羟化酶的活性均显著减弱。可见，依诺沙星对大鼠肝药酶具有广泛的抑制作用。     

Comparison of mRNA expression profiles of drug-metabolizing enzymes and transporters in fresh and cryopreserved cynomolgus monkey hepatocytes

In this study, freshly isolated and cryopreserved cynomolgus monkey hepatocytes were seeded on Cell-able^® plates with feeder cells to form spheroids and were cultured for 28 days. As a control, hepatocytes were also cultured with or without feeder cells on collagen-coated plates. We verified the mRNA expression levels of drug-metabolizing enzyme-related genes and the leakage of enzymes (AST, ALT, LDH, and γ-GTP) as indicators of cell survival. As a result, the patterns of target mRNA expression in fresh and cryopreserved hepatocytes were very similar during the culture period between culture methods. mRNA expression levels were highly maintained at day 28 using the 3D spheroid and co-culture methods, demonstrating that these methods are useful for maintenance of liver function. Leakage of AST and ALT was higher at day 3 but decreased at day 14. LDH was not detected, suggesting that the cell viability was also maintained during the culture period. Furthermore, the functional differences between fresh and cryopreserved hepatocytes were not clearly detected. The co-culture method was useful for long-term culture not requiring 3D structure, and the 3D spheroid culture method was effective as well. With these techniques, cynomolgus monkey hepatocytes are expected to exhibit smaller individual differences and high reproducibility.     

大黄酸对药物转运体和代谢酶影响的研究进展

药物转运体和药物代谢酶是影响药物体内处置过程中至关重要的因素。大黄酸作为传统中药大黄的主要活性成分,具有广泛药理活性。研究发现,大黄酸与药物转运体和代谢酶密切相关,能够直接激活或抑制多种转运体的功能及其蛋白表达。而且大黄酸对药物代谢酶细胞色素P450（CYP450）的功能及其蛋白表达同样有抑制作用。因此,大黄酸与其他药物合用时,可能发生基于药动学的药物相互作用（drug-drug interaction,DDI）。从药物转运体和代谢酶的体内分布、大黄酸对转运体及代谢酶的影响等方面进行综述。     

Lactate transporters as therapeutic targets in cancer and inflammatory diseases

Introduction: Inflammation is associated with the accumulation of lactate at sites of tumor-growth and inflammation. Lactate initiates tissue-responses contributing to disease. We discuss the potential of targeting lactate transporters in the treatment of cancer and inflammatory conditions.

Areas covered: Lactate is the end product of glycolysis, often considered a waste metabolite but also a fuel for oxidative cells. It is however an active signaling molecule with immunomodulatory and angiogenic properties. They are the consequence of lactate binding to membrane receptor(s) or being transported through specific carrier-mediated-transporters across the cellular membrane. Carriers are distinct in proton-linked-monocarboxylate-transporters (MCTs) and Na⁺-coupled- electrogenic-transporters, expressed by several tissues including immune-system, endothelium and epithelium. Several tumors and inflammatory sites show accumulation of lactate and altered expression of its transporters, thus suggesting a role of this metabolite in cancer and inflammation. We review the most recent evidence on lactate biology, focusing on transporter expression and function in health and disease.

Expert opinion: Lactate-initiated signaling is gaining attention for its implications in cancer and inflammation. This review discusses the therapeutic potential of targeting lactate transporters and drugs that are already in clinical use for cancer and discusses the opportunity to develop new therapeutics for inflammation and cancer.     

Expression and inducibility of drug-metabolizing enzymes in preneoplastic and neoplastic lesions of rat liver during nitrosamine-induced hepatocarcinogenesis

The expression, inducibility, and regulation of four different cytochrome (cyt.) P-450 isoenzymes (PB₁, PB₂, MC₁, and MC₂) NADPH-cytochrome P-450 reductase, the glutathione transferases (GSTs) B and C and microsomal epoxide hydrolase (mEH_b) have been studied during nitrosamine-induced hepatocarcinogenesis using immunohistochemical techniques. The investigations revealed basic differences in the expression of the individual drug metabolizing enzymes in the course of neoplastic development. While the two GSTs and mEH_b were increased in all preneoplastic and benign neoplastic lesions, the levels of the distinct cyt. P-450 isoenzymes were characteristically different from each other. Following initial changes in the expression of these enzymes in early preneoplastic lesions (i. e., increase of cyt. P-450 PB₁ versus slight decrease of the other cyt. P-450 isoenzymes), a continuous reduction of all cyt. P-450 isoenzymes was observed during the further course of hepatocarcinogenesis. In progressed neoplastic nodules, all cyt. P-450-isoenzymes and NADPH cyt. P-450 reductase were decreased to varying extents.Treatment of animals with inducers of the monooxygenase system, such as phenobarbital, 3-methylcholanthrene and polychlorinated biphenyls, led to a rather heterogenous pattern of enzyme alterations in preneoplastic and neoplastic lesions. Following administration of phenobarbital, some islets responded to the same degree as the surrounding tissue, others were less or not at all inducible and a few of the lesions showed a prominent increase in cyt. P-450 PB₂ and NADPH-cyt. P-450 reductase levels. The interesting finding that these two enzymes always showed concurrent changes may be indicative of a common regulation. Similar to phenobarbital, an induction of cyt. P-450 isoenzymes within carcinogen-induced lesions was also observed following administration of 3-methyl-cholanthrene or polychlorinated biphenyls.The results demonstrate that drug-metabolizing enzymes are abnormally regulated in carcinogen-induced lesions. The multiplicity of enzyme deviations within individual lesions and especially the enzyme inducibility strongly suggest that the focal enzyme alterations result from genotoxic effects of the carcinogen on regulatory systems of a higher order rather than from mutational events in individual structural genes.Dedicated to Professor Dr. Herbert Remmer on the occasion of his 65th birthday     

Nuclear receptors in the multidrug resistance through the regulation of drug-metabolizing enzymes and drug transporters

Chemotherapy is one of the three most common treatment modalities for cancer. However, its efficacy is limited by multidrug resistant cancer cells. Drug metabolizing enzymes (DMEs) and efflux transporters promote the metabolism, elimination, and detoxification of chemotherapeutic agents. Consequently, elevated levels of DMEs and efflux transporters reduce the therapeutic effectiveness of chemotherapeutics and, often, lead to treatment failure. Nuclear receptors, especially pregnane X receptor (PXR, NR1I2) and constitutive androstane activated receptor (CAR, NR1I3), are increasingly recognized for their role in xenobiotic metabolism and clearance as well as their role in the development of multidrug resistance (MDR) during chemotherapy. Promiscuous xenobiotic receptors, including PXR and CAR, govern the inducible expressions of a broad spectrum of target genes that encode phase I DMEs, phase II DMEs, and efflux transporters. Recent studies conducted by a number of groups, including ours, have revealed that PXR and CAR play pivotal roles in the development of MDR in various human carcinomas, including prostate, colon, ovarian, and esophageal squamous cell carcinomas. Accordingly, PXR/CAR expression levels and/or activation statuses may predict prognosis and identify the risk of drug resistance in patients subjected to chemotherapy. Further, PXR/CAR antagonists, when used in combination with existing chemotherapeutics that activate PXR/CAR, are feasible and promising options that could be utilized to overcome or, at least, attenuate MDR in cancer cells.     

microRNA介导低氧对药物代谢酶和转运体的调控

低氧条件下机体的循环系统、神经系统、内分泌系统等的功能发生显著改变,这些变化影响药物在体内的吸收、分布、代谢和排泄。药物代谢酶和转运体是影响药物代谢的主要因素,微小RNA (microRNA, miRNA)除调控与药物代谢相关的基因如缺氧诱导因子、炎症因子、核受体等,还可直接作用于药物代谢酶和转运体,影响药物的体内代谢。本文通过综述低氧对miRNA及药物代谢酶和转运体的调节, miRNA调控药物代谢酶和转运体及药物代谢相关基因,低氧调节药物代谢酶和转运体的相关机制等,探讨miRNA在低氧调节药物代谢酶和转运体中的作用,提出以miRNA为核心的低氧影响药物代谢的分子机制。     

大蒜对药物代谢酶及转运体活性影响研究进展

作为天然药物的一种,大蒜包含多种具有药理活性的有效成分,在临床应用日益广泛。然而,大蒜长期应用,可能对肝脏药物代谢酶及体内药物转运体的活性产生诱导或抑制效应,从而引起自身或其他合用药物代谢的改变和导致药物相互作用的发生。     

Regulation of drug-metabolizing enzymes by local and systemic liver injuries

Introduction: Drug metabolism and disposition are critical in maintaining the chemical and functional homeostasis of xenobiotics/drugs and endobiotics. The liver plays an essential role in drug metabolism and disposition due to its abundant expression of drug-metabolizing enzymes (DMEs) and transporters. There is growing evidence to suggest that many hepatic and systemic diseases can affect drug metabolism and disposition by regulating the expression and/or activity of DMEs and transporters in the liver.

Areas covered: This review focuses on the recent progress on the regulation of DMEs by local and systemic liver injuries. Liver ischemia and reperfusion (I/R) and sepsis are used as examples of local and systemic injury, respectively. The reciprocal effect of the expression and activity of DMEs on animals’ sensitivity to local and systemic liver injuries is also discussed.

Expert opinion: Local and systemic liver injuries have a major effect on the expression and activity of DMEs in the liver. Understanding the disease effect on DMEs is clinically important due to the concern of disease-drug interactions. Future studies are necessary to understand the mechanism by which liver injury regulates DMEs. Human studies are also urgently needed in order to determine whether the results in animals can be replicated in human patients.     

Effect of commercially available green and black tea beverages on drug-metabolizing enzymes and oxidative stress in Wistar rats

The effect of commercially available green tea (GT) and black tea (BT) drinks on drugmetabolizing enzymes (DME) and oxidative stress in rats was investigated. Male Wistar rats were fed a laboratory chow diet and GT or BT drink for 5 weeks. Control rats received de-ionized water instead of the tea drinks. Rats received the GT and BT drinks treatment for 5 weeks showed a significant increase in hepatic microsomal cytochrome P450 (CYP) 1A1 and CYP1A2, and a significant decrease in CYP2C, CYP2E1 and CYP3A enzyme activities. Results of immunoblot analyses of enzyme protein contents showed the same trend with enzyme activity. Significant increase in UDP-glucuronosyltransferase activity and reduced glutathione content in liver and lungs were observed in rats treated with both tea drinks. A lower lipid peroxide level in lungs was observed in rats treated with GT drink. Electrophoretic mobility shift assay revealed that both tea drinks decreased pregnane X receptor binding to DNA and increased nuclear factor-erythroid 2 p45-related factor 2 binding to DNA. These results suggest that feeding of both tea drinks to rats modulated DME activities and reduced oxidative stress in liver and lungs. GT drink is more effective on reducing oxidative stress than BT drink.     

Proteomic analysis of small intestinal epithelial cells in antibiotic-treated mice: Changes in drug transporters and metabolizing enzymes

Antibiotics act on bacterial flora originally present in the intestine, and changes in the intestinal flora have various effects on the host. This study investigated changes in the protein levels of drug transporters and metabolizing enzymes in the small intestines of antibiotic-treated mice by proteomic analysis. After the oral administration of non-absorbable antibiotics (vancomycin and polymyxin B) for 5 days, 15 drug transporter or metabolizing enzyme proteins had significantly changed levels among 1780 proteins identified in small intestinal epithelial cells. Of these, the levels of peptide transporter 1 (Pept1), multidrug resistance protein 1a (Mdr1a), and multidrug resistance-associated protein 2 (Mrp2) were increased approximately 2-fold. In addition, the levels of two Cyp4f proteins were decreased and those of Cyp4b1, Ces1d, and three glutathione S-transferase (Gst) proteins were increased. Our results indicate that the oral administration of antibiotics changes the levels of proteins related to the absorption and metabolism of drugs in the small intestine, and suggest that substrate drugs of these proteins have a risk for indirect drug interactions with antibacterial drugs via the intestinal flora.     

Emerging role of NRF2 in chemoresistance by regulating drug-metabolizing enzymes and efflux transporters

Chemoresistance is a disturbing barrier in cancer therapy, which always results in limited therapeutic options and unfavorable prognosis. Nuclear factor E2-related factor 2 (NRF2) controls the expression of genes encoding cytoprotective enzymes and transporters that protect against oxidative stress and electrophilic injury to maintain intrinsic redox homeostasis. However, recent studies have demonstrated that aberrant activation of NRF2 due to genetic and/or epigenetic mutations in tumor contributes to the high expression of phase I and phase II drug-metabolizing enzymes, phase III transporters, and other cytoprotective proteins, which leads to the decreased therapeutic efficacy of anticancer drugs through biotransformation or extrusion during chemotherapy. Therefore, a better understanding of the role of NRF2 in regulation of these enzymes and transporters in tumors is necessary to find new strategies that improve chemotherapeutic efficacy. In this review, we summarized the recent findings about the chemoresistance-promoting role of NRF2, NRF2-regulated phase I and phase II drug-metabolizing enzymes, phase III drug efflux transporters, and other cytoprotective genes. Most importantly, the potential of NRF2 was proposed to counteract drug resistance in cancer treatment.     

Nuclear receptors and the regulation of drug-metabolizing enzymes and drug transporters: implications for interindividual variability in response to drugs

Erratic or unpredictable response to drugs remains a challenge of modern drug therapy. An important determinant of such interindividual differences in drug response is variability in the expression of drug-metabolizing enzymes and/or transporters at sites of absorption and/or tissue distribution. Variable drug-metabolizing enzyme and transporter expression can result in unpredictable exposure and tissue distribution of drugs and may manifest as adverse effects or therapeutic failure. In the past decade, important new insights have been made relating to the regulatory mechanisms governing the expression of drug-metabolizing enzymes and transporters by ligand-activated nuclear receptors. Specifically, there is compelling evidence to demonstrate that PXR, CAR, FXR, LXR, VDR, HNF4alpha, and AhR form a battery of nuclear receptors that regulate the expression of many important drug-metabolizing enzyme and transporters. In this review, the authors focus on clinically important drug-metabolizing enzymes such as CYP3A4, CYP2B6, CYP2C9, CYP2C19, UGT1A1, SULT2A1, and glutathione S-transferases and their regulation by nuclear receptors. They also review the nuclear receptor-mediated regulation of drug transporters such as MDR1, MRP2, MRP4, BSEP, BCRP, NTCP, OATP1B3, and OATP1A2. Finally, they outline how the drug development process has been affected by the current understanding of the involvement of nuclear receptors in the regulation of drug disposition genes.     

Coordinate regulation of human drug-metabolizing enzymes, and conjugate transporters by the Ah receptor, pregnane X receptor and constitutive androstane receptor

Coordinate regulation of Phase I and II drug-metabolizing enzymes and conjugate transporters by nuclear receptors suggests that these proteins evolved to an integrated biotransformation system. Two major groups of ligand-activated nuclear receptors/xenosensors evolved: the Ah receptor (activated by aryl hydrocarbons and drugs such as omeprazole) and type 2 steroid receptors such as PXR and CAR, activated by drugs such as rifampicin, carbamazepin and phenytoin. It is increasingly recognized that there is considerable cross-talk between these xenosensors. Therefore, an attempt was made to discuss biotransformation by the Ah receptor together with that of PXR and CAR. Due to considerable species differences the emphasis is on human biotransformation. Agonists coordinately induce biotransformation due to common xenosensor-binding response elements in the regulatory region of target genes. However, whereas different groups of xenobiotics appear to more selectively stimulate CYPs (Phase I), their regulatory control largely converged in modulating Phase II metabolism and transport. Biotransformation appears to be tightly controlled to achieve efficient homeostasis of endobiotics and detoxification of dietary phytochemicals, but nuclear receptor agonists may also lead to potentially harmful drug interactions.     

Effects of atorvastatin metabolites on induction of drug-metabolizing enzymes and membrane transporters through human pregnane X receptor

BACKGROUND AND PURPOSE

Atorvastatin metabolites differ in their potential for drug interaction because of differential inhibition of drug-metabolizing enzymes and transporters. We here investigate whether they exert differential effects on the induction of these genes via activation of pregnane X receptor (PXR) and constitutive androstane receptor (CAR).

EXPERIMENTAL APPROACH

Ligand binding to PXR or CAR was analysed by mammalian two-hybrid assembly and promoter/reporter gene assays. Additionally, surface plasmon resonance was used to analyse ligand binding to CAR. Primary human hepatocytes were treated with atorvastatin metabolites, and mRNA and protein expression of PXR-regulated genes was measured. Two-hybrid co-activator interaction and co-repressor release assays were utilized to elucidate the molecular mechanism of PXR activation.

KEY RESULTS

All atorvastatin metabolites induced the assembly of PXR and activated CYP3A4 promoter activity. Ligand binding to CAR could not be proven. In primary human hepatocytes, the para-hydroxy metabolite markedly reduced or abolished induction of cytochrome P450 and transporter genes. While significant differences in co-activator recruitment were not observed, para-hydroxy atorvastatin demonstrated only 50% release of co-repressors.

CONCLUSIONS AND IMPLICATIONS

Atorvastatin metabolites are ligands of PXR but not of CAR. Atorvastatin metabolites demonstrate differential induction of PXR target genes, which results from impaired release of co-repressors. Consequently, the properties of drug metabolites have to be taken into account when analysing PXR-dependent induction of drug metabolism and transport. The drug interaction potential of the active metabolite, para-hydroxy atorvastatin, might be lower than that of the parent compound.     

孕烷X受体介导的代谢酶和转运体转录调控及其在化疗药物耐药中的作用

孕烷X受体(PXR,NR1I2)是生物体内药物代谢酶和转运体基因表达的主要调控因子之一.近来研究发现,PXR介导的药物代谢酶和转运体的过表达,与化疗药物多药耐药的产生密切相关.鉴于PXR在药物代谢酶和转运体调控中的重要性和PXR转录调控的多样性,有必要对其导致的多药耐药形成机制进行更深入的研究.本文综述了PXR介导的代谢酶和转运体基因表达调控机制,及其引起化疗药物多药耐药的相关研究进展,为提高化疗药物敏感性、逆转化疗药物的多药耐药提供有效的治疗策略.     

Effects of renal diseases on the regulation and expression of renal and hepatic drug-metabolizing enzymes: a review

1. The activity of drug-metabolizing enzymes (DMEs) in extrahepatic organs is highest in the kidneys. Generally, the kidneys contain most, if not all, of the DMEs found in the liver. Surprisingly, some of these DMEs show higher activity in the kidneys than in the liver. 2. Most of the renal DMEs are localized in the cortex of the kidneys, especially in the proximal tubules. DMEs are also found in the distal tubules and collecting ducts. 3. Renal diseases such as acute and chronic renal failure and renal cell carcinoma alter the regulation of both hepatic and extrahepatic phase I and II DMEs. Changes in the expression of these DMEs seem to be tissue and species specific. 4. Generally, there is significant down-regulation of most of the phase I and a few of phase II DMEs at the protein, mRNA and activity levels. Unfortunately, the mechanisms leading to the alteration in DMEs in renal diseases remain unclear, although many theories have been made. 5. The presence of some circulating factors such as cytokines, nitric oxide, parathyroid hormones and increased intracellular calcium play a role in the regulation of DMEs in renal diseases.     

药物转运体和代谢酶在抗生素药动学中的作用

近年来,对体内药物转运体的研究取得了重大进展,越来越多的转运体被发现及研究,其对药物的跨膜转运,具有重要的意义。各种转运体包括摄取转运体和外排转运体对药物的体内过程以及药物相互作用均有着重要影响。研究表明大多数抗生素的体内过程都与转运体和代谢酶有关,因此,归纳总结了转运体和代谢酶在抗生素的药动学和药物相互作用中的最新研究进展,为临床合理用药提供参考。