Expression Of Drug-metabolizing Enzymes Research Articles

Circadian Clock-Controlled Drug Metabolism: Implications for Chronotherapeutics.

Dependence of drug metabolism on dosing time has long been recognized. However, only recently are the underlying mechanisms for circadian drug metabolism being clarified. Diurnal rhythmicity in expression of drug-metabolizing enzymes is believed to be a key factor determining circadian metabolism. Supporting the notion that biological rhythms are generated and maintained by the circadian clock, a number of diurnal enzymes are under the control of the circadian clock. In general, circadian clock genes generate and regulate diurnal rhythmicity in drug-metabolizing enzymes via transcriptional actions on one or two of three cis-elements (i.e., E-box, D-box, and Rev-erb response element or RAR-related orphan receptor response element). Additionally, cycling or clock-controlled nuclear receptors such as hepatocyte nuclear factor 4α and peroxisome proliferator-activated receptor γ are contributors to diurnal enzyme expression. These newly discovered mechanisms for each of the rhythmic enzymes are reviewed in this article. We also discuss how the rhythms of enzymes are translated to circadian pharmacokinetics and drug chronotoxicity, which has direct implications for chronotherapeutics. Our discussion is also extended to two diurnal transporters (P-glycoprotein and multidrug resistance-associated protein 2) that have an important role in drug absorption. Although the experimental evidence is lacking in metabolism-based chronoefficacy, circadian genes (e.g., Rev-erbα) as drug targets are shown to account for diurnal variability in drug efficacy. SIGNIFICANCE STATEMENT: Significant progress has been made in understanding the molecular mechanisms for generation of diurnal rhythmicity in drug-metabolizing enzymes. In this article, we review the newly discovered mechanisms for each of the rhythmic enzymes and discuss how the rhythms of enzymes are translated to circadian pharmacokinetics and drug chronotoxicity, which has direct implications for chronotherapeutics.

Whole-cell dependent biosynthesis of N- and S-oxides using human flavin containing monooxygenases expressing budding yeast

Flavin containing monooxygenases (FMOs) represent one of the predominant types of phase I drug metabolizing enzymes (DMEs), and thus play an important role in the metabolism of xeno- and endobiotics for the generation of their corresponding oxides. These oxides often display biological activities, however they are difficult to study since their chemical or biological synthesis is generally challenging even though only small amounts are required to evaluate their efficacy and safety. Previously, we constructed a DME expression system for cytochrome P450, UDP-glucuronosyltransferase (UGT), and sulfotransferase (SULT) using yeast cells, and successfully produced xenobiotic metabolites in a whole-cell dependent manner. In this study, we developed a heterologous expression system for human FMOs, including FMO1–FMO5, in Saccharomyces cerevisiae and examined its N- and S-oxide productivity. The recombinant yeast cells expressed each of the FMO successfully, and the FMO4 transformant produced N- and S-oxide metabolites at several milligrams per liter within 24 h. This whole-cell dependent biosynthesis enabled the production of N- and S-oxides without the use of the expensive cofactor NADPH. Such novel yeast expression system could be a powerful tool for the production of oxide metabolites.

Acetaminophen-Induced Liver Injury Alters Expression and Activities of Cytochrome P450 Enzymes in an Age-Dependent Manner in Mouse Liver.

Drug-induced liver injury (DILI) is a global medical problem. The risk of DILI is often related to expression and activities of drug-metabolizing enzymes, especially cytochrome P450s (P450s). However, changes on expression and activities of P450s after DILI have not been determined. The aim of this study is to fill this knowledge gap. Acetaminophen (APAP) was used as a model drug to induce DILI in C57BL/6J mice at different ages of days 10 (infant), 22 (child), and 60 (adult). DILI was assessed by levels of alanine aminotransferase and aspartate aminotransferase in plasma with a confirmation by H&E staining on liver tissue sections. The expression of selected P450s at mRNA and protein levels was measured by real-time polymerase chain reaction and liquid chromatography-tandem mass spectrometry, respectively. The activities of these P450s were determined by the formation of metabolites from probe drugs for each P450 using ultraperformance liquid chromatography-quadrupole time of flight mass spectrometry. DILI was induced at mild to severe levels in a dose-dependent manner in 200, 300, and 400 mg/kg APAP-treated groups at child and adult ages, but not at the infant age. Significantly decreased expression at mRNA and protein levels as well as enzymatic activities of CYP2E1, 3A11, 1A2, and 2C29 were found at child and adult ages. Adult male mice were more susceptible to APAP-induced liver injury than female mice with more decreased expression of P450s. These results suggest that altered levels of P450s in livers severely injured by drugs may affect the therapeutic efficacy of drugs, which are metabolized by P450s, more particularly for males. SIGNIFICANCE STATEMENT: The current study in an animal model demonstrates that acetaminophen-induced liver injury results in decreased expression and enzyme activities of several examined drug-metabolizing cytochrome P450s (P450s). The extent of such decreases is correlated to the degree of liver injury severity. The generated data may be translated to human health for patients who have drug-induced liver injury with decreased capability to metabolize drugs by certain P450s.

Coordinated Regulation of UGT2B15 Expression by Long Noncoding RNA LINC00574 and hsa-miR-129-5p in HepaRG Cells.

Recent studies have shown that microRNAs and long noncoding RNAs (lncRNAs) regulate the expression of drug metabolizing enzymes (DMEs) in human hepatic cells and that a set of DMEs, including UDP glucuronosyltransferase (UGT) 2B15, is down-regulated dramatically in liver cells by toxic acetaminophen (APAP) concentrations. In this study we analyzed mRNA, microRNA, and lncRNA expression profiles in APAP-treated HepaRG cells to explore noncoding RNA-dependent regulation of DME expression. The expression of UGT2B15 and lncRNA LINC00574 was decreased in APAP-treated HepaRG cells. UGT2B15 levels were diminished by LINC00574 suppression using antisense oligonucleotides or small interfering RNA. Furthermore, we found that hsa-miR-129-5p suppressed LINC00574 and decreased UGT2B15 expression via LINC00574 in HepaRG cells. In conclusion, our results indicate that LINC00574 acts as an important regulator of UGT2B15 expression in human hepatic cells, providing experimental evidence and new clues to understand the role of cross-talk between noncoding RNAs. SIGNIFICANCE STATEMENT: We showed a molecular network that displays the cross-talk and consequences among mRNA, micro RNA, long noncoding RNA, and proteins in acetaminophen (APAP)-treated HepaRG cells. APAP treatment increased the level of hsa-miR-129-5p and decreased that of LINC00574, ultimately decreasing the production of UDP glucuronosyltransferase (UGT) 2B15. The proposed regulatory network suppresses UGT2B15 expression through interaction of hsa-miR-129-5p and LINC00574, which may be achieved potentially by recruiting RNA binding proteins.

Abstract P4-04-08: Histological and epigenetic analyses of placenta tissue from breast cancer patients and healthy participants

Abstract Background: Breast cancer is one of the most common malignancies during pregnancy. Guidelines for breast cancer treatment during pregnancy demonstrating that breast cancer during pregnancy can be treated similarly to non-pregnancy-associated breast cancer, except for hormonal and anti-HER2 therapies. Nonetheless, a decreased birth weight is often observed in newborns. Therefore, this project aims to analyze the effects of chemotherapy and the impact of cancer progression on the placenta. Methods: Placentas from breast cancer patients (n=63) and non-cancer participants (n=20) enrolled in Breast Cancer in Pregnancy (BCP) study were collected after delivery and embedded in paraffin. Sections were stained with Hematoxylin-Eosin (HE) and IHC (Ki-67, cPARP, p27Kip1); trophoblast morphology was evaluated by an established scoring system and distribution of immunohistochemical markers was assessed. Additionally, epigenetic analyses (cytosine methylation) for the following genes were performed: LINE-1 (general methylation), IGF2-H19 (birth weight), EPO (hypoxia), BDNF (neurobiological development), glucocorticoid receptor, HSD11B2 (inactivation of glucocorticoids), estrogen receptor, P-glycoprotein, CYP-3A4 (drug metabolism). Results: HE staining revealed significant damage of trophoblast nuclei and membranes in placentas from breast cancer patients compared to controls (mean score of damage 1.9/1.8 vs 0.8/0.7, p<0.001). Simultaneously, mean proliferation index (Ki-67 positive cells/ 1000 trophoblasts) was reduced (36.3 vs 58.0, p<0.001). In contrast, nuclear and cytoplasmic expression of the negative cell cycle regulator p27Kip1 was reduced (mean IRS score 1.0/0.9 vs 4.3/4.8, p<0.001). No evidence of enhanced apoptosis was found. Epigenetic analyses showed significant differences in mean percentage of cytosine methylation of EPO (68.4% vs 71.1%, p<0.05) and hCyp-3A4 (87.8% vs 90.0%, p<0.01) gene. Altered methylation of CpG positions of LINE-1, IGF2-H19, HSD11B2, estrogen receptor and P-glycoprotein gene were found. Conclusions: Placentas from breast cancer patients seem to be harmed in contrast to placentas from normal pregnancies, shown by morphologic abnormalities and a decreased proliferation index. Nevertheless, no increase of apoptotic cells could be demonstrated. Altered expression of efflux pumps or drug-metabolizing enzymes might be a reason for good fetal tolerability of chemotherapy during pregnancy as methylation patterns were changed in P-glycoprotein and CYP-3A4 gene. Citation Format: Karolin Fröhlich, Torsten Plösch, Fenja Seither, Volkmar Müller, Thomas Karn, Elmar Stickeler, Christian Schem, Christine Solbach, Amelie Lupp, Rikst N Verkaik-Schakel, Gitta Turowski, Peter Mallmann, Marion van Mackelenbergh, Bruno Sinn, Valentina Nekljudova, Carsten Denkert, Pamela Schittler, Udo Markert, Sibylle Loibl. Histological and epigenetic analyses of placenta tissue from breast cancer patients and healthy participants [abstract]. In: Proceedings of the 2019 San Antonio Breast Cancer Symposium; 2019 Dec 10-14; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2020;80(4 Suppl):Abstract nr P4-04-08.

Kaempferol modulates TCDD- and t-BHQ-induced drug-metabolizing enzymes and luteolin enhances this effect.

The expression of drug-metabolizing enzymes is deeply involved in chemical-induced cancer progression and prevention. The aryl hydrocarbon receptor (AhR) induces phase I, and certain phase II drug-metabolizing enzymes after the binding of ligands, such as 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD). We have previously demonstrated that luteolin inhibited TCDD-induced AhR transformation, and modulated the expression of drug-metabolizing enzymes through not only the AhR, but also the nuclear factor-erythroid-2-related factor 2 (Nrf2). We have examined the effect of kaempferol on the expression of drug-metabolizing enzymes through modulation of the AhR- and Nrf2-pathways, and the effect of co-treatment with kaempferol and luteolin. Kaempferol dose-dependently inhibited not only the TCDD-induced expression of phase I and phase II drug-metabolizing enzymes, but also the tertiary butylhydroquinone (t-BHQ)-induced expression of phase II drug-metabolizing enzymes, by modulating the AhR- and Nrf2-pathways. Co-treatment with kaempferol and luteolin enhanced the inhibitory effect on the expression of drug-metabolizing enzymes, compared with either kaempferol or luteolin alone. Moreover, co-treatment with kaempferol and luteolin increased the cellular levels of kaempferol without affecting the levels of luteolin. An in vivo study was also performed and the results demonstrated that co-treatment with kaempferol and luteolin enhanced the inhibition of benzo[a]pyrene-induced drug-metabolizing enzymes compared with either kaempferol or luteolin alone, in the liver of ICR mice. These results suggest that luteolin promoted the incorporation of kaempferol into hepatocytes and enhanced the inhibitory effect of kaempferol on chemical-induced drug-metabolizing enzymes. Thus, luteolin enhances the kaempferol-inhibited expression of drug-metabolizing enzymes.

P16 - Drug-induced liver injury alters expression and activities of drug-metabolizing cytochrome P450 enzymes in mouse liver at different ages

P16 - Drug-induced liver injury alters expression and activities of drug-metabolizing cytochrome P450 enzymes in mouse liver at different ages

P13 - Assessment of drug metabolizing enzyme expression after high-fat feedings, and after a switch back to a normal diet in rats

P13 - Assessment of drug metabolizing enzyme expression after high-fat feedings, and after a switch back to a normal diet in rats

Impact of obese levels on the hepatic expression of nuclear receptors and drug-metabolizing enzymes in adult and offspring mice

The prevalence of obesity-associated conditions raises new challenges in clinical medication. Although altered expression of drug-metabolizing enzymes (DMEs) has been shown in obesity, the impacts of obese levels (overweight, obesity, and severe obesity) on the expression of DMEs have not been elucidated. Especially, limited information is available on whether parental obese levels affect ontogenic expression of DMEs in children. Here, a high-fat diet (HFD) and three feeding durations were used to mimic different obese levels in C57BL/6 mice. The hepatic expression of five nuclear receptors (NRs) and nine DMEs was examined. In general, a trend of induced expression of NRs and DMEs (except for Cyp2c29 and 3a11) was observed in HFD groups compared to low-fat diet (LFD) groups. Differential effects of HFD on the hepatic expression of DMEs were found in adult mice at different obese levels. Family-based dietary style of an HFD altered the ontogenic expression of DMEs in the offspring older than 15 days. Furthermore, obese levels of parental mice affected the hepatic expression of DMEs in offspring. Overall, the results indicate that obese levels affected expression of the DMEs in adult individuals and that of their children. Drug dosage might need to be optimized based on the obese levels.

Strategies for developing pregnane X receptor antagonists: Implications from metabolism to cancer.

Pregnane X receptor (PXR) is a ligand-activated nuclear receptor (NR) that was originally identified as a master regulator of xenobiotic detoxification. It regulates the expression of drug-metabolizing enzymes and transporters to control the degradation and excretion of endobiotics and xenobiotics, including therapeutic agents. The metabolism and disposition of drugs might compromise their efficacy and possibly cause drug toxicity and/or drug resistance. Because many drugs can promiscuously bind and activate PXR, PXR antagonists might have therapeutic value in preventing and overcoming drug-induced PXR-mediated drug toxicity and drug resistance. Furthermore, PXR is now known to have broader cellular functions, including the regulation of cell proliferation, and glucose and lipid metabolism. Thus, PXR might be involved in human diseases such as cancer and metabolic diseases. The importance of PXR antagonists is discussed in the context of the role of PXR in xenobiotic sensing and other disease-related pathways. This review focuses on the development of PXR antagonists, which has been hampered by the promiscuity of PXR ligand binding. However, substantial progress has been made in recent years, suggesting that it is feasible to develop selective PXR antagonists. We discuss the current status, challenges, and strategies in developing selective PXR antagonists. The strategies are based on the molecular mechanisms of antagonism in related NRs that can be applied to the design of PXR antagonists, primarily driven by structural information.

Evaluation of mRNA expression of drug-metabolizing enzymes in acetaminophen-induced hepatotoxicity using a three-dimensional hepatocyte culture system

1. The expression and activity of drug-metabolizing enzymes are known to affect the pharmacokinetics of drugs metabolized in the liver. Here, we assessed the effect of acetaminophen (APAP)-induced hepatotoxicity on the mRNA expression of drug-metabolizing enzymes and elucidated the underlying mechanism using three-dimensional (3D) cultures of HepG2 cells.2. 3D culture cells enabled us to establish an in vitro model of APAP-induced hepatotoxicity which showed the increase in N-acetyl-p-benzoquinone imine production, reactive oxygen species (ROS) generation and cellular injury.3. In this 3D culture model, APAP treatment significantly increased the mRNA expression of drug-metabolizing enzymes (cytochrome P450 [CYP]3A4, CYP2E1 and UDP-glucuronosyltransferase 1A6) and their nuclear receptors (pregnane X receptor and constitutive androstane receptor) compared with untreated cells. Treatment with N-acetylcysteine, a therapeutic agent for APAP-induced hepatotoxicity, suppressed these increases. In addition, the mRNA expression of drug-metabolizing enzymes and nuclear receptors were elevated depending on the concentration of H2O2, one of ROS involved in the development of APAP-induced hepatotoxicity. The mRNA expression of nuclear receptors increased before that of drug-metabolizing enzymes.4. In conclusion, ROS may induce the mRNA expression of nuclear receptors and promote the transcription of drug-metabolizing enzymes in the in vitro model of APAP-induced hepatotoxicity.

Toward precision medicine in pediatric population using cytochrome P450 phenotyping approaches and physiologically based pharmacokinetic modeling.

The activity of drug-metabolizing enzymes (DME) shows high inter- and intra-individual variability. Genetic polymorphisms, exposure to drugs, and environmental toxins are known to significantly alter DME expression. In addition, the activity of these enzymes is highly age-dependent due to maturation processes that occur during development. Currently, there is a vast choice of phenotyping methods in adults using exogenous probes to characterize the activity of these enzymes. However, this can hardly be applied to children since it requires the intake of non-therapeutic xenobiotics. In addition, sampling may be challenging in the pediatric population for a variety of reasons: limited volume (e.g., blood), inappropriate sampling methods for age (e.g., urine), and metric requiring invasive or multiple blood samples. This review covers the main existing methods that can be used in the pediatric population to determine DME activity, with a particular focus on cytochrome P450 enzymes. Less invasive tools are described, including phenotyping using endogenous probes. Finally, the potential of pediatric model-informed precision dosing using physiologically based pharmacokinetic modeling is discussed.

Interaction of cigarette smoke condensate and some of its components with chlorpromazine toxicity on Saccharomyces cerevisiae

Chlorpromazine (CPZ) is an antipsychotic phenothiazine which is still commonly prescribed though it causes idiosyncratic toxicity such as cholestasis. CPZ toxicity mechanisms involve oxidative stress among others. Cigarette smoke (CS) causes deleterious effects through diverse mechanisms such as oxidative stress. CS alters drug metabolizing enzymes expression and drug transporters expression and activity in animal cell models as well as in Saccharomyces cerevisiae. CS therefore alters pharmacokinetic and pharmacodynamics of many drugs including CPZ and caffeine whose toxicity is promoted by CS condensate (CSC). CSC interaction with CPZ toxicity deserves investigation. In this study, CSC exerted mild toxicity on Saccharomyces cerevisiae which resisted to this chemical stress after several hours. CPZ toxicity on yeast was dose-dependent and the cells resisted to CPZ up to 40 µM after 24 h of treatment. Yeast cells treated simultaneously with CPZ and a nontoxic CSC dose were less sensitive to CPZ. CSC probably triggers cross-resistance to CPZ. Using Sod1 mutant strain, we showed that this gene is potentially involved in the potential cross-resistance. Other genes encoding stress-related transcription factors could be involved in this process. Nicotine and cadmium chloride, which caused a dose-dependent toxicity individually, acted with CPZ in an additive or synergistic manner in terms of toxicity. Although our results cannot be extrapolated to humans, they clearly show that CSC and its components interact with CPZ toxicity.

Effects of dexamethasone to reverse decreased hepatic midazolam metabolism in rats with acute renal failure

The inductive effects of dexamethasone on hepatic midazolam metabolism were examined in Wistar rats with acute renal failure (ARF) to clarify whether the ARF-related decrease in the hepatic expression of drug-metabolizing enzymes is caused by an impairment in the translation/polypeptide formation process.ARF was induced with intramuscular glycerol injection. Dexamethasone was orally administered. Pooled liver microsomes from five rats were prepared with ultracentrifugation for each of four groups, namely, control and ARF rats, control rats with dexamethasone treatment and ARF rats with dexamethasone treatment.Hepatic drug-metabolizing activity was examined in an incubation study with the microsomes, where midazolam was employed as a substrate of cytochrome P450 (CYP) 3A enzymes. The hepatic protein and mRNA expressions of CYP3A23/3A1 and 3A2 enzymes were also evaluated.With dexamethasone treatment, the hepatic metabolic rate of midazolam increased 1.4 times in control rats, while it increased 19.6 times in ARF rats, reflecting the greater induction of hepatic protein expressions of CYP3A enzymes in ARF rats than in control rats.The hepatic protein expression process for CYP3A23/3A1 and 3A2 responds well to dexamethasone treatment in ARF rats, indicating that the translation/polypeptide formation process is not impaired in the presence of ARF.

A non-lethal malarial infection results in reduced drug metabolizing enzyme expression and drug clearance in mice

BackgroundGiven the central importance of anti-malarial drugs in the treatment of malaria, there is a need to understand the effect of Plasmodium infection on the broad spectrum of drug metabolizing enzymes. Previous studies have shown reduced clearance of quinine, a treatment for Plasmodium infection, in individuals with malaria.MethodsThe hepatic expression of a large panel of drug metabolizing enzymes was studied in the livers of mice infected with the AS strain of Plasmodium chabaudi chabaudi, a nonlethal parasite in most strains of mice with several features that model human Plasmodium infections. C57BL/6J mice were infected with P. chabaudi by intraperitoneal injection of infected erythrocytes and sacrificed at different times after infection. Relative hepatic mRNA levels of various drug metabolizing enzymes, cytokines and acute phase proteins were measured by reverse transcriptase-real time PCR. Relative levels of cytochrome P450 proteins were measured by Western blotting with IR-dye labelled antibodies. Pharmacokinetics of 5 prototypic cytochrome P450 substrate drugs were measured by cassette dosing and high-resolution liquid chromatography-mass spectrometry. The results were analysed by MANOVA and post hoc univariate analysis of variance.ResultsThe great majority of enzyme mRNAs were down-regulated, with the greatest effects occurring at the peak of parasitaemia 8 days post infection. Protein levels of cytochrome P450 enzymes in the Cyp 2b, 2c, 2d, 2e, 3a and 4a subfamilies were also down-regulated. Several distinct groups differing in their temporal patterns of regulation were identified. The cassette dosing study revealed that at the peak of parasitaemia, the clearances of caffeine, bupropion, tolbutamide and midazolam were markedly reduced by 60–70%.ConclusionsThese findings in a model of uncomplicated human malaria suggest that changes in drug clearance in this condition may be of sufficient magnitude to cause significant alterations in exposure and response of anti-malarial drugs and co-medications.

Adenosine Deaminases Acting on RNA Downregulate the Expression of Constitutive Androstane Receptor in the Human Liver-Derived Cells by Attenuating Splicing.

Adenosine deaminases acting on RNA (ADARs) enzymes-catalyzing adenosine-to-inosine RNA editing possibly modulates gene expression and function. In this study, we investigated whether ADARs regulate the expression of human constitutive androstane receptor (CAR), which controls the expression of various drug-metabolizing enzymes. CAR mRNA and protein levels in human hepatocellular carcinoma-derived HepG2 cells were increased by knockdown of ADAR1 and slightly increased by ADAR2, indicating that ADARs negatively regulate CAR expression. Increased luciferase activity of a reporter plasmid containing the CYP3A4 promoter region by phenobarbital was augmented by transfection of siRNA for ADAR1 (siADAR1) but not by siADAR2. In addition, the knockdown of ADAR1 resulted in the enhanced induction of CYP2B6 and CYP3A4 mRNA by 6-(4-chlorophenyl)imidazo[2,1-b][1,3]thiazole-5-carbaldehyde O-(3,4-dichlorobenzyl)oxime and phenobarbital, respectively. These results suggest that ADAR1-mediated downregulation of CAR affects its downstream cytochrome P450 expression. When the transcription was inhibited by α-amanitin, the degradation of CAR mRNA was attenuated by knockdown of ADAR1, suggesting that the increase in CAR mRNA level by ADAR1 knockdown is a post-transcriptional event. Finally, we found that ADAR1 knockdown promotes the splicing of CAR as a mechanism of the increased expression of CAR by ADAR1 knockdown. In conclusion, this study revealed that ADAR1 plays a role in modulating xenobiotic metabolism potency via regulation of CAR. SIGNIFICANCE STATEMENT: This study revealed that adenosine deaminase acting on RNA 1 (ADAR1) and ADAR2, which catalyze adenosine-to-inosine RNA editing, downregulate the expression of constitutive androstane receptor (CAR) in human liver-derived cells by attenuating splicing. The downregulation of CAR by ADARs affected its downstream cytochrome P450 expression. ADARs would play a role in modulating xenobiotic metabolism potency via regulation of CAR.

Comparison of three human liver cell lines for in vitro drug-induced liver injury assessment: Huh7, HepaRG, and stem cell-derived hepatocytes

Backgrounds: Drug-induced liver injury (DILI) is a major causal factor for failure in clinical trials and withdrawal of drugs from pharmaceutical markets. Therefore, a human cell-based in vitro system has been established to overcome the limitations of preclinical trials. However, previous studies focused on the drug metabolism in each cell line or described to be superior in one single cell line for assessing DILI. In this study, we used Huh7, HepaRG, and Hepatosight-S cells to evaluate the liver toxicity and dysfunction driven by DILI-inducing drugs. Methods: Cytotoxicity of 17 DILI-inducing drugs was assessed by LDH release assay and the drug-induced liver dysfunction was confirmed by albumin secretion. Furthermore, we analyzed the expression levels of phase I and phase II enzymes and nuclear receptors in each cell lines. Results: DILI-inducing drugs were differently detected in each cell line because each cell line differs on the expression of drug metabolic enzymes which associated with reactivity on DILI inducing drugs. Conclusion: To predict the responses of DILI-inducing drugs in human liver cells, using diverse cell lines having different drug metabolic capabilities is more suitable for predicting DILI.

Activation of Pregnane X Receptor Sensitizes Mice to Hemorrhagic Shock-Induced Liver Injury.

Hemorrhagic shock (HS) is a life-threatening condition associated with tissue hypoperfusion and often leads to injury of multiple organs including the liver. Pregnane X receptor (PXR) is a species-specific xenobiotic receptor that regulates the expression of drug-metabolizing enzymes (DMEs) such as the cytochrome P450 (CYP) 3A. Many clinical drugs, including those often prescribed to trauma patients, are known to activate PXR and induce CYP3A. The goal of this study is to determine whether PXR plays a role in the regulation of DMEs in the setting of HS and whether activation of PXR is beneficial or detrimental to HS-induced hepatic injury. PXR transgenic, knockout, and humanized mice were subject to HS, and the liver injury was assessed histologically and biochemically. The expression and/or activity of PXR and CYP3A were manipulated genetically or pharmacologically in order to determine their effects on HS-induced liver injury. Our results showed that genetic or pharmacological activation of PXR sensitized wild-type and hPXR/CYP3A4 humanized mice to HS-induced hepatic injury, whereas knockout of PXR protected mice from HS-induced liver injury. Mechanistically, the sensitizing effect of PXR activation was accounted for by PXR-responsive induction of CYP3A and increased oxidative stress in the liver. The sensitizing effect of PXR was attenuated by ablation or pharmacological inhibition of CYP3A, treatment with the antioxidant N-acetylcysteine amide, or treatment with a PXR antagonist. Conclusion: We have uncovered a function of PXR in HS-induced hepatic injury. Our results suggest that the unavoidable use of PXR-activating drugs in trauma patients has the potential to exacerbate HS-induced hepatic injury, which can be mitigated by the coadministration of antioxidative agents, CYP3A inhibitors, or PXR antagonists.

Effects of Radiation on Drug Metabolism: A Review.

Radiation is the fourth most prevalent type of pollution following the water, air and noise pollution. It can adversely affect normal bodily functions. Radiation alters the protein and mRNA expression of drugmetabolizing enzymes and drug transporters and the pharmacokinetic characteristics of drugs, thereby affecting drug absorption, distribution, metabolism, and excretion. Therefore, it is important to study the pharmacokinetic changes in drugs under radiation. To update data on the effects of ionizing radiation and non-ionizing radiation caused by environmental pollution or clinical treatments on the protein and mRNA expression of drug-metabolizing enzymes and drug transporters. Data and information on pharmacokinetic changes in drugs under radiation were analyzed and summarized. The effect of radiation on cytochrome P450 is still a subject of debate. The widespread belief is that higherdose radiation increased the expression of CYP1A1 and CYP1B1 of rat, zebrafish or human, CYP1A2, CYP2B1, and CYP3A1 of rat, and CYP2E1 of mouse or rat, and decreased that of rat's CYP2C11 and CYP2D1. Radiation increased the expression of multidrug resistance protein, multidrug resistance-associated protein, and breast cancer resistance protein. The metabolism of some drugs, as well as the clearance, increased during concurrent chemoradiation therapy, whereas the half-life, mean residence time, and area under the curve decreased. Changes in the expression of cytochrome P450 and drug transporters were consistent with the changes in the pharmacokinetics of some drugs under radiation. The findings of this review indicated that radiation caused by environmental pollution or clinical treatments can alter the pharmacokinetic characteristics of drugs. Thus, the pharmacokinetics of drugs should be rechecked and the optimal dose should be re-evaluated after radiation.

Similarities in mRNA expression of peripheral blood drug metabolizing enzymes and cancer marker genes with biopsy samples of head and neck cancer patients

Purpose: To develop peripheral blood mRNA expression profiles of drug metabolizing enzymes (DMEs) as a surrogate to monitor tobacco induced head and neck squamous cell carcinoma (HNSCC), attempts were made to investigate (i) similarities in alterations with the cancer marker genes in biopsy samples and (ii) if alterations similar to that seen in biopsy samples are reflected in peripheral blood.Methods: Total RNA from eight soft gingival tissues and eight biopsy samples of HNSCC patients and total DNA and RNA from blood of healthy controls (n = 150) and HNSCC patients (n = 150) was processed for expression and genotyping studies. Blood from patients receiving chemo-radiotherapy was processed for follow-up study.Results: qRT-PCR revealed significant increase in mRNA expression of DMEs in biopsy and blood samples of HNSCC patients when compared to controls. Similar alterations were observed in cancer marker genes in these samples. Patients with variant genotypes of DMEs showed greater magnitude of alterations in mRNA expression when compared to wild type controls. Responders of chemo-radiotherapy showed significant decline in induction of mRNA expression of DMEs and cancer marker genesConclusions: The data suggest that peripheral blood expression profiles could be used to monitor tobacco-induced HNSCC as well as the treatment response.