Activity Of Drug-metabolizing Enzymes Research Articles

The Effect of Thymoquinone on the Activities of Drug Metabolizing Enzymes Following Gentamicin - Induced Nephrotoxicity in Rats

Objectives: The present study investigated the effects of thymoquinone on the renal activities of drug-metabolizing enzyme following gentamicin-induced nephrotoxicity in Wistar breed rats. Intraperitoneal administration of thymoquinone alone in rats at the dose rate of 10 mg/kg daily for ten consecutive days did not produce toxic effects in the kidney or change its drug metabolizing capacity. Results: However, nephrotoxicity was produced in Rats injected intraperitoneally with gentamicin daily at the dose rate of 80 mg/kg body weight for ten consecutive days, where there was significant increase in the concentrations of serum creatinine and blood urea compared to control rats (P < 0.0005) and those treated with thymoquinone alone (P < 0.0005), or those given thymoquinone and gentamicin concomitantly (10 mg/kg and 80 mg/kg intraperitoneally daily for ten consecutive days, respectively) (P < 0.0005). In addition, injection of gentamicin illustrated a tendency to decrease, although not statistically significant, in the kidney concentration of reduced glutathione. Furthermore, gentamicin administration resulted in a significant decrease in the renal concentration of cytochrome P-450 (P < 0.05), while it could not produce significant changes in the renal activities of phase II drug metabolizing enzymes namely, UDP- glucuronyltransferase and glutathione-S-transferase compared to values obtained for control rats and those treated with thymoquinone alone or coadministered with gentamicin. These findings confirm the nephrotoxic effects of gentamicin and its ability to decrease the renal activities of phase I drug metabolizing enzymes as shown by the significant reduction in the concentration of cytochrome P-450 in the kidneys of treated rats. Conclusion: It can be concluded that thymoquinone coadministration with gentamicin can induce protective effects against gentamicin nephrotoxicity accompanied with restoration of the concentration of cytochrome P-450 to normal levels in the kidneys of treated rats.

Pharmacokinetic and Pharmacodynamic Interactions of Ethanol and Propofol in Rabbits

Propofol is a short-acting intravenous anesthetic which is rapidly metabolized by glucuronidation and ring hydroxylation catalyzed by cytochrome P450. In clinical trials, it was found that patients receiving both ethanol and propofol injection in the surgery awoke significantly earlier than those who were injected propofol alone. To investigate pharmacokinetic and pharmacodynamic interactions of propofol and ethanol, propofol was injected alone and in combination with ethanol to rabbits. Propofol was injected intravenously at a dose rate of 15 mg kg−1. Ethanol was injected intraperitoneally at a dose rate of 2.5 mg kg−1. An LC assay with native fluorescence detection was developed and validated for determining the concentration of propofol in rabbit plasma and the effect of co-injected ethanol on the plasma pharmacokinetics of propofol in rabbits. Co-injection of ethanol decreased the maximum plasma concentration (Cmax) and area under the plasma concentration–time curve (AUC0–∞) of propofol by 53.28 and 53.42%, respectively, as compared to the control (P < 0.001). Concomitant ethanol also caused a 66.67% increase in plasma clearance (CL) (P < 0.001). These may partially explain the finding that co-injected ethanol reduced the anesthetic effect of propofol by introducing hepatic microsomal drug-metabolizing enzymes activity in rabbits.

Evaluation of drug toxicity with hepatocytes cultured in a micro-space cell culture system

A micro-space cell culture system was recently developed in which cells such as hepatocytes can be cultured and formed into a multicellular three-dimensional (3D) architecture. In this study, we assessed the performance of HepG2 cells cultured in this micro-space cell culture system in a drug toxicity test, and evaluated the effects of micro-space culture on their hepatocyte-specific functions. The micro-space cell culture facilitated the formation of 3D HepG2 cell architecture. HepG2 cells cultured in a micro-space culture plate exhibited increased albumin secretion and enhanced mRNA expression levels of cytochrome P450 (CYP) enzyme compared to those cultured in a monolayer culture. When the cells were exposed to acetaminophen, a hepatotoxic drug, the damage to the HepG2 cells grown in micro-space culture was greater than the damage to the HepG2 cells grown in monolayer culture. In addition, human primary hepatocytes grown in micro-space culture also exhibited increased albumin secretion, enhanced CYP mRNA expression levels and increased sensitivity to acetaminophen compared to those grown in monolayer culture. These results suggest that this micro-space culture method enhances the hepatocyte-specific functions of hepatocytes, including drug-metabolizing enzyme activities, making hepatocytes grown in the micro-space culture system a useful tool for evaluating drug toxicity in vitro.

Liu Wei Di Huang Wan, a well-known traditional Chinese medicine, induces CYP1A2 while suppressing CYP2A6 and N-acetyltransferase 2 activities in man

Liu wei di huang wan (LDW), a well-known traditional Chinese medicine, is frequently combined with other prescription or non-prescription drugs in China. This study was designed to investigate the effects of LDW on the activities of CYP1A2, CYP2A6, N-acetyltransferase 2 (NAT2) and xanthine oxidase (XO) in healthy subjects, using caffeine as a probe drug. Twelve unrelated healthy males were enrolled in a single-blind, randomized, placebo-controlled, two-phase crossover study. Placebo or LDW (12 pills, 0.2 g/pill, twice daily) was given to each participant for 14 continuous days with a wash-out period of 2 weeks. A dose of 100 mg caffeine was given afterwards to test the activities of drug-metabolizing enzymes of interest. Compared to placebo, LDW significantly induced the CYP1A2 activity, as determined by an increase in the ratio of (AFMU+1U+1X)/17U and the formation of 17X and 1X after taking caffeine. Interestingly, LDW significantly decreased the ratio of 17U/(17U+17X+1X+1U+AFMU) and the formation of 17U (CYP2A6-mediated) (by 39.2%; 95%CI: 23.1-55.3%; P=0.026), and decreased the ratio of AFMU/(AFMU+1U+1X) and the formation of AFMU (NAT2-catalyzed) (by 26.2%; 95%CI: 9.2-61.6%; P=0.038), suggesting a marked inhibition of CYP2A6 and NAT2, respectively. LDW can induce CYP1A2 and suppress CYP2A6 and NAT2 activities, and affect caffeine metabolism in vivo.

Fermented wheat aleurone induces enzymes involved in detoxification of carcinogens and in antioxidative defence in human colon cells

Dietary fibre is fermented by the human gut flora resulting mainly in the formation of SCFA, for example, acetate, propionate and butyrate. SCFA, in particular butyrate, may be important for secondary cancer prevention by inducing apoptosis and inhibiting cell growth of cancer cells, thereby inhibiting the promotion and/or progression of cancer. Furthermore, SCFA could also act on primary cancer prevention by activation of detoxifying and antioxidative enzymes. We investigated the effects of fermented wheat aleurone on the expression of genes involved in stress response and toxicity, activity of drug-metabolising enzymes and anti-genotoxic potential. Aleurone was digested and fermented in vitro to obtain samples that reflect the content of the colon. HT29 cells and colon epithelial stripes were incubated with the resulting fermentation supernatant fractions (fs) and effects on mRNA expression of CAT, GSTP1 and SULT2B1 and enzyme activity of glutathione S-transferase (GST) and catalase (CAT) were measured. Fermented aleurone was also used to study the protection against H2O2-induced DNA damage in HT29 cells. The fs of aleurone significantly induced the mRNA expression of CAT, GSTP1 and SULT2B1 (HT29) and GSTP1 (epithelial stripes), respectively. The enzyme activities of GST (HT29) and CAT (HT29, epithelial stripes) were also unambiguously increased (1.4- to 3.7-fold) by the fs of aleurone. DNA damage induced by H2O2 was significantly reduced by the fs of aleurone after 48 h, whereupon no difference was observed compared with the faeces control. In conclusion, fermented aleurone is able to act on primary prevention by inducing mRNA expression and the activity of enzymes involved in detoxification of carcinogens and antioxidative defence.

Effects of Hepatic Drug-metabolizing Enzyme Induction on Clinical Pathology Parameters in Animals and Man

Hepatic drug-metabolizing enzyme (DME) induction is an adaptive response associated with changes in preclinical species; this response can include increases in liver weight, hepatocellular hyperplasia and hypertrophy, and upregulated tissue expression of DMEs. Effects of DME induction on clinical pathology markers of hepatobiliary injury and function in animals as well as humans are not well established. This component of a multipart review of the comparative pathology of xenobiotically mediated induction of hepatic metabolizing enzymes reviews pertinent data from retrospective and prospective preclinical and clinical studies. Particular attention is given to studies with confirmation of DME induction and concurrent evaluation of liver and/or serum hepatobiliary marker enzyme activities and histopathology. These results collectively indicate that in the rat, when histologic findings are limited to hepatocellular hypertrophy, DME induction is not expected to be associated with consistent or substantive changes in serum or plasma activity of hepatobiliary marker enzymes such as alanine aminotransferase, alkaline phosphatase, and gamma glutamyltransferase. In the dog and the monkey, published studies also do not demonstrate a consistent relationship across DME-inducing agents and changes in these clinical pathology parameters. However, increased liver alkaline phosphatase or gamma glutamyltransferase activity in dogs treated with phenobarbital or corticosteroids suggests that direct or indirect induction of select hepatobiliary injury markers can occur both in the absence of liver injury and independently of induction of DME activity. Although correlations between tissue and serum levels of these hepatobiliary markers are limited and inconsistent, increases in serum/plasma activities that are substantial or involve changes in other markers generally reflect hepatobiliary insult rather than DME induction. Extrahepatic effects, including disruption of the hypothalamic-pituitary-thyroid axis, can also occur as a direct outcome of hepatic DME induction in humans and animals. Importantly, hepatic DME induction and associated changes in preclinical species are not necessarily predictive of the occurrence, magnitude, or enzyme induction profile in humans.

Chemopreventive effects of zinc on prostate carcinogenesis induced by N-methyl-N-nitrosourea and testosterone in adult male Sprague-Dawley rats

Zinc is an important micronutrient involved in structural and regulatory functions in mammalian cells. It inhibits proliferation of both androgen-dependent and -independent prostate cancer in vitro. However, no report is available on the chemopreventive role of zinc on prostate cancer initiation in in vivo model. The main purpose of this study was to assess the chemopreventive effects of zinc on prostate carcinogenesis induced by a single dose of N-methyl-N-nitrosourea (MNU) and continuous testosterone administration in Sprague-Dawley rats. In this study, prostate cancer was induced in Sprague-Dawley rats using MNU+ testosterone (MNU + T). Rats were simultaneously treated with zinc (100 ppm) thrice a week. Serum and tissue activity of prostatic acid phosphatase (PAcP) was measured using biochemical analysis. Serum and tissue zinc levels were assessed by atomic absorption spectrophotometry. The ventral prostatic citrate level, phase I drug-metabolizing enzymes such as cytochrome P450, cytochrome b(5), cytochrome b(5) reductase, cytochrome C reductase, phase II enzyme like glutathione-S-transferase, lipid peroxidation, hydrogen peroxide (H(2)O(2)), and reduced glutathione were also analyzed by biochemical assays. Protein expressions of p53, proliferating cell nuclear antigen (PCNA), caspase-3, and B-cell lymphoma protein-X(L) (Bcl-X(L)) were detected by Western blot analysis. Histopathological evaluation of ventral prostate was studied using hematoxylin and eosin staining method. MNU + T-treated rats showed 60, 50, and 30% of hyperplastic, dysplastic, and prostatic intraepithelial neoplastic changes, respectively, in ventral prostate, whereas MNU + T along with zinc-treated rats showed an incidence of each 10% of hyperplasia, dysplasia, and prostatic intraepithelial neoplasia in the ventral prostate. Serum zinc level and PAcP activity were significantly increased in MNU + T-treated rats, whereas these were decreased in zinc-treated rats. The ventral prostatic PAcP and glutathione-S-transferase activities, zinc, citrate, reduced glutathione levels, and protein levels of p53, caspase-3 were significantly decreased in MNU + T-treated rats, whereas increased in zinc-treated rats. Phase I drug-metabolizing enzyme activities, lipid peroxidation, H(2)O(2) levels, PCNA, and Bcl-X(L) levels were increased in MNU + T-treated rats, whereas these levels were restored to within normal limits in zinc-treated rats. This study suggests that zinc may have a beneficial effect against MNU and testosterone-induced prostate carcinogenesis. Thus, it may act as a potential chemopreventive agent in targeting the prostate cancer.

The effects of CYP2D6 and CYP3A activities on the pharmacokinetics of immediate release oxycodone

There is high interindividual variability in the activity of drug-metabolizing enzymes catalysing the oxidation of oxycodone [cytochrome P450 (CYP) 2D6 and 3A], due to genetic polymorphisms and/or drug-drug interactions. The effects of CYP2D6 and/or CYP3A activity modulation on the pharmacokinetics of oxycodone remains poorly explored. A randomized crossover double-blind placebo-controlled study was performed with 10 healthy volunteers genotyped for CYP2D6 [six extensive (EM), two deficient (PM/IM) and two ultrarapid metabolizers (UM)]. The volunteers randomly received on five different occasions: oxycodone 0.2 mg x kg(-1) and placebo; oxycodone and quinidine (CYP2D6 inhibitor); oxycodone and ketoconazole (CYP3A inhibitor); oxycodone and quinidine+ketoconazole; placebo. Blood samples for plasma concentrations of oxycodone and metabolites (oxymorphone, noroxycodone and noroxymorphone) were collected for 24 h after dosing. Phenotyping for CYP2D6 (with dextromethorphan) and CYP3A (with midazolam) were assessed at each session. CYP2D6 activity was correlated with oxymorphone and noroxymorphone AUCs and C(max) (-0.71 < Spearman correlation coefficient rhos < -0.92). Oxymorphone C(max) was 62% and 75% lower in PM than EM and UM. Noroxymorphone C(max) reduction was even more pronounced (90%). In UM, oxymorphone and noroxymorphone concentrations increased whereas noroxycodone exposure was halved. Blocking CYP2D6 (with quinidine) reduced oxymorphone and noroxymorphone C(max) by 40% and 80%, and increased noroxycodone AUC(infinity) by 70%. Blocking CYP3A4 (with ketoconazole) tripled oxymorphone AUC(infinity) and reduced noroxycodone and noroxymorphone AUCs by 80%. Shunting to CYP2D6 pathway was observed after CYP3A4 inhibition. Drug-drug interactions via CYP2D6 and CYP3A affected oxycodone pharmacokinetics and its magnitude depended on CYP2D6 genotype.

Plant Polyphenol Curcumin Significantly Affects CYPIA2 and CYP2A6 Activity in Healthy, Male Chinese Volunteers

Curcumin is a kind of plant polyphenol that is extracted from the rhizome of Curcuma longa. Studies about the effect of curcumin on the activity of drug-metabolizing enzymes in humans are lacking. To investigate the effect of curcumin on the activities of CYP1A2, CYP2A6, N-acetyltransferase (NAT2), and xanthine oxidase (XO) in vivo, using caffeine as a probe drug. Sixteen unrelated, healthy Chinese men were recruited for the study. There were 2 phases in the study. In the first phase, volunteers orally received 100 mg caffeine and 0- to 12-hour blood and urine samples were collected. In the second phase, volunteers received 1000 mg curcumin once daily for 14 continuous days, and blood and urine samples were collected on day 15, following the same procedure used on the first day. Urinary caffeine metabolite ratios were used as the indicators of the activities of CYP1A2, CYP2A6, NAT2, and XO. The pharmacokinetics of caffeine and its metabolites were determined by high-performance liquid chromatography. In the curcumin-treated group, CYP1A2 activity was decreased by 28.6% (95% CI 15.6 to 41.8; p < 0.000), while increases were observed in CYP2A6 (by 48.9%; 95% CI 25.3 to 72.4; p < 0.000). Plasma area under the curve from zero to 12 hours of 1,7-dimethylxanthine (17X) was decreased by 27.2% (95% CI 6.1 to 48.3; p = 0.014). The urinary excretion of 17X and 1-methylxanthine was significantly decreased by 36.4% (95% CI 19.4 to 53.6; p < 0.000) and 31.2% (95% CI 8.5 to 54.1; p = 0.010), respectively. The excretion of 1,7-dimethylurate (17U) was significantly increased by 77.3% (95% CI 5.6 to 148.8; p = 0.036). No significant changes were observed for caffeine, 1-methylurate, and 5-acetylamino-6-formylamino-3-methyluracil between the 2 study phases. The results indicated that curcumin inhibits CYP1A2 function but enhances CYP2A6 activity. Simultaneously, some pharmacokinetic parameters relating to 17X were affected by curcumin. If this finding is confirmed by other studies, the possibility of herb-drug interactions associated with curcumin should be considered in clinical practice.

Abstract 3604: Significant effect of polymorphisms in CYP2D6 and DT-1 on clinical outcomes of adjuvant tamoxifen therapy for breast cancer patients

Abstract PURPOSE: The clinical efficacy of tamoxifen is suspected to be influenced by the activity of drug-metabolizing enzymes and transporters involved in the formation, metabolism and elimination of its active forms. We investigated relations of polymorphisms in transporter genes as well as CYP2D6 to clinical outcome of patients with tamoxifen treatment. PATIENTS AND METHODS: We enrolled 282 hormone receptor-positive, invasive breast cancer patients receiving tamoxifen monotherapy. We investigated the effects of allelic variants of CYP2D6 and tag-SNPs of DT-1, DT-2 and DT-3 on recurrence-free survivals using Kaplan-Meier and Cox regression analyses. Plasma concentrations of tamoxifen metabolites were measured in 98 patients taking 20 mg/day of tamoxifen. RESULTS: CYP2D6 variants were significantly associated with shorter recurrence-free survival (P = 3.6×10−5; hazard ratio (HR) 9.52 [95% CI 2.79-32.45] in patients with 2 variant alleles vs without variant allele). Among 51 tag-SNPs in transporter genes, a significant association was found at SNPY in DT-1(P = 1.7×10−4; HR 10.64 [95% CI 1.44-78.88] in patients with AA vs GG genotypes). The number of risk alleles of CYP2D6 and DT-1 showed cumulative effects on recurrence-free survivals (P = 5.5×10−8). The patients carrying 4 risk alleles had 45.25-times higher risk compared to those with ≤1 risk allele. CYP2D6 variants were associated with lower plasma levels of endoxifen and 4-hydroxytamoxifen (P = 4.3×10−6 and 5.2×10−4), whereas no significant difference was found among DT-1 genotype groups. CONCLUSION: Our results indicated that polymorphisms in CYP2D6 and DT-1 are important predictors for the prognosis of breast cancer patients treated with tamoxifen. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 3604.

Significant Effect of Polymorphisms in CYP2D6 and ABCC2 on Clinical Outcomes of Adjuvant Tamoxifen Therapy for Breast Cancer Patients

The clinical efficacy of tamoxifen is suspected to be influenced by the activity of drug-metabolizing enzymes and transporters involved in the formation, metabolism, and elimination of its active forms. We investigated relationships of polymorphisms in transporter genes and CYP2D6 to clinical outcome of patients receiving tamoxifen. We studied 282 patients with hormone receptor-positive, invasive breast cancer receiving tamoxifen monotherapy, including 67 patients who have been previously reported. We investigated the effects of allelic variants of CYP2D6 and haplotype-tagging single nucleotide polymorphisms (tag-SNPs) of ABCB1, ABCC2, and ABCG2 on recurrence-free survival using the Kaplan-Meier method and Cox regression analysis. Plasma concentrations of tamoxifen metabolites were measured in 98 patients receiving tamoxifen 20 mg/d. CYP2D6 variants were significantly associated with shorter recurrence-free survival (P = .000036; hazard ratio [HR] = 9.52; 95% CI, 2.79 to 32.45 in patients with two variant alleles v patients without variant alleles). Among 51 tag-SNPs in transporter genes, a significant association was found at rs3740065 in ABCC2 (P = .00017; HR = 10.64; 95% CI, 1.44 to 78.88 in patients with AA v GG genotypes). The number of risk alleles of CYP2D6 and ABCC2 showed cumulative effects on recurrence-free survival (P = .000000055). Patients carrying four risk alleles had 45.25-fold higher risk compared with patients with <or= one risk allele. CYP2D6 variants were associated with lower plasma levels of endoxifen and 4-hydroxytamoxifen (P = .0000043 and .00052), whereas no significant difference was found among ABCC2 genotype groups. Our results suggest that polymorphisms in CYP2D6 and ABCC2 are important predictors for the prognosis of patients with breast cancer treated with tamoxifen.

Protection by Osbeckia aspera against carbon tetrachloride-mediated alterations in microsomal drug metabolizing enzyme activity.

Previous investigations have confirmed the protective effect of Osbeckia aspera leaf extract on carbon tetrachloride-mediated liver injury in rat models. It is well known that the earliest alterations in liver cell structure and function following carbon tetrachloride poisoning involve the endoplasmic reticulum and its drug metabolizing enzymes. Therefore, we investigated whether an aqueous leaf extract of O. aspera could offer protection against carbon tetrachloride-induced changes in the microsomal drug metabolizing enzymes aniline hydroxylase and p-aminopyrine N-demethylase. This enzyme activity was compared with phenobarbital-induced righting reflex and lipid peroxidation. Treatment of rats with the aqueous leaf extract of O. aspera (before or after the administration of carbon tetrachloride) resulted in a marked decrease in carbon tetrachloride-mediated alterations in aniline hydroxylase and p-aminopyrine N-demethylase activity, phenobarbital-induced loss of righting reflex and malondialdehyde formation due to lipid peroxidation. The Km value of these enzymes in control and Osbeckia-treated rats were the same. These results show that the plant extract can markedly decrease the carbon tetrachloride-mediated reduction in aniline hydroxylase and p-aminopyrine N-demethylase activity and inhibit peroxidative damage to the cell membrane. Phenobarbital-induced sleeping time in rats and kinetic enzyme studies suggested that the effects of the plant extract was neither due to an induction of the drug-metabolizing enzymes under investigation, nor due to an alteration in the Km values of these enzymes.

Do 5-fluorouracil therapies alter CYP2C19 metaboliser status?

To report a case of altered CYP2C19 metaboliser status following 5-fluorouracil treatment. A 78-year-old male with stage III colorectal adenocarcinoma was prescribed with weekly iv 5-fluorouracil and folinic acid (FU/FA). Fourteen weeks after starting FU/FA, the patient was enrolled in a clinical study to investigate the role of tumour burden on drug metabolising enzyme activity. CYP2C19 genotype was determined and the activity of CYP2C19 was measured using proguanil (PG) as a probe substrate. A metabolic ratio (PG/CG) for CYP2C19 activity was determined on three separate occasions, 7 days apart. The patient was homozygous wild type (CYP2C19*1/*1), and on the first test, the metabolic ratio was concordant with the extensive metaboliser genotype. However, at day 14 and day 21, the metabolic capacity of this enzyme had decreased, and the subject had become a poor metaboliser (PG/CG > 30). The patient developed grade III hand and foot syndrome at day 10 of the study during the period of null CYP2C19 activity. Although 5FU is not a substrate for hepatic drug metabolising CYP enzymes, it may interfere with the synthesis of CYP2C19. Decreased activity of a related enzyme, CYP2C9, following 5FU has been reported previously. Down regulation of CYP2C9 and CYP2C19 synthesis by 5FU therapies may explain the adverse effect of 5FU on the clinical disposition of warfarin and phenytoin.

Variability in drug metabolizing enzyme activity in HIV-infected patients

To evaluate variability in cytochrome P450 (CYP) 1A2, CYP2D6, CYP3A, N-acetyltransferase 2 (NAT2), and xanthine oxidase (XO) activity in HIV-infected patients and compare this with data from uninfected, healthy volunteers. Ten HIV-infected men and seven women on medication affecting CYP enzyme activity were phenotyped four times over 2 months using caffeine, dextromethorphan, and midazolam. Urinary caffeine and dextromethorphan metabolite ratios were used to phenotype CYP1A2, NAT2, XO, and CYP2D6 activity and midazolam plasma clearance was used to phenotype CYP3A activity. Plasma and urine samples were analyzed by validated LC/UV or LC/MS methods for midazolam, caffeine, and dextromethorphan. Noncompartmental pharmacokinetics and nonparametric statistical analyses were performed, and the data compared with those of healthy volunteer historic controls. Compared with age and sex-matched healthy volunteers, HIV-infected subjects had 18% lower hepatic CYP3A4 activity, 90% lower CYP2D6 activity, 53% lower NAT2 activity, and 22% higher XO activity. No significant difference was found in CYP1A2 activity. Additionally, 25% genotype-phenotype discordance in CYP2D6 activity was noted in HIV-infected subjects. Intraindividual variability in enzyme activity increased by 42-62% in HIV-infected patients for CYP1A2, NAT2, and XO, and decreased by 33% for CYP2D6. Interindividual variability in enzyme activity increased by 27-63% in HIV-infected subjects for CYP2D6, CYP1A2, and XO, and decreased by 38% for NAT2. Higher plasma TNFalpha concentrations correlated with lower CYP2D6 and CYP3A4 activity. Infection with HIV or stage of HIV infection may alter Phase I and II drug metabolizing enzyme activity. HIV infection was related to an increase in variability of these drug-metabolizing enzymes. Altered metabolism may be a consequence of immune activation and cytokine exposure.

Variability in drug metabolizing enzyme activity in HIV-infected patients

Variability in drug metabolizing enzyme activity in HIV-infected patients

Epigenetic changes improve differentiation of hepatozyte-like cells from adipose tissue: possible application for cell therapies in surgery

Question Due to donor organ scarcity, researchers nowadays focus on cell transplantation as alternative method to orthotopic liver transplantation. For this purpose, several groups attempt to generate hepatocyte-like cells from various adult stem or precursor cells such as mesenchymal stem cells (MSCs), fat tissue (Ad-) or liver. Aim of this study was to improve hepatic-function and amount of hepatocyte-like cells via epigenetic changes. Methods Human Ad-MSCs were isolated from different patients, with their informed consent according to ethical guidelines of the MRI. For hepatic differentiation several supplement combinations of 5-Azacytidine, FGF-4, Dexamethasone, Nicotinamid, ITS, HGF and EGF were used. The generated hepatocyte-like cells were stained for Glycogen, Glucose-6-phosphatase and neutral lipids. We further investigated glucose and urea metabolism as well as several phase I and II drug metabolizing enzyme activities. Expression of pluripotency-, mesoderm- and endoderm-markers was analyzed by RT-PCR. Results The isolated Ad-MSCs were tested on their pluripotency via Oct3/4, KLF4, Sox2 and c-Myc. After 14 days of differentiation, Ad-MSCs show similar morphological features than primary human hepatocytes and gain the ability to accumulate glycogen and express glucose-6-phophatase. To investigate the metabolic ability Urea- and Glucose metabolism was analyzed. Phase I and II enzyme activities reached levels up to 70% of primary human hepatocytes. Pre-treatment with 5-Azacytidine further increased both metabolic and enzymatic activities of the cells significantly. Conclusion Our work shows, inhibition of DNA-methyltransferase leads to a better hepatic differentiation of Ad-MSCs. Furthermore, a large number of AD-MSC´s can be generated. Hence, these cells may be used for alternative autologous therapies in surgery to bridge liver dys-functions.

Toxicodynamics of subacute co-exposure to groundwater contaminant arsenic and analgesic–antipyretic drug acetaminophen in rats

Arsenic is an environmental contaminant, while acetaminophen is an extensively used nonsteroidal analgesic–antipyretic drug. We evaluated whether subacute co-exposure to arsenic and acetaminophen would produce more toxicity than that caused by exposure to either of the xenobiotics in rats. Toxicity was evaluated through changes in body weight, feed consumption, liver weight and microsomal drug-metabolizing enzymes, lipid peroxidation and antioxidants in liver. Arsenic had no effect on body weight and feed consumption. Acetaminophen-mediated decrease in body weight was attenuated in the co-exposed rats. Acetaminophen alone or its co-administration with arsenic decreased feed consumption. Arsenic reduced acetaminophen-mediated increase in the activities of drug-metabolizing enzymes. The co-exposure caused lesser lipid peroxidation than the individual exposure. Arsenic or acetaminophen given alone depleted GSH and decreased the activities of superoxide dismutase, catalase, glutathione peroxidase, glutathione reductase and glutathione- S-transferase and these effects remained mostly unaffected after co-exposure. The results suggest that co-exposure to arsenic and acetaminophen may be less hazardous than their independent exposure in rats.

Drug metabolizing enzyme activity of in vitro human dermal (EpiDerm™) and airway (EpiAirway™) epithelial models: Relationship to genotoxicity of chemicals as determined by in vitro skin micronucleus assays

Drug metabolizing enzyme activity of in vitro human dermal (EpiDerm™) and airway (EpiAirway™) epithelial models: Relationship to genotoxicity of chemicals as determined by in vitro skin micronucleus assays

Global Gene Expression Analysis in the Livers of Rat Offspring Perinatally Exposed to Low Doses of 2,2′,4,4′-Tetrabromodiphenyl Ether

BackgroundPolybrominated diphenyl ethers are a group of flame-retardant chemicals appearing increasingly in the environment. Their health effects and mechanisms of toxicity are poorly understood.ObjectivesWe screened for the sensitive effects and mechanisms of toxicity of 2,2′,4,4′-tetrabromodiphenyl ether (BDE-47) by analyzing the gene expression profile in rats exposed to doses comparable to human exposure.MethodsWistar dams were exposed to vehicle or BDE-47 (0.002 and 0.2 mg/kg body weight) every fifth day from gestation day 15 to postnatal day 20 by injections to caudal vein. Total RNA was extracted from the livers of pups and hybridized to the whole-genome RNA expression microarrays. The list of genes 2-fold differentially expressed was exported to PANTHER and Ingenuity Systems for analysis of enriched ontology groups and molecular pathways.ResultsOxidoreductase and transferase protein families were enriched in exposed rats as were these biological process categories: carbohydrate metabolism; electron transport; and lipid, fatty acid, and steroid metabolism. Four signaling pathways (cascades of activation of drug-metabolizing enzymes) and 10 metabolic pathways were significantly enriched. Drug-metabolizing enzymes appear to be regulated by BDE-47 through an aryl hydrocarbon receptor–independent mechanism. Direct interaction with retinoid X receptor or its upstream cascade may be involved. The main metabolic effects consisted of activation of metabolic pathways: α- and ω-oxidation of fatty acids, glycolysis, and starch hydrolysis.ConclusionsAltered expression of genes involved in metabolic and signaling pathways and functions of the organism occurs after perinatal exposure of rat offspring to BDE-47 at doses relevant for the general population.

SIMULTANEOUS ACTION OF THE FLAVONOID QUERCETIN ON CYTOCHROME P450 (CYP) 1A2, CYP2A6, N‐ACETYLTRANSFERASE AND XANTHINE OXIDASE ACTIVITY IN HEALTHY VOLUNTEERS

1. Quercetin, one of the most abundant natural flavonoids, has been reported to modulate the activity of several drug-metabolising enzymes. The aim of the present study was to investigate the effects of quercetin on cytochrome P450 (CYP) 1A2, CYP2A6, N-acetyltransferase (NAT2) and xanthine oxidase (XO) activity in healthy volunteers using caffeine as a probe drug. 2. Twelve unrelated, healthy volunteers were recruited to the study. There were two phases to the study; in the first phase, each subject was given a single oral dose of caffeine (one 100 mg capsule) with 150 mL water; in the second phase, each subject was give a 500 mg quercetin capsule once daily for 13 continuous days and was coadministered a 100 mg caffeine capsule on the 13th day. Urinary caffeine metabolite ratios were used as indicators of the activity of CYP1A2, CYP2A6, NAT2 and XO. The pharmacokinetics of caffeine and its metabolites were determined by HPLC. 3. In the quercetin-treated group, CYP1A2 activity was decreased by 10.4% (95% confidence interval (CI), 1.1-29.8%; P = 0.039), whereas increases were observed in CYP2A6 (by 25.3%; 95% CI, 6.2-34.5%; P = 0.002), NAT2 (by 88.7%; 95% CI, 7.1-160.2%; P = 0.010) and XO activity (by 15.0%; 95% CI, 1.6-21.6%; P = 0.007). Plasma C(max) and the AUC((0-24 h)) of 1,7-dimethylxanthine were decreased by 17.2% (95% CI, 6.4-28.0%; P = 0.024) and 16.2% (95% CI, 3.9-28.5%; P = 0.032), respectively. The urinary excretion of 1,7-dimethylxanthine and 1-methylxanthine was significantly decreased by 32.4% (95% CI, 2.5-62.1%; P = 0.036) and 156.1% (95% CI, 53.3-258.9%; P = 0.004), respectively. The urinary excretion of 1,7-dimethylurate and 1-methylurate was increased by 82.9% (95% CI, 56.0-165.4%; P = 0.030) and 97.8% (95% CI, 12.1-183.5%; P = 0.029), respectively. No changes were observed in the urinary excretion of caffeine and 5-acetylamino-6-formylamino-3-methyluracil between the two study phases. 4. The results of the present study indicate that quercetin inhibits CYP1A2 function, but enhances CYP2A6, NAT2 and XO activity. Simultaneously, some pharmacokinetic parameters relating to 1,7-dimethylxanthine were affected by quercetin. Thus, we conclude that quercetin affects CYP1A2, CYP2A6, NAT2 and XO activity in vivo.