The therapeutic role of lactobacillus and montelukast in combination with metformin in diabetes mellitus complications through modulation of gut microbiota and suppression of oxidative stress.

Male reproductive dysfunction is one of the overlooked findings of diabetes mellitus (DM) that deserves greater scientific attention. This study is designed to explore the therapeutic potential of metformin and montelukast, in combination with Lactobacillus, for modulation of intestinal flora and suppression of oxidative stress in testicular and liver damage in diabetic male rats. A DM model was induced by streptozotocin (STZ)which caused functional, biochemical, and inflammatory injuries to the testicular and liver tissues. The experimental panel included nine rat groups: normal control, normal control plus metformin, normal control plus montelukast, DM control, DM plus montelukast, DM plus a combination of metformin and Lactobacillus, DM plus a combination of montelukast and Lactobacillus, and DM plus a combination of metformin and montelukast. In parallel, clinical evaluation of microscopic examination scoring, and hepatic and testicular injuries, were evaluated. Biochemical markers including glucose level, lipid profile, inflammatory markers (tumor necrosis factor- (TNF-α) and interleukin-17 (IL-17), Caspase-3, and Bax proteins expressions were measured. The change in the microbiota abundance was investigated using conventional and real-time PCR. The current study revealed a significant difference in the relative abundance of microbiota, where DM is associated with an enormous increase of Bacteroides spp., Clostridium spp., E. coli, and Fusobacterium spp., and a significant decrease in Bifidobacteria spp., and Lactobacillus spp., in contrast with normal control. Metformin and montelukast, in combination with Lactobacillus, significantly reversed the testicular and liver damage caused by STZ. Moreover, the drugs significantly reduced the oxidative, inflammatory, and apoptotic activities induced by STZ.

Antiviral activity against Middle East Respiratory Syndrome coronavirus by Montelukast, an anti-asthma drug.

Middle East Respiratory Syndrome (MERS) is a respiratory disease caused by a coronavirus (MERS-CoV). Since its emergence in 2012, nosocomial amplifications have led to its high epidemic potential and mortality rate of 34.5%. To date, there is an unmet need for vaccines and specific therapeutics for this disease. Available treatments are either supportive medications in use for other diseases or those lacking specificity requiring higher doses. The viral infection mode is initiated by the attachment of the viral spike glycoprotein to the human Dipeptidyl Peptidase IV (DPP4). Our attempts to screen antivirals against MERS led us to identify montelukast sodium hydrate (MSH), an FDA-approved anti-asthma drug, as an agent attenuating MERS-CoV infection. We showed that MSH directly binds to MERS-CoV-Receptor-Binding Domain (RBD) and inhibits its molecular interaction with DPP4 in a dose-dependent manner. Our cell-based inhibition assays using MERS pseudovirions demonstrated that viral infection was significantly inhibited by MSH and was further validated using infectious MERS-CoV culture. Thus, we propose MSH as a potential candidate for therapeutic developments against MERS-CoV infections.

Evaluation of Montelukast for the Prevention of Infusion-related Reactions With Daratumumab.

BACKGROUND: Daratumumab is an anti-CD38 monoclonal antibody indicated for the treatment of multiple myeloma. Infusion-related reactions (IRRs) are among the most common adverse events associated with daratumumab. IRRs are most common with the first infusion of daratumumab. Recommended premedications to be given prior to the daratumumab dose include acetaminophen, diphenhydramine, and a corticosteroid. There is emerging data to suggest that the addition of montelukast to this premedication regimen can lower the incidence of daratumumab-related IRRs. PATIENTS AND METHODS: This was a single-center, retrospective chart review conducted at a large, multistate health system with several different hematology/oncology practice sites. Eligible patients included those with a primary diagnosis of a plasma cell disorder who received at least 1 dose of daratumumab. The primary outcome was the incidence of IRRs with the first daratumumab infusion. RESULTS: A total of 141 patients receiving daratumumab-based therapy were included in this study. All patients received acetaminophen, diphenhydramine, and a corticosteroid as premedications prior to the first infusion of daratumumab. Overall, 46 (33%) patients experienced an IRR with the first infusion of daratumumab. The incidence of IRR was lower in patients that received montelukast as a premedication compared with those that did not (montelukast, n = 25 [27%]; no montelukast, n = 21 [45%]; P = .0371). Patients in each arm experienced similar rates of overall, composite pulmonary, gastrointestinal, and systemic IRR manifestations. CONCLUSION: The use of montelukast prior to the first daratumumab infusion led to a reduction in the incidence of IRRs in our experience.

Effect of High-Dose Esomeprazole on CYP1A2, CYP2C19, and CYP3A4 Activities in Humans: Evidence for Substantial and Long-lasting Inhibition of CYP2C19.

In vitro, esomeprazole is a time-dependent inhibitor of CYP2C19. Additionally, racemic omeprazole induces CYP1A2 and omeprazole and its metabolites inhibit CYP3A4 in vitro. In this 5-phase study, 10 healthy volunteers ingested 20 mg pantoprazole, 0.5 mg midazolam, and 50 mg caffeine as respective index substrates for CYP2C19, 3A4, and 1A2 before and 1, 25, 49 (pantoprazole only), and 73 hours after an 8-day pretreatment with 80 mg esomeprazole twice daily. The area under the plasma concentration-time curve (AUC) of R-pantoprazole increased 4.92-fold (90% confidence interval (CI) 3.55-6.82), 2.31-fold (90% CI 1.85-2.88), and 1.33-fold (90% CI 1.06-1.68) at the 1-hour, 25-hour, and 73-hour phases, respectively, consistent with a substantial and persistent inhibition of CYP2C19. The AUC of midazolam increased up to 1.44-fold (90% CI 1.22-1.72) and the paraxanthine/caffeine metabolic ratio up to 1.19-fold (90% CI 1.04-1.36), when the index substrates were taken 1 hour after esomeprazole. Based on the recovery of R-pantoprazole oral clearance, the turnover half-life of CYP2C19 was estimated to average 53 hours. Pharmacokinetic simulation based on the observed concentrations of esomeprazole and its metabolites as well as their published CYP2C19 inhibitory constants was well in line with the observed changes in R-pantoprazole pharmacokinetics during the course of the study. Extrapolations assuming linear pharmacokinetics of esomeprazole suggested weak to moderate inhibition at 20 and 40 mg twice daily dosing. In conclusion, high-dose esomeprazole can cause strong inhibition of CYP2C19, but only weakly inhibits CYP3A4 and leads to minor induction of CYP1A2. The enzymatic activity of CYP2C19 recovers gradually in ~ 3-4 days after discontinuation of esomeprazole treatment.

Comparison of the toxicity of sulfur mustard and its oxidation products in vitro.

The molecular toxicology of the chemical warfare agent sulfur mustard (SM) is still not completely understood. It has been suggested that in addition to SM itself also biotransformation products thereof mediate cytotoxicity. In the current study, we assessed this aspect by exposing a human hepatocyte cell line (HepG2) to SM or to its oxidation products sulfur mustard sulfoxide (SMO), sulfur mustard sulfone (SMO2), and divinyl sulfone (DVS). Cytotoxicity, determined with the XTT assay, revealed a significant higher toxicity of SMO2 and DVS compared to SM while SMO had no effect at any concentration. The exact biotransformation of SM leading to SMO, SMO2 and finally DVS is unknown so far. Involvement of the CYP450 system is discussed and was also investigated in the presented study. Modulation of CYP1A2 activity, taken as a model enzyme for CYP450, affected cytotoxicity of SM, SMO2 or DVS significantly. Induction of CYP1A2 with omeprazole led to decreased cytotoxicity for all compounds whereas inhibition with cimetidine resulted in an increased cytotoxicity for SM, but not for SMO2 and DVS. Our results indicate a distinctive role of the CYP450 system in SM poisoning. Future studies should address the metabolic conversion of SM in more detail. Our data may suggest the well-tolerated drug omeprazole as a potential co-treatment after contact to SM.

A New Approach of Mitigating CYP3A4 Induction Led to the Discovery of Potent Hepatitis B Virus (HBV) Capsid Inhibitor with Optimal ADMET Profiles.

Described herein is a new approach to mitigate CYP3A4 induction. In this unconventional approach, a fine-tuning of the dihedral angle between the C4 phenyl and the dihydropyrimidine core of the heteroaryldihydropyrimidine (HAP) class of capsid inhibitors successfully altered the structure-activity-relationships (SARs) of the unwanted CYP3A4 induction and the desired HBV capsid inhibition to more favorable values. This eventually led to the discovery of a new capsid inhibitor with significantly reduced CYP3A4 induction, excellent anti-HBV activity, favorable preclinical PK/PD profiles, and no early safety flags.

Is there any effect of montelukast on prevention of myringosclerosis after myringotomy in a rat model?

OBJECTIVES: In this study, our aim was to identify the possible effects of montelukast sodium (ML) on the prevention of experimentally induced myringosclerosis. MATERIALS AND METHODS: Twenty-eight female Wistar albino rats were used and they were divided into four groups randomly. Tympanic membranes (TM) of all animals were perforated and then group 1 received no treatment (control group), group 2 was treated with a topical saline solution, group 3 received topically ML and group 4 received orally ML. On the 15th day, all animals were euthanized. Tympanic membranes were evaluated otomicroscopically and histopathologically. RESULTS: The histopathological findings, compared against a control and saline groups, showed the topically and orally ML groups had statistically significant differences of degree of myringosclerosis (p < 0.002) and median thickness of the TMs (p < 0.001). Suppression of inflammation was statistically significant only in the oral ML treatment group (p < 0.002). CONCLUSION: Oral and topically administration of ML reduced myringosclerosis formation in myringotomies rats.

Discovery and characterization of N-(1,3-dialkyl-1H-indazol-6-yl)-1H-pyrazolo[4,3-b]pyridin-3-amine scaffold as mGlu4 positive allosteric modulators that mitigate CYP1A2 induction liability.

Previous reports from our laboratory disclosed the structure and activity of a novel 1H-pyrazolo[4,3-b]pyridine-3-amine scaffold (VU8506) which showed excellent potency, selectivity and in vivo efficacy in preclinical rodent models of Parkinson's disease. Unfortunately, this compound suffered from significant CYP1A2 induction as measured through upstream AhR activation (125-fold) and thus was precluded from further advancement in chronic studies. Herein, we report a new scaffold developed recently which was systematically studied in order to mitigate the CYP1A2 liabilities presented in the earlier scaffolds. We have identified a novel structure that maintains the potency and selectivity of other mGlu4 PAMs, leading to 9i (hmGlu4 EC50 = 43 nM; AhR activation = 2.3-fold).

In Vivo Assessment of the Effect of CYP1A2 Inhibition and Induction on Pomalidomide Pharmacokinetics in Healthy Subjects.

Pomalidomide is an immunomodulatory drug, and the dosage of 4 mg per day taken orally on days 1-21 of repeated 28-day cycles has been approved in the European Union and the United States to treat patients with relapsed/refractory multiple myeloma. In vitro data showed that pomalidomide is a substrate of multiple cytochrome P450 (CYP) isozymes and that its oxidative metabolism is mediated primarily by CYP1A2 and CYP3A4, with minor contributions from CYP2C19 and CYP2D6. The effect of CYP1A2 inhibition by fluvoxamine (a strong CYP1A2 inhibitor) and CYP1A2 induction by smoking on pomalidomide pharmacokinetics in healthy subjects has been assessed in 2 separate phase 1 open-label, single-dose studies. Following administration of a single oral dose of 4 mg pomalidomide, the plasma exposure when coadministered with fluvoxamine was 225.1% and 123.7% of that when administered alone for the total plasma exposure (AUC0-inf ) and the plasma peak exposure (Cmax ), respectively. In smokers with elevated CYP1A2 activity demonstrated by high caffeine clearance (a marker of CYP1A2 induction), the AUC0-inf was 32.3% lower, whereas the Cmax was 14.4% higher than that in nonsmokers. In addition, pomalidomide was safe and well tolerated as a single oral dose of 4 mg in healthy male smokers and nonsmokers ≥ 40 to ≤ 80 years old, and a single oral dose of 4 mg pomalidomide coadministered with multiple oral 50-mg doses of the CYP1A2 inhibitor fluvoxamine compared with pomalidomide alone was safe and well tolerated by the healthy male subjects.

Hydroxystilbenes and methoxystilbenes activate human aryl hydrocarbon receptor and induce CYP1A genes in human hepatoma cells and human hepatocytes.

Natural polyphenol resveratrol (trihydroxystilbene) is a partial agonist of human aryl hydrocarbon receptor AhR, thereby, displaying a plethora of biological effects. Biological activities of metoxylated and hydroxylated stilbenes were studied in the past. The aim of the current study was to describe the effects of 13 different hydroxy- and methoxystilbenes, including their cis/trans isomers on the transcriptional activity of AhR and the expression of CYP1A genes in hepatic cancer cells HepG2 and in primary human hepatocytes. Techniques of gene reporter assays, qRT-PCR, Simple Western blotting by Sally Sue™ and electrophoretic mobility shift assay EMSA were employed. All compounds activated AhR, but their efficacies, potencies and dose-response profiles differed substantially. The strongest activators of AhR and inducers of CYP1A1 in HepG2 cells were DMU-212 ((E)-3,4,5,4´-tetramethoxystilbene), trans-piceatannol, cis-piceatannol, trans-trismethoxyresveratrol and trans-pinostilbene. While DMU-212 and trans-trismethoxyresveratrol also induced CYP1A1 and CYP1A2 in primary human hepatocytes, the effects of trans-piceatannol, cis-piceatannol and trans-pinostilbene weaned off. On the other hand, trans-4-methoxystilbene was strong CYP1A inducer in hepatocytes but not in HepG2 cells. Differences between effects of stilbenes in HepG2 cells and human hepatocytes are probably due to the extensive phase I and phase II xenobiotic metabolism in human hepatocytes. The data obtained may be of toxicological relevance.

Muscone Induces CYP1A2 and CYP3A4 Enzyme Expression in L02 Human Liver Cells and CYP1A2 and CYP3A11 Enzyme Expression in Kunming Mice.

AIM: To examine the effect of synthetic muscone on the expression of CYP1A2 and CYP3A4 enzymes in human liver L02 cells and in the liver tissue of Kunming mice. METHODS: The L02 hepatic cell line was used to study the effect of low (10-4 µmol/L), middle (10-3 µmol/L), and high concentrations (10-2 µmol/L) of muscone on the expression of CYP1A2 and CYP3A4 enzymes. In addition, the cytochrome P450 (CYP) expression was investigated in Kunming mice after the administration of 10 mg/kg (low), 50 mg/kg (middle), and 100 mg/kg (high) dose of muscone for 6 days. A mixture of phenobarbital (30 mg/kg) and ß-napthoflavone (80 mg/kg) was used as positive control and the effects of the compounds on CYP expression were investigated at the end of 6- and 12-day periods. RESULTS: Muscone induced the expression of CYP1A2 (middle and low concentrations) and of CYP3A4 (high concentration) enzymes in L02 cells. In vivo, administration of muscone in Kunming mice revealed significant weight reduction at the end of 6- and 12-day periods (middle and high doses, respectively), compared to the control group (p < 0.05). Liver toxicity scores indicated that the liver injuries in the positive control and high doses of muscone group were significantly higher in the 6- and 12-day periods, compared to those in the blank control group (p < 0.05). Furthermore, muscone induced CYP1A2 and CYP3A11 expressions in Kunming mice at the middle dose and all doses during the 12-day period as demonstrated by immunoblotting experiments. A low dose of mucone induced the CYP enzyme expression more rapidly, whereas a high dose of muscone caused the longest inductive effect. The results were confirmed by immunohistochemistry experiments and real-time PCR studies, where similar patterns of muscone-mediated inductive effects were noted. CONCLUSIONS: Muscone induces CYP1A2 and CYP3A4 expression in liver cells in vitro and in vivo. In addition, it exhibits liver toxicity in Kunming mice at concentrations higher than 50 mg/kg. The CYP-inductive effect that is caused by muscone encompasses a 6- to 12-day period of activity after drug administration as demonstrated by follow-up in vivo studies.

Evaluation of the in vitro and in vivo metabolic pathway and cytochrome P450 inhibition/induction profile of Huperzine A.

Huperzine A (HupA), one of the reversible and selective acetylcholinesterase inhibitors derived from Chinese herb Huperzia Serrata, possesses affirmative action of ameliorating cognitive dysfunction of Alzheimer's disease. Up to now, the effects of HupA on human cytochrome P450s (CYPs) have not been fully elucidated. The purpose of the present study was to clarify the metabolic pathway of HupA in vitro and in vivo, and to evaluate the CYPs inhibition/induction profile of HupA in vitro. The catalytic activity of CYP enzymes (CYP1A2, 2A6, 2C9, 2C19, 2D6, 2E1 and 3A4) was measured by the quantification of specific enzyme substrates using validated liquid chromatography-tandem mass spectrometry (LC/MS/MS) methods. The in vivo metabolic pathway evaluation was performed in an open, single-dose pharmacokinetic study of HupA in fourteen elderly subjects, with urine collecting at certain intervals. In human liver microsomes, HupA (10 ng/mL) was not metabolized within 90 min, and it showed negligible inhibition against these CYP isoforms within 0.2-100 ng/mL. In human liver hepatocytes, the activities of CYP1A2 and CYP3A4 were not significantly altered when incubated at 2 or 20 ng/mL of HupA. After oral administration of 0.1 mg HupA, the total proportion of HupA excreted through urine was relatively high, accounting to 35± 9% at the limited time period of 48 h. These results suggest that HupA is substantially excreted by kidney unchanged rather than metabolized by human liver, and is unlikely to cause clinically relevant drug-drug interaction (DDI) when co-administrated with drugs that are metabolized by CYP isoenzyme system.

[Effect of xenobiotics on microRNA expression in rat liver].

Using bioinformatics analysis we selected microRNAs which could bind 3'-UTR-region of cytochrome P450 (CYP) genes. Three microRNA miR-21, -221, -222, their potential targets might be mRNA for CYP1A1, and two microRNA miR-143, miR-152 for CYP2B1 accordingly were selected for experimental verification. Expression level of these microRNAs in rat liver upon benzo(a)pyrene (BP), phenobarbital (PB), and DDT induction was determined using RT-qPCR method. In rats treated by both BP, and DDT the hepatic content of miR-21, -221, -222 significantly demonstrated a 2-3-fold decrease. The decrease in miR expression was accompanied by a considerable (5.5-8.7-fold) increase in the CYP1A1-mediated EROD activity. The expression of miR-143 remained unchanged after the PB treatment, while the expression of miR-152 increased by 2 times, however, the (10.5-fold) increase in PROD activity of CYP2B was much higher. In the DDT-treated liver PROD activity increased by 20 times, the expression of miR-152 didn't change, and the expression of miR-143 increased by 2 times. The bioinformatics analysis of interactions between microRNAs and targets showed that the studied miRs can potentially bind 3'-end of AhR, ESR1, GR, CCND1, PTEN mRNA. Thus, the expression profile of miR-21, -221, -222, -143, -152 might change under the xenobiotics exposure. In silico analysis confirmed, that microRNAs target not only cytochrome P450 mRNA but also other genes, including those involved in hormonal carcinogenesis, they also can be regulated with studied miRs.

Pharmacokinetic Drug Interactions with Tobacco, Cannabinoids and Smoking Cessation Products.

Tobacco smoke contains a large number of compounds in the form of metals, volatile gases and insoluble particles, as well as nicotine, a highly addictive alkaloid. Marijuana is the most widely used illicit drug of abuse in the world, with a significant increase in the USA due to the increasing number of states that allow medical and recreational use. Of the over 70 phytocannabinoids in marijuana, Δ9-tetrahydrocannabinol (Δ9THC), cannabidiol (CBD) and cannibinol are the three main constituents. Both marijuana and tobacco smoking induce cytochrome P450 (CYP) 1A2 through activation of the aromatic hydrocarbon receptor, and the induction effect between the two products is additive. Smoking cessation is associated with rapid downregulation of CYP1A enzymes. On the basis of the estimated half-life of CYP1A2, dose reduction of CYP1A drugs may be necessary as early as the first few days after smoking cessation to prevent toxicity, especially for drugs with a narrow therapeutic index. Nicotine is a substrate of CYP2A6, which is induced by oestrogen, resulting in lower concentrations of nicotine in females than in males, especially in females taking oral contraceptives. The significant effects of CYP3A4 inducers and inhibitors on the pharmacokinetics of Δ9THC/CBD oromucosal spray suggest that CYP3A4 is the primary enzyme responsible for the metabolism of Δ9THC and CBD. Limited data also suggest that CBD may significantly inhibit CYP2C19. With the increasing use of marijuana and cannabis products, clinical studies are needed in order to determine the effects of other drugs on pharmacokinetics and pharmacodynamics.

Is montelukast as effective as N-acetylcysteine in hepatic injury due to acetaminophen intoxication in rats?

This study aims to investigate the acute protective effect of montelukast sodium in hepatic injury secondary to acetaminophen (APAP) intoxication. This study used 60 rats. The rats were grouped into 6 groups. The control group was administered oral distilled water 10 ml/kg, the APAP group oral APAP 1 g/kg, the montelukast sodium (MK) group oral MK 30 mg/kg, the acetaminophen+N-acetylcysteine (APAP+NAC) group oral APAP 1 g/kg, followed by a single dose of intraperitoneal NAC 1.5 g/kg three hours later, the acetaminophen+montelukast sodium (APAP+MK) group oral APAP 1 g/kg, followed by oral MK 30 mg/kg 3 h later, the acetaminophen+N-acetylcysteine+montelukast sodium (APAP+NAC+MK) group oral APAP 1 g/kg, followed by a single intraperitoneal NAC 1.5 g/kg plus oral MK 30 mg/kg 3 h later. Blood and liver tissue samples were taken 24h after drug administration. Aspartate aminotransferase (AST), alanine aminotransferase (ALT), alkaline phosphatase (ALP), and total bilirubin were studied from the blood samples. Liver tissue samples were used for histopathological examination. Compared with the control group, serum AST and ALT activities were higher in the APAP and APAP+NAC groups. APAP+NAC, APAP+MK, and APAP+NAC+MK groups had reduced serum ALT and AST activities than the group administered APAP alone. APAP+MK and APAP+NAC+MK groups had a lower serum ALP activity than the control group. Histopathologically, there was a difference between the group administered APAP alone and the APAP+MK and APAP+NAC+MK groups. MK is as protective as NAC in liver tissue in APAP intoxication in rats.

Auto-Induction Effect of Chloroxoquinoline on the Cytochrome P450 Enzymes of Rats Associated with CYP 3A and 1A.

To investigate the auto-induction of cytochrome P450 (CYP450) by Chloroxoquinoline (CXL), a novel anticancer drug. Three experiments related to the induction of CYP450 were performed: a) In vitro use of the rat fresh hepatocytes model; b) In vivo 'cocktail' of CYP450 probe model; c) Pharmacokinetic (PK) study of the single and multiple doses. Some typical CYP enzyme probes and inducers were used in these experiments and were all determined by HPLC-MS/MS. The expression levels of CYP3A and CYP1A mRNA were analyzed by the real time polymerase chain reaction (RT-PCR) technique. The PK studies showed that the area under the curve (AUC0-t) and the peak concentration (Cmax) of the multiple doses were approximately 2.4-fold and 1.9-fold lower than those of the single dose, respectively (p < 0.05). Subsequent studies were conducted to study the possible induction of CXL on CYP 450. The in vivo 'cocktail' administration of CYP450 probe model indicated that 5 d pretreatment with CXL resulted in a mean 4.6 times increase in the metabolites/probe plasma ratios for CYP 3A and a 336% increase for CYP 1A than those of the negative control (p < 0.05). The induction effect of CXL on CYP450 was further evaluated on rat hepatocytes with four concentrations (1, 10, 50 and 100 µmol/L). Compared with the negative control, the mRNA levels of CYP 1A2 increased significantly in rat hepatocytes after treatment with 10, 50 and 100 µmol/L CXL (p < 0.05). While significant inductions of CYP 3A1 were observed in the entire treated groups. The results of the present study demonstrate enhanced and induced expression of CYP 3A and CYP 1A in response to CXL exposure in rats, suggesting that CXL is an auto-inducer of CYP 3A and CYP 1A.

Potentially Unsafe Herb-drug Interactions Between a Commercial Product of Noni Juice and Phenytoin- A Case Report.

PURPOSE: To report the unsafe herb-drug interactions between a commercial product of noni juice and phenytoin in a human case. CASE REPORT: A 49-year-old-male has been treated with phenytoin for epilepsy for more than ten years. In spite of his medication adherence, persistent sub-therapeutic phenytoin levels, which were sometimes from low to undetectable, with the result of having poor seizure control were noted as the noni fruit juice was co-administered daily. The possible mechanism is speculated to be due to noni juiceinduced cytochrome P-450 2C9 metabolism of phenytoin. Owing to many beneficial effects of noni juice, the patient was unwilling to accept our advice to quit taking it. Clobazam treatment was added, and with gradually reducing the amount of juice drunk over six months, the patient's epilepsy has been well controlled. Now only auras along with sometimes minor partial seizures occur, but no major attack has been reported for more than one year. CONCLUSION: Phenytoin had been commonly used for seizure control worldwide and nearly half of patients with epilepsy had received complementary and alternative medicine in Taiwan. Thus, this report is significantly important for clinicians to be aware of the interaction between antiepileptic drugs and some herbs like noni juice. Moreover, as far as we know, this is a rare human case that is reported to disclose this unfavorable herb-drug interaction.

Evaluation of 309 molecules as inducers of CYP3A4, CYP2B6, CYP1A2, OATP1B1, OCT1, MDR1, MRP2, MRP3 and BCRP in cryopreserved human hepatocytes in sandwich culture.

1. Regulation of hepatic metabolism or transport may lead to increase in drug clearance and compromise efficacy or safety. In this study, cryopreserved human hepatocytes were used to assess the effect of 309 compounds on the activity and mRNA expression (using qPCR techniques) of CYP1A2, CYP2B6 and CYP3A4, as well as mRNA expression of six hepatic transport proteins: OATP1B1 (SCLO1B1), OCT1 (SLC22A1), MDR1 (ABCB1), MRP2 (ABCC2), MRP3 (ABCC3) and BCRP (ABCG2). 2. The results showed that 6% of compounds induced CYP1A2 activity (1.5-fold increase); 30% induced CYP2B6 while 23% induced CYP3A4. qPCR data identified 16, 33 or 32% inducers of CYP1A2, CYP2B6 or CYP3A4, respectively. MRP2 was induced by 27 compounds followed by MDR1 (16)>BCRP (9)>OCT1 (8)>OATP1B1 (5)>MRP3 (2). 3. CYP3A4 appeared to be down-regulated (≥2-fold decrease in mRNA expression) by 53 compounds, 10 for CYP2B6, 6 for OCT1, 4 for BCRP, 2 for CYP1A2 and OATP1B1 and 1 for MDR1 and MRP2. 4. Structure-activity relationship analysis showed that CYP2B6 and CYP3A4 inducers are bulky lipophilic molecules with a higher number of heavy atoms and a lower number of hydrogen bond donors. Finally, a strategy for testing CYP inducers in drug discovery is proposed.

Cooperation of structurally different aryl hydrocarbon receptor agonists and ß-catenin in the regulation of CYP1A expression.

The ligand-activated nuclear receptor AhR (aryl hydrocarbon receptor) mediates the response of hepatocytes to various exogenous compounds. AhR is classically activated by planar, aromatic hydrocarbons, but also by other, structurally rather unrelated compounds. Recent data show that the canonical Wnt/ß-catenin signaling pathway is also involved in the regulation of hepatic zonal gene expression and drug metabolism in mammalian liver. Previous studies indicate that the loss of ß-catenin in hepatocytes diminishes the response to the AhR agonists 3-methylcholanthrene (3MC) in vivo and to 2,3,7,8-tetrachlorodibenzo-[p]-dioxin in vitro. The knockout of ß-catenin also impairs the zonal pattern of AhR target gene induction by 3MC. However, it is presently unknown whether the chemical nature of the AhR agonist influences the AhR/ß-catenin interaction. Moreover, no information is available about the dose-response curves of AhR activation in the absence or presence of Wnt/ß-catenin signaling. In the present study, we have analyzed AhR-dependent responses to different concentrations of structurally unrelated AhR agonists in vivo and in vitro. The results demonstrate that ß-catenin is essential to obtain the maximum AhR response. Moreover, using transgenic mouse models which allow for the ablation of ß-catenin at different age of mice, we demonstrate that the presence of ß-catenin, not postnatal developmental effects in ß-catenin-deficient livers, is responsible for the observed interplay of ß-catenin and the AhR.

Aggregate culture of human embryonic stem cell-derived hepatocytes in suspension are an improved in vitro model for drug metabolism and toxicity testing.

Early phase drug development relies on primary human hepatocytes for studies of drug metabolism, cytotoxicity, and drug-drug interactions. However, primary human hepatocytes rapidly lose metabolic functions ex vivo and are refractory to expansion in culture and thus are limited in quantity. Hepatocytes derived from human pluripotent stem cells (either embryonic stem (ES) or induced pluripotent stem (iPS) cells), have the potential to overcome many of the limitations of primary human hepatocytes, but to date the use of human pluripotent stem cell-derived hepatocytes has been limited by poor enzyme inducibility and immature metabolic function. Here, we present a simple suspension culture of aggregates of ES cell-derived hepatocytes that compared to conventional monolayer adherent culture significantly increases induction of CYP 1A2 by omeprazole and 3A4 by rifampicin. Using liquid chromatography-tandem mass spectrometry, we further show that ES cell-derived hepatocytes in aggregate culture convert omeprazole and rifampicin to their human-specific metabolites. We also show that these cells convert acetaminophen (APAP) to its cytotoxic metabolite (N-acetyl-p-benzoquinone imine (NAPQI)), although they fail to perform APAP glucuronidation. In summary, we show that human pluripotent stem cell-derived hepatocytes in aggregate culture display improved enzymatic inducibility and metabolic function and is a promising step toward a simple, scalable system, but nonetheless will require further improvements to completely replace primary human hepatocytes in drug development.