Drug interaction potential of toremifene and N-desmethyltoremifene with multiple cytochrome P450 isoforms

1. Toremifene is an effective agent for the treatment of breast cancer in postmenopausal women and is being evaluated for its ability to prevent bone fractures in men with prostate cancer taking androgen deprivation therapy.

2. Due to the potential for drug–drug interactions, the ability of toremifene and its primary circulating metabolite N-desmethyltoremifene (NDMT) to inhibit nine human cytochrome P450 (CYP) enzymes was determined using human liver microsomes. Induction of CYP1A2 and 3A4 by toremifene was also investigated in human hepatocytes.

3. Toremifene did not significantly inhibit CYP1A2 or 2D6. However, toremifene is a competitive inhibitor of CYP3A4, non-competitive inhibitor of CYP2A6, 2C8, 2C9, 2C19 and 2E1 and mixed-type inhibitor of CYP2B6. CYP inhibition by NDMT was similar in magnitude to toremifene. Toremifene did not induce CYP1A2 but increased CYP3A4 monooxygenase activity and gene expression in drug-exposed human primary hepatocytes.

4. Although clinical doses of toremifene produce steady state exposures to toremifene and NDMT that may be sufficient to cause pharmacokinetic drug–drug interactions with other drugs metabolised by CYP2B6, CYP2C8, CYP3A4, CYP2C9 and CYP2C19, these data indicate that toremifene is unlikely to play a role in clinical drug–drug interactions with substrate drugs of CYP1A2 and CYP2D6.

     

Equally potent inhibitors of cholesterol synthesis in human hepatocytes have distinguishable effects on different cytochrome P450 enzymes

Six 3-hydroxy-3-methylglutaryl coenzyme A (HMG-CoA) reductase inhibitors (the present cholesterol-lowering drugs known as statins), lovastatin (L), simvastatin (S), pravastatin (P), fluvastatin (F), atorvastatin (A) and cerivastatin (C) are shown to be potent inhibitors of cholesterol synthesis in human hepatocytes, the target tissue for these drugs in man. All six inhibited in the nM range (IC(50) values: 0.2-8.0 nM). As daily used cholesterol-lowering drugs they are likely coadministered with other drugs. While several cytochrome P450 (CYP) enzymes are involved in drug metabolism in the liver and thus play an important role in drug-drug interaction it was investigated which of these enzymes are influenced by the active forms of the six statins. These enzyme activities were studied in human liver microsomal preparations, and in simian and human hepatocytes in primary culture. The following CYP reactions were used: nifedipine aromatization (CYP3A4), testosterone 6beta-hydroxylation (CYP3A4), tolbutamide methylhydroxylation (CYP2C9), S-mephenytoin 4-hydroxylation (CYP2C19), bufuralol 1'-hydroxylation (CYP2D6), aniline 4-hydroxylation (CYP2E1), coumarin 7-hydroxylation (CYP2A6) and 7-ethoxyresorufin O-dealkylation (CYP1A1/2). In the human liver microsomes the statins (concentrations up to 400 microM) did not influence the CYP1A1/2 activity and hardly the CYP2A6 and CYP2E1 activities. Except P, the other five statins were stronger inhibitors of the CYP2C19 activity with IC(50) values around 200 microM and the same holds for the effect of A, C and F on the CYP2D6 activity. L and S were weaker inhibitors of the latter enzyme activity, whereas P did not influence both activities. About the same was observed for the statin effect on CYP2C9 activity, except that F was a strong inhibitor of this activity (IC(50) value: 4 microM). Using the assay of testosterone 6beta-hydroxylation the CYP3A4 activity was decreased by L, S and F with IC(50) values of about 200 microM and a little more by C and A (IC(50) around 100 microM). P had hardly an effect on this activity. To a somewhat less extent the same trend was seen when CYP3A4 activity was measured using nifedipine as substrate. The inhibitory effects observed in microsomes were verified in suspension culture of freshly isolated hepatocytes from Cynomolgus monkey (as a readily available model) and of human hepatocytes. In general the same trends were seen as in the human microsomes, except that in some cases the inhibition of the CYP activity was less, possibly by the induction of the particular CYP enzyme by incubation of the cells with a particular statin. F remained a strong inhibitor of CYP2C9 activity in human and monkey hepatocytes. A induced the CYP2C9 in monkey hepatocytes but was an inhibitor of the CYP2C9 in human hepatocytes. A, S, L and C were moderate inhibitors in both cellular systems of CYP3A4. P was not affecting any of the CYP activities in the three systems studied. It is concluded that different CYP enzymes interact with different statins and therefore differences in between these drugs are to be expected when drug-drug interaction is considered.     

Preclinical evaluation of the potential for cytochrome P450 inhibition and induction of the selective ALK inhibitor,alectinib

1.?A novel selective anaplastic lymphoma kinase (ALK) inhibitor, alectinib, has shown remarkable efficacy and safety in patients with ALK-positive non-small-cell lung cancer (NSCLC). The purpose of this study was to evaluate in vitro the potential to inhibit and induce cytochrome P450 (CYP) isoforms for alectinib and its major metabolite M4.

2.?Alectinib and M4 did not show the meaningful direct inhibition of six major CYP isoforms (CYP1A2, 2B6, 2C9, 2C19, 2D6 and 3A4) in human liver microsomes (HLM). Alectinib, but not M4, competitively inhibited CYP2C8, by which few marketed drugs are exclusively metabolized, with an inhibition constant of 1.98?μM.

3.?Out of the seven CYP isoforms in HLM, alectinib and M4 showed time-dependent inhibition (TDI) of only CYP3A4, which suggests low TDI potential due to low inactivation efficiency.

4.?Alectinib exhibited quite smaller induction of mRNA expression of CYP1A2, 2B6 and 3A4 genes in human hepatocytes compared to the respective positive controls, suggesting a low potential of enzyme induction.

5.?In summary, the risk of alectinib causing drug-drug interactions with coadministered drugs is expected to be low due to the weak potential of CYP inhibition and induction estimated in the preclinical studies.     

In vitro inhibition and induction of human hepatic cytochrome P450 enzymes by modafinil.

The ability of modafinil to affect human hepatic cytochrome P450 (CYP) activities was examined in vitro. The potential for inhibition of CYP1A2, CYP2A6, CYP2B6, CYP2C9, CYP2C19, CYP2D6, CYP2E1, CYP3A4/5, and CYP4A9/11 by modafinil (5-250 microM) was evaluated with pooled human liver microsomes. Modafinil exhibited minimal capacity to inhibit any CYP enzyme, except CYP2C19. Modafinil inhibited the 4'-hydroxylation of S-mephenytoin, a marker substrate for CYP2C19, reversibly and competitively with a K(i) value of 39 microM, which approximates the steady-state C(max) value of modafinil in human plasma at a dosage of 400 mg/day. No irreversible inhibition of any CYP enzyme was observed, and there was no evidence of metabolism-dependent inhibition. The potential for induction of CYP activity was evaluated by exposing primary cultures of human hepatocytes to modafinil (10-300 microM). Microsomes were then prepared and assayed for CYP1A2, CYP2A6, CYP2B6, CYP2C8, CYP2C9, CYP2C19, CYP2D6, and CYP3A4/5 activities. The mean activities of microsomal CYP1A2, CYP2B6, and CYP3A4/5 from modafinil-treated hepatocytes were higher (up to 2-fold) than those in the solvent-treated controls but were less than those produced by reference inducers of these enzymes. At high concentrations of modafinil (>/=100 microM), the mean activity of CYP2C9 was decreased (up to 60%) relative to that in the solvent controls. Overall, modafinil was shown to have effects on human hepatic CYP1A2, CYP2B6, CYP2C9, CYP2C19, and CYP3A4/5 activities in vitro. Although effects obtained in vitro are not always predictive of effects in vivo, such results provide a rational basis for understanding drug-drug interactions that are observed clinically and for planning subsequent investigations.     

Inhibitory effects of nicardipine to cytochrome P450 (CYP) in human liver microsomes

To anticipate drug-drug interactions by nicardipine in vivo, cytochrome P450 (CYP) forms responsible for the metabolism of nicardipine and inhibition of CYP-dependent drug metabolism by nicardipine were investigated. Microsomes of human B-lymphoblastoid cells expressing each human CYP form were used for the metabolism of nicardipine. Inhibitory effects of nicardipine on drug metabolism were studied using human liver microsomes. CYP2C8, CYP2D6 and CYP3A4 were identified as major CYP forms for the metabolism of nicardipine in human liver microsomes. Nicardipine strongly inhibited two-pathways of triazolam hydroxylation both catalyzed by CYP3A4. Comparison of three Ca(2+) antagonists, nicardipine, nifedipine, and diltiazem revealed that only nicardipine showed such a strong inhibitory potency on the typical CYP2D6-catalyzed drug metabolism. Furthermore, nicardipine inhibited other reactions catalyzed by CYP1A, CYP2A6, CYP2C8, CYP2C9 and CYP2C19 with K(i) values ranging from 1.1 to 29.4 microM. In conclusion, nicardipine was a relatively potent inhibitor of human CYP2D6, CYP3A4 and CYP2C (especially for CYP2C8 and CYP2C19) in vitro, suggesting that drug-drug interactions between nicardipine and other drugs metabolized mainly by these CYP forms appear to occur in vivo.     

In vitro evaluation of the interaction potential of irosustat with drug-metabolizing enzymes

Irosustat is a first-generation, irreversible, steroid sulfatase inhibitor currently in development for hormone-dependent cancer therapy. To predict clinical drug-drug interactions between irosustat and possible concomitantly administered medications, the inhibition/induction potential of irosustat with the main drug-metabolizing enzymes was investigated in vitro. The interaction of aromatase inhibitors in the in vitro metabolism of irosustat was also studied. Irosustat inhibited CYP1A2 activity in human liver microsomes through the formation of its desulfamoylated degradation product and metabolite 667-coumarin. CYP1A2 inhibition by 667-coumarin was competitive, with a K(i) of 0.77 μM, a concentration exceeding by only 5-fold the maximal steady-state concentration of 667-coumarin in human plasma with the recommended dose of irosustat. In addition, 667-coumarin metabolites enhanced the inhibition of CYP1A2 activity. Additional clinical interaction studies of irosustat with CYP1A2 substrate drugs are strongly recommended. 667-Coumarin also appeared to be a competitive inhibitor of CYP2C19 (K(i) = 5.8 μM) in human liver microsomes, and this inhibition increased with assessment in human hepatocytes. Inhibition of CYP2C19 enzyme activity was not caused by repression of CYP2C19 gene expression. Therefore, additional mechanistic experiments or follow-up studies with clinical evaluation are recommended. Irosustat neither inhibited CYP2A6, CYP2B6, CYP2C8, CYP2C9, CYP2D6, CYP2E1, CYP3A4/5, or UDP-glucuronosyltransferase 1A1, 1A4, or 2B7 activities nor induced CYP1A2, CYP2C9, CYP2C19, or CYP3A4/5 at clinically relevant concentrations. Results from human liver microsomes indicated that no changes in irosustat pharmacokinetics in vivo are expected as a result of inhibition of irosustat metabolism in cases of concomitant medication administration or irosustat-aromatase inhibitor combination therapy with letrozole, anastrozole, or exemestane.     

Drug-drug interaction of antifungal drugs

This article reviews the in vitro metabolic and the in vivo pharmacokinetic drug-drug interactions with antifungal drugs, including fluconazole, itraconazole, micafungin, miconazole, and voriconazole. In the in vitro interaction studies, the effects of antifungal drugs on specific activities of cytochrome P450s (CYPs), including CYP1A2, CYP2C9, CYP2C19, CYP2D6, CYP2E1, and CYP3A4, in human liver microsomes are compared to predict the possibility of drug interactions in vivo. Fluconazole, micafungin, and voriconazole have lower inhibitory effects on CYP3A4 activities than itraconazole and miconazole, and IC(50) and/or K(i) values against CYP2C9 and CYP2C19 activities are the lowest for miconazole, followed by voriconazole and fluconazole. In in vivo pharmacokinetic studies, it is well known that itraconazole is a potent clinically important inhibitor of the clearance of CYP3A4 substrates, and fluconazole and voriconazole are reported to increase the blood or plasma concentrations of not only midazolam and cyclosporine (CYP3A4 substrates) but also of phenytoin (CYP2C9 substrate) and/or omeprazole (CYP2C19/CYP3A4 substrate). On the other hand, no inhibition of CYP activities except for CYP3A4 activity by micafungin is observed in vitro, and the blood concentrations of cyclosporine and tacrolimus are not affected by coadministration of micafungin in vivo, suggesting that micafungin would not cause clinically significant interactions with drugs that are metabolized by CYPs via the inhibition of metabolism. Miconazole is a potent inhibitor of all CYPs investigated in vitro, although there are few detailed studies on the clinical significance of this except for CYP2C9. Therefore the differential effects of these antifungal drugs on CYP activities must be considered in the choice of antifungal drugs in patients receiving other drugs.     

P450酶介导的EGFR-TKIs药物与其他药物的相互作用

目的探讨细胞色素P450酶介导的表皮生长因子受体酪氨酸激酶抑制剂(EGFR-TKIs)与其他药物的相互作用。方法查阅文献,统计国家药品监督管理局批准用于治疗晚期非小细胞肺癌的7个EGFR-TKIs与P450酶相关的药物相互作用。结果 7个EGFR-TKIs中,阿法替尼的代谢不经过P450酶途径,也并非CYP酶系的诱导剂或抑制剂;其余EGFR-TKIs主要经CYP3A4酶、CYP2D6酶及CYP1A2酶代谢,并可明显抑制CYP2D6酶、CYP3A4酶、CYP2C8/9酶的活性,药物相互作用较多。结论 EGFR-TKIs类药物中,除阿法替尼与P450酶底物发生相互作用的概率较低外,其余药物对P450酶均可产生明显的抑制作用,临床医师应结合患者病情合理选用该类药物,避免不合理合用。     

Role of human cytochrome P450 3A4 in the metabolism of DA-8159, a new erectogenic

The purpose of this paper was to characterize cytochrome P450 (CYP) enzymes involved in N-dealkylation of a new oral erectogenic, DA-8159 to DA-8164, a major circulating active metabolite, in human liver microsomes and to investigate the inhibitory potential of DA-8159 on CYP enzymes. CYP3A4 was identified as the major enzyme responsible for DA-8159 N-dealkylation to DA-8164 based on correlation analysis and specific CYP inhibitor and antibody-mediated inhibition study in human liver microsomes, and DA-8159 metabolism in cDNA expressed CYP enzymes. There is the possibility of drug-drug interactions when prescribing DA-8159 concomitantly with known inhibitors or inducers of CYP3A4. DA-8159 was found to be only a very weak inhibitor of eight major CYPs (1A2, 2A6, 2C8, 2C9, 2C19, 2D6, 2E1 and 3A4), the largest inhibition occurring against CYP2D6 (IC5o 67.7 microM) in human liver microsomes. Drug-drug interactions would not be predicted on the basis of DA-8159 inhibiting the metabolism of coadministered drugs.     

Multiple cytochrome P-450s involved in the metabolism of terbinafine suggest a limited potential for drug-drug interactions.

Biotransformation pathways and the potential for drug-drug interactions of the orally active antifungal terbinafine were characterized using human liver microsomes and recombinant human cytochrome P-450s (CYPs). The terbinafine metabolites represented four major pathways: 1) N-demethylation, 2) deamination, 3) alkyl side chain oxidation, and 4) dihydrodiol formation. Michaelis-Menten kinetics for the pathways revealed mean K(m) values ranging from 4.4 to 27.8 microM, and V(max) values of 9.8 to 82 nmol/h/mg protein. At least seven CYP enzymes are involved in terbinafine metabolism. Recombinant human CYPs predict that CYP2C9, CYP1A2, and CYP3A4 are the most important for total metabolism. N-demethylation is primarily mediated by CYP2C9, CYP2C8, and CYP1A2; dihydrodiol formation by CYP2C9 and CYP1A2; deamination by CYP3A4; and side chain oxidation equally by CYP1A2, CYP2C8, CYP2C9, and CYP2C19. Additionally, characteristic CYP substrates inhibited pathways of terbinafine metabolite formation, confirming the involvement of multiple enzymes. The deamination pathway was mainly inhibited by CYP3A inhibitors, including troleandomycin and azole antifungals. Dihydrodiol formation was inhibited by the CYP1A2 inhibitor furafylline. Terbinafine had little or no effect on the metabolism of many characteristic CYP substrates. Terbinafine, however, is a competitive inhibitor of the CYP2D6 reaction, dextromethorphan O-demethylation (K(i) = 0.03 microM). In summary, terbinafine is metabolized by at least seven CYPs. The potential for terbinafine interaction with other drugs is predicted to be insignificant with the exception that it may inhibit the metabolism of CYP2D6 substrates. Clinical trials are needed to assess the relevance of these findings.     

In vitro assessment of metabolic drug–drug interaction potential of AZD2624, neurokinin-3 receptor antagonist, through cytochrome P(450) enzyme identification, inhibition, and induction studies

AZD2624 was pharmacologically characterized as a NK3 receptor antagonist intended for treatment of schizophrenia. The metabolic drug-drug interaction potential of AZD2624 was evaluated in in vitro studies. CYP3A4 and CYP3A5 appeared to be the primary enzymes mediating the formation of pharmacologically active ketone metabolite (M1), whereas CYP3A4, CYP3A5, and CYP2C9 appeared to be the enzymes responsible for the formation of the hydroxylated metabolite (M2). The apparent K(m) values were 1.5 and 6.3 μM for the formation of M1 and M2 in human liver microsomes, respectively. AZD2624 exhibited an inhibitory effect on microsomal CYP3A4/5 activities with apparent IC(50) values of 7.1 and 19.8 μM for midazolam and testosterone assays, respectively. No time-dependent inactivation of CYP3A4/5 activity (midazolam 1'-hydroxylation) by AZD2624 was observed. AZD2624 demonstrated weak to no inhibition of CYP1A2, CYP2B6, CYP2C8, CYP2C9, CYP2C19, and CYP2D6. AZD2624 was not an inducer of CYP1A2 or CYP2B6. Although AZD2624-induced CYP3A4 activity in hepatocytes, the potential of AZD2624 to cause inductive drug interactions of this enzyme was low at relevant exposure concentration. Together with targeted low efficacious concentration, the results of this study demonstrated AZD2624 has a relatively low metabolic drug-drug interaction potential towards co-administered drugs. However, metabolism of AZD2624 might be inhibited when co-administrated with potent CYP3A4/5 inhibitors.     

Evaluation of drug-drug interaction potential of beraprost sodium mediated by P450 in vitro

Beraprost sodium (BPS), a chemically stable and orally active prostacyclin analogue used for the treatment of chronic occlusive disease and primary pulmonary hypertension, was investigated in terms of its drug-drug interaction mediated by cytochrome P450. In a metabolic enzyme characterization study using P450-expressing insect cell microsomes, beraprost (BP) was slightly metabolized in the presence of CYP2C8, but not metabolized by the other P450 isoforms (CYP1A1, CYP2A6, CYP2B6, CYP2C9, CYP2C19, CYP2D6, CYP2E1, CYP3A4, and CYP4A11) at a concentration of 20 microM. These results suggest that none of the P450 isoforms is a major metabolic enzyme of BP. In a P450 induction study using human hepatocytes, BP did not induce any P450 isoform (CYP1A2, CYP2C9, CYP2C19, and CYP3A4) at concentrations of 1-100 microM. Furthermore, in a P450 inhibition study using human liver microsomes, BP did not inhibit any P450 isoform (CYP1A2, CYP2A6, CYP2C8, CYP2C9, CYP2C19, CYP2D6, and CYP3A4) at concentrations of 0.05-1 microM. Therefore it is concluded that BP is not involved in drug-drug interaction mediated by P450 isoforms.     

Determination of the human cytochrome P450 isoforms involved in the metabolism of zolmitriptan

1. Zolmitriptan was extensively metabolized by freshly isolated human hepatocytes to a number of components including the three main metabolites observed in vivo (N-desmethyl-zolmitriptan, zolmitriptan N-oxide and the indole acetic acid derivative). In contrast, metabolism of zolmitriptan by human hepatic microsomes was extremely limited with only small amounts of the N-desmethyl and indole ethyl alcohol metabolites being produced. 2. Furafylline, a selective inhibitor of CYP1A2, almost completely abolished the hepatocellular metabolism of zolmitriptan and markedly inhibited formation of the N-desmethyl metabolite in microsomes. Chemical inhibitors, selective against other major human cytochrome P450 (CYP2C9, 2C19, 2D6 and 3A4), had no obvious effects. In addition, expressed human CYP1A2 was the only cytochrome P450 to form the N-desmethyl metabolite. 3. N-desmethyl-zolmitriptan was extensively metabolized by both human hepatocytes and microsomes. The indole acetic acid and ethyl alcohol derivatives were the major metabolites formed by hepatocytes, whereas only the indole ethyl alcohol derivative was produced by microsomes. Metabolism of N-desmethyl-zolmitriptan was not inhibited by cytochrome P450-selective chemical inhibitors nor was it observed following incubation with expressed human cytochrome P450. Clorgyline, a selective inhibitor of monoamine oxidase A (MAO-A), markedly inhibited the microsomal formation of the indole ethyl alcohol derivative. 4. Primary metabolism of zolmitriptan is dependent mainly on CYP1A2, whereas MAO-A is responsible for further metabolism of N-desmethyl-zolmitriptan, the active metabolite. Since the in vivo clearance of zolmitriptan is primarily dependent on metabolism, interactions with drugs that induce or inhibit CYP1A2 or MAO-A may be anticipated.     

Metabolism of vanoxerine, 1-[2-[bis(4-fluorophenyl)methoxy]ethyl]-4-(3-phenylpropyl)piperazine, by human cytochrome P450 enzymes.

Vanoxerine (1-[2-[bis(4-fluorophenyl)methoxy]ethyl]-4-(3-phenylpropyl)piperazine; GBR12909) is a promising agent for the treatment of cocaine dependence. Knowledge of the major pathway for GBR12909 metabolism is important for prediction of the likelihood of drug-drug interactions, which may affect the therapeutic clinical outcome, when this agent is used in cocaine-dependent individuals receiving multiple drug therapy. We studied biotransformation of GBR12909 in human liver microsomes (n = 4), human hepatocytes, and microsomes containing cDNA-expressed human P450 isoforms with GBR12909 concentrations within the range of steady-state plasma concentrations detected in healthy volunteers. A high-pressure liquid chromatography assay was used to measure parent GBR12909 and its primary metabolite. GBR12909 was metabolized by human liver microsomes, hepatocytes, and cDNA-expressed human P450s to a single metabolite. Ketoconazole, a selective inhibitor of CYP3A, reduced GBR12909 biotransformation in human liver microsomes and primary hepatocytes by 92 +/- 2 and 92.4 +/- 0.4%, respectively. Quercetin (an inhibitor of CYP2C8/3A4) was a less effective inhibitor producing 62 +/- 22% inhibition in human liver microsomes and 54 +/- 35% in hepatocytes. Other P450 selective inhibitors did not decrease GBR12909 biotransformation more than 29% in either human liver microsomes or hepatocytes with the exception of chlorzoxazone (CYP2E1), which inhibited GBR12909 biotransformation by 71.4 +/- 18.5% in primary human hepatocytes. Ciprofloxacin (CYP1A2), sulfaphenazole (CYP2C9), quinidine (CYP2D6), chlorzoxazone (CYP2E1), and mephenytoin (CYP2C19) did not demonstrate statistically significant inhibition (p > 0.05) of GBR12909 biotransformation in liver microsomes. cDNA-expressed P450 3A4 metabolized GBR12909 to a greater extent than 2C8 and 2E1. These data suggest the possibility that multiple P450 isoforms may be involved in human GBR12909 metabolism but that CYP3A appears to be the major enzyme responsible for human GBR12909 biotransformation.     

豆腐果苷对人肝微粒体CYP450酶体外抑制作用研究

目的:通过评价豆腐果苷在体外对人肝微粒体CYP450酶的7种亚型酶活性的影响,预测服用豆腐果苷可能出现的食物-药物及药物-药物代谢的影响。方法:将豆腐果苷与CYP450酶7种亚型的特异性探针底物咖啡因(CYP1A2)、右美沙芬(CYP2D6)、甲苯磺丁脲(CYP2C9)、S-美芬妥因(CYP2C19)、氯唑沙宗(CYP2E1)、香豆素(CYP2A6)及咪达唑仑(CYP3A4)与人肝微粒体进行孵育反应,采用HPLC和LC-MS/MS法测定对应的7种代谢产物(1,7-二甲基黄嘌呤、去甲右美沙芬、4-羟基甲苯磺丁脲、4-羟基美芬妥因、6-羟基氯唑沙宗、7-羟基香豆素和1-羟基咪达唑仑)的浓度,与对照组比较,确定豆腐果苷对以上7种亚酶活性的影响。结果:豆腐果苷在1～100μmol.L-1时对7种酶的抑制作用均无明显统计学意义(P>0.05)。结论:豆腐果苷可能不会引起有临床意义的CYP450酶抑制现象的发生。     

In vitro studies investigating the interactions between degarelix, a decapeptide gonadotropin-releasing hormone blocker, and cytochrome P450

The decapeptide degarelix is a novel competitive gonadotropin-releasing hormone receptor antagonist that has been approved for the treatment of advanced prostate cancer by the FDA and the EU authorities. In this study, the interaction of degarelix with human cytochrome P450 (CYP450) enzymes was investigated in vitro. Inhibition of CYP450 was performed in human liver microsomes using documented marker substrates for the CYP450 isozymes CYP1A2, CYP2B6, CYP2C8, CYP2C9, CYP2C19, CYP2D6, CYP3A4 and CYP2E1. The inhibitory effects on selected P450 enzyme activities were investigated with degarelix concentrations representing the range of 2-200 times of expected clinical concentrations. No inhibition of any isozyme-catalysed biotransformations studied was detected. Induction of CYP450 enzyme activity by degarelix was investigated using primary human hepatocytes. Cryopreserved plateable hepatocytes and fresh hepatocytes in culture were treated for two-three consecutive days with degarelix at concentrations of 0.1, 1.0 and 10 μM. The cultured hepatocytes were also treated with three prototypical CYP450 inducers: omeprazole, phenobarbital and rifampin as positive controls for CYP450 enzyme induction. No induction of the activity of CYP1A2, CYP2B6, CYP2C8, CYP2C9, CYP2C19 and CYP3A4 isozymes was observed. Degarelix appears to be a poor substrate of the CYP450 enzyme system, and the in vitro results indicate that the interaction between CYP450 and degarelix is low. These results indicate that degarelix is unlikely to cause any clinically significant drug-drug interactions in vivo.     

甲基原薯蓣皂苷对人肝微粒体中7种CYP450酶活性的影响

目的研究甲基原薯蓣皂苷对CYP450酶的7种亚型酶活性的影响。方法将MPD和CYP450酶7种亚型的特异性探针底物咖啡因(CYP1A2)、右美沙芬(CYP2D6)、甲苯磺丁脲(CYP2C9)、s-美芬妥因(CYP2C19)、氯唑沙宗(CYP2E1)、香豆素(CYP2A6)及咪达唑仑(CYP3A4)与人肝微粒体进行孵化反应,采用HPLC和LC-MS/MS法测定对应的7种代谢产物(1,7-二甲基黄嘌呤、去甲右美沙芬、4-羟基甲苯磺丁脲、4-羟基美芬妥因、6-羟基氯唑沙宗、7-羟基香豆素和1-羟基咪达唑仑)的浓度,通过与对照组比较,确定MPD对以上7种酶活性的影响。结果MPD在1~10μmol.L-1时对7种酶均无明显抑制作用,在100μmol.L-1时对CYP2D6有抑制趋势,但对其他6种酶无抑制作用,均无统计学意义(P>0.05)。结论MPD在与以上6种酶(CYP1A2、CYP2E1、CYP2C19、CYP3A4、CYP2C9和CYP2A6)代谢的药物联合用药时,发生药物相互作用的可能性较小。     

Role of human cytochrome P450 3A4 in the metabolism of DA–8159, a new erectogenic

1.?The purpose of this paper was to characterize cytochrome P450 (CYP) enzymes involved in N-dealkylation of a new oral erectogenic, DA-8159 to DA-8164, a major circulating active metabolite, in human liver microsomes and to investigate the inhibitory potential of DA-8159 on CYP enzymes.

2.?CYP3A4 was identified as the major enzyme responsible for DA-8159 N-dealkylation to DA-8164 based on correlation analysis and specific CYP inhibitor and antibody-mediated inhibition study in human liver microsomes, and DA-8159 metabolism in cDNA expressed CYP enzymes. There is the possibility of drug-drug interactions when prescribing DA-8159 concomitantly with known inhibitors or inducers of CYP3A4.

3.?DA-8159 was found to be only a very weak inhibitor of eight major CYPs (1A2, 2A6, 2C8, 2C9, 2C19, 2D6, 2E1 and 3A4), the largest inhibition occurring against CYP2D6 (IC₅₀ 67.7?μM) in human liver microsomes. Drug–drug interactions would not be predicted on the basis of DA-8159 inhibiting the metabolism of coadministered drugs.     

Drug interaction studies with esomeprazole, the (S)-isomer of omeprazole

Esomeprazole, the (S)-isomer of omeprazole, is the first proton pump inhibitor (PPI) developed as a single isomer for the treatment of patients with acid related diseases. Because of the extensive use of PPIs, the documentation of the potential for drug interactions with esomeprazole is of great importance. Altered absorption or metabolism are 2 of the major mechanisms for drug-drug interactions. Since intragastric pH will increase with esomeprazole treatment, it can be hypothesised that the absorption of drugs with pH-sensitive absorption (e.g. digoxin and ketoconazole) may be affected. Esomeprazole does not seem to have any potential to interact with drugs that are metabolised by cytochrome P450 (CYP) 1 A2, 2A6, 2C9, 2D6 or 2E1. In drug interaction studies with diazepam, phenytoin and (R)-warfarin, it was shown that esomeprazole has the potential to interact with CYP2C19. The slightly altered metabolism of cisapride was also suggested to be the result of inhibition of a minor metabolic pathway for cisapride mediated by CYP2C19. Esomeprazole did not interact with the CYP3A4 substrates clarithromycin (2 studies) or quinidine. Since the slightly increased area under the concentration-time curve (AUC) of cisapride could be explained as an inhibition of CYP2C19, the data on these 3 CYP3A4 substrates indicate that esomeprazole does not have the potential to inhibit this enzyme. The minor effects reported for diazepam, phenytoin, (R)-warfarin, and cisapride are unlikely to be of clinical relevance. Clarithromycin interacts with the metabolism of esomeprazole resulting in a doubling of the AUC of esomeprazole. The increased plasma concentrations of esomeprazole are unlikely to have any safety implications. It can be concluded that the potential for drug-drug interactions with esomeprazole is low, and similar to that reported for omeprazole.     

Determination of major human cytochrome P450s activities in 96-well plates using liquid chromatography tandem mass spectrometry.

At the early stage of drug discovery, thousands of new chemical entities (NCEs) may be screened before a single candidate can be identified for development. Evaluation of the effect of NCEs on human CYP450 enzyme activities is a key issue in pharmaceutical development as it may explain inter-subject variability, drug-drug interactions, non-linear pharmacokinetics and toxic effects. A liquid chromatography tandem mass spectrometry (HPLC-MS/MS) method has been developed for the fast and routine analysis of major human CYP450s enzyme activities (CYP1A2, CYP2A6, CYP2B6, CYP2C9, CYP2C19, CYP2D6, CYP2E1 and CYP3A4) in primary hepatocyte cell cultures. The high sensitivity and selectivity of mass spectrometry has allowed traditional assays to be minimized, thus enabling the use of 96-well plate format which markedly reduced the number of hepatocytes needed for each cytochrome CYP450 activity measurement, a fact that is particularly critical concerning human hepatocytes.