Potent cytochrome P450 2C19 genotype-related interaction between voriconazole and the cytochrome P450 3A4 inhibitor ritonavir

OBJECTIVES: Cytochrome P450 (CYP) 2C19 and CYP3A4 are the major enzymes responsible for voriconazole elimination. Because the activity of CYP2C19 is under genetic control, the extent of inhibition with a CYP3A4 inhibitor was expected to be modulated by the CYP2C19 metabolizer status. This study thus assessed the effect of the potent CYP3A4 inhibitor ritonavir after short-term administration on voriconazole pharmacokinetics in extensive metabolizers (EMs) and poor metabolizers (PMs) of CYP2C19. METHODS: In a randomized, placebo-controlled crossover study, 20 healthy participants who were stratified according to CYP2C19 genotype received oral ritonavir (300 mg twice daily) or placebo for 2 days. Together with the first ritonavir or placebo dose, a single oral dose of 400 mg voriconazole was administered. Voriconazole was determined in plasma and urine by liquid chromatography-mass spectrometry, and pharmacokinetic parameters were estimated by noncompartmental analysis. RESULTS: When given alone, the apparent oral clearance of voriconazole after single oral dosing was 26%+/-16% (P > .05) lower in CYP2C19*1/*2 individuals and 66%+/-14% (P < .01) lower in CYP2C19 PMs. The addition of ritonavir caused a major reduction in voriconazole apparent oral clearance (354+/-173 mL/min versus 202+/-139 mL/min, P = .0001). This reduction occurred in all CYP2C19 genotypes (463+/-168 mL/min versus 305+/-112 mL/min [P = .023] for *1/*1, 343+/-127 mL/min versus 190+/-93 mL/min [P = .008] for *1/*2, and 158+/-54 mL/min versus 22+/-11 mL/min for *2/*2) and is probably caused by inhibition of CYP3A4-mediated voriconazole metabolism. CONCLUSIONS: Coadministration of a potent CYP3A4 inhibitor leads to a higher and prolonged exposure with voriconazole that might increase the risk of the development of adverse drug reactions on a short-term basis, particularly in CYP2C19 PM patients.     

Clinical consequences of cytochrome P450 2C9 polymorphisms

The gene coding for the cytochrome P450 (CYP) enzyme 2C9 (CYP2C9) carries numerous inherited polymorphisms. Those coding for R144C (*2) and I359L (*3) amino acid substitutions have both significant functional effects and appreciable high population frequencies, and their in vivo consequences have been studied in humans with regard to drug metabolism. This review summarizes present knowledge about the pharmacokinetics, drug responses, and outcomes of clinical studies in individuals with different CYP2C9 genotypes. Tentative estimates of how CYP2C9 genotyping might be applied to dose adjustments in clinical therapy were based on dose-related pharmacokinetic parameters such as clearance or trough drug concentrations. Mean clearances in homozygous carriers of the *3 allele were below 25% of that of the wild type for S -warfarin, tolbutamide, glipizide, celecoxib, and fluvastatin. In the more frequent heterozygous carriers (genotype *1/*3), the clearances were between 40% and 75%. In these cases in which individual dosages are derived from clinical drug effects, such as for the oral anticoagulants, the pharmacogenetics-based dose adjustments showed a good correlation with the genotype-specific empirically derived doses. In addition to its role in pharmacokinetics, CYP2C9 contributes to the metabolism of fatty acids, prostanoids, and steroid hormones, and it may catalyze potentially toxic bioactivation reactions. However, our current understanding of the role of CYP2C9 in biotransformation of endogenous signaling molecules and in drug toxicity is relatively meager.     

Acenocoumarol pharmacokinetics in relation to cytochrome P450 2C9 genotype

BACKGROUND AND OBJECTIVES: Cytochrome P450 (CYP) 2C9 is one of the major CYP enzymes involved in the biotransformation of drugs, among others, the oral anticoagulant acenocoumarol. The enzyme has several polymorphisms, with the CYP2C9*2 and CYP2C9*3 variants most commonly present in white patients. Patients with the CYP2C9*3 variant are known to require a lower maintenance dose of racemic acenocoumarol. We investigated the impact of the polymorphisms CYP2C9*2 and CYP2C9*3 on the pharmacokinetics of R- and S-acenocoumarol. METHODS AND RESULTS: In the first study 26 healthy volunteers with the genotype *1/*1 (n = 9), *1/*2 (n = 7), *1/*3 (n = 6), *2/*3 (n = 3), and *2/*2 (n = 1) were given 8 mg of racemic acenocoumarol as a single oral dose. Plasma R- and S-acenocoumarol concentrations were assayed at 4, 7, and 24 hours. Mean plasma S-acenocoumarol concentrations at 7 hours were higher in subjects with a variant allele; the differences were significant (P =.01) for the *1/*3 and *2/*3 genotypes. In the second study, the oral pharmacokinetics of acenocoumarol was investigated in 6 subjects (*1/*1 [n = 3] and *1/*3 [n = 3]). The mean oral clearance of S-acenocoumarol was 45% lower in the CYP2C9*1/*3 genotypes (10.9 +/- 3.0 L/h versus 19.8 +/- 3.1 L/h, P =.02). Plasma half-life was prolonged from 1.0 +/- 0.2 hours to 2.0 +/- 0.7 hours (P =.09). R-acenocoumarol pharmacokinetics did not differ between the genotypes. There was no difference in mean international normalized ratio at 24 hours, which was 1.2 in both groups. In vitro enzyme kinetics showed reduced (85%) intrinsic activity of the *3 enzyme to catalyze the hydroxylations of S-acenocoumarol. The lower activity resulted from higher Michaelis-Menten constant (2-fold) and lower maximum rate of metabolism by an enzyme-mediated reaction (by 70%). The activity of the *2 enzyme was 50% of the wild-type one. CONCLUSION: The results show S-acenocoumarol pharmacokinetics to be dependent on CYP2C9 polymorphism. In particular, the presence of the CYP2C9*3 allele impairs oral clearance of the coumarin.     

妊娠大鼠肾脏血流动力学变化与肾脏微血管细胞色素P450表达的关系

目的探讨肾脏微血管细胞色素P450表达与妊娠大鼠肾脏血流动力学变化的关系。方法比较妊娠晚期与非妊娠大鼠肾脏血流动力学指标差异，运用Western blot和HPLC比较妊娠各个时期与非妊娠大鼠肾脏微血管细胞色素P450表达与活性。结果与非妊娠大鼠相比，妊娠晚期大鼠平均动脉压和肾微血管阻力明显下降（P〈0．05），妊娠期CYP表达增加，产生EETs活性增加。结论妊娠期肾脏血流动力学变化可能与肾脏微血管细胞色素P450表达有关。     

建立检测细胞色素P450酶与紫杉醇代谢相关突变位点的基因芯片分型方法

目的 建立针对与临床抗癌药物紫杉醇代谢相关的细胞色素P450(CYP450)酶基因多态性位点的快速、准确、高通量的基因芯片基因分型方法.方法 选取CYP450酶2C8*3、3A4* 18、3A5*3C等3个与紫杉醇代谢相关突变位点,根据基因库中报道的序列,设计每个基因多态性位点的野生型和突变型探针,在突变位点2侧设计PCR扩增引物,PCR片段长度应＜200 bp,并构建标准质粒.探针在3'端氨基修饰,下游引物标记荧光素Cy3,将探针按一定顺序点样于经醛基化处理的玻片制备成基因芯片.人体血液DNA样本分别通过3对引物扩增后与基因芯片上的探针杂交,通过扫描图像和配套软件对结果进行分析和判断.同时,对50份血液样本进行检测.结果 通过标准质粒杂交结果显示,每个位点对应的1对探针均能将野生型和突变型质粒进行准确地区分,无非特异性杂交信号出现;检测50份血液样本,CYP2C8 * 3位点的突变率为2%,CYP3A4 * 18位点均为野生型,CYP3A5 * 3C位点的突变率为62%.同时,通过测序法进行验证,基因芯片方法与测序方法的结果完全一致.结论 建立的同时检测抗癌药物紫杉醇代谢相关的CYP450酶基因多态性位点CYP2C8*3、CYP3A4 * 18、CYP3A5 * 3C的基因芯片分型方法快速、准确,结果可靠,重复性好,可用于指导紫杉醇患者的个体化用药,并为分析个体患者对于紫杉醇药物体内代谢提供了一个高通量技术平台.     

Effects of cytochrome P450 2C9 polymorphisms on phenprocoumon anticoagulation status

OBJECTIVE: Our objective was to assess whether there is an association between the presence of allelic variants of the gene for cytochrome P450 (CYP) 2C9 and anticoagulation problems during the initial phase of phenprocoumon treatment. METHODS: A prospective follow-up study was performed at 2 anticoagulation clinics in The Netherlands. Included subjects started phenprocoumon during the study period, had their first check of the international normalized ratio (INR) on the third or fourth day of therapy, and had an indication for the low therapeutic range (INR, 2.0-3.5). CYP2C9 genotypes ( CYP2C9*1 , CYP2C9*2 , and CYP2C9*3 ) were assessed, and data on indication, INR checks, comedication, and comorbidity were collected. RESULTS: After genotyping, 284 subjects were available for analysis. Of these, 186 (65.5%) were homozygous carriers of the CYP2C9 wild-type allele ( CYP2C9*1/*1 ), 61 (21.5%) were carriers of the CYP2C9*2 allele, and 37 (13.0%) were carriers of the CYP2C9*3 allele. Compared with homozygous CYP2C9*1/*1 subjects, carriers of CYP2C9*2 or *3 had an increased risk of severe overanticoagulation (INR >6.0). The hazard ratio for CYP2C9*2 versus CYP2C9*1/*1 was 3.09 (95% confidence interval [CI], 1.56 to 6.13; P=.001), and the hazard ratio for CYP2C9*3 versus CYP2C9*1/*1 was 2.40 (95% CI, 1.03 to 5.57; P=.042). Carriers of CYP2C9*2 also had a lower chance to achieve stability in the follow-up period. The hazard ratio for CYP2C9*2 versus CYP2C9*1/*1 was 0.61 (95% CI, 0.43 to 0.85; P=.003). Carriers of the CYP2C9*2 or *3 allele needed a significantly lower phenprocoumon dosage compared with homozygous CYP2C9*1/*1 subjects. CONCLUSION: The presence of at least 1 CYP2C9*2 or *3 allele in phenprocoumon users is associated with an increased risk of severe overanticoagulation. Similar to warfarin and acenocoumarol, phenprocoumon had a lower dosage requirement in carriers of CYP2C9*2 or *3 compared with that in CYP2C9 wild-type subjects.     

Initial 48-hour acid inhibition by intravenous infusion of omeprazole, famotidine, or both in relation to cytochrome P450 2C19 genotype status

BACKGROUNDS AND AIMS: Faster and stronger acid inhibition is required for the treatment of hemorrhage from peptic ulcers. We compared the effects of intravenous infusion regimens of a proton pump inhibitor (PPI) alone, a histamine 2 receptor antagonist (H2RA) alone, and the combination of a PPI with an H2RA on acid inhibition in the early postadministration phase in relation to cytochrome P450 (CYP) 2C19 genotype status. METHODS: Fifteen Helicobacter pylori-positive volunteers with 3 different CYP2C19 genotypes were administered twice-daily intravenous infusions of 20 mg famotidine alone, 20 mg omeprazole alone, 10 mg omeprazole plus 10 mg famotidine (half-concomitant regimen), and 20 mg omeprazole plus 20 mg famotidine (full-concomitant regimen) for 2 days. The subjects underwent 48-hour intragastric pH monitoring. RESULTS: In homozygous extensive metabolizers (EMs) the median first 24-hour intragastric pH with the full-concomitant regimen was 4.8 (range, 4.5-5.4), which was significantly higher than that with omeprazole alone (3.9 [range, 2.6-4.7], P=.043) or famotidine alone (4.4 [range, 3.8-4.9], P=.043). In heterozygous EMs and poor metabolizers the respective median first 24-hour pH values attained with omeprazole alone (5.8 [range, 4.3-6.3] and 6.1 [range, 5.3-7.4]) and the full-concomitant regimen (5.8 [range, 5.1-6.4] and 5.8 [range, 5.4-6.2]) were significantly higher than those attained with famotidine alone (4.1 [range, 3.9-6.5] and 4.7 [range, 3.7-5.7], P=.043 for all). CONCLUSIONS: For faster and stronger acid inhibition, the concomitant infusion regimen of a PPI and an H2RA appears to be therapeutically useful in homozygous and heterozygous EMs, but omeprazole alone appears to be sufficient in poor metabolizers of CYP2C19.     

Influence of cytochrome P450 polymorphisms on drug therapies: pharmacogenetic, pharmacoepigenetic and clinical aspects

The polymorphic nature of the cytochrome P450 (CYP) genes affects individual drug response and adverse reactions to a great extent. This variation includes copy number variants (CNV), missense mutations, insertions and deletions, and mutations affecting gene expression and activity of mainly CYP2A6, CYP2B6, CYP2C9, CYP2C19 and CYP2D6, which have been extensively studied and well characterized. CYP1A2 and CYP3A4 expression varies significantly, and the cause has been suggested to be mainly of genetic origin but the exact molecular basis remains unknown. We present a review of the major polymorphic CYP alleles and conclude that this variability is of greatest importance for treatment with several antidepressants, antipsychotics, antiulcer drugs, anti-HIV drugs, anticoagulants, antidiabetics and the anticancer drug tamoxifen. We also present tables illustrating the relative importance of specific common CYP alleles for the extent of enzyme functionality. The field of pharmacoepigenetics has just opened, and we present recent examples wherein gene methylation influences the expression of CYP. In addition microRNA (miRNA) regulation of P450 has been described. Furthermore, this review updates the field with respect to regulatory initiatives and experience of predictive pharmacogenetic investigations in the clinics. It is concluded that the pharmacogenetic knowledge regarding CYP polymorphism now developed to a stage where it can be implemented in drug development and in clinical routine for specific drug treatments, thereby improving the drug response and reducing costs for drug treatment.     

Genetic polymorphisms of cytochrome P450 among patients with Balkan endemic nephropathy (BEN)

OBJECTIVES: The concept of multifactorial etiology of BEN anticipates that a combination of polymorphic gene variants and various environmental factors causes an increased risk for the disease. CYP enzymes play a key role in the metabolic activation of environmental chemicals and toxins. CYP3A enzymes are particularly relevant for xenobiotic metabolism because of their broad substrate specificity and abundant expression in the human liver, intestine, and kidney. Previous phenotyping analysis on CYP2D6 enzyme activity in BEN patients proposed a modifying effect of CYP2D6 gene variants on BEN risk, but it was not approved with molecular-genetic methods. The aim of the current case-control study was to compare the frequency of CYP2D6 and CYP3A5 polymorphisms, as well as one CYP3A4 promoter variant in BEN patients and controls in order to investigate a possible association between individual genetic variations in these genes and susceptibility to BEN. DESIGN AND METHODS: Ninety-six nonrelated Bulgarian BEN patients from endemic villages in the Vratza district and 112 healthy Bulgarians from nonendemic areas (controls) were genotyped. Identification of alleles was done by allele-specific PCR or by rapid-cycle amplification on the LightCycler, followed by sequence-specific detection. RESULTS: The UM, PM, and EM + IM genotype frequencies of CYP2D6 did not differ significantly between the two groups (P > 0.05). The CYP3A4*1B allele was only found in the heterozygous form, with allelic frequencies of 5.21% in the patients and 2.23% in the healthy individuals (P = 0.11). The CYP3A5*1 allele was more prevalent in BEN patients with a frequency of 9.38% compared to 5.36% in the controls and was associated with a higher risk for BEN (OR 2.41, 95% CI 1.09-5.33) (P = 0.02). CONCLUSIONS: Our results demonstrate that the CYP3A5*1 allele, previously reported as a marker for CYP3A5 expression in human kidney, is associated with increased risk for BEN, while CYP3A4*1B and CYP2D6 genotypes do not significantly modify the risk for the disease.     

Inhibition of cytochrome P450 2A6 increases nicotine's oral bioavailability and decreases smoking

BACKGROUND: Nicotine establishes and maintains tobacco dependence. Individuals with genetically deficient CYP2A6 nicotine metabolism are at lower risk to become smokers and, if dependent, will smoke fewer cigarettes. Hepatic CYP2A6 accounts for nicotine's low systemic bioavailability, precluding oral nicotine replacement to treat dependence. OBJECTIVE: We sought to determine whether CYP2A6 inhibition via oral methoxsalen decreases nicotine clearance, increases nicotine bioavailability, and decreases smoking. METHODS: Two within-subject designs in healthy tobacco-dependent volunteers were conducted: a singleblind kinetic study (n = 17) of methoxsalen 30, 10, or 3.5 mg or placebo given with nicotine 4 mg orally to abstinent smokers; and a double-blind randomized crossover study (n = 11) of methoxsalen 30 mg or placebo crossed with nicotine 4 mg given orally or placebo before 60 minutes' abstinence and 90 minutes' free smoking. RESULTS: Placebo plus nicotine 4 mg orally increased the mean 3-hour plasma nicotine level by 4 ng/mL over residual baseline nicotine level, whereas methoxsalen 10 or 30 mg plus nicotine increased it by 9 ng/mL (P<.01), demonstrating in vivo inhibition of CYP2A6 nicotine metabolism. Methoxsalen 30 mg plus nicotine 4 mg given orally decreased breath carbon monoxide concentration at the end of free smoking by 47% (4.6 versus 8.7 ppm; P<.01) and cigarettes smoked by 24% (3.1 versus 4.1, P<.01) compared with placebo plus placebo. CONCLUSIONS: Methoxsalen inhibits nicotine first-pass metabolism of orally administered nicotine, and the combination directly reduces smoking in a laboratory setting. CYP2A6 inhibitors may have an important role in smoking cessation and tobacco exposure reduction.     

Organization of multiple cytochrome P450s with NADPH-cytochrome P450 reductase in membranes

Microsomal P450-mediated monooxygenase activity supported by NADPH requires an interaction between flavoprotein NADPH-cytochrome P450 reductase and cytochrome P450. These proteins have been identified as the simplest system (with the inclusion of a phospholipid (PL) component) that possesses monooxygenase function; however, little is known about the organization of these proteins in the microsomal membrane. Although reductase and P450 are known to form a 1:1 functional complex, there exists a 10- to 20-fold excess of P450 over the reductase. This raises several questions including "How are the enzymes of the P450 system organized in the microsomal membrane?" and "Can one P450 enzyme affect the functional characteristics of another P450?" This review summarizes evidence supporting the potential for enzymes involved in the P450 system to interact, focusing on the interactions between reductase and P450 and interactions between multiple P450 enzymes. Studies on the aggregation characteristics of P450 as well as on rotational diffusion are detailed, with a special emphasis on the potential for P450 enzymes to produce oligomeric complexes and to suggest the environment in which P450 exists in the endoplasmic reticulum. Finally, more recent studies describing the potential for multiple P450s to exist as complexes and their effect on P450 function are presented, including studies using reconstituted systems as well as systems where two P450s are coexpressed in the presence of reductase. An understanding of the interactions among reductase and multiple P450s is important for predicting conditions where the drug disposition may be altered by the direct effects of P450-P450 complex formation. Furthermore, the potential for one P450 enzyme to affect the behavior of another P450 may be extremely important for drug screening and development, requiring metabolic screening of a drug with reconstituted systems containing multiple P450s rather than simpler systems containing only a single form.     

芳香化酶细胞色素P450在子宫内膜异位症的表达

目的探讨芳香化酶细胞色素P450(以下简称芳香化酶)在子宫内膜异位症(EMs)发病机制中的作用。方法采用免疫组化方法检测36例EMs患者在位及异位子宫内膜组织芳香化酶的表达,并与22例正常子宫内膜进行比较。结果芳香化酶在EMs组异位内膜及在位内膜的表达明显高于对照组(P<0.05),增殖期与分泌期无明显差异(P>0.05),且与修订的美国生育协会标准(rAFS)分期无相关性(P>0.05)。结论芳香化酶在异位内膜及在位内膜的的表达可能在EMs的发病中起重要作用。芳香化酶表达与EMs的严重程度无关。     

阿尔茨海默病患者细胞色素P450 1A1基因与载脂蛋白E基因多态性的关联分析

目的：探讨细胞色素P450 1A1（CP1A1）和载脂蛋白E基因多态性及其交互作用与阿尔茨海默病的关系。 方法：实验于2004—11／2005—05在南方医科大学珠江医院中心实验室进行。应用聚合酶链反应-限制性片段长度多态性方法检测了135例散发性阿尔茨海默病患者和138例正常老年人细胞色素P450 1A1基因和载脂蛋白E基因多态性分布特征。 结果：阿尔茨海默病组135例，健康对照组138例，均进入结果分析。①细胞色素P450 1A1等位基因m1，m2频率：散发性阿尔茨海默病组分别为63％，37％，对照组分别为65．1％，31．9％，各等位基因频率分布差异无显著性（r=1．605，P〉0．05）。②些散发性阿尔茨海默病组载脂蛋白Eε4等位基因频率与对照组比较有统计学意义（10．74％，5．80％，χ^2=4．41，P〈0．05，OR=1．95），散发性阿尔茨海默病组和对照组各基因型分布差异无统计学意义（P〉0．05）。③研究对象按携带载脂蛋白E84状况分层后，在带有ε4等位基因的人群中，散发性阿尔茨海默病组细胞色素P450 1A1m2等位基因频率与对照组比较差异有显著性意义（7．4％，2．2％，r=6．825，P=0．009），患病风险增加3．417倍（OR=3．417）。 结论：细胞色素P450 1A1基因MSP1多态性与中国汉族人散发性阿尔茨海默病发病无明显关联。但在携带有载脂蛋白讲基因型的个体，细胞色素P450 1A1m2等位基因可能是散发性阿尔茨海默病的危险基因。     

Drug-associated disease: cytochrome P450 interactions

Critically ill patients generally are older, frequently have organ failure, and commonly receive multiple medications, all of which make them susceptible to adverse effects of drugs. Drug interactions are a common adverse effect, and many are predictable based on understanding the mechanisms that underlie drug interactions. This article identifies commonly used medications in critically ill patients and the associated drug interactions that may occur with emphasis on the cytochrome P450 enzyme system.     

Effect of voriconazole on the pharmacokinetics and pharmacodynamics of intravenous and oral midazolam

OBJECTIVE: Our objective was to assess the effect of the antimycotic voriconazole on the pharmacokinetics and pharmacodynamics of oral and intravenous midazolam. METHODS: We used a randomized, crossover study design. Ten healthy male volunteers were given either no pretreatment (control phase) or voriconazole (voriconazole phase) orally, 400 mg twice daily on the first day and 200 mg twice daily on the second day. Midazolam was given, either 0.05 mg/kg intravenously or 7.5 mg orally, 1 hour after the last dose of voriconazole and during the control phase. Plasma concentrations of midazolam, alpha-hydroxymidazolam, and voriconazole were determined for 24 hours and pharmacodynamic variables measured for 12 hours. RESULTS: Voriconazole reduced the clearance of intravenous midazolam by 72% (P < .001) and increased its elimination half-life from 2.8 to 8.3 hours (P < .001). Voriconazole increased the peak concentration and the area under the plasma concentration-time curve of oral midazolam by 3.8- and 10.3-fold, respectively (P < .001). The bioavailability of oral midazolam was increased from 31% to 84% (P < .001). Voriconazole profoundly increased the psychomotor effects of oral midazolam (P < .001) but only weakly increased the effects of intravenous midazolam. CONCLUSION: When midazolam is given as small intravenous bolus doses, its effect is not increased to a clinically significant degree by voriconazole. The use of large midazolam doses increases the risk of clinically significant interactions also after its intravenous administration. The use of oral midazolam with voriconazole should be avoided, or substantially lower doses should be used.