Disposition of debrisoquine and nortriptyline in Korean subjects in relation to CYP2D6 genotypes, and comparison with Caucasians.

AIMS: To study the influence of the CYP2D6*10 allele on the disposition of debrisoquine and nortriptyline. METHODS: The pharmacokinetics of debrisoquine and nortriptyline and their main metabolites were determined in ten Koreans with the CYP2D6*1/*1 (n = 5) and CYP2D6*1/*10 (n = 5) genotypes after single oral doses of 20 mg debrisoquine and 25 mg nortriptyline, respectively. The data were compared with previously published findings from 21 Caucasians with 0, one, two, three, four or 13 functional CYP2D6 genes. RESULTS: The AUC0-8 of 4-hydroxydebrisoquine was significantly lower in Koreans with CYP2D6*1/*10 genotype compared with CYP2D6*1/*1[95% confidence interval (CI) for the ratio between means 1.17, 1.85]. No other genotype-related differences were found in the plasma kinetics of nortriptyline and debrisoquine, or their hydroxy metabolites. The AUCnortriptyline/AUC10-hydroxynortriptyline ratio did not differ between the *1/*1 and *1/*10 genotype groups (95% CI for the ratio of means 0.60, 1.26). Similarly, there was no difference between these genotypes with respect to the AUCdebrisoquine/AUC4-hydroxydebrisoquine ratio (95% CI for the ratio of mean values 0.38, 1.46). Both Korean genotype groups had similar AUCs and parent compound/metabolite AUC ratios of debrisoquine and nortriptyline to Caucasians with two functional CYP2D6 genes. CONCLUSIONS: Heterozygosity for CYP2D6*10 decreases the CYP2D6-dependent elimination of nortriptyline and debrisoquine to only a limited degree. Further studies in subjects homozygous for CYP2D6*10 are required to elucidate fully the pharmacokinetic consequences of this CYP2D6 genotype in Orientals.

Characterization of testosterone metabolism and 7-hydroxycoumarin conjugation by rat and human liver slices after storage in liquid nitrogen for 1 h up to 6 months.

1. Slices of human and rat liver were cryopreserved in 18% dimethyl sulphoxide (DMSO) and subsequently stored in liquid nitrogen for periods up to as long as 6 months. After thawing, the metabolism of testosterone to hydroxylated products and conjugation of 7-hydroxycoumarin were investigated. 2. Rat liver slices stored in liquid nitrogen for 6 months exhibited rates of formation of 7alpha-, 6beta- 16alpha- and 2alpha-hydroxytestosterone, and of androstenedione that did not differ significantly from those observed with fresh slices. 3. No formation of 2alpha-hydroxytestosterone was detected with slices of human liver. However, in contrast with the rat, human slices produced 2beta-hydroxytestosterone. The rates of formation of 7alpha-, 6beta-, 16alpha- and 2beta-hydroxytestosterone and of androstenedione by human liver slices after 6 months of storage in liquid nitrogen were 82, 71, 236, 66 and 92%, respectively, of the corresponding rates by fresh slices. 4. The rates of sulphation and glucuronidation of 7-hydroxycoumarin by slices from rat liver were 97 and 119%, respectively, of the corresponding fresh values after 6 months of storage in liquid nitrogen. 5. 7-Hydroxycoumarin glucuronidation by human liver slices was 53% of the corresponding fresh values after 6 months of storage. However, human slices showed little or no capacity to conjugate 7-hydroxycoumarin with sulphate. 6. It was demonstrated that slices of both human and rat liver can be cryopreserved and stored in liquid nitrogen for at least 6 months without major changes in their rates of metabolism of testosterone to its hydroxylated products and of 7-hydroxycoumarin conjugation. These findings further emphasize that cryopreservation of liver slices can be an effective tool in the use of biological material of limited availability.

Does hepatic metabolism of melatonin affect the endogenous serum melatonin level in man?

Melatonin (MT) is metabolized in the liver by cytochrome P450 (CYP) 1A2 but its importance for the metabolic process has not been fully elucidated. Therefore, the objective of this investigation was to study whether patients with different CYP1A2 activity would have different nocturnal serum MT levels. For that purpose serum MT concentrations were determined every second hour during the night in 12 healthy subjects and their MT areas under the curve (MT-AUCs) were calculated. Caffeine (CA) clearance was determined in advance. It is generally accepted that CA clearance reflects CYP1A2 activity. This made it possible to evaluate whether a relationship prevails between endogenous MT-AUCs and CYP1A2 activity. If CYP1A2 is of importance for the metabolism of MT one would expect to find an inverse correlation between the MT-AUCs and the CA clearance. However, such correlation did not exist in the current study (Rs=-0.021, NS). Since endogenous MT-AUC is dependent not only on MT elimination by CYP1A2 but also on MT secretion, it is possible that an increased MT secretion counter-balances an increased hepatic MT metabolism. If so, this could explain why the MT-AUCs and the CA clearance values were not inversely correlated in this study.

Inhibition of cytochrome P4502D6 activity with paroxetine normalizes the ultrarapid metabolizer phenotype as measured by nortriptyline pharmacokinetics and the debrisoquin test.

BACKGROUND: The ultrarapid metabolizer phenotype of the cytochrome P4502D6 (CYP2D6) enzyme has been considered a relevant cause of nonresponse to antidepressant drug therapy. Prescribing high doses of antidepressants to such patients leads to high concentrations of potentially toxic metabolites and an increased risk for adverse reactions. Normalization of the metabolic status of ultrarapid metabolizers by inhibition of CYP2D6 activity could offer a clinically acceptable method to successfully treat such patients with antidepressants. METHODS: Five ultrarapid metabolizers with a CYP2D6 gene duplication or triplication were treated with 25 mg nortriptyline twice a day for 3 consecutive weeks, alone during the first week and concomitantly with the CYP2D6 inhibitor paroxetine 10 mg or 20 mg twice a day, respectively, during the second and third weeks. After the third week, nortriptyline was discontinued and the subjects were treated with paroxetine 20 mg twice a day during the fourth study week. At the end of each study week, the steady-state pharmacokinetic parameters of nortriptyline or paroxetine were determined within the dose interval. In addition, the CYP2D6 phenotype was determined by debrisoquin (INN, debrisoquine) test at baseline and at the end of each study phase. Treatment-related adverse events were recorded during drug administration and for 1 week thereafter. RESULTS: All 5 subjects had very low (subtherapeutic) nortriptyline concentrations after 7 days' treatment with nortriptyline only. Addition of paroxetine 10 mg twice a day to the nortriptyline regimen resulted in a change in all individuals to the "normal" extensive debrisoquine metabolizer phenotype, and therapeutic plasma nortriptyline concentrations were achieved in 4 of 5 subjects after a 3 times mean increase in nortriptyline trough concentration (P =.0011). Doubling the paroxetine dose caused a 15 times mean increase in paroxetine trough concentration (P <.001), indicating strong inhibition by paroxetine of its own metabolism. The high paroxetine concentrations in 2 subjects caused them to have the poor debrisoquine metabolizer phenotype and resulted in a further increase in plasma nortriptyline trough concentration (P =.0099). A strong correlation (rank correlation coefficient [r(s)] = 0.89; P <.0001) was observed between paroxetine and nortriptyline trough concentrations. Paroxetine also significantly decreased the fluctuation of nortriptyline concentrations within the dose interval. One subject discontinued the study after the second study week because of adverse effects; otherwise, the study drugs were well tolerated. CONCLUSIONS: Paroxetine, with a daily dosage from 20 to 40 mg, is an effective tool in normalizing the metabolic status of CYP2D6 ultrarapid metabolizers.

Different enantioselective 9-hydroxylation of risperidone by the two human CYP2D6 and CYP3A4 enzymes.

The antipsychotic agent risperidone, is metabolized by different cytochrome P-450 (CYP) enzymes, including CYP2D6, to the active 9-hydroxyrisperidone, which is the major metabolite in plasma. Two enantiomers, (+)- and (-)-9-hydroxyrisperidone might be formed, and the aim of this study was to evaluate the importance of CYP2D6 and CYP3A4/CYP3A5 in the formation of these two enantiomers in human liver microsomes and in recombinantly expressed enzymes. The enantiomers of 9-hydroxyrisperidone were analyzed with high pressure liquid chromatography using a chiral alpha-1 acid glycoprotein column. A much higher formation rate was observed for (+)-9-hydroxyrisperidone than for (-)-9-hydroxyrisperidone in microsomes prepared from six individual livers. The formation of (+)-9-hydroxyrisperidone was strongly inhibited by quinidine, a potent CYP2D6 inhibitor, whereas ketoconazole, a CYP3A4 inhibitor, strongly inhibited the formation of (-)-9-hydroxyrisperidone. Recombinant human CYP2D6 produced only (+)-9-hydroxyrisperidone, whereas a lower formation rate of both enantiomers was detected with expressed CYP3A4 and CYP3A5. In vivo data from 18 patients during treatment with risperidone indicate that the plasma concentration of the (+)-enantiomer is higher than that of the (-)-enantiomer in extensive metabolizers of CYP2D6. These findings clearly suggest that CYP2D6 plays a predominant role in (+)-9-hydroxylation of risperidone, the major metabolic pathway in clinical conditions, whereas CYP3A catalyzes the formation of the (-)-9-hydroxymetabolite. Further studies are required to evaluate the pharmacological/toxic activity of both enantiomers.

Orally given melatonin may serve as a probe drug for cytochrome P450 1A2 activity in vivo: a pilot study.

BACKGROUND: Melatonin is a hormone that is metabolized by cytochrome P450 (CYP) 1A2 to its main primary metabolite 6-hydroxymelatonin. We therefore evaluated the utility of oral melatonin as a marker of hepatic CYP1A2 activity. METHODS: Twenty-five milligrams of melatonin was given at 9:30 am to 12 healthy Swedish volunteers, who had previously been phenotyped for CYP1A2 with caffeine. Melatonin and conjugated 6-hydroxymelatonin were analyzed by liquid chromatography-mass spectrometry in blood samples taken between 0.5 and 6.5 hours after drug intake. Serum concentrations of melatonin and conjugated 6-hydroxymelatonin, or their ratio at different time points, and the apparent melatonin clearance were tested for correlation with caffeine clearance. RESULTS: We found a significant correlation between apparent clearance of melatonin and caffeine clearance with a Spearman rank correlation coefficient (Rs) of 0.75 (P =.005). The melatonin concentration 1.5 hours after administration also closely correlated with the caffeine clearance (Rs = -0.62; P =.03). Inclusion of conjugated 6-hydroxymelatonin gave no closer correlations. CONCLUSION: Melatonin might be developed as an alternative to caffeine as a probe drug for CYP1A2 phenotyping.

Role of CYP2C9 polymorphism in losartan oxidation.

Losartan, an angiotensin II receptor antagonist, is oxidized by hepatic cytochromes P450 to an active carboxylic acid metabolite, E-3174. The aim of the present investigation was to study the contribution of CYP2C9 and CYP3A4 in losartan oxidation in vitro and to evaluate the role of CYP2C9 polymorphism. Kinetic properties of different genetic CYP2C9 variants were compared both in a yeast expression system and in 25 different samples of human liver microsomes where all known genotypes of CYP2C9 were represented. Microsomes were incubated with losartan (0.05-50 microM), and the formation of E-3174 was analyzed by high-performance liquid chromatography to estimate V(max), K(m), and intrinsic clearance for all individual samples. Sulfaphenazole, a CYP2C9 inhibitor, blocked the formation of E-3174 at low losartan concentrations (<1 microM), whereas the inhibitory effect of triacetyloleandomycin, a CYP3A4 inhibitor, was significant only at high concentrations of losartan (>25 microM). In comparison to the CYP2C9.1 variant, oxidation of losartan was significantly reduced in yeast expressing the rare CYP2C9.2 or CYP2C9.3 variants. Moreover, the rate of losartan oxidation was lower in liver microsomes from individuals hetero- or homozygous for the CYP2C9*3 allele, or homozygous for the CYP2C9*2 allele. The difference between the common and rare CYP2C9 variants was mainly explained by a lower V(max), both in yeast and human liver microsomes. In summary, these in vitro results indicate that CYP2C9 is the major human P450 isoenzyme responsible for losartan oxidation and that the CYP2C9 genotype contributes to interindividual differences in losartan oxidation and activation.

Determination of exogenous melatonin and its 6-hydroxy metabolite in human plasma by liquid chromatography-mass spectrometry.

SUMMARY: Melatonin has recently garnered interest as a possible treatment for sleep disorders, and this has created a desire for appropriate pharmacokinetic studies. No method has yet been published that can measure the concentrations of both melatonin and its main metabolite, 6-hydroxymelatonin, in plasma or serum. Therefore, a liquid chromatography-mass spectrometry (LC/MS) method including enzymatic hydrolysis and one-step liquid-liquid extraction was developed for this purpose. The mean extraction recovery was 59% for melatonin and 42% for 6-hydroxymelatonin. The mean precision of the method as calculated from the interassay coefficient of variation of quality control samples at high and low concentrations was 18% for 6-hydroxymelatonin and 15% for melatonin. The inaccuracy was always less than 2%. The limit of detection was approximately 2 ng/mL for 6-hydroxymelatonin (signal/noise ratio = 4) and less than 2 ng/mL for melatonin (signal/noise ratio at 2 ng/mL = 10). The method was applied to the analysis of plasma from a healthy volunteer who had received a single oral dose of 25 mg melatonin.

Cytochrome P450 isoforms involved in melatonin metabolism in human liver microsomes.

OBJECTIVE: The present study was carried out to identify the cytochrome P450 enzyme(s) involved in the 6-hydroxylation and O-demethylation of melatonin. METHODS: The formation kinetics of 6-hydroxymelatonin and N-acetylserotonin were determined using human liver microsomes and cDNA yeast-expressed human enzymes (CYP1A2, 2C9 and 2C19) over the substrate concentration range 1-1000 microM. Selective inhibitors and substrates of various cytochrome P450 enzymes were also employed. RESULTS: Fluvoxamine was a potent inhibitor of 6-hydroxymelatonin formation, giving 50 +/- 5% and 69 +/- 9% inhibition at concentrations of 1 microM and 10 microM, respectively, after incubation with 50 microM melatonin. Furafylline, sulphaphenazole and omeprazole used at low and high concentrations substantially inhibited both metabolic pathways. cDNA yeast-expressed CYP1A2, CYP2C9 and CYP2C19 catalysed the formation of the two metabolites, confirming the data obtained with specific inhibitors and substrates. CONCLUSIONS: Our results strongly suggest that 6-hydroxylation, the main metabolic pathway of melatonin, is mediated mainly, but not exclusively, by CYP1A2, the high-affinity enzyme involved in melatonin metabolism, confirming the observation that a single oral dose of fluvoxamine increases nocturnal serum melatonin levels in healthy subjects. Furthermore, the results indicate that there is a potential for interaction with drugs metabolised by CYP1A2 both at physiological levels and after oral administration of melatonin, while CYP2C19 and CYP2C9 are assumed to be less important.

The role of CYP2C9 genotype in the metabolism of diclofenac in vivo and in vitro.

INTRODUCTION: The polymorphic cytochrome P450 enzyme 2C9 (CYP2C9) catalyses the metabolism of many drugs including S-warfarin, acenocoumarol, phenytoin, tolbutamide, losartan and most of the non-steroidal anti-inflammatory drugs. Diclofenac is metabolised to 4'-hydroxy (OH), the major diclofenac metabolite, 3'-OH and 3'-OH-4'-methoxy metabolites by CYP2C9. The aim of the present study was to clarify the impact of the CYP2C9 polymorphism on the metabolism of diclofenac both in vivo and in vitro. SUBJECTS, MATERIALS AND METHODS: Twenty healthy volunteers with different CYP2C9 genotypes [i.e. CYP2C9*1/ *1 (n = 6), *1/*2 (n = 3), *1,/*3 (n = 5), *2/*3 (n = 4), *21*2 (n = 1), *31*3 (n = 1)] received a single 50-mg oral dose of diclofenac. Plasma pharmacokinetics [peak plasma concentration (Cmax), half-life (t 1/2) and area under the plasma concentration-time curve (AUCtotal)] and urinary recovery of diclofenac and its metabolites were compared between the genotypes. Diclofenac 4'-hydroxylation was also analysed in vitro in 16 different samples of genotyped [i.e. CYP2C9*1/*1 (n = 7), *1/*2 (n=2), *1/*3 (n = 2), *2/*3 (n = 2), *2/*2 (n = 2), *31/*3 (n = 1)] human liver microsomes. RESULTS: Within each genotype group, a high variability was observed in kinetic parameters for diclofenac and 4'-OH-diclofenac (6- and 20-fold, respectively). No significant differences were found between the different genotypes either in vivo or in human liver microsomes. No correlation was found between the plasma AUC ratio of diclofenac/4'-OH-diclofenac and that of losartan/ E-3174, previously determined in the same subjects. CONCLUSION: No relationship was found between the CYP2C9 genotype and the 4'-hydroxylation of diclofenac either in vivo or in vitro. This, together with the lack of correlation between losartan oxidation and diclofenac hydroxylation in vivo raises the question about the usefulness of diclofenac as a CYP2C9 probe.

Effects of omeprazole on intragastric pH and plasma gastrin are dependent on the CYP2C19 polymorphism.

BACKGROUND & AIMS: Omeprazole is metabolized by cytochrome P450 (CYP2C19). The activity of this enzyme is polymorphic, with incidences of poor metabolizers (PMs), heterozygous extensive metabolizers (EMs), and homozygous EMs in white populations of 3%, 30%, and 67%, respectively. The importance of the CYP2C19 polymorphism for the effects of omeprazole on intragastric pH and plasma gastrin concentrations has been investigated. METHODS: Twenty-five white patients were genotyped for CYP2C19 by allele-specific polymerase chain reaction amplification, and their Helicobacter pylori status was assessed by serology and with immunoblot analysis. Intragastric pH was monitored over 24 hours, and meal-stimulated plasma gastrin concentration was measured over 4 hours (AUC 4h) before (day 0) and during (day 8) treatment with 20 mg omeprazole once daily. RESULTS: Eleven patients were homozygous for the wild-type allele (wt/wt), 12 were heterozygous EMs (wt/mut), and 2 were PMs (mut/mut). Median (95% confidence interval) 24-hour intragastric pH in the heterozygous EM group was 5.5 (range, 5.1-5. 9) compared with 3.1 (range, 2.7-3.6) in homozygous EMs (P < 0.0001) at day 8. The percentage of time with intragastric pH > 4 at day 8 was significantly higher in the wt/mut than wt/wt group (72.4% vs. 37.1%; P < 0.0001). H. pylori status had less influence than CYP2C19 on intragastric acidity. Omeprazole treatment increased meal-stimulated plasma gastrin concentrations from day 0 to day 8 in the homozygous EMs and heterozygous EMs by 16% (NS) and 157% (P = 0. 002), respectively. In heterozygous EMs, the gastrin increase was more pronounced in the H. pylori-positive group (226%) than H. pylori-negative group (80%; P = 0.02). CONCLUSIONS: The effects of omeprazole on intragastric pH and plasma gastrin are dependent on the CYP2C19 polymorphism in patients with acid-related disorders.

No sex-related differences but significant inhibition by oral contraceptives of CYP2C19 activity as measured by the probe drugs mephenytoin and omeprazole in healthy Swedish white subjects.

BACKGROUND: Although it is known that the use of oral contraceptives inhibits oxidative drug metabolism, there is little information regarding their effect on CYP2C19 activity. Moreover, earlier reports suggest that there may be differences in CYP2C19 activity between men and women. OBJECTIVE: We sought to assess the effect of sex and intake of oral contraceptives on CYP2C19 activity as measured by the probe drugs mephenytoin and omeprazole. METHODS: To determine CYP2C19 activity in white Swedish subjects, 644 subjects previously phenotyped with mephenytoin and 175 subjects phenotyped with omeprazole were investigated. The 8-hour urinary mephenytoin S/R ratio after ingestion of 100 mg mephenytoin and the plasma concentration ratio of omeprazole/hydroxyomeprazole at 3 hours after ingestion of 20 mg omeprazole were used as measures of CYP2C19 activity. Differences in these ratios and in their frequency distributions were then examined among women with and without oral contraceptives and men. In addition, nearly all subjects in the omeprazole group had been genotyped with regard to the CYP2C19*2 (ml) allele. Subjects homozygous for the CYP2C19*2 allele were excluded from the study. RESULTS: The median mephenytoin S/R ratio was 2.5-fold higher in the subgroup of women taking oral contraceptives compared with either women not taking oral contraceptives (P < .001) or men (P < .001). Similarly, the mean omeprazole/hydroxyomeprazole ratio was twice as high in the oral contraceptive group compared with women not taking oral contraceptives (P < .001) or men (P < .001). However, no differences were evident between women not taking oral contraceptives and men in either the mephenytoin group (P = .48) or the omeprazole group (P = .77). The oral contraceptive-induced inhibitory effect on CYP2C19 activity was similar between the CYP2C19*1/*1 and *1/*2 genotypes, and they were independent of age. CONCLUSIONS: Intake of oral contraceptives significantly inhibits CYP2C19 activity, but there is no true sex-related difference in CYP2C19 activity in healthy, white, Swedish subjects.

Fluvoxamine but not citalopram increases serum melatonin in healthy subjects-- an indication that cytochrome P450 CYP1A2 and CYP2C19 hydroxylate melatonin.

OBJECTIVE: The nocturnal serum melatonin (MT) level increases after ingestion of fluvoxamine (FLU)-- a selective serotonin re-uptake inhibitor (SSRI) with antidepressive properties. The mechanism behind the MT increase is unknown. Citalopram (CIT) is another SSRI. It is not known whether CIT affects the serum MT level. It may well be that these two compounds affect serum MT levels differently, inasmuch as the ways they inhibit cytochrome P450 (CYP) enzymes in the liver differ markedly. FLU inhibits CYP1A2 potently, and to some extent also CYP2C19, whereas CIT is without such an effect. CYP enzymes are probably involved in the hepatic metabolism of MT. If FLU, but not CIT, inhibits liver enzymes involved in the metabolism of MT, different serum MT concentrations should probably ensue. The objective of this investigation was to test this hypothesis. METHODS: Seven healthy subjects participated in three different experiments, which were performed in random order 6-8 days apart. In experiment A, placebo was given, in experiment B 40 mg CIT and in experiment C 50 mg FLU. All doses were given orally at 1600 hours. Serum MT concentrations were determined at regular intervals between 1600 hours and noon next day (20 h). Plasma concentrations of CIT were measured repeatedly in experiment B, and plasma FLU concentrations in experiment C. MT areas under the curve representing the 20-h period (MT-AUC(0-20)) were compared in the three experiments, and differences were statistically evaluated. RESULTS: FLU augmented the MT-AUC(0-20) by a factor of 2.8 compared with the effect of placebo (P < 0.01), whereas CIT was without significant effect. More MT was excreted in the urine after ingestion of FLU than after placebo. In contrast, CIT did not influence the MT excretion. A clear relationship was found between serum levels of MT and plasma concentrations of FLU. CONCLUSION: The serum MT level increased markedly after ingestion of FLU but not after CIT. The exact mechanism behind this finding is unknown, but decreased hepatic metabolism of MT by either CYP1A2 or CYP2C19, or both, is probable. Although exogenous MT, causing high MT concentration in plasma, has sleep-promoting properties in man, it is at this stage unknown whether serum MT concentrations in the range found in this study have similar effects. This has to be given further attention in additional studies.

Stereospecific analysis of omeprazole supports artemisinin as a potent inducer of CYP2C19.

The purpose of the study was to determine the enantiomer pharmacokinetics of omeprazole and 5-hydroxy-omeprazole before and after administration of the antimalarial artemisinin to confirm artemisinin's ability to induce CYP2C19. Nine healthy male Vietnamese subjects were given a single 20 mg dose of omeprazole orally 1 week before (day - 7) artemisinin administration. Artemisinin was then given orally (500 mg) for 7 days (days 1-7). On days 1 and 7, a single 20 mg dose of omeprazole was coadministered with artemisinin. After a washout period of 6 days, a single 20 mg dose of omeprazole was again administered together with a single 500 mg of artemisinin (day 14). Stereoselective pharmacokinetics of omeprazole and 5-hydroxyomeprazole was determined on days of omeprazole administration. Seven days of artemisinin administration significantly decreased the AUC of both omeprazole enantiomers (day 7), compared with day 1 (P < 0.001). All values were normalized after the washout period. Artemisinin increased the AUC ratio of R-5-hydroxyomeprazole/R-omeprazole significantly (P < 0.01) on day 7. The AUC ratio of omeprazole sulphone/S-omeprazole did not differ between study days. Artemisinin decreased the AUC of S-omeprazole to the same extent as that of R-omeprazole in extensive CYP2C19 metabolizers. suggesting that artemisinin induces a different enzyme in addition to CYP2C19. These results support and strengthen earlier findings that artemisinin induces CYP2C19 as well as at least one enzyme other than CYP3A4.

Pharmacokinetics of nortriptyline and its 10-hydroxy metabolite in Chinese subjects of different CYP2D6 genotypes.

OBJECTIVES: To study the impact of the CYP2D6*10 allele on the disposition of nortriptyline in Chinese subjects. METHODS: A single dose of 25 mg nortriptyline was given orally to 15 healthy Chinese volunteers who were classified as extensive metabolizers after phenotyping with debrisoquin (INN, debrisoquine) and who were genotyped by allele-specific polymerase chain reaction. Five subjects were homozygous for CYP2D6*1, 5 subjects were homozygous for CYP2D6*10, and 5 subjects were heterozygous for these 2 alleles. Plasma concentrations of nortriptyline and its main metabolite 10-hydroxynortriptyline were measured by liquid chromatography-mass spectrometry, and the pharmacokinetics were studied during 168 hours after the dose. RESULTS: Subjects who were homozygous for CYP2D6*10 had significantly higher total areas under the plasma concentration-time curve (AUC), lower apparent oral clearances, and longer mean plasma half-life of nortriptyline than subjects in the CYP2D6*1/*1 and the heterozygous groups. For 10-hydroxynortriptyline, the AUC was lower and the plasma half-life was longer in subjects who were homozygous for CYP2D6*10 than in subjects in the other 2 groups. CONCLUSION: The CYP2D6*10 allele in Chinese subjects was associated with significantly higher plasma levels of nortriptyline compared with the CYP2D6*1 allele because of an impaired metabolism of nortriptyline to 10-hydroxynortriptyline, particularly in the subjects with the CYP2D6*10/*10 genotype. The results suggest that genotyping of CYP2D6 may be a useful tool in predicting the pharmacokinetics of nortriptyline.

Bantu Tanzanians have a decreased capacity to metabolize omeprazole and mephenytoin in relation to their CYP2C19 genotype.

OBJECTIVE: To investigate the CYP2C19 polymorphism in Tanzanians because this enzyme shows large interindividual differences in activity and metabolizes several drugs of importance in Africa, especially the antimalarial agent chloroguanide (INN, proguanil). METHODS: Two hundred fifty-one Tanzanian healthy volunteers were phenotyped with respect to CYP2C19 with use of a single oral dose of mephenytoin (n = 106), a single oral dose of omeprazole (n = 207), or both. Sixty-two were phenotyped with both probe drugs. The urinary 0- to 8-hour S/R-mephenytoin ratio and the plasma omeprazole metabolic ratio (MR) (omeprazole/hydroxyomeprazole) 3 hours after drug intake were determined. The genotype was determined by analysis for CYP2C19*1 (wt), CYP2C19*2 (m1), and CYP2C19*3 (m2). Ten subjects with high omeprazole MR were screened for new mutations in the CYP2C19 gene by searching for single-strand conformation polymorphisms (SSCP). RESULTS: Eight subjects were classified as mephenytoin poor metabolizers (7.5%). Only 5 of these were homozygous for mutated alleles. The S/R ratio was skewed to the right (lower CYP2C19 activity) compared with other ethnic groups studied previously. No new mutations were found with polymerase chain reaction (PCR)-SSCP. We found 30 volunteers (14.5%) with an MR > 7, which is the antimode found previously in white subjects and Asian subjects. Of the 251 volunteers genotyped, 3.2% were homozygous for mutated alleles and 66.1% were homozygous for the wild-type allele. The allele frequencies of CYP2C19*1, *2, and *3 were 81.5%, 17.9%, and 0.6%, respectively. The correlation between the S/R-mephenytoin ratio and the omeprazole MR was significant (Spearman r = 0.59; P < .01). CONCLUSION: Tanzanians have a decreased capacity to metabolize both omeprazole and mephenytoin when their genotype is compared with metabolic capacity and genotype in other previously studied populations. We identified a low frequency of the Asian allele (CYP2C19*3). Although we did not find any new mutations, our results may be consistent with the presence of yet-unidentified mutations of CYP2C19 that causes decreased CYP2C19 activity in the Tanzanian population.

High-performance liquid chromatography-mass spectrometry for the quantification of nortriptyline and 10-hydroxynortriptyline in plasma.

A highly sensitive and selective method for the quantification of nortriptyline and its major 10-hydroxy metabolite in plasma is described. The method is based on liquid-liquid extraction in combination with acid dehydration of the 10-hydroxy metabolite to the less polar 10,11-dehydronortriptyline. Deuterium labelled internal standards ([2H4]NT and [2H3]10-OH-NT) were used and the compounds were separated by reversed-phase HPLC and detected using atmospheric pressure chemical ionisation and mass spectrometry. The limit of quantification was 0.8 ng/ml for both compounds. A 1-ml volume of plasma was used for analysis in the concentration range 0.8-32 ng/ml. The within- and between-day coefficients of variation were 11% in the low, 1.6 ng/ml range, and 7% at 8 ng ml/ml. Using this method it was possible to quantify plasma concentrations for 168 h following a single oral dose of 25 mg of nortriptyline with good accuracy and precision.

CYP2C19 genotype and phenotype determined with omeprazole in patients with acid-related disorders with and without Helicobacter pylori infection.

BACKGROUND: Omeprazole is to a major extent metabolized by cytochrome P450 isozyme CYP2C19. The aims of this study were to compare the phenotype of CYP2C19 determined by omeprazole with the genotype and to determine the effect of Helicobacter pylori infection on the metabolism of omeprazole. METHODS: One-hundred and forty-three Caucasian patients with acid-related disorders assessed with a combination of gastrointestinal symptoms and upper endoscopic findings were given 20 mg omeprazole orally. Three hours after intake, omeprazole and 5-hydroxyomeprazole plasma concentrations were determined with high-performance liquid chromatography, and the phenotype for metabolic capacity was expressed as metabolic ratio (MR). Genotyping of defect alleles (CYP2C19*2 and *3) was performed by polymerase chain reaction amplification. One hundred eleven patients were tested after the first dose of omeprazole and 32 patients after repetitive administration (median time, 30 days). H. pylori serology was determined with enzyme-linked immunosorbent assay at the time of phenotyping. RESULTS: Genotypically, 2.8% had two mutated alleles and were poor metabolizers (PM), and 22.4% were heterozygous extensive metabolizers (EM). Among the 111 patients who received the first omeprazole dose, 4 patients had MR >5--that is, belonged to the PM phenotype. Two of these had PM genotype (both CYP2C19*2/*2), and two had an EM genotype (CYP2C19*11*1 and *1/*3), indicating that they have still unidentified mutations. In the heterozygous EM group the mean MR was higher in patients who had been on continued omeprazole treatment than in those given the first dose (5.7 versus 2.5, P = 0.02). There were no significant differences in MR and omeprazole concentrations between H. pylori-negative (43%) and -positive (57%) patients. CONCLUSION: In all but two patients with probable unidentified mutations there was agreement between the CYP2C19 phenotype determined by omeprazole and the genotype. The metabolism of omeprazole in patients with acid-related disorders is genetically determined and without relation to H. pylori infection.

Correlation between steady-state plasma concentrations of mianserin and trazodone in depressed patients.

OBJECTIVE: The correlations between steady-state plasma concentrations of mianserin and its active metabolite desmethylmianserin and those of trazodone and its active metabolite m-chlorophenylpiperazine (m-CPP) were examined in 19 depressed patients. METHODS: Ten patients received first mianserin (30 mg per day) and second trazodone (150 mg per day), while 9 patients received these treatments in the opposite sequence, with at least 2-week intervals between the two phases. Blood was sampled at steady state, 1-3 weeks after initiation of each treatment. Plasma concentrations of mianserin, the separate enantiomers S(+)- and R(-)-mianserin, desmethylmianserin, trazodone and m-CPP were measured by means of high-performance liquid chromatography. RESULTS: There was a significant correlation between steady-state plasma concentrations of trazodone and total mianserin (r = 0.59) or S(+)-mianserin (r = 0.57), but not R(-)-mianserin (r = 0.33). CONCLUSION: The present study thus suggests that the metabolic capacity of mianserin, especially the more active S(+)-enantiomer, and that of trazodone correlate to each other. This finding supports the previous suggestions that cytochrome P4502D6 is involved in the metabolism of mianserin and trazodone.

The CYP2D6 genotype and plasma concentrations of mianserin enantiomers in relation to therapeutic response to mianserin in depressed Japanese patients.

The relationship between therapeutic response to racemic mianserin and steady-state plasma concentrations of S(+)- and R(-)-mianserin was studied in 26 Japanese patients with major depression. The daily dose of mianserin was 30 mg, and the duration of treatment was 3 weeks. Regarding S-mianserin, the proportion of responders (final Montgomery-Asberg Depression Rating Scale score of 10 or less) was significantly higher in the plasma concentration range of 10 to 23 ng/mL than outside (10 of 11 vs. 3 of 15, p = 0.0005). Such a plasma concentration difference between responders and nonresponders was not found for R-mianserin. In 15 patients, the relationships between the CYP2D6 genotype, determined by allele-specific polymerase chain reaction analysis and Escherichia coli RI restriction fragment length polymorphism, plasma concentrations of the enantiomers, and the therapeutic response were studied. Five patients were homozygous for the wild type (wt) allele (wt/wt), nine were heterozygous for the CYP2D6Ch (Ch) allele causing decreased CYP2D6 activity (Ch/wt), and one patient was heterozygous for the Ch allele and the defect allele CYP2D6D (D) (Ch/D). The Ch/wt group showed significantly higher plasma concentrations of S-mianserin (mean +/- SD: 15 +/- 6 vs. 8 +/- 1 ng/mL, p = 0.007) and proportion of responders (8 of 9 vs. 1 of 5, p = 0.023) than the wt/wt group. The patient with the Ch/D genotype had the highest plasma concentration of S-mianserin (37 ng/mL) and a poor response. No significant relationship was found between the CYP2D6 genotype and plasma concentration of R-mianserin. The study presented here thus suggests that the CYP2D6 genotype plays a major role in controlling plasma concentration of the S-enantiomer of mianserin, which contributes to a major extent to the antidepressant effect during mianserin treatment.