Genetic polymorphism of CYP2D6 and CYP2C19 metabolism determined by phenotyping Israeli ethnic groups

Genetic polymorphism of the cytochrome P450 isoenzymes CYP2D6 and CYP2C19 was determined by phenotyping four ethnic groups of the Israeli population. The groups consisted of Ethiopian subjects, Yemenite subjects, and Russian subjects representing first-generation new immigrants and an Israeli Arab group. Dextromethorphan was used as the probe for CYP2D6 activity and mephenytoin was used for CYP2C19 activity. The two drugs were administered simultaneously and urine samples were collected over a period of 8 hours. The CYP2D6 phenotype was determined from the ratio of dextromethorphan conversion to dextrorphan and the CYP2C19 phenotype from the ratio of S-mephenytoin and R-mephenytoin. The used liquid chromatographic method was able to completely separate dextrorphan and dextromethorphan. Fluorescence detection allowed dextromethorphan quantification at 1 ng/mL. Mephenytoin enantiomers were completely separated in high-performance liquid chromatography and the respective fractions were collected and analyzed using a gas chromatography/mass spectrometry system with selective ion monitoring. The prevalence of poor metabolizer phenotype of dextromethorphan (CYP2D6) in the Yemenite (0%) and Ethiopian groups (0%) was significantly different from the prevalence in the Russian (17%) and Israeli Arab (9%) groups. A significant difference was also found in the distribution of the metabolic ratio of the extensive metabolizer phenotype between the Ethiopian group and the Russian and Yemenite groups. No significant difference was found in the prevalence of poor mephenytoin metabolizer phenotype (CYP2C19) between the Yemenite (8%), Ethiopian (6%), Russian (9%), and Israeli Arab (8%) groups. No difference was observed in the distribution of metabolic ratio within the extensive metabolizer phenotype subgroups of the four ethnic groups.     

The effect of perazine on the CYP2D6 and CYP2C19 phenotypes as measured by the dextromethorphan and mephenytoin tests in psychiatric patients

There is evidence that the antipsychotic drug perazine is an inhibitor of CYP2D6. This study aimed at evaluating its effect on CYP2D6 and CYP2C19 activities in submitting psychiatric patients to phenotyping with dextromethorphan and mephenytoin, respectively, substrates of these enzymes, before and during a treatment with perazine. A total of 31 patients were phenotyped with dextromethorphan (CYP2D6) and mephenytoin (CYP2C19) before and after a 2‐week treatment with 450 ± 51 mg/day (mean ± sd) perazine. At baseline, five patients appeared to be poor metabolizers (PM) of dextromethorphan and two patients of mephenytoin. The metabolic ratio (MR) of dextromethorphan/dextrorphan as determined in collected urine increased significantly (Wilcoxon; P < .0001) from baseline (0.39 ± 1.38 [mean ± sd]) till day 14 (1.46 ± 2.22). In 19 out of 26 extensive metabolizers (EM) of dextromethorphan, the phenotype changed from EM to PM. This suggests an almost complete inhibition of CYP2D6 by perazine and/or its metabolites. On the other hand, perazine (or some of its metabolites) did seemingly not inhibit CYP2C19. In conclusion, this study suggests that in patients treated with perazine and co‐medicated with CYP2D6 substrates, there could be an increased risk of adverse effects as a consequence of a pharmacokinetic interaction.     

Influence of gender and oral contraceptives on CYP2D6 and CYP2C19 activity in healthy volunteers

AIMS: The study was carried out in order to assess the effects of gender and the use of oral contraceptives (OCs) on CYP2D6 and CYP2C19 activities in healthy volunteers. METHODS: Six hundred and eleven Caucasian volunteers (330 males and 281 females; age range 18-49 years) were phenotyped with respect to CYP2D6 and CYP2C19 by means of the probe drugs dextromethorphan and mephenytoin, respectively. Extensive metabolisers were selected for this study. RESULTS: The median dextromethorphan/dextrorphan metabolic ratio in non-OC using females was significantly lower than in males (0.067 vs 0.080; P = 0.033) (mean difference in ln dextromethorphan/dextrorphan metabolic ratio 0.023, 95% CI 0.03-0.43). For the mephenytoin S/R ratio, no such difference was observed. However, OC using females had a significantly higher median mephenytoin S/R ratio than non-OC using females (0.230 vs 0.090; P < 0.001) (mean difference in ln mephenytoin S/R ratio 0.082, 95% CI 0.60-1.04). Moreover, females using combined OCs had a significantly higher median ratio than females using OCs with progestins only (median 0.258 vs 0.135; P = 0.008) (mean difference in ln mephenytoin S/R ratio 0.82, 95% CI 0.21-1.34). CONCLUSIONS: Given certain assumptions, the study indicates that females in the fertile age have a slightly higher CYP2D6 activity compared with males. There was no evidence of a gender difference in CYP2C19 activity. The use of combined OCs reduces the activity of CYP2C19, an effect that seems to be related to the ethinyloestradiol component.     

Lack of interaction of milnacipran with the cytochrome p450 isoenzymes frequently involved in the metabolism of antidepressants

OBJECTIVE: To compare the pharmacokinetics of milnacipran in extensive metabolisers (EMs) and poor metabolisers (PMs) of sparteine and mephenytoin, and to assess the influence of multiple administrations of milnacipran on the activity of cytochrome P450 (CYP) isoenzymes through its own metabolism and through various probes, namely CYP2D6 (sparteine/dextromethorphan), CYP2C19 (mephenytoin), CYP1A2 (caffeine) and CYP3A4 (endogenous 6-beta-hydroxy-cortisol excretion). METHODS: Twenty-five healthy subjects, 12 EMs for both sparteine/dextromethorphan and mephenytoin, nine EMs for mephenytoin and PMs for sparteine/dextromethorphan (PM(2D6)) and four PMs for mephenytoin and EMs for sparteine/dextromethorphan (PM(2C19)) were administered milnacipran as a single 50 mg capsule on day 1 followed by a 50 mg capsule twice daily for 7 days. The pharmacokinetics of milnacipran and its oxidative metabolites were assessed after the first dose (day 1) and after multiple administration (day 8), and were compared for differences between CYP2D6 and CYP2C19 PMs and EMs. Metabolic tests were performed before (day -2), during (days 1 and 8) and after (day 20) milnacipran administration. RESULTS: Milnacipran steady state was rapidly achieved. Metabolism was limited: approximately 50% unchanged drug, 30% as glucuronide and 20% as oxidative metabolite (mainly F2800 the N-dealkyl metabolite). Milnacipran administration to PM2D6 and PM2C19 subjects did not increase parent drug exposure or decrease metabolite exposure. Milnacipran oxidative metabolism is not mediated through CYP2D6 or CYP2C19 polymorphic pathways nor does it significantly interact with CYP1A2, CYP2C19, CYP2D6 or CYP3A4 activities. CONCLUSION: Limited reciprocal pharmacokinetic interaction between milnacipran and CYP isoenzymes would confer flexibility in the therapeutic use of the drug when combined with antidepressants. Drug-drug interaction risk would be low, even if the combined treatments were likely to inhibit CYP2D6 and CYP2C19 isoenzyme activities.     

Steady state plasma levels of the enantiomers of trimipramine and of its metabolites in CYP2D6-, CYP2C19- and CYP3A4/5-phenotyped patients

Steady state plasma concentrations of the (L)- and (D)-enantiomers of trimipramine (TRI), desmethyltrimipramine (DTRI), 2-hydroxytrimipramine (TRIOH) and 2-hydroxydesmethyl-trimipramine (DTRIOH) were measured in 27 patients receiving between 300 and 400 mg/day racemic TRI. The patients were phenotyped with dextromethorphan and mephenytoin, and the 8-hour urinary ratios of dextromethorphan/dextrorphan, dextromethorphan/3-methoxymorphinan, and (S)-mephenytoin/(R)mephenytoin were used as markers of cytochrome P-450IID6 (CYP2D6), CYP3A4/5 and CYP2C19 activities, respectively. One patient was a CYP2D6 and one was a CYP2C19 poor metabolizer. A stereoselectivity in the metabolism of TRI has been found, with a preferential N-demethylation of (D)-TRI and a preferential hydroxylation of (L)-TRI. CYP2D6 appears to be involved in the 2-hydroxylation of (L)-TRI, (L)DTRI and (D)-DTRI, but not of (D)-TRI, as significant correlations were measured between the dextromethorphan/dextrorphan ratios and the (L)-TRI/(L)-TRIOH (r = 0.45, p = 0.019), the (L)-DTRI/(L)-DTRIOH (r = 0.47, p = 0.014), and the (D)-DTRI/(D)-DTRIOH (r = 0.51, p = 0.006), but not with the (D)-TRI/(D)-TRIOH ratios (r = 0.29, NS). CYP2C19, but not CYP2D6, appears to be involved in the demethylation pathway, with a stereoselectivity toward the (D)-enantiomer of TRI, as a significant positive correlation was calculated between the mephenytoin (S)/(R) ratios and the concentrations to dose-to-weight ratios of (D)-TRI (r = 0.69, p = 0.00006). CYP3A4/5 appears to be involved in the metabolism of (L)-TRI to a presently not determined metabolite. The CYP2D6 poor metabolizer had the highest (L)-DTRI and (D)-DTRI concentrations to dose-to-weight ratios, and the CYP2C19 poor metabolizer had the highest (L)-TRI and (D)-TRI concentrations to dose-to-weight ratios of the group.     

No effect of the new antidepressant reboxetine on CYP2D6 activity in healthy volunteers.

The effect of the new antidepressant reboxetine on the activity of the cytochrome P450 (CYP) 2D6 isoenzyme was investigated in 10 healthy volunteers using dextromethorphan as a model CYP2D6 substrate. Each volunteer received a single 30 mg oral dose of dextromethorphan on three different occasions separated by an interval of at least 4 weeks: a) in a control session; b) after 1 week of treatment with reboxetine, 8 mg/day; and c) after 1 week of treatment with paroxetine (an inhibitor of CYP2D6 activity) 20 mg/day. Urine was collected over the next 8 hours for the determination of the dextromethorphan/dextrorphan metabolic ratio. All subjects were classified as extensive metabolizers (EM) with a dextromethorphan/dextrorphan ratio < 0.3. There were no notable changes in the urinary dextromethorphan/dextrorphan ratio in the reboxetine phase as compared to the control session. By contrast, there was a statistically significant increase in the metabolic ratio in the paroxetine phase (p < 0.001), with 4 subjects switching to poor metabolizer (PM) phenotype. These results suggest that reboxetine is unlikely to cause clinically significant interactions with substrates of CYP2D6.     

An optimized methodology for combined phenotyping and genotyping on CYP2D6 and CYP2C19

A method for simultaneous phenotyping and genotyping for CYP2D6 and CYP2C19 was tested. Six healthy volunteers were selected (three extensive and three poor metabolisers for CYP2D6). CYP2D6 was probed with dextromethorphan and metoprolol and CYP2C19 was probed with omeprazole. Blood samples were collected and analysed for dextromethorphan, dextrorphan, metoprolol, alpha-hydroxymetoprol, omeprazole and 5-hydroxyomeprazole by HPLC. Genotyping was performed for both CYP2D6 and CYP2C19. Generally, plasma levels could be measured up to 8 h post-dose except for alpha-hydroxymetoprolol in poor metabolizers (PMs) and dextromethorphan in extensive metabolizers (EMs) (35% below quantification limit). The correlation between the metabolic ratio based on timed individual measurements and the metabolic ratio based on the AUC0-12 values was significant at 3 h post-dose for all probes. In conclusion, the following procedure is suggested: administer metoprolol (100 mg) and omeprazole (40 mg); after 3 h, take a blood sample to assess the genotype and the metabolic ratio for CYP2D6 (metoprolol over alpha-hydroxymetoprolol) and CYP2C19 (omeprazole over 5-hydroxyomeprazole) in plasma. With this procedure, all necessary information on the individual CYP2D6 and CYP2C19 metabolising capacity can be obtained in a practical, single-sample approach.     

CYP2D6 phenotyping with dextromethorphan

Genetically determined individual differences in the ability to oxidize certain drugs have raised recently a considerable interest because of clinical importance of this problem. Determination of CYP2D6 oxidation phenotype is used to obtain more efficient pharmacotherapy and to explain lower efficacy of some drugs and presentation of adverse effects in particular patients. The aim of this study was to identify the CYP2D6 oxidation phenotype with dextromethorphan (DM) as a probe drug. The study included 85 healthy volunteers of Polish origin. DM (40 mg) was given orally to healthy adults and 10-h urine samples were collected. DM and the metabolite dextrorphan (DX) were analyzed by the HPLC method. Phenotyping was performed using the metabolic ratio (MR) calculated as the urinary DM/DX output. Based on the metabolic ratio, we can distinguish extensive (EM) and poor (PM) metabolizers in human population. Individuals with a dextromethorphan MR greater than 0.3 (log > -0.5) were classified as PMs. In our study, the frequency of the PM phenotype was 9.4%, which is in the range found in other Caucasian populations (3-10%).     

Influence of donor and recipient genotypes on CYP2D6 phenotype after liver transplantation: a study of mutations CYP2D6*3 and CYP2D6*4

Objective: After liver transplantation (LT), genotypic differences between the recipient and the transplanted liver, medications and post-LT complications may all affect drug metabolism. We have studied the effect of two CYP2D6 mutations in the donor and the recipient on post-LT CYP2D6 phenotype. Method: The CYP2D6 phenotype was assessed in 48 patients before and after LT with debrisoquine or␣dextromethorphan. CYP2D6*3 (CYP2D6A) and CYP2D6*4 (CYP2D6B) mutations were detected in the donor and the recipient using polymerase chain reaction. Results: Before LT, 40 subjects were classified as extensive metabolisers (EM) and 8 as poor metabolisers (PM); after transplantation, 41 were EMs and 7 were PMs. Genotype and phenotype were in agreement in 100% of EMs and 40% of PMs. The low percentage of agreement in PMs could not be explained by severely altered liver function. The phenotype of 13 subjects was apparently changed by LT: 6 EMs became PMs and 7 PMs became EMs. All four subjects in whom genotype changed following LT had a corresponding change in phenotype: two EM subjects became PMs and two PM subjects became EMs. Conclusion: The low percentage of agreement in PMs may be partly explained by mutations other than CYP2D6*3 and CYP2D6*4. Nevertheless, our study shows that the CYP2D6 genotype of the donor controls the phenotype of the recipient of a liver transplantation. Received: 2 June 1997 / Accepted in revised form: 9 October 1997     

Evaluation of dextromethorphan metabolism using hepatocytes from CYP2D6 poor and extensive metabolizers

It is important to estimate the defective metabolism caused by genetic polymorphism of drug metabolizing enzymes before the clinical stage. We evaluated the utility of cryopreserved human hepatocytes of CYP2D6 poor metabolizer (PM) for the estimation of the metabolism in PM using dextromethorphan (DEX) as the probe drug for CYP2D6 substrate. The results of low formations of dextrorphan (DXO) and 3-hydroxymorphinan (3-HM) in CYP2D6 PM hepatocytes incubated with dextromethorphan reflected the clinical data. Formation of 3-methoxymorphinan (3-MEM) normalized by CYP3A4/5 activity in the PM hepatocytes reached about 2.8-fold higher than that in CYP2D6 extensive metabolizer (EM) hepatocytes, which clearly showed the compensatory metabolic pathway of O-demethylation catalyzed by CYP2D6 as seen in clinical study. On the contrary, in the condition of the EM hepatocytes with CYP2D6 inhibitors, the enhancement of 3-MEM formation was not observed. In phase II reaction, the glucuronide formation rate of DXO in the PM hepatocytes was lower than that in the EM hepatocytes, which was consistent with clinical data of DXO-glucuronide (DXO-glu) concentration. These results would suggest that CYP2D6 PM hepatocytes could be a good in vitro tool for estimating CYP2D6 PM pharmacokinetics.     

Interferon-beta treatment in patients with multiple sclerosis does not alter CYP2C19 or CYP2D6 activity

AIMS: To determine CYP2C19 and CYP2D6 activity in patients with multiple sclerosis (MS) before and during interferon (IFN)-beta treatment. METHODS: CYP2C19 and CYP2D6 activities were assessed using the probe drugs mephenytoin and debrisoquine, respectively. Urinary mephenytoin (S/R) and debrisoquine (debrisoquine/hydroxy-debrisoquine) metabolic ratios (MR) were determined in 10 otherwise healthy Caucasian multiple sclerosis (MS) patients in the initial stage of the disease, prior to and 1 month after commencing treatment with IFN-beta (Avonex, Rebif or Betaferon). In addition, CYP2C19*2, CYP2C19*3, CYP2D6*3, CYP2D6*4, and CYP2D6*5 genotyping was performed. RESULTS: There was no significant difference in the (S)/(R) mephenytoin ratio (mean difference 0.04; 95% CI -0.03, 0.11) or the debrisoquine MR (mean difference 0.29; 95% CI -0.44, 1.02) before and during regular IFN-beta treatment in extensive metabolizers (EM) (P = 0.5 and P = 0.4 for the respective probe drugs; n = 9 subjects). There were also no differences between the different IFN-beta treatments (P = 0.6 for the (S)/(R) mephenytoin ratio and P = 0.7 for the debrisoquine MR; anova; n = 10). CONCLUSIONS: IFN-beta treatment did not affect the activity of CYP2C19 or CYP2D6. The results suggest that it is safe to administer CYP2C19 or CYP2D6 substrates, without dose adjustment, to patients treated with IFN-beta.     

Multiple Human Cytochromes Contribute to Biotransformation of Dextromethorphan In-vitro: Role of CYP2C9, CYP2C19, CYP2D6, and CYP3A

Cytochromes mediating the biotransformation of dextromethorphan to dextrorphan and 3-methoxymorphinan, its principal metabolites in man, have been studied by use of liver microsomes and microsomes containing individual cytochromes expressed by cDNA-transfected human lymphoblastoid cells. In-vitro formation of dextrorphan from dextromethorphan by liver microsomes was mediated principally by a high-affinity enzyme (K_m (substrate concentration producing maximum reaction velocity) 3–13 μM). Formation of dextrorphan from 25 μM dextromethorphan was strongly inhibited by quinidine (IC50 (concentration resulting in 50% inhibition) = 0.37 μm); inhibition by sulphaphenazole was approximately 18% and omeprazole and ketoconazole had minimal effect. Dextrorphan was formed from dextromethorphan by microsomes from cDNA-transfected lymphoblastoid cells expressing CYP2C9, ?2C19, and ?2D6 but not by those expressing CYP1A2, ?2E1 or ?3A4. Despite the low in-vivo abundance of CYP2D6, this cytochrome was identified as the dominant enzyme mediating dextrorphan formation at substrate concentrations below 10 μM. Formation of 3-methoxy-morphinan from dextromethorphan in liver microsomes proceeded with a mean K_m of 259 μM. For formation of 3-methoxymorphinan from 25 μM dextromethorphan the IC50 for ketoconazole was 1.15 μM; sulphaphenazole, omeprazole and quinidine had little effect. 3-Methoxymorphinan was formed by microsomes from cDNA-transfected lymphoblastoid cells expressing CYP2C9, ?2C19, ?2D6, and ?3A4, but not by those expressing CYP1A2 or ?2E1. CYP2C19 had the highest affinity (K_m = 49 μM) whereas CYP3A4 had the lowest (K_m = 1155 μM). Relative abundances of the four cytochromes were determined in liver microsomes by use of the relative activity factor approach. After adjustment for relative abundance, CYP3A4 was identified as the dominant enzyme mediating 3-methoxymorphinan formation from dextromethorphan, although CYP2C9 and ?2C19 were estimated to contribute to 3-methoxymorphinan formation, particularly at low substrate concentrations. Although formation of dextrorphan from dextromethorphan appears to be sufficiently specific to be used as an in-vitro or in-vivo index reaction for profiling of CYP2D6 activity, the findings raise questions about the specificity of 3-methoxymorphinan formation as an index of CYP3A activity.     

Mephenytoin as a probe for CYP2C19 phenotyping:effect of sample storage, intra-individual reproducibility and occurrence of adverse events

AIMS: To further evaluate mephenytoin as a probe for CYP2C19 phenotyping. METHODS: Healthy subjects (n = 2638) were phenotyped using the urinary (S)-mephenytoin to (R)-mephenytoin ratio. This method was evaluated for (a) the stability of the S/R-ratio following sample storage, (b) the intraindividual reproducibility of the ratio, and (c) the occurrence of adverse events. RESULTS: After prolonged storage, the S/R-ratio of samples from extensive metabolisers (EM) increased up to 85%. In 1.5% of the cases (1 out 66), this led to incorrect classification of phenotype. In EMs, but not in poor metabolisers (PMs), the S/R-ratio increased after acid treatment. The intraindividual reproducibility of the mephenytoin phenotyping procedure was 28%. No major side-effects were observed and there was no relationship between the incidence of side-effects and the phenotype of the subject. CONCLUSIONS: After prolonged storage the S/R-ratio significantly increased in EMs and, although low, the risk of incorrect classification should not be ignored. Our data support the use of mephenytoin as a safe drug for CYP2C19 phenotyping.     

The pharmacokinetics of ketobemidone are not affected by CYP2D6 or CYP2C19 phenotype

The pharmacokinetics of orally administered ketobemidone were investigated in three groups of healthy volunteers with respect to cytochrome P450 (CYP) 2D6 and CYP2C19 phenotypes: extensive metabolisers (EMs) of debrisoquine and mephenytoin (EMdeb/EMmeph), poor metabolisers (PMs) of debrisoquine (PMdeb/EMmeph) and poor metabolisers of mephenytoin (PMmeph/EMdeb). Peak plasma concentration, oral clearance, area under the plasma concentration-time curve, half-life, volume of distribution and mean residence time were not significantly different among the three groups. There was no correlation between oral clearance of ketobemidone and debrisoquine or mephenytoin metabolic ratios. Further, the urinary excretion of ketobemidone and norketobemidone was not affected by the phenotype for either CYP2D6 or CYP2C19. However, a substantial variation in plasma concentration was observed within all three groups. The results indicate that the metabolism of ketobemidone is not dependent on CYP2D6 or CYP2C19. PMs of debrisoquine or mephenytoin, as well as patients who are concomitantly treated with inhibitors of CYP2D6 or CYP2C19, are not expected to be at higher risk of adverse effects. However, due to the interindividual variability in plasma levels of ketobemidone, independent of phenotype, individual dosing based on the clinical response and therapeutic drug monitoring is recommended.     

Inhibition of CYP2D6 activity by bupropion

The purpose of this study was to assess the effect of bupropion on cytochrome P450 2D6 (CYP2D6) activity. Twenty-one subjects completed this repeated-measures study in which dextromethorphan (30-mg oral dose) was administered to smokers at baseline and after 17 days of treatment with either bupropion sustained-release (150 mg twice daily) or matching placebo. Subjects quit smoking 3 days before the second dextromethorphan administration. To assess CYP2D6 activity, urinary dextromethorphan/dextrorphan metabolic ratios were calculated after an 8-hour urine collection. Thirteen subjects received bupropion, and 8 received placebo. In those receiving active medication, the dextromethorphan/dextrorphan ratio increased significantly at the second assessment relative to the first (0.012 +/- 0.012 vs. 0.418 +/- 0.302; P < 0.0004). No such change was observed in those randomized to placebo (0.009 +/- 0.010 vs. 0.017 +/- 0.015; P = NS). At baseline, all subjects were phenotypically extensive CYP2D6 metabolizers (metabolic ratio <0.3); after treatment, 6 of 13 subjects receiving bupropion, but none of those receiving placebo, had metabolic ratios consistent with poor CYP2D6 metabolizers. Bupropion is therefore a potent inhibitor of CYP2D6 activity, and care should be exercised when initiating or discontinuing bupropion use in patients taking drugs metabolized by CYP2D6.     

Concordance of P450 2D6 (debrisoquine hydroxylase) phenotype and genotype: inability of dextromethorphan metabolic ratio to discriminate reliably heterozygous and homozygous extensive metabolizers.

Debrisoquine-hydroxylase (P450 2D6) not equal to phenotype was determined in 116 individuals using dextromethorphan as the substrate probe. Polymerase chain reaction and restriction fragment length polymorphism analyses were used to detect inactivating mutations in the CYP2D6 gene and assign genotype in all 116 individuals. Using a urinary metabolic ratio (DM/DT) of > or = 0.3 to define poor metabolizer (PM) phenotypes, 96 subjects were extensive metabolizers (EM) and 20 were PMs. The CYP2D6(B) mutation was the most common mutation, present in 18% of phenotypic EM alleles and 66% of the alleles in PM phenotypes. The CYP2D6(A) mutation (8% of PM alleles) and the CYP2D6 gene deletion (2.6% of PM alleles) were found less frequently. Seven different variants of the CYP2D6 gene were found. In subjects with two mutant alleles, genotype correctly predicted the PM phenotype in 100% (n = 13). Overall, genotype agreed with phenotype assignments in 109 of 116 (94%) subjects. Seven subjects with a wild-type allele at the CYP2D6(A) and CYP2D6(B) loci were phenotypic PMs, representing the only discrepant results. These discrepancies could be due to the imprecision of phenotype assignment or to as yet unknown mutations in CYP2D6. Although the median urinary metabolic ratio was significantly lower in homozygous EMs compared with heterozygous EMs, there was extensive overlap in metabolic ratios in these two groups, indicating that the DM/DT metabolic ratio cannot reliably discriminate homozygous EMs from heterozygous EMs.     

The prevalence of CYP2D6 and CYP2C19 genotypes in a population of healthy Dutch volunteers

AIM: This study was performed in a sample of the Dutch population to estimate the prevalence of noncoding mutations of CYP2D6 and CYP2C19 as obtained by genotyping. In addition, the predictability of the genotyping strategy was assessed. METHODS: The CYP2D6 and CYP2C19 status of 765 unrelated healthy volunteers was evaluated. Dextromethorphan and mephenytoin were used for determining the phenotypes. Genotyping was performed by PCR on the most common null alleles for CYP2D6 (except for CYP2D6*5) and CYP2C19. RESULTS: For CYP2D6, the most frequently observed null allele was CYP2D6*4, which accounted for 89% of all null alleles. The prevalence of poor metabolizers (PMs) in healthy volunteers was 5.5%, which was lower than that found previously by phenotyping (8.0%; chi2 test P = 0.009). For CYP2C19*2 and CYP2C19*3, the frequencies were 13.3% and 0.2%, respectively. The S:R ratio was significantly higher in heterozygous subjects (S:R ratio 0.22) than in homozygous wild type subjects (S:R ratio 0.11). Comparison of all subjects below 45 years showed a significantly higher S:R ratio in the female ones compared to the male ones, especially in heterozygous subjects (S:R ratio 0.39 vs. 0. 19; P < 0.001). CONCLUSIONS: The frequencies of CYP2D6 and CYP2C19 allelic variants were in accordance with other European populations. Assessment of *3, *4, *6, *7, and *8 alleles for CYP2D6, and *2 and *3 for CYP2C19, predicted the phenotype with an accuracy of over 98.6%. A gene-dose effect was found for CYP2C19. CYP2C19 heterozygous female subjects had a decreased CYP2C19 activity that may be at least partly due to the use of oral contraceptives.     

Genetic polymorphism of CYP2D6 in North Indian subjects

Objectives: CYP2D6 polymorphism of clinical relevance occurs with variable frequency in different ethnic groups. Since this polymorphism has not been studied in a North Indian population, the present study was undertaken. Methods: One hundred healthy unrelated North Indian subjects received 30?mg dextromethorphan (DM) orally at bed-time. The amounts of DM and its metabolite, dextrorphan (DR), excreted in 8?h urine were estimated by high performance liquid chromatography. Metabolic ratio (DM/DR excreted in 8?h) was used as an index of the metabolic status of an individual. Results: The analysis of the data by frequency distribution histogram, probit and NTV plots demonstrated bimodal distribution of the North Indian subjects with respect to hepatic CYP2D6. Out of 100 subjects, 97 were extensive metabolizers (EMs), whereas three were poor metabolizers (PMs). EMs and PMs excreted 29.82 and 2.67?μ mol DR (mean value) and 2.59 and 8.82?μ mol DM (mean value) in 8?h, respectively. MR and log MR was 197- and 2.2-fold higher in PMs versus EMs. The antimode value of zero was determined by visual observation in frequency distribution histogram and inflection point in probit plot. Conclusion: From this study, it can be concluded that the PM phenotype of CYP2D6 occurs with a frequency of 3% (95% confidence interval of 0.33%–6.33%) in North Indians.     

Debrisoquin and S-Mephenytoin Hydroxylation Phenotypes and CYP2D6 Genotypes in an Estonian Population

Abstract: The polymorphisms of debrisoquin (CYP2D6) and S-mephenytoin (CYP2C19) hydroxylation were studied in 210 unrelated healthy native Estonians by coadministration of mephenytoin and debrisoquin or dextromethorphan. Among the 210 volunteers 21 (10%) were poor metabolizers of debrisoquin/dextromethorphan and two (0.95%) were poor metabolizers of S-mephenytoin. By pooling these data with an earlier study on 156 Estonians, the prevalences of poor metabolizers of debrisoquin/dextromethorphan and poor metabolizers of S-mephenytoin were 7.6% and 2.2%, respectively. The CYP2D6 genotype of 151 subjects was analysed by allele-specific PCR amplification for the defect alleles CYP2D6A and CYP2D6B. All poor metabolizers of debrisoquin carried two defect CYP2D6-alleles. The phenotype (extensive or poor metabolizer) was in all subjects correctly predicted by the genotype. The frequencies of the defect alleles CYP2D6B and CYP2D6A among these 151 subjects (including 14 poor metabolizers - 9.3%) were 21.5% and 2.3%, respectively. DNA from 6 subjects with very high CYP2D6 activity (debrisoquin MR<0.1) was analysed by EcoRI RFLP to identify duplicated or amplified CYP2D6-genes. Two of the subjects were found to carry a duplicated CYP2D6L-gene. In conclusion, the distribution of genetically determined metabolic capacities of CYP2D6 and CYP2C19 in Estonian unrelated subjects did not differ significantly from that in other Caucasian populations. The CYP2D6 phenotype was predicted by PCR-based amplification for the CYP2D6A and CYP2D6B-aellels.     

Effect of honey on CYP3A4, CYP2D6 and CYP2C19 enzyme activity in healthy human volunteers

Honey is a common food supplement but not many studies have studied honey and drug interaction. This study investigates the influence of 7 days of honey administration on the activity of CYP3A4, CYP2D6 and CYP2C19 drug-metabolizing enzymes in healthy volunteers by using appropriate biomarker and probe drugs. A within-group pharmacokinetic study was done in 12 healthy volunteers. Urine samples (0-8 hr) were collected after administration of 30 mg of oral dextromethorphan (probe drug for CYP2D6) for analysis of dextromethorphan and dextrorphan. A plasma sample (4 hr) was collected after administration of 200 mg of oral proguanil (probe drug for CYP2C19) for the analysis of proguanil and cycloguanil. Urine samples (0-24 hr) were collected for the analysis of 6beta-hydroxycortisol (biomarker for CYP3A4). The volunteers were administered honey for 7 days. Subsequently blood and urine samples were collected after drug dosing as before. These samples were analysed for drug and metabolite concentrations in urine and plasma using high performance liquid chromatography method. Seven days of honey administration resulted in statistically significant increase in 24-hr urinary excretion of 6beta-hydroxycortisol. However, the metabolic ratios of dextromethorphan and proguanil were not significantly altered after 7 days of honey administration. Honey obtained from Western Ghats of southern India may induce CYP3A4 enzyme activity but not CYP2D6 and CYP2C19 enzyme activities.