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Tamoxifen is a mainstay in the treatment of estrogen receptor-positive breast cancer and is metabolized to more than 30 different compounds. Little is known about in vivo concentrations of estrogenic metabolites E-metabolite E, Z-metabolite E, and bisphenol and their relevance for tamoxifen efficacy. Therefore, we developed a highly sensitive HPLC-ESI-MS/MS quantification method for tamoxifen metabolites bisphenol, E-metabolite E, and Z-metabolite E as well as for the sex steroid hormones estradiol, estrone, testosterone, androstenedione, and progesterone. Plasma samples were subjected to protein precipitation followed by solid phase extraction. Upon derivatization with 3-[(N-succinimide-1-yl)oxycarbonyl]-1-methylpyridinium iodide, all analytes were separated on a sub-2-μm column with a gradient of acetonitrile in water with 0.1 % of formic acid. Analytes were detected on a triple-quadrupole mass spectrometer with positive electrospray ionization in the multiple reaction monitoring mode. Our method demonstrated high sensitivity, accuracy, and precision. The lower limits of quantification were 12, 8, and 25 pM for bisphenol, E-metabolite E, and Z-metabolite E, respectively, and 4 pM for estradiol and estrogen, 50 pM for testosterone and androstenedione, and 25 pM for progesterone. The method was applied to plasma samples of postmenopausal patients taken at baseline and under tamoxifen therapy. Graphical Abstract Sample preparation and derivatization for highly sensitive quantification of estrogenic tamoxifen metabolites and steroid hormones by HPLC-MS/MS.

The therapeutic effect of tamoxifen depends on active metabolites, e.g., cytochrome P450 2D6 (CYP2D6) mediated formation of endoxifen. To test for additional relationships, 236 breast cancer patients were genotyped for CYP2D6, CYP2C9, CYP2B6, CYP2C19, CYP3A5, UGT1A4, UGT2B7, and UGT2B15; also, plasma concentrations of tamoxifen and 22 of its metabolites, including the (E)-, (Z)-, 3-, and 4′-hydroxymetabolites as well as their glucuronides, were quantified using liquid chromatography–tandem mass spectrometry (MS). The activity levels of the metabolites were measured using an estrogen response element reporter assay; the strongest estrogen receptor inhibition was found for (Z)-endoxifen and (Z)-4-hydroxytamoxifen (inhibitory concentration 50 (IC50) 3 and 7 nmol/l, respectively). CYP2D6 genotypes explained 39 and 9% of the variability of steady-state concentrations of (Z)-endoxifen and (Z)-4-hydroxytamoxifen, respectively. Among the poor metabolizers, 93% had (Z)-endoxifen levels below IC90 values, underscoring the role of CYP2D6 deficiency in compromised tamoxifen bioactivation. For other enzymes tested, carriers of reduced-function CYP2C9 (*2, *3) alleles had lower plasma concentrations of active metabolites (P < 0.004), pointing to the role of additional pathways. Clinical Pharmacology & Therapeutics (2011) 89 5, 708–717. doi:10.1038/clpt.2011.27

The impact of thyroid dysfunction on the regulation, expression, and function of ABCB1 remains unclear. We therefore investigated ABCB1 mRNA expression and function in patients with thyroid dysfunction and studied the disposition of the ABCB1 substrate digoxin before and after treatment for thyroid disease. In patients with hypothyroidism, normalization of thyroid function was associated with a 1.8-fold increase in mRNA expression and a 26% increase in rhodamine efflux from CD56+ cells. In hypothyroidism, digoxin clearance was significantly decreased, whereas bioavailability, volume of distribution, half-life time, and protein binding were unaltered. In hyperthyroidism, ABCB1 mRNA expression, rhodamine efflux, and disposition of digoxin were not significantly affected other than in relation to renal clearance. Experiments using the LS174T cell line indicated that the gene is a direct target of thyroid hormone receptors. In conclusion, thyroid abnormalities can exert significant effects on the expression of P-glycoprotein, thereby altering the disposition and action of drugs that are substrates of P-glycoprotein. Clinical Pharmacology & Therapeutics (2010) 88 5, 685–694. doi: 10.1038/clpt.2010.176

We studied the efficacy of propafenone in preventing atrial tachyarrhythmias after cardiac surgery, and the possible relationships between CYP2D6 polymorphism and the efficacy, pharmacokinetics, and tolerability of propafenone. One hundred and sixty patients were randomized (double blind) to receive propafenone (n= 78) or placebo (n= 82) for 1 week after cardiac surgery. The patients who were assigned to the propafenone group received 1 mg/kg infused in 1 h, followed by a continuous infusion at a rate of 4 mg/kg/24 h until the following morning, and subsequently 450 mg/day orally until the sixth postoperative day. Thirty-seven patients completed the trial in the propafenone group and 45 in the placebo group. The frequency of occurrence of atrial tachyarrhythmia was lower in the propafenone group than in the placebo group (29.7% vs. 53.3%, P< 0.05; relative risk, 0.56). Plasma propafenone concentrations were markedly influenced by CYP2D6 genotype-derived phenotype.

The multidrug resistance protein 4 (MRP4) is an efflux transporter involved in the transport of endogenous substrates and xenobiotics. We measured MRP4 mRNA and protein expression in human livers and found a 38- and 45-fold variability, respectively. We sequenced 2 kb of the 5'-flanking region, all exons and intron/exon boundaries of the MRP4 gene in 95 patients and identified 74 genetic variants including 10 non-synonymous variations, seven of them being located in highly conserved regions. None of the detected polymorphisms was significantly associated with changes in the MRP4 mRNA or protein expression. Immunofluorescence microscopy indicated that none of the non-synonymous variations affected the cellular localization of MRP4. However, in cholestatic patients the MRP4 mRNA and protein expression both were significantly upregulated compared to non-cholestatic livers (protein: 299+/-138 vs 100+/-60a.u., P

Response to treatment with the antiestrogen tamoxifen is variable and at least partially due to its highly complex metabolism. Tamoxifen is transformed by polymorphic and inducible cytochrome P450 enzymes to a large number of metabolites with varying biological activities. The estrogen receptor dependent growth inhibitory effect of antiestrogens is mediated by activation of antiproliferative Transforming Growth Factor beta (TGFbeta) signal transduction pathways. The aim of the present study was to establish if TGFbeta2 or TGFbeta receptor II (TbetaRII), could be used as markers to assess the pharmacological potency of tamoxifen and its metabolites. Consequently, we analyzed the growth inhibitory effect of tamoxifen and its major metabolites and explored whether it correlated with their capacity to induce TGFbeta2 and TbetaRII expression. Human breast cancer cells (MCF-7 and T47D) were treated with tamoxifen and tamoxifen metabolites and mRNA expression of TGFbeta2 and TbetaRII was analyzed by quantitative RT-PCR. Only two metabolites 4-hydroxytamoxifen and N-desmethyl-4-hydroxytamoxifen had significant antiproliferative activity and were able to induce TGFbeta2 and TbetaRII. Plasma concentrations of these metabolites are usually very low in patients. However, even minor growth inhibitory effects at concentrations which are below the limit of quantification in plasma samples resulted in clearly discernible effects on expression of TGFbeta2 and TbetaRII. Taken together, our data demonstrate that TGFbeta2 and TbetaRII are very specific and sensitive biomarkers for the antiestrogenic activity of tamoxifen metabolites in breast cancer.

To characterise the comparative potency of optically pure (R)- and (S)-verapamil as regards negative dromotropic effects on atrioventricular (AV) node conduction and to compare the hemodynamic effects of single doses of the enantiomers in healthy volunteers.Eight healthy volunteers received a single oral dose of 120 mg (S)-verapamil, 480 mg (R)-verapamil, 240 mg racemic verapamil (rac-verapamil) or placebo on 4 separate occasions. Serum concentrations of (R)- and (S)-verapamil were measured up to 24 h. Cardiovascular effects were assessed by electrocardiography, measurement of blood pressure and transthoracic impedance cardiography (cardiac output and total peripheral resistance). The comparative potency of (R)- and (S)-verapamil with regard to prolongation of the PR interval in the surface ECG was estimated by use of the areas under the effect-time and serum concentration-time curves and linear regression analyses of per cent change in PR interval from baseline versus the logarithm of serum (R)- or (S)-verapamil concentration.The PR interval was significantly prolonged after all verapamil treatments as compared with placebo. (S)-verapamil was 20.6-21.8 times more potent than (R)-verapamil with regard to negative dromotropic effects. (R)-verapamil caused a significantly greater maximum reduction in the mean arterial pressure (MAP) than placebo [15.9+/-6.8 versus 8.7+/-3.2 mmHg (mean+/-SD); 95% CI on the difference, 0.79-13.7 mmHg; p0.05], whereas MAP was not affected by the other verapamil treatments. No significant changes were observed in heart rate, cardiac output and total peripheral resistance after any verapamil treatment as compared with placebo.(S)-verapamil was about 20 times more potent than (R)-verapamil with regard to negative dromotropic effects on AV node conduction. (R)-verapamil but not (S)-verapamil significantly reduced the MAP as compared with placebo.

Background and Objective On the basis of in vitro studies indicating that ursodeoxycholic acid (UDCA) is a cytochrome P450 (CYP) 3A4 inducer and a pregnane X receptor activator and because the pregnane X receptor is a transcriptional regulator of multidrug resistance 1 (MDR1)/P-glycoprotein (P-gp), we postulated that UDCA might decrease the bioavailability of CYP3A4 and P-gp probe drugs in humans. The main objective of this study was to determine whether UDCA alters the pharmacokinetics of digoxin and midazolam. The secondary objective was to determine whether the intestinal expression of P-gp and CYP3A4 is increased by UDCA. Methods The effect of UDCA on MDR1 and CYP3A4 messenger ribonucleic acid (mRNA) expression was investigated in human colon carcinoma cell lines LS174T and Caco-2. Eight healthy volunteers participated in this open, nonrandomized 2-period study, in which the effects of UDCA (13 mg · kg−1 · d−1 during 2 weeks) versus control on the pharmacokinetics of digoxin (0.5-mg single intravenous infusion), d3-digoxin (3-fold deuterated digoxin, 0.5-mg single oral dose), and midazolam (7.5-mg single oral dose) were compared. Duodenal biopsy specimens were obtained during both periods to quantify MDR1/P-gp and CYP3A4 expression. Results In vitro UDCA induced MDR1 and CYP3A4 mRNA in Caco-2 cells but not in LS174T cells. In humans UDCA significantly decreased the extent of digoxin absorption from 0.77 to 0.70 and the oral d3-digoxin area under the plasma concentration-time curve from 0 to 4 hours from 6.4 ± 1.7 ng · h · mL−1 to 5.3 ± 1.5 ng · h · mL−1 (P=.01 and P=.05, respectively). UDCA had no detectable effects on the pharmacokinetics of midazolam or the intestinal mRNA and protein expression levels of MDR1/P-gp and CYP3A4. Conclusion Under the conditions in our study, UDCA only modestly decreased digoxin disposition without detectable changes in midazolam pharmacokinetics. The clinical relevance of these findings remains to be determined. Clinical Pharmacology & Therapeutics (2006) 79, 449–460; doi: 10.1016/j.clpt.2006.01.005

Background and Objective The polymorphism of SLCO1B1 (solute carrier organic anion transporter family, member 1B1), encoding the hepatic uptake transporter organic anion transporting polypeptide 1B1, has been associated with increased pravastatin concentrations in single-dose studies. We have investigated whether this polymorphism influences the pharmacokinetics and lipid-lowering efficacy of multiple-dose pravastatin. Methods A prospective, parallel-group study of 16 healthy volunteers, including 8 carriers of an SLCO1B1 variant haplotype and 8 control subjects, was carried out. Pravastatin pharmacokinetics and reduction in total and low-density lipoprotein (LDL) cholesterol concentrations were analyzed after treatment with 40 mg pravastatin daily for 3 weeks. Results The mean area under the plasma concentration-time curve of pravastatin was 110% higher (305.0 ± 149.4 ng · h/mL [mean ± SD] versus 144.9 ± 48.2 ng · h/mL, P = .012) and the mean peak concentration in plasma was 231% higher (174.5 ± 98.6 ng/mL versus 52.7 ± 22.1 ng/mL, P = .0042) in the SLCO1B1 variant haplotype group than in the control group. Pravastatin significantly reduced the concentrations of total and LDL cholesterol in both groups. The mean percentage reduction in total cholesterol concentration was 13.1% ± 9.1% and 19.1% ± 8.3% in the variant and control groups, respectively (P = .19; 95% confidence interval of difference between groups, −15.3% to 3.3%). The mean percentage reduction in LDL cholesterol concentration was 27.7% ± 8.3% in the variant group and 32.3% ± 11.2% in the control group (P = .37; 95% confidence interval for difference, −15.1% to 6.0%). Conclusions Despite considerably higher plasma pravastatin concentrations in carriers of an SLCO1B1 variant haplotype, there was no significant difference in the lipid-lowering efficacy of pravastatin between the variant haplotype and control groups. However, this pilot study had sufficient statistical power to detect only a large difference in lipid response between the 2 groups. Further clinical studies are warranted to characterize the impact of the SLCO1B1 polymorphism on the lipid response to pravastatin. Clinical Pharmacology & Therapeutics (2006) 79, 419–426; doi: 10.1016/j.clpt.2006.01.010

Objective Our objective was to study the impact of the cytochrome P450 (CYP) 2D6 polymorphism on the tolerability of metoprolol in a real-life primary care setting. The adverse effects studied comprised effects related to the central nervous system, cardiovascular effects, and sexual dysfunction. Methods Patients in whom treatment with metoprolol was considered were enrolled into this prospective, 6-week multicenter study. The dosage of metoprolol was determined on an individual basis and could be freely adjusted on clinical grounds. The indication for treatment was hypertension in about 90% of cases. Systolic and diastolic blood pressure, resting heart rate, and plasma metoprolol and α-hydroxymetoprolol concentrations were measured. CYP2D6 genotyping covered alleles *3 to *10 and *41 and the duplications. Possible adverse effects of metoprolol were systematically assessed over a 6-week period by means of standardized rating scales and questionnaires. Results The final study population comprised 121 evaluable patients (all white patients); among them, there were 5 ultrarapid metabolizers (UMs) (4.1%), 91 extensive metabolizers (EMs) (75%), 21 intermediate metabolizers (IMs) (17%), and 4 poor metabolizers (PMs) (3.3%). Plasma metoprolol concentrations normalized for the daily dose and metoprolol/α–hydroxymetoprolol ratios at steady state were markedly influenced by CYP2D6 genotype and displayed a gene–dose effect. The median of the dose–normalized metoprolol concentration was 0.0088 ng/mL, 0.047 ng/mL, 0.34 ng/mL, and 1.34 ng/mL among UMs, EMs, IMs, and PMs, respectively (P< .0001). There was no significant association between CYP2D6 genotype–derived phenotype (EMs and UMs combined versus PMs and IMs combined) and adverse effects during treatment with metoprolol. There was a tendency toward a more frequent occurrence of cold extremities in the PM plus IM group as compared with the EM plus UM group (16.0% versus 4.2%, P = .056; relative risk, 3.8 [95% confidence interval, 1.03–14.3]). Conclusions CYP2D6 genotype–derived phenotype was not significantly associated with a propensity for adverse effects to develop during treatment with metoprolol. However, the results concerning tolerability of metoprolol in PMs were inconclusive because of the small number of PMs enrolled. Clinical Pharmacology & Therapeutics (2005) 78, 378–387; doi: 10.1016/j.clpt.2005.07.004

Background and Objective A large interindividual variability exists in the plasma concentrations of repaglinide. Our aim was to investigate possible associations between the pharmacokinetics of repaglinide and single nucleotide polymorphisms (SNPs) in the genes encoding for the drug transporters organic anion transporting polypeptide 1B1 (OATP1B1) (SLCO1B1) and P-glycoprotein (MDR1, ABCB1) and the drug-metabolizing enzymes cytochrome P450 (CYP) 2C8 and CYP3A5. Methods A total of 56 healthy volunteers ingested a single 0.25-mg dose of repaglinide. Plasma repaglinide and blood glucose concentrations were measured for up to 7 hours. All subjects were genotyped for the −11187G>A and 521T>C SNPs in SLCO1B1 and the 3435C>T and 2677G>T/A SNPs in ABCB1, as well as for the CYP2C8*3 (416G>A, 1196A>G), CYP2C8*4 (792C>G), and CYP3A5*3 (6986A>G) alleles. Results The area under the plasma concentration-time curve from time 0 to infinity [AUC(0-∞)] and peak concentration in plasma (Cmax) of repaglinide varied 16.9-fold and 10.7-fold, respectively, between individual subjects. Multiple regression analyses indicated that the SLCO1B1 521T>C SNP and the CYP2C8*3 allele were independent predictors of the AUC(0-∞) and Cmax of repaglinide (adjusted multiple R2 = 45% and 36%, respectively). In subjects with the SLCO1B1 521CC genotype, the AUC(0-∞) of repaglinide was 107% and 188% higher, respectively, than in subjects with the SLCO1B1 521TC or 521TT (reference) genotype (P < .0001). In subjects with the CYP2C8*1/*3 genotype, the AUC(0-∞) and Cmax of repaglinide were 48% and 44% lower, respectively, than in those with the CYP2C8*1/*1 genotype (P < .05). The pharmacokinetics of repaglinide was not associated with the studied ABCB1 SNPs or the CYP3A5*3 allele. The elimination half-life of repaglinide was not associated with any SNP. Only the SLCO1B1 −11187GA genotype was significantly associated with an enhanced effect of repaglinide on blood glucose. Conclusions Genetic polymorphism in SLCO1B1 is a major determinant of interindividual variability in the pharmacokinetics of repaglinide. The effect of SLCO1B1 polymorphism on the pharmacokinetics of repaglinide may be clinically important. Clinical Pharmacology & Therapeutics (2005) 77, 468–478; doi: 10.1016/j.clpt.2005.01.018

Objective: The aim of this study was to address the presently controversial question of whether cytochrome P450 (CYP) 3A5 polymorphism is associated with hypertension. Method: We studied 373 elderly (age ≥75 years) Finnish (Caucasian) patients from the ongoing DEBATE (Drugs and Evidence Based Medicine in the Elderly) trial. The patients were classified into those with a history of hypertension (n = 229) and those without a history of hypertension (n = 144) on the basis of a detailed questionnaire on each patient’s medical history and an interview. The patients were genotyped for the CYP3A5 6986A/G single nucleotide polymorphism (SNP) [CYP3A5*1/*3 alleles]. Results: The proportion of individuals with the CYP3A5*1/*3 genotype, i.e. CYP3A5 expressors, was significantly higher among patients with a diagnosis of hypertension than among patients without (18.3% vs 9.0%, p = 0.016). The corresponding odds ratio was 2.26 (95% CI 1.17, 4.38). The allele and genotype frequencies for the two control SNPs, ABCB1 (MDR1) 3435C/T and SLCO1B1 521T/C, did not differ between the two groups. Conclusion: This work lends support to the theory that the polymorphic CYP3A5 enzyme may be involved in regulation of blood pressure. The possible role of CYP3A5 as a genetic contributor to hypertension susceptibility warrants further study.

Background Individuals with the cytochrome P450 (CYP) 2D6 intermediate metabolizer (IM) phenotype have low residual enzyme activity and compose about 10% to 15% of white populations. Their identification is clinically relevant but remains unsatisfactory because of incomplete characterization of the major allele involved, termed CYP2D6*41 (−1584C, R296C, S486T). Methods To search for novel mutations, resequencing of the entire CYP2D6 gene was performed in selected individuals. Genotype-phenotype correlation analysis was done in a population sample of 308 white subjects phenotyped with sparteine and previously genotyped for all major alleles. Results A total of 16 novel polymorphic positions were identified, of which 7 were located within 2.4 kilobases of previously uncharacterized 2D7–2D6 intergenic sequence and 9 were located within intronic regions. The novel mutation 2988G>A in intron 6 appeared to be specifically associated with the IM phenotype. Further analysis in the population sample demonstrated that 2988G>A was strongly linked to allele *41 but not to any other alleles including *1, *2, *2×N, *4, *6, *7, *8, *9, *10, and *35. The overall frequency of the novel polymorphism was 8.4% in the normal white population. Compared with conventionally determined *41, 2988G>A was shown to have improved predictivity for the IM phenotype. With 2988G>A being taken into account, alleles *1, *2, and *35 (−1584G, V11M, R296C, S486T) were found to be phenotypically equivalent. Conclusions CYP2D6 genotyping can be considerably simplified by using 2988G>A as a marker for *41 and by omitting genotyping for the functionally equivalent alleles *2 and *35. Clinical Pharmacology & Therapeutics (2004) 76, 128–138; doi: 10.1016/j.clpt.2004.04.009

Objectives Our objectives were to determine the content of cytochrome P450 (CYP) 3A4, CYP3A5, and P-glycoprotein and to measure CYP3A4-dependent catalytic activity in paired human small intestinal and liver specimens. Methods Samples of duodenum or proximal jejunum and liver wedge biopsy specimens were obtained from 15 patients undergoing a gastrointestinal operation. Enterocytes were isolated from the intestinal samples. The contents of CYP3A4, CYP3A5, and P-glycoprotein and CYP3A4-mediated catalytic activities were determined in homogenized enterocyte and liver samples. Results The CYP3A4 protein content was about 3 times (P < .01) and the P-glycoprotein content about 7 times (P < .0001) higher in the enterocyte homogenates than in the liver homogenates. CYP3A5 protein was detected in all samples, but the levels were too low in most cases to allow quantification. The 2 cases with a quantifiable hepatic CYP3A5 content had the CYP3A5*1/*3 genotype; all other cases were homozygous for the CYP3A5*3 allele. No intraindividual correlations between the intestine and liver with respect to CYP3A4 content, P-glycoprotein content, or the measured catalytic activities were present. Values for the maximum rate of metabolism (Vmax) of verapamil N-dealkylation (formation of D-617) and N-demethylation (formation of norverapamil) activities correlated with the CYP3A4 protein content in both organs. Conclusions This work demonstrated a much higher content of both CYP3A4 protein and P-glycoprotein in enterocytes isolated from human duodenal or jejunal mucosa than in paired specimens of liver tissue. These results lend support to the view that biotransformation in the gut wall substantially contributes to the overall first-pass metabolism of many CYP3A4 substrates. Furthermore, the high content of P-glycoprotein on the apical surface of enterocytes supports the theory that this efflux transporter may act in concert with CYP3A4 to limit oral drug bioavailability. Finally, these results indicate that neither CYP3A4 nor MDR1 (P-glycoprotein) is coordinately regulated in the liver and intestine. Clinical Pharmacology & Therapeutics (2004) 75, 172–183; doi: 10.1016/j.clpt.2003.10.008

Trotz vergleichbarer Plasmakonzentrationen ist die Wirkung der antiviralen Therapie der HIV-Infektion im Hinblick auf die Suppression der Virusreplikation zwischen den einzelnen Patienten sehr variabel. Bisher erklarte man solche Unterschiede unter anderem durch Mutationen des HI-Virus,die zur Resistenzentwicklung fuhren. Es ist allerdings auch vorstellbar, dass Wirtsfaktoren, die den Arzneimittelmetabolismus (Mutationen in Genen von Cytochrom-P450-Enzymen) sowie den Transfer des Wirkstoffes aus dem Blut an den Wirkort kontrollieren, fur diese Variabilitat mitverantwortlich sind. Einige der in der antiretroviralen Therapie eingesetzten Arzneimittel werden von arzneimittelmetabolisierenden Enzymen verstoffwechselt, die einem genetischen Polymorphismus unterliegen (z. B. Nelfinavir durch CYP2D6). Fur die HIV-Proteaseinhibitoren ist auch bekannt, dass sie nicht nur einem ausgepragten Metabolismus unterliegen, sondern auch sehr gute Substrate des Efflux-Transporters P-Glykoprotein sind, der in Dunndarm, Leber, Niere sowie in Lymphozyten exprimiert wird. So konnte kurzlich gezeigt werden, dass Mutationen im MDR1-Gen mit einer reduzierten P-Glykoproteinexpression und -funktion beim Menschen assoziiert sind. Erste Daten belegen,dass das Vorhandensein dieser Mutation den Erfolg der Therapie mit HIV-Proteaseinhibitoren beeinflusst. Zudem sind nukleosidische Reverse-Transkriptase-Inhibitoren Substrate von Transporterproteinen (z. B. MRP4), wobei in vitro die MRP4-Uberexpression die antivirale Aktivitat dieser Arzneimittel erheblich einschrankt. Diese Ubersichtsarbeit stellt anhand ausgewahlter Beispiele die mogliche Relevanz genetischer Polymorphismen in arzneimittelmetabolisierenden Enzymen sowie von Transporterproteinen fur eine masgeschneiderte HIV-Therapie dar.

This study investigates the impact of dose escalation and of doxorubicin and cyclophosphamide coadministration on the pharmacokinetics of etoposide (ETO). Pharmacokinetics of ETO were analyzed in seven patients with breast cancer receiving 3-4 cycles of conventional-dose (CD) and one final course of high-dose (HD) chemotherapy including ETO (450 mg/m(2) and 2100 mg/m(2), respectively, fractionated over 3 consecutive days). ETO was given as monoinfusion apart from day 1 of CD, where cyclophosphamide and doxorubicin were coadministered. Plasma samples obtained on day 1 and day 2 of CD- and HD-therapy, respectively, were analyzed for ETO by HPLC. Data from a total of 25 cycles of CD- and 7 cycles of HD-therapy are given as means +/- SD for CD-day 1, CD-day 2, HD-day 1 and HD-day 2, respectively. Following administration of 210+/-29, 278+/-41, 1143+/-79 and 1143+/-79 mg ETO, the AUC (0-24 h, normalized to 150 mg/m(2)) was 123+/-23, 113+/-22, 92+/-11 and 100+/-22 microgxh/ml. The AUC and CL of single-agent ETO were not significantly different between CD (day 2) and both days of HD ETO. However, we observed a modest but significant difference for AUC and CL between day 1 of CD (coadministration of doxorubicin and cyclophosphamide) and day 2 of CD (ETO monoinfusion), the AUC and CL being 9% higher (see above) and 10% lower (21.1 vs. 23.3 ml/minxm(2)) ( P

Background and Aims The contribution of the gastrointestinal tract in comparison with the liver for the low and variable bioavailability of orally administered drugs is still poorly understood. Here we report on a new intestinal perfusion technique for the direct assessment of absorption, metabolism, and transport of drugs by the intestinal wall. Methods In 6 healthy volunteers a multilumen perfusion catheter was used to generate a 20-cm isolated jejunal segment that was perfused with 80 mg verapamil. Simultaneously, 5 mg [2H7]verapamil was given intravenously. Blood, perfusate, and bile samples were analyzed for parent verapamil and its major metabolites. Results The mean fraction of the verapamil dose absorbed from the 20-cm segment was 0.76 but substantial interindividual variability (0.51–0.96) was shown. Bioavailability was low (19.3%). The intestinal wall contributed to the same extent as the liver to extensive first-pass metabolism (mean extraction ratio, 0.49 versus 0.48). Substantial transport of verapamil metabolites from the systemic circulation via the enterocytes into the intestinal lumen was observed. Compared with biliary excretion, intestinal secretion into a 20-cm jejunal segment contributed to drug elimination to a similar extent. Conclusion First-pass metabolism by the intestinal wall is extensive and contributes to the same extent as the liver to low bioavailability of some drugs such as verapamil. Moreover, intestinal secretion is as important as biliary excretion for the elimination of metabolites. Clinical Pharmacology & Therapeutics (2001) 70, 217–227; doi: 10.1067/mcp.2001.117937

Background Racemic (R/S)-verapamil is widely used in the management of chronic atrial fibrillation. The negative dromotropic effect is mainly mediated by the S-enantiomer, which is preferentially metabolized. Previous studies report an accumulation of R/S-verapamil during long-term oral treatment of patients with chronic atrial fibrillation. However, the specific disposition of S-verapamil and the pharmacologic effects were not assessed. Therefore uncertainties about the need for dose adjustments remain. Methods Using stable isotope technology and a stereospecific assay, we compared the pharmacokinetics and pharmacodynamics of intravenous (10 mg of d7-R/S-verapamil) and oral (240 mg of slow release (SR) d0-R/S-verapamil) R–verapamil and S-verapamil after the first dose (day 1) and after 3 weeks (day 21) of continuous oral therapy in 8 patients with long-term atrial fibrillation. On both study days, serum samples were obtained for the analysis of d7- and d0-R-verapamil and S-verapamil. Heart rate (HR) was monitored with electrocardiography (with each blood sample) and Holter electrocardiography (before the study, on day 1, and on day 21). Results Compared with day 1, clearance of oral R-verapamil and S-verapamil was significantly reduced on day 21 (1007 ± 380 versus 651 ± 253 mL/min [–35%] and 5481 ± 2731 versus 2855 ± 1097 mL/min [–48%], respectively; P < .05), whereas only a moderate decrease was observed for intravenous R-verapamil and S-verapamil (–23% and −14%, respectively, not significant). Mean HR (89 ± 11 bpm before verapamil) was effectively reduced, with the same effects on day 1 (68 ± 8 bpm) and day 21 (68 ± 8 bpm). Compared with day 1, the HR reduction per ng/mL of S-verapamil (calculated by the area under the curve [from 0–24 hours] ratio of HR reduction and S-verapamil concentration) was significantly lower on day 21 (0.7 ± 0.4 versus 1.2 ± 0.7 [bpm] · [ng/mL]−1, for day 21 versus day 1; P < .01). Conclusions In patients with chronic atrial fibrillation, clearance of oral, but not intravenous, S-verapamil and R-verapamil is significantly reduced with multiple doses compared with a single dose, thereby indicating predominant impairment of prehepatic rather than hepatic metabolism as the underlying mechanism. However, this kinetic change is clinically compensated by a decrease in the responsiveness to S-verapamil observed with regular dosing. The data suggest that despite accumulation of the drug individual verapamil doses can be maintained during long-term oral rate control therapy. Clinical Pharmacology & Therapeutics (2001) 69, 324–332; doi: 10.1067/mcp.2001.115125

Microsomal epoxide hydrolase is a biotransformation enzyme which is involved in the hydrolysis of various epoxides and epoxide intermediates. In the present study, its distribution was investigated in both normal human tissues and human tumours of different histogenetic origin using immunohistochemical techniques. In normal tissue, epithelial cells were more often and more intensely immunostained than mesenchymal cells. The main epithelial cell types expressing microsomal epoxide hydrolase were hepatocytes, acinus cells of the pancreas, and cells of salivary and adrenal glands. Immunostained cells of mesenchymal origin included monocytes, fibrocytes, fibroblasts, vessel endothelium, muscle cells, and cells of the reproductive system. Three patterns of expression were observed in tumour tissues: (1) moderate or strong in hepatocellular carcinomas, tumours of the adrenal gland, and theca-fibromas of the ovary; (2) inhomogeneous staining pattern of variable intensity in breast cancer, lung cancer, colorectal carcinomas, carcinoid tumours, and some tumours of mesenchymal origin; and (3) no expression in malignant melanomas, malignant lymphomas, and renal carcinomas. These data indicate that microsomal epoxide hydrolase expression is not restricted to tissue of any particular histogenetic origin. Nonetheless, immunohistochemical identification of microsomal epoxide hydrolase may be helpful in some well-defined histological settings, for example, confirmation of hepatocellular carcinoma.

In addition to genetically variable metabolic enzymes such as Cyp p450 proteins, blood and tissue levels of many drugs are influenced by controlled transport across compartmental boundaries. Major determinants in these transport processes are ATP-dependent efflux pumps such as P-glycoprotein and related proteins (eg MRPs), which can influence the bioavailability and CNS concentrations, as well as disposition of drugs. In addition to its recognized role in the development of multiple chemotherapy resistances, experimental evidence for the relevant influence of the MDR1 gene encoded P-glycoprotein, on the pharmacology of many other drugs has been gathered by the analyses of knockout mice, as well as in clinical studies. Recently, functional genetic polymorphisms in the MDR1 gene have been identified which influence the distribution and bioavailability of PGP substrates.

The debrisoquine/sparteine or CYP2D6 genetic polymorphism of drug oxidation is a common cause for interindividual variability in drug response. We recently identified a mutant allele, designated CYP2D6-E or CYP2D6*7, which is associated with the poor metabolizer phenotype and occurs in Caucasian populations with a frequency of about 1%. In contrast to other loss-of-function alleles, a full length protein with a single amino acid substitution. His324Pro, is encoded by the CYP2D6*7 allele. To functionally analyze this mutant protein form of CYP2D6, recombinant baculoviruses were constructed to express the CYP2D6 cDNA. Up to 0.33 nmol of spectrally detected P450/mg of cell protein were produced in Spodoptera frugiperda cells, whereas Trichoplusia ni 5B1-4 cells reproducibly produced 0.8 nmol/mg (4% of total cell protein). Insect cell membranes were functionally characterized with cumene hydroperoxide or after reconstitution with purified rat NADPH:cytochrome P450 reductase. Km values for the substrates bufuralol and sparteine and other enzymatic properties were almost identical to those of human liver microsomes. The H324P mutation was introduced into the cDNA by site-directed mutagenesis and recombinant baculovirus was obtained. Expression under a variety of conditions demonstrated that mutant protein amounts comparable to the wild-type enzyme were produced. However, no spectrally detectable P450 was formed and no catalytic activity was detected. Furthermore, in contrast to the wild-type protein the mutant protein was almost exclusively located in a detergent-insoluble insect cell fraction. These results demonstrate that the H324P mutation is responsible for the in vivo poor metabolizer phenotype associated with the CYP2D6*7 allele by preventing normal protein folding and heme incorporation.

In the pathogenesis of isoniazid-induced hepatic injury, cytochrome P450-dependent metabolic activation of the metabolite, acetylhydrazine (AcHz), is the crucial step. Exhalation of [14C]-carbon dioxide has previously been used to quantify indirectly this pathway. In contrast, according to the current concept of AcHz bioactivation, molecular nitrogen is produced directly, but has not yet been identified. Here, we measured [15N]-nitrogen and 14CO2 exhalation, after the administration of [15N2]-[14C]-AcHz, in rats. Laser magnetic resonance (LMR) spectroscopy, a new sensitive and specific technique for the measurement of 15N and 14N in gas samples, was used. To demonstrate the involvement of cytochrome P450, rats were treated with phenobarbital (PB) or PB+cobalt(II) chloride (CoCl2) (n = 3 in each group). Time-dependent 15N2 exhalation differed significantly between treatment groups (p0.001). At 240 min, cumulative exhalation of 15N was 1.92 +/- 0.43% (mean +/- SE) of the dose in the control group, 2.53 +/- 0.23% in the PB group, and 1.00 +/- 0.15% in the PB+CoCl2 group (p0.05 compared to controls, p0.01 compared to PB). Cumulative exhalation of 14CO2 in 24 h ranged from 15.1 to 21.9%, with no significant difference between treatment groups. In conclusion, N2 is a metabolite of AcHz. N2 formation reflects the cytochrome P450-mediated activation of AcHz and can be used as an index of this pathway. Generally, LMR spectroscopy is valuable for monitoring any N2-liberating process in vivo.

Objectives The cardiovascular and biochemical effects of R- and S-nitrendipine were studied in six healthy subjects in a single-blind placebo-controlled study. Methods After received oral doses of placebo, 20 mg R-, 80 mg R- (n = 5), 20 mg S-, and 20 mg racemic nitrendipine, heart rate, systolic, diastolic, and mean arterial blood pressure, leg blood flow, peripheral vascular resistance, plasma renin activity, norepinephrine, epinephrine, dopamine, and aldosterone plasma levels were measured before and up to 3 hours after administration. Results Neither placebo nor 20 or 80 mg R-nitrendipine caused significant changes of cardiovascular and biochemical parameters. After 20 mg S-nitrendipine and 20 mg racemic nitrendipine, significant changes in diastolic blood pressure (−9.1/−7.4 mm Hg), heart rate (+21.9/+17.3 beats/min), leg blood flow (+6.8 ml · min−1 · gm tissue−1), peripheral vascular resistance (−16.9 mm Hg · min · gm tissue · ml−1), norepinephrine (+476/+281 ng · L−1), and plasma renin activity (+9.5/+3.6 ng · ml−1 · hr−1) were observed. The changes in cardiovascular and biochemical parameters were closely related to the serum S-nitrendipine concentrations. Conclusions It can be concluded that, after administration of the racemate, the S-enantiomer is responsible for the cardiovascular and biochemical effects observed and that S-nitrendipine is at least an order of magnitude more potent than the R-enantiomer. Clinical Pharmacology & Therapeutics (1995) 57, 52–61; doi

A novel TPMT missense mutation associated with TPMT deficiency in a 5-year-old boy with ALL

We studied the influence of cytochrome P450 2D6 (CYP2D6) on the steady-state disposition kinetics and the electrocardiographic effects of flecainide at two doses and during combination with amiodarone. Seven extensive and five poor metabolizers of dextromethorphan were studied during a three-period crossover study. All subjects received 50 mg flecainide every 12 hours, alone or together with 200 mg amiodarone every 12 hours, and 100 mg flecainide every 12 hours for 5 days. Mean steady-state plasma concentration of flecainide and QRS change from predrug value did not differ significantly among extensive and poor metabolizer subjects during each study period. Except for a shortened elimination half-life and nonlinear kinetics in extensive metabolizer subjects, phenotype had no significant influence on flecainide pharmacokinetics. Combination with amiodarone resulted in an increase in mean flecainide plasma concentration and effect in subjects with both phenotypes. Our findings indicate that CYP2D6 phenotype predicts flecainide nonlinear kinetics and flecainide half-life but has no influence on electrocardiographic effects during repeated administration of flecainide or on the extent of the amiodaroneflecainide interaction. Clinical Pharmacology and Therapeutics (1994) 55, 256–269; doi:10.1038/clpt.1994.26

Modulators for the reversal of multidrug resistance such as R-verapamil and B8509-035, a dihydropyridine, effectively overcome multidrug resistance in vitro and are currently undergoing clinical trial. One problem with their use is the application protocol; the question as to whether they should be given by continuous administration or in sequential doses in combination with the cytotoxic drugs has to be addressed. Therefore, we examined the influence of the exposure time and the sequence of modulator administration on the active transport of the fluorescent dye rhodamine 123 (R123), a substrate for the P-glycoprotein, in the resistant lymphoblastoid cell line VCR1000 and the parental nonresistant cell line CCRF-CEM. Our results demonstrate the importance of coadministration of R-verapamil and the cytotoxic agent for the modulation of multidrug resistance, whereas the exposure sequence does not seem to be such an essential parameter in the case of B8509-035. This observation should be considered for the further design of clinical studies.

Both HPLC separation with fluorometric detection and 13C NMR spectrometry were used to quantify carbocysteine (CMC) and its major metabolites directly in the urine of thirteen human subjects following oral administration of the isotopomer S-carboxy-[13C]methyl-l-cysteine (13C-CMC). Comparison of the data obtained by these two independent analytical methods provides unequivocal evidence that the excretion of significant amounts of previously described cysteinyl sulphoxide metabolites cannot be verified. Instead, the identification and reproducible quantitation of renally eliminated CMC and its major non-cysteinyl metabolites, thiodiglycolic acid and thiodiglycolic acid sulphoxide, has been accomplished. It has to be concluded from this study, therefore, that the proposed genetically determined polymorphism of CMC sulphoxidation does not exist.

Most drugs are metabolized by CYP3A enzymes, and variations in expression levels of these enzymes are believed to determine whether patients will have a positive or adverse drug response. Little is known about the mechanisms that underlie inter-individual differences in CYP3A expression, but the mapping of human genome sequence variations will facilitate the search for answers.

The mammalian cytochrome P450-dependent monooxygenase system is involved in the metabolism of drugs and chemical carcinogens. The role of these enzymes in toxicological response is exemplified by an autosomal recessive polymorphism at the cytochrome P450 CYP2D6 debrisoquine hydroxylase locus which results in the severely compromised metabolism of at least 25 drugs, and which in some cases can lead to life-threatening side-effects. In addition, this polymorphism, which affects 8-10% of the caucasian population, has been associated with altered susceptibility to lung and bladder cancer. Here we report the identification of the primary mutation responsible for this metabolic defect and the development of a simple DNA-based genetic assay to allow both the identification of most individuals at risk of drug side-effects and clarification of the conflicting reports on the association of this polymorphism with cancer susceptibility.

The haemodynamic effects of an i.v. infusion of 2 mg nitrendipine have been studied in six healthy volunteers. Nitrendipine significantly decreased the systolic (-8.3%) diastolic (-19.9%) and mean arterial (-11.6%) blood pressures and the peripheral vascular resistance (-57.8%), and significantly increased leg blood flow (+128%). Stroke volume did not change. Due to the increase in heart rate (+28.5%), the cardiac output (2.8.2%) rose significantly. The haemodynamic effects were closely related to the serum nitrendipine concentration. The sigmoidal Emax-model was appropriate to describe the data. Pronounced interindividual differences in the serum nitrendipine concentrations required to elicit 50% of the maximum haemodynamic effect (EC50) were observed. The EC50 for the increase in leg blood flow ranged from 2.9 to 30.9 ng/ml and for the reduction in peripheral vascular resistance from 2.1 to 25.7 ng/ml. Interindividual differences in EC50 values were less pronounced if based on unbound serum nitrendipine levels. The fraction of nitrendipine not bound to serum proteins showed a three-fold difference between subjects, with free fractions ranging from 0.011 to 0.036. The unbound EC50 values for the increase in leg blood flow varied between 0.06 and 0.44 ng/ml and for the reduction in peripheral vascular resistance from 0.07 to 0.35 ng/ml. Based on the serum concentrations associated with comparable haemodynamic effects nitrendipine was at least three-times more potent than nifedipine.

The constitutive androstane receptor (CAR) plays a key role in the control of drug metabolism and transport by mediating the phenobarbital-type induction of many phase I and II drug metabolizing enzymes and drug transporters. We identified transcripts generated by four different alternative splicing events in the human CAR gene. Two of the corresponding ligand binding domain isoforms demonstrated novel functional properties: First, CAR(SV3), which is encoded by a transcript containing an lengthened exon 7, differentially transactivated target gene promoters. Second, CAR(SV2), which results from the use of an alternative 3' splice site lengthening exon 8, showed ligand-dependent instead of constitutive interaction with coactivators. Furthermore, alternatively spliced transcripts demonstrated a tissue-specific expression pattern. In most tissues, only transcripts generated by alternative splicing within exon 9 were expressed. The encoded variant demonstrated a loss-of-function phenotype. Correct splicing of exon 8 to exon 9 is restricted to only a few tissues, among them liver and small intestine for which CAR function has been demonstrated, and is associated with the induction of CAR expression during differentiation of intestinal cells. Due to their specific activities, CAR variant proteins SV2 and SV3 may modulate the activity of reference CAR(SV1). Furthermore, we propose that transcriptional activation and regulation of splicing of exon 9 may be coupled to ensure appropriate tissue- and differentiation state-specific expression of transcripts encoding functional CAR protein. Altogether, alternative splicing seems to be of utmost importance for the regulation of CAR expression and function.

Following i. v. administration, the plasma concentration-time curve of verapamil could best be described by either a mono- or biexponential equation. Total plasma clearance (1.26 l/min) approached liver blood flow (1.5 l/min), so it can be concluded that its clearance is liver blood flow-dependent. Although absorption was almost complete after oral administration, absolute bioavailability (20%) was low, due to extensive hepatic first-pass metabolism. The approach using stable isotope-labelled and unlabelled drug permits simultaneous administration by the intravascular and extravascular routes, thus allowing determination of absolute bioavailability in a single experiment.

Sparteine, an antiarrhythmic and oxytocic drug, is metabolised by N1-oxidation. The sparteine-N1-oxide rearranges with loss of water to 2- and 5-dehydrosparteine. 18 (i.e., 5%) out of 360 subjects were unable to metabolise the drug. These persons, who were designated as nonmetabolisers, excreted almost 100% of the administered dose in urine as unchanged drug. The defective metabolism of sparteine was found to have a genetic basis. Sparteine-N1-oxidation appears to be determined by two allelic genes at a single locus where nonmetabolisers are homozygous for an autosomal recessive gene.

Sparteine metabolism was studied in human liver microsomes from nine extensive ([EM] urinary metabolic ratio [MR] less than 20) and four poor metabolizers ([PM] MR greater than 20). The formation of 2- and 5-dehydrospartein displayed monophasic Michaelis-Menten kinetics. In livers from PMs the formation of the major sparteine metabolite 2-dehydrosparteine was characterized by more than a thirtyfold increase in Michaelis-Menten constant (Km)(1880 +/- 1044 mumol/L) as compared with EM subjects with an MR less than 1 (58.3 +/- 38.8 mumol/L). EM subjects with an MR greater than 3 who may constitute heterozygous metabolizers showed Km values in between (658 +/- 301 mumol/L). There was no difference in maximum rate of metabolism (Vmax) of 2-dehydrosparteine formation between EM and PM subjects (101 +/- 39 vs. 86 +/- 52 pmol/min/mg). The formation of 5-dehydrosparteine exhibited a Km of 100 +/- 123 mumol/L similar to the Km of 2-dehydrosparteine formation. The urinary MR correlated positively with the Km for 2-dehydrosparteine formation. The intrinsic clearance for 2-dehydrosparteine showed a highly significant negative correlation with the MR. The pronounced differences in Km together with the significant correlation between Km and MR with no marked differences in Vmax between phenotypes suggest that the impaired oxidation capacity in PM subjects is more likely the result of a P-450 isozyme with altered catalytic properties rather than a decreased amount of enzyme.

Theophylline plasma clearance (Clp) and clearance to its metabolites (Clm), as well as antipyrine saliva clearance (Clsal) and its Clm were compared in a crossover study in 25 healthy subjects. They were selected with regard to smoking status (nine smokers, 16 nonsmokers) and oxidation phenotype of debrisoquine and sparteine (six poor metabolizers [PMs] and 19 extensive metaboUzers [EMs]). Clm of theophylline (1,3-dimethyluric acid, 1-methyluric acid, and 3-methylxanthine) correlated (r ≥ 0.92) to each other and to total theophylline Clp (r ≥ 0.97). Smokers had higher Clm to all metabolites, particularly by the N-demethylation pathways. After correction for the effect of smoking, there was no difference between EMs and PMs with regard to theophylline Clp or Clm. Antipyrine clearances by EMs and PMs (Clsal and Clm of 4-OH-antipyrine, 3-OH-methylantipyrine, or norantipyrine) also did not differ. Antipyrine Clsal and Clm correlated to theophylline Clp (r between 0.50 and 0.69). It is concluded that theophylline metabolism (N-demethylations and C-oxidation) is not under the same genetic control as sparteine and debrisoquine oxidations, and that there may be a partial overlap in factors that regulate the metabolism of theophylline and antipyrine. Clinical Pharmacology and Therapeutics (1984) 35, 815–821; doi:10.1038/clpt.1984.118

Carbamazepine and its epoxide in plasma were measured by liquid chromatography in 25 patients treated with a mean dose of carbamazepine of 12.5 +/- 3.3 mg/kg body weight. The mean concentrations of parent drug and metabolite were 5.4 +/- 2.5 mug/ml and 1.10 +/- 0.42 mug/ml, respectively. A singificant correlation was found between the plasma concentrations of the two compounds (r = 0.64; p less than 0.001), but marked interindividual variation existed in the ratio of carbamazepine to carbamazepine to epoxide. Based on simultaneous measurements in plasma and cerebrospinal fluid, the unbound fraction of carbamazepine in plasma was of the order of 20% as compared to 45% for the epoxide. Thirteen ambulant patients suffering from partial epilepsy with complex symptomatology, who were already being treated with phenytoin in optimal doses (plasma level 14-20 mug/ml) were also given carbamazepine. At plasma levels of the latter of about 5 mug/ml there was no further reduction in the frequency of partial or generalized epileptic seizures. In five patients the dose was increased to produce plasma concentrations of 7 - 8 mug/ml. There was still no improvement and side-effects were seen in three patients.

In artificially ventilated patients, intratracheal aminoglycoside administration is used as a form of prophylaxis of broncho-pulmonary infections. In artificially ventilated patients with multiple organ failure, serum gentamicin concentrations were measured dependent on renal function after endotracheal administration. A standard commercial ampule of 40 mg gentamicin in a 1 ml solution was injected in undiluted form, intratracheally through the tubus , every 6 h. In the patients without renal failure, values over 1 microgram/ml were only found in certain individual cases and reached a maximum of 1.5 micrograms/ml. In patients with renal failure even after prolonged application, the average serum concentrations were between 2 and 3.5 micrograms/ml. In a very few cases, however, levels of up to 10.5 micrograms/ml were measured. The daily serum pattern revealed a distinct dependence on the administration; 1 h after administration there was an increase in the serum concentrations which decreased to the initial levels after 6 h. When patients with renal impairment are given an aminoglycoside intratracheally, serum levels of up to 10.5 micrograms/ml may be reached and thus additional systemic aminoglycoside therapy should be avoided.

The calcium antagonists diltiazem, nifedipine and verapamil are widely used in the treatment of coronary heart disease, arterial hypertension, certain supraventricular tachyarrhythmias and obstructive hypertrophic cardiomyopathy. During recent years their pharmacokinetic properties and metabolism have been studied in more detail. Although these 3 calcium antagonists exhibit great diversity in chemical structure, they exhibit common pharmacokinetic properties. These drugs are extensively metabolised and only traces of unchanged drugs are eliminated in urine. Their systemic plasma clearances are high and dependent on liver blood flow. Therefore, their bioavailabilities (diltiazem 40 to 50%; nifedipine 40 to 50%; verapamil 10 to 30%) are low despite almost complete absorption following oral administration. During long term treatment, oral clearance decreases and bioavailability increases due to saturation of hepatic first-pass metabolism. Pronounced intra- and inter-individual variations in clearance and bioavailability are observed. In patients with liver cirrhosis the various pharmacokinetic parameters are grossly altered. Clearance decreases, elimination half-life is substantially prolonged, and bioavailability more than doubles. In addition, the volume of distribution increases. Whereas renal disease has no impact on the pharmacokinetics of diltiazem and verapamil, elimination half-life of nifedipine increases in relation to the degree of renal impairment due to an increase in volume of distribution. Systemic clearance, however, remains unchanged. The data so far available indicate that the plasma concentrations of these drugs correlate with both their electrophysiological and haemodynamic effects. However, no effective therapeutic plasma concentration range has been firmly established. As reliable clinical end-points are available for dose titration of calcium antagonists, it is doubtful whether therapeutic drug monitoring will be of great value. Calcium antagonists are often administered in combination with a variety of other drugs. Thus, the potential for both pharmacodynamic and pharmacokinetic drug interaction exists. The interaction between digoxin and these drugs is of clinical importance. Verapamil and diltiazem cause a significant increase in plasma digoxin concentrations. In contrast, nifedipine does not lead to a significant increase in the plasma digoxin concentration. The mechanism responsible for this interaction is inhibition of both renal and non-renal digoxin clearance.(ABSTRACT TRUNCATED AT 400 WORDS)

Sparteine is metabolized by N1-oxidation, which in some subjects is defective. The defect has a pronounced effect on the kinetics of the drug. In nonmetabolisers elimination of sparteine proceeds entirely via renal excretion by a capacity-limited process, 99,9% of the dose being excreted as unchanged drug. In metabolisers the drug is mainly eliminated by metabolic degradation. Pronounced differences in beta-phase half-life and total plasma clearance were observed between metabolisers (156 min; 535 ml . min-1) and nonmetabolisers (409 min; 180 ml . min-1).

For drugs with a high hepatic clearance, bioavailability is low due to the so-called "first pass effect". Prediction of the bioavailability for these drugs has been only loosely tested. It is proposed that by plotting the reciprocal of bioavailability versus the oral clearance, a straight line with intercept of unity and slope of reciprocal of hepatic blood flow should ensue. For lignocaine and verapamil, this relationship was found to be strong and gave good predictability, whereas for propranolol this relationship was weak and gave poor predictability. The proposed method may be of value in determining whether the low bioavailability of a drug is due to hepatic first pass metabolism.

Thyroid dysfunction can influence the physiological disposition of drugs. Depending on the pharmacokinetic properties of the individual drug, changes in the rate of metabolism ranging from profound to moderate or negligible have been observed. Since renal function is also influenced by thyroid disease, changes in renal elimination of drugs which are excreted in the urine mainly as unchanged drugs have to be considered as another reason for altered drugs disposition in thyroid disease. In patients with thyrotoxicosis lower, and in patients with myxoedema, higher, digitalis plasma levels have been observed. The altered disposition of cardiac glycosides in thyroid dysfunction can be attributed to changes in renal elimination and metabolism. These findings may be the kinetic correlate for the clinical observation that larger than the usual dose of digitalis is required in thyrotoxic patients and lower in hypothyroid patients. Antipyrene half-lives are very much shortened during hyperthyroidism and prolonged appreciably during hypothyroidism. The alterations in the disposition of these drugs seen during thyroid dysfunction can be ascribed to changes in its rate of metabolism which is controlled by the levels of circulating thyroid hormones. N-demethylation of aminopyrine is depressed both in hyper- and hypothyroid patients as compared with euthyroid subjects. Changes in the half-life of this drug were observed only during hypothyroidism. The physiological disposition of the antithyroid drug propylthiouracil is not changed during thyrotoxicosis. A decrease in plasma half-life of methimazole is however, observed during hyperthyroidism, whereas in hypothyroid patients half-life is increased. The few data available so far do not allow general prediction of how thyroid disease could alter drug metabolism in man.

In order to determine whether the metabolism of the antiarrhythmic drug N-propylajmaline is under the same genetic control as sparteine metabolism, the pharmacokinetics of this antiarrhythmic drug were studied in a groups of six extensive and four poor metabolizers of sparteine. Pronounced differences in terminal half-life, total plasma clearance, metabolic clearance and urinary excretion of N-propylajmaline were observed between extensive and poor metabolizers. A close relationship between the total clearance and metabolic clearance of N-propylajmaline and sparteine could be demonstrated. Clinically available N-propylajmaline is a 55% to 45% mixture of the i- and n-diastereomers. The extensive metabolizers exhibited stereoselective metabolism; the i-diastereomer was preferentially metabolized. Poor metabolizers were characterized by a loss of this stereoselective metabolism. Five subjects were treated for 7 days with a daily N-propylajmaline dosage of either 60 mg or 20 mg. Since a close relationship between the clearance of N-propylajmaline and the metabolic ratio of sparteine had been observed after single dosing the metabolic ratio of sparteine was used to predict N-propylajmaline steady-state plasma concentrations during multiple dosing. Only in two extensive metabolizers with a metabolic ratio less than 0.4 predicted and observed, steady-state plasma concentrations were in good agreement. In the other three subjects observed steady-state plasma concentrations were appreciably higher than predicted. In these three subjects metabolic N-propylajmaline clearance decreased indicating saturation N-propylajmaline metabolism during multiple dosing. The data indicate that N-propylajmaline metabolism is subject to a genetic polymorphism controlled by the sparteine/debrisoquine gene locus.

Bei 95 Patienten mit aktiver Tbc untersuchten wir prospektive den Einflus des Acetyliererphanotyps auf die hepatotoxische Nebenwirkung der tuberkulostatischen Kombination Isoniazid (INH) 10 mg/kg, Rifampicin (RMP) 10 mg/kg, Myambutol (EMB) 25 mg/kg. Neben einer viel hoheren Inzidenz der Isoniazidhepatitis (SGOT, SGPT > 200 U/l) von 12,6% der Behandelten — verglichen zur Haufigkeit der Isoniazidhepatitis wahrend Chemoprophylaxe mit Isoniazid als Monotherapie von 0,5–1% (IUAT 1969, U.S.P.H.S. 1971) — stellten wir bei der Kombinationstherapie ein signifikant hoheres Risiko schwerer INH-induzierter Leberschadigungen bei Langsamacetylierern fest (p 50 U/l, von 30 Schnellacetylierern dagegen nur 4 (=13,3%). Unter den 12 Patienten mit Isoniazidhepatitis befanden sichnur Langsamacetylierer. Frauen waren von der Isoniazidhepatitis haufiger betroffen als Manner (p < 0,05). Bei der Isoniazidhepatitis wurde entweder die Therapie vorubergehend abgesetzt oder als Zweierkombination RMP, EMB fortgesetzt. Bei leichterem Verlauf der Leberschadigung wurde die Therapie unverandert fortgesetzt. In allen Fallen normalisierten sich die Transaminasen innerhalb 2–4 Wochen. Die anschliesende Wiederaufnahme der Dreifachtherapie ohne Dosisreduktion fuhrte zu keinem erneuten Transaminasenanstieg. Das konstante Auftreten der INH-Hepatitis in der 2.–4. Woche (19±7 Tage) sowie die ohne Reaktion vertragene spatere Reexposition der vollen Dreifachtherapie sprechen fur eine zeitlich begrenzte Interaktion des Rifampicin mit dem Isoniazid-Metabolismus in der Anfangsphase der tuberkulostatischen Therapie.

The pharmacokinetics and metabolism of quinidine were investigated in extensive and poor metabolisers of sparteine. No differences in plasma clearance, terminal half life, volume of distribution or cumulative urinary excretion of quinidine, 3-hydroxyquinidine and quinidine-N-oxide were observed between phenotypes. Thus, it is unlikely that quinidine metabolism is controlled by the sparteine/debrisoquine gene locus.

The potential to inhibit drug metabolism of the new antifungal agent terbinafine has been studied using antipyrine (single oral dose of 10 mg/kg) as a probe drug. In a cross-over study in 8 healthy volunteers, antipyrine was administered prior to, during and after 8 days of oral terbinafine 125 mg b.d. Antipyrine, its major metabolites 4-hydroxyantipyrine (4-OH-AP), 3-hydroxymethylantipyrine (3-OH-CH3-AP) and norantipyrine (Nor-AP) were analyzed by specific HPLC assays in multiple plasma and urine samples. During all three parts of the study, the pharmacokinetics of antipyrine viz. t1/2 (11.7 h), total plasma (38.5 ml.h-1.kg-1) and renal clearance (1.6 ml.h-1.kg-1), and its clearance rates to metabolites (CLM), eg. CLM for 4-OH-AP (12.3 ml.h-1.kg-1), CLM for 3-OH-CH3-AP (4.2 ml.h-1.kg-1) and CLM for Nor-AP (6.7 ml.h-1.kg-1) did not differ from the control values. Thus, all the cytochrome P-450-dependent isozymes involved in the metabolism of antipyrine and many other drugs should not be affected by therapeutic doses of terbinafine.

Two subjects from each of the three groups of homozygous rapid, heterozygous, and homozygous non-metabolizers (N-oxidation) of sparteine received a single oral dose of debrisoquine. The urinary ratio of debrisoquine/4-hydroxy-debrisoquine, reflecting the individual's capacity to C-hydroxylate debrisoquine, was closely related to his phenotype for sparteine metabolism. This indicates that the two metabolic reactions are controlled by similar if not identical genetic factors.