Sequence diversity in CYP3A promoters and characterization of the genetic basis of polymorphic CYP3A5 expression.

Variation in the CYP3A enzymes, which act in drug metabolism, influences circulating steroid levels and responses to half of all oxidatively metabolized drugs. CYP3A activity is the sum activity of the family of CYP3A genes, including CYP3A5, which is polymorphically expressed at high levels in a minority of Americans of European descent and Europeans (hereafter collectively referred to as 'Caucasians'). Only people with at least one CYP3A5*1 allele express large amounts of CYP3A5. Our findings show that single-nucleotide polymorphisms (SNPs) in CYP3A5*3 and CYP3A5*6 that cause alternative splicing and protein truncation result in the absence of CYP3A5 from tissues of some people. CYP3A5 was more frequently expressed in livers of African Americans (60%) than in those of Caucasians (33%). Because CYP3A5 represents at least 50% of the total hepatic CYP3A content in people polymorphically expressing CYP3A5, CYP3A5 may be the most important genetic contributor to interindividual and interracial differences in CYP3A-dependent drug clearance and in responses to many medicines.

The role of GABAergic modulation in motor function related neuronal network activity.

At rest, the primary motor cortex (M1) exhibits spontaneous neuronal network oscillations in the beta (15-30 Hz) frequency range, mediated by inhibitory interneuron drive via GABA-A receptors. However, questions remain regarding the neuropharmacological basis of movement related oscillatory phenomena, such as movement related beta desynchronisation (MRBD), post-movement beta rebound (PMBR) and movement related gamma synchronisation (MRGS). To address this, we used magnetoencephalography (MEG) to study the movement related oscillatory changes in M1 cortex of eight healthy participants, following administration of the GABA-A modulator diazepam. Results demonstrate that, contrary to initial hypotheses, neither MRGS nor PMBR appear to be GABA-A dependent, whilst the MRBD is facilitated by increased GABAergic drive. These data demonstrate that while movement-related beta changes appear to be dependent upon spontaneous beta oscillations, they occur independently of one other. Crucially, MRBD is a GABA-A mediated process, offering a possible mechanism by which motor function may be modulated. However, in contrast, the transient increase in synchronous power observed in PMBR and MRGS appears to be generated by a non-GABA-A receptor mediated process; the elucidation of which may offer important insights into motor processes.

Association of genotypes of the CYP3A cluster with midazolam disposition in vivo.

The genes that encode for CYP3A4 and CYP3A5 are located in the same region (CYP3A cluster) on chromosome 7. Midazolam (MDZ) is a substrate for both CYP3A4 and CYP3A5. We hypothesize that MDZ disposition in vivo is associated with genotypes of the CYP3A cluster. A meta-analysis of the pharmacokinetic (PK) parameters from seven clinical trials was carried out, in which MDZ was administered both intravenously and orally. DNA samples were available from 116 patients. There were significant ethnic differences in the allelic frequencies of these four common single-nucleotide polymorphisms (SNPs) in the CYP3A cluster. Significant linkage disequilibrium was found between CYP3A5(*)3 and CYP3A4(*)1A in Caucasians, and between CYP3A5(*)1 and CYP3A4(*)1B in African Americans. There were no differences in MDZ disposition in vivo between different genotypes, haplotypes and diplotypes in the CYP3A cluster (P>0.05). No significant differences in MDZ PK parameters were observed between Caucasians and African Americans. Women had higher weight-corrected systemic and oral clearance than men, but dose-adjusted AUC and bioavailability differences were not observed between sexes. The clinical importance of elevated CYP3A activity in women remains to be determined. The r(GC)'s of MDZ PK parameters were between 0.3 and 13.6%. In conclusion, the meta-analysis of seven studies suggests that environmental factors explain the majority of CYP3A activity variation. Further studies are necessary to define the functional significance of SNPs in the CYP3A cluster and the effects of CYP3A genotypes on MDZ disposition in vivo.

Pharmacologically induced and stimulus evoked rhythmic neuronal oscillatory activity in the primary motor cortex in vitro.

Parkinson's disease (PD) is associated with enhanced synchronization of neuronal network activity in the beta (15-30 Hz) frequency band across several nuclei of the basal ganglia (BG). Deep brain stimulation of the subthalamic nucleus (STN) appears to reduce this pathological oscillation, thereby alleviating PD symptoms. However, direct stimulation of primary motor cortex (M1) has recently been shown to be effective in reducing symptoms in PD, suggesting a role for cortex in patterning pathological rhythms. Here, we examine the properties of M1 network oscillations in coronal slices taken from rat brain. Oscillations in the high beta frequency range (layer 5, 27.8+/-1.1 Hz, n=6) were elicited by co-application of the glutamate receptor agonist kainic acid (400 nM) and muscarinic receptor agonist carbachol (50 microM). Dual extracellular recordings, local application of tetrodotoxin and recordings in M1 micro-sections indicate that the activity originates within deep layers V/VI. Beta oscillations were unaffected by specific AMPA receptor blockade, abolished by the GABA type A receptor (GABA(A)R) antagonist picrotoxin and the gap-junction blocker carbenoxolone, and modulated by pentobarbital and zolpidem indicating dependence on networks of GABAergic interneurons and electrical coupling. High frequency stimulation (HFS) at 125 Hz in superficial layers, designed to mimic transdural/transcranial stimulation, generated gamma oscillations in layers II and V (incidence 95%, 69.2+/-7.3 Hz, n=17) with very fast oscillatory components (VFO; 100-250 Hz). Stimulation at 4 Hz, however, preferentially promoted theta activity (incidence 62.5%, 5.1+/-0.6 Hz, n=15) that effected strong amplitude modulation of ongoing beta activity. Stimulation at 20 Hz evoked mixed theta and gamma responses. These data suggest that within M1, evoked theta, gamma and fast oscillations may coexist with and in some cases modulate pharmacologically induced beta oscillations.

Altered first-pass effects in a liver transplant recipient explained intraindividual variation in calcineurin inhibitor concentrations: a case report.

BACKGROUND: The significant interindividual and intraindividual variability in the blood concentrations of the most commonly used calcineurin inhibitors such as tacrolimus and cyclosporine makes the exact dosing of these agents in transplant recipients very challenging. As both of these drugs have narrow therapeutic index and are metabolized by hepatic and intestinal cytochrome P450 3A, we tested the hypothesis that these variations are secondary to varying first-pass effects in the gut and the liver over a period of time. CASE REPORT: A liver transplant recipient, who had previously presented with tacrolimus toxicity on his usual dosing regimen and intolerant to standard doses of cyclosporine, was selected to undergo the study. Oral and intravenous midazolam was used as the probe to measure hepatic and intestinal CYP3A4 activities at two different time points (phases one and two). Small intestinal biopsies were also obtained for measuring CYP3A4 activity for in vitro studies. On serially determining the patient's hepatic and intestinal CYP3A activities, we concluded that the variability in the dosing requirements is due to altered first-pass effects in the intestine. DISCUSSION: Transplant recipients receive multiple medications that may inhibit or induce these metabolizing enzymes, which eventually determine the concentrations of these narrow therapeutic agents. If no obvious etiology of intolerance to calcineurin inhibitors in a transplant recipient is identified, one should consider altered first-pass effects in the gut and the liver contributing to intraindividual variations in the blood concentrations.

Electron microscopic visualization of RecT protein and its complexes with DNA.

Electron microscopy has been used to examine Escherichia coli RecT protein alone and in the complexes it forms with DNA substrates, with which it catalyzes strand exchange in vitro. Negative staining has revealed that the 33 kDa RecT protein monomers form open C-shaped and closed O-shaped particles. RecT protein monomers assemble into donut-shaped oligomers containing seven or eight protein monomers and rod-like structures. When bound to single-stranded DNA, RecT forms highly twisted nucleoprotein filaments that are 18 nm in diameter and have a helical pitch of 10 nm. When added to linear duplex DNA in the presence of active RecE protein (exonuclease VIII), filamentous nucleoprotein complexes are formed on the DNA ends and the DNA molecules are frequently cyclized through protein-protein interactions.

Physiologically based pharmacokinetic modeling in drug discovery and development: a pharmaceutical industry perspective.

The application of physiologically based pharmacokinetic (PBPK) modeling has developed rapidly within the pharmaceutical industry and is becoming an integral part of drug discovery and development. In this study, we provide a cross pharmaceutical industry position on "how PBPK modeling can be applied in industry" focusing on the strategies for application of PBPK at different stages, an associated perspective on the confidence and challenges, as well as guidance on interacting with regulatory agencies and internal best practices.

Understanding Disease-Drug Interactions in Cancer Patients: Implications for Dosing Within the Therapeutic Window.

The human inflammatory response can result in the alteration of drug clearance through effects on metabolizing enzymes or transporters. In this article we briefly review the theory of how cancer can lead to indirect changes in drug metabolism, review acute phase proteins and cytokines as markers of changes in cytochrome P450 (CYP) activity in cancer patients, and provide clinical case examples of how the inflammation in advanced cancer patients can lead to altered CYP-mediated drug clearance.

Human cytochrome P450 3A (CYP3A) mediated midazolam metabolism: the effect of assay conditions and regioselective stimulation by alpha-naphthoflavone, terfenadine and testosterone.

The effect of ionic strength, assay constituents, alpha-naphthoflavone (aNF), terfenadine and testosterone on human CYP3A mediated midazolam (MDZ) 1'-hydroxylation (MDZ 1'-OH) and 4-hydroxylation (MDZ 4-OH) in vitro was examined. Increasing concentration of Tris-HCl (Tris) and sodium phosphate (PO4) buffers differentially affected MDZ 1'-OH and MDZ 4-OH formation rates and had a different effect on MDZ metabolism mediated by microsomes containing CYP3A4 versus CYP3A4 and CYP3A5. MDZ metabolism was not affected by PO4 buffer concentration when cumene hydroperoxide (CUOOH) was used as the source of reactive oxygen. Interestingly, the ammonium ion present in the solution of glucose 6-phosphate dehydrogenase was found to inhibit MDZ metabolism. The addition of MgCl2 up to 50 mM and CaCl2 (5-30 mM) had no affect or inhibited MDZ metabolism, respectively. Formation of MDZ 1'-OH by microsomes from adult and fetal liver and expressed CYP3A4 was regioselectively stimulated by aNF (10 microM). In human hepatocytes, aNF stimulated MDZ 1'-OH formation (up to 100%). Terfenadine (20 microM) regioselectively stimulated MDZ 1'-OH formation in Tris (1-200 mM) and PO4 (1-10 mM) buffers by up to 159%. Surprisingly, with expressed CYP3A4, terfenadine (20 microM) inhibited MDZ 1'-OH formation. Terfenadine (20 microM) had little effect on MDZ 1'-OH formation by fetal liver microsomes. Testosterone (10 and 100 microM) regioselectively stimulated (up to 269%) MDZ 4-OH formation by adult liver microsomes and expressed CYP3A4. Testosterone (100 microM) inhibited (> 40%) MDZ 1'-OH and MDZ 4-OH formation by fetal liver microsomes. With adult liver microsomes, aNF and terfenadine had little effect on the Km for MDZ 1'-OH formation. However, the Km for MDZ 4-OH formation was decreased (up to 94%) by 100 microM testosterone. In the presence of CUOOH, no stimulation of MDZ metabolism was observed by aNF, terfenadine or testosterone in adult liver microsomes. These studies indicate that because assay conditions can substantially alter the catalytic activity of CYP3A, caution should be exerted when extrapolating results between in vitro and in vivo, and when results from different laboratories are compared. Further, these results suggest that the stimulation of CYP3A4 may also occur in vivo and, consequently, may have clinical importance.

Tissue distribution and interindividual variation in human UDP-glucuronosyltransferase activity: relationship between UGT1A1 promoter genotype and variability in a liver bank.

The variability in a liver bank and tissue distribution of three probe UDP-glucuronosyltransferase (UGT) activities were determined as a means to predict interindividual differences in expression and the contribution of extrahepatic metabolism to presystemic and systemic clearance. Formation rates of acetaminophen-O-glucuronide (APAPG), morphine-3-glucuronide (M3G), and oestradiol-3-glucuronide (E3G) as probes for UGT1A6, 2B7, and 1A1, respectively, were determined in human kidney, liver, and lung microsomes, and in microsomes from intestinal mucosa corresponding to duodenum, jejunum and ileum. While formation of E3G and APAPG were detectable in human kidney microsomes, M3G formation rates from kidney microsomes approached the levels seen in liver, indicating significant expression of UGT2B7. Interestingly, rates of E3G formation in human intestine exceeded the hepatic rates by several fold, while APAPG and M3G formation rates were low. The intestinal apparent Km value for E3G formation was essentially identical to that seen in liver, consistent with intestinal UGT1A1 expression. No UGT activities were observed in lung. Variability in APAPG and M3G activity across a bank of 20 human livers was modest (< or = 7-fold), compared to E3G formation, which varied approximately 30-fold. The E3G formation rates were found to segregate by UGT1A1 promoter genotype, with wild-type (TA)6 rates significantly greater than homozygous mutant (TA)7 individuals. Kinetic analyses were performed to demonstrate that the promoter mutation altered apparent Vmax without significantly affecting apparent Km. These results suggest that glucuronidation, and specifically UGT1A1 activity, can profoundly contribute to intestinal first pass metabolism and interindividual variability due to the expression of common allelic variants.

The human pregnane X receptor: genomic structure and identification and functional characterization of natural allelic variants.

The pregnane X receptor (PXR)/steroid and xenobiotic receptor (SXR) transcriptionally activates cytochrome P4503A4 (CYP3A4) when ligand activated by endobiotics and xenobiotics. We cloned the human PXR gene and analysed the sequence in DNAs of individuals whose CYP3A phenotype was known. The PXR gene spans 35 kb, contains nine exons, and mapped to chromosome 13q11-13. Thirty-eight single nucleotide polymorphisms (SNPs) were identified including six SNPs in the coding region. Three of the coding SNPs are non-synonymous creating new PXR alleles [PXR*2, P27S (79C to T); PXR*3, G36R (106G to A); and PXR*4, R122Q (4321G to A)]. The frequency of PXR*2 was 0.20 in African Americans and was never found in Caucasians. Hepatic expression of CYP3A4 protein was not significantly different between African Americans homozygous for PXR*1 compared to those with one PXR*2 allele. PXR*4 was a rare variant found in only one Caucasian person. Homology modelling suggested that R122Q, (PXR*4) is a direct DNA contact site variation in the third alpha-helix in the DNA binding domain. Compared with PXR*1, and variants PXR*2 and PXR*3, only the variant PXR*4 protein had significantly decreased affinity for the PXR binding sequence in electromobility shift assays and attenuated ligand activation of the CYP3A4 reporter plasmids in transient transfection assays. However, the person heterozygous for PXR*4 is normal for CYP3A4 metabolism phenotype. The relevance of each of the 38 PXR SNPs identified in DNA of individuals whose CYP3A basal and rifampin-inducible CYP3A4 expression was determined in vivo and/or in vitro was demonstrated by univariate statistical analysis. Because ligand activation of PXR and upregulation of a system of drug detoxification genes are major determinants of drug interactions, it will now be useful to extend this work to determine the association of these common PXR SNPs to human variation in induction of other drug detoxification gene targets.

Polymorphic metabolism of mephenytoin in man: pharmacokinetic interaction with a co-regulated substrate, mephobarbital.

The simultaneous dosing of two drugs with co-regulated genetic polymorphisms determined by a single cytochrome P-450 isozyme could result in competitive inhibition of metabolism. We investigated this hypothesis in vivo by studying the interaction of mephobarbital and mephenytoin in eight normal subjects with wide variability in S-mephenytoin 4-hydroxylation. Each received oral racemic mephenytoin (100 mg) alone and, on a separate occasion, 1 hour after oral racemic mephobarbital (200 mg). After mephenytoin dosing alone, the 8-hour urinary enantiomeric (R/S) ratio indicated one poor (PM), one intermediate (IM), and six extensive (EM) metabolizers. Total intrinsic clearance of S-mephenytoin varied more than 100-fold, whereas the range for R-mephenytoin was only twofold. The urinary R/S ratio correlated (r = 0.92) with the enantiomeric ratio of the plasma AUCs over the same period, indicating no stereoselectivity in renal clearance. When mephenytoin was taken in the presence of mephobarbital, peak levels and AUC of S-mephenytoin increased while those of the R-enantiomer remained unchanged. Accordingly, the R/S ratios in both plasma and urine were reduced, with the change rank order-related to the control value of the total intrinsic clearance of S-mephenytoin (i.e., greatest in the most extensive EM). Thus the urinary R/S ratio can be used as a measure of the enantiomeric ratio of the plasma concentrations over the same time period of collection. Moreover, this ratio may be used to detect drug interactions that involve the cytochrome P-450 isozyme(s) responsible for the polymorphic 4-hydroxylation of mephenytoin.(ABSTRACT TRUNCATED AT 250 WORDS)

The interaction of diltiazem with simvastatin.

BACKGROUND: Simvastatin is an inhibitor of 3-hydroxy-3-methylglutaryl coenzyme A (HMG-CoA) reductase that is used as a cholesterol-lowering agent and is metabolized by cytochrome P450 3A (CYP3A) enzymes. Diltiazem is a substrate and an inhibitor of CYP3A enzymes and is commonly coadministered with cholesterol-lowering agents such as simvastatin. The objective of this study was to quantify the effect of diltiazem on the pharmacokinetics of simvastatin. METHOD: A fixed-order study was conducted in 10 healthy volunteers with a 2-week washout period between the phases. In one arm of the study, a single 20-mg dose of simvastatin was administered orally; the second arm entailed administration of a single 20-mg dose of simvastatin orally after 2 weeks of treatment with 120 mg diltiazem twice a day. RESULTS: Diltiazem significantly increased the mean peak serum concentration of simvastatin by 3.6-fold (P < .05) and simvastatin acid by 3.7-fold (P < .05). Diltiazem also significantly increased the area under the serum concentration-time curve of simvastatin 5-fold (P < .05) and the elimination half-life 2.3-fold (P < .05). There was no change in the time to peak concentration for simvastatin and simvastatin acid. CONCLUSION: Diltiazem coadministration resulted in a significant interaction with simvastatin, probably by inhibiting CYP3A-mediated metabolism. Concomitant use of diltiazem or other potent inhibitors of CYP3A with simvastatin should be avoided, or close clinical monitoring should be used.

Effect of clofibrate on the chiral inversion of ibuprofen in healthy volunteers.

OBJECTIVES: To determine the influence of the hypolipidemic drug clofibrate on the stereoselective metabolism of ibuprofen in humans. METHODS: Healthy male subjects (n = 12) ingested a dose of 400 mg pseudoracemic ibuprofen (200 mg R-ibuprofen, 160 mg S-ibuprofen, and 40 mg 13C-S-ibuprofen) on two occasions after either pretreatment with clofibrate (2 gm/day over 1 week) or no pretreatment in a randomized order. RESULTS: When subjects were pretreated with clofibrate, clearances of R-ibuprofen and 13C-S-ibuprofen increased significantly from 55.0 and 66.4 ml/min to 186.2 and 106.7 ml/min (p < 0.01), respectively. This increase was similarly reflected in the clearance by inversion of R-ibuprofen (control, 36.0 ml/min; treated, 118.8 ml/min; p < 0.01), as well as in the clearance by noninversion (control, 19.0 ml/min; treated, 67.4 ml/min; p < 0.01). Unbound clearance values significantly increased for R-ibuprofen (control, 19.5 L/min; treated, 38.7 L/min) but not for 13C-S-ibuprofen (11.8 versus 10.6 L/min, respectively). The fractional inversion of ibuprofen calculated from the urinary metabolite data was increased after clofibrate pretreatment (clofibrate group, 66.4%; control, 53.5%; p < 0.01). However, this was not evident when fractional inversion was calculated from the plasma concentration-time data for the unmetabolized drug. CONCLUSIONS: Clofibrate altered the stereoselective disposition of ibuprofen in healthy volunteers by increased formation of R-ibuprofenoyl-coenzyme A rather than by an effect on oxidative metabolism of ibuprofen. This interaction has potential therapeutic implications.

Furosemide absorption in patients with cirrhosis.

Twelve patients with cirrhosis (seven mild and five severe) were administered intravenous and oral furosemide in random order to assess its absorption and disposition. Total serum clearance (113 +/- 49 ml/min), volume of distribution (11.9 +/- 4.5 L), and elimination half-life (166 +/- 149 minutes) were similar to those reported previously in both healthy control subjects and patients with cirrhosis. Bioavailability of 58% +/- 17% (range, 37% to 82%) was comparable to that of previous studies, and there was no difference between patients with mild and those with severe cirrhosis. In 9 of 12 patients the mean absorption time was longer than the mean residence time determined after intravenous administration (mean for all patients, 203 +/- 86 versus 134 +/- 101 minutes; p less than 0.05), indicating that furosemide followed a "flip-flop" model in these patients. In all patients the mean absorption time was prolonged relative to normal subjects irrespective of the presence of edema. As such, the slower absorption of furosemide in edematous states, such as congestive heart failure and cirrhosis, does not appear to be a consequence of edema per se. Moreover, because similar changes occur in patients with congestive heart failure, it seems that diseases with diverse pathophysiology can slow furosemide absorption.

The effect of hormone replacement therapy on CYP3A activity.

BACKGROUND: The effect of menopause and hormone replacement therapy on hepatic and intestinal wall CYP3A activity is poorly defined. This study was therefore designed to determine the effect of menopause and estrogen replacement therapy on hepatic and intestinal CYP3A activity with a specific CYP3A substrate, midazolam. METHODS: Twelve young women (27 +/- 5 years), 10 elderly women receiving estrogen replacement therapy (71 +/- 6 years), and 14 elderly women not receiving estrogen replacement therapy (71 +/- 5 years) received simultaneous intravenous (0.05 mg/kg over 30 minutes) and oral (3 to 4 mg of a stable isotope, 15N3-midazolam) doses of midazolam. Serum and urine samples were assayed for midazolam, 15N3-midazolam, and metabolites by use of gas chromatography-mass spectrometry. RESULTS: No significant (P > .05) differences were observed in systemic clearance and oral clearance between the three groups. Likewise, no differences were observed in oral, hepatic, or intestinal availability. A significant correlation was observed between oral and intestinal availability and not hepatic availability. CONCLUSION: Neither menopause nor menopause with estrogen replacement therapy altered intestinal or hepatic CYP3A activity relative to that in a control group of young women.

The effect of rifampin administration on the disposition of fexofenadine.

OBJECTIVE: Our objective was to assess the effect of rifampin (INN, rifampicin) on the pharmacokinetics of fexofenadine and to assess the influence of advanced age and sex. METHODS: Twelve young volunteers (6 men and 6 women; age range, 22 to 35 years) and twelve elderly volunteers (6 men and 6 women; age range, 65 to 76 years) received a 60-mg oral dose of fexofenadine before and after treatment with 600 mg of oral rifampin for 6 days. Blood and urine were collected for 48 hours and assayed for fexofenadine, azacyclonol, and rifampin by HPLC with either fluorescence or mass spectrometry detection. RESULTS: All of the groups had a significant increase (P <.05) in the oral clearance of fexofenadine after rifampin treatment: young men, 2955 +/- 1516 versus 5524 +/- 3410 mL/min; young women, 2632 +/- 996 versus 7091 +/- 5,379 mL/min; elderly men, 1760 +/- 850 versus 4608 +/- 1159 mL/min; and elderly women, 2210 +/- 554 versus 4845 +/- 1600 mL/min. The peak serum concentration of fexofenadine was also significantly reduced (P <.05) by rifampin treatment: young men, 77 +/- 31 versus 52 +/- 17 ng/mL; young women, 72 +/- 19 versus 36 +/- 14 ng/mL; elderly men, 106 +/- 42 versus 52 +/- 14 ng/mL; elderly women, 76 +/- 23 versus 46 +/- 19 ng/mL. Half-life (150 to 230 minutes), time to maximum concentration (130 to 205 minutes), renal clearance (95 to 153 mL/min), and fraction unbound (2.9% to 3.7%) of fexofenadine showed no significant difference between control and treatment. The amount of azacyclonol, a CYP3A4 mediated metabolite of fexofenadine, eliminated renally increased on average 2-fold after rifampin dosing; however, this pathway accounted for less than 0.5% of the dose. No effect of age or sex on fexofenadine disposition or serum trough rifampin concentration (0.2 microg/mL to 1.8 microg/mL) was observed before or after rifampin treatment. CONCLUSION: This study showed that rifampin effectively increased fexofenadine oral clearance and that this effect was independent of age and sex. We conclude that the cause of the increased oral clearance of fexofenadine is a reduced bioavailability caused by induction of intestinal P-glycoprotein.

Determination of cytochrome P450 3A4/5 activity in vivo with dextromethorphan N-demethylation.

Dextromethorphan is used widely in vivo to phenotype the polymorphically expressed cytochrome P450 (CYP) 2D6. Dextromethorphan is N-demethylated in vitro to 3-methoxymorphinan by human CYP3A4/5. We examined whether the dextromethorphan/3-methoxymorphinan urinary metabolic ratio (MR) could be used as an in vivo probe of CYP3A. Urinary excretion of 3-methoxymorphinan was excretion rate-limited in extensive metabolizers of CYP2D6, which necessitated a longer urine collection, 0 to 72 hours, to obtain true MR values for CYP3A. The urine excretion of dextromethorphan and 3-methoxymorphinan was delayed in poor metabolizers of CYP2D6 but appeared to be formation rate-limited. The delayed excretion in poor metabolizers necessitated longer urine collection intervals, 0 to 11 days, to estimate the true CYP3A MR and 0 to 8 days to estimate the true CYP2D6 MR. However, a 72-hour collection in poor metabolizers was used as an index of the true dextromethorphan/3-methoxymorphinan MR. Rifampin (300 mg b.i.d. for 7 days) significantly reduced the 0- to 72-hour dextromethorphan/3-methoxymorphinan MR consistent with an 830% (+/- 1808%) induction of CYP3A activity (n = 8), whereas erythromycin (250 mg q.i.d. for 7 days) significantly increased the dextromethorphan/3-methoxymorphinan MR, corresponding to a 34% +/- 44% inhibition of activity (n = 7) in extensive metabolizers and poor metabolizers. The changes in CYP3A activity were independent of CYP2D6 phenotype and were also observed after 24- and 48-hour urine collections in extensive metabolizers and poor metabolizers. In addition, MRs reflecting CYP2D6 and CYP3A were not significantly correlated. We conclude that the commonly used antitussive dextromethorphan can be used as an in vivo marker of CYP3A and CYP2D6 activity.

The interaction of diltiazem with lovastatin and pravastatin.

BACKGROUND: Lovastatin is oxidized by cytochrome P4503A to active metabolites but pravastatin is active alone and is not metabolized by cytochrome P450. Diltiazem, a substrate and a potent inhibitor of cytochrome P4503A enzymes, is commonly coadministered with cholesterol-lowering agents. METHODS: This was a balanced, randomized, open-label, 4-way crossover study in 10 healthy volunteers, with a 2-week washout period between the phases. Study arms were (1) administration of a single dose of 20 mg lovastatin, (2) administration of a single dose of 20 mg pravastatin, (3) administration of a single dose of lovastatin after administration of 120 mg diltiazem twice a day for 2 weeks, and (4) administration of a single dose of pravastatin after administration of 120 mg diltiazem twice a day for 2 weeks. RESULTS: Diltiazem significantly (P < .05) increased the oral area under the serum concentration-time curve (AUC) of lovastatin from 3607 +/- 1525 ng/ml/min (mean +/- SD) to 12886 +/- 6558 ng/ml/min and maximum serum concentration (Cmax) from 6 +/- 2 to 26 +/- 9 ng/ml but did not influence the elimination half-life. Diltiazem did not affect the oral AUC, Cmax, or half-life of pravastatin. The average steady-state serum concentrations of diltiazem were not significantly different between the lovastatin (130 +/- 58 ng/ml) and pravastatin (110 +/- 30 ng/ml) study arms. CONCLUSION: Diltiazem greatly increased the plasma concentration of lovastatin, but the magnitude of this effect was much greater than that predicted by the systemic serum concentration, suggesting that this interaction is a first-pass rather than a systemic event. The magnitude of this effect and the frequency of coadministration suggest that caution is necessary when administering diltiazem and lovastatin together. Further studies should explore whether this interaction abrogates the efficacy of lovastatin or enhances toxicity and whether it occurs with other cytochrome P4503A4-metabolized 3-hydroxy-3-methylglutaryl-coenzyme A reductase inhibitors, such as simvastatin, fluvastatin, and atorvastatin.

Lack of presystemic inversion of (R)- to (S)-ibuprofen in humans.

Presystemic inversion of (R)- to (S)-ibuprofen has been proposed but not directly examined in humans. We investigated the bioavailability of the enantiomers of ibuprofen in 10 healthy volunteers. Low-dose racemic ibuprofen (400 mg) was administered orally and intravenously (60-minute infusion), in random order. There were no significant differences between oral and intravenous doses for the area under the curve values, terminal rate constants, clearances, metabolite formation clearances, and serum protein binding for (R)- and (S)-ibuprofen. The bioavailabilities of (R)-ibuprofen and total ibuprofen were 0.92 +/- 0.11 and 0.95 +/- 0.08, respectively. The fractional inversion of (R)-ibuprofen was determined by two methods (stable isotope method and from the stereochemical composition of the urinary metabolites) that gave similar estimates of inversion for oral dosing (0.56 +/- 0.12 and 0.60 +/- 0.07, respectively) and intravenous dosing (0.56 +/- 0.09 and 0.60 +/- 0.06, respectively). We conclude that the bioavailability of both enantiomers of ibuprofen is complete and find no evidence of significant presystemic inversion.