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G R Wilkinson

Publications and source records attributed to G R Wilkinson.

At least 55 records · Page 3Linked to original sources
PubMed1996

Comparison of the dapsone recovery ratio and the erythromycin breath test as in vivo probes of CYP3A activity in patients with rheumatoid arthritis receiving cyclosporine.

INTRODUCTION: Cytochrome P4503A (CYP3A) is primarily responsible for the metabolism of cyclosporine and that of many other drugs. Several substrates of CYP3A have been investigated for use as pharmacologic probes to predict the CYP3A-metabolizing capacity of an individual and, therefore, the disposition of other CYP3A substrate drugs. One such measure of CYP3A activity is the 14C erythromycin breath test, which has been applied to the prediction of cyclosporine disposition. However, the test has practical limitations. Because of this, the 0- to 8-hour urinary dapsone recovery ratio has been studied as an alternative and more practical probe of CYP3A activity. METHODS: The dapsone recovery ratio and the 14C erythromycin breath test were correlated with cyclosporine concentrations in 16 patients with rheumatoid arthritis to determine the usefulness of the dapsone recovery ratio as an alternative to the 14C erythromycin breath test. The erythromycin breath test showed a fourfold variation between subjects and correlated weakly with trough cyclosporine concentrations (r = -0.50, p < 0.05), whereas the dapsone recovery ratio varied only approximately twofold between subjects and did not correlate with trough cyclosporine concentrations (r = 0.02, p = 0.94). The correlation between the dapsone recovery ratio and the erythromycin breath test (r = 0.22, p = 0.41) was not significant. CONCLUSIONS: These data suggest that results obtained with one probe in vivo may not apply to another CYP3A substrate. The poor quantitative relationship between cyclosporine concentrations and the erythromycin breath test limits its usefulness in the prediction of an individual's cyclosporine dose requirement.

Adult↗
PubMed1996

Oral first-pass elimination of midazolam involves both gastrointestinal and hepatic CYP3A-mediated metabolism.

OBJECTIVE: To determine in humans the relative roles of intestinal and hepatic metabolism in the oral first-pass elimination of a CYP3A substrate using midazolam as a model compound. METHODS: Midazolam was administered intravenously (1 mg) or orally (2 mg) to 20 healthy young subjects (10 men and 10 women) in a random fashion, and the disposition of the drug and its 1'-hydroxy metabolite were determined. In separate in vitro studies, the CYP3A-mediated formation of 1'-hydroxymidazolam by human hepatic and intestinal microsomes was investigated. RESULTS: No gender-related differences were noted in either the systemic (370 +/- 114 ml/min [mean +/- SD]) or oral (1413 +/- 807 ml/min) clearance values of midazolam. Despite complete oral absorption, measured oral bioavailability was on average about 50% less than that predicted on the assumption that only the liver contributed to first-pass metabolism. Pharmacokinetic estimation of the intestinal component indicated an extraction ratio (0.43 +/- 0.24) that was similar to that of the liver (0.44 +/- 0.14). 1'-Hydroxymidazolam was extensively but variably formed in vitro by both hepatic and intestinal microsomes and, although the intrinsic clearance (V(max)/Km) was higher in the liver preparations (540 +/- 747 versus 135 +/- 92 microliters/min/mg protein), this difference was not statistically significant. CONCLUSIONS: These results show that the small intestine can be a major site for presystemic, CYP3A-mediated metabolism after oral administration. Moreover, it appears that this represents a true first-pass effect. In addition, intestinal and hepatic metabolism may be important factors in interindividual variability in disposition after oral administration of midazolam and similar CYP3A substrates. Finally, intestinal localization of CYP3A may be significant in metabolism-based drug-drug interactions.

Administration, Oral↗
PubMed1996

Differences between white subjects and Chinese subjects in the in vivo inhibition of cytochrome P450s 2C19, 2D6, and 3A by omeprazole.

OBJECTIVES: To determine the effects of omeprazole on indexes of CYP2D6, CYP2C19 and 3A in vivo activity and to compare these in white subjects and Chinese subjects. METHODS: Omeprazole, 40 mg/day, or placebo were administered in a double-blind crossover study for 3 weeks to eight healthy white and seven Chinese male extensive metabolizers of mephenytoin and debrisoquin. Debrisoquin (10 mg), dapsone (100 mg), and mephenytoin (100 mg) were given 1 week before administration, on the last day of administration, and 3 weeks after administration, and urine was collected over 8 hours. Phenotypic trait values were obtained from the urinary recoveries of the probe drugs or their metabolites. RESULTS: In the white subjects, omeprazole significantly inhibited CYP2C19-mediated S-mephenytoin metabolism as indicated by decreases in the urinary R/S enantiomeric ratio (63% +/- 13%; p < 0.02; mean +/- SD) and the excretion of 4'-hydroxymephenytoin (39% +/- 13%; p < 0.001). Similar but smaller changes were also noted in Chinese subjects, 22% +/- 25% (p = 0.08) and 29% +/- 13% (p < 0.002), respectively. The interracial differences in the extent of inhibition of metabolism were statistically significant (p < 0.01 and p < 0.05, respectively). In contrast, the debrisoquin urinary metabolic ratio, a measure of CYP2D6, was unaffected. The excretion of hydroxylamine dapsone-a putative probe of CYP3A activity-was reduced by 40% +/- 30% (p < 0.03) in white subjects but not in Chinese subjects. CONCLUSIONS: Omeprazole selectively inhibits the in vivo metabolism of S-mephenytoin, consistent with the predictions based on in vitro studies. The extent of interaction is greater in subjects of white European ancestry. It is to be expected that similar situations would also occur when omeprazole is coadministered with other substrates of CYP2C19.

Adult↗
PubMed1996

Disposition of diazepam in young and elderly subjects after acute and chronic dosing.

1. The pharmacokinetics of diazepam were examined in seven young (20-30 years) and six elderly (60-75 years) males prior to and also after chronic oral dosing of diazepam. 2. Following intravenous administration, the half-life and volume of distribution of 14C-labelled diazepam in the elderly were approximately twofold greater than corresponding estimates in younger subjects (mean +/- s.d., 71.5 +/- 27.6 vs 44.5 +/- 16.5 h and 1.39 +/- 0.32 vs 0.88 +/- 0.30 1 kg-1, respectively). Clearance did not differ between the two groups (0.26 +/- 0.09 vs 0.29 +/- 0.09 ml min-1 kg-1). 3. The accumulation of diazepam and its major metabolite, desmethyldiazepam, were extensive during chronic administration. A radioreceptor assay that measured total benzodiazepine activity, including diazepam and its active metabolites, indicated that the accumulation of 'benzodiazepine equivalents' was similar to the sum of the accumulated diazepam and desmethyldiazepam concentration levels. However, the level of 'benzodiazepine equivalents' on multiple-dosing was about double that of the predicted steady-state 'equivalent' concentration from single-dose studies. This was due to the insensitivity of the radioreceptor assay for desmethyldiazepam following single-dose diazepam administration. 4. There were no age- or dosing-related differences in diazepam clearance (0.37 +/- 0.22 vs 0.32 +/- 0.18 ml min-1 kg-1, young vs elderly, single-dose; 0.37 +/- 0.11 vs 0.27 +/- 0.12 ml min-1 kg-1, young vs elderly, multiple-dose) and no age-related differences in the levels of accumulated 'benzodiazepine equivalents' (243.7 +/- 60.1 vs 288.0 +/- 125.8 ng ml-1, young vs elderly). 5. Thus, changes that occur in diazepam disposition with ageing after acute administration do not appear to be important during chronic dosing. On the other hand, accumulation of diazepam and desmethyldiazepam are considerable and would be expected to be clinically relevant.

Adult↗
PubMed1996

In vivo and in vitro characterization of CYP2E1 activity in Japanese and Caucasians.

Chlorzoxazone's disposition after oral administration was determined in 20 young healthy Caucasian men and a similar group of Japanese men. The drug's plasma concentrations were significantly higher and its rate of elimination slower in Japanese compared to Caucasian men. Accordingly, chlorzoxazone's oral clearance was smaller (40%) in Japanese men and a similar difference (30%) was still apparent after normalizing for body weight (3.74 +/- 1.23 versus 5.05 +/- 1.41 ml.min-1.kg-1, P < .05). This slower elimination was associated with a reduced (fractional) clearance by 6-hydroxylation (2.34 +/- 1.04 ml.min-1.kg-1 versus 3.23 +/- 1.10, P < .05). Because such metabolism is mediated by cytochrome P4502E1 (CYP2E1), these findings suggest a lower level of the enzyme's catalytic activity in Japanese men. This was confirmed by in vitro studies with microsomes prepared from livers of individuals representative of the two racial groups. CYP2E1 levels were lower (61% P < .002) and CYP2E1-mediated chlorzoxazone 6-hydroxylase (22%, P < .001) and aniline 4-hydroylase (35%, P < .0001) activities were reduced in Japanese preparations compared to those from Caucasians. No relationships were found between measures of CYP2E1 activity, both in vivo and in vitro, and genomic polymorphisms in the CYP2E1 gene identified by Rsal/Pstl and Dral restriction fragment length polymorphisms. Collectively, these data show an interracial difference in CYP2E1 activity. Because this enzyme is importantly involved in the activation of environmental procarcinogens, such a difference may account, in part, for the lower rate of some cancers, e.g., lung cancer, in Japanese compared to Caucasians men.

Adolescent↗
PubMed1995

The procarcinogen hypothesis for bladder cancer: activities of individual drug metabolizing enzymes as risk factors.

Bladder cancer provides the most definitive example for an association between environmental agents and cancer. However, in the absence of industrial occupational exposure, the primary carcinogen is rarely identified, and the mechanisms involved in cancer formation are poorly understood. The environmental procarcinogen hypothesis of tumour pathogenesis proposes that many carcinogens require metabolic activation by drug metabolizing enzymes to form the proximate carcinogen. A balance of exposure to the carcinogen, the activity of the enzymes involved in either formation of proximate carcinogen, or production of non-toxic metabolites, will determine tumour risk. We have used mephenytoin, debrisoquine and dapsone as selective probes for the phenotypic measures of activity of CYP2C19, CYP2D6, and CYP3A4, respectively. Within subject reproducibility of phenotypic measures, and the lack of cross-inhibition when the three drugs are given in a concurrent cocktail, have been confirmed. We have applied the cocktail drug approach in two, non-overlapping series of cases with bladder cancer and matched controls. In both series, patients with aggressive bladder cancer (GIII histopathology) had a history of excess alcohol intake, an under-representation of poor metabolizers of debrisoquine, a significant mean reduction in dapsone recovery ratio, but no difference in mephenytoin phenotype. Collectively, these observations involving multiple routes of drug metabolism support the procarcinogen environmental hypothesis for bladder cancer and suggest that measurement of activity of selected individual drug metabolizing enzymes involved in the pathogenesis of this tumour can be used to identify subjects at high risk of developing bladder cancer.

Aryl Hydrocarbon Hydroxylases↗
PubMed1994

The major genetic defect responsible for the polymorphism of S-mephenytoin metabolism in humans.

The metabolism of the anticonvulsant drug mephenytoin exhibits a genetic polymorphism in humans, with the poor metabolizer trait being inherited in an autosomal recessive fashion. There are large interracial differences in the frequency of the poor metabolizer phenotype, with Oriental populations having a 5-fold greater frequency compared to Caucasians. Impaired metabolism of mephenytoin and a number of other currently used drugs results from a defect in a cytochrome P450 enzyme recently identified as CYP2C19. Attempts over the past decade to define the molecular genetic basis of the polymorphism have, however, been unsuccessful. We now report that the principal defect in poor metabolizers is a single base pair (G-->A) mutation in exon 5 of CYP2C19, which creates an aberrant splice site. This change alters the reading frame of the mRNA starting with amino acid 215 and produces a premature stop codon 20 amino acids downstream, which results in a truncated, non-functional protein. We further demonstrate that 7/10 Caucasian and 10/17 Japanese poor metabolizers are homozygous for this defect, indicating that this is the major defect responsible for the poor metabolizer phenotype. Finally, the familial inheritance of the deficient allele was found to be concordant with that of the phenotypic trait.

Alternative Splicing↗
PubMed1994

Effect of fasting and obesity in humans on the 6-hydroxylation of chlorzoxazone: a putative probe of CYP2E1 activity.

BACKGROUND AND OBJECTIVES: The hepatic 6-hydroxylation of chlorzoxazone in vitro is mediated primarily by CYP2E1, and measurement of this metabolic pathway may provide an in vivo probe of the enzyme. In animals, such as the rat, levels of CYP2E1 are induced by both fasting and obesity. This study investigated whether these two physiologic factors are determinants of the metabolism and disposition of chlorzoxazone in humans. METHODS: The plasma concentration-time profiles of chlorzoxazone and its 6-hydroxy metabolite were determined after oral administration of parent drug (250 mg). The urinary excretion of the metabolite was also determined. In one study, the disposition profiles were obtained in six healthy white men, first after an overnight fast, and on a separate occasion after a 38-hour fast. The second study investigated the disposition of chlorzoxazone in nine obese women and in nine age-matched women. RESULTS: Prolonged fasting produced a significant increase in circulating ketone bodies. This was associated with a reduction in the oral clearance of chlorzoxazone (mean +/- SD, 5.79 +/- 1.04 to 3.69 +/- 1.54 ml.min-1.kg-1; p < 0.03). The 0- to 24-hour urinary recovery of the 6-hydroxy metabolite was extensive (50% to 80%), and the reduced clearance reflected a lower 6-hydroxylating ability after fasting. The elimination half-life of the drug was increased by a similar extent to clearance (1.00 +/- 0.09 versus 1.50 +/- 0.42 hours; p < 0.004), whereas its apparent volume of distribution was unaffected by fasting. By contrast, obesity resulted in significant increases in the oral clearance and distribution of chlorzoxazone on both an absolute and weight-normalized basis; for example, 4.15 +/- 0.81 versus 6.23 +/- 1.72 ml.min-1.kg-1 and 0.50 +/- 0.28 versus 0.82 +/- 0.19 L.kg-1. Half-life of elimination was similar in both groups of subjects. Estimation of the fractional clearance of 6-hydroxychlorzoxazone showed that obesity increased this parameter to a similar extent as oral clearance. The difference in the oral clearance and 6-hydroxylating ability of nonobese men and women was also statistically different. CONCLUSIONS: A discordancy was observed between the reported effect of fasting in rodents and that observed in humans. This may reflect an interspecies difference in CYP2E1 regulation or, more likely, destruction of the enzyme by lipid peroxidation resulting from the prolonged period of fasting. However, serious to morbid obesity in humans is associated with increased 6-hydroxylation of chlorzoxazone, consistent with induction of CYP2E1. Accordingly, such individuals may be at increased risk of CYP2E1-mediated toxicities and adverse effects caused by the formation of CYP2E1-mediated metabolites of environmental agents. In addition, the efficacy of an active drug that is a CYP2E1 substrate may be reduced in obese patients.

Adult↗
PubMed1994

Frequency distribution of dapsone N-hydroxylase, a putative probe for P4503A4 activity, in a white population.

Phenotypic trait values in 166 healthy white subjects (age range, 18 to 88 years) were determined for dapsone N-hydroxylation, dapsone N-acetylation, debrisoquin 4-hydroxylation, and S-mephenytoin 4'-hydroxylation after single oral dose administration of the probe drugs dapsone (100 mg), debrisoquin (10 mg), and mephenytoin (100 mg). No associations or evidence of cosegregation were found between the individual routes of metabolism. Dapsone N-hydroxylation exhibited a unimodal distribution, with marked (tenfold) intersubject variability, and aging was associated with reduced N-oxidation. However, the other measured routes of metabolism were age independent, but intersubject variability in all of the trait measurements increased with age. In subjects younger than 50 years, S-mephenytoin 4'-hydroxylation was modestly (34%) less in men than in women. In contrast, dapsone N-acetylation, dapsone N-hydroxylation, and debrisoquin 4-hydroxylation were not influenced by gender. Previous smoking habit and alcohol consumption were not associated with a difference in any of the four routes of metabolism. Accordingly, the measured phenotypic traits of drug oxidation and N-acetylation appear to be quite robust in regard to some common demographic variabilities present in population studies, with the exception of dapsone N-hydroxylase, which is affected by aging.

Acetylation↗
PubMed1994

Correlation of polymorphic expression of CYP2D6 mRNA in bladder mucosa and tumor tissue to in vivo debrisoquine hydroxylase activity.

Debrisoquine hydroxylase activity has been attributed to CYP2D6 and poor metabolizers of debrisoquine have a reduced relative risk of developing aggressive bladder cancer. Production of a proximate carcinogen could occur in liver or bladder mucosa. However, it is not known if CYP2D6 is expressed in human bladder mucosa. In vivo whole body debrisoquine hydroxylase activity was measured as the debrisoquine recovery ratio (DBRR) following single dose oral administration of debrisoquine (10 mg) in 10 normal subjects and 20 patients with bladder cancer prior to diagnostic cystoscopy. Semi-quantitative PCR was used to measure mRNA for CYP2D6 in bladder tissue obtained at cystoscopy. Of the 30 subjects, three were phenotypically and genotypically poor metabolizers. Among the extensive metabolizers, there were extensive intersubject variations in DBRR. A 10-fold variation in CYP2D6 mRNA levels was observed in bladder tissue. There was a highly significant association between DBRR and CYP2D6 mRNA expression (r2 = 0.702, P < 0.001). These results demonstrate the presence of CYP2D6 mRNA in bladder mucosa. Furthermore, they are consistent with debrisoquine hydroxylation being mediated by CYP2D6 and suggest that differences in mRNA concentration are rate limiting for enzyme activity and that bladder mucosal regulation reflects total body regulation for this enzyme. The expression of CYP2D6 in bladder mucosa suggests that this enzyme could be involved in the local production of a proximate carcinogen in this tissue and contribute to the pathogenesis of bladder cancer in man.

Base Sequence↗
PubMed1994

Low activity of dapsone N-hydroxylation as a susceptibility risk factor in aggressive bladder cancer.

N-arylamines involved in the pathogenesis of bladder cancer, require metabolic activation via N-hydroxylation. The efficiency of in vivo N-hydroxylation of dapsone, a non-carcinogenic arylamine, may, therefore, provide a host susceptibility measure of risk of developing bladder cancer. To investigate this possibility, the dapsone recovery ratio, a phenotypic measure of the efficiency of dapsone hydroxylation, has been measured in a case control study in an urban UK population, comparing patients with aggressive bladder cancer (n = 33), non-aggressive bladder cancer (n = 60) and controls (n = 108). Dapsone recovery ratio in controls exhibited a unimodal distribution. Patients with aggressive bladder cancer had a similar distribution but significantly lower mean value (p < 0.005). Logistic regression analysis, controlling for sex, age, smoking habit and alcohol consumption confirmed a significant (p < 0.05) association between the dapsone recovery ratio and aggressive bladder cancer. Subjects in the lowest tertile of dapsone recovery ratio had a relative risk to 5.4-fold greater than subjects in the upper tertile (p < 0.009), and a trends test was significant (p < 0.001). There was no significant association between dapsone recovery ratio and non-aggressive bladder cancer. These results do not support the hypothesis that the drug metabolizing enzymes involved in dapsone N-hydroxylation are involved in causing bladder cancer. Instead, they suggest the opposite, the observation that low enzyme activity was associated with increased risk is consistent with this enzyme providing a detoxification mechanism for environmental procarcinogens.

Case-Control Studies↗
PubMed1994

Identification of a new genetic defect responsible for the polymorphism of (S)-mephenytoin metabolism in Japanese.

A genetic polymorphism in the metabolism of the anticonvulsant drug (S)-mephenytoin has been well documented in humans. There are marked interracial differences in the frequency of the poor metabolizer phenotype, which comprises 2-5% of Caucasian but 18-23% of Asian populations. We have recently reported that the principal genetic defect responsible for the poor metabolizer phenotype is a single-base pair mutation in exon 5 of CYP2C19 (CYP2C19m), which accounts for approximately 75-83% of the defective alleles in both Japanese and Caucasians subjects. In the present study, we have identified a new mutation (CYP2C19m2) in Japanese poor metabolizers, consisting of a guanine to adenine mutation at position 636 of exon 4 of CYP2C19, which creates a premature stop codon. Genotyping of seven Japanese poor metabolizers who were not homozygous for the previously described CYP2C19m defect (now designated CYP2C19m1) indicated that they were either homozygous for the new defect (CYP2C19m2/CYP2C19m2) or heterozygous (CYP2C19m1/CYP2C19m2) for the two defects. CYP2C19m1 accounts for 25 of 34 alleles in Japanese poor metabolizers, whereas CYP2C19m2 accounts for the remaining nine alleles. Hence, CYP2C19m1 and CYP2C19m2 explain 100% of the available Japanese poor metabolizers (34 alleles). In contrast, the CYP2C19m2 defect was not detected in nine Caucasian poor metabolizers (83% of available poor metabolizer alleles were CYP2C19m1), indicating the existence of another, as yet unidentified, mutation. Genetic testing of the families of two Japanese poor metabolizer probands showed that coinheritance of the CYP2C19m1 and CYP2C19m2 alleles was concordant with the autosomal recessive inheritance of the poor metabolizer phenotype.

Amino Acid Sequence↗
PubMed1994

Stereoselective disposition of hexobarbital and its metabolites: relationship to the S-mephenytoin polymorphism in Caucasian and Chinese subjects.

In vitro studies with human liver preparations suggest that the metabolism of hexobarbital involves CYP2CMP--the determinant of the S-mephenytoin 4'-hydroxylation polymorphism, but no in vivo evidence of interphenotypic differences exist. The pharmacokinetics and urinary excretion of hexobarbital and its metabolites were, therefore, investigated following oral administration of a differentially labelled pseudoracemate that allowed determination of the fate of the individual enantiomers. Studies were undertaken in 10 Caucasian and nine Chinese healthy subjects known to be either extensive (EM) or poor (PM), metabolizers of mephenytoin. No inter-racial differences were observed in any of the measured parameters within a given phenotype. However, pronounced stereoselectivity in disposition was noted in EMs with R-(-)-hexobarbital's oral clearance being five- to six-fold greater than that for the S-(+)-enantiomer. By contrast, the S-(+)-isomer was eliminated twice as fast as R-(-) hexobarbital in PMs and, in addition, the oral clearances of both enantiomers were significantly reduced compared with their values in EMs. Formation of 3'-hydroxy- and 3'-ketohexobarbital and 1,5-dimethylbarbituric acid were the major identified routes of metabolism for each enantiomer in both phenotypes. Furthermore, these pathways were found to co-segregate with the mephenytoin polymorphism and in EMs they were primarily responsible for the observed stereoselectivity in disposition. These findings, therefore, confirm the stereoselectivity in hexobarbital's disposition in humans and identify the major pathways of metabolism involved. Additionally, the results indicate that CYP2CMP is a major determinant of the in vivo metabolism of both of hexobarbital's enantiomers but especially that of the R-(-)-enantiomer.

Administration, Oral↗