Juliette Säwe

Importance of genetic factors in the regulation of diazepam metabolism: Relationship to S‐mephenytoin, but not debrisoquin, hydroxylation phenotype

Single oral 10 mg doses of diazepam and demethyldiazepam were given on different occasions to 16 healthy subjects. The subjects included four poor hydroxylators of debrisoquin and three poor hydroxylators of mephenytoin. There was a correlation between the total plasma clearance of diazepam and demethyldiazepam (rs = 0.83; p < 0.01). There was no relationship between benzodiazepine disposition and debrisoquin hydroxylation. Poor hydroxylators of mephenytoin had less than half the plasma clearance of both diazepam (p = 0.0008) and demethyldiazepam (p = 0.0001) compared with extensive hydroxylators of mephenytoin. The plasma half‐lives were longer in poor hydroxylators than they were in extensive hydroxylators of mephenytoin for both diazepam (88.3 ± SD 17.2 and 40.8 ± 14.0 hours; p = 0.0002) and demethyldiazepam (127.8 ± 23.0 and 59.0 ± 16.8 hours; p = 0.0001). There was no significant difference in volume of distribution of the benzodiazepines between the phenotypes. This study shows that the metabolism of both diazepam (mainly demethylation) and demethyldiazepam (mainly hydroxylation) is related to the mephenytoin, but not to the debrisoquin, hydroxylation phenotype.

Morphine pharmacokinetics and metabolism in humans : enterohepatic cycling and relative contribution of metabolites to active opioid concentrations

SummaryMorphine, morphine-6-glucuronide (M6G), morphine-3-glucuronide (M3G) and normorphine were analysed with high performance liquid chromatography in plasma and urine, collected over 72h after administration of single intravenous 5mg and oral 20mg doses of morphine to 7 healthy volunteers. Systemic plasma clearance of morphine was on average 21.1 ± 3.4 ml/min/kg (1.27 ± 0.20 L/h/kg), volume of distribution was 2.9 ± 0.8 L/kg and oral bioavailability was 29.2 ± 7.2%. Clearance of morphine to form M3G and M6G comprised 57.3% and 10.4%, respectively, and renal clearance comprised 10.9% of total systemic plasma clearance; hence, more than one-fifth of a dose (20.8%) remained as unidentified residual clearance. On the basis of the area under the plasma concentration-time curves determined after oral and intravenous administration, the ratios of M6G: morphine were 3.6 ± 1.2 and 0.7 ± 0.3, respectively. The corresponding figures for M3G: morphine were 29.9 ± 6.8 and 7.7 ± 1.4. Differences in metabolic ratios between the parenteral and oral routes could be attributed solely to differences in morphine concentrations as evidenced both by plasma concentrations and amounts excreted in urine. An oral: parenteral potency ratio of 1: 3 may, thus, be due to differences in circulating amounts of morphine since the proportions of an administered dose found as M6G and M3G after administration by both routes were equal.A major finding was a slowly declining terminal phase of morphine and metabolites that was evident both in plasma and in urinary excretion versus time curves, where the half-lives of morphine, M3G and M6G were 15.1 ± 6.5h, 11.2 ± 2.7h and 12.9 ± 4.5h, respectively. The terminal half-life of normorphine was 23.9 ± 10.1 h after oral administration. Comparison of oral with intravenous excretion curves showed that a greater part of morphine and metabolites were excreted during the slowly declining phase after the oral dose than the intravenous dose, which is highly suggestive of enterohepatic cycling. The renal clearance of M6G and morphine was seen to exceed creatinine clearance, possibly due to an active secretion process.

E- and Z-10-hydroxylation of nortriptyline: Relationship to polymorphic debrisoquine hydroxylation

Eight healthy subjects [who were phenotyped with a debrisoquine (D) hydroxylation test] were selected to cover a wide range in the ratio between D and 4‐hydroxydebrisoquine (4‐OH‐D) in the urine. After a single oral dose of nortriptyline (NT) the metabolic clearance by 10‐hydroxylation in the E‐position, but not in the Z‐position, correlated closely to the metabolic ratio D/4‐0H‐D (rs = ‐0.88, p < 0.01). This indicates that common enzymatic mechanisms are involved in the hydroxylation of D and the E‐ but not the Z‐10‐hydroxylation of NT. Slow hydroxylators of NT and D excreted less 10‐hydroxynortriptyline in urine and had lower plasma clearance of NT than the rapid hydroxylators. The strong correlation (r = 0.96) between the total plasma clearance of NT and the metabolic clearance by E‐10‐hydroxylation shows that this metabolic reaction is important in the disposition of the drug.

Morphine kinetics in cancer patients.

Oral and intravenous morphine kinetics were studied in seven patients with cancer who needed continuous treatment with morphine because of severe chronic pain. Single oral (20 to 30 mg) and intravenous (4 mg) doses were given on separate days, followed by repetitive blood sampling for morphine analysis by gas chromatography. Volume of distribution ranged from 0.95 to 3.75 l/kg and serum clearance from 5.0 to 16.1 ml/min/kg. Oral morphine in doses that were more than five times the intravenous dose gave concentrations (at 10 and 120 min after dose) between 38 and 112 ng/ml. During the 0.25‐ to 8‐hr period after the oral dose serum concentrations were higher than after the intravenous dose. There was a variation in oral bioavailability of 15% to 64% and an interindividual variation in terminal half‐life from 58 to 467 min. These data warrant careful adjustment of the oral dose under close supervision of the patient at the onset of therapy.

Polymorphic debrisoquin hydroxylation in 757 Swedish subjects

The metabolic ratios (MRs) between debrisoquin and 4‐hydroxydebrisoquin in urine after a single oral dose of 10 mg debrisoquin were bimodally distributed in 757 healthy, white Swedish volunteers. Forty‐one subjects (5.4%) had an MR >12.6 and were classified as slow debrisoquin hydroxylators. The MR was reproducible in urine stored at + 8° C for 1 week and at − 20° C over a period of 5 yers. Collection intervals of 6 or 12 hours gave the same MR. Intraindividual repeatability of the debrisoquin phenotyping test was established in 37 subjects examined twice at least 2 weeks apart. The calculated frequency of the single allele that is believed to control deficient debrisoquin hydroxylation is similar among white Swedish people, as among other white groups examined so far; however, it is significantly different from the frequency in certain Oriental groups. Detailed comparisons of the prevalence of slow debrisoquin hydroxylation in different ethnic groups are not possible due to shortcomings in current epidemiologie techniques used (small materials, the location of the antimode distinguishing rapid and slow hydroxylators unknown, and family studies missing).

Morphine glucuronidation in human fetal and adult liver

SummaryThe glucuronyltransferase activity towards morphine was measured in microsomes isolated from liver specimens obtained from human fetuses and cancer patients. All the fetal livers investigated had measurable UDP-glucuronyltransferase activity towards morphine. There was no correlation between the gestational age (15 to 27 weeks) and the glucuronidation rate. The mean value of the enzymatic activities was higher in fetal livers obtained by hysterotomy (0.20 nmoles×min−1×mg−1) than in livers obtained after induced abortion (0.11 nmoles×min−1×mg−1). The average rate of glucuronidation in microsomes from adult liver (mean 1.15 nmoles×mint-1×mg−1) was 6 to 10 times higher than in the fetal liver microsomes. Together with previous investigations on human adult and fetal liver glucuronidation, the present results support the theory of heterogeneity of human UDP-glucuronyltransferase.

Nortriptyline and antipyrine clearance in relation to debrisoquine hydroxylation in man

Abstract Eight healthy volunteers with a broad range in their ability to hydroxylate debrisoquine, a metabolic reaction which is under polymorphic genetic control, were given single oral doses of nortriptyline and antipyrine. There was a close correlation between an individuals ability to hydroxylate debrisoquine and nortriptyline (measured as plasma clerance of the drug) (Spearman rs = 0.83; p = 0.01). This indicates that these two benzylic hydroxylations are controlled by similar if not identical genetic factors. In contrast to nortriptyline the clearance of antipyrine (assessed in saliva) was not positively related to the ability to hydroxylate debrisoquine.

Genetic analysis of the CYP2D locus in relation to debrisoquine hydroxylation capacity in Korean, Japanese and Chinese subjects.

The CYP2D6 genotype and the debrisoquine and mephenytoin hydroxylation phenotypes were studied in 63 Oriental subjects including 21 Chinese, 21 Japanese and 21 Koreans. All subjects were extensive metabolizers of debrisoquine. The incidence of the S-mephenytoin poor metabolizer phenotype was 14% in the Chinese, 24% in the Japanese and 24% in the Korean population, respectively, which is similar to previous reports. The CYP2D6 genotype was analysed by Xba I and Eco RI RFLP, and by allele-specific PCR analysis for the presence of several allelic variants of the CYP2D locus. No CYP2D6A or CYP2D6B alleles, two of the most common defect alleles among Caucasians, were found among the Oriental subjects. The frequency of the CYP2D6D allele was similar to that in Caucasian populations and consistent with the low incidence of the poor metabolizer phenotype in all three Oriental populations. The CYP2D6L2-allele with duplication of an active CYP2D6L gene was identified in one Korean and one Chinese allele in association with high CYP2D6 activity. The CYP2D6Ch alleles CYP2D6Ch1 and Ch2, identified by RFLP and PCR for the -1338C-->T and 188C-->T mutations, were the most frequent allelic variants in all three populations studied, and were related to a decreased CYP2D6 activity as previously shown in Chinese. In conclusion, the present pilot study revealed major similarities in the polymorphic CYP2D locus between Korean, Japanese and Chinese populations.

Genetic analysis of the interethnic difference between Chinese and Caucasians in the polymorphic metabolism of debrisoquine and codeine

SummaryThe Far Eastern and Caucasian populations are strikingly different with respect to the debrisoquine/sparteine hydroxylation polymorphism. The number of poor metabolizers, as defined for Caucasians, is very low among Chinese and Japanese. We investigated the molecular basis for this difference by analysis of the CYP2D6 gene in 115 Chinese subjects, combined with phenotypic classification of codeine and debrisoquine metabolism.A correlation between the rates of metabolism of these two drugs and genotype, as analyzed by RFLP using XbaI, was observed among the Chinese. A high frequency (37%) of alleles indicative of gene insertions (reflected by Xba I 44kb fragments) was recorded in the Chinese, but was not associated with the poor metabolizer phenotype, as it is in Caucasians. PCR amplification of part of the CYP2D6 gene with mutation specific primers for CYP2D6A (29A) and CYP2D6B (29B) allelic variants revealed that the XbaI 44kb fragment in Chinese apparently contains a functional CYP2D6 gene, in contrast to the situation among Caucasians.The results provide a molecular explanation of the interethnic difference in the metabolism of drugs affected by the debrisoquine hydroxylation polymorphism.

Amitriptyline metabolism: Association with debrisoquin hydroxylation in nonsmokers

Eleven healthy nonsmokers with wide variation in the ability to hydroxylate debrisoquin (D) were given single oral doses of amitriptyline and nortriptyline on different occasions. The urinary D/4‐hydroxy‐D ratio correlated significantly (P < 0.01) with all three parameters of amitryptyline disposition measured (total plasma clearance, clearance by demethylation, and clearance by pathways other than demethylation), with rs= − 0.89, − 0.78, and − 0.83, respectively. In contrast, we failed to demonstrate such correlations in a previous sample of smokers. Our data suggest that there may be a common regulation of the hydroxylation of D and the oxidative metabolism of amitriptyline in nonsmokers. It is hypothesized that an additional demethylase/hydroxylase is induced in smokers that is not involved in D hydroxylation.