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Biomedical subjects

W Kalow

Publications and source records attributed to W Kalow.

At least 19 recordsLinked to original sources
PubMed1995

Pharmacokinetics of dextromethorphan and metabolites in humans: influence of the CYP2D6 phenotype and quinidine inhibition.

Dextromethorphan is primarily metabolized to dextrorphan by cytochrome P450 2D6 (CYP2D6), a genetically polymorphic enzyme in humans. Dextrorphan is an active metabolite that produces phencyclidine-like behavioral effects in animals and exhibits anticonvulsant and neuroprotective properties in a variety of experimental models. In these studies, we examined the effects of CYP2D6 phenotype and quinidine inhibition on the pharmacokinetics of dextromethorphan and its metabolites in humans. After a single oral dose of dextromethorphan HBr (30 mg), the major metabolites in the plasma of extensive metabolizers (N = 5) were conjugated dextrorphan and conjugated 3-hydroxymorphinan. Free dextrorphan concentrations were about 100-fold less than the conjugated dextrorphan, and dextromethorphan was not detectable. Pretreatment of these subjects with 100 mg of quinidine, a selective inhibitor of CYP2D6, significantly suppressed the formation of dextrorphan and elevated the concentrations of dextromethorphan (t1/2, 16.4 hours). In poor metabolizers (N = 4) given the same dose, dextromethorphan was the major component in the plasma with a t1/2 of 29.5 hours. Present at concentrations 5- to 10-fold less were conjugated dextrorphan and the other two metabolites. Urinary recovery studies indicated that the inhibition by quinidine was reversible and that the elimination of dextromethorphan primarily depends on CYP2D6 activity rather than renal elimination. These data demonstrated that the CYP2D6 phenotype and the concurrent administration of quinidine significantly affect the disposition of dextromethorphan and the formation of the active metabolite dextrorphan and are important factors to be considered in studies of the pharmacologic and behavioral effects of dextromethorphan.

Adult↗
PubMed1995

Effects of route of administration on dextromethorphan pharmacokinetics and behavioral response in the rat.

One of the potential problems of using dextromethorphan as a neuroprotective or anticonvulsant agent is the phencyclidine-like behavioral effects that have been attributed to its major metabolite dextrorphan. Because previous behavioral studies of dextromethorphan have generally failed to consider metabolic conversion to this metabolite, the present studies were conducted to examine the effects of route of administration on dextromethorphan pharmacokinetics and locomotor activity in the rat. The bioavailability of dextromethorphan was 1.3-fold lower and the formation of dextrorphan and other metabolites was 3-fold greater after i.p. injection of 30 mg/kg of dextromethorphan as compared to the s.c. route, indicating substantial effect of first-pass metabolism. Plasma dextromethorphan was correlated with brain dextromethorphan (r = 0.84, P < .001), and the brain/plasma concentration ratio was about 6.5. Plasma-free dextrorphan, but not conjugated dextrorphan, was correlated with brain dextrorphan (r = 0.97, P < .001). Tmax of brain dextrorphan was earlier, and Cmax was higher after i.p. injection of dextromethorphan than s.c. administration (60 min vs. 120 min and 1.0 nmol/g vs. 0.2 nmol/g). Dextromethorphan (60 mg/kg i.p.) increased locomotor activity in the rat 60 min postdose, whereas the same dose of dextromethorphan administered by s.c. injection was without effect. These data demonstrate the route-specific effects on the disposition of dextromethorphan and dextrorphan in rat plasma and brain, as well as the behavioral consequence of the difference.

Animals↗
PubMed1992

Variability of acetaminophen metabolism in Caucasians and Orientals.

Acetaminophen (paracetamol) is extensively conjugated with glucuronic acid and sulfate prior to renal excretion. A minor metabolic route involves microsomal oxidation of acetaminophen to a hepatotoxic reactive intermediate, which subsequently undergoes glutathione (GSH) conjugation, yielding cysteine and mercapturate conjugates, both of which are excreted in the urine (Slattery et al., 1987). Data collected by de Morais et al. (1989) indicated that in comparison with normal subjects, glucuronidation of acetaminophen was impaired in subjects with Gilbert's syndrome, a genetically-based impairment of bilirubin glucuronidation. Thus, inter-subject and ethnic differences in acetaminophen disposition have pharmacogenetic and toxicological implications. This study was conceived to explore these differences. Urinary excretion of acetaminophen and its metabolites was observed in 125 Caucasian and 33 Oriental subjects. No appreciable difference was noted in the mean fraction of drug excreted as glucuronide between the two groups (51.5% in Caucasians vs 51.8% in Orientals). However, the data strongly indicated that the excretion of acetaminophen glucuronide was not normally distributed. Bimodality was apparent in both groups, with 20% of Caucasian and 33% of Oriental subjects displaying relatively extensive glucuronidation. In addition, glucuronidation displayed a strong negative correlation with sulfation (r = -0.97), suggesting the existence of a compensatory mechanism between the two metabolic pathways. The mean fractional excretions of cysteine and mercapturate conjugates did show significant differences between Caucasians and Orientals (p < 0.005). In addition, the ratio of mercapturate to total GSH-derived conjugates recovered appeared to be bimodal, indicating possible heterogeneity in the conversion of the cysteine conjugate to mercapturate via N-acetylation.

Acetaminophen↗
PubMed1992

Biotransformation of caffeine, paraxanthine, theobromine and theophylline by cDNA-expressed human CYP1A2 and CYP2E1.

Six human cytochrome P450s expressed in HepG2 cells using vaccinia virus cDNA-directed expression, were used to study the biotransformation of caffeine and its metabolites. CYP1A2 alone was responsible for caffeine 3-demethylation and paraxanthine 7-demethylation; in addition, 1A2 catalysed virtually all reactions related to caffeine and its metabolites. The metabolic profile of caffeine biotransformation by CYP1A2 averaged 81.5% for paraxanthine, 10.8% for theobromine and 5.4% for theophylline formation. It remained quite uniform when caffeine concentrations were varied. The most striking finding was that CYP2E1 (the ethanol-inducible form) had major influences upon caffeine metabolism: in particular, it catalysed the formation of theophylline and theobromine from caffeine. Thus, the in vivo metabolite profiling of caffeine may reveal CYP2E1 activities in addition to the previously documented activities of CYP1A2, polymorphic N-acetyltransferase and xanthine oxidase.

Biotransformation↗
PubMed1992

Diazepam metabolism by rat and human liver in vitro: inhibition by mephenytoin.

1. Diazepam metabolism and its association with mephenytoin hydroxylase were studied in vitro using human and rat livers. 2. Enzyme kinetic parameters were obtained for the formation of p-hydroxydiazepam (p-hydroxy-DZP), N-desmethyldiazepam (NDZ), and temazepam (TMZ) from diazepam (DZP) in rat liver fractions. The Km values for formation in rat of p-hydroxy-DZP, NDZ and TMZ were 14 +/- 3 (SEM) microM, 44 +/- 4 and 63 +/- 8, respectively; clearance values calculated from Vmax/Km were 5.7, 3.2 and 4.9 ml/g per min, respectively. 3. Mephenytoin (MP) competitively inhibited, in rat liver, the formation of NDZ, but not the formation of p-hydroxy-DZP or TMZ; in human liver neither NDZ nor TMZ formation was inhibited by MP. 4. In seven different human livers the formation of p-hydroxy-DZP represented a minor pathway compared to the formation of NDZ and TMZ.

Animals↗
PubMed1991

Intramuscular desferrioxamine in patients with Alzheimer's disease.

Although epidemiological and biochemical evidence suggests that aluminium may be associated with Alzheimer's disease (AD), there is no convincing proof of a causal link for aluminium in disease progression. We have completed a two year, single-blind study to investigate whether the progression of dementia could be slowed by the trivalent ion chelator, desferrioxamine. 48 patients with probable AD were randomly assigned to receive desferrioxamine (125 mg intramuscularly twice daily, 5 days per week, for 24 months), oral placebo (lecithin), or no treatment. No significant differences in baseline measures of intelligence, memory, or speech ability existed between groups. Activities of daily living were assessed and videorecorded at 6, 12, 18, and 24 month intervals. There were no differences in the rate of deterioration of patients receiving either placebo or no treatment. Desferrioxamine treatment led to significant reduction in the rate of decline of daily living skills as assessed by both group means (p = 0.03) and variances (p less than 0.04). The mean rate of decline was twice as rapid for the no-treatment group. Appetite (n = 4) and weight (n = 1) loss were the only reported side-effects. We conclude that sustained administration of desferrioxamine may slow the clinical progression of the dementia associated with AD.

Aged↗
PubMed1991

Interethnic variation of drug metabolism.

There are many examples of differences between ethnically defined populations with regard to drug-metabolizing enzymes i.e. enzymes that serve as defences against exogenous chemicals. The prevalence of variants of enzymes such as monooxygenases, dehydrogenases, esterases and some transferases is not uniform, and this leads to differences in metabolism of drugs such as aspirin and diazepam, among others. Werner Kalow explains why the mere counting of a genetic variant may not be sufficient to uncover more major problems such as may be posed by multiple allelism or heterozygosity, and why it may be predicted that a large number of drugs will display interethnic differences in function.

Alleles↗
PubMed1991

Use of caffeine metabolite ratios to explore CYP1A2 and xanthine oxidase activities.

Caffeine was used as a metabolic probe to screen healthy subjects for their activities of two enzymes, deduced to be CYP1A2 (an inducible cytochrome P450) and xanthine oxidase. A longitudinal study revealed modest effects of caffeine dose, ethanol intake, and time-of-day on the CYP1A2 index, without any effect on the xanthine oxidase index. The coefficients of intraindividual variation not accounted for were 5.0% for the xanthine oxidase and 17.2% for the CYP1A2 index. In a population study, both indexes showed a log normal distribution, with CYP1A2 values of most subjects covering a 6.3-fold range but only a 1.7-fold range with xanthine oxidase. The CYP1A2 index was 33% decreased in women who used oral contraceptives and substantially increased in cigarette smokers. Neither the CYP1A2 nor the xanthine oxidase index differed between volunteers of Chinese and European extraction. Four of 178 subjects showed unexplained low xanthine oxidase values (i.e., values several standard deviations below the mean).

Adult↗
PubMed1991

Caffeine as a metabolic probe: exploration of the enzyme-inducing effect of cigarette smoking.

It has been realized recently that the primary metabolism of caffeine in humans is catalyzed by P-450IA2 and that the rate of caffeine metabolism can be estimated from a metabolic ratio in a single urine sample. A population of 178 students including 19 smokers were subjected to this caffeine test to establish their P-450IA2 index. Both stated numbers of cigarettes smoked per day and urinary cotinine levels as a confirmatory measure correlated significantly with enzyme activity showing dose-effect relationships (r = 0.62 and 0.89, respectively). Nevertheless, more nonsmokers than smokers had the highest enzyme indexes, suggesting that dietary elements or other factors may determine P-450IA2 activities in populations. Because P-450IA2 is a monooxygenase that may be confined to the liver, caffeine reveals directly the Ah-receptor-dependent enzyme induction only in the liver, but it may also be a signal of induction elsewhere.

Caffeine↗
PubMed1991

Caffeine as a metabolic probe: validation of its use for acetylator phenotyping.

The use of two caffeine metabolite ratios for acetylator phenotyping was validated by demonstrating concordance with two sulfamethazine tests in 178 unrelated healthy subjects. The caffeine metabolites used for this purpose were 5-acetylamino-6-amino-3-methyluracil (AAMU), 1-methylxanthine (1X), and 1-methylurate (1U). The ratio AAMU/(AAMU + 1X + 1U), referred to as molar ratio or N-acetyltransferase, was compared with the ratio AAMU/1X. The results indicated that, for screening purposes, the acetylator phenotype can be determined by analysis of a 6-hour urine sample after a cup of coffee or strong tea or a can of caffeine-containing soft drink. The ratio AAMU/1X is the ratio of choice for the study of subjects in whom variability of xanthine oxidase can be neglected; use of the ratio AAMU/(AAMU + 1X + 1U) appears appropriate for special purposes. Gender, ethnic origin, habitual or moderate consumption of coffee, tea, soft drinks, or ethanol, or cigarette smoking have little if any effect on the caffeine tests for acetylator phenotyping.

Acetylation↗
PubMed1991

Identification of a new variant CYP2D6 allele lacking the codon encoding Lys-281: possible association with the poor metabolizer phenotype.

A variant CYP2D6(C) P450 protein was found in a liver characterized by deficient microsomal metabolism of bufuralol and sparteine, prototypical substrates for the debrisoquine-sparteine drug oxidation polymorphism. This protein was present at decreased levels in liver and had a slightly different relative mobility on SDS-polyacrylamide gels. The cDNA cloning and sequencing of the variant, designated CYP2D6(C), revealed that its mRNA lacked a single codon resulting in deletion of Lys281. This was the result of a three base pair deletion at the 3' end of CYP2D6 exon 5. The CYP2D6(C) P450, produced in HepG2 cells using vaccinia virus mediated cDNA expression displayed Km values toward bufuralol, debrisoquine and sparteine that were not significantly different from wild type CYP2D6. These data suggest that the poor metabolizer phenotype in livers expressing CYP2D6(C) is not due to a catalytically defective enzyme but perhaps due to decreased levels of the P450 protein in microsomal membranes. Low microsomal CYP2D6(C) contents could result from deficient membrane insertion or decreased stability of the P450 protein. A polymerase chain reaction-based procedure, developed to detect CYP2D6(C) alleles, indicates that this variant probably represents less than 1.5% of all CYP2D6 alleles.

Alleles↗
PubMed1991

Pharmacogenetics of caffeine and caffeine-halothane contractures in biopsies of human skeletal muscle.

In vitro pharmacological responses of fresh biopsy specimens of human skeletal muscle were used as indicators of some intrinsic muscle properties. The measured parameters that were utilized for the current study were contractures induced in vitro by caffeine or by caffeine plus halothane. The opportunity to study such specimens arose from clinical testing for diagnosing the genetic predisposition to malignant hyperthermia, a potentially fatal complication of anaesthesia. The current analysis covers data from over 1,000 subjects, most of whom were clinical suspects and relatives of these. Responsiveness of the muscle specimens varied over two orders of magnitude. The frequency distribution curves suggest that the variation does not represent a continuum but that there are three or more clusters of functional variants. Muscle specimens from males were on average more responsive to caffeine than were those from females. Correlations within father-son and brother-brother pairs indicated complete heritability of responsiveness; this might have been expected but the surprise was a lack of correlation within mother-daughter pairs. There was an intermediate correlation in father-daughter pairs. The sex difference in heritability could be due to gender-related modifying genes or due to secondary modification of the muscle response in females by sex-related, perhaps hormonal factors. Among the effects of age appeared to be poor development in early childhood of the potentiation of the caffeine contracture by halothane.

Adolescent↗
PubMed1991

Oxidation of reduced haloperidol to haloperidol: involvement of human P450IID6 (sparteine/debrisoquine monooxygenase).

1. The conversion of haloperidol (HAL) to reduced haloperidol (RHAL) and then back to HAL has been established in vivo and observed in psychiatric patients. The reduction of HAL to RHAL is known to be catalysed by a ketone reductase, while the nature of oxidation back to HAL is the subject of the present study. 2. We examined the in vitro oxidation of RHAL to HAL in human livers. The activity was microsomal and evidence is presented to suggest that the sparteine/debrisoquine metabolizing isoenzyme P450IID6 contributes to this oxidation. 3. Reciprocal inhibition studies between RHAL and sparteine, a specific substrate for cytochrome P450IID6, indicated that both compounds compete for the same binding site. Quinidine, the most specific inhibitor for this cytochrome P450 potently inhibited the oxidative conversion of reduced haloperidol to haloperidol. A significant correlation (rs = 0.62, P less than 0.01) was found between RHAL oxidation and sparteine oxidation in a study involving 17 human liver samples.

Cytochrome P-450 CYP2D6↗
PubMed1991

Neuronal cytochrome P450IID1 (debrisoquine/sparteine-type): potent inhibition of activity by (-)-cocaine and nucleotide sequence identity to human hepatic P450 gene CYP2D6.

Catalytic, pharmacological, and molecular criteria have been used to identify cytochrome P450IID1 in mammalian brain (enzyme, P450IID; gene, CYP2D). Sparteine metabolism in canine striatal membranes was shown to be inhibited in a concentration-dependent and stereoselective manner by quinidine (Ki, approximately 51 nM), quinine (Ki, approximately 5.9 microM), and various other known substrates and inhibitors of hepatic P450IID1 activity. In addition, canine striatal P450IID1 was inhibited with high affinity by dopamine uptake blockers, such as (-)-cocaine (Ki, approximately 74 nM), d-amphetamine (Ki, approximately 4.5 microM), and methylphenidate (Ki, approximately 15 microM). Inhibitory constants (Ki) of numerous compounds for inhibition of sparteine metabolism in canine striatal membranes correlated well with (a) Ki values observed in human liver microsomes (r = 0.95), (b) [3H]GBR-12935 binding to P450IID1 in canine striatal membranes (r = 0.85), and (c) the inhibition (IC50) of sparteine metabolism in HepG2 cells expressing human CYP2D6 cDNA (r = 0.93). Moreover, antibodies raised against rat hepatic enzyme inhibited, in a concentration-dependent manner, sparteine metabolism in canine striatal membranes. Enzymatic activity was unevenly distributed throughout the canine brain and ranged from 0.5 to 21 pmol/mg of protein/hr in cerebellum and supraorbital cortex, respectively, with the striatum displaying moderate levels of activity (8 pmol/mg of protein/hr). The polymerase chain reaction was used to amplify cDNA from a human caudate lambda gt11 library encoding exons 6-9 of the human CYP2D6 gene, which revealed, upon sequencing, 100% nucleic acid sequence identity. These data indicate that P450IID1 is expressed centrally and is similar, at the functional and molecular levels, to the human hepatic P450IID1 enzyme. Because the debrisoquine/sparteine mono-oxygenase is a polymorphic enzyme, in which 5-10% of caucasians are deficient in metabolism of various drugs, a genetic difference in human brain metabolism of P450IID1 substrates may possibly lead to differences in drug response and toxicity.

Animals↗
PubMed1990

The dopamine transporter and cytochrome P45OIID1 (debrisoquine 4-hydroxylase) in brain: resolution and identification of two distinct [3H]GBR-12935 binding proteins.

Two [3H]GBR-12935 binding proteins, identified as the dopamine transporter and cytochrome P45OIID1, were solubilized in digitonin from canine striatal membranes, and were resolved following wheat germ agglutinin (WGA)-lectin column chromatography. Protein adsorbed to and specifically eluted from WGA-lectin with N-acetylglucosamine displayed saturable, high affinity (KD approximately 3 nM), and sodium-dependent binding of [3H]GBR-12935, which was inhibited in a concentration-dependent and stereoselective manner by dopamine uptake blockers and substrates with a pharmacological profile indicative of the dopamine uptake site. Protein not adsorbed to WGA-lectin also bound [3H]-GBR-12935 with high affinity (approximately 7 nM), in a sodium-independent manner, and was insensitive to classical dopamine uptake blockers and substrates such as mazindol or dopamine, corresponding to the so-called "piperazine acceptor" site seen in native membranes. [3H]GBR-12935 binding to this latter protein was, however, inhibited by various compounds with a pharmacological profile indicative of a form of cytochrome P450 designated P45OIID1 (debrisoquine/sparteine monooxygenase) with the following rank order of inhibitory potency: GBR-12909 greater than budipine greater than alpha-lobeline greater than quinidine greater than alpha flupenthixol greater than SKF-525A greater than sparteine greater than quinine. Ki values obtained for inhibition of [3H]-GBR-12935 binding to neuronal WGA passthrough fractions by these drugs correlate well with their respective Ki values for liver P45OIID1 activity. Western blotting and immunoprecipitation analysis with rabbit anti-rat P45OIID1 antibody also supported the identity of the mazindol-insensitive [3H]GBR-12935 binding site (or piperazine acceptor site) as P45OIID1. Furthermore, a [3H]GBR-12935 binding protein with pharmacological and immunological characteristics similar to those of P45OIID1 was solubilized from both bovine and human liver membranes, and GBR-12909 was found to be a potent competitive inhibitor (Ki approximately 100 nM) of sparteine monooxygenase activity in human liver microsomes. These data clearly indicate that [3H]GBR-12935 and its analogs display similar affinities for both the dopamine transporter and neuronal P45OIID1, and that this radioligand may be a useful probe of P45OIID1 activity in brain and liver. The exact molecular and functional association (if any) between these two distinct binding protein populations remains to be established; however, it is tempting to speculate that P45OIID1 is involved in the catabolism and processing of neurotransmitters subsequent to their reuptake into target cells.

Animals↗
PubMed1990

Sparteine metabolism capacity in human liver: structural variants of human P450IID6 as assessed by immunochemistry.

An antibody raised against rat P450dbl was used to examine the heterogeneity of the human enzyme involved in the sparteine/debrisoquine polymorphism. The extent to which the antibody was able to inhibit sparteine metabolism varied in different human livers (10-80%, n = 9) and reflected the amount of sparteine metabolism carried out by the polymorphic P450IID6 in individual liver specimens. The individual sample variation in inhibition by the antibody correlated with the inhibition caused by quinidine, a prototype competitive inhibitor of the P450IID6 enzyme active site. Western immunoblots of the liver microsomes confirmed that the variation in the inhibition of sparteine metabolism by this antibody reflected the amount of P450IID6 protein. In addition, a detailed study of one of the livers (K19) which demonstrated a lack of inhibition by the antibody was performed which confirmed the lack of P450IID6 in this liver specimen and suggested that the nascent sparteine metabolism activity was due to other forms of P450.

Animals↗