Biomedical subjects

Matthias Schwab

Publications and source records attributed to Matthias Schwab.

At least 73 records · Page 4Linked to original sources

PubMed2004

Identification of budesonide and prednisone as substrates of the intestinal drug efflux pump P-glycoprotein.

Steroid resistance is a major problem in the management of patients with inflammatory bowel disease. In Crohn disease, poor response to corticosteroids has been related to increased expression of the drug efflux pump, P-glycoprotein. However, it has not been investigated thoroughly whether corticosteroids commonly used for drug therapy in inflammatory bowel disease are substrates of P-glycoprotein. We tested the hypothesis that budesonide and prednisone are substrates of P-glycoprotein thereby possibly contributing to variable therapeutic effects. Polarized, basal to apical transport of [3H]budesonide and [3H]prednisone was studied in monolayers of L-MDR1 cells (LLC-PK1 cells stably transfected with human MDR1 cDNA) and Caco-2 cells, both of which express P-glycoprotein in their apical membrane. Drug transport was measured during 4 hours at substrate concentrations of 5 microM. Net transport rates and permeability coefficients were calculated. Inhibition of P-glycoprotein-mediated transport across Caco-2 monolayers was determined after addition of the P-glycoprotein inhibitor PSC-833. The net transport rate from the basolateral to the apical side was significantly higher in L-MDR1 than in LLC-PK1 cells for both budesonide and prednisone. Apparent permeability coefficients of budesonide and prednisone reflected polarized transport from basal to apical. PSC-833 inhibited the polarized transport of both corticosteroids. In conclusion, budesonide and prednisone were identified as substrates of the intestinal drug efflux pump, P-glycoprotein. Therefore, drug secretion via P-glyco-protein into gut lumen might play a more important role in pharmacokinetics and pharmacodynamics of these corticosteroids than currently appreciated in gastroenterological practice.

ATP Binding Cassette Transporter, Subfamily B, Mem↗

PubMed2004

Comprehensive analysis of thiopurine S-methyltransferase phenotype-genotype correlation in a large population of German-Caucasians and identification of novel TPMT variants.

The thiopurine S-methyltransferase (TPMT) genetic polymorphism has a significant clinical impact on the toxicity of thiopurine drugs. It has been proposed that the identification of patients who are at high risk for developing toxicity on the basis of genotyping could be used to individualize drug treatment. In the present study, phenotype-genotype correlation of 1214 healthy blood donors was investigated to determine the accuracy of genotyping for correct prediction of different TPMT phenotypes. In addition, the influence of gender, age, nicotine and caffeine intake was examined. TPMT red blood cell activity was measured in all samples and genotype was determined for the TPMT alleles *2 and *3. Discordant cases between phenotype and genotype were systematically sequenced. A clearly defined trimodal frequency distribution of TPMT activity was found with 0.6% deficient, 9.9% intermediate and 89.5% normal to high methylators. The frequencies of the mutant alleles were 4.4% (*3A), 0.4% (*3C) and 0.2% (*2). All seven TPMT deficient subjects were homozygous or compound heterozygous carriers for these alleles. In 17 individuals with intermediate TPMT activity discordant to TPMT genotype, four novel variants were identified leading to amino acid changes (K119T, Q42E, R163H, G71R). Taking these new variants into consideration, the overall concordance rate between TPMT genetics and phenotypes was 98.4%. Specificity, sensitivity and the positive and negative predictive power of the genotyping test were estimated to be higher than 90%. Thus, the results of this study provide a solid basis to predict TPMT phenotype in a Northern European Caucasian population by molecular diagnostics.

Adolescent↗

PubMed2004

High plasma pravastatin concentrations are associated with single nucleotide polymorphisms and haplotypes of organic anion transporting polypeptide-C (OATP-C, SLCO1B1).

This study aimed to characterize possible relationships between polymorphisms in the drug transporter genes organic anion transporting polypeptide-C (OATP-C, SLCO1B1), OATP-B (SLCO2B1), multidrug resistance-associated protein 2 (MRP2, ABCC2) and multidrug resistance transporter (MDR1, ABCB1) and the pharmacokinetics of pravastatin. We studied 41 healthy Caucasian volunteers who had previously participated in pharmacokinetic studies with pravastatin. Six volunteers had a very high pravastatin AUC value and were defined as outliers according to statistical criteria. The OATP-C gene was sequenced completely in all subjects, and they were also genotyped for selected single nucleotide polymorphisms (SNP) in the OATP-B, MDR1 and MRP2 genes. Of the six outliers, five were heterozygous for the OATP-C 521T>C (Val174Ala) SNP (allele frequency 42%) and three were heterozygous for a new SNP in the promoter region of OATP-C (-11187G>A, allele frequency 25%). Among the remaining 35 subjects, two were homozygous and six were heterozygous carriers of the 521T>C SNP (allele frequency 14%, P = 0.0384 versus outliers) and three were heterozygous carriers of the -11187G>A SNP (allele frequency 4%, P = 0.0380 versus outliers). In subjects with the -11187GA or 521TC genotype, the mean pravastatin AUC0-12 was 98% (P = 0.0061) or 106% (P = 0.0034) higher, respectively, compared to subjects with the reference genotype. These results were substantiated by haplotype analysis. In heterozygous carriers of *15B (containing the 388A>G and 521T>C variants), the mean pravastatin AUC0-12 was 93% (P = 0.024) higher compared to non-carriers and, in heterozygous carriers of *17 (containing the -11187G>A, 388A>G and 521T>C variants), it was 130% (P = 0.0053) higher compared to non-carriers. No significant associations were found between OATP-B, MRP2 or MDR1 polymorphisms and the pharmacokinetics of pravastatin. These results suggest that haplotypes are more informative in predicting the OATP-C phenotype than single SNPs.

Adult↗

PubMed2004

Lipid-lowering response to statins is affected by CYP3A5 polymorphism.

Individuals expressing the polymorphic CYP3A5 enzyme might show a more than average efficiency in the metabolism of lovastatin, simvastatin and atorvastatin. We studied whether the expression of CYP3A5 is associated with an impaired lipid-lowering response to statins in 69 Caucasian patients. Lovastatin, simvastatin and atorvastatin were significantly less effective in CYP3A5 expressors than in non-expressors. The mean serum total cholesterol concentration at 1 year was 23% higher (P = 0.0014) and the mean serum low-density lipoprotein cholesterol concentration was 24% higher (P = 0.036) in subjects possessing the CYP3A5*1 allele (CYP3A5 expressors, n = 7) than in subjects homozygous for the CYP3A5*3 allele (non-expressors, n = 39). The mean percentage reduction in serum total cholesterol from baseline was significantly smaller in CYP3A5 expressors than in non-expressors (17% versus 31%, P = 0.026). No association between hypolipidemic efficacy and CYP3A5 polymorphism was observed among 23 subjects taking statins that are not dependent on CYP3A5 (fluvastatin, pravastatin). These findings suggest that CYP3A5 may be a genetic determinant of interindividual differences in response to certain statins.

Aged↗

PubMed2004

CYP2C19 polymorphism and proton pump inhibitors.

Proton pump inhibitors such as omeprazole (esomeprazole), lansoprazole, pantoprazole and rabeprazole are eliminated by the hepatic route and the polymorphic CYP2C19 is mainly involved in their metabolism. In different populations three phenotypes have been identified: extensive metabolizers, poor metabolizers and individuals carrying one wild type and one mutant allele (het extensive metabolizers). Systemic exposure to the proton pump inhibitors as expressed by the AUC (area under the plasma level time profiles) is 5-12-times higher in poor metabolizers than in extensive metabolizers. As the pharmacodynamic response (elevation of intragastric pH) to the proton pump inhibitors is related directly to their AUC, a much higher pH can be monitored over 24 hr in poor metabolizers than in extensive metabolizers. Furthermore, clinical efficacy of all proton pump inhibitors depend on maintaining intragastric pH above certain threshold levels and significantly higher eradication rates of Helicobacter pylori have been observed in patients of the poor metabolizers and het extensive metabolizers phenotype if compared to extensive metabolizers. Likewise, limited data suggest that proton pump inhibitors-induced healing rates in gastro-oesophageal reflux disease are apparently higher in poor metabolizers/het extensive metabolizers than in extensive metabolizers of CYP2C19. Therefore initial genotyping for this enzyme and higher dosage in extensive metabolizers is likely to improve the clinical efficacy of proton pump inhibitors.

Anti-Ulcer Agents↗

PubMed2004

[Rationale and methods of the UNAMET study (dose- and CYP2D6-genotype-dependent adverse drug reactions of metoprolol)--a contribution to quality improvement in pharmacotherapy].

Metoprolol has not yet been systematically studied in terms of quality of life and incidence of adverse drug reactions (ADRs). Metoprolol is metabolized by polymorphic CYP2D6, therefore poor CYP2D6 metabolizers may be at higher risk of ADRs. Therefore, it is to be proven whether genotyping is useful to guide initial dose selection. In the ongoing UNAMET study, nonrandomized out-patients start treatment with metoprolol for various disorders. With the use of standard questionnaires, the patients are prospectively evaluated for common ADRs (headache, dizziness, tiredness, sleep disturbances, dyspnea, cold extremities, sexual dysfunction) and quality of life. The questionnaires are filled out before and until 6 weeks after initiating therapy; blood pressure and heart rate are also measured. The acquired data are then related to the patients' metoprolol dose and plasma concentrations, as well as to their metabolic ratio of metoprolol/alpha-OH-metoprolol and CYP2D6 genotype.

Adrenergic beta-Antagonists↗

PubMed2003

Potent mechanism-based inhibition of human CYP2B6 by clopidogrel and ticlopidine.

The thienopyridine derivatives ticlopidine and clopidogrel are inhibitors of ADP-induced platelet aggregation. Pharmacological activity of these prodrugs depends on cytochrome P450 (P450)-dependent oxidation to the active antithrombotic agent. In this study, we investigated the interaction potential of clopidogrel and ticlopidine by using human liver microsomes and recombinantly expressed P450 isoforms. Both clopidogrel and ticlopidine inhibited CYP2B6 with highest potency and CYP2C19 with lower potency. Clopidogrel also inhibited CYP2C9, and ticlopidine also inhibited CYP1A2, with lower potency. Inhibition of CYP2B6 was time- and concentration-dependent, and as shown by dialysis experiments, it was irreversible and dependent on NADPH, suggesting a mechanism-based mode of action. Inactivation was of nonpseudo-firstorder type with maximal rates of inactivation (K(inact)) for clopidogrel and ticlopidine in microsomes (recombinant CYP2B6) of 0.35 (1.5 min(-1)) and 0.5 min(-1) (0.8 min(-1)), respectively, and half-maximal inactivator concentrations (KI) were 0.5 microM (1.1 microM) for clopidogrel and 0.2 microM (0.8 microM) for ticlopidine. Inhibition was attenuated by the presence of alternative active site ligands but not by nucleophilic trapping agents or reactive oxygen scavengers, further supporting mechanism-based action. A chemical mechanism is discussed based on the known metabolic activation of clopidogrel and on the finding that hemoprotein integrity of recombinant CYP2B6 was not affected by irreversible inhibition. These results suggest the possibility of drug interactions between thienopyridine derivates and drug substrates of CYP2B6 and CYP2C19.

Aryl Hydrocarbon Hydroxylases↗

PubMed2003

Enantioselective quantitation of the ecstasy compound (R)- and (S)-N-ethyl-3,4-methylenedioxyamphetamine and its major metabolites in human plasma and urine.

An enantioselective HPLC method has been developed and validated for the stereospecific analysis of N-ethyl-3,4-methylenedioxyamphetamine (MDE) and its major metabolites N-ethyl-4-hydroxy-3-methoxyamphetamine (HME) and 3,4-methylenedioxyamphetamine (MDA). These compounds have been analyzed both from human plasma and urine after administration of 70 mg pure MDE-hydrochloride enantiomers to four subjects. The samples were prepared by hydrolysis of the o-glucuronate and sulfate conjugates using beta-glucuronidase/arylsulfatase and solid-phase extraction with a cation-exchange phase. A chiral stationary protein phase (chiral-CBH) was used for the stereoselective determination of MDE, HME and MDA in a single HPLC run using sodium dihydrogenphosphate, ethylendiaminetetraacetic acid disodium salt and isopropanol as the mobile phase (pH 6.44) and fluorimetric detection (lambda(ex) 286 nm, lambda(em) 322 nm). Moreover, a suitable internal standard (N-ethyl-3,4-methylenedioxybenzylamine) was synthesized and qualified for quantitation purposes. The method showed high recovery rates (>95%) and limits of quantitation for MDE and MDA of 5 ng/ml and for HME of 10 ng/ml. The RSDs for all working ranges of MDE, MDA and HME in plasma and urine, respectively, were less than 1.5%. After validation of the analytical methods in plasma and urine samples pharmacokinetic parameters were calculated. The plasma concentrations of (R)-MDE exceeded those of the S-enantiomer (ratio R:S of the area under the curve, 3.1) and the plasma half time of (R)-MDE was longer than that of (S)-MDE (7.9 vs. 4.0 h). In contrast, the stereochemical disposition of the MDE metabolites HME and MDA was reversed. Concentrations of the (S)-metabolites in plasma of volunteers were much higher than those of the (R)-enantiomers.

3,4-Methylenedioxyamphetamine↗

PubMed2003

Telomerase activity in cell lines of pediatric soft tissue sarcomas.

Telomeres and their maintenance by telomerase have been implicated to play an important role in carcinogenesis. As almost all malignant tumors express telomerase (in contrast to normal somatic cells), assessment of its activity has been proposed as a diagnostic and prognostic tool. To test the prognostic value of telomerase in pediatric soft tissue sarcoma (STS), we analyzed telomere length (by telomere restriction fragment analysis), telomerase activity (by modified telomerase repeat amplification protocol assay), and expression of human telomerase reverse transcriptase (hTERT) mRNA (by TaqMan technique) in cell lines of different types of STS from 12 children and adolescents. Telomere length (3.7-9.0 kb) showed a very heterogeneous pattern, independent of subtype of STS or the age of the patients, and it was not associated with expression of hTERT mRNA. In contrast, there was a trend of an association between hTERT and telomerase activity. The three tested cell lines of embryonal rhabdomyosarcomas demonstrated no or low (n = 2) telomerase activity, which was confirmed in two cases by a very low expression of hTERT mRNA. Thus, we suggest that the significant difference (p < 0.01) in the less aggressive clinical behavior of embryonal rhabdomyosarcomas in comparison to other subtypes may be due to differences in telomerase expression. Taken together, our cell line experiments imply that telomerase activity might be a biologic marker for stratification between STS with different clinical prognosis.

Adolescent↗

PubMed2003

Sensitive method for the quantification of urinary pyrimidine metabolites in healthy adults by gas chromatography-tandem mass spectrometry.

Enzyme deficiencies in pyrimidine metabolism are associated with a risk for severe toxicity against the antineoplastic agent 5-fluorouracil. To assess whether urinary levels of pyrimidines and their metabolites can be used for predicting patients' individual phenotype, a new gas chromatographic-tandem mass spectrometric method was developed which allows the simultaneous determination of uracil and thymine and their metabolites dihydrouracil, dihydrothymine, beta-ureidopropionic acid, beta-ureidoisobutyric acid, and the amino acids beta-alanine and beta-aminoisobutyric acid in human urine. Small aliquots (2-20 microl) of the urine samples were evaporated and derivatized to the tert.-butyldimethylsilyl derivatives before quantification, using the respective stable isotope-labelled analogues as internal standards. Analytical variation was acceptable with an intra-day imprecision (RSD) below 10%, for beta-ureidoisobutyric acid below 15%. The method was used for investigating the stability of urine samples and the influence of urine collection at different times.

Adult↗

PubMed2003

Population pharmacokinetics of oral busulfan in children.

PURPOSE: To characterize the population pharmacokinetics of oral busulfan in 48 children including pooled data from three transplantation centres with the aim of estimating the variability in the kinetics of busulfan and to identify covariates that could be used for dose calculation. METHODS: A total of 508 plasma samples from 250 administrations (mean 9 samples per patient over 4 days of treatment) were collected from 48 children receiving busulfan orally every 6 h. The dosing varied between 13 and 20 mg/kg with seven patients receiving a dose of 600 mg/m(2). The busulfan formulations administered varied considerably. They included 2-mg tablets (Myleran), gelatine capsules, crushed tablets suspended in water and suspension for administration via nasogastric tube. Samples were analysed for busulfan either by HPLC using postcolumn photolysis or by LC-MS. Plasma concentration-time data were analysed by population pharmacokinetic modelling using NONMEM. RESULTS: Busulfan kinetics were best described by a one-compartment model (subroutine ADVAN 2 TRANS 2). Residual variability was modelled using a combined additive and proportional error model. The influence of different covariates on the pharmacokinetic parameters was tested. The best results were obtained by inclusion of body surface area (BSA) as a covariate for clearance (Cl/F) and volume of distribution (V/F). The final population estimates were: Cl/F 4.13 l/h per m(2) +/-26%, V/F 21.3 l/m(2) +/-31% and ka 1.31 h(-1) +/-110% (population mean +/- interindividual variability, IIV). Variability in one patient during the 4 days of treatment (interoccasion variability, IOV) for Cl/F (10%) and V/F (19%) were calculated to be less than interindividual variability, fulfilling the condition for individualization of busulfan dosage regimens. CONCLUSIONS: In our paediatric population, BSA, not body weight, is the best predictor of Cl/F and V/F. Our final estimations reflect the wide interpatient variability after oral administration of busulfan with an IIV for ka of 110%.

Administration, Oral↗

PubMed2003

Frequent co-occurrence of the TATA box mutation associated with Gilbert's syndrome (UGT1A128) with other polymorphisms of the UDP-glucuronosyltransferase-1 locus (UGT1A62 and UGT1A7*3) in Caucasians and Egyptians.

Polymorphisms of drug metabolizing enzymes are frequently associated with diseases and side effects of drugs. Recently, a TATA box mutation of UGT1A1 (UGT1A1*28), a common genotype leading to Gilbert's syndrome, and several missense mutations of other UDP-glucuronosyltransferase 1 (UGT1) family members have been described. Furthermore, co-occurrence of UGT1A1*28 and UGT1A6*2 has been observed. In order to elucidate the basis for co-occurrence of UGT1 mutations, fluorescence resonance energy transfer techniques were developed for rapid determination of polymorphisms of three UGT isoforms (UGT1A1*28, 1A6*2, and 1A7*2/*3). Hundred healthy Caucasians and 50 Egyptians were genotyped. All genotypes followed the Hardy-Weinberg equilibrium. Only three major haplotypes were found, including a haplotype consisting of allelic variants of all three isoforms (29% in Caucasians and 22% in Egyptians), all leading to reduced UGT activity. Frequent haplotypes containing several UGT1 allelic variants should be taken into account in studies on the association between diseases, abnormal drug reactions, and UGT1 family polymorphisms.

Egypt↗

PubMed2003

CYP2D6 genotyping strategy based on gene copy number determination by TaqMan real-time PCR.

The genetic polymorphism of the cytochrome P450 monooxygenase, CYP2D6, comprises at least 43 alleles giving rise to distinct drug metabolism phenotypes termed ultrarapid, extensive, intermediate, and poor metabolizers. As a consequence, drug side effects or lack of drug effect may occur if standard doses are applied. Genetic prediction of drug oxidation phenotype as a basis for dose selection requires analysis of single nucleotide polymorphisms and of alleles with duplicated or deleted genes. Here we developed a novel method to determine the CYP2D6 gene dose per genome. A TaqMan real-time PCR assay to specifically amplify genomic CYP2D6 was established by using a specific set of amplification primers and probe, located in exon 9, which effectively prevent amplification of CYP2D7 and CYP2D8 pseudogenes. Quantitative CYP2D6 amplification data were normalized to albumin as an internal reference gene which was coamplified simultaneously in a single-tube biplex assay. The assay was validated with a selection of previously genotyped DNA samples containing none, one, two, or three CYP2D6 gene copies. The results were highly reproducible and closely matched the number of genes with no overlap between the groups. Analysis of DNA samples comprising all major alleles and genotypes revealed high sensitivity and specificity of the assay, as demonstrated by agreement of the determined gene dose with the presence of CYP2D6(*)2 x 2 (gene duplication) and CYP2D6(*) 5 (gene deletion) alleles. The predictability of the new strategy was systematically evaluated. The semiautomatic TaqMan assay allows high sample throughput and will be useful for pharmacogenetic studies and in the clinical setting.

Amino Acid Substitution↗

PubMed2003

Expression and localization of the multidrug resistance protein 5 (MRP5/ABCC5), a cellular export pump for cyclic nucleotides, in human heart.

The multidrug resistance protein 5 (MRP5/ABCC5) has been recently identified as cellular export pump for cyclic nucleotides with 3',5'-cyclic GMP (cGMP) as a high-affinity substrate. In view of the important role of cGMP for cardiovascular function, expression of this transport protein in human heart is of relevance. We analyzed the expression and localization of MRP5 in human heart [21 auricular (AS) and 15 left ventricular samples (LV) including 5 samples of dilated and ischemic cardiomyopathy]. Quantitative real-time polymerase chain reaction normalized to beta-actin revealed expression of the MRP5 gene in all samples (LV, 38.5 +/- 12.9; AS, 12.7 +/- 5.6; P < 0.001). An MRP5-specific polyclonal antibody detected a glycoprotein of approximately 190 kd in crude cell membrane fractions from these samples. Immunohistochemistry with the affinity-purified antibody revealed localization of MRP5 in cardiomyocytes as well as in cardiovascular endothelial and smooth muscle cells. Furthermore, we could detect MRP5 and ATP-dependent transport of [(3)H]cGMP in sarcolemma vesicles of human heart. Quantitative analysis of the immunoblots indicated an interindividual variability with a higher expression of MRP5 in the ischemic (104 +/- 38% of recombinant MRP5 standard) compared to normal ventricular samples (53 +/- 36%, P < 0.05). In addition, we screened genomic DNA from our samples for 20 single-nucleotide polymorphisms in the MRP5 gene. These results indicate that MRP5 is localized in cardiac and cardiovascular myocytes as well as endothelial cells with increased expression in ischemic cardiomyopathy. Therefore, MRP5-mediated cellular export may represent a novel, disease-dependent pathway for cGMP removal from cardiac cells.

Aged↗

PubMed2003

Association between the C3435T MDR1 gene polymorphism and susceptibility for ulcerative colitis.

BACKGROUND & AIMS: The human multidrug resistance 1 (MDR1) gene product P-glycoprotein is highly expressed in intestinal epithelial cells, where it constitutes a barrier against xenobiotics. The finding that mdr1a knockout mice develop a form of colitis that is similar to ulcerative colitis, which can be prevented by antibiotics, indicates a barrier function for P-glycoprotein against the invasion of bacteria or toxins. Because the MDR1 single nucleotide polymorphism C3435T is associated with lower intestinal P-glycoprotein expression, we tested whether this polymorphism predisposes to development of ulcerative colitis. METHODS: Allele frequencies and genotype distributions of the C3435T single nucleotide polymorphism were investigated in 149 patients with ulcerative colitis, 126 patients with Crohn's disease, and sex-matched healthy controls. RESULTS: Significantly increased frequencies of the 3435T allele and the 3435TT genotype were observed in patients with ulcerative colitis compared with controls (3435T: P = 0.049; odds ratio, 1.4; 95% confidence interval, 1.02-1.94; 3435TT: P = 0.045; odds ratio, 2.03; 95% confidence interval, 1.04-3.95). In contrast, frequencies of the T allele and the TT genotype were the same in patients with Crohn's disease as in controls (P = 0.66 and P = 0.59, respectively). In comparison to 998 non-sex-matched controls, the effect for the TT genotype in ulcerative colitis patients was more pronounced (P = 0.0055; odds ratio, 2.1). CONCLUSIONS: The higher frequency of the 3435TT genotype in patients with ulcerative colitis corroborates the findings from the mdr1a knockout mice. The results support the notion that P-glycoprotein plays a major role in the defense against intestinal bacteria or toxins. Impairment of barrier function in 3435TT subjects could render this genotype more susceptible to the development of ulcerative colitis.

Adolescent↗

PubMed2003

Sex is a major determinant of CYP3A4 expression in human liver.

Many drugs that are substrates of CYP3A4, the major human drug-metabolizing cytochrome P450 (CYP), show higher clearance in women than in men. Although this effect is believed to be related to drug metabolism, the underlying cause has not been elucidated. We investigated CYP3A4 in a large collection (n = 94) of well-characterized surgical liver samples and found 2-fold higher CYP3A4 levels in female compared with male samples (P <.0001) and a corresponding 50% increase in the CYP3A-dependent N-dealkylation of verapamil (P <.01). This expression difference was not due to preferential induction in women following higher drug exposure because it was even larger in a subgroup not previously exposed to drugs. Higher expression in women was also found for CYP3A4 messenger RNA (mRNA) transcripts, suggesting a pretranslational mechanism. Expression of the pregnane X receptor (PXR), which is crucially involved in CYP3A4 induction by xenobiotics, was strongly correlated to CYP3A4 at the mRNA level in all individuals as well as in the subgroup not exposed to drugs (r = 0.81; P <.0001), but no sex-dependent expression of PXR mRNA was found. The ABC transporter P-glycoprotein, which has been proposed to be implicated in the mechanism of sex-dependent drug clearance, was also not differentially expressed. The influence of drug treatment on expression was examined from patient drug histories, and strong induction of CYP3A4 by carbamazepine and St. John's wort was found. In conclusion, sex, in addition to PXR and drug exposure, is a major factor for CYP3A4 expression in humans, thus explaining many of the previous observations of sex-dependent drug clearance.

ATP Binding Cassette Transporter, Subfamily B, Mem↗

PubMed2003

Mutational analysis of the human dihydropyrimidine dehydrogenase gene by denaturing high-performance liquid chromatography.

Mutations in the DPYD gene, which encodes dihydropyrimidine dehydrogenase (DPD), the rate-limiting enzyme in the catabolism of pyrimidines, are responsible for an inborn error of metabolism associated with thymine-uraciluria and neurological symptoms. Because the antimetabolite 5-fluorouracil (5-FU) is metabolized by the same enzyme, deficient DPYD alleles may also constitute a risk factor for severe toxicity following treatment with this anticancer drug. The aim of this study was to develop a comprehensive and rapid method to detect sequence variations within the DPYD gene. Using polymerase chain reaction (PCR) amplification and denaturing high-performance liquid chromatography (DHPLC), we established a protocol that makes it possible to screen all 23 exons of the DPYD gene and their exon-intron boundaries for both known and unknown mutations under identical conditions. A novel one-step PCR mutagenesis procedure was developed to generate heterozygous mutant amplicons as positive controls to optimize DHPLC detection of any sequence variation. DHPLC analysis was shown to result in mutation-specific elution profiles and to be able to distinguish different base changes within the same exon or different heterozygous combinations of mutations within the same exon. By analyzing the DPYD gene in 16 affected individuals, a total of 47 base changes were detected, representing eight known mutations and three novel intronic base changes. Sequence analysis confirmed all base changes detected. This method will be useful in identifying patients at risk for toxicity prior to 5-FU treatment, as well as in the analysis of individual patients with thymine-uraciluria.

Alleles↗

PubMed2003

Proximal femur fracture in older patients--rehabilitation and clinical outcome.

BACKGROUND: hip fracture is a major cause of morbidity and mortality in older people; optimal post-surgical treatment is a matter of controversy. OBJECTIVE: to examine the effects of rehabilitation on the clinical outcome following surgical treatment of hip fracture. DESIGN: prospective longitudinal study in three groups of patients with different post-surgical care. METHODS: initial screening of 283 elderly patients with proximal femur fracture; documentation of medical and social history and clinical data; geriatric assessments (Activities of Daily Living and Instrumental Activities of Daily Living) during hospital stay and follow-up for 12 months. One hundred and forty-five patients (>or=65 years) of normal mental status were eligible for the study. Successful follow-up could be monitored in 120 and 117 patients for 6 and 12 months, respectively. Sixty-nine and 39 patients underwent supervised inpatient rehabilitation in an orthopaedic or geriatric hospital, respectively (intervention groups A and B, respectively) whereas 34 patients received no special rehabilitation as they were directly discharged home (control group C). RESULTS: initially a fall-/surgical-induced reduction (P<0.001) of the main outcome measure (Activities of Daily Living) was observed in all patients. Within 6 months of rehabilitation there was an improvement (P<0.01) in Activities of Daily Living; however the pre-fracture scores were not reached. The same time pattern was seen in group C. Therefore no significant differences between the three groups of patients in approaching the baseline status was visible. Moreover, the one-year total mortality in the studied population with normal mental status averaged 11.7% and did not differ between the three groups. CONCLUSION: based on our measured outcome variables institutional rehabilitation after surgical treatment of hip fracture apparently had no significant impact on mortality and morbidity in older patients of normal mental status.

Accidental Falls↗