Biomedical subjects
Publications and source records attributed to B Lacarelle.
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The genetic polymorphism affecting the CYP3A5 enzyme is responsible for interindividual and interethnic variability in the metabolism of CYP3A5 substrates. The full extent of the CYP3A5 genetic polymorphism was analysed in French Caucasian, Gabonese and Tunisian populations using a polymerase chain reaction-single strand conformational polymorphism (PCR-SSCP) strategy. In the three populations, eight, 17 and ten single nucleotide polymorphisms (SNPs), respectively, were identified, among which nine correspond to rare new mutations. Also identified were 16 alleles including eight new allelic variants. Significant differences were observed in the distribution of these alleles. Particularly, the frequency of the CYP3A5*3C null allele in French Caucasians (81.3%) and in Tunisians (80.0%) is higher than in the Gabonese population (12.5%) (p < 0.001). Considering the CYP3A5 genotypes of the tested individuals, only 10.4% of French Caucasians and 30.0% of Tunisians were identified as CYP3A5 expressors. In contrast, 90.0% of Gabonese subjects appear to express the CYP3A5 protein.
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BACKGROUND AND OBJECTIVES: Taxol (paclitaxel), is an antimicrotubule agent widely prescribed for the treatment of many tumoral diseases. Taxol must be used in non-polyvinyl chloride bags, diluted to concentrations of 0.3-1.2 mg/mL in 5% dextrose or in 0.9% sodium chloride. Under these conditions, Taxol is chemically and physically stable for 27 h at 25 degrees C. The aim of the study was to evaluate the 72-h stability of Taxol under common clinical use conditions. METHODS: Taxol was diluted with 5% dextrose and 0.9% sodium chloride to final concentrations of 0.3 and 1.2 mg/mL in four polyolefin bags (Viaflo, Freeflex, Ecoflac and Macoflex N). Taxol-stability, was assessed by turbidimetry and by high-performance liquid chromatography using solutions stored in the dark, over 72 h at +4 degrees C. RESULTS: No haze, turbidity, or precipitate was observed. Paclitaxel concentration remained above 95% of the initial value whatever the solvent or container used. CONCLUSION: Paclitaxel at 0.3 and 1.2 mg/mL in 5% dextrose and in 0.9% sodium chloride is stable in Viaflo, Freeflex, Ecoflac and Macoflex N non-PVC bags for 72 h in the dark at +4 degrees C. The longer stability should make the use of Taxol in clinical practice easier.
BACKGROUND AND AIMS: Cytochrome P450 1A1 catalyzes the degradation of endobiotics (estradiol, fatty acids, and so on) and the bioactivation of numerous environmental procarcinogens, such as arylamines and polycyclic aromatic hydrocarbons, that are found in food. Several peroxisome proliferators and arachidonic acid derivatives enhance cytochrome P450 1A1 activity, but the mechanisms involved remain unknown. The aim of this work was to study the role of peroxisome proliferator-activated receptors in cytochrome P450 1A1 gene induction. METHODS: The role of peroxisome proliferator-activated receptor transcription factors in cytochrome P450 1A1 induction was assessed by means of enzymatic activities, quantitative real-time polymerase chain reaction, gene reporter assays, mutagenesis, and electrophoretic mobility shift assay. RESULTS: We show that peroxisome proliferator-activated receptor-alpha agonists (WY-14643, bezafibrate, clofibrate, and phthalate) induce human cytochrome P450 1A1 gene expression, whereas 2,4-thiazolidinedione, a specific peroxisome proliferator-activated receptor-gamma agonist, represses it. The induction of cytochrome P450 1A1 transcripts by WY-14643 was associated with a marked increase of ethoxyresorufin O -deethylase activity (10-fold at 200 mumol/L). Transfection of peroxisome proliferator-activated receptor-alpha complementary DNA enhanced cytochrome P450 1A1 messenger RNA induction by WY-14643, although WY-14643 failed to activate xenobiotic responsive element sequences. Two peroxisome proliferator response element sites were located at positions -931/-919 and -531/-519 of the cytochrome P450 1A1 promoter. Their inactivation by directed mutagenesis suppressed the inductive effect of WY-14643 on cytochrome P450 1A1 promoter activation. Electrophoretic mobility shift assay and chromatin immunoprecipitation assay experiments showed that the 2 cytochrome P450 1A1 peroxisome proliferator response element sites bind the peroxisome proliferator-activated receptor-alpha/retinoid X receptor-alpha heterodimer. CONCLUSIONS: We describe here a new cytochrome P450 1A1 induction pathway involving peroxisome proliferator-activated receptor-alpha and 2 peroxisome proliferator response element sites, indicating that peroxisome proliferator-activated receptor-alpha ligands, which are common environmental compounds, may be involved in carcinogenesis.
BACKGROUND: Dihydropyrimidine dehydrogenase (DPD) gene polymorphism may lead to severe toxicity with 5-fluorouracil (5-FU), a major anticancer drug extensively used in clinical oncology. Drug monitoring combined with early detection of patients at risk would enable timely dose adaptation so as to maintain drug concentrations within a therapeutic window. However, the best method to identify such patients remains to be determined. OBJECTIVE: The aim of this study was to develop a rapid and simple high-performance liquid chromatographic (HPLC) method for estimating uracil/dihydrouracil (U/UH2) ratio in plasma, as an index of DPD status, and for assaying 5-FU as part of drug level monitoring. METHOD: Assay of 5-FU, and U/UH2 detection were performed on a HPLC system equipped with UV detector. Analytes were separated at room temperature using a 5 microm particles, 25 cm RP-18 X-Terra column. The mobile-phase consisted of a KH(2)PO(4) salt solution (0.05 m) + 0.1% triethylamine (TEA) pumped at 0.4 mL/min. Detection of 5-FU and 5-bromouracil were performed at 254 nm; U and UH2 elution was monitored at 210 nm. RESULTS: The method was sensitive and specific for assaying 5-FU within the 5-500 ng/mL concentration range, which covers exposure levels currently met in clinical practice. The method was simple, and relatively cheap, and rapid, with an analytical run time of about 30 min. Data from a patient with 5-FU toxicity suggest that the method was capable of identifying DPD metabolic phenotype in cancer patients, based on measurement of plasma U/UH2 ratio. CONCLUSION: The method described should be suitable both for detecting patients at high risk of 5-FU toxicity, and for drug level monitoring during chemotherapy.
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Low doses of ritonavir, a strong inhibitor of cytochrome P450 3A4, enhances the pharmacokinetic profile of indinavir with increased serum levels. We assessed the indinavir-ritonavir 400/200 twice daily combination in 17 HIV-infected patients focusing on the pharmacokinetic data and the tolerance of this regimen. IDV trough and peak concentrations were measured by high-performance liquid chromatography. Median indinavir trough and peak concentrations were 553 ng/ml and 3626 ng/ml, respectively. A good tolerance was observed except for three patients who experienced a major toxicity. Only one dose adjustment was related to indinavir toxicity. Considering the fact that Cmax is mainly responsible of the adverse effects, particularly renal stones, the indinavir-ritonavir 400/200 mg twice daily regimen offers a well-tolerated combination with an increased Cmin but a lower Cmax compared with both the standard tid regimen and higher dose of IDV-RTV regimens.
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A French survey of 325 HIV-infected subjects with a history of injecting drugs allowed us to study the recognition of patients' injection drug use (IDU) by physicians providing HIV-infection care, and to analyze the correlation between patient demographics and incorrect IDU identification. Kappa for concordance of physician's reports of their patient's IDU with patient's declaration was 0.37; concordance was lower among socially vulnerable patients. This contrasted with a nested study of validity of patient's self-report of opioid use: Kappa for patient's declaration of opioid use within the past two days against a biological assay was 0.61, and concordance was higher among socially vulnerable patients. Concordance of physicians' ratings and patients' reports of IDU was not more than fair, even though physicians were knowledgeable about their patient's IDU history. This concordance varied with social status in a way that did not correspond with variations in self-reported opioid use validity, suggesting that identification of active IDU might be partly based on incorrect interpretation of subjective cues.
AIMS: The primary objective of this study was to determine how the pharmacokinetics of sabeluzole, an investigational drug with specific effects on memory and learning abilities, are affected by chronic liver disease. Since sabeluzole is metabolised by CYP2D6, a secondary objective was to study the correlation between CYP2D6 activity (as assessed by the dextromethorphan dextrorphan metabolic ratio) and hepatic dysfunction. METHODS: The single-dose pharmacokinetics of sabeluzole (10 mg) was compared in 10 healthy Caucasian subjects and 10 patients with severe hepatic dysfunction. The urinary dextromethorphan/dextrorphan (DMP/DRP) metabolic ratio was determined after intake of 20 mg dextromethorphan (NODEX capsules). RESULTS: The terminal half-life of sabeluzole was significantly prolonged in subjects with severe hepatic dysfunction vs healthy subjects (respectively 39.3 +/- 11.5 h; 17.5 +/- 10.2 h (mean +/- s.d.)). The areas under the curve (AUC) were significantly higher in subjects with severe hepatic dysfunction than in healthy volunteers (681 +/- 200 ng ml(-1) h vs 331 +/- 282 ng ml(-1) h). There was a significant correlation between the AUC(0,infinity) and the DMP/DRP metabolic ratio in healthy volunteers and subjects with severe hepatic dysfunction. AUC was greater and elimination of sabeluzole slower in poor metabolizers compared with extensive metabolizers. CONCLUSIONS: These results suggest that a) sabeluzole dose should be reduced in patients with severe hepatic dysfunction and b) the AUC of sabeluzole is linked to individual CYP2D6 activity.
In order to explore the effect of Panax vietnamensis on carbon tetrachloride-induced hepatotoxicity, mice were pretreated for 7 days with either crude extract or total saponins. Crude extract and total saponins dramatically decreased carbon tetrachloride-induced increase of serum GST alpha level (-50.0%, -49.5% respectively). Serum AST level was significantly decreased only with total saponins (-52.2%) and ALT level was slightly modified. In vitro experiments shown that both preparations at high concentrations (> 2000 micrograms/ml) are able to inhibit CYP2E1 enzymatic activity in mouse and human microsomes. However, we did not observe any modification of Cyp2e1 gene expression (enzymatic activity, protein and mRNA levels) in mice treated with either crude extract or total saponins. Taken together, these data demonstrated that Panax vietnamensis could be used as an hepatoprotectant. However, the mechanism of action is not associated with CYP2E1 expression, as previously suggested in vitro in rat for total saponins from Panax ginseng.
We studied the pharmacokinetics of cefepime (2 g bd) in six burns patients. Blood, urine and skin samples were collected to measure cefepime concentrations. A two-compartment model was fitted to the data. At day 1, t(1/2beta) was 2.45 +/- 0.56 h, V(ss) 0.36 +/- 0.1 L/kg, total clearance 152 +/- 25.2 mL/min, and AUC 217 +/- 34 mg*h/L. There was no statistical difference between day 1 and day 3 for any of the pharmacokinetic parameters. We demonstrated good penetration of cefepime in skin. These results show that it is not necessary to change the standard dosage of cefepime in burns patients.
The relationship between ritonavir plasma concentration, efficacy, and tolerance was evaluated in 31 children with advanced HIV infection who were receiving a triple therapy with ritonavir as protease inhibitor. Median CD4+ lymphocyte count and median viral load before the initiation of ritonavir-containing combination therapy were 1320 cells/mL and 5 log10 copies/mL, respectively. Ritonavir was given at a dose ranging from 300 to 450 mg/m2 twice daily. The median follow-up of triple therapy was 19 months. Response was defined as a drop of viremia of more than 1 log. Plasma drug levels were determined twice during the observation period: after at least 4 weeks and after 3 months of combined treatment. Samples were collected before (residual) and 2 hours (T2) after drug intake. Cholesterol, triglycerides, alanine transaminase, aspartate transaminase, and gamma-glutamyl transpeptidase were assessed at the same time. The median values of ritonavir residual and T2 levels were 1.64 mg/L and 5.9 mg/L at observation 1 and 3.35 mg/L and 6.29 mg/L at observation 2, respectively. According to virologic response, median residual concentrations of ritonavir were 3.17, 2.52, and 1.04 mg/L for the complete, the partial, and the no-response groups. The authors observed a wide intersubject variability of ritonavir concentrations with an increase in residual levels between the two observation periods. Residual levels were correlated with virologic response whereas there was no direct association between T2 levels and long-term response. Patients with complete or partial response displayed statistically significantly higher residual concentrations than the no-response group. No correlation could be demonstrated between elevated plasma drug concentrations and abnormal cholesterol or triglycerides values. These results emphasize the importance of a sustained high ritonavir concentration to achieve optimal treatment efficacy. Furthermore, these results prove the clinical benefit of therapeutic drug monitoring and could potentially improve patient evaluation in terms of treatment efficacy, compliance, and viral resistance.
Cerebrospinal fluid (CSF) penetration and the pharmacokinetics of vancomycin were studied after continuous infusion (50 to 60 mg/kg of body weight/day after a loading dose of 15 mg/kg) in 13 mechanically ventilated patients hospitalized in an intensive care unit. Seven patients were treated for a sensitive bacterial meningitis and the other six patients, who had a severe concomitant neurologic disease with intracranial hypertension, were treated for various infections. Vancomycin CSF penetration was significantly higher (P < 0.05) in the meningitis group (serum/CSF ratio, 48%) than in the other group (serum/CSF ratio, 18%). Vancomycin pharmacokinetic parameters did not differ from those obtained with conventional dosing. No adverse effect was observed, in particular with regard to renal function.
Bupivacaine is used to provide prolonged anesthesia and postoperative analgesia. The human cytochrome P450 (CYP) involved in bupivacaine degradation into pipecolylxylidine (PPX), its major metabolite, has, to our knowledge, never been described. Microsome samples were prepared from six human livers and incubated in the presence of bupivacaine. The concentrations of PPX in the microsomal suspensions were assessed, and K(m) and V(max) values were calculated. Bupivacaine incubations were then performed with specific CYP substrates and inhibitors. For each sample of hepatic microsomes, the correlation between the rate of PPX formation and the corresponding erythromycin N-demethylase activity was analyzed. Finally, an immunoinhibition study using an anti-rabbit CYP3A6 antibody and assays with cDNA-expressed human CYP were conducted. The apparent K(m) and V(max) values of bupivacaine were, respectively, 125 microM and 4.78 nmol/min/mg of microsomal protein. The strongest inhibition of bupivacaine metabolism was obtained for troleandomycin (-95% at 50 microM), a specific CYP3A inhibitor. The correlation between PPX formation and erythromycin N-demethylase activity showed an R value of 0.99 whereas anti-rabbit CYP3A6 antibody inhibited the degradation of bupivacaine into PPX by 99%. Finally, CYP1A2 and CYP2E1 cDNA-expressed forms of human CYP did not allow PPX formation, CYP2C19 and CYP2D6 produced only small amounts whereas CYP3A4 most efficiently metabolized bupivacaine into PPX. These results demonstrated that bupivacaine degradation into PPX was mediated in humans by CYP3A.
Ethanol is a well-known hepatotoxicant inducing steatosis and membrane lipoperoxidation. The aim of the present study was to investigate in rats, whether the protective effect of UDC on ethanol-induced lipid peroxidation may be related with CYP2E1 and CYP3A1/2 gene expression. We showed that UDC treatment in ethanol-fed rats induced a significant decrease in liver triglyceride concentration which was closely correlated with a reduction in malondialdehyde and hydroxyalkenal levels. In chronically ethanol-fed rats, CYP2E1 and CYP3A1/2 gene expressions were increased by a post-transcriptional mechanism. These inductions, mainly of CYP2E1, could take part in alcohol-induced hepatic lipoperoxidation. UDC modified neither the specific activity, nor the protein level, nor the mRNA level of CYP2E1 when compared with control. UDC supplementation to alcohol diet did not prevent the increase in CYP2E1 expression of ethanol-fed rats. Furthermore, CYP3A1/2 protein levels were similarly increased by ethanol and ethanol plus UDC treatment. Therefore, UDC protective effect against ethanol-induced lipoperoxidation was not associated with a modification of CYP2E1 and CYP3A1/2 expression.
A strong overlap exists between gp170 and CYP3A substrates and inducers. In order to investigate a putative coregulation of MDR and CYPA gene expression, we measured their transcripts in human liver and after dexamethasone treatment in HepG2 cells or in different mouse tissues. In human liver, we observed no correlation between MDR1 and CYP3A4 expression, whereas these genes were coinduced by dexamethasone in HepG2 cells. In mouse liver treated with dexamethasone, mdr1b and Cyp3a were induced (5- and 2-fold, respectively). In adrenals, the main expressing gp170 tissue, Cyp3a, was increased while mdr1b was repressed (-51%). The expression of mdr1b increased in heart, brain, and colon and decreased in lung and kidney but Cyp3a was not detectable. In conclusion, human hepatic CYP3A4 and MDR1 are not corregulated but are coinducible. In vivo murine mdr1b and Cyp3a are coregulated by dexamethasone in liver and inversely regulated in adrenals.