Search

This study aimed at characterizing the interspecies absorption, distribution, metabolism and elimination (ADME) profile of N-geranyl-N'-(2-adamantyl)ethane-1,2-diamine (SQ109), a new diamine-based antitubercular drug. Single doses of SQ109 were administered (intravenously (i.v.) and per os (p.o.)) to rodents and dogs and blood samples were analyzed by liquid chromatography tandem mass spectrometry (LC/MS/MS). Based on i.v. equivalent body surface area dose, the terminal half-life (t1/2) of SQ109 in dogs was longer than that in rodents, reflected by a larger volume of distribution (Vss) and a higher clearance rate of SQ109 in dogs, compared to that in rodents. The oral bioavailability of SQ109 in dogs, rats and mice were 2.4-5, 12 and 3.8%, respectively. After oral administration of [14C]SQ109 to rats, the highest level of radioactivity was in the liver, followed by the lung, spleen and kidney. Tissue-to-blood ratios of [14C]SQ109 were greater than 1. Fecal elimination of [14C]SQ109 accounted for 22.2% of the total dose of [14C]SQ109, while urinary excretion accounted for only 5.6%. The binding of [14C]SQ109 (0.1-2.5 microg ml-1) to plasma proteins varied from 6 to 23% depending on the species (human, mouse, rat and dog). SQ109 was metabolized by rat, mouse, dog and human liver microsomes, resulting in 22.8, 48.4, 50.8 or 58.3%, respectively, of SQ109 remaining after a 10-min incubation at 37 degrees C. The predominant metabolites in the human liver microsomes gave intense ion signals at 195, 347 and 363m/z, suggesting the oxidation, epoxidation and N-dealkylation of SQ109. P450 reaction phenotyping using recombinant cDNA-expressed human CYPs in conjunction with specific CYP inhibitors indicated that CYP2D6 and CYP2C19 were the predominant CYPs involved in SQ109 metabolism.

There is circumstantial evidence suggesting that 5-hydroxytryptamine (5-HT) could be biotransformed by enzymatic systems other than monoamino oxidase A, and that the isoforms of cytochrome P450 may be a source of nitric oxide. This study aimed to assess whether cytochrome P450 contributes to 5-HT biotransformation, and to provide evidence that 5-HT metabolism generates nitric oxide. Addition of 5-HT to cultured hepatocytes yielded 5-hydroxyindol acetic acid, a formation modulated by cytochrome P450 enzyme inducers and inhibitors. Recombinant human CYP2B6, 2C9 and 2C19 biotransformed 5-HT in 5-hydroxyindol acetic acid, but not CYP1A2, 2D6 or 3A4. Cultured hepatocytes with 5-HT generated nitric oxide, the amount of which was altered by cytochrome P450 enzyme inducers and inhibitors. In the presence of CYP2B6, 2C9 and 2C19, 5-HT relaxed precontracted isolated aortic rings, with or without endothelium, an effect prevented by the addition of methylene blue and an inhibitor of catalase, but not by myoglobin. In the absence of catalase, hydroxylamine was always assayed as a byproduct of 5-HT metabolism. In conclusion, CYP2B6, 2C9 and 2C19 biotransform 5-HT, yielding hydroxylamine, which is converted to nitric oxide in the presence of catalase. British Journal of Pharmacology (2004) 141, 407-414. doi:10.1038/sj.bjp.0705632

Background: Many commonly ingested substances such as grapefruit juice and Hypericum perforatum (St John's wort) have been found to interact with important therapeutic agents such as cyclosporine (INN, ciclosporin). The mechanism for these interactions is thought to involve modulation of the activity of the drug-metabolizing enzyme cytochrome P4503A4 (CYP3A4) and/or the drug transport protein Pglycoprotein. In vitro data suggest that red wine may interact with CYP3A4 substrates such as cyclosporine., Methods: We conducted a randomized, 2-way crossover study of 12 healthy individuals. Subjects received a single 8-mg/kg dose of oral cyclosporine with water (control) and with 12 oz of red wine (Blackstone Merlot, 1996; Blackstone Winery, Graton, Calif). Whole blood was analyzed for cyclosporine and 6 metabolites by specific fluorescence polarization immunoassay and tandem liquid chromatography-mass spectrometry. Blood levels of cyclosporine were compared between the 2 arms., Results: Red wine caused a 50% increase in the oral clearance of cyclosporine. Systemic exposure as measured by the area under the concentration-versus-time curve (AUC) and peak concentration (C(max)) were significantly decreased by red wine. However, half-life was not affected, suggesting that red wine decreased cyclosporine absorption. In vitro, the solubility of cyclosporine in red wine appeared to be lower than in water., Conclusions: Administration of cyclosporine with red wine causes a significant decrease in cyclosporine exposure. Because cyclosporine is a narrow therapeutic range compound, caution may be warranted with concomitant intake of red wine and cyclosporine.

The authors assessed the in vitro contribution of cytochrome P450 (CYP) isoforms 1A2, 3A4, 2C9, 2C19, and 2D6 to the N-demethylation of clozapine mediated by human liver microsomal preparations (HLM). In contrast to previous studies, the authors focused on a relatively low hepatic concentration level, 5 microM, to assess the conditions at a therapeutically relevant hepatic concentration level of clozapine. The optimal concentrations of specific inhibitors were initially established using cDNA-expressed CYP isoforms. The mean contributions of CYPs 1A2, 2C19, 3A4, 2C9, and 2D6 amounted to 30%, 24%, 22%, 12%, and 6%, respectively, with regard to the total HLM-mediated N-demethylation. Thus, the present in vitro study on clozapine N-demethylation suggests that CYP1A2 is the most important form at low concentrations, which is in agreement with clinical findings. CYP2C19 is also of considerable importance, while the roles of CYP2C9 and 2D6 are more modest. CYP3A4 attained a dominating role with an average contribution of 37% at a high clozapine concentration (50 microM). The rate of other metabolic routes mediated by CYP2D6 only corresponded to about one fifth of the CYP2D6 catalyzed N-demethylation rate.

Background: Moclobemide, an antidepressant with selective monoamine oxidase-A inhibitory action, is known to be metabolized by CYP2C19 and is also reported to be an inhibitor of CYP2C19, CYP2D6, and CYP1A2. To confirm the involvement of CYP2C19, we performed a pharmacokinetic interaction study., Methods: The effect of omeprazole on the pharmacokinetics of moclobemide was studied in 16 healthy volunteers. The volunteer group comprised 8 extensive metabolizers and 8 poor metabolizers of CYP2C19, which was confirmed by genotyping. Subjects were randomly allocated into two sequence groups, and a single-blind, placebo-controlled, two-period crossover study was performed. In study I, a placebo was orally administered for 7 days. On the eighth morning, 300 mg of moclobemide and 40 mg of placebo were coadministered with 200 mL of water, and a pharmacokinetic study was performed. During study II, 40 mg of omeprazole was given each morning instead of placebo, and pharmacokinetic studies were performed on the first and eighth day with 300 mg of moclobemide coadministration., Results: The inhibition of moclobemide metabolism was significant in extensive metabolizers even after a single dose of omeprazole. After daily administration of omeprazole for 1 week, the pharmacokinetic parameters of moclobemide and its metabolites in extensive metabolizers changed to values similar to those in poor metabolizers. In poor metabolizers, no remarkable changes in the pharmacokinetic parameters were observed., Conclusion: Our results show that CYP2C19 is an important enzyme in the elimination of moclobemide and that it is extensively inhibited by omeprazole in extensive metabolizers, but not in poor metabolizers.

We attempted to predict the in vivo metabolic clearance of alprazolam from in vitro metabolic studies using human liver microsomes and human CYP recombinants. Good correlations were observed between the intrinsic clearance (CL(int)) for 4-hydroxylation and CYP3A4 content and between the CL(int) for alpha-hydroxylation and CYP3A5 content in ten human liver microsomal samples. Using the recombinant CYP isoforms expressed in insect cells, the CL(int) for CYP3A4 was about 2-fold higher than the CL(int) for CYP3A5 in the case of 4-hydroxylation. However, the CL(int) for CYP3A5 was about 3-fold higher than the CL(int) for CYP3A4 in the case of alpha-hydroxylation. The metabolic rates for 4- and alpha-hydroxylation increased as the added amount of cytochrome b(5) increased, and their maximum values were 3- to 4-fold higher than those without cytochrome b(5). The values of CL(int), in vivo predicted from in vitro studies using human liver microsomes and CYP3A4 and CYP3A5 recombinants were within 2.5 times of the observed value calculated from literature data. The average CL(int) value (sum of 4- and alpha-hydroxylation) obtained using three human liver microsomal samples was 4-fold higher than that obtained using three small intestinal microsomal samples from the same donors, indicating the minor contribution of intestinal metabolism to alprazolam disposition. The area under the plasma concentration-time curve (AUC) of alprazolam is reported to increase following co-administration of ketoconazole and the magnitude of the increase predicted from the in vitro K(i) values and reported pharmacokinetic parameters of ketoconazole was 2.30-2.45, which is close to the value observed in vivo (3.19). A quantitative prediction of the AUC increase by cimetidine was also successful (1.73-1.79 vs 1.58-1.64), considering the active transport of cimetidine into the liver. In conclusion, we have succeeded in carrying out an in vitro/in vivo scaling of alprazolam metabolism using human liver microsomes and human CYP3A4 and CYP3A5 recombinants., (Copyright 2001 John Wiley & Sons, Ltd.)

Drug interaction studies were carried out to ensure that hypoglycemia due to inhibition of repaglinide elimination or chronic hyperglycemia due to inhibition of repaglinide absorption was avoided. Conversely, the effects of repaglinide on the pharmacokinetics of drugs with only a narrow margin between effective and toxic concentrations, such as digoxin or theophylline, were determined. The studies reported here compared monotherapy with combined therapies in healthy volunteers. There were no significant differences between the pharmacokinetic parameters of repaglinide when given as monotherapy and when administered concurrently with cimetidine. Similarly, the coadministration of repaglinide and digoxin did not influence the pharmacokinetics of digoxin administered alone. When repaglinide was coadministered with theophylline, the only pharmacokinetic change was that the peak plasma theophylline concentration was slightly reduced. No direct drug-drug interactions were found in these studies, suggesting that repaglinide may be coprescribed with cimetidine, digoxin, or theophylline at the dosage used for monotherapy.

Previous studies have demonstrated an important role for the cytochrome P450 (CYT-P450) pathway in afferent arteriole autoregulatory responses but the involvement of specific pathways remains unknown. Experiments were performed to determine the role of CYT-P450 epoxygenase and hydroxylase pathways in pressure mediated preglomerular autoregulatory responses. Afferent arteriolar diameter was measured as renal perfusion pressure was increased from 80-160 mmHg. Afferent arteriolar diameter averaged 19+/-2 microm at a renal perfusion pressure of 80 mmHg and decreased by 15+/-2% when pressure was increased to 160 mmHg. Inhibition of the epoxygenase pathway with 6-(2-proparglyloxyphenyl)hexanoic acid (PPOH), enhanced the microvascular response to increasing renal perfusion pressure. In the presence of 50 microM PPOH, afferent arteriolar diameter decreased by 29+/-4% when pressure was increased from 80-160 mmHg. Likewise, the sulphonimide derivative of PPOH, N-methylsulphonyl-6-(2-proparglyloxyphenyl) hexanamide (MS-PPOH, 50 microM), enhanced the afferent arteriolar response to increasing renal perfusion pressure. In contrast, the selective CYT-P450 hydroxylase inhibitor, N-methylsulphonyl-12,12-dibromododec-11-enamide (DDMS) attenuated the vascular response to increasing renal perfusion pressure. In the pressure of 25 microM DDMS, afferent arteriolar diameter decreased by 4+/-2% when pressure was increased from 80-160 mmHg. These results suggest that CYT-P450 metabolites of the epoxygenase pathway alter afferent arteriolar responsiveness and thereby modify the ability of the preglomerular vasculature to autoregulate renal blood flow. Additionally, these results provide further support to the concept that a metabolite of the hydroxylase pathway is an integral component of the afferent arteriolar response to elevations in perfusion pressure.

Objective: To further substantiate the role of CYP1A2 and CYP3A4 for the N-demethylation in vivo. At least three different P450s appear to be responsible for the N-demethylation of imipramine to desipramine in vivo: CYP1A2, CYP2C19, and CYP3A4. The role of CYP2C19 in this regard is well documented, but for the two other P450s the evidence is either indirect or based on in vitro studies., Methods: Phenotypic tests for imipramine N-demethylation, CYP1A2 (caffeine testing), CYP2C19 (mephenytoin and chloroguanide [proguanil] testing), and CYP3A4 (hydrocortisone and quinidine testing) were carried out in 32 healthy young Danes; all were poor (n = 31) or extremely slow extensive metabolizers (n = 1) of sparteine., Results: By exclusion of the insignificant log-transformed variables, multiple regression analysis for In (desipramine/imipramine) showed that only in (mephenytoin S/R) correlated (p = 0.013; r2 = 0.19). For in (2-hydroxydesipramine/2-hydroxyimipramine) we found that in (mephenytoin S/R) and in (4-chlorophenylbiguanide/chloroguanide) correlated (p = 0.001; r2 = 0.41)., Conclusion: We did not find in vivo evidence of either CYP1A2 or CYP3A4 activity in the N-demethylation of imipramine. This could be due in part to inadequate CYP1A2 and CYP3A4 in vivo function tests.

1. We have proposed that contractile tension of the ductus arteriosus is sustained by a cytochrome P450-linked mechanism acting as a limiting step in the synthesis of endothelin-1 (ET-1). In the present study, we have used the isolated ductus from near-term foetal lambs and guinea-pigs to investigate the effect on both muscle tone and ET-1 formation of 1-aminobenzotriazole (ABT), a suicide substrate for mono-oxygenase reactions. 2. ABT relaxed the lamb ductus at rest (2.5% O2) and during the oxygen contraction (15 to 95% O2). The effect was seen at 40 microM, and at 0.8 mM active tone was almost completely abolished. ABT (1 mM) also reversed the oxygen contraction in the guinea-pig ductus. 3. In the lamb ductus, the ABT response was not affected by removal of the endothelium or by treatment with 2.8 microM indomethacin (at 2.5% O2) and the ensuing contraction. 4. At both low and high concentration, ABT relaxed marginally, or not at all, the potassium-contracted (55 mM) ductus from either species. 5. ET-1 release from either the intact or endothelium-denuded lamb ductus tended to decrease in the presence of ABT (1 mM), whilst during the same treatment cyclic GMP content of the tissue remained unchanged. 6. We conclude that ABT relaxation is due to suppression of a contractile mechanism and not to activation of prostaglandin- and NO-mediated relaxing mechanisms. This contractile mechanism has a cytochrome P450-based mono-oxygenase reaction as a key component.

Propafenone, a sodium channel blocking antiarrhythmic drug with beta-blocking properties, is metabolized to non-beta-blocking metabolites in part by cytochrome P4502D6. Subtherapeutic doses of quinidine inhibit P4502D6 and increase plasma propafenone in extensive metabolizer subjects, in whom the active enzyme is present. In this study we tested the hypothesis that quinidine would enhance beta-blockade in extensive metabolizers receiving propafenone. Seven extensive and two poor metabolizers received propafenone (225 mg orally every 8 hours) plus quinidine sulfate (60 mg orally every 8 hours) or propafenone plus placebo for 7 days in a randomized, double-blind, crossover fashion. In extensive metabolizers, the coadministration of quinidine significantly increased the extent of propafenone-induced beta-blockade, assessed by a decrease in exercise heart rate and by sensitivity to isoproterenol. We conclude that low-dose quinidine enhances propafenone-induced beta-blockade in extensive metabolizers. Thus the polymorphic patterns of drug metabolism can result in clinically significant drug interactions on a genetic basis.

The formation of three oxidative metabolites of imipramine, N-desmethylimipramine (desipramine), 2-hydroxyimipramine, and 10-hydroxyimipramine was studied in microsomes of an extensive metabolizer liver (KDL 26) and of a poor metabolizer liver (KDL 31) and in a homogenate of COS-1 cells in which the P450IID6 complementary deoxyribonucleic acid had been expressed. The following data support the role of P450IID6 in the 2-hydroxylation of imipramine: (1) The formation of 2-hydroxyimipramine was reduced to less than 20% of the control value when microsomes were incubated with serum containing inhibitory antibodies against P450IID6 (anti-LKM1), whereas no effect was seen with regard to formation of desipramine and 10-hydroxyimipramine, (2) quinidine and levomepromazine were potent competitive inhibitors of 2-hydroxylation of imipramine (ki approximately 70 nmol/L, and ki approximately 1 mumol/L, respectively) but had no effect on N-demethylation and 10-hydroxylation, and (3) in the COS-1 cell, homogenate, 10-hydroxyimipramine, 2-hydroxyimipramine, and desipramine were formed at rates of 48, 164, and 256 pmol per hour per milligram of homogenate protein, respectively. The P450 isozymes that are responsible for N-demethylation and 10-hydroxylation of imipramine have not yet been identified.

Hepatic dysfunction is a frequent complication of total parenteral nutrition (TPN), indicated by derangement of standard liver function tests. However, such changes are variable and nonspecific, and represent hepatic injury rather than changes in hepatic function. Antipyrine (Phenazone) clearance is a sensitive indicator of hepatic microsomal enzyme activity and provides a more specific indication of hepatic function. This was used to investigate the effect of different TPN regimens. Patients receiving a postoperative 2000 kcal TPN regimen providing all nonprotein calories as dextrose (n = 16) showed a 34% reduction of mean antipyrine clearance after 7 days of TPN compared to controls (n = 13, p less than 0.05). This effect was seen also in patients receiving a 1600 kcal dextrose-based regimen (n = 8). In patients receiving a 2000 kcal TPN regimen in which 500 kcal were provided as lipid (n = 10), mean antipyrine clearance was not significantly different from that of the control group. This study indicates the sensitivity of hepatic microsomal oxidative function, an important route of drug metabolism, to different TPN regimens.