Absorption, distribution and excretion of the anti-tuberculosis drug delamanid in rats: Extensive tissue distribution suggests potential therapeutic value for extrapulmonary tuberculosis.

Delamanid (OPC-67683, Deltyba™, nitro-dihydro-imidazooxazoles derivative) is approved for the treatment of adult pulmonary multidrug-resistant tuberculosis. The absorption, distribution and excretion of delamanid-derived radioactivity were investigated after a single oral administration of 14 C-delamanid at 3 mg/kg to rats. In both male and female rats, radioactivity in blood and all tissues reached peak levels by 8 or 24 h post-dose, and thereafter decreased slowly. Radioactivity levels were 3- to 5-fold higher in lung tissue at time to maximum concentration compared with plasma. In addition, radioactivity was broadly distributed in various tissues, including the central nervous system, eyeball, placenta and fetus, indicating that 14 C-delamanid permeated the brain, retinal and placental blood barriers. By 168 h post-dose, radioactivity in almost all the tissues was higher than that in the plasma. Radioactivity was also transferred into the milk of lactating rats. Approximately 6% and 92% of radioactivity was excreted in the urine and feces, respectively, indicating that the absorbed radioactivity was primarily excreted via the biliary route. No significant differences in the absorption, distribution and excretion of 14 C-delamanid were observed between male and female rats. The pharmacokinetic results suggested that delamanid was broadly distributed to the lungs and various tissues for a prolonged duration of time at concentrations expected to effectively target tuberculosis bacteria. These data indicate that delamanid, in addition to its previously demonstrated efficacy in pulmonary tuberculosis, might be an effective therapeutic approach to treating extrapulmonary tuberculosis. Copyright © 2017 John Wiley & Sons, Ltd.

Antitubercular Agent Delamanid and Metabolites as Substrates and Inhibitors of ABC and Solute Carrier Transporters.

Delamanid (Deltyba, OPC-67683) is the first approved drug in a novel class of nitro-dihydro-imidazooxazoles developed for the treatment of multidrug-resistant tuberculosis. Patients with tuberculosis require treatment with multiple drugs, several of which have known drug-drug interactions. Transporters regulate drug absorption, distribution, and excretion; therefore, the inhibition of transport by one agent may alter the pharmacokinetics of another, leading to unexpected adverse events. Therefore, it is important to understand how delamanid affects transport activity. In the present study, the potencies of delamanid and its main metabolites as the substrates and inhibitors of various transporters were evaluated in vitro Delamanid was not transported by the efflux ATP-binding cassette (ABC) transporters P-glycoprotein (P-gp; MDR1/ABCB1) and breast cancer resistance protein (BCRP/ABCG2), solute carrier (SLC) transporters, organic anion-transporting polypeptides, or organic cation transporter 1. Similarly, metabolite 1 (M1) was not a substrate for any of these transporters except P-gp. Delamanid showed no inhibitory effect on ABC transporters MDR1, BCRP, and bile salt export pump (BSEP; ABCB11), SLC transporters, or organic anion transporters. M1 and M2 inhibited P-gp- and BCRP-mediated transport but did so only at the 50% inhibitory concentrations (M1, 4.65 and 5.71 µmol/liter, respectively; M2, 7.80 and 6.02 µmol/liter, respectively), well above the corresponding maximum concentration in plasma values observed following the administration of multiple doses in clinical trials. M3 and M4 did not affect the activities of any of the transporters tested. These in vitro data suggest that delamanid is unlikely to have clinically relevant interactions with drugs for which absorption and disposition are mediated by this group of transporters.

A synthetic cannabinoid FDU-NNEI, two 2H-indazole isomers of synthetic cannabinoids AB-CHMINACA and NNEI indazole analog (MN-18), a phenethylamine derivative N-OH-EDMA, and a cathinone derivative dimethoxy-α-PHP, newly identified in illegal products.

Six new psychoactive substances were identified together with two other substances (compounds 1-8) in illegal products by our ongoing survey in Japan between January and July 2014. A new synthetic cannabinoid, FDU-NNEI [1-(4-fluorobenzyl)-N-(naphthalen-1-yl)-1H-indole-3-carboxamide, 2], was detected with the newly distributed synthetic cannabinoid FDU-PB-22 (1). Two 2H-indazole isomers of synthetic cannabinoids, AB-CHMINACA 2H-indazole analog (3) and NNEI 2H-indazole analog (4), were newly identified with 1H-indazoles [AB-CHMINACA and NNEI indazole analog (MN-18)]. In addition, 2-methylpropyl N-(naphthalen-1-yl) carbamate (5) and isobutyl 1-pentyl-1H-indazole-3-carboxylate (6) were detected in illegal products. Compound 6 is considered to be a by-product of the preparation of NNEI indazole analog from compound 5 and 1-pentyl-1H-indazole. A phenethylamine derivative, N-OH-EDMA [N-hydroxy-3,4-ethylenedioxy-N-methylamphetamine, 7], and a cathinone derivative, dimethoxy-α-PHP (dimethoxy-α-pyrrolidinohexanophenone, 8), were newly identified in illegal products. Among them, compounds 1 and 8 have been controlled as designated substances (Shitei-Yakubutsu) under the Pharmaceutical Affairs Law in Japan since August and November 2014, respectively.

Inhibitory Potential of Twenty Five Anti-tuberculosis Drugs on CYP Activities in Human Liver Microsomes.

The direct inhibitory potential of twenty five anti-tuberculosis drugs on eight CYP-specific reactions in human liver microsomes was investigated to predict in vivo drug-drug interactions (DDIs) from in vitro data. Rifampicin, rifabutin, and thioacetazone inhibited one CYP reaction. Isoniazid and clofazimine had inhibitory effects on four CYP reactions, and rifapentine, ethionamide, and prothionamide widely inhibited CYP reactions. Based on the inhibition constant (Ki) and the therapeutic total inhibitor concentrations [I]max of eight drugs in human plasma, [I]max/Ki values were calculated to evaluate clinical DDIs. The [I]max/Ki values were 0.20 or less for rifampicin, rifabutin, and thioacetazone; 0.15-2.0 for isoniazid; 0.14-1.5 for rifapentine; 0.29-1.4 for ethionamide; 0.41-2.2 for prothionamide; and 0.12-6.3 for clofazimine. The highest [I]max/Ki values were 2.0 for isoniazid on CYP3A4 [testosterone (T)]; 1.5 for rifapentine on CYP3A4 [midazolam (M)]; 1.4 for ethionamide on CYP2C8; 2.2, 1.8, and 1.3 for prothionamide on CYP2B6, CYP2C19, and CYP2C8, respectively; and 6.3 and 5.7 for clofazimine on CYP3A4 (M) and CYP3A4 (T), respectively. These drugs with high [I]max/Ki values lead to clinical DDIs. Considering the drug regimens for tuberculosis (TB) and co-infection with TB and human immunodeficiency virus, the inhibitory potential for CYP3A4 and CYP2B6 is particularly important. These results suggest that clofazimine and prothionamide are likely to cause clinically relevant DDIs when co-administered with products metabolized by CYP3A4 and CYP2B6, respectively. Isoniazid and rifapentine may cause DDIs with drugs metabolized by CYP3A4.

Pharmacokinetics and Metabolism of Delamanid, a Novel Anti-Tuberculosis Drug, in Animals and Humans: Importance of Albumin Metabolism In Vivo.

Delamanid, a new anti-tuberculosis drug, is metabolized to M1, a unique metabolite formed by cleavage of the 6-nitro-2,3-dihydroimidazo[2,1-b] oxazole moiety, in plasma albumin in vitro. The metabolic activities in dogs and humans are higher than those in rodents. In this study, we characterized the pharmacokinetics and metabolism of delamanid in animals and humans. Eight metabolites (M1-M8) produced by cleavage of the imidazooxazole moiety of delamanid were identified in the plasma after repeated oral administration by liquid chromatography-mass spectrometry analysis. Delamanid was initially catalyzed to M1 and subsequently metabolized by three separate pathways, which suggested that M1 is a crucial starting point. The major pathway in humans was hydroxylation of the oxazole moiety of M1 to form M2 and then successive oxidation to the ketone form (M3) mainly by CYP3A4. M1 had the highest exposure among the eight metabolites after repeated oral dosing in humans, which indicated that M1 was the major metabolite. The overall metabolism of delamanid was qualitatively similar across nonclinical species and humans but was quantitatively different among the species. After repeated administration, the metabolites had much higher concentrations in dogs and humans than in rodents. The in vitro metabolic activity of albumin on delamanid probably caused the species differences observed. We determined that albumin metabolism is a key component of the pharmacokinetics and metabolism of delamanid. Nonhepatic formation of M1 and multiple separate pathways for metabolism of M1 suggest that clinically significant drug-drug interactions with delamanid and M1 are limited.

Metabolic Mechanism of Delamanid, a New Anti-Tuberculosis Drug, in Human Plasma.

The metabolism of delamanid (OPC-67683, Deltyba), a novel treatment of multidrug-resistant tuberculosis, was investigated in vitro using plasma and purified protein preparations from humans and animals. Delamanid was rapidly degraded by incubation in the plasma of all species tested at 37°C, with half-life values (hours) of 0.64 (human), 0.84 (dog), 0.87 (rabbit), 1.90 (mouse), and 3.54 (rat). A major metabolite, (R)-2-amino-4,5-dihydrooxazole derivative (M1), was formed in the plasma by cleavage of the 6-nitro-2,3-dihydroimidazo(2,1-b)oxazole moiety of delamanid. The rate of M1 formation increased with temperature (0-37°C) and pH (6.0-8.0). Delamanid was not converted to M1 in plasma filtrate, with a molecular mass cutoff of 30 kDa, suggesting that bioconversion is mediated by plasma proteins of higher molecular weight. When delamanid was incubated in plasma protein fractions separated by gel filtration chromatography, M1 was observed in the fraction consisting of albumin, γ-globulin, and α1-acid glycoprotein. In pure preparations of these proteins, only human serum albumin (HSA) metabolized delamanid to M1. The formation of M1 followed Michaelis-Menten kinetics in both human plasma and the HSA solution, with similar Km values: 67.8 µM in plasma and 51.5 µM in HSA. The maximum velocity and intrinsic clearance values for M1 were also comparable in plasma and HSA. These results strongly suggest that albumin is predominantly responsible for metabolizing delamanid to M1. We propose that delamanid degradation by albumin begins with a nucleophilic attack of amino acid residues on the electron-poor carbon at the 5 position of nitro-dihydro-imidazooxazole, followed by cleavage of the imidazooxazole moiety to form M1.

Delamanid does not inhibit or induce cytochrome p450 enzymes in vitro.

Delamanid is a new drug for the treatment of multidrug-resistant tuberculosis. Individuals who are co-infected with human immunodeficiency virus and Mycobacterium tuberculosis may require treatment with a number of medications that might interact significantly with the CYP enzyme system as inhibitors or inducers. It is therefore important to understand how drugs in development for the treatment of tuberculosis will affect CYP enzyme metabolism. The ability of delamanid to inhibit or induce CYP enzymes was investigated in vitro using human liver microsomes or human hepatocytes. Delamanid (100 µM) had little potential for mechanism-based inactivation on eight CYP isoforms (CYP1A2, CYP2A6, CYP2B6, CYP2C8, CYP2C9, CYP2C19, CYP2D6, and CYP3A4). Delamanid's metabolites were noted to inhibit the metabolism of some CYP isoforms, but these effects were observed only at metabolite concentrations that were well above those observed in human plasma during clinical trials. Delamanid (≤10 µM) did not induce CYP1A2, CYP2C9, and CYP3A4 activities in human hepatocytes, and there were no increases in CYP1A2, CYP2B6, CYP2C9, and CYP3A4 mRNA levels. Taken together, these data suggest that delamanid is unlikely to cause clinically relevant drug-drug interactions when co-administered with products that are metabolized by the CYP enzyme system.

Characterization of four new designer drugs, 5-chloro-NNEI, NNEI indazole analog, α-PHPP and α-POP, with 11 newly distributed designer drugs in illegal products.

Our continuous survey of illegal products in Japan revealed the new distribution of 15 designer drugs. We identified four synthetic cannabinoids, i.e., NNEI (1), 5-fluoro-NNEI (2), 5-chloro-NNEI (3) and NNEI indazole analog (4), and seven cathinone derivatives, i.e., MPHP (5), α-PHPP (6), α-POP (7), 3,4-dimethoxy-α-PVP (8), 4-fluoro-α-PVP (9), α-ethylaminopentiophenone (10) and N-ethyl-4-methylpentedrone (11). We also determined LY-2183240 (12) and its 2'-isomer (13), which were reported to inhibit endocannabinoid uptake, a methylphenidate analog, 3,4-dichloromethylphenidate (14), and an MDA analog, 5-APDB (15). No chemical and pharmaceutical data for compounds 3, 4, 6 and 7 had been reported, making this the first report on these compounds.

Cloning and structure-function analyses of quinolone- and acridone-producing novel type III polyketide synthases from Citrus microcarpa.

Two novel type III polyketide synthases, quinolone synthase (QNS) and acridone synthase (ACS), were cloned from Citrus microcarpa (Rutaceae). The deduced amino acid sequence of C. microcarpa QNS is unique, and it shared only 56-60% identities with C. microcarpa ACS, Medicago sativa chalcone synthase (CHS), and the previously reported Aegle marmelos QNS. In contrast to the quinolone- and acridone-producing A. marmelos QNS, C. microcarpa QNS produces 4-hydroxy-N-methylquinolone as the "single product" by the one-step condensation of N-methylanthraniloyl-CoA and malonyl-CoA. However, C. microcarpa ACS shows broad substrate specificities and produces not only acridone and quinolone but also chalcone, benzophenone, and phloroglucinol from 4-coumaroyl-CoA, benzoyl-CoA, and hexanoyl-CoA, respectively. Furthermore, the x-ray crystal structures of C. microcarpa QNS and ACS, solved at 2.47- and 2.35-Å resolutions, respectively, revealed wide active site entrances in both enzymes. The wide active site entrances thus provide sufficient space to facilitate the binding of the bulky N-methylanthraniloyl-CoA within the catalytic centers. However, the active site cavity volume of C. microcarpa ACS (760 Å(3)) is almost as large as that of M. sativa CHS (750 Å(3)), and ACS produces acridone by employing an active site cavity and catalytic machinery similar to those of CHS. In contrast, the cavity of C. microcarpa QNS (290 Å(3)) is significantly smaller, which makes this enzyme produce the diketide quinolone. These results as well as mutagenesis analyses provided the first structural bases for the anthranilate-derived production of the quinolone and acridone alkaloid by type III polyketide synthases.

Benzalacetone synthase.

Benzalacetone synthase, from the medicinal plant Rheum palmatum (RpBAS), is a plant-specific chalcone synthase (CHS) superfamily of type III polyketide synthase (PKS). RpBAS catalyzes the one-step, decarboxylative condensation of 4-coumaroyl-CoA with malonyl-CoA to produce the C(6)-C(4) benzalacetone scaffold. The X-ray crystal structures of RpBAS confirmed that the diketide-forming activity is attributable to the characteristic substitution of the conserved active-site "gatekeeper" Phe with Leu. Furthermore, the crystal structures suggested that RpBAS employs novel catalytic machinery for the thioester bond cleavage of the enzyme-bound diketide intermediate and the final decarboxylation reaction to produce benzalacetone. Finally, by exploiting the remarkable substrate tolerance and catalytic versatility of RpBAS, precursor-directed biosynthesis efficiently generated chemically and structurally divergent, unnatural novel polyketide scaffolds. These findings provided a structural basis for the functional diversity of the type III PKS enzymes.

Benzophenone synthase from Garcinia mangostana L. pericarps.

The cDNA of a benzophenone synthase (BPS), a type III polyketide synthase (PKS), was cloned and the recombinant protein expressed from the fruit pericarps of Garcinia mangostana L., which contains mainly prenylated xanthones. The obtained GmBPS showed an amino acid sequence identity of 77-78% with other plant BPSs belonging to the same family (Clusiaceae). The recombinant enzyme produced 2,4,6-trihydroxybenzophenone as the predominant product with benzoyl CoA as substrate. It also accepted other substrates, such as other plant PKSs, and used 1-3 molecules of malonyl CoA to form various phloroglucinol-type and polyketide lactone-type compounds. Thus, providing GmBPS with various substrates in vivo might redirect the xanthone biosynthetic pathway.

Structure-based engineering of benzalacetone synthase.

Benzalacetone synthase (BAS) and chalcone synthase (CHS) are plant-specific type III polyketide synthases (PKSs), sharing 70% amino acid sequence identity and highly homologous overall protein structures. BAS catalyzes the decarboxylative coupling of 4-coumaroyl-CoA with malonyl-CoA to produce the diketide benzalacetone, whereas CHS produces the tetraketide chalcone by iterative condensations with three molecules of malonyl-CoA, and folding the resulting intermediate into a new aromatic ring system. Recent crystallographic analyses of Rheum palmatum BAS revealed that the characteristic substitution of Thr132 (numbering of Medicago sativa CHS2), a conserved CHS residue lining the active-site cavity, with Leu causes steric contraction of the BAS active-site to produce the diketide, instead of the tetraketide. To test this hypothesis, we constructed a set of R. palmatum BAS site-directed mutants (L132G, L132A, L132S, L132C, L132T, L132F, L132Y, L132W and L132P), and investigated the mechanistic consequences of the point mutations. As a result, the single amino acid substitution L132T restored the chalcone-forming activity in BAS, whereas the Ala, Ser, and Cys substitutions expanded the product chain length to produce 4-coumaroyltriacetic acid lactone (CTAL) after three condensations with malonyl-CoA, but without the formation of the aromatic ring system. Homology modeling suggested that this is probably caused by the restoration of the 'coumaroyl binding pocket' in the active-site cavity. These findings provide further insights into the structural details of the catalytic mechanism of the type III PKS enzymes.

A structure-based mechanism for benzalacetone synthase from Rheum palmatum.

Benzalacetone synthase (BAS), a plant-specific type III polyketide synthase (PKS), catalyzes a one-step decarboxylative condensation of malonyl-CoA and 4-coumaroyl-CoA to produce the diketide benzalacetone. We solved the crystal structures of both the wild-type and chalcone-producing I207L/L208F mutant of Rheum palmatum BAS at 1.8 A resolution. In addition, we solved the crystal structure of the wild-type enzyme, in which a monoketide coumarate intermediate is covalently bound to the catalytic cysteine residue, at 1.6 A resolution. This is the first direct evidence that type III PKS utilizes the cysteine as the nucleophile and as the attachment site for the polyketide intermediate. The crystal structures revealed that BAS utilizes an alternative, novel active-site pocket for locking the aromatic moiety of the coumarate, instead of the chalcone synthase's coumaroyl-binding pocket, which is lost in the active-site of the wild-type enzyme and restored in the I207L/L208F mutant. Furthermore, the crystal structures indicated the presence of a putative nucleophilic water molecule which forms hydrogen bond networks with the Cys-His-Asn catalytic triad. This suggested that BAS employs novel catalytic machinery for the thioester bond cleavage of the enzyme-bound diketide intermediate and the final decarboxylation reaction to produce benzalacetone. These findings provided a structural basis for the functional diversity of the type III PKS enzymes.

Structure function analysis of novel type III polyketide synthases from Arabidopsis thaliana.

The genome sequencing project revealed presence of two active chalcone synthase (CHS) homologues (At1g02050 and At4g34850) in the model plant Arabidopsis thaliana. We report herein the two genes encode closely related novel plant-specific type III polyketide synthases (PKSs) that produces long-chain alkyl alpha-pyrones. PKS-A (At1g02050) and PKS-B (At4g34850) share significantly low amino acid sequence identity (20-40%) with other type III PKSs, and the phylogenetic tree analysis revealed that they form a separate cluster located closely to those of bacterial type III PKSs. When expressed in Escherichia coli, both PKS-A and PKS-B accepted unusually long (up to the C(20) chain-length) fatty acyl CoAs as a starter substrate, and carried out sequential condensations with malonyl-CoA to produce triketide and tetraketide alpha-pyrones. Interestingly, despite the low sequence identity, homology modeling revealed that the active-site architecture of PKS-A and PKS-B showed similarity to that of a bacterial type III PKS from Mycobacterium tuberculosis.

OPC-67683, a nitro-dihydro-imidazooxazole derivative with promising action against tuberculosis in vitro and in mice.

BACKGROUND: Tuberculosis (TB) is still a leading cause of death worldwide. Almost a third of the world's population is infected with TB bacilli, and each year approximately 8 million people develop active TB and 2 million die as a result. Today's TB treatment, which dates back to the 1970s, is long and burdensome, requiring at least 6 mo of multidrug chemotherapy. The situation is further compounded by the emergence of multidrug-resistant TB (MDR-TB) and by the infection's lethal synergy with HIV/AIDS. Global health and philanthropic organizations are now pleading for new drug interventions that can address these unmet needs in TB treatment. METHODS AND FINDINGS: Here we report OPC-67683, a nitro-dihydro-imidazooxazole derivative that was screened to help combat the unmet needs in TB treatment. The compound is a mycolic acid biosynthesis inhibitor found to be free of mutagenicity and to possess highly potent activity against TB, including MDR-TB, as shown by its exceptionally low minimum inhibitory concentration (MIC) range of 0.006-0.024 microg/ml in vitro and highly effective therapeutic activity at low doses in vivo. Additionally, the results of the post-antibiotic effect of OPC-67683 on intracellular Mycobacterium tuberculosis showed the agent to be highly and dose-dependently active also against intracellular M. tuberculosis H37Rv after a 4-h pulsed exposure, and this activity at a concentration of 0.1 microg/ml was similar to that of the first-line drug rifampicin (RFP) at a concentration of 3 microg/ml. The combination of OPC-67683 with RFP and pyrazinamide (PZA) exhibited a remarkably quicker eradication (by at least 2 mo) of viable TB bacilli in the lung in comparison with the standard regimen consisting of RFP, isoniazid (INH), ethambutol (EB), and PZA. Furthermore, OPC-67683 was not affected by nor did it affect the activity of liver microsome enzymes, suggesting the possibility for OPC-67683 to be used in combination with drugs, including anti-retrovirals, that induce or are metabolized by cytochrome P450 enzymes. CONCLUSIONS: We concluded that based on these properties OPC-67683 has the potential to be used as a TB drug to help combat the unmet needs in TB treatment.

High performance liquid chromatographic methods for the determination of aripiprazole with ultraviolet detection in rat plasma and brain: application to the pharmacokinetic study.

High performance liquid chromatographic (HPLC) methods were validated for the determination of aripiprazole (OPC-14597, Abilify) in rat plasma and brain. Separation was by Nova-pak phenyl column; flow rate, 1.0 ml/min; mobile phase, acetonitrile-methanol-20 mM sodium sulfate-acetic acid (27:25:48:1, v/v/v/v); UV detection at 254 nm. Reproducibility in plasma and brain showed excellent precision (within 7.8 and 10.6%) and accuracy (96.0-102.4% and 99.0-108.7%) with calibration curve ranges 10.0-2000 ng/ml and 30.0-6000 ng/g, respectively. Validated HPLC methods were successfully applied to pharmacokinetic study of aripiprazole in rats, demonstrating brain concentrations after oral administration five times higher than plasma concentrations.

Effect of gamma-oryzanol on cytochrome P450 activities in human liver microsomes.

The effects of gamma-oryzanol, a drug mainly used for the treatment of hyperlipidaemia, on several cytochrome P450 (CYP) specific reactions in human liver microsomes were investigated to predict drug interactions with gamma-oryzanol in vivo from in vitro data. The following eight CYP catalytic reactions were used in this study: CYP1A1/2-mediated 7-ethoxyresorufin O-deethylation, CYP2A6-mediated coumarin 7-hydroxylation, CYP2B6-mediated 7-benzyloxyresorufin O-debenzylation, CYP2C8/9-mediated tolbutamide methylhydroxylation, CYP2C19-mediated S-mephenytoin 4'-hydroxylation, CYP2D6-mediated bufuralol 1'-hydroxylation, CYP2E1-mediated chlorzoxazone 6-hydroxylation, and CYP3A4-mediated testosterone 6beta-hydroxylation. gamma-Oryzanol had little inhibitory effects on CYP activities, indicating that this compound would not be expected to cause clinically significant interactions with other CYP-metabolized drugs at expected therapeutic concentrations.

Effect of probucol on cytochrome P450 activities in human liver microsomes.

The effects of probucol, a cholesterol-lowering agent, on several cytochrome P450 (CYP) isoform-specific reactions in human liver microsomes were investigated to predict drug interactions with probucol in vivo from in vitro data. The following eight CYP catalytic reactions were used in this study: CYP1A1/2-mediated 7-ethoxyresorufin O-deethylation, CYP2A6-mediated coumarin 7-hydroxylation, CYP2B6-mediated 7-benzyloxyresorufin O-debenzylation, CYP2C8/9-mediated tolbutamide methylhydroxylation, CYP2C19-mediated S-mephenytoin 4'-hydroxylation, CYP2D6-mediated bufuralol 1'-hydroxylation, CYP2E1-mediated chlorzoxazone 6-hydroxylation, and CYP3A4-mediated testosterone 6beta-hydroxylation. Probucol had neither stimulatory nor inhibitory effects on CYP1Al/2, 2A6, 2B6, 2C8/9, 2C19, 2D6, 2E1, and 3A4 activities at concentrations up to 300 microM, indicating that probucol, at the expected therapeutic concentrations, would not be predicted to cause clinically significant interactions with other CYP-metabolized drugs.

Inhibition of human drug metabolizing cytochrome P450 by buprenorphine.

The effects of buprenorphine, a powerful mixed agonist/antagonist analgesic, on several cytochrome P450 (CYP) isoform specific reactions in human liver microsomes were investigated to predict drug interaction of buprenorphine in vivo from in vitro data. The following eight CYP-catalytic reactions were used in this study: CYPlA1/2-mediated 7-ethoxyresorufin O-deethylation, CYP2A6-mediated coumarin 7-hydroxylation, CYP2B6-mediated 7-benzyloxyresorufin O-debenzylation, CYP2C8/9-mediated tolbutamide methylhydroxylation, CYP2C19-mediated S-mephenytoin 4-hydroxylation, CYP2D6-mediated bufuralol 1'-hydroxylation, CYP2E1-mediated chlorzoxazone 6-hydroxylation, and CYP3A4-mediated testosterone 6beta-hydroxylation. Buprenorphine strongly inhibited the CYP3A4- and CYP2D6-catalyzed reactions with Ki values of 14.7 microM and 21.4 microM, respectively. The analgesic also weakly inhibited specific reactions catalyzed by CYP1A1/2 (Ki=132 microM), CYP2B6 (Ki=133 microM), CYP2C19 (Ki=146 microM), CYP2C8/9 (IC50>300 microM), and CYP2E1 (IC50>300 microM), but not CYP2A6 mediated pathway. In consideration of the Ki values obtained in this study and the therapeutic concentration of buprenorphine in human plasma, buprenorphine would not be predicted to cause clinically significant interactions with other CYP-metabolized drugs.