Inhibition Of Cytochrome P450 Enzymes Research Articles

Phenethyl isocyanate is not the metabolite of phenethyl isothiocyanate responsible for mechanism-based inhibition of cytochrome P450

Phenethyl isothiocyanate is a chemopreventive phytochemical present in cruciferous vegetables where it exists as the glucosinolate gluconasturtiin. It is a mechanism-based inhibitor of both rat and human cytochrome P450 enzymes. The principal objective of the present study was to ascertain whether phenethyl isocyanate, formed by the cytochrome P450-mediated oxidative desulphuration of phenethyl isothiocyanate, is the metabolite responsible for the mechanism-based inhibition. Phenethyl isothiocyanate, following incubation with Aroclor 1254-induced rat liver microsomes in the presence of NADPH, markedly suppressed the CYP1A-mediated O-deethylation of ethoxyresorufin; extent of inhibition was directly related to the pre-incubation time and was antagonised by reduced glutathione. When human liver microsomes were used, the inhibitory effect of phenethyl isothiocyanate, which was once again related to the pre-incubation time, was even more pronounced. When the ability of phenethyl isothiocyanate and phenethyl isocyanate to directly inhibit the O-deethylation of ethoxyresorufin in rat microsomes was compared, the latter compound was only moderately more effective. In human microsomes, both compounds were equipotent. In phenobarbital-induced lung microsomes, phenethyl isothiocyanate was a direct and potent inhibitor of the O-depentylation of pentoxyresorufin; pre-incubation of the isothiocyanate had no impact. Human precision-cut liver slices were more effective than rat slices in metabolising phenethyl isothiocyanate. Pre-treatment of rats, however, with phenobarbitone significantly enhanced the metabolism of isothiocyanate. It may be inferred from the present studies that: (a) phenethyl isocyanate is not the metabolite of phenethyl isothiocyanate responsible for its mechanism-based inhibition, and (b) CYP2B is an important catalyst of the metabolism of phenethyl isothiocyanate.

Long-term use of rosuvastatin: a critical risk benefit appraisal and comparison with other antihyperlipidemics

Rosuvastatin represents the latest inhibitor of 3-hydroxy-3-methylglutaryl coenzyme A (HMG-CoA) reductase introduced in clinical practice for the treatment of hypercholesterolemia. In comparative trials, across dose ranges this statin reduced low-density lipoprotein (LDL) cholesterol and total cholesterol significantly more than atorvastatin, simvastatin, and pravastatin, and triglycerides significantly more than simvastatin and pravastatin. In healthy subjects with normal LDL cholesterol and elevated C-reactive protein, rosuvastatin treatment significantly decreased the incidence of cardiovascular events. Its chemical and pharmacokinetic properties (with a low lipophilicity and poor capacity to inhibit cytochrome P450 enzymes) suggest a very limited penetration in extrahepatic tissues with a lower risk of muscle toxicity and unlike metabolically mediated drug-drug interactions. This article reviews the most recent data on the pharmacologic and clinical properties of rosuvastatin, in order to enable the correct use of this statin for the treatment of hypercholesterolemia.

Isolation and Functional Characterization of CYP71AJ4 Encoding for the First P450 Monooxygenase of Angular Furanocoumarin Biosynthesis

The biosynthesis of linear and angular furanocoumarins is still poorly understood at the molecular level, with only psoralen synthase (CYP71AJ1) identified from Ammi majus. Using cDNA probes inferred from CYP71AJ1, three orthologs were isolated from Apium graveolens (CYP71AJ2) and Pastinaca sativa (CYP71AJ3 and -4) and functionally expressed in yeast cells. CYP71AJ2 and CYP71AJ3 displayed psoralen synthase activity, whereas CYP71AJ4 only catalyzed the conversion of (+)-columbianetin to angelicin and negligible amounts of a hydroxylated columbianetin by-product. CYP71AJ4 thus constitutes the first fully characterized P450 monooxygenase specific for the angular furanocoumarin pathway. The angelicin synthase exhibited an apparent K(m) of 2.1 +/- 0.4 microm for (+)-columbianetin and a k(cat) of 112 +/- 14 min(-1). Moreover, the use of 3'-deuterated (+)-columbianetin as substrate led to an almost complete "metabolic switch," resulting in the synthesis of anti-3'-hydroxy-3'-deuterated(+)-columbianetin. This confirms that angelicin synthase attacks columbianetin by syn-elimination of hydrogen from C-3'. Sequence comparison between psoralen synthase (CYP71AJ3) and angelicin synthase (CYP71AJ4) showed 70% identity, whereas the identity dropped to 40% in those regions thought to provide the substrate recognition sites. Accordingly, CYP71AJ3 and CYP71AJ4 might be derived from a common ancestor of unknown functionality by gene duplication and subsequent molecular evolution.

Accelerated Metabolism of Voriconazole and Its Partial Reversal by Cimetidine

We report a case of accelerated metabolism of voriconazole during therapy for invasive pulmonary aspergillosis, resulting in subtherapeutic levels. Target voriconazole levels were restored with high dosages of voriconazole (up to 40 mg/kg of body weight/day) and the addition of cimetidine as a cytochrome P450 enzyme inhibitor.

Reaction of Mycobacterium tuberculosis Cytochrome P450 Enzymes with Nitric Oxide

During the initial growth infection stage of Mycobacterium tuberculosis (Mtb), (*)NO produced by host macrophages inhibits heme-containing terminal cytochrome oxidases, inactivates iron/sulfur proteins, and promotes entry into latency. Here we evaluate the potential of (*)NO as an inhibitor of Mtb cytochrome P450 enzymes, as represented by CYP130, CYP51, and the two previously uncharacterized enzymes CYP125 and CYP142. Using UV-visible absorption, resonance Raman, and stopped-flow spectroscopy, we investigated the reactions of (*)NO with these heme proteins in their ferric resting form. (*)NO coordinates tightly to CYP125 and CYP142 (submicromolar) and with a lower affinity (micromolar) to CYP130 and CYP51. Anaerobic reduction of the ferric-NO species with sodium dithionite led to the formation of two spectrally distinct classes of five-coordinate ferrous-NO complexes. Exposure of these species to O(2) revealed that the ferrous-NO forms of CYP125 and CYP142 are labile and convert back to the ferric state within a few minutes, whereas ferrous CYP130 and CYP51 bind (*)NO almost irreversibly. This work clearly indicates that, at physiological concentrations (approximately 1 microM), (*)NO would impair the activity of CYP130 and CYP51, whereas CYP125 and CYP142 are more resistant. Selective P450 inhibition may contribute to the inhibitory effects of (*)NO on Mtb growth.

In Vitro and In Vivo Studies To Characterize the Clearance Mechanism and Potential Cytochrome P450 Interactions of Anidulafungin

Anidulafungin is a novel semisynthetic echinocandin with potent activity against Candida (including azole-resistant isolates) and Aspergillus spp. and is used for serious systemic fungal infections. The purpose of these studies was to characterize the clearance mechanism and potential for drug interactions of anidulafungin. Experiments included in vitro degradation of anidulafungin in buffer and human plasma, a bioassay for antifungal activity, in vitro human cytochrome P450 inhibition studies, in vitro incubation with rat and human hepatocytes, and mass balance studies in rats and humans. Clearance of anidulafungin appeared to be primarily due to slow chemical degradation, with no evidence of hepatic-mediated metabolism (phase 1 or 2). Under physiological conditions, further degradation of the primary degradant appears to take place. The primary degradation product does not retain antifungal activity. Anidulafungin was not an inhibitor of cytochrome P450 enzymes commonly involved in drug metabolism. Mass balance studies showed that anidulafungin was eliminated in the feces predominantly as degradation products, with only a small fraction (10%) eliminated as unchanged drug; fecal elimination likely occurred via biliary excretion. Only negligible renal involvement in the drug's elimination was observed. In conclusion, the primary biotransformation of anidulafungin is mediated by slow chemical degradation, with no evidence for hepatic enzymatic metabolism or renal elimination.

Comprehensive Textbook of AIDS Psychiatryedited by Mary Ann Cohen and Jack M. Gorman. New York, Oxford University Press, 2007, 656 pp., $98.50.

Back to table of contents Previous article Next article Book ForumFull AccessComprehensive Textbook of AIDS PsychiatryJOYCE SEIKO KOBAYASHI M.D.,JOYCE SEIKO KOBAYASHI M.D.Search for more papers by this author,Published Online:1 Nov 2008https://doi.org/10.1176/appi.ajp.2008.08050719AboutSectionsPDF/EPUB ToolsAdd to favoritesDownload CitationsTrack Citations ShareShare onFacebookTwitterLinked InEmail The Comprehensive Textbook of AIDS Psychiatry will serve as a standard reference for psychiatrists and other mental health professionals providing care for HIV-positive individuals. This is a scholarly work informed by the experience of many pioneers in AIDS care and research, including Dr. Mary Ann Cohen, who first wrote about psychiatric care of people with AIDS when it was a deadly new epidemic without medical treatment. She and her colleagues return to chronicle the vast range of knowledge that has been gained through nearly three decades of research and experience in caring for people with AIDS. The scope of this text ranges from initial biopsychosocial assessment to end-of-life care. The reviews are all extensively researched, and the information is useful for both general practice and HIV specialty care. The clinician will learn to recognize HIV-related cognitive, motor, and psychiatric manifestations of HIV infection, reconsider the treatment of psychiatric disorders of patients who are infected with HIV, identify psychotherapeutic modalities appropriate for persons at various stages of HIV disease, and understand the particular concerns about medication adherence, provider “burnout,” and ethical dilemmas in the treatment of HIV-infected individuals. The chapter on substance use disorders emphasizes the role of specific substances in HIV transmission and high-risk behaviors, and the review of hepatitis C is essential knowledge for treating the nearly 30% of HIV-positive patients who are co-infected. For those interested in more specific comorbidities, there are chapters on HIV-associated renal disease and endocrinopathies, antiretroviral-related metabolic side effects, including lipodystrophies and cardiovascular disease, opportunistic infections associated with AIDS, and two excellent chapters on the neuropathological manifestations of HIV infection. The chapter by editor Jack Gorman and colleagues summarizes advances in psychoneuroimmunology.The drug-drug interactions between antiretroviral and psychotropic medications, such as the impact of ritonavir on levels of psychiatric medications through cytochrome P450 enzyme inhibition or the withdrawal symptoms that can be caused by enzyme induction when nevirapine is administered in conjunction with methadone, are thoroughly reviewed. There are also multiple reference tables summarizing these interactions, although these should be used cautiously and with the understanding that clinical effects can vary significantly between individuals. There is also a brief review of the mechanisms of action of antiretroviral medications, such as interference with viral transcription and replication, which includes protease inhibitors, reverse transcriptase inhibitors, and the newer fusion and integrase inhibitors. However, decisions about specific antiretroviral combination therapies by likely viral resistance based on genotype mutations are beyond the scope of this book.This text establishes a standard of excellence for the care of the individual HIV-positive patient by the individual psychiatrist within the context of the biopsychosocial model. There are also chapters on HIV and youth, serodiscordant couples, and the elderly from a developmental life cycle perspective. However, there is little analysis from a population perspective, such as the myriad issues faced by the population of HIV-positive women of reproductive age. The notable exceptions are a chapter on the homeless HIV-positive population and a provocative chapter suggesting that the disruption of already disadvantaged communities through the "engine" of incarceration in the nation"s prisons accounts for the disproportionate impact of HIV on racial and ethnic minorities. There are estimates that one in five HIV-positive individuals in this country passes through the correctional system, and the chapter proposes system interventions targeting this population. Disparities of care and problems in the health care system are discussed in a chapter on health services that includes an interesting historical perspective on syringe exchange programs as a community intervention.Finally, while there is a rich discussion of the role of psychiatric disorders in HIV transmission and high-risk behaviors in this book, there is little discussion of the role of the psychiatrist in HIV prevention. HIV psychiatry is a prime example of the need for a public mental health perspective that addresses populations as well as individual patients, explores sociocultural roots of illness as well as the medical and psychological, and targets interventions for communities as well as for individuals. Psychiatrists should be engaged in disease prevention as well as diagnosis and treatment.Denver, Colo.Book review accepted for publication May 2008 (doi: 10.1176/appi.ajp.2008.08050719). FiguresReferencesCited byDetailsCited ByNone Volume 165Issue 11 November, 2008Pages 1493-1493THE AMERICAN JOURNAL OF PSYCHIATRY November 2008 Volume 165 Number 11 Metrics PDF download History Published online 1 November 2008 Published in print 1 November 2008

Impact of Multiple Inhibitors or Substrates of Cytochrome P450 2D6 on Plasma Risperidone Levels in Patients on Polypharmacy

Studies that focus on multidrug interactions in natural settings are sparse. In this investigation, data from therapeutic drug monitoring (TDM) were used to study the impact of multiple cytochrome P450 enzyme (CYP) 2D6 substrates and inhibitors on plasma risperidone levels. CYP2D6 catalyzes the conversion of risperidone to the active metabolite 9-OH-risperidone. The question whether CYP2D6 activity is important for the level of the "active moiety" (ie, the sum of risperidone and 9-OH-risperidone) is controversial. Concentration-to-dose (C:D) ratios of risperidone and 9-OH-risperidone in 218 patients were associated with the number of concomitantly used substrates or inhibitors of CYP2D6. The C:D ratios of risperidone in patients with 0, 1, and >1 numbers of CYP2D6 inhibitors were 2.6, 8.5, and 17 nmol L mg, respectively. Differences between the groups were highly significant (P < 0.001). All patients with >1 CYP2D6 inhibitors were administered at least 1 potent CYP2D6 inhibitor, that is fluoxetine, paroxetine, thioridazine, and/or levomepromazine. The C:D ratios of the active moiety (risperidone + 9-OH-risperidone) in patients with 0, 1, and >1 numbers of concomitant CYP2D6 inhibitors were 17, 24, and 30 nmol L mg, respectively (P = 0.001), which was explained by higher levels of risperidone without any change in the levels of 9-OH-risperidone. Concomitant use of 1 or several drugs recognized as substrates for CYP2D6, without any proven inhibitory effect, had no apparent influence on the levels of risperidone or 9-OH-risperidone, suggesting that the risk of drug-drug interactions between different substrates of CYP2D6 is low when used in therapeutic doses. In conclusion, the results suggest that an increase in the number of concomitant inhibitors may be associated with a lower CYP2D6 activity, although the type of inhibitor is probably more important. Drug-dependent inhibition of CYP2D6 increases the active moiety of risperidone. An indication for risperidone TDM should therefore include concomitant medication with established CYP inhibitors.

Inhibition of cytochrome P450 enzymes by rhein in rat liver microsomes

Rhein, an active ingredient extensively found in plants such as Aloe, Cassitora L., rhubarb and so on, has been used for a long time in China. Pharmacological tests revealed that rhein not only had a strong antibacterial action, but also may be useful in cancer chemotherapy as a biochemical modulator. Its therapeutic action and toxicity is still the subject of considerable research. With microsome incubation assays in vitro and HPLC methods, the inhibition of rat liver CYP1A2, CYP2C9, CYP2D6, CYP2E1 and CYP3A enzymes by rhein were studied kinetically. The results showed the most inhibition of CYP2E1 by rhein (K(i) = 10 microm, mixed); CYP3A and CYP2C9 were also inhibited by rhein, K(i) = 30 microm (mixed) and K(i) = 38 microm (mixed), respectively; rhein revealed some inhibition of CYP1A2 (K(i) = 62 microm, uncompetitive) and CYP2D6 (K(i) = 74 microm, mixed). Drug-drug interactions, especially cytochrome P450 (CYP)-mediated interactions, cause an enhancement or attenuation in the efficacy of co-administered drugs. Inhibition of the five major CYP enzymes observed for rhein suggested that changes in pharmacokinetics of co-administered drugs were likely to occur. Therefore, caution should be paid to the possible drug interaction of medicinal plants containing rhein and CYP substrates.

Failure in drug development: the role of inhibition and induction of cytochrome P450 enzymes

For pharmaceuticals, inappropriate pharmacokinetics can result in wide variations in clinical response (Hewitt et al. 2007). Unsatisfactory pharmacokinetic properties are a major reason for the failure of new chemical entities in drug development (Hengstler et al. 2000; Gebhardt et al. 2003). Key mechanisms of compromised pharmacokinetics are inhibition or downregulation (Gueguen et al. 2006; Masubuchi et al. 2006) as well as induction of cytochrome P450 enzymes (Kafert-Kasting et al. 2006; Souma et al. 2006; Davila-Borja et al. 2007; Dorrenhaus et al. 2007; Skupinska et al. 2007). Therefore, besides mechanisms of metabolic activation (Kurebayashi and Ohno 2006; Carmo et al. 2006, 2007; Hermansson et al. 2007; Moustafa et al. 2007), polymorphisms (Hengstler et al. 1998; Kamimura 2006; Ulusoy et al. 2007) and tissue speciWc expression patterns (Hengstler et al. 2005; Bernshausen et al. 2006; Roos et al. 2006; Brulport et al. 2007; Hohme et al. 2007; Nannelli et al. 2007), induction and inhibition of cytochrome P450 enzymes belong to the hottest topics in toxicology (Becker et al. 2006; Klingmuller et al. 2006; Ito et al. 2007; Mitchell et al 2007; Saussele et al. 2007). Considering the outstanding relevance of this topic, the editors are happy to present a comprehensive review of Olavi Pelkonen, Miia Turpeinen, Jukka Hakkola, Paavo Honkakoski, Janne Hukkanen and Hannu Raunio about the current status of inhibition and induction of cytochrome P450 enzymes in this issue of the Archives (Pelkonen et al. 2008). Olavi Pelkonen and Jukka Hakkola, both from the University of Oulu in Finland, are among the most recognized experts in the Weld of cytochrome P450 research. Paavo Honkakoski is Professor of Biopharmacy and Hannu Raunio Professor of Pharmacology at Kuopio University. Janne Hukkanen is vice head physician at Internal Medicine at Oulu University Hospital. Miia Turpeinen defended her PhD thesis in 2006, already received her docentship in 2007 and is now with the Margarete-Fischer-Bosch-Institute in Stuttgart. The authors give a comprehensive overview on inhibitors of CYP1A2, CYP2A6, CYP2B6, CYP2C8, CYP2C9, CYP2C19, CYP2D6, CYP2E1 as well as CYP3A4, and also explain the mechanisms involved and describe strategies of in vitroin vivo extrapolation of inhibitory potencies. After a thorough review of CYP inhibitors, the authors move on to cytochrome P450 induction, presenting the current status of AhR, PXR, CAR mediated mechanisms, and describe the experimental tools to study induction. Having explained the basic mechanisms, the authors present a systematic overview of the induced enzymes and inducers and their clinical relevance. One of the strengths of this review is comprehensive tables of all relevant cytochrome P450 inhibitors and inducers. Reading the review is a must for all scientists working in the Weld of cytochrome P450.

In vitro evaluation of reversible and irreversible cytochrome P450 inhibition: current status on methodologies and their utility for predicting drug-drug interactions.

It is widely accepted that today's practice of polypharmacy inevitably increases the incidence of drug-drug interactions (DDIs). Serious DDI is a major liability for any new chemical entity (NCE) entering the pharmaceutical market. As such, pharmaceutical companies employ various strategies to avoid problematic compounds for clinical development. A key cause for DDIs is the inhibition of cytochrome P450 enzymes (CYPs) that are responsible for metabolic clearance of many drugs. Screening for inhibition potency of CYPs by NCEs has therefore become a routine practice during the drug discovery stage. However, in order to make proper use of DDI data, an understanding of the strengths and weaknesses of the various experimental systems in current use is required. An illustrated review of experimental practices is presented with discussion of likely future developments. The combination of high quality in vitro data generation and the application of in vivo CYP inhibition modelling approaches should allow more informed decisions to be made in the search for drug molecules with acceptable DDI characteristics.

Pathways of Carbamazepine Bioactivation in Vitro. III. The Role of Human Cytochrome P450 Enzymes in the Formation of 2,3-Dihydroxycarbamazepine

Conversion of the carbamazepine metabolite 3-hydroxycarbamazepine (3-OHCBZ) to the catechol 2,3-dihydroxycarbamazepine (2,3-diOHCBZ) followed by subsequent oxidation to a reactive o-quinone species has been proposed as a possible bioactivation pathway in the pathogenesis of carbamazepine-induced hypersensitivity. Initial in vitro phenotyping studies implicated CYP3A4 as a primary catalyst of 2,3-diOHCBZ formation: 2-hydroxylation of 3-OHCBZ correlated significantly (r(2) > or = 0.929, P < 0.001) with CYP3A4/5 activities in a panel of human liver microsomes (n = 14) and was markedly impaired by CYP3A inhibitors (>80%) but not by inhibitors of other cytochrome P450 enzymes (< or = 20%). However, in the presence of troleandomycin, the rate of 2,3-diOHCBZ formation correlated significantly with CYP2C19 activity (r(2) = 0.893, P < 0.001) in the panel of human liver microsomes. Studies with a panel of cDNA-expressed enzymes revealed that CYP2C19 and CYP3A4 were high (S50 = 30 microM) and low (S50 = 203 microM) affinity enzymes, respectively, for 2,3-diOHCBZ formation and suggested that CYP3A4, but not CYP2C19, might be inactivated by a metabolite formed from 3-OHCBZ. Subsequent experiments demonstrated that preincubation of 3-OHCBZ with human liver microsomes or recombinant CYP3A4 led to decreased CYP3A4 activity, which was both preincubation time- and concentration-dependent, but not inhibited by inclusion of glutathione or N-acetylcysteine. CYP3A4, CYP3A5, CYP3A7, CYP2C19, and CYP1A2 converted [14C]3-OHCBZ into protein-reactive metabolites, but CYP3A4 was the most catalytically active enzyme. The results of this study suggest that CYP3A4-dependent secondary oxidation of 3-OHCBZ represents a potential carbamazepine bioactivation pathway via formation of reactive metabolites capable of inactivating CYP3A4, potentially generating a neoantigen that may play a role in the etiology of carbamazepine-induced idiosyncratic toxicity.

Update on Echinocandin Antifungals

Echinocandins are semisynthetic lipopeptides that competitively inhibit an essential cell wall component of Candida and aspergillus. They are generally inactive against other fungi. Resistance to these agents is infrequent to date. Echinocandins exhibit low oral bioavailability and are available only as parenteral formulations that are dosed once daily. None of the echinocandins serve as major substrates, inducers, or inhibitors of cytochrome P450 enzymes or the P-glycoprotein transport system; thus they have a low potential for serious drug-drug interactions. In candidemia trials, all echinocandins showed similar rates of success and were as efficacious as fluconazole, amphotericin B, or lipid formulations of amphotericin B. Caspofungin and micafungin have been studied as single-agent therapy in patients with invasive aspergillosis and will likely remain as second-line agents. However, because of their unique mechanism of action, echinocandins are ideally suited for use in combination with polyenes or azoles and are likely to be used increasingly in immunosuppressed hosts despite a current lack of controlled trials demonstrating efficacy. Limited experience suggests that caspofungin and micafungin are safe to use in pediatric patients. Hospital formulary committees are likely to view the three echinocandins as equivalent agents and place the least expensive agent on formulary.

Dabigatran Etexilate

Dabigatran etexilate is an orally administered prodrug of dabigatran, which is a potent, concentration-dependent inhibitor of thrombus formation and thrombin-induced platelet aggregation. Dabigatran etexilate pharmacokinetics were linear across a wide dosage range. There were no clinically important pharmacokinetic interactions with digoxin (a P-glycoprotein substrate), pantoprazole (a proton-pump inhibitor) or drugs that are substrates and/or inhibitors of hepatic cytochrome P450 enzymes. In two large, randomized, double-blind trials of the prevention of venous thromboembolism (VTE) in patients undergoing total hip or total knee replacement surgery, orally administered dabigatran etexilate 220 mg/day was noninferior to subcutaneous enoxaparin sodium 40 mg/day for the primary composite endpoint of total VTE events or all-cause mortality during the treatment period. There were no significant differences between dabigatran etexilate and enoxaparin sodium in major VTE events and VTE-related mortality. Across trials, < or =0.5% of patients experienced a symptomatic pulmonary embolus or died. Dabigatran etexilate was generally well tolerated. In patients undergoing total hip or total knee replacement surgery, there was no significant difference between dabigatran etexilate and enoxaparin sodium recipients in the incidence of major or minor bleeding.

Targeted chronic myeloid leukemia therapy: Seeking a cure

This article focuses on imatinib, how it has altered CML therapy, clinical trials that are the basis for its efficacy, and adverse effects associated with its current clinical use. Maintaining patients with CML in chronic phase (CP) yields the prospect of improved long-term survival. As recently as 1993, CML was limited to treatment with standard cytoreductive therapies. These therapies provide temporary disease control but do not alter progression to advanced disease with a median survival ranging 45 to 55 months from diagnosis. In the 1990s, immunologically based therapy with interferon alpha (IFNalpha) therapy was shown to be superior to cytoreductive therapies with a median survival of 60 to 90 months. Allogeneic hematopoietic stem cell transplant (HSCT) has offered curative potential for patients with CML; however, the median age of diagnosis of 55 years, the lack of suitable donors, and the morbidity of the procedure precludes widespread applicability of this treatment. Imatinib, the first approved tyrosine kinase inhibitor, functions by blocking the ATP binding site on the BCR-ABL kinase. It was first shown to be efficacious in patients who failed IFNalpha and then tested as a front line therapy (the International Randomized Study of Interferon [IRIS] trial). The five year follow up on the IRIS trial found that the responses were durable with progression free survival estimated at 93%. Imatinib has been found to have a lower rate of hematologic response and shorter duration of response in patients with advanced disease. Currently patients in blast crisis (BC) have the option to undergo a number of induction chemotherapies, such as etoposide, cytarabine, carboplatin (VAC) with the hope of temporarily restoring the patient to CP in preparation for HSCT. Imatinib, when administered at the standard dose of 400 mg/day is relatively well tolerated with major toxicities limited to myelo-suppression, edema, GI upset, rash, and muscle pain. Many of these toxicities are managed by decreasing the dose until the toxicity resolves. Imatinib is an inhibitor of cytochrome P450 enzymes necessitating careful monitoring of concomitant medications metabolized by these enzymes. Resistance may develop to imatinib most often caused by the evolution of mutations blocking imatinib interactions with the BCR-ABL adenosine triphosphate (ATP) binding site. The second generation BCR-ABL inhibitor, dasatinib, can block the activity of many of these mutations; however, the T315I mutation, at present, is resistant to all available kinase inhibitors. Experimental drugs that block this mutation are just entering phase two clinical trials. The development of therapeutic agents targeting BCR-ABL has revolutionized the treatment of chronic myeloid leukemia (CML). Imatinib has successfully allowed CML patients to remain in CP for at least five years in 90% of patients. Dasatinib has activity against a number of Imatinib-resistant mutants providing an additional therapeutic option for these patients.

SAFETY EVALUATION OF A COMBINATION, DEFINED EXTRACT OF SCUTELLARIA BAICALENSIS AND ACACIA CATECHU

ABSTRACT A combined extract containing primarily baicalin from Scutellaria baicalensis and (+)-catechin from Acacia catechu used in both joint supplements and a prescription medical food was tested for safety. Cytotoxicity testing in THP-1 monocytes showed limited cell death compared to nonsteroidal anti-inflammatory drugs (NSAIDs). Acute and subchronic toxicity testing demonstrated no abnormalities in any toxicological end points examined including animal body weights, gross organ pathology and tissue histology, and blood chemistries or serology. The extract, when dosed in Fischer 344 rats, a model for gastric toxicity of NSAIDs, showed no evidence of ulceration. No mutagenicity or drug interactions were seen by AMES and cytochrome P450 enzyme inhibition, respectively. When the extract was compared with placebo after administration to a healthy human population, no changes in blood chemistry or serology were observed. Based on these findings, the combined extract with baicalin and catechin appears to possess a safety profile that justifies further testing in humans. PRACTICAL APPLICATIONS A specific ratio of Scutellaria baicalensis and Acacia catechu metabolically balances the inhibition of cyclooxygenase-1 and cyclooxygenase-2 (COX-2) enzymes and provides a comparable amount of 5-lipoxygenase (5-LOX) enzyme inhibition in the conversion of arachidonic acid (AA) to inflammatory metabolites. This mechanism of action may yield fewer side effects than traditional nonsteroidal anti-inflammatory drugs and selective COX-2 inhibitors produced by their generally specific inhibition of either COX enzyme. Simultaneous inhibition of 5-LOX may also avoid a “shunt” of arachidonic acid metabolism down the LOX pathway, thus reducing side effects related to excess, unopposed production of inflammatory vasoconstrictive as well as chemoattractive leukotrienes. The net result should be an effective anti-inflammatory metabolic product for the management of osteoarthritis with an improved toxicity profile and lower requirement for use with gastroprotective therapies.

In Vitro Metabolism of Isoliquiritigenin by Human Liver Microsomes

Isoliquiritigenin (2',4',4-trihydroxychalcone), a chalcone found in licorice root and other plants, has shown potent antitumor, antioxidant, and phytoestrogenic activity in vitro. In preparation for in vivo studies, the metabolism of isoliquiritigenin by human liver microsomes was investigated, and seven phase 1 metabolites were identified. In addition to aromatic hydroxylation that occurred on the A or B ring to form 2',4,4',5'-tetrahydroxychalcone or butein, respectively, reduction of the carbon-carbon double bond of an alpha,beta-unsaturated ketone and cyclization occurred to form 2',4,4'-trihydroxydihydrochalcone and (Z/E)-6,4'-dihydroxyaurone. All metabolites were characterized and identified by using liquid chromatography-tandem mass spectrometry with comparison to authenticated compounds. Finally, monoclonal antibody inhibitors of specific human cytochrome P450 (P450) enzymes and recombinant human P450 enzymes were used to identify the enzymes responsible for the formation of the major mono-oxygenated metabolites, and P450 2C19 was found to be a significant enzyme in the formation of butein from isoliquiritigenin, which also has anticancer activity. Cytochromes P450, reactive oxygen species, and peroxidases can all contribute to the formation of (Z/E)-6,4'-dihydroxyaurone in human liver microsomes.

A Comparison of 2-Phenyl-2-(1-piperidinyl)propane (PPP), 1,1′,1″-Phosphinothioylidynetrisaziridine (ThioTEPA), Clopidogrel, and Ticlopidine as Selective Inactivators of Human Cytochrome P450 2B6

The use of selective chemical inhibitors of human cytochrome P450 (P450) enzymes represents a powerful method by which the relative contributions of various human P450 enzymes to the metabolism of drugs can be determined. However, the identification of CYP2B6 in the metabolism of drugs has been more challenging because of the lack of a well established inhibitor of this enzyme. In this report, we describe the selectivity of 2-phenyl-2-(1-piperidinyl)propane (PPP) as an inactivator of CYP2B6 and compare this selectivity versus other CYP2B6 inactivators: 1,1',1''-phosphinothioylidynetrisaziridine (thioTEPA), clopidogrel, and ticlopidine. Values of K(I) and k(inact) for PPP were 5.6 microM and 0.13/min for bupropion hydroxylase catalyzed by pooled human liver microsomes, and values for thioTEPA were similar (4.8 microM and 0.20/min, respectively). Intrinsic inactivation capability was considerably greater for clopidogrel because of a greater k(inact) value (1.9/min). Ticlopidine was potent with K(I) and k(inact) values of 0.32 microM and 0.43/min, respectively. The selectivity of these four agents for CYP2B6 was determined by testing their effects on other human P450 enzyme activities using conditions that yield approximately 90% inactivation of CYP2B6 activity. The results showed that preincubation of human liver microsomes with PPP at 30 microM for 30 min provided more selective inhibition for CYP2B6 than thioTEPA, clopidogrel, and ticlopidine. Furthermore, the use of clopidogrel is complicated by the observation that this agent is not stable in the presence of human liver microsomes, even without addition of NADPH. Therefore, PPP can serve as a selective chemical inactivator of CYP2B6 and be used to define the role of CYP2B6 in the metabolism of drugs.

Tricyclic antidepressant pharmacology and therapeutic drug interactions updated

New data on the pharmacology of tricyclic antidepressants (TCAs), their affinities for human cloned CNS receptors and their cytochrome P450 enzyme inhibition profiles, allow improved deductions concerning their effects and interactions and indicate which of the TCAs are the most useful. The relative toxicity of TCAs continues to be more precisely defined, as do TCA interactions with selective serotonin reuptake inhibitors (SSRIs). TCA interactions with monoamine oxidase inhibitors (MAOIs) have been, historically, an uncertain and difficult question, but are now well understood, although this is not reflected in the literature. The data indicate that nortriptyline and desipramine have the most pharmacologically desirable characteristics as noradrenaline reuptake inhibitors (NRIs), and as drugs with few interactions that are also safe when coadministered with either MAOIs or SSRIs. Clomipramine is the only available antidepressant drug that has good evidence of clinically relevant serotonin and noradrenaline reuptake inhibition (SNRI). These data assist drug selection for monotherapy and combination therapy and predict reliably how and why pharmacodynamic and pharmacokinetic interactions occur. In comparison, two newer drugs proposed to have SNRI properties, duloxetine and venlafaxine, may have insufficient NRI potency to be effective SNRIs. Combinations such as sertraline and nortriptyline may therefore offer advantages over drugs like venlafaxine that have fixed ratios of SRI/NRI effects that are not ideal. However, no TCA/SSRI combination is sufficiently safe to be universally applicable without expert knowledge. Standard texts (e.g. the British National Formulary) and treatment guidelines would benefit by taking account of these new data and understandings.

Protective effects of xanthohumol against the genotoxicity of benzo(a)pyrene (BaP), 2-amino-3-methylimidazo[4,5-f]quinoline (IQ) and tert-butyl hydroperoxide ( t-BOOH) in HepG2 human hepatoma cells

Xanthohumol is the major prenylated flavonoid present in the hop plant Humulus lupulus L. (Cannabinaceae) and a common ingredient of beer. Recently, xanthohumol has gained considerable interest due to its potential cancer chemo-preventive effect. The aim of this study was to reveal the possible anti-genotoxic activity of xanthohumol in metabolically competent human hepatoma HepG2 cells, by use of the comet assay. Xanthohumol by itself was neither cytotoxic nor genotoxic to the cells at concentrations below 10 μM. However, a significant protective effect against the pro-carcinogens benzo(a)pyrene (BaP) and 2-amino-3-methylimidazo[4,5-f]quinoline (IQ) was observed at concentrations as low as 0.01 μM. In cells treated with xanthohumol in combination with tert-butyl hydroperoxide ( t-BOOH) – an inducer of reactive oxygen species (ROS) – no protective effect was observed and xanthohumol also showed no significant scavenging activity against 1,1-diphenyl-2-picrylhydrazyl (DPPH) radicals. On the other hand, HepG2 cells pre-treated with xanthohumol showed significantly reduced levels of t-BOOH-induced DNA strand breaks, indicating that its protective effect is mediated by induction of cellular defence mechanisms against oxidative stress. As xanthohumol is known to be an effective inhibitor of cytochrome P450 enzymes and an inducer of NAD(P)H: quinone reductase (QR), our findings can be explained by an inhibition of metabolic activation of pro-carcinogens and/or by induction of carcinogen-detoxifying and anti-oxidative enzymes by xanthohumol. These results provide evidence that xanthohumol displays anti-genotoxic activity in metabolically competent human cells.