The Exeter Contemporary flanged cemented acetabular component in primary total hip arthroplasty.

AIMS: We report on the outcome of the Exeter Contemporary flanged cemented all-polyethylene acetabular component with a mean follow-up of 12 years (10 to 13.9). This study reviewed 203 hips in 194 patients. 129 hips in 122 patients are still in situ; 66 hips in 64 patients were in patients who died before ten years, and eight hips (eight patients) were revised. Clinical outcome scores were available for 108 hips (104 patients) and radiographs for 103 hips (100 patients). PATIENTS AND METHODS: A retrospective review was undertaken of a consecutive series of 203 routine primary cemented total hip arthroplasties (THA) in 194 patients. RESULTS: There were no acetabular component revisions for aseptic loosening. Acetabular revision was undertaken in eight hips. In four hips revision was necessitated by periprosthetic femoral fractures, in two hips by recurrent dislocation, in one hip for infection and in one hip for unexplained ongoing pain. Oxford and Harris hip scores demonstrated significant clinical improvement (all p < 0.001). Radiolucent lines were present in 37 (36%) of the 103 acetabular components available for radiological evaluation. In 27 of these, the line was confined to zone 1. No component had migrated. CONCLUSION: Kaplan-Meier survivorship, with revision for aseptic loosening as the endpoint, was 100% at 12.5 years and for all causes was 97.8% (95% confidence interval 95.6 to 100) when 40 components remained at risk. The Exeter Contemporary flanged cemented acetabular component demonstrates excellent survivorship at 12.5 years. TAKE HOME MESSAGE: The Exeter Contemporary flanged cemented acetabular component has excellent clinical outcomes and survivorship when used with the Exeter stem in total hip arthroplasty.

Role of reactive metabolites in drug-induced hepatotoxicity.

Drugs are generally converted to biologically inactive forms and eliminated from the body, principally by hepatic metabolism. However, certain drugs undergo biotransformation to metabolites that can interfere with cellular functions through their intrinsic chemical reactivity towards glutathione, leading to thiol depletion, and functionally critical macromolecules, resulting in reversible modification, irreversible adduct formation, and irreversible loss of activity. There is now a great deal of evidence which shows that reactive metabolites are formed from drugs known to cause hepatotoxicity, such as acetaminophen, tamoxifen, isoniazid, and amodiaquine. The main theme of this article is to review the evidence for chemically reactive metabolites being initiating factors for the multiple downstream biological events culminating in toxicity. The major objectives are to understand those idiosyncratic hepatotoxicities thought to be caused by chemically reactive metabolites and to define the role of toxic metabolites.

Measurement of cortisol metabolites in faeces of ruminants.

Twenty-one metabolites were detected in faecal samples collected after infusion of (14C)cortisol into the jugular vein of sheep. Using high-performance liquid chromatography/radiometric analysis plus mass spectrometry. One group of metabolites had molecular weights of between 302 and 308, and another group of 350, which indicates that the substances have a C19O3 or a C21O4 structure. Therefore, an enzyme immunoassay against 5beta-androstane-3alpha-ol-11,17-dione-17-CMO:BSA was established. Faecal samples were collected from 10 cows immediately after transport and then during a course in which non-invasive diagnostic procedures were being taught (course 1). For comparison, faeces were sampled from another 5 cows that were being used for teaching invasive procedures (course 2). Six cows from a university farm served as controls. In the animals used in course 1, the highest concentrations of cortisol metabolites were measured immediately after transport to the university (median value: 2.2 micromol/kg faeces). During the first 5 days at the university, the concentrations decreased to 0.52 micromol/kg (median) and remained at this level during the rest of the course. The median concentration in the samples that were taken during coursc 2 (collected about 2 months after transport) was 0.48 micromol/kg. There was no significant difference in the excretion of cortisol metabolites between these cows and the controls. We conclude from these data that, using the enzyme immunoassay against 5beta-androstane-3alpha-ol-11,17-dione-17-CMO, we were able to detect transport/novel environment stress but not the potential disturbance that cows experience during diagnostic procedures.

Enzyme-induction dependent bioactivation of troglitazone and troglitazone quinone in vivo.

Troglitazone (TGZ), a 2,4-thiazolidinedione antidiabetic, causes hepatotoxicity in 1.9% of patients. TGZ is an inducer of, and substrate for, hepatic P450 3A. Microsomal metabolism yields a benzoquinone (TGZQ) and reactive intermediates. Kassahun et al. [Kassahun et al. (2001) Chem. Res. Toxicol. 14, 62-70] have trapped the intermediates as thioester, thioether, and disulfide conjugates of glutathione and found five conjugates in rat bile. The thioether was substituted in the chromane moiety. We have investigated the effect of the P450 3A inducer, dexamethasone (DEX), on metabolism of TGZ and TGZQ in rats and assessed the compounds' cytotoxicity. TGZ-glucuronide and sulfonate were confirmed as principal biliary metabolites of TGZ (50 mg/kg, iv). Bile from noninduced animals also contained a TGZ-glutathione thioether adduct (ML3) but it was substituted in the thiazolidinedione moiety. Pretreatment with DEX (50 mg/kg/day for 3 days) resulted in a 2-5-fold increase in the biliary concentration of ML3 and a 2-fold increase in the concentration of TGZQ, which was commensurate with the induction of hepatic P450 3A. Three of the known glutathione-conjugated metabolites were also found. TGZQ (50 mg/kg, iv) was metabolized to an analogue of one of the TGZ-glutathione thioesters and a glutathione adduct of TGZQ hydroquinone after DEX pretreatment. TGZ quinol glucuronide was a biliary metabolite of TGZ and TGZQ. Its formation would represent deactivation of TGZQ. TGZ was toxic to rat hepatocytes and Hep-G2 cells at concentrations exceeding 50 and 25 microM, respectively, after 24 h. In contrast, TGZQ was nontoxic to rat hepatocytes and toxic to Hep G2 cells only at concentrations exceeding 100 microM. Our results show that TGZQ as well as TGZ yields reactive metabolites in vivo, and that bioactivation is enhanced by induction of P450 3A. However, hepatotoxicity is unlikely to be due to either TGZQ or its metabolites.

Efficient preparations of the beta-glucuronides of dihydroartemisinin and structural confirmation of the human glucuronide metabolite.

New and greatly improved preparations of the 12alpha,1'beta- (5) and 12beta,1'beta- (6) glucuronides of dihydroartemisinin (DHA, 2) are reported using anomeric hydroxy and imidate glucuronate intermediates. Comparison of the synthetic and natural materials shows that the human metabolite of DHA is the 12alpha-epimer 5.

The problem of college drinking: insights from a developmental perspective.

This article represents the proceedings of a symposium at the 2000 RSA Meeting in Denver, Colorado. John Schulenberg and Jennifer L. Maggs were Organizers. Stephen W. Long was Chair and provided opening remarks. The presentations were: (1) I'm not a drunk, just a college student: Binge drinking during college as a developmental disturbance, by John Schulenberg; (2) Course of alcohol use disorders during college, by Kenneth J. Sher; (3) How do students experience alcohol and its effects? Positive versus negative expectancies and consequences, by Jennifer L. Maggs; and (4) Brief intervention in the context of developmental trends in college drinking, by G. Alan Marlatt. Critique and commentary were provided by Robert A. Zucker.

Comparison of the modulatory effects of human and rat liver microsomal metabolism on the estrogenicity of bisphenol A: implications for extrapolation to humans.

Bisphenol A [BPA, 2,2-bis(4-hydroxyphenyl)propane], a xenoestrogen, is a monomer for the synthesis of polycarbonate plastics, epoxy resins, and composites. Metabolism of BPA to the monoglucuronide will determine the extent of its estrogenicity in vivo. Investigation of the metabolism of BPA (500 microM) by isolated female rat hepatocytes confirmed the formation of BPA glucuronide as the major metabolite. There was a significant difference (p < 0.05) between the V(max) (mean +/- S.E.M., n = 4) of glucuronidation by pooled male or female human (four livers in each case) and immature female rat liver microsomes (5.9 +/- 0.4, 5.2 +/- 0.3, and 31.6 +/- 8.1 nmol/min/mg of protein, respectively). Estrogenic activity of BPA, assessed in a coupled microsomal metabolism-yeast estrogenicity assay, was decreased 3- and 7-fold following glucuronidation by human female and immature female rat liver microsomes, respectively. Incubations of BPA with pooled human or rat liver microsomes, in the presence of NADPH, resulted in the formation of 5-hydroxybisphenol A [2-(4,5-dihydroxyphenyl)-2-(4-hydroxyphenyl)propane], which was 10-fold less potent than BPA in the yeast estrogenicity assay. However, there was insufficient turnover to achieve a significant effect on the estrogenic activity of BPA. Because human liver microsomes did not glucuronidate BPA as extensively as the rat liver microsomes, estrogen target tissues in humans may be subject to greater exposure to BPA than the tissues of the immature female rats used for assessing estrogenicity of xenobiotics.

Assessment of the effects of metabolism on the estrogenic activity of xenoestrogens: a two-stage approach coupling human liver microsomes and a yeast estrogenicity assay.

Concern that the reproductive health of humans is being affected by exposure to xenoestrogens has led to the development of various in vitro and in vivo screening assays for the identification of suspected xenoestrogens. However, the estrogenic activity of a chemical determined in vitro may not necessarily predict its activity in vivo if the chemical is metabolized during the assay and/or in vivo. Therefore, to investigate the role of metabolism in modulating the estrogenic activity of suspected xenoestrogens, we have devised a two-stage approach coupling incubations with either human or rat hepatic microsomes with a yeast estrogenicity (transcription) assay. We have assessed the activity of the proestrogenic pesticide 99.5% methoxychlor [1,1,1-trichloro-2,2-bis-(4-methoxyphenyl)ethane, MXC] (EC(50) = 4.45 +/- 1.9 ,icroM, n = 6) and a structural analog, methoxybisphenol A [2,2-bis-(4-methoxyphenyl) propane, MBPA], in the yeast estrogenicity assay and also established that yeast (Saccharomyces cerevisiae), unlike human liver microsomes, are not able to demethylate MXC or MBPA to estrogenic metabolites. This indicates that the proestrogen MXC has weak intrinsic estrogenic activity. Using 99.5% MXC and 17beta-estradiol as paradigms, we have demonstrated how metabolism can enhance or suppress, respectively, estrogenic activity. The effect of metabolism on the activities of the weak xenoestrogens 3,17beta-bisdesoxyestradiol [1,3,5(10)-estratriene] and 6-hydroxytetralin (5,6,7,8-tetrahydro-2-naphthol) was also assessed. This two-stage approach can distinguish the estrogenic activity of a suspect chemical from the activity due to its more, or less, active metabolites and will aid in the evaluation of novel xenoestrogens and, more importantly, proestrogens.

Synthesis, antimalarial activity, biomimetic iron(II) chemistry, and in vivo metabolism of novel, potent C-10-phenoxy derivatives of dihydroartemisinin.

The combination of TMSOTf and AgClO(4) promotes the efficient C-10-phenoxylation of dihydroartemisinin (3) in good chemical yield and excellent stereoselectivity. All of the new phenoxy derivatives have potent in vitro antimalarial activity. On the basis of the excellent yield and stereoselectivity obtained for the p-trifluoromethyl derivative 7b, this compound and the parent phenyl-substituted derivative 5b were selected for in vivo biological evaluation against Plasmodium berghei in the mouse model and for metabolism studies in rats. Compound 7b demonstrated excellent in vivo antimalarial potency with an ED(50) of 2.12 mg/kg (cf. artemether = 6 mg/kg) versus P. berghei. Furthermore, from preliminary metabolism studies, this compound was not metabolized to dihydroartemisinin; suggesting it should have a longer half-life and potentially lower toxicity than the first-generation derivatives artemether and arteether. From biomimetic Fe(II)-catalyzed decomposition studies and ESR spectroscopy, the mechanism of action of these new lead antimalarials is proposed to involve the formation of both primary and secondary C-centered cytotoxic radicals which presumably react with vital parasite thiol-containing cellular macromolecules.

Metabolism of lamotrigine to a reactive arene oxide intermediate.

Lamotrigine [3,5-diamino-6-(2,3-dichlorophenyl)-1,2,4-triazine] is an antiepileptic drug associated with hypersensitivity reactions which are thought to be an immunological response to metabolically generated drug-protein adducts. The o-dichlorophenyl moiety is a potential site for bioactivation of the drug to an arene oxide. The metabolites of [(14)C]lamotrigine (78 micromol/kg, iv) in adult male Wistar rats were characterized with particular reference to thioether derivatives of an epoxide intermediate. Biliary recovery of radioactivity from anesthetized and cannulated animals was 7.3 +/- 3.0% (mean +/- SD, n = 4) of the dose over 4 h; 5.5 +/- 0.5% was recovered in bladder urine after 4 h. Bile contained [(14)C]lamotrigine (1.4 +/- 0.3%), a glutathione adduct of [(14)C]dihydrohydroxylamotrigine (1.8 +/- 0.3%), i.e., an adduct of an arene oxide, and the glutathione (1.5 +/- 0.7%), cysteinylglycine (1.9 +/- 0.5%), and N-acetylcysteine (0.4 +/- 0.2%) adducts of [(14)C]lamotrigine. Formation of the thioether metabolites was partially blocked by the cytochrome P450 inhibitor, ketoconazole. Urine contained [(14)C]lamotrigine (4.5 +/- 0.5%) and [(14)C]lamotrigine N-oxide (0.9 +/- 0.2%). The radiolabeled material in skin (15.6 +/- 1.4%) was almost entirely [(14)C]lamotrigine. Isolated rat hepatocytes achieved a low rate of turnover to the glutathione adduct and N-oxide. However, neither rat nor human liver microsomes catalyzed NADPH-dependent irreversible binding. Lamotrigine can be bioactivated to an arene oxide by rat hepatocytes in the absence of a major competing pathway such as N-glucuronidation. Inhibition of N-glucuronidation has been associated with an increased risk of skin reactions in patients.

Obstacles to the prediction of estrogenicity from chemical structure: assay-mediated metabolic transformation and the apparent promiscuous nature of the estrogen receptor.

Information on structure-activity relationships (SAR) and pathways of metabolic activation would facilitate the preliminary screening of chemicals for estrogenic potential. Published crystallographic studies of the estrogen receptor (ER) imply an essential role of the two hydroxyl groups on estradiol (17beta-E(2)) for its binding to ER. The influence of these hydroxyl groups on ER binding and estrogenicity was evaluated by the study of 17beta-E(2) with one or both of these hydroxyl groups removed (17beta-desoxyestradiol and 3, 17beta-bisdesoxyestradiol, respectively). 6-Hydroxytetralin (17beta-E(2) with its C- and D-rings removed) and other synthetic estrogens were also studied. The estrogenicity assays comprised a yeast ER-mediated transcription assay, mammalian cell transcription assays incorporating either ER alpha or ER beta, and the immature rat uterotrophic assay. With the exception of 6-hydroxytetralin in the uterotrophic assay, all the chemicals were active in all the assays. Hydroxylation of the two desoxy compounds to estradiol was shown to occur in immature female rats, but metabolism was not implicated in the responses observed in the ER-binding and yeast systems. It is concluded that the 3-hydroxyl and 17beta-hydroxyl groups of 17beta-E(2) are not absolute requirements for estrogenicity. It would therefore be of value to the derivation of SAR for estrogenicity were the crystal structure of the bisdesoxy-E(2)/ER complex to be evaluated.

Major inter-species differences in the rates of O-sulphonation and O-glucuronylation of alpha-hydroxytamoxifen in vitro: a metabolic disparity protecting human liver from the formation of tamoxifen-DNA adducts.

Tamoxifen is a hepatic genotoxin in rats and mice but a hepatocarcinogen only in rats. It is not associated with DNA adducts and liver tumours in patients. The proposed major pathway for its bioactivation in rats involves alpha-hydroxylation, O-sulphonation and generation of a carbocation that reacts with DNA. Rat liver microsomes catalyse alpha-hydroxylation at approximately 2- and 4-fold the rate achieved by human and murine liver microsomes, respectively. O-glucuronylation will deactivate alpha-hydroxytamoxifen and compete with sulphonation. Rates of O-sulphonation of alpha-hydroxytamoxifen in hepatic cytosol have been determined by a HPLC assay of substrate-dependent 3'-phosphoadenosine 5'-phosphate production. The rank order of O-glucuronylation in hepatic microsomes was estimated by HPLC-mass spectrometry. The rate of sulphonation of trans-alpha-hydroxytamoxifen (25 microM) in cytosol from adult female Sprague-Dawley rats and CD1 mice was 5.3 +/- 0.8 and 3.9 +/- 0.5 pmol/min/mg protein (mean +/- SD, n = 3), respectively. In cytosol fractions from women aged 40-65 years, the rate was 1.1 +/- 0.4 pmol/min/mg protein (mean +/- SD, n = 6). The K(m) for trans-alpha-hydroxytamoxifen in rat, mouse and human cytosol was 84. 6 +/- 3.8, 81.4 +/- 4.6 and 104.3 +/- 5.6 microM (mean +/- SD, n = 3), respectively; the corresponding V:(max) values were 22.4 +/- 3.4, 17.1 +/- 3.1 and 6.3 +/- 1.9 pmol/min/mg protein. These K:(m) were similar to a value obtained by others using purified rat liver hydroxysteroid sulphotransferase a. Turnover of the cis epimer was too slow for accurate determination of rates. Sulphonation of trans-alpha-hydroxytamoxifen in human uterine cytosol was undetectable. The rank order of O-glucuronylation of trans-alpha-hydroxy- tamoxifen in liver microsomes was human > > mouse > rat. In combination, lower rates of alpha-hydroxylation and O-sulphonation and a higher rate of O-glucuronylation in human liver would protect patients from the formation of tamoxifen-DNA adducts.

Preventing alcohol misuse: the impact of refusal skills and norms.

The purpose of this study was to determine the extent to which refusal skills and norm setting mediated the impact of a school-based prevention program from the Alcohol Misuse Prevention Study (AMPS) on adolescent alcohol overindulgence. The AMPS is a randomized, pre-post, experimental-control study. Respondents in the present study included 6th through 10th graders (ns ranged from 232 to 371). Structural equation modeling analyses using EQS indicated that norm setting mediated the effect of the intervention on alcohol overindulgence at the 7th through the 8th grade and at the 8th through the 10th grade. In contrast, although the prevention program served to increase refusal skills, refusal skills did not mediate the effect of the program on alcohol misuse.

Biliary metabolites of beta-artemether in rats: biotransformations of an antimalarial endoperoxide.

beta-Artemether (AM), the O-methyl ether prodrug of dihydroartemisinin (DHA), is an endoperoxide antimalarial. The biliary metabolites of AM in adult male Wistar rats were characterized with particular reference to potential antimalarial compounds and stable derivatives of free radical intermediates. [13-(14)C]-AM (35 micromol kg(-1), i.v.) was administered to anesthetized rats. Within 0 to 3 h, 38.6 +/- 4.8% (mean +/- S.D., n = 6) of the radiolabel was recovered in bile; the 0- to 5-h recovery was 42.3 +/- 4.3%. The major metabolites (0-3 h) were the glucuronides of 9alpha-hydroxyAM (33.4 +/- 6.8% biliary radioactivity) and alpha-DHA (22.5 +/- 4.4%); four stereochemically unassigned monohydroxyAM glucuronides (II, 3.1 +/- 0.9; IV, 4.4 +/- 1.7%; V, 21.4 +/- 3.0%; VI, 3.0 +/- 1.1%) and a dihydroxyAM glucuronide (6.0 +/- 2.1%) were also identified. A sixth monohydroxyAM glucuronide (VIIa) and desoxyDHA glucuronide were detected in trace amounts. The furano acetate isomer of DHA glucuronide, indicative of the formation of a radical intermediate, was also found in trace amounts. O-methyl substitution of DHA favors ring hydroxylation in vivo. However, the principal hydroxylated metabolite, 9alpha-hydroxyAM, is unlikely to possess significant antimalarial activity.

Methapyrilene hepatotoxicity is associated with increased hepatic glutathione, the formation of glucuronide conjugates, and enterohepatic recirculation.

The mechanisms by which acute administration of methapyrilene, an H(1)-receptor antihistamine causes periportal necrosis to rats are unknown. This study investigated the role of the hepato-biliary system in methapyrilene hepatotoxicity following daily administration of 150 mg/kg per day over 3 consecutive days. Biliary metabolites of methapyrilene were tentatively identified. In male Han Wistar rats administration of methapyrilene significantly increased hepatic reduced glutathione (GSH) to 140% of control levels 24 h following the last dose. There were no significant changes in the activities of glutathione-related enzymes, glutathione peroxidase (GPx) and reductase (GSH), glutathione S-transferase (GST), and gamma-glutamyl cysteine synthetase (gamma-GCS) over 3 days of methapyrilene administration. Methapyrilene treatment resulted in no significant increase in excretion of biliary oxidized glutathione (GSSG), a sensitive marker of oxidative stress in vivo, following the third dose. [3H]Methapyrilene-derived radioactivity was detected in bile, to a greater extent than in feces, indicating that methapyrilene and/or metabolites underwent enterohepatic recirculation. Cannulation and exteriorization of the bile duct (to interrupt enterohepatic recirculation) afforded some protection against the hepatotoxicity, assessed by clinical chemistry and histopathology. Liquid chromatography-mass spectrometry (LC-MS) analysis of bile indicated the presence of unmetabolized methapyrilene, methapyrilene O-glucuronide and desmethyl methapyrilene O-glucuronide. These data demonstrate that acute methapyrilene hepatotoxicity in vivo is not a consequence of GSH depletion, or oxidative stress, but that enterohepatic recirculation of biliary metabolites may be important. Progressive exposure to non-oxidizing, reactive metabolic intermediates may be responsible for hepatotoxicity.

Metabolism of the antimalarial endoperoxide Ro 42-1611 (arteflene) in the rat: evidence for endoperoxide bioactivation.

Ro 42-1611 (arteflene) is a synthetic endoperoxide antimalarial. The antimalarial activity of endoperoxides is attributed to iron(II)-mediated generation of carbon-centered radicals. An alpha, beta-unsaturated ketone (enone; 4-[2',4' bis(trifluoromethyl)phenyl]-3-buten-2-one), obtained from arteflene by reaction with iron(II), was identified previously as the stable product of a reaction that, by inference, also yields a cyclohexyl radical. The activation of arteflene in vivo has been characterized with particular reference to enone formation. [14C]Arteflene (35 micromol/kg) was given i.v. to anesthetized and cannulated male rats: 42.2 +/- 7.0% (mean +/- S.D., n = 7) of the radiolabel was recovered in bile over 5 h. In the majority of rats, the principal biliary metabolites were 8-hydroxyarteflene glucuronide (14.2 +/- 3. 9% dose, 0-3 h) and the cis and trans isomers of the enone (13.5 +/- 4.6% dose, 0-3 h). In conscious rats, 15.3 +/- 1.6% (mean +/- S.D., n = 8) of the radiolabel was recovered in urine over 24 h. The principal urinary metabolite appeared to be a glycine conjugate of a derivative of the enone. Biliary excretion of the glucuronide, but not of the enones, was inhibited by ketoconazole. 8-Hydroxyarteflene was formed extensively by rat and human liver microsomes but no enone was found. Bioactivation is a major pathway of arteflene's metabolism in the rat. Although the mechanism of in vivo bioactivation is unclear, the reaction is not catalyzed by microsomal cytochrome P-450 enzymes.

Alpha-hydroxytamoxifen, a genotoxic metabolite of tamoxifen in the rat: identification and quantification in vivo and in vitro.

The metabolic formation of a-hydroxytamoxifen, a reactive metabolite of tamoxifen in rat liver, was characterized and quantified in vitro (hepatic microsomal incubations) and in vivo (bile-duct cannulated animals). This minor metabolite was identified by chromatographic and mass spectral comparisons with the authentic compound. The rates of formation of alpha-hydroxytamoxifen in incubations (30 min) of tamoxifen (25 microM) with liver microsomal preparations from women (pool of six), female CD1 mice or female Sprague-Dawley rats, as quantified by liquid chromatography-mass spectrometry (LC-MS), were 1.15+/-0.03, 0.30+/-0.05 and 2.70+/-0.35 pmol/min/mg protein, respectively. Selective inhibition of microsomal P450 indicated that alpha-hydroxylation was catalysed predominantly by CYP3A in humans. Bile-duct cannulated and anaesthetized female rats and mice given [14C]tamoxifen (43 micromol/kg, i.v.) excreted, respectively, 24 and 21% of the administered radioactivity in bile over 5 and 3.5 h. The major radiolabelled biliary metabolite in rats, characterized by LC-MS after enzymic hydrolysis of conjugates, was the glucuronide of 4-hydroxytamoxifen (10% of dose) and only 0.1% of the dose was recovered as alpha-hydroxytamoxifen. After administration of alpha-hydroxytamoxifen (43 micromol/kg, i.v.) to rats, only 1.19% of the administered compound was recovered from a glucuronide metabolite in bile, indicating a possible 0.84% alpha-hydroxylation of tamoxifen in vivo. There was, however, no indication of the presence in bile of either O-sulphonate or glutathione conjugates derived from alpha-hydroxytamoxifen. This study shows for the first time that alpha-hydroxytamoxifen can be glucuronylated in rat liver. Whereas sulphonation results in electrophilic genotoxic intermediates, glucuronidation may represent a means of detoxifying alpha-hydroxytamoxifen.

Novel, potent, semisynthetic antimalarial carba analogues of the first-generation 1,2,4-trioxane artemether.

Ten novel, second-generation, fluorinated ether and ester analogues of the potent first-generation analogues artemether (4a) and arteether (4b) have been designed and synthesized. All of the compounds demonstrate high antimalarial potency in vitro against the chloroquine-sensitive HB3 and -resistant K1 strains of Plasmodium falciparum. The most potent derivative 8 was 15 times more potent than artemisinin (2) against the HB3 strain of P. falciparum. In vivo, versus Plasmodium berghei in the mouse, selected derivatives were generally less potent than dihydroartemisinin with ED(50) values of between 5 and 8 mg/kg. On the basis of the products obtained from the in vitro biomimetic Fe(II)-mediated decomposition of 8, the radical mediator of biological activity of this series may be different from that of the parent drug, artemisinin (2).

Metabolism-dependent neutrophil cytotoxicity of amodiaquine: A comparison with pyronaridine and related antimalarial drugs.

Life-threatening agranulocytosis and hepatotoxicity during prophylactic administration of amodiaquine have led to its withdrawal. Agranulocytosis is thought to involve bioactivation to a protein-reactive quinoneimine metabolite. The toxicity of amodiaquine and the lack of cheap drugs have prompted a search for alternative antimalarial agents. The aim of this study was to determine the metabolism and neutrophil toxicity of amodiaquine, pyronaridine, and other related antimalarial agents. Horseradish peroxidase and hydrogen peroxide were used to activate drugs to their respective quinoneimine metabolites. Metabolites were trapped as stable glutathione conjugates, prior to analysis by LC/MS. Amodiaquine was metabolized to a polar metabolite (m/z 661), identified as a glutathione adduct. Tebuquine was converted to two polar metabolites. The principal metabolite (m/z 686) was derived from glutathione conjugation and side chain elimination, while the minor metabolite gave a protonated molecule (m/z 496). Only parent ions were identified when chloroquine, cycloquine, or pyronaridine was incubated with the activating system and glutathione. Calculation of the heat of formation of the drugs, however, demonstrated that amodiaquine, tebuquine, cycloquine, and pyronaridine readily undergo oxidation to their quinoneimine. None of the antimalarial compounds depleted the level of intracellular glutathione (1-300 microM) when incubated with neutrophils alone. Additionally, with the exception of tebuquine, no cytotoxicity below 100 microM was observed. In the presence of the full activating system, however, all compounds except chloroquine resulted in depletion of the level of glutathione and were cytotoxic. Pretreating the cells with glutathione and other antioxidants inhibited metabolism-dependent cytotoxicity. In summary, our data show that amodiaquine and related antimalarials containing a p-aminophenol moiety undergo bioactivation in vitro to chemically reactive and cytotoxic intermediates. In particular, pyronaridine, which is currently being investigated in humans, was metabolized to a compound which was toxic to neutrophils. Thus, the possibility that it will cause agranulocytosis in clinical practice cannot be excluded, and will require careful monitoring.