Drug Metabolism Group
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Br. J. Cancer (1985), 52, 409-468 Twenty-Sixth Annual General Meeting of the British Association for Cancer Research* (in conjunction with the European Organization for Research and Treatment for Cancer -Pharmacokinetics and Metabolism Group and the Drug Metabolism Group) (Incorporating Symposia on 'Reactive intermediates in drug metabolism and carcinogenicity', 'New directions in the therapy of cancer', 'The early clinical trials of novel antitumour agents'; the 1985 Walter Hubert Lecture: and the West Midlands Oncology Association Guest Lecture) March 24-27, 1985. Held at Aston University, Birmingham, UK. Abstracts of invited paperst Symposium: aromatase, as potential therapeutic agents in the treatment of oestrogen-dependent tumours. Reactive intermediates in drug metabolism and carcinogenicity The role of reactive electrophiles in biological activity The generation and fate of free radicals in intact and toxicity cells S.D. Nelson S. Orrenius Department of Medicinal Chemistry BG-20, Department of Toxicology, Karolinska Institutet, University of Washington, Seattle, WA 98195 USA. S-104 01 Stockholm, Sweden. In the last two decades it has become apparent that There is increasing evidence that organic radicals as toxic manifestations of many chemicals arise from well as oxygen radicals may cause acute cell injury covalent interactions of either the compounds and also be involved in the initiation and themselves or more commonly their metabolites promotion of tumor formation. In the cell, with tissue macromolecules. Identification of the peroxidase-mediated metabolism of various drugs products of covalent interaction has demonstrated and carcinogens, and one-electron reduction of that the reactive forms are generally electrophilic. many quinones, e.g. menadione, can be a source of The nature of several classes of electrophiles will be generation of both types of radicals. Their briefly described in regard to mechanisms of subsequent interaction with reduced glutathione formation and reaction with nucleophiles. Ongoing (GSH) results in the formation of either glutathione studies in my laboratory on the pathogenesis of S-conjugates or glutathione disulfide (GSSG); the toxicities caused by a halogenated alkyl phosphate latter may occur in part by dimerization of and a terpene will illustrate procedures that are glutathionyl (GS ) radicals generated during the used to define mechanisms of reactive metabolite interaction of organic free radicals with GSH. Since formation. Acetaminophen metabolism will be both glutathione S-conjugates and GSSG are discussed to highlight the complex nature of actively excreted by the cell, this may result in GSH interactions of some electrophiles with tissue depletion and, if the production of free radicals nucleophiles. And, finally, our knowledge of continues, their attack on other cellular nucleophilic reactive electrophiles will be applied to describe the groups, including those in various proteins and preparation of suicide substrates of the enzyme, DNA. This may in turn result in either acute or chronic cell damage. Thus, depletion of protein *Enquiries to the BACR Secretariat c/o Institute of thiols appears to be intimately associated with the Biology, 20 Queensberry Place, London SW7 2DZ. development of menadione-induced cytotoxicity in tReprints of these abstracts are not available - Ed. hepatocytes, possibly via a perturbation of their tThis issue pp. 239-302. Ca2+ homeostasis. 410 AGM BACR ABSTRACTS The role of glutathione in the detoxication of enabled us to make some progress towards electrophilic, peroxy and free radical metabolites determining enzyme mechanisms in vivo. These B. Ketterer results throw much new light on the biochemistry of formaldehyde. More importantly they Cancer Research Campaign Molecular Toxicology demonstrate that these techniques are applicable to Research Group, Middlesex Hospital Medical any xenobiotic which can be labelled and handled School, London WIP 7PN, UK. at the millimolar level, and to any organism which The oxidation of xenobiotics may give rise to can be put into an NMR spectrometer. electrophiles, and by-products of oxygen utilization such as phospholipid hydroperoxides both of which are potentially cytotoxic, and may be detoxified by Symposium: glutathione (GSH) which is a nucleophile and a New Directions reducing agent. in the Therapy of GSH reacts poorly with hard electrophiles, but Cancer well with soft electrophiles, and a family of GSH Oncogenes in cancer therapy transferase enzymes has the capacity to catalyse some if not all these reactions. Such catalysis is an K. Sikora important determinant of the fate of hard Ludwig Institute for Cancer Research, MRC Centre, electrophiles which tend to be genotoxic. On the Hills Road, Cambridge CB2 2QH, UK. other hand if good substrates are present at high The central problem in cancer therapy is the poor doses catalysis itself may cause toxicity due to GSH selectivity of current systemic agents against the depletion. The capacity of different tissues to common solid tumours. The demonstration that detoxify electrophiles varies: it depends on their unique segments of DNA; constant in location and GSH content, the qualitative and quantitative conserved in evolution are involved in growth distribution of GSH transferase isoenzymes in each control opens new avenues for basic and clinical tissue, and the nature of the electrophiles research. The functions of the products of these themselves. genes need to be elucidated. Examples of growth Reduction of phospholipid hydroperoxides by control functions include homology to growth GSH requires the concerted action of factors; surface receptors; protein kinases and cell phospholipase A2 and a range of GSH peroxidases cycle control proteins. From DNA sequence data (the latter utilize free fatty acid hydroperoxides as peptides predicted to be exposed within the intact substrates). molecule can be constructed and used to produce In principle GSH might act as a trap for oxidising monoclonal antibodies to oncogene products. Such free radicals and also free radicals resulting from antibodies have now been successfully used to one electron oxidations of xenobiotics. This has yet demonstrate the intracellular localisation of gene to be demonstrated incontrovertibly because the products as well as the cell cycle regulatory role of resultant thiyl free radical is so difficult to detect. the c-myc protein in vitro. By having a battery of antibodies against the different gene products their direct clinical application for diagnosis and Observing xenobiotic metabolism by in vivo NMR prognosis has become a reality. Immunohistology J.K.M. Sanders and flow cytometry permit the geographical and University Chemical Laboratory, Lensfield Road, quantitative analysis of function in normal and Cambridge CB2 JEW, UK. neoplastic tissues. Furthermore by purification and biochemical analysis the molecular basis for their Carbon, deuterium and proton NMR spectroscopy action can be elucidated. It is likely that by the end have been used to monitor the metabolism of of the decade new drugs that inhibit oncoprotein labelled formaldehyde by a variety of bacterial and function will be available for clinical trial. plant cell preparations. Depending on circumstances, the timescale per spectrum may be as short as 2 min. The rate of metabolism has been Clinical correlates of in vitro studies of small cell correlated with growth and experimental lung cancer conditions, and the metabolism has been shown to D.N. Carney be a detoxifying process in competition with lethal chemistry. The structures of several metabolic Mater Hospital and Saint Lukes Hospital, Dublin, products have been determined in these living Eire. cultures, and have been shown to include A panel of 52 cell lines (CL) derived from patients incorporation, oxidation and reduction products. (pts) with small cell lung cancer (SCLC) have been NMR observations using deuterium NMR have analysed for morphology, cloning efficiency (CE), AGM BACR ABSTRACTS 411 radiation sensitivity, and the expression of a panel facilities at Birkbeck were then used to construct a of biomarkers including L-dopa decarboxylase stereochemically-sensible model for the ATP- (DDC), bombesin (BN), neuron specific enolase binding site of v-src. The model may prove useful (NSE), and creatine kinase BB (CK-BB). Many CL for the design of inhibitors as clinical agents. have been analysed for the c-myc and N-myc (Stemnberg & Taylor, 1984, FEBS Lett., 175, 387). oncogenes. Results of these studies show that SCLC CL can be subdivided into 3 groups: (1) Multipotent CL (N= 2) which in vitro undergo The dependence of cancer cells on growth factors as simultaneous differentiation into SCLC, adeno and a target for chemotherapy squamous cell carcinoma providing evidence for a P. Alexander common stem cell for all types of lung cancer: (2) CRC Medical Oncology Unit, University of Classic SCLC CL (N= 35, 70%) grow as tight Southampton, Southampton General Hospital, aggregates of floating cells, have a low CE (1-5%); Southampton S09 4XY, UK. a long doubling time (DT) (72h); express elevated levels of all 4 biomarkers, and are radiosensitive. Evidence is accumulating that for cancer cells as for (3) Variant SCLC CL (N= 15, 30%) grow as loose normal cells proliferation is dependent on a aggregates; have a high CE (10-30%) a short DT sequence of interactions between receptors on the (26 h), are radioresistant, and lack DDC and BN. outside of the cell membrane and polypeptide Amplified