Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: EC:1.6.5.2 (NQO1)
 6,196 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

3-Amino-1,2,4-benzotriazine-1,4-dioxide (SR 4233; WIN 59075) is a highly selective hypoxic cell cytotoxin soon to enter phase I clinical trial. The compound is thought to exert its action through a toxic one-electron reduced free radical intermediate. Preliminary data have suggested that SR 4233 may be metabolized by DT-diaphorase [NAD(P)H: (quinone acceptor) oxidoreductase (EC 1.6.99.2)] to both two- and four-electron reduced products and that this route of biotransformation may represent a bioprotection pathway. In this study, a highly purified enzyme preparation was employed in order to investigate further the metabolism of SR 4233 by DT-diaphorase and to examine the mechanism of reduction in more detail. Spectrophotometric analysis showed that SR 4233 underwent reduction by DT-diaphorase with an apparent Km of 1.23 +/- 0.27 mM and Vmax of 8.55 +/- 1.67 nmol/min/microgram protein. This reaction was inhibited completely by dicoumarol (100 microM) and partially by an antiserum raised against the purified enzyme. Characterization of the products of SR 4233 reduction by reverse-phase HPLC confirmed that both two- (SR 4317) and four- (SR 4330) electron reduction products were generated, the latter being the predominant metabolite, particularly in prolonged incubations. Further experiments showed that the four-electron reduction product, but not the two-electron reduction product, was also a substrate for DT-diaphorase with an apparent Km of 1.14 mM and a Vmax of 57.12 nmol/min/micrograms protein. The results presented confirm that SR 4233 is indeed a substrate for DT-diaphorase and that a mixture of two-, four- and six-electron reduced products may be formed. The possible toxicological and pharmacodynamic significance of this metabolism is discussed.
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PMID:Enzymology of the reduction of the potent benzotriazine-di-N-oxide hypoxic cell cytotoxin SR 4233 (WIN 59075) by NAD(P)H: (quinone acceptor) oxidoreductase (EC 1.6.99.2) purified from Walker 256 rat tumour cells. 173 5

The benzotriazine di-N-oxide SR 4233 (tirapazamine, WIN 59075) is currently in phase I clinical trials as the lead compound in a series of novel and highly selective antitumour hypoxic cytotoxins. Reductive bioactivation is thought to proceed via a one-electron reduced, oxidizing nitroxide radical and also forms the inactive single N-oxide SR 4317 via radical disproportionation or a second one-electron reduction. In mouse liver microsomes reductive metabolism is catalysed predominantly by cytochrome P450 (70%) and cytochrome P450 reductase (30%). The aim of the present study was to examine which cytochrome P450 isozymes may be involved. Reduction of SR 4233 to SR 4317 was monitored by HPLC analysis. Metabolism by microsomes from both control and dexamethasone-induced BALB/c male mice was 70% inhibited by carbon monoxide. The cytochrome P450 inhibitor SKF 525A, following aerobic preincubation, also inhibited SR 4233 reduction by 58%. Reduction was induced 2-3-fold by dexamethasone and was not accountable by increases in cytochrome P450 reductase or DT-diaphorase. The induction data and the greater degree of inhibition of SR 4233 reduction by metyrapone compared to alpha-naphthoflavone suggested a possible involvement of Cyp2b, Cyp2c and Cyp3a cytochrome P450 subfamilies. Both Cyp3a (7.4-fold) and Cyp2b (1.8-fold) type enzymes were shown by western immunoblot analysis to be induced by dexamethasone, the latter correlating more closely with increased SR 4233 reductase activity and also with the 2-fold induction of benzphetamine N-demethylase, a Cyp2b-type enzyme. No inhibition of SR 4233 reduction was seen with erythromycin or cyclosporin A which act as substrates/inhibitors for Cyp3a-type enzymes, but inhibition was seen with p-nitrophenol and tolbutamide which are substrates for Cyp2el- and Cyp2c-type enzymes, respectively (11% and 25% inhibition in induced microsomes). SR 4233 itself inhibited benzphetamine N-demethylase, which is catalysed by Cyp2b-type enzymes but not erythromycin N-demethylase which is catalysed by Cyp3a-type isoforms. Immunoinhibition studies with epitope specific monoclonal antibodies were consistent with the major involvement of phenobarbitone- and steroid-inducible products of the Cyp2b and Cyp2c subfamilies. These forms contributed at least 53% and 26%, respectively, of the cytochrome P450-associated SR 4233 reductase activity in the induced microsomes. The findings support our earlier conclusion that cytochrome P450 is the major SR 4233 reductase in mouse liver and provides leads as to the possible involvement of specific isoforms in human tumours and normal tissues.
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PMID:Initial characterization of the major mouse cytochrome P450 enzymes involved in the reductive metabolism of the hypoxic cytotoxin 3-amino-1,2,4-benzotriazine-1,4-di-N-oxide (tirapazamine, SR 4233, WIN 59075). 846 Oct 36

Tirapazamine (TPZ, 3-amino-1,2,4-benzotriazine 1,4-di-N-oxide, SR 4233, WIN 59075) is a bioreductive antitumor agent with a high selective toxicity for hypoxic cells. The selective hypoxic toxicity of TPZ results from the rapid reoxidation of the one-electron reduction product, the TPZ radical, in the presence of molecular oxygen with the concomitant production of superoxide radical. Under hypoxia the TPZ radical kills cells by causing DNA double-strand breaks and chromosome aberrations. However, the mechanism of aerobic cytotoxicity is still a matter of debate. In this study, we investigated the mechanism of aerobic cytotoxicity by adapting human lung adenocarcinoma A549 cells to aerobic TPZ exposure and characterizing the changes associated with drug resistance. The adapted cells were resistant to aerobic TPZ exposures (with dose-modifying factors of up to 9.2), although hypoxic sensitivity was largely unchanged. Relative to the parental A549 cell line, adaptation to continuous aerobic TPZ exposure resulted in increased levels of manganese superoxide dismutase (up to 9.4-fold), moderate increases in glutathione reductase (up to 2.1-fold), and loss of both quinone oxidoreductase (DT-diaphorase) activity and NADPH cytochrome P450 reductase activity. There was essentially no change in the activity of the cytoplasmic form of superoxide dismutase (CuZnSOD), catalase, or glutathione peroxidase. The increased activity of antioxidant enzymes in the resistant cell lines (in particular MnSOD) strongly suggests that reactive oxygen species are, in large part, responsible for the toxicity of TPZ under aerobic conditions, and is consistent with aerobic and hypoxic drug cytotoxicity resulting from different mechanisms.
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PMID:Adaptation of human tumor cells to tirapazamine under aerobic conditions: implications of increased antioxidant enzyme activity to mechanism of aerobic cytotoxicity. 927 29