This Article Full Text (PDF) Alert me when this article is cited Alert me if a correction is posted Services Similar articles in this journal Similar articles in PubMed Alert me to new issues of the journal Download to citation manager Citing Articles Citing Articles via HighWire Citing Articles via Google Scholar Google Scholar Articles by Siegel, D. Articles by Ross, D. Search for Related Content PubMed PubMed Citation Articles by Siegel, D. Articles by Ross, D.

Metabolism of Mitomycin C by DT-Diaphorase: Role in Mitomycin C-induced DNA Damage and Cytotoxicity in Human Colon Carcinoma Cells¹

Molecular Toxicology and Environmental Health Sciences Program, School of Pharmacy, University of Colorado, Boulder, Colorado 80309 [D. S., D. R.]; Division of Pharmaceutics, School of Pharmacy and Comprehensive Cancer Center, University of Southern California, Los Angeles, California 90033 [N. W. G.]; and Department of Chemistry, University of Akron, Akron, Ohio 44325 [P. C. P.]

The role of DT-diaphorase in bioreductive activation of mitomycin C was examined using HT-29 and BE human carcinoma cells which have high and low levels of DT-diaphorase activity, respectively. HT-29 cells were more sensitive to mitomycin C-induced cytotoxicity than the DT-diaphorase-deficient BE cell line. Mitomycin C induced DNA interstrand cross-linking in HT-29 cells but not in BE cells. Both mitomycin C-induced cytotoxicity and induction of DNA interstrand cross-links could be inhibited by pretreatment of HT-29 cells with dicoumarol. Metabolism of mitomycin C by HT-29 cell cytosol was pH dependent and increased as the pH was lowered to 5.8, the lowest pH tested. Metabolism of mitomycin C by HT-29 cytosol was inhibited by prior boiling of cytosol or by the inclusion of dicoumarol. Little metabolism was detected in BE cytosols. When purified rat hepatic DT-diaphorase was used, metabolism of mitomycin C increased as the pH was decreased and could be detected at pH 5.8, 6.4, 7.0, 7.4, but not at 7.8. Metabolism of mitomycin C was NADH dependent and inhibited by dicoumarol or by prior boiling of enzyme. An approximate 1:1 stoichiometry between NADH and mitomycin C removal was demonstrated and no oxygen consumption could be detected. Metabolism of mitomycin C by purified HT-29 DT-diaphorase was also dicoumarol inhibitable and pH dependent. The major metabolite formed during metabolism of mitomycin C by HT-29 cytosol, purified HT-29, and rat hepatic DT-diaphorase was characterized as 2,7-diaminomitosene. These data suggest that two-electron reduction of mitomycin C by DT-diaphorase may be an important determinant of mitomycin C-induced genotoxicity and cytotoxicity.

¹ Supported by grant RO1CA 51210 from the National Cancer Institute. Parts of this work have been presented in abstract form at the Annual Meeting of the American Association for Cancer Research, Washington, DC, 1990 (Proc. Am. Assoc. Cancer Res., 31: 2367, 1990).

² To whom requests for reprints should be addressed, at School of Pharmacy, Campus Box 297, University of Colorado, Boulder, CO 80309-0297.

Received 6/11/90. Accepted 8/30/90.

This article has been cited by other articles:

				E. Mrozek, J. Kolesar, D. Young, J. Allen, M. Villalona-Calero, and C. L. Shapiro Phase II study of sequentially administered low-dose mitomycin-C (MMC) and irinotecan (CPT-11) in women with metastatic breast cancer (MBC) Ann. Onc., August 1, 2008; 19(8): 1417 - 1422. [Abstract] [Full Text] [PDF]

				W. Guo, P. Reigan, D. Siegel, J. Zirrolli, D. Gustafson, and D. Ross The Bioreduction of a Series of Benzoquinone Ansamycins by NAD(P)H:Quinone Oxidoreductase 1 to More Potent Heat Shock Protein 90 Inhibitors, the Hydroquinone Ansamycins Mol. Pharmacol., October 1, 2006; 70(4): 1194 - 1203. [Abstract] [Full Text] [PDF]

				W. Guo, P. Reigan, D. Siegel, J. Zirrolli, D. Gustafson, and D. Ross Formation of 17-Allylamino-Demethoxygeldanamycin (17-AAG) Hydroquinone by NAD(P)H:Quinone Oxidoreductase 1: Role of 17-AAG Hydroquinone in Heat Shock Protein 90 Inhibition Cancer Res., November 1, 2005; 65(21): 10006 - 10015. [Abstract] [Full Text] [PDF]

				T. H. Ward, S. Danson, A. T. McGown, M. Ranson, N. A. Coe, G. C. Jayson, J. Cummings, R. H.J. Hargreaves, and J. Butler Preclinical Evaluation of the Pharmacodynamic Properties of 2,5-Diaziridinyl-3-Hydroxymethyl-6-Methyl-1,4-Benzoquinone Clin. Cancer Res., April 1, 2005; 11(7): 2695 - 2701. [Abstract] [Full Text] [PDF]

				M. Michael and M.M. Doherty Tumoral Drug Metabolism: Overview and Its Implications for Cancer Therapy J. Clin. Oncol., January 1, 2005; 23(1): 205 - 229. [Abstract] [Full Text] [PDF]

				H. A. Seow, P. G. Penketh, M. F. Belcourt, M. Tomasz, S. Rockwell, and A. C. Sartorelli Nuclear Overexpression of NAD(P)H:Quinone Oxidoreductase 1 in Chinese Hamster Ovary Cells Increases the Cytotoxicity of Mitomycin C under Aerobic and Hypoxic Conditions J. Biol. Chem., July 23, 2004; 279(30): 31606 - 31612. [Abstract] [Full Text] [PDF]

				D. L. Dehn, S. L. Winski, and D. Ross Development of a New Isogenic Cell-Xenograft System for Evaluation of NAD(P)H:Quinone Oxidoreductase-Directed Antitumor Quinones: Evaluation of the Activity of RH1 Clin. Cancer Res., May 1, 2004; 10(9): 3147 - 3155. [Abstract] [Full Text] [PDF]

				Y. Han, H. Shen, B. I. Carr, P. Wipf, J. S. Lazo, and S.-s. Pan NAD(P)H:Quinone Oxidoreductase-1-Dependent and -Independent Cytotoxicity of Potent Quinone Cdc25 Phosphatase Inhibitors J. Pharmacol. Exp. Ther., April 1, 2004; 309(1): 64 - 70. [Abstract] [Full Text]

				D. L. Gustafson, D. Siegel, J. C. Rastatter, A. L. Merz, J. C. Parpal, J. K. Kepa, D. Ross, and M. E. Long Kinetics of NAD(P)H:Quinone Oxidoreductase I (NQO1) Inhibition by Mitomycin C in Vitro and in Vivo J. Pharmacol. Exp. Ther., June 1, 2003; 305(3): 1079 - 1086. [Abstract] [Full Text] [PDF]

				F. Zappa, T. Ward, E. Pedrinis, J. Butler, and A. McGown NAD(P)H:Quinone Oxidoreductase 1 Expression in Kidney Podocytes J. Histochem. Cytochem., March 1, 2003; 51(3): 297 - 302. [Abstract] [Full Text] [PDF]

				K. M. Holtz, S. Rockwell, M. Tomasz, and A. C. Sartorelli Nuclear Overexpression of NADH:Cytochrome b5 Reductase Activity Increases the Cytotoxicity of Mitomycin C (MC) and the Total Number of MC-DNA Adducts in Chinese Hamster Ovary Cells J. Biol. Chem., February 7, 2003; 278(7): 5029 - 5034. [Abstract] [Full Text] [PDF]

				R. P. Baumann, W. F. Hodnick, H. A. Seow, M. F. Belcourt, S. Rockwell, D. H. Sherman, and A. C. Sartorelli Reversal of Mitomycin C Resistance by Overexpression of Bioreductive Enzymes in Chinese Hamster Ovary Cells Cancer Res., November 1, 2001; 61(21): 7770 - 7776. [Abstract] [Full Text] [PDF]

				R. M. Phillips, A. M. Burger, P. M. Loadman, C. M. Jarrett, D. J. Swaine, and H.-H. Fiebig Predicting Tumor Responses to Mitomycin C on the Basis of DT-Diaphorase Activity or Drug Metabolism by Tumor Homogenates: Implications for Enzyme-directed Bioreductive Drug Development Cancer Res., November 1, 2000; 60(22): 6384 - 6390. [Abstract] [Full Text]

				S. Y. Sharp, L. R. Kelland, M. R. Valenti, L. A. Brunton, S. Hobbs, and P. Workman Establishment of an Isogenic Human Colon Tumor Model for NQO1 Gene Expression: Application to Investigate the Role of DT-Diaphorase in Bioreductive Drug Activation In Vitro and In Vivo Mol. Pharmacol., November 1, 2000; 58(5): 1146 - 1155. [Abstract] [Full Text]

				W. Lang, J. Mao, T. W. Doyle, and B. Almassian Isolation and Identification of Urinary Metabolites of Porfiromycin in Dogs and Humans Drug Metab. Dispos., August 1, 2000; 28(8): 899 - 904. [Abstract] [Full Text]

				C. Nesti, F. Trippi, R. Scarpato, L. Migliore, and G. Turchi Cytokinesis-block micronucleus assay in primary human liver fibroblasts exposed to griseofulvin and mitomycin C Mutagenesis, March 1, 2000; 15(2): 143 - 147. [Abstract] [Full Text] [PDF]

				J. J. Pink, S. M. Planchon, C. Tagliarino, M. E. Varnes, D. Siegel, and D. A. Boothman NAD(P)H:Quinone Oxidoreductase Activity Is the Principal Determinant of beta -Lapachone Cytotoxicity J. Biol. Chem., February 25, 2000; 275(8): 5416 - 5424. [Abstract] [Full Text] [PDF]

				M. F. Belcourt, P. G. Penketh, W. F. Hodnick, D. A. Johnson, D. H. Sherman, S. Rockwell, and A. C. Sartorelli Mitomycin resistance in mammalian cells expressing the bacterial mitomycin C resistance protein MCRA PNAS, August 31, 1999; 96(18): 10489 - 10494. [Abstract] [Full Text] [PDF]

				S. Chen, K. Wu, D. Zhang, M. Sherman, R. Knox, and C. S. Yang Molecular Characterization of Binding of Substrates and Inhibitors to DT-Diaphorase: Combined Approach Involving Site-Directed Mutagenesis, Inhibitor-Binding Analysis, and Computer Modeling Mol. Pharmacol., August 1, 1999; 56(2): 272 - 278. [Abstract] [Full Text]

				D. Siegel, E. M. Bolton, J. A. Burr, D. C. Liebler, and D. Ross The Reduction of alpha -Tocopherolquinone by Human NAD(P)H:Quinone Oxidoreductase: The Role of alpha -Tocopherolhydroquinone as a Cellular Antioxidant Mol. Pharmacol., August 1, 1997; 52(2): 300 - 305. [Abstract] [Full Text]

				K.-S. Yao, A. Hageboutros, P. Ford, and P. J. O'Dwyer Involvement of Activator Protein-1 and Nuclear Factor-kappa B Transcription Factors in the Control of the DT-Diaphorase Expression Induced by Mitomycin C Treatment Mol. Pharmacol., March 1, 1997; 51(3): 422 - 430. [Abstract] [Full Text] [PDF]

				S. Chen, R. Knox, K. Wu, P. S.-K. Deng, D. Zhou, M. A. Bianchet, and L. M. Amzel Molecular Basis of the Catalytic Differences among DT-diaphorase of Human, Rat, and Mouse J. Biol. Chem., January 17, 1997; 272(3): 1437 - 1439. [Abstract] [Full Text] [PDF]

				M. Faig, M. A. Bianchet, P. Talalay, S. Chen, S. Winski, D. Ross, and L. M. Amzel Structures of recombinant human and mouse NAD(P)H:quinone oxidoreductases: Species comparison and structural changes with substrate binding and release PNAS, March 28, 2000; 97(7): 3177 - 3182. [Abstract] [Full Text] [PDF]