Cytotoxicity and DNA lesions produced by mitomycin C and porfiromycin in hypoxic and aerobic EMT6 and Chinese hamster ovary cells (original) (raw)
Related papers
Mechanisms of hypoxic and aerobic cytotoxicity of mitomycin C in Chinese hamster V79 cells
Cancer research, 1991
Mitomycin C (MMC) induced aerobic and hypoxic cytotoxicity in Chinese hamster V79 cells was studied to evaluate the role of the 1-electron versus 2-electron reductive bioactivation. Superoxide dismutase, catalase, and desferal had no protective effects on the aerobic or hypoxic cytotoxicity of MMC, whereas Tempol and Tempol-H, which are known to interrupt and terminate radical reactions, provided partial protection under aerobic conditions. However, under hypoxic conditions, Tempol provided complete protection whereas Tempol-H was ineffective. Electron paramagnetic resonance and spin-trapping investigations, designed to study the mechanisms of such protective effects, confirmed that MMC is activated by the human NADPH:cytochrome P-450 oxidoreductase to its semiquinone radical and that, under aerobic conditions, the semiquinone radical reduces molecular oxygen. Under hypoxic conditions, the semiquinone of MMC reduces H2O2 to produce OH radicals as detected by electron paramagnetic re...
Journal of Biological Chemistry, 2004
The effects of the subcellular localization of overexpressed bioreductive enzyme NAD(P)H:quinone oxidoreductase 1 (NQO1) on the activity of the antineoplastic agent mitomycin C (MC) under aerobic and hypoxic conditions were examined. Chinese hamster ovary (CHO-K1/dhfr(-)) cells were transfected with NQO1 cDNA to produce cells that overexpressed NQO1 activity in the nucleus (148-fold) or the cytosol (163-fold) over the constitutive level of the enzyme in parental cells. Subcellular localization of the enzyme was confirmed using antibody-assisted immunofluorescence. Nuclear localization of transfected NQO1 activity increased the cytotoxicity of MC over that produced by overexpression in the cytosol under both aerobic and hypoxic conditions, with greater cytotoxicity being produced under hypoxia. The greater cytotoxicity of nuclear localized NQO1 was not attributable to greater metabolic activation of MC but instead was the result of activation of the drug in close proximity to its target, nuclear DNA. A positive relationship existed between the degree of MC-induced cytotoxicity and the number of MC-DNA adducts produced. The findings indicate that activation of MC proximal to nuclear DNA by the nuclear localization of transfected NQO1 increases the cytotoxic effects of MC regardless of the degree of oxygenation and support the concept that the mechanism of action of MC involves alkylation of DNA.
Cancer research, 1981
The cytotoxicities of a number of antineoplastic agents to oxygenated and hypoxic EMT6 mouse mammary tumor cells in culture were examined. Based on the relative sensitivities of cells under aerobic and hypoxic conditions, drugs were placed into three categories. Drugs that were preferentially toxic to cells under oxygenated conditions were classified as type 1 agents; this group includes bleomycin, procarbazine, streptonigrin, actinomycin D, and vincristine. Type 2 agents were those preferentially toxic to cells under hypoxic conditions. These include mitomycin C and Adriamycin. On the basis of other published reports, the glucose analogs, 5-thio-D-glucose and 2-deoxy-D-glucose, and the radiosensitizers, misonidazole and metronidazole, can also be placed in this category. Several antineoplastic agents showed no major preferential toxicity to cells under the conditions of oxygenation or hypoxia used in these experiments and were placed in a third class. This group (type 3) includes 1...
Generation of Reactive Oxygen Radicals through Bioactivation of Mitomycin Antibiotics
Cancer Research, 1986
Mitomycin C (MC) is a naturally occurring anticancer agent which has been shownto be more cytotoxicto hypoxictumor cells than to their aerobic counterparts. The mechanism of action of this agent is thought to involvebiologicalreductiveactivation,to a speciesthat alkylutes DNA. A comparison of the cytotoxicityof MC to EMT6 tumor cells with that of the structural analogues porfiromycin(PM), .•V-(,V',Ai"-diiiielhylaniinomethylene)amineanalogueof mitomycinC (BMY-25282),and ;\'-(.!\", .V'-ilimeth>laniinoinctliylciie (aminoanalogue of porfiromycin(BL-6783) has demonstrated that PM is considerably less cytotoxic to aerobic EMT6 cells than MC, whereas BMY-25282 and BL-6783 are signifi cantly more toxic. The relative abilities of each of these compounds to generate oxygen free radicals followingbiologicalactivation were mea sured. Tumor cell sonicates, reduced nicotinamideadenine dinucleotide phosphate-cytochromec reducÃ-ase, xanthine oxidase, and mitochondria were used as the biologicalreducing systems. All four mitomycinanti bioticsproducedoxygenradicals followingbiologicalreduction,a process that may account for the aerobic cytotoxicityof agents of this class. The generation of relame amounts of Superoxideand hydroxyl radical were also measured in EMT6 cell sonicates. BMY-2S282and BL-6783 pro duced significantly greater quantities of oxygen free radicals with the EMT6 cell sonicate, reduced nicotinamide adenine dinucleotide phos phate-cytochromec reducÃ-ase, and mitochondriathan did MC and PM. In contrast, BMY-25282and BL-6783 did not generate detectable levels of free radicals in the presence of xanthine oxidase, whereas this enzyme was capable of generating free radicals with MC and PM as substrates. MC consistentlyproducedgreater amounts of free radicals than PM with all of the reducingsystems. BMY-25282,BL-6783,and MC all generated hydroxyl radicals, while PM did not appear to form these radicals. The findings indicate that a correlation exists between the ability of the mitomycinantibiotics to generate oxygen radicals and their cytotoxicity to aerobic EMT6 tumor cells.
Bioorganic & Medicinal Chemistry, 2010
The heterocyclic N-oxide, 3-amino-1,2,4-benzotriazine 1,4-dioxide (tirapazamine, 1), shows promising antitumor activity in preclinical studies, but there is a continuing need to explore new compounds in this general structural category. In the work described here, we examined the properties of 7-chloro-2-thienylcarbonyl-3-trifluoromethylquinoxaline 1,4-dioxide (9h). We find that 9h causes redox-activated, hypoxia-selective DNA cleavage that mirrors the lead compound, tirapazamine, in both mechanism and potency. Furthermore, we find that 9h displays hypoxiaselective cytotoxicity against human cancer cell lines. antitumor agents that is exemplified by the lead compound 3-amino-1,2,4-benzotriazine 1,4dioxide (tirapazamine, 1, Scheme 1). 4,8-11 Tirapazamine's promising medicinal properties stem from its ability to cause lethal DNA damage in hypoxic tumor cells. 12-18 Tirapazamine-induced DNA damage is initiated by intracellular enzymatic one-electron reduction of the drug. 9,12,13,19-23 In normal aerobic cells, the resulting radical 2 undergoes back-oxidation to regenerate the parent drug and superoxide radical (Scheme 1). 8,9,12,13,19-22 This type of redox cycling can be cytotoxic; 6 however, in the context of tirapazamine, reactions of the drug radical 2 in the absence of molecular oxygen evidently are more lethal. 9,12,13,22 Under hypoxic conditions, the drug radical decomposes to release either the well known DNA-damaging cytotoxin, hydroxyl radical 21,24-27 or a highly reactive benzotriazinyl radical 4. 28-31 The antitumor activity of tirapazamine has been examined in
Role of NADPH:cytochrome c reductase and DT-diaphorase in the biotransformation of mitomycin C1
Cancer research, 1984
Hypoxic cells of solid tumors are difficult to eradicate by X-irradiation or chemotherapy; as an approach to this problem, our laboratories are investigating the effects of the bioreductive alkylating agent mitomycin C (MC) on hypoxic cells. This antibiotic was preferentially toxic to EMT6 mouse mammary tumor cells and V79 Chinese hamster lung fibroblasts under hypoxic conditions, but it was equitoxic to Chinese hamster ovary cells in the presence and absence of oxygen. All cell lines catalyzed the formation of reactive metabolites under hypoxic conditions and contained NADPH:cytochrome c reductase and DT-diaphorase, two enzymes which may be responsible for the cellular activation of MC. Although a correlation existed between enzymatic activities and the formation of reactive metabolites from MC, there was no correspondence between these parameters and the degree of cytotoxicity expressed by MC under hypoxic conditions. Purified NADPH:cytochrome c reductase reduced MC in the absence...
Cancer Research
The clinical utility of antineoplastic agents is limited by the develop- ment of drug resistance by tumors. Mitomycin C (MC) is a bacterial product that must be enzymatically reduced to exert anticancer activity. We have demonstrated that expression of the bacterial MC resistance- associated (MCRA) protein in Chinese hamster ovary (CHO) cells confers profound resistance to this antibiotic under aerobic conditions, but not under hypoxia. MCRA produces resistance to MC by redox cycling of the activated hydroquinone intermediate back to the prodrug form. A CHO cell line developed by stepwise exposure to increasing concentrations of MC likewise expressed high level resistance to MC in air, but not under hypoxia. The overexpression of DT-diaphorase and NADPH:cytochrome c (P-450) reductase, two enzymes known to activate MC, restored sensitivity to MC in both MCRA-transfected and drug-selected cell lines. The level of sensitization was proportional to the quantity of enzyme activity ex- pr...