Relationship between NAD(P)H:quinone oxidoreductase 1 (NQO1) levels in a series of stably transfected cell lines and susceptibility to antitumor quinones.
Affiliation
Department of Pharmaceutical Sciences, School of Pharmacy and Cancer Center, Campus Box C238, University of Colorado Health Sciences Center, Denver, CO 80262, USA.Issue Date
2001-06-15
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To investigate the importance of NAD(P)H:quinone oxidoreductase 1 (or DT-diaphorase; NQO1) in the bioactivation of antitumor quinones, we established a series of stably transfected cell lines derived from BE human colon adenocarcinoma cells. BE cells have no NQO1 activity due to a genetic polymorphism. The new cell lines, BE-NQ, stably express wild-type NQO1. BE-NQ7 cells expressed the highest level of NQO1 and were more susceptible [determined by the thiazolyl blue (MTT) assay] to known antitumor quinones and newer clinical candidates. Inhibition of NQO1 by pretreatment with an irreversible inhibitor, ES936 [5-methoxy-1,2-dimethyl-3-[(4-nitrophenoxy)methyl]indole-4,7-dione], protected BE-NQ7 cells from toxicity induced by streptonigrin, ES921 [5-(aziridin-1-yl)-3-(hydroxymethyl)-1,2-dimethylindole-4,7-dione], and RH1 [2,5-diaziridinyl-3-(hydroxymethyl)-6-methyl-1,4-benzoquinone]. RH1 was evaluated further by clonogenic assay for cytotoxic response and was more cytotoxic to BE-NQ7 cells than to BE cells. Cytotoxicity was abrogated by inhibition of NQO1 with ES936 pretreatment. Using a comet assay to evaluate DNA cross-linking, BE-NQ7 cells demonstrated significantly higher DNA cross-links than did BE cells in response to RH1 treatment. DNA cross-linking in BE-NQ7 cells was observed at very low concentrations of RH1 (5 nM), confirming that NQO1 activates RH1 to a potent cross-linking species. Further studies using streptonigrin, ES921, and RH1 were undertaken to analyze the relationship between NQO1 activity and quinone toxicity. Toxicity of these compounds was measured in a panel of BE-NQ cells expressing a range of NQO1 activity (23-433 nmol/min/mg). Data obtained suggest a threshold for NQO1-induced toxicity above 23 nmol/min/mg and a sharp dose-response curve between the no effect level of NQO1 (23 nmol/min/mg) and the maximal effect level (>77 nmol/min/mg). These data provide evidence that NQO1 can bioactivate antitumor quinones in this system and suggest that a threshold level of NQO1 activity is required to initiate toxic events.Citation
Relationship between NAD(P)H:quinone oxidoreductase 1 (NQO1) levels in a series of stably transfected cell lines and susceptibility to antitumor quinones. 2001, 61 (12):1509-16 Biochem. Pharmacol.Journal
Biochemical PharmacologyPubMed ID
11377380Type
ArticleLanguage
enISSN
0006-2952Collections
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