Relationship between NAD(P)H:quinone oxidoreductase 1 (NQO1) levels in a series of stably transfected cell lines and susceptibility to antitumor quinones.
Relationship between NAD(P)H:quinone oxidoreductase 1 (NQO1) levels in a series of stably transfected cell lines and susceptibility to antitumor quinones.
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DOI：
10.1016/s0006-2952(01)00631-1
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发表时间：
2001-06
影响因子：
5.8
通讯作者：
S. Winski;E. Swann;R. Hargreaves;Donna L. Dehn;John Butler;Christopher J. Moody;David Ross
中科院分区：
文献类型：
--
作者：
S. Winski;E. Swann;R. Hargreaves;Donna L. Dehn;John Butler;Christopher J. Moody;David Ross
为了研究NAD（P）H：醌氧化还原酶1（或DT-黄递酶; NQO 1）在抗肿瘤醌类生物活化中的重要性，我们建立了一系列稳定转染的BE人结肠腺癌细胞系。BE细胞由于遗传多态性而没有NQO 1活性。新的细胞系BE-NQ稳定表达野生型NQO 1。BE-NQ 7细胞表达最高水平的NQO 1，并且对已知的抗肿瘤醌类和较新的临床候选药物更敏感[通过噻唑蓝（MTT）测定]。用不可逆抑制剂ES 936 [5-甲氧基-1，2-二甲基-3-[（4-硝基苯氧基）甲基]吲哚-4，7-二酮]，保护BE-NQ 7细胞免受链黑菌素诱导的毒性，ES 921 [5-（氮丙啶-1-基）-3-（羟甲基）-1，2-二甲基吲哚-4，7-二酮]和RH 1 [2，5-二氮丙啶基-3-（羟甲基）-6-甲基-1，4-苯醌]。通过克隆形成试验进一步评价了RH 1的细胞毒性反应，并且RH 1对BE-NQ 7细胞的细胞毒性大于BE细胞。通过用ES 936预处理抑制NQO 1来消除细胞毒性。使用彗星试验来评估DNA交联，BE-NQ 7细胞表现出比BE细胞对RH 1处理的响应显著更高的DNA交联。在非常低浓度的RH 1（5 nM）下观察到BE-NQ 7细胞中的DNA交联，证实NQO 1将RH 1激活为有效的交联物质。进一步研究使用链黑菌素，ES 921，和RH 1进行分析NQO 1活性和醌毒性之间的关系。在一组表达一定范围NQO 1活性（23-433 nmol/min/mg）的BE-NQ细胞中测量这些化合物的毒性。获得的数据表明NQO 1诱导毒性的阈值高于23 nmol/min/mg，并且在NQO 1的无作用水平（23 nmol/min/mg）和最大作用水平（>77 nmol/min/mg）之间存在尖锐的剂量反应曲线。这些数据提供的证据表明，NQO 1可以在该系统中生物激活抗肿瘤醌类，并表明需要阈值水平的NQO 1活性来启动毒性事件。
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.