Indolequinone Antitumor Agents:  Correlation between Quinone Structure, Rate of Metabolism by Recombinant Human NAD(P)H:Quinone Oxidoreductase, and in Vitro Cytotoxicity

Beall, Howard D.; Winski, Shannon L.; Swann, E.; Hudnott, Anna R.; Cotterill, Ann S.; O'Sullivan, Noeleen; Green, Stephen J.; Bien, Richard; Siegel, David; Ross, David; Moody, Christopher J.

doi:10.1021/jm980328r

Cited by 83 publications

(112 citation statements)

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“…The indolequinones ES936 [1 (Beall et al, 1998)], 6-methoxy-1,2-dimethyl-3-[(4-nitrophenoxy)methyl]indole-4,7-dione [2 ], 2-hydroxymethyl-5-methoxy-1-methyl-3-[(4-nitrophenoxy)methyl]indole-4,7-dione [3 (Newsome, 2004)], 2-hydroxymethyl-5-methoxy-1-methyl-3-[(2,4,6-trifluorophenoxy)methyl]indole-4,7-dione [4 (A.C., unpublished results)], 2-hydroxymethyl-6-methoxy-1-methyl-3-[(4-nitrophenoxy)methyl]indole-4,7-dione [5 (Colucci, 2007)], 6-methoxy-1-methyl-3-[(4-nitrophenoxy)methyl]indole-4,7-dione [6 (Colucci, 2007)], 6-methoxy-1-methyl-3-[(2,4,6-trifluorophenoxy) methyl]indole-4,7-dione [7 (Colucci, 2007)], 5-methoxy-1-methyl-3-[(4-nitrophenoxy)methyl]indole-4,7-dione [8 (Everett et al, 2001)], 5-methoxy-1-methyl-3-[(2,4,6-trifluorophenoxy)methyl]indole-4,7-dione [9 (A.C., unpublished results)], and 5-methoxy-1,2-dimethyl-3-[1-oxo-2-(2,4,6-trifluorophenyl)ethyl]indole-4,7-dione [ACH983 (A.C., unpublished results)] were synthesized according to methods previously developed. Recombinant human NQO2 was obtained from Sigma (St. Louis, MO).…”

Section: Methodsmentioning

confidence: 99%

Potent Activity of Indolequinones against Human Pancreatic Cancer: Identification of Thioredoxin Reductase as a Potential Target

Yan¹,

Shieh²,

Reigan³

et al. 2009

Mol Pharmacol

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The indolequinone ES936 {5-methoxy-1,2-dimethyl-3-[(4-nitrophenoxy)methyl]indole-4,7-dione} was previously developed in our lab as an antitumor agent against pancreatic cancer. The objective of this study was to identify indolequinones with improved potency against pancreatic cancer and to define their mechanisms of action. Pancreatic cancer cell lines PANC-1, MIA PaCa-2, and BxPC-3 were used in in vitro assays [3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium (MTT) and clonogenic assays]; indolequinones displayed potent cytotoxicity against all three cell lines, and two specific classes of indolequinone were particularly potent agents. These indolequinones induced caspase-dependent apoptosis but no redox cycling or oxidative stress in MIA PaCa-2 and BxPC-3 cells. Selected indolequinones were also screened against the NCI-60 cell line panel and were found to be particularly effective against colon, renal, and melanoma cancer cells. A potential target of these indolequinones was identified as thioredoxin reductase. Indolequinones were found to be potent inhibitors of thioredoxin reductase activity both in pancreatic cancer cells and in cellfree systems. The mechanism of action of the indolequinones was shown to involve metabolic reduction, loss of a leaving group to generate a reactive electrophile resulting in alkylation of the selenocysteine residue in the active site of thioredoxin reductase. In vivo efficacy of the indolequinones was also tested in the MIA PaCa-2 pancreatic tumor xenograft in nude mice, and lead indolequinones demonstrated high efficacy and low toxicity. Inhibition of thioredoxin reductase represents a potential novel target in pancreatic cancer and may provide a biomarker of effect of lead indolequinones in this type of cancer.Pancreatic cancer is the fourth leading cause of cancer death in the United States (Jemal et al., 2008), with a 5-year survival rate of Ͻ5%. Current treatment options of radiation therapy, chemotherapy, and surgery have been ineffective at improving the survival rate (Ghaneh et al., 2007). Development of novel targeted therapeutic approaches is desperately needed.We have reported previously the development of an indolequinone, 5-methoxy-1,2-dimethyl-3-[(4-nitrophenoxy)methyl]-indole-4,7-dione (ES936, 1), that exhibited potent growth inhibition effects against human pancreatic cancer cell lines (Dehn et al., 2006). The antitumor activity of ES936 was originally attributed to its role as a mechanism-based inhibitor of human NQO1 [NAD(P)H:quinone oxidoreductase 1 (DT-diaphorase; EC 1.6.99.2)] (Winski et al., 2001). NQO1 inhibition by dicumarol, a nonspecific inhibitor, has been shown to be cytotoxic in human pancreatic cancer cells (Cullen et al., 2003;Lewis et al., 2004). However, when a series of indolequinone compounds

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Section: Methodsmentioning

confidence: 99%

Potent Activity of Indolequinones against Human Pancreatic Cancer: Identification of Thioredoxin Reductase as a Potential Target

Yan¹,

Shieh²,

Reigan³

et al. 2009

Mol Pharmacol

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“…2-Ethoxycarbonyl-2-methyl-3,4-dihydro-2H-pyrrole-1-oxide (EMPO) was obtained from Alexis Corporation (Lä ufelfingen, Switzerland). 5-Methoxy-1,2-dimethyl-3-[(4-nitrophenoxy)methyl] indole-4,7-dione (ES936) was synthesized as described previously (Beall et al, 1998). Recombinant human NQO1 was purified from Escherichia coli by Cibacron blue affinity chromatography as described previously (Beall et al, 1994).…”

Section: Methodsmentioning

confidence: 99%

NAD(P)H:Quinone Oxidoreductase 1: Role as a Superoxide Scavenger

Siegel

Gustafson²,

Dehn³

et al. 2004

Mol Pharmacol

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Experiments using purified recombinant human NAD(P)H:quinone oxidoreductase 1 (NQO1) revealed that the auto-oxidation of fully reduced protein resulted in a 1:1 stoichiometry of oxygen consumption to NADH oxidation with the production of hydrogen peroxide. The rate of auto-oxidation of fully reduced NQO1 was markedly accelerated in the presence of superoxide (O 2 . ), whereas the addition of superoxide dismutase greatly inhibited the rate of auto-oxidation.

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“…All other chemicals were purchased from SigmaAldrich. ES936 and RH1 were synthesized as previously described (Beall et al, 1998;Winski et al, 1998), with stock solutions made up in dimethyl sulfoxide. Cell culture medium and supplements were obtained from Invitrogen (Carlsbad, CA) unless otherwise specified.…”

Section: Methodsmentioning

confidence: 99%

RH1 Induces Cellular Damage in an NAD(P)H:Quinone Oxidoreductase 1-Dependent Manner: Relationship between DNA Cross-linking, Cell Cycle Perturbations, and Apoptosis

Dehn

Inayat-Hussain

Ross

2005

J Pharmacol Exp Ther

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Structure-based development of NAD(P)H:quinone oxidoreductase (NQO1)-directed antitumor quinones resulted in development of RH1 [2,5-diaziridinyl-3-(hydroxymethyl)-6-methyl-1,4-benzoquinone], a methyl-substituted diaziridinyl quinone. We conducted experiments to evaluate the mechanism of RH1-induced cytotoxicity and the inter-relationship between DNA cross-linking, cell cycle changes, and apoptosis using an isogenic cell line pair developed from the human breast cancer cell line MDA-MB-468 differing only in expression of wtNQO1 (NQ16 cells). Statistically significant DNA cross-linking was detected using a modified comet assay in cells with wtNQO1 within 1 h of dosing, whereas in parental cells, only marginal DNA cross-linking was observed and required a concentration up to 50 times higher. Cross-linking in NQ16 cells could be abrogated with 5-methoxy-1,2-dimethyl-3-[(4-nitrophenoxy)methyl]indole-4,7-dione, a mechanismbased inhibitor of NQO1. RH1 prolonged S phase and caused a G 2 /M block. Cell cycle changes were observed up to 10-fold lower in RH1 concentrations in NQ16 cells relative to parental cells. Apoptosis was similarly observed morphologically in both cell lines after RH1 treatment but was induced preferentially in NQ16 cells at lower concentrations and earlier time points. Marked cleavage of caspase-3 was observed in NQ16 cells relative to parental cells using lower concentrations of RH1. Temporally, low doses of RH1-induced rapid DNA cross-linking in NQ16 cells followed by induction of apoptosis at times when a G 2 /M block was not observed. This suggests that cell cycle arrest is not required for RH1-induced apoptosis and that DNA damage may directly initiate apoptotic events. In summary, RH1-induced preferential DNA cross-linking, cell cycle changes, and apoptosis in an NQO1-dependent manner.The bioactivation of antitumor quinones by the two-electron reductase NQO1 has attracted considerable interest after it was found that NQO1 was expressed at high levels throughout many human solid tumors (Cresteil and Jaiswal, 1991;Malkinson et al., 1992;Mikami et al., 1998). The structure-based development of NQO1-directed antitumor quinones led to the identification and testing of a number of compounds, including the aziridinylbenzoquinone RH1 [2,5-diaziridinyl-3-(hydroxymethyl)-6-methyl-1,4-benzoquinone], which is currently in clinical trials in the UK. Upon twoelectron reduction of the quinone by NQO1, the two aziridinyl functional groups undergo facile protonation, leading to aziridine ring opening. The resulting bifunctional alkylating species can covalently bind to DNA (forming inter-and intrastrand cross-links) and other cellular nucleophiles (Hargreaves et al., 1999). It is presumed that the cytotoxicity of RH1 is related to its ability to cross-link DNA (Ward et al., 2000) after activation by NQO1 because it does not appreciably redox cycle , nor does one-electron reduction appear to contribute significantly to the cytotoxicity of RH1. Kim et al. (2004b) did not find a difference in RH1 toxicit...

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Indolequinone Antitumor Agents: Correlation between Quinone Structure, Rate of Metabolism by Recombinant Human NAD(P)H:Quinone Oxidoreductase, and in Vitro Cytotoxicity

Cited by 83 publications

References 28 publications

Potent Activity of Indolequinones against Human Pancreatic Cancer: Identification of Thioredoxin Reductase as a Potential Target

Potent Activity of Indolequinones against Human Pancreatic Cancer: Identification of Thioredoxin Reductase as a Potential Target

NAD(P)H:Quinone Oxidoreductase 1: Role as a Superoxide Scavenger

RH1 Induces Cellular Damage in an NAD(P)H:Quinone Oxidoreductase 1-Dependent Manner: Relationship between DNA Cross-linking, Cell Cycle Perturbations, and Apoptosis

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