Abstract
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 cell-free 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.
Footnotes
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This research was supported by the National Institutes of Health National Cancer Institute [Grant R01-CA114441] (to D.R.); by FORCE (Friends of the Oncology and Radiotherapy Centre, Exeter, UK); and by the Association for International Cancer Research (to C.J.M.). C.J.M., D.R., and D.S. are scientific cofounders and stockholders in QGenta Inc., which holds an option to license molecules described in this article.
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ABBREVIATIONS: ES936, 5-methoxy-1,2-dimethyl-3-[(4-nitrophenoxy)methyl]indole-4,7-dione; NQO1, NAD(P)H:quinone oxidoreductase 1; MTT, 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide; Trx, thioredoxin; TrxR, thioredoxin reductase; ACH983, 5-methoxy-1,2-dimethyl-3-[1-oxo-2-(2,4,6-trifluorophenyl)ethyl]indole-4,7-dione; NRH, dihydronicotinamide riboside; NQO2, NRH:quinone oxidoreductase2; DMSO, dimethyl sulfoxide; DTNB, 5,5′-dithiobis(2-nitrobenzoic acid); IQ, indolequinone; BIAM, biotin-conjugated iodoacetamide; z-VAD-fmk, N-benzyloxycarbonyl-Val-Ala-Asp-fluoromethyl ketone.
- Accepted April 13, 2009.
- Received February 25, 2009.
- The American Society for Pharmacology and Experimental Therapeutics
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