Inhibition of mitochondrial respiration as a source of adaphostin-induced reactive oxygen species and cytotoxicity

Son B. Le, M. Katie Hailer, Sarah Buhrow, Qi Wang, Karen Flatten, Peter Pediaditakis, Keith C. Bible, Lionel D. Lewis, Edward A. Sausville, Yuan Ping Pang, Matthew M. Ames, John J. Lemasters, Ekhson L. Holmuhamedov, Scott H. Kaufmann

Research output: Contribution to journalArticlepeer-review

56 Scopus citations

Abstract

Adaphostin is a dihydroquinone derivative that is undergoing extensive preclinical testing as a potential anticancer drug. Previous studies have suggested that the generation of reactive oxygen species (ROS) plays a critical role in the cytotoxicity of this agent. In this study, we investigated the source of these ROS. Consistent with the known chemical properties of dihydroquinones, adaphostin simultaneously underwent oxidation to the corresponding quinone and generated ROS under aqueous conditions. Interestingly, however, this quinone was not detected in intact cells. Instead, high performance liquid chromatography demonstrated that adaphostin was concentrated by up to 300-fold in cells relative to the extracellular medium and that the highest concentration of adaphostin (3000-fold over extracellular concentrations) was detected in mitochondria. Consistent with a mitochondrial site for adaphostin action, adaphostin-induced ROS production was diminished by > 75% in MOLT-4 rho0 cells, which lack mitochondrial electron transport, relative to parental MOLT-4 cells. In addition, inhibition of oxygen consumption was observed when intact cells were treated with adaphostin. Loading of isolated mitochondria to equivalent adaphostin concentrations caused inhibition of uncoupled oxygen consumption in mitochondria incubated with the complex I substrates pyruvate and malate or the complex II substrate succinate. Further analysis demonstrated that adaphostin had no effect on pyruvate or succinate dehydrogenase activity. Instead, adaphostin inhibited reduced decylubiquinone-induced cytochrome c reduction, identifying complex III as the site of inhibition by this agent. Moreover, adaphostin enhanced the production of ROS by succinate-charged mitochondria. Collectively, these observations demonstrate that mitochondrial respiration rather than direct redox cycling of the hydroquinone moiety is a source of adaphostin-induced ROS and identify complex III as a potential target for antineoplastic agents.

Original languageEnglish (US)
Pages (from-to)8860-8872
Number of pages13
JournalJournal of Biological Chemistry
Volume282
Issue number12
DOIs
StatePublished - Mar 23 2007

ASJC Scopus subject areas

  • Biochemistry
  • Molecular Biology
  • Cell Biology

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