TY - JOUR
T1 - Inhibition of mitochondrial respiration as a source of adaphostin-induced reactive oxygen species and cytotoxicity
AU - Le, Son B.
AU - Hailer, M. Katie
AU - Buhrow, Sarah
AU - Wang, Qi
AU - Flatten, Karen
AU - Pediaditakis, Peter
AU - Bible, Keith C.
AU - Lewis, Lionel D.
AU - Sausville, Edward A.
AU - Pang, Yuan Ping
AU - Ames, Matthew M.
AU - Lemasters, John J.
AU - Holmuhamedov, Ekhson L.
AU - Kaufmann, Scott H.
PY - 2007/3/23
Y1 - 2007/3/23
N2 - 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.
AB - 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.
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U2 - 10.1074/jbc.M611777200
DO - 10.1074/jbc.M611777200
M3 - Article
C2 - 17213201
AN - SCOPUS:34247851007
SN - 0021-9258
VL - 282
SP - 8860
EP - 8872
JO - Journal of Biological Chemistry
JF - Journal of Biological Chemistry
IS - 12
ER -