Tumor cell death induced by the inhibition of mitochondrial electron transport: the effect of 3-hydroxybakuchiol

Abstract

Changes in mitochondrial ATP synthesis can affect the function of tumor cells due to the dependence of the first step of glycolysis on mitochondrial ATP. The oxidative phosphorylation (OXPHOS) system is responsible for the synthesis of approximately 90% of the ATP in normal cells and up to 50% in most glycolytic cancers; therefore, inhibition of the electron transport chain (ETC) emerges as an attractive therapeutic target. We studied the effect of a lipophilic isoprenylated catechol, 3-hydroxybakuchiol (3-OHbk), a putative ETC inhibitor isolated from Psoralea glandulosa. 3-OHbk exerted cytotoxic and anti-proliferative effects on the TA3/Ha mouse mammary adenocarcinoma cell line and induced a decrease in the mitochondrial transmembrane potential, the activation of caspase-3, the opening of the mitochondrial permeability transport pore (MPTP) and nuclear DNA fragmentation. Additionally, 3-OHbk inhibited oxygen consumption, an effect that was completely reversed by succinate (an electron donor for Complex II) and duroquinol (electron donor for Complex III), suggesting that 3-OHbk disrupted the electron flow at the level of Complex I. The inhibition of OXPHOS did not increase the level of reactive oxygen species (ROS) but caused a large decrease in the intracellular ATP level. ETC inhibitors have been shown to induce cell death through necrosis and apoptosis by increasing ROS generation. Nevertheless, we demonstrated that 3-OHbk inhibited the ETC and induced apoptosis through an interaction with Complex I. By delivering electrons directly to Complex III with duroquinol, cell death was almost completely abrogated. These results suggest that 3-OHbk has antitumor activity resulting from interactions with the ETC, a system that is already deficient in cancer cells.

Keywords: 3-OHbk; 3-hydroxybakuchiol; ANOVA; Apoptosis; CsA; Cyclosporine A; DCFDA; DEVDase; DMEM; DMSO; DQ; Dulbecco's Modified Eagle Medium; EGTA; ETC; Electron transport chain; FACS; FITC; G+M; HKII; MPTP; Mitochondria; NADH; OXPHOS; PBS; PI; RFU; ROS; SD; SEM; TMRM; TUNEL; VDAC; analysis of variance; caspase-3-like enzyme; dichlorofluorescein diacetate; dimethyl sulfoxide; duroquinol; electron transport chain; ethylene glycol tetraacetic acid; fluorescein isothiocyanate; fluorescence-activated cell sorting; glucose-6-P; glucose-6-phosphate; glutamate plus malate; hexokinase II; mitochondrial permeability transport pore; oxidative phosphorylation; phosphate buffered saline; propidium iodide; reactive oxygen species; reduced nicotinamide adenine dinucleotide; relative fluorescence units; standard deviation; standard error of the mean; terminal deoxyuridine triphosphate nick end labeling; tetramethylrhodamine methyl ester; voltage-dependent anion channel.