RAS-RAF-MEK-dependent oxidative cell death involving voltage-dependent anion channels. Nature

Department of Biological Sciences, Fairchild Center, 1212 Amsterdam Avenue, MC 2406, New York, New York 10027, USA.
Nature (Impact Factor: 41.46). 07/2007; 447(7146):864-8. DOI: 10.1038/nature05859
Source: PubMed


Therapeutics that discriminate between the genetic makeup of normal cells and tumour cells are valuable for treating and understanding cancer. Small molecules with oncogene-selective lethality may reveal novel functions of oncoproteins and enable the creation of more selective drugs. Here we describe the mechanism of action of the selective anti-tumour agent erastin, involving the RAS-RAF-MEK signalling pathway functioning in cell proliferation, differentiation and survival. Erastin exhibits greater lethality in human tumour cells harbouring mutations in the oncogenes HRAS, KRAS or BRAF. Using affinity purification and mass spectrometry, we discovered that erastin acts through mitochondrial voltage-dependent anion channels (VDACs)--a novel target for anti-cancer drugs. We show that erastin treatment of cells harbouring oncogenic RAS causes the appearance of oxidative species and subsequent death through an oxidative, non-apoptotic mechanism. RNA-interference-mediated knockdown of VDAC2 or VDAC3 caused resistance to erastin, implicating these two VDAC isoforms in the mechanism of action of erastin. Moreover, using purified mitochondria expressing a single VDAC isoform, we found that erastin alters the permeability of the outer mitochondrial membrane. Finally, using a radiolabelled analogue and a filter-binding assay, we show that erastin binds directly to VDAC2. These results demonstrate that ligands to VDAC proteins can induce non-apoptotic cell death selectively in some tumour cells harbouring activating mutations in the RAS-RAF-MEK pathway.

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Available from: Sudhir Sahasrabudhe
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    • "Downregulation of GK in pancreatic beta cells has been shown to decrease interactions between GK and mitochondria, leading to apoptosis [58]. According to our VGC model, a decrease in the OMP due to the GK detachment (decrease in VGC%, Fig. 7C) allowed higher concentrations of calcium in the MIMS (calculations not shown) that should contribute to the reported death induction [54] due to the activation of mitochondrial permeability transition pore [59]. In general, the VGC model of the OMP generation is consistent with the concept of the critical role of glucokinase in pancreatic beta cell metabolism and cell survival [60]. "
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    ABSTRACT: The simplest mechanism of the generation of the mitochondrial outer membrane potential (OMP) by the VDAC (voltage-dependent anion channel)-hexokinase complex (VHC), suggested earlier, and by the VDAC-glucokinase complex (VGC), was computationally analyzed. Even at less than 4 % of VDACs bound to hexokinase, the calculated OMP is high enough to trigger the electrical closure of VDACs beyond the complexes at threshold concentrations of glucose. These results confirmed our previous hypothesis that the Warburg effect is caused by the electrical closure of VDACs, leading to global restriction of the outer membrane permeability coupled to aerobic glycolysis. The model showed that the inhibition of the conductance and/or an increase in the voltage sensitivity of a relatively small fraction of VDACs by factors like tubulin potentiate the electrical closure of the remaining free VDACs. The extrusion of calcium ions from the mitochondrial intermembrane space by the generated OMP, positive inside, might increase cancer cell resistance to death. Within the VGC model, the known effect of induction of ATP release from mitochondria by accumulated glucose-6-phosphate in pancreatic beta cells might result not only of the known effect of GK dissociation from the VDAC-GK complex, but also of a decrease in the free energy of glucokinase reaction, leading to the OMP decrease and VDAC opening. We suggest that the VDAC-mediated electrical control of the mitochondrial outer membrane permeability, dependent on metabolic conditions, is a fundamental physiological mechanism of global regulation of mitochondrial functions and of cell death.
    Preview · Article · Jan 2014 · Biochimica et Biophysica Acta
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    • "One notable exception is that the mitochondrial outer membrane protein, porin 2, was up-regulated in CA1a cells whereas porins 1 and 3 were not. Mitochondrial porins 2 and 3 overexpression has been implicated in Ras-activated cancers as the functional target of the anti-cancer drug, erastin [38], which cuts off the supply of ATP from the mitochondria to hexokinase. "
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    ABSTRACT: One of the most persistent hallmarks of cancer biology is the preference of tumor cells to derive energy through glycolysis as opposed to the more efficient process of oxidative phosphorylation (OXPHOS). However, little is known about the molecular cascades by which oncogenic pathways bring about this metabolic switch. We carried out a quantitative proteomic and metabolic analysis of the MCF10A derived cell line model of breast cancer progression that includes parental cells and derivatives representing three different tumor grades of Ras-driven cancer with a common genetic background. A SILAC (Stable Isotope Labeling by Amino acids in Cell culture) labeling strategy was used to quantify protein expression in conjunction with subcellular fractionation to measure dynamic subcellular localization in the nucleus, cytosol and mitochondria. Protein expression and localization across cell lines were compared to cellular metabolic rates as a measure of oxidative phosphorylation (OXPHOS), glycolysis and cellular ATP. Investigation of the metabolic capacity of the four cell lines revealed that cellular OXPHOS decreased with breast cancer progression independently of mitochondrial copy number or electron transport chain protein expression. Furthermore, glycolytic lactate secretion did not increase in accordance with cancer progression and decreasing OXPHOS capacity. However, the relative expression and subcellular enrichment of enzymes critical to lactate and pyruvate metabolism supported the observed extracellular acidification profiles. This analysis of metabolic dysfunction in cancer progression integrated with global protein expression and subcellular localization is a novel and useful technique for determining organelle-specific roles of proteins in disease.
    Full-text · Article · Sep 2013 · PLoS ONE
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    • "Therefore, a major oncological target is the HK-VDAC complex (e.g., Galluzzi et al., 2008; Simamura et al., 2008; Fulda et al., 2010; Mathupala and Pedersen, 2010). HK2 can be dissociated from mitochondria by peptides interfering with HK-VDAC association, by erastin (Yagoda et al., 2007) and by 3-bromopyruvate (e.g., Cardaci et al., 2012; Ko et al., 2012; Pedersen, 2012; Shoshan, 2012). Antifungal drugs clotrimazole and bifonazole and the plant hormone methyl jasmonate (MJ) are also effective. "
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    ABSTRACT: Several types of channels play a role in the maintenance of ion homeostasis in subcellular organelles including endoplasmatic reticulum, nucleus, lysosome, endosome, and mitochondria. Here we give a brief overview of the contribution of various mitochondrial and other organellar channels to cancer cell proliferation or death. Much attention is focused on channels involved in intracellular calcium signaling and on ion fluxes in the ATP-producing organelle mitochondria. Mitochondrial K(+) channels (Ca(2+)-dependent BKCa and IKCa, ATP-dependent KATP, Kv1.3, two-pore TWIK-related Acid-Sensitive K(+) channel-3 (TASK-3)), Ca(2+) uniporter MCU, Mg(2+)-permeable Mrs2, anion channels (voltage-dependent chloride channel VDAC, intracellular chloride channel CLIC) and the Permeability Transition Pore (MPTP) contribute importantly to the regulation of function in this organelle. Since mitochondria play a central role in apoptosis, modulation of their ion channels by pharmacological means may lead to death of cancer cells. The nuclear potassium channel Kv10.1 and the nuclear chloride channel CLIC4 as well as the endoplasmatic reticulum (ER)-located inositol 1,4,5-trisphosphate (IP3) receptor, the ER-located Ca(2+) depletion sensor STIM1 (stromal interaction molecule 1), a component of the store-operated Ca(2+) channel and the ER-resident TRPM8 are also mentioned. Furthermore, pharmacological tools affecting organellar channels and modulating cancer cell survival are discussed. The channels described in this review are summarized on Figure 1. Overall, the view is emerging that intracellular ion channels may represent a promising target for cancer treatment.
    Full-text · Article · Sep 2013 · Frontiers in Physiology
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