Multiple studies indicate that iron chelators enhance their anti-cancer properties by inducing NDRG1, a known tumor and metastasis suppressor. However, the exact role of NDRG1 remains controversial, as newer studies have shown that NDRG1 can also act as an oncogene. Our group recently introduced mitochondrially targeted iron chelators deferoxamine (mitoDFO) and deferasirox (mitoDFX) as effective anti-cancer agents. In this study, we evaluated the ability of these modified chelators to induce NDRG1 and the role of NDRG1 in breast cancer. We demonstrated that both compounds specifically increase NDRG1 without inducing other NDRG family members. We have documented that the effect of mitochondrially targeted chelators is at least partially mediated by GSK3α/β, leading to phosphorylation of NDRG1 at Thr346 and to a lesser extent on Ser330. Loss of NDRG1 increases cell death induced by mitoDFX. Notably, MDA-MB-231 cells lacking NDRG1 exhibit reduced extracellular acidification rate and grow slower than parental cells, while the opposite is true for ER+ MCF7 cells. Moreover, overexpression of full-length NDRG1 and the N-terminally truncated isoform (59112) significantly reduced sensitivity towards mitoDFX in ER+ cells. Furthermore, cells overexpressing full-length NDRG1 exhibited a significantly accelerated tumor formation, while its N-terminally truncated isoforms showed significantly impaired capacity to form tumors. Thus, overexpression of full-length NDRG1 promotes tumor growth in highly aggressive triple-negative breast cancer.

• Keywords
• GSK3α/β, NDRG1, breast cancer, mitoDFO, mitoDFX, mitochondrial iron chelation, oncogene, tumor suppressor,
• Publication type
• Journal Article MeSH

The mitochondrion has emerged as a promising therapeutic target for novel cancer treatments because of its essential role in tumorigenesis and resistance to chemotherapy. Previously, we described a natural compound, 10-((2,5-dihydroxybenzoyl)oxy)decyl) triphenylphosphonium bromide (GA-TPP+C10), with a hydroquinone scaffold that selectively targets the mitochondria of breast cancer (BC) cells by binding to the triphenylphosphonium group as a chemical chaperone; however, the mechanism of action remains unclear. In this work, we showed that GA-TPP+C10 causes time-dependent complex inhibition of the mitochondrial bioenergetics of BC cells, characterized by (1) an initial phase of mitochondrial uptake with an uncoupling effect of oxidative phosphorylation, as previously reported, (2) inhibition of Complex I-dependent respiration, and (3) a late phase of mitochondrial accumulation with inhibition of α-ketoglutarate dehydrogenase complex (αKGDHC) activity. These events led to cell cycle arrest in the G1 phase and cell death at 24 and 48 h of exposure, and the cells were rescued by the addition of the cell-penetrating metabolic intermediates l-aspartic acid β-methyl ester (mAsp) and dimethyl α-ketoglutarate (dm-KG). In addition, this unexpected blocking of mitochondrial function triggered metabolic remodeling toward glycolysis, AMPK activation, increased expression of proliferator-activated receptor gamma coactivator 1-alpha (pgc1α) and electron transport chain (ETC) component-related genes encoded by mitochondrial DNA and downregulation of the uncoupling proteins ucp3 and ucp4, suggesting an AMPK-dependent prosurvival adaptive response in cancer cells. Consistent with this finding, we showed that inhibition of mitochondrial translation with doxycycline, a broad-spectrum antibiotic that inhibits the 28 S subunit of the mitochondrial ribosome, in the presence of GA-TPP+C10 significantly reduces the mt-CO1 and VDAC protein levels and the FCCP-stimulated maximal electron flux and promotes selective and synergistic cytotoxic effects on BC cells at 24 h of treatment. Based on our results, we propose that this combined strategy based on blockage of the adaptive response induced by mitochondrial bioenergetic inhibition may have therapeutic relevance in BC.

• Keywords
• decyl polyhydroxybenzoate triphenylphosphonium derivatives, doxycycline, inhibition of alpha-ketoglutarate dehydrogenase complex, inhibition of the electron transport chain, mitochondrial ribosome inhibition, mitochondrially targeted,
• MeSH
• Apoptosis drug effects   MeSH
• Doxycycline pharmacology   MeSH
• Gentisates chemistry   pharmacology   MeSH
• Heterocyclic Compounds chemistry   pharmacology   MeSH
• Ketoglutarate Dehydrogenase Complex antagonists & inhibitors   genetics   MeSH
• AMP-Activated Protein Kinase Kinases MeSH
• Humans MeSH
• Mitochondria drug effects   pathology   MeSH
• Breast Neoplasms drug therapy   genetics   pathology   MeSH
• Organophosphorus Compounds chemistry   pharmacology   MeSH
• Oxidative Phosphorylation drug effects   MeSH
• Cell Proliferation drug effects   MeSH
• Protein Kinases genetics   MeSH
• Protein Biosynthesis drug effects   MeSH
• Antineoplastic Agents pharmacology   MeSH
• Ribosomes drug effects   MeSH
• Drug Synergism MeSH
• Check Tag
• Humans MeSH
• Female MeSH
• Publication type
• Journal Article MeSH
• Research Support, Non-U.S. Gov't MeSH
• Names of Substances
• 2,5-dihydroxybenzoic acid MeSH Browser
• Doxycycline MeSH
• Gentisates MeSH
• Heterocyclic Compounds MeSH
• Ketoglutarate Dehydrogenase Complex MeSH
• AMP-Activated Protein Kinase Kinases MeSH
• Organophosphorus Compounds MeSH
• Protein Kinases MeSH
• Antineoplastic Agents MeSH
• tris(o-phenylenedioxy)cyclotriphosphazene MeSH Browser

Cellular senescence is a form of cell cycle arrest that limits the proliferative potential of cells, including tumour cells. However, inability of immune cells to subsequently eliminate senescent cells from the organism may lead to tissue damage, inflammation, enhanced carcinogenesis and development of age-related diseases. We found that the anticancer agent mitochondria-targeted tamoxifen (MitoTam), unlike conventional anticancer agents, kills cancer cells without inducing senescence in vitro and in vivo. Surprisingly, it also selectively eliminates both malignant and non-cancerous senescent cells. In naturally aged mice treated with MitoTam for 4 weeks, we observed a significant decrease of senescence markers in all tested organs compared to non-treated animals. Mechanistically, we found that the susceptibility of senescent cells to MitoTam is linked to a very low expression level of adenine nucleotide translocase-2 (ANT2), inherent to the senescent phenotype. Restoration of ANT2 in senescent cells resulted in resistance to MitoTam, while its downregulation in non-senescent cells promoted their MitoTam-triggered elimination. Our study documents a novel, translationally intriguing role for an anticancer agent targeting mitochondria, that may result in a new strategy for the treatment of age-related diseases and senescence-associated pathologies.

• MeSH
• Apoptosis drug effects   genetics   MeSH
• Gene Knockdown Techniques MeSH
• Antineoplastic Agents, Hormonal pharmacology   MeSH
• Humans MeSH
• MCF-7 Cells MeSH
• Mitochondria drug effects   metabolism   MeSH
• Mice, Inbred NOD MeSH
• Mice, SCID MeSH
• Mice, Transgenic MeSH
• Mice MeSH
• Cell Proliferation drug effects   MeSH
• Cellular Senescence drug effects   MeSH
• Tamoxifen pharmacology   MeSH
• Transfection MeSH
• Adenine Nucleotide Translocator 2 genetics   metabolism   MeSH
• Cell Survival drug effects   genetics   MeSH
• Xenograft Model Antitumor Assays MeSH
• Animals MeSH
• Check Tag
• Humans MeSH
• Mice MeSH
• Animals MeSH
• Publication type
• Journal Article MeSH
• Research Support, Non-U.S. Gov't MeSH
• Names of Substances
• Antineoplastic Agents, Hormonal MeSH
• Tamoxifen MeSH
• Adenine Nucleotide Translocator 2 MeSH

AIMS: Expression of the HER2 oncogene in breast cancer is associated with resistance to treatment, and Her2 may regulate bioenergetics. Therefore, we investigated whether disruption of the electron transport chain (ETC) is a viable strategy to eliminate Her2high disease. RESULTS: We demonstrate that Her2high cells and tumors have increased assembly of respiratory supercomplexes (SCs) and increased complex I-driven respiration in vitro and in vivo. They are also highly sensitive to MitoTam, a novel mitochondrial-targeted derivative of tamoxifen. Unlike tamoxifen, MitoTam efficiently suppresses experimental Her2high tumors without systemic toxicity. Mechanistically, MitoTam inhibits complex I-driven respiration and disrupts respiratory SCs in Her2high background in vitro and in vivo, leading to elevated reactive oxygen species production and cell death. Intriguingly, higher sensitivity of Her2high cells to MitoTam is dependent on the mitochondrial fraction of Her2. INNOVATION: Oncogenes such as HER2 can restructure ETC, creating a previously unrecognized therapeutic vulnerability exploitable by SC-disrupting agents such as MitoTam. CONCLUSION: We propose that the ETC is a suitable therapeutic target in Her2high disease. Antioxid. Redox Signal. 26, 84-103.

• Keywords
• HER2, breast cancer, mitochondria, mitochondrially targeted tamoxifen, respirasome,
• MeSH
• Biomarkers MeSH
• Cell Death drug effects   MeSH
• Cell Respiration drug effects   MeSH
• Molecular Targeted Therapy MeSH
• Electron Transport Chain Complex Proteins antagonists & inhibitors   chemistry   metabolism   MeSH
• Inhibitory Concentration 50 MeSH
• Humans MeSH
• Membrane Potential, Mitochondrial drug effects   MeSH
• Mitochondria drug effects   metabolism   MeSH
• Molecular Conformation MeSH
• Models, Molecular MeSH
• Cell Line, Tumor MeSH
• Breast Neoplasms drug therapy   metabolism   pathology   MeSH
• Antineoplastic Agents chemistry   pharmacology   MeSH
• Reactive Oxygen Species metabolism   MeSH
• Receptor, ErbB-2 antagonists & inhibitors   metabolism   MeSH
• Electron Transport Complex I antagonists & inhibitors   chemistry   metabolism   MeSH
• Tamoxifen pharmacology   MeSH
• Protein Binding MeSH
• Check Tag
• Humans MeSH
• Female MeSH
• Publication type
• Journal Article MeSH
• Names of Substances
• Biomarkers MeSH
• Electron Transport Chain Complex Proteins MeSH
• Antineoplastic Agents MeSH
• Reactive Oxygen Species MeSH
• Receptor, ErbB-2 MeSH
• Electron Transport Complex I MeSH
• Tamoxifen MeSH

Respiratory complex II (CII, succinate dehydrogenase, SDH) inhibition can induce cell death, but the mechanistic details need clarification. To elucidate the role of reactive oxygen species (ROS) formation upon the ubiquinone-binding (Qp) site blockade, we substituted CII subunit C (SDHC) residues lining the Qp site by site-directed mutagenesis. Cell lines carrying these mutations were characterized on the bases of CII activity and exposed to Qp site inhibitors MitoVES, thenoyltrifluoroacetone (TTFA) and Atpenin A5. We found that I56F and S68A SDHC variants, which support succinate-mediated respiration and maintain low intracellular succinate, were less efficiently inhibited by MitoVES than the wild-type (WT) variant. Importantly, associated ROS generation and cell death induction was also impaired, and cell death in the WT cells was malonate and catalase sensitive. In contrast, the S68A variant was much more susceptible to TTFA inhibition than the I56F variant or the WT CII, which was again reflected by enhanced ROS formation and increased malonate- and catalase-sensitive cell death induction. The R72C variant that accumulates intracellular succinate due to compromised CII activity was resistant to MitoVES and TTFA treatment and did not increase ROS, even though TTFA efficiently generated ROS at low succinate in mitochondria isolated from R72C cells. Similarly, the high-affinity Qp site inhibitor Atpenin A5 rapidly increased intracellular succinate in WT cells but did not induce ROS or cell death, unlike MitoVES and TTFA that upregulated succinate only moderately. These results demonstrate that cell death initiation upon CII inhibition depends on ROS and that the extent of cell death correlates with the potency of inhibition at the Qp site unless intracellular succinate is high. In addition, this validates the Qp site of CII as a target for cell death induction with relevance to cancer therapy.

• MeSH
• Cell Death physiology   MeSH
• Protein Conformation MeSH
• Humans MeSH
• Mitochondria metabolism   physiology   MeSH
• Molecular Sequence Data MeSH
• Mutagenesis, Site-Directed MeSH
• Electron Transport Complex II chemistry   genetics   metabolism   physiology   MeSH
• Amino Acid Sequence MeSH
• Ubiquinone chemistry   genetics   metabolism   MeSH
• Binding Sites MeSH
• Check Tag
• Humans MeSH
• Publication type
• Journal Article MeSH
• Research Support, Non-U.S. Gov't MeSH
• Names of Substances
• Electron Transport Complex II MeSH
• respiratory complex II MeSH Browser
• Ubiquinone MeSH