bims-mibica Biomed News
on Mitochondrial bioenergetics in cancer
Issue of 2020‒10‒04
forty-five papers selected by
Kelsey Fisher-Wellman
East Carolina University


  1. Cancer Treat Res Commun. 2020 Sep 17. pii: S2468-2942(20)30046-0. [Epub ahead of print]25 100210
    Cheng G, Hardy M, Zielonka J, Weh K, Zielonka M, Boyle KA, Abu Eid M, McAllister D, Bennett B, Kresty LA, Dwinell MB, Kalyanaraman B.
      INTRODUCTION: Melanoma is an aggressive form of skin cancer for which there are no effective drugs for prolonged treatment. The existing kinase inhibitor antiglycolytic drugs (B-Raf serine/threonine kinase or BRAF inhibitors) are effective for a short time followed by a rapid onset of drug resistance.PRESENTATION OF CASE: Here, we show that a mitochondria-targeted analog of magnolol, Mito-magnolol (Mito-MGN), inhibits oxidative phosphorylation (OXPHOS) and proliferation of melanoma cells more potently than untargeted magnolol. Mito-MGN also inhibited tumor growth in murine melanoma xenografts. Mito-MGN decreased mitochondrial membrane potential and modulated energetic and mitophagy signaling proteins.
    DISCUSSION: Results indicate that Mito-MGN is significantly more potent than the FDA-approved OXPHOS inhibitor in inhibiting proliferation of melanoma cells.
    CONCLUSION: These findings have implications in the treatment of melanomas with enhanced OXPHOS status due to metabolic reprogramming or drug resistance.
    Keywords:  Bioenergetic metabolism; Melanoma; Mitochondria-targeted agents; Mitophagy; Oxidative phosphorylation
    DOI:  https://doi.org/10.1016/j.ctarc.2020.100210
  2. Redox Biol. 2020 Sep 19. pii: S2213-2317(20)30938-1. [Epub ahead of print]37 101733
    Milliken AS, Kulkarni CA, Brookes PS.
      Generation of mitochondrial reactive oxygen species (ROS) is an important process in triggering cellular necrosis and tissue infarction during ischemia-reperfusion (IR) injury. Ischemia results in accumulation of the metabolite succinate. Rapid oxidation of this succinate by mitochondrial complex II (Cx-II) during reperfusion reduces the co-enzyme Q (Co-Q) pool, thereby driving electrons backward into complex-I (Cx-I), a process known as reverse electron transport (RET), which is thought to be a major source of ROS. During ischemia, enhanced glycolysis results in an acidic cellular pH at the onset of reperfusion. While the process of RsET within Cx-I is known to be enhanced by a high mitochondrial trans-membrane ΔpH, the impact of pH itself on the integrated process of Cx-II to Cx-I RET has not been fully studied. Using isolated mouse heart and liver mitochondria under conditions which mimic the onset of reperfusion (i.e., high [ADP]), we show that mitochondrial respiration (state 2 and state 3) as well as isolated Cx-II activity are impaired at acidic pH, whereas the overall generation of ROS by Cx-II to Cx-I RET was insensitive to pH. Together these data indicate that the acceleration of Cx-I RET ROS by ΔpH appears to be cancelled out by the impact of pH on the source of electrons, i.e. Cx-II. Implications for the role of Cx-II to Cx-I RET derived ROS in IR injury are discussed.
    Keywords:  Acidosis; Complex I; Ischemia; Metabolism; Mitochondria; ROS
    DOI:  https://doi.org/10.1016/j.redox.2020.101733
  3. Int J Mol Med. 2020 Nov;46(5): 1695-1706
    Liu Y, Wang J, Qiao J, Liu S, Wang S, Zhao D, Bai X, Liu M.
      20(S)‑Ginsenoside Rh2 [20(S)‑GRh2], one of the main active components of Panax ginseng, induces apoptosis in a wide range of cancer cell types. The present study found that 20(S)‑GRh2 reduces mitochondrial membrane potential, decreases adenosine triphosphate generation and induces reactive oxygen species in HeLa cervical cancer cells. In addition, 20(S)‑GRh2 activated mitochondrion‑dependent apoptosis and inhibited both mitochondrial oxidative phosphorylation and glycolysis in HeLa cells. It was found that voltage‑dependent anion channel 1 (VDAC1) expression was significantly upregulated by 20(S)‑GRh2 treatment, while hexokinase 2 expression was downregulated and segregated from the mitochondria. Furthermore, 20(S)‑GRh2 promoted Bax transport from the cytoplasm to the mitochondria, and knockdown of VDAC1 inhibited Bax transport and apoptosis. These results suggest that VDAC1 is a novel target of 20(S)‑GRh2. The present study provides a better understanding of the mechanistic link between cervical cancer metabolism and growth control, and these results may facilitate the development of new treatments for cervical cancer.
    DOI:  https://doi.org/10.3892/ijmm.2020.4725
  4. Biomolecules. 2020 Sep 30. pii: E1395. [Epub ahead of print]10(10):
    Aminzadeh-Gohari S, Weber DD, Catalano L, Feichtinger RG, Kofler B, Lang R.
      Drastically elevated glycolytic activity is a prominent metabolic feature of cancer cells. Until recently it was thought that tumor cells shift their entire energy production from oxidative phosphorylation (OXPHOS) to glycolysis. However, new evidence indicates that many cancer cells still have functional OXPHOS, despite their increased reliance on glycolysis. Growing pre-clinical and clinical evidence suggests that targeting mitochondrial metabolism has anti-cancer effects. Here, we analyzed mitochondrial respiration and the amount and activity of OXPHOS complexes in four melanoma cell lines and normal human dermal fibroblasts (HDFs) by Seahorse real-time cell metabolic analysis, immunoblotting, and spectrophotometry. We also tested three clinically approved antibiotics, one anti-parasitic drug (pyrvinium pamoate), and a novel anti-cancer agent (ONC212) for effects on mitochondrial respiration and proliferation of melanoma cells and HDFs. We found that three of the four melanoma cell lines have elevated glycolysis as well as OXPHOS, but contain dysfunctional mitochondria. The antibiotics produced different effects on the melanoma cells and HDFs. The anti-parasitic drug strongly inhibited respiration and proliferation of both the melanoma cells and HDFs. ONC212 reduced respiration in melanoma cells and HDFs, and inhibited the proliferation of melanoma cells. Our findings highlight ONC212 as a promising drug for targeting mitochondrial respiration in cancer.
    Keywords:  BRAF; NRAS; ONC212; Warburg effect; anti-parasitic drug; antibiotic; melanoma; mitochondrial respiration
    DOI:  https://doi.org/10.3390/biom10101395
  5. Aging Cell. 2020 Sep 29. e13248
    Oblong JE, Bowman A, Rovito HA, Jarrold BB, Sherrill JD, Black MR, Nelson G, Kimball AB, Birch-Machin MA.
      Alterations in metabolism in skin are accelerated by environmental stressors such as solar radiation, leading to premature aging. The impact of aging on mitochondria is of interest given their critical role for metabolic output and the finding that environmental stressors cause lowered energy output, particularly in fibroblasts where damage accumulates. To better understand these metabolic changes with aging, we performed an in-depth profiling of the expression patterns of dermal genes in face, forearm, and buttock biopsies from females of 20-70 years of age that encode for all subunits comprising complexes I-V of the mitochondrial electron transport chain. This complements previous preliminary analyses of these changes. "Oxidative phosphorylation" was the top canonical pathway associated with aging in the face, and genes encoding for numerous subunits had decreased expression patterns with age. Investigations on fibroblasts from older aged donors also showed decreased gene expression of numerous subunits from complexes I-V, oxidative phosphorylation rates, spare respiratory capacity, and mitochondrial number and membrane potential compared to younger cells. Treatment of older fibroblasts with nicotinamide (Nam) restored these measures to younger cell levels. Nam increased complexes I, IV, and V activity and gene expression of representative subunits. Elevated mt-Keima staining suggests a possible mechanism of action for these restorative effects via mitophagy. Nam also improved mitochondrial number and membrane potential in younger fibroblasts. These findings show there are significant changes in mitochondrial functionality with aging and that Nam treatment can restore bioenergetic efficiency and capacity in older fibroblasts with an amplifying effect in younger cells.
    DOI:  https://doi.org/10.1111/acel.13248
  6. J Clin Med. 2020 Sep 26. pii: E3113. [Epub ahead of print]9(10):
    Fritzen AM, Andersen SP, Qadri KAN, Thøgersen FD, Krag T, Ørngreen MC, Vissing J, Jeppesen TD.
      Mitochondrial dysfunction is thought to be involved in age-related loss of muscle mass and function (sarcopenia). Since the degree of physical activity is vital for skeletal muscle mitochondrial function and content, the aim of this study was to investigate the effect of 6 weeks of aerobic exercise training and 8 weeks of deconditioning on functional parameters of aerobic capacity and markers of muscle mitochondrial function in elderly compared to young individuals. In 11 healthy, elderly (80 ± 4 years old) and 10 healthy, young (24 ± 3 years old) volunteers, aerobic training improved maximal oxygen consumption rate by 13%, maximal workload by 34%, endurance capacity by 2.4-fold and exercise economy by 12% in the elderly to the same extent as in young individuals. This evidence was accompanied by a similar training-induced increase in muscle citrate synthase (CS) (31%) and mitochondrial complex I-IV activities (51-163%) in elderly and young individuals. After 8 weeks of deconditioning, endurance capacity (-20%), and enzyme activity of CS (-18%) and complex I (-40%), III (-25%), and IV (-26%) decreased in the elderly to a larger extent than in young individuals. In conclusion, we found that elderly have a physiological normal ability to improve aerobic capacity and mitochondrial function with aerobic training compared to young individuals, but had a faster decline in endurance performance and muscle mitochondrial enzyme activity after deconditioning, suggesting an age-related issue in maintaining oxidative metabolism.
    Keywords:  aerobic exercise training; deconditioning; elderly; endurance; mitochondria; sarcopenia; skeletal muscle
    DOI:  https://doi.org/10.3390/jcm9103113
  7. J Genet Genomics. 2020 Aug 01. pii: S1673-8527(20)30114-4. [Epub ahead of print]
    Nguyen H, LaFramboise T.
      Mutations in the human mitochondrial genome have been observed in all types of human cancer, indicating that mutations might contribute to tumorigenesis, metastasis, recurrence, or drug response. This possibility is appealing because of the known shift from oxidative metabolism to glycolysis, known as the Warburg effect, that occurs in malignancy. Mitochondrial DNA (mtDNA) mutations could either be maternally inherited and predispose to cancer (germ line mutations) or occur sporadically in the mtDNA of specific tissues (tissue- or tumor-specific somatic mutations) and contribute to the tumor initiation and progression process. High-throughput sequencing technologies now enable comprehensive detection of mtDNA variation in tissues and bodily fluids, with the potential to be used as an early detection tool that may impact the treatment of cancer. Here, we discuss insights into the roles of mtDNA mutations in carcinogenesis, highlighting the complexities involved in the analysis and interpretation of mitochondrial genomic content, technical challenges in studying their contribution to pathogenesis, and the value of mtDNA mutations in developing early detection, diagnosis, prognosis, and therapeutic strategies for cancer.
    Keywords:  Cancer; Genetic variation; Mitochondrial DNA
    DOI:  https://doi.org/10.1016/j.jgg.2020.04.007
  8. Am J Physiol Endocrinol Metab. 2020 Sep 28.
    Miller VJ, LaFountain RA, Barnhart E, Sapper TS, Short J, Arnold WD, Hyde PN, Crabtree CD, Kackley ML, Kraemer WJ, Villamena F, Volek JS.
      Animal data indicate that ketogenic diets are associated with improved mitochondrial function, but human data are lacking. We aimed to characterize skeletal muscle mitochondrial changes in response to a ketogenic diet combined with exercise training in healthy individuals. Twenty-nine physically active adults completed a 12-week supervised exercise program after self-selection into a ketogenic diet (KD, n=15) group or maintenance of their habitual mixed diet (MD, n=14). Measures of metabolic health and muscle biopsies (Vastus lateralis) were obtained before and after the intervention. Mitochondria were isolated from muscle and studied after exposure to carbohydrate (pyruvate), fat (palmitoyl-L-carnitine), and ketone (β-hydroxybutyrate+acetoacetate) substrates. Compared to MD, the KD resulted in increased whole-body resting fat oxidation (p<0.001) and decreased fasting insulin (p=0.019), insulin resistance (HOMA-IR, p=0.022), and visceral fat (p<0.001). The KD altered mitochondrial function as evidenced by increases in mitochondrial respiratory control ratio (19%, p=0.009), ATP production (36%, p=0.028), and ATP/H2O2 (36%, p=0.033) with the fat-based substrate. ATP production with the ketone-based substrate was 4 to 8 times lower than with other substrates, indicating minimal oxidation. The KD resulted in a small decrease in muscle glycogen (14%, p=0.035) and an increase in muscle triglyceride (81%, p=0.006). These results expand our understanding of human adaptation to a ketogenic diet combined with exercise. In conjunction with weight loss, we observed altered skeletal muscle mitochondrial function and efficiency, an effect that may contribute to the therapeutic use of ketogenic diets in various clinical conditions, especially those associated with insulin resistance.
    Keywords:  exercise; fat oxidation; ketogenic diet; mitochondria; skeletal muscle
    DOI:  https://doi.org/10.1152/ajpendo.00305.2020
  9. Cells. 2020 Sep 29. pii: E2202. [Epub ahead of print]9(10):
    Chun KS, Jang JH, Kim DH.
      Signal transducer and activator of transcription 3 (STAT3) functions as a major molecular switch that plays an important role in the communication between cytokines and kinases. In this role, it regulates the transcription of genes involved in various biochemical processes, such as proliferation, migration, and metabolism of cancer cells. STAT3 undergoes diverse post-translational modifications, such as the oxidation of cysteine by oxidative stress, the acetylation of lysine, or the phosphorylation of serine/threonine. In particular, the redox modulation of critical cysteine residues present in the DNA-binding domain of STAT3 inhibits its DNA-binding activity, resulting in the inactivation of STAT3-mediated gene expression. Accumulating evidence supports that STAT3 is a key protein that acts as a mediator of metabolism and mitochondrial activity. In this review, we focus on the post-translational modifications of STAT3 by oxidative stress and how the modification of STAT3 regulates cell metabolism, particularly in the metabolic pathways in cancer cells.
    Keywords:  STAT3; cancer metabolism; mitochondria; oxidative stress; post-translational modification; redox regulation
    DOI:  https://doi.org/10.3390/cells9102202
  10. Elife. 2020 Sep 29. pii: e58108. [Epub ahead of print]9
    Cheng CC, Wooten J, Gibbs ZA, McGlynn K, Mishra P, Whitehurst AW.
      Cancer testis antigens (CTAs) are proteins whose expression is normally restricted to the testis but anomalously activated in human cancer. In sperm, a number of CTAs support energy generation, however whether they contribute to tumor metabolism is not understood. We describe human COX6B2, a component of cytochrome c oxidase (complex IV). COX6B2 is expressed in human lung adenocarcinoma (LUAD) and expression correlates with reduced survival time. COX6B2, but not its somatic isoform COX6B1, enhances activity of complex IV, increasing oxidative phosphorylation (OXPHOS) and NAD+ generation. Consequently, COX6B2-expressing cancer cells display a proliferative advantage, particularly in low oxygen. Conversely, depletion of COX6B2 attenuates OXPHOS and collapses mitochondrial membrane potential leading to cell death or senescence. COX6B2 is both necessary and sufficient for growth of human tumor xenografts in mice. Our findings reveal a previously unappreciated, tumor specific metabolic pathway hijacked from one of the most ATP-intensive processes in the animal kingdom: sperm motility.
    Keywords:  cancer biology; human
    DOI:  https://doi.org/10.7554/eLife.58108
  11. Trends Cancer. 2020 Sep 23. pii: S2405-8033(20)30236-3. [Epub ahead of print]
    Tasdogan A, McFadden DG, Mishra P.
      Mitochondria play an essential role in cellular metabolism, generation of reactive oxygen species (ROS), and the initiation of apoptosis. These properties enable mitochondria to be crucial integrators in the pathways of tumorigenesis. An open question is to what extent variation in the mitochondrial genome (mtDNA) contributes to the biological heterogeneity observed in human tumors. In this review, we summarize our current understanding of the role of mtDNA genetics in relation to human cancers.
    Keywords:  cancer; haplotypes; mitochondria; mtDNA; oncocytoma
    DOI:  https://doi.org/10.1016/j.trecan.2020.08.004
  12. FASEB J. 2020 Sep 30.
    Høgh RI, Møller SH, Jepsen SD, Mellergaard M, Lund A, Pejtersen M, Fitzner E, Andresen L, Skov S.
      SCFAs are primarily produced in the colon by bacterial fermentation of nondigestible carbohydrates. Besides providing energy, SCFAs can suppress development of colon cancer. The mechanism, however, remains elusive. Here, we demonstrate that the SCFA propionate upregulates surface expression of the immune stimulatory NKG2D ligands, MICA/B by imposing metabolic changes in dividing cells. Propionate-mediated MICA/B expression did not rely on GPR41/GPR43 receptors but depended on functional mitochondria. By siRNA-directed knockdown, we could further link phosphoenolpyruvate carboxykinase (PEPCK), the rate-limiting enzyme in gluconeogenesis to propionate regulation of MICA/B expression. Moreover, knockdown of Rictor and specific mTOR inhibitors implicated mTORC2 activity with metabolic changes that control MICA/B expression. SCFAs are precursors to short-chain acyl-CoAs that are used for histone acylation thereby linking the metabolic state to chromatin structure and gene expression. Propionate increased the overall acetylation and propionylation and inhibition of lysine acetyltransferases (KATs) that are responsible for adding acyl-CoAs to histones reduced propionate-mediated MICA/B expression, suggesting that propionate-induced acylation increases MICA/B expression. Notably, propionate upregulated MICA/B surface expression on colon cancer cells in an acylation-dependent manner; however, the impact of mitochondrial metabolism on MICA/B expression was different in colon cancer cells compared with Jurkat cells, suggesting that continuous exposure to propionate in the colon may provide an enhanced capacity to metabolize propionate. Together, our findings support that propionate causes metabolic changes resulting in NKG2D ligand surface expression, which holds potential as an immune activating anticancer therapy.
    Keywords:  NKG2D ligands; cancer; immunometabolism; short-chain fatty acids
    DOI:  https://doi.org/10.1096/fj.202000162R
  13. Clin Transl Med. 2020 Sep;10(5): e172
    Ou T, Yang W, Li W, Lu Y, Dong Z, Zhu H, Sun X, Dong Z, Weng X, Chang S, Li H, Li Y, Qiu Z, Hu K, Sun A, Ge J.
      BACKGROUND: Mesenchymal stem cells (MSCs) have therapeutic potential for multiple ischemic diseases. However, in vitro expansion of MSCs before clinical application leads to metabolic reprogramming from glycolysis to oxidative phosphorylation, drastically impairing their proliferative and therapeutic capacities. This study aimed to define the regulatory effects of Sirtuin 5 (SIRT5) on the proliferative and therapeutic functions of adipose-derived MSCs (ADMSCs) during in vitro expansion.METHODS: ADMSCs were isolated from wild-type (WT) and Sirt5-knockout (Sirt5-/- ) mice. Cell counting assay was used to investigate the proliferative capacities of the ADMSCs. Dihydroethidium and senescence-associated β-galactosidase stainings were used to measure intracellular ROS and senescence levels. Mass spectrometry was used to analyze protein succinylation. Oxygen consumption rates and extra cellular acidification rates were measured as indicators of mitochondrial respiration and glycolysis. Metabolic-related genes expression were verified by quantitative PCR and western blot. Hind limb ischemia mouse model was used to evaluate the therapeutic potentials of WT and Sirt5-/- ADSMCs.
    RESULTS: SIRT5 protein levels were upregulated in ADMCs during in vitro expansion. Sirt5-/- ADMSCs exhibited a higher proliferation rate, delayed senescence, and reduced ROS accumulation. Furthermore, elevated protein succinylation levels were observed in Sirt5-/- ADMSCs, leading to the reduced activity of tricarboxylic acid cycle-related enzymes and attenuated mitochondrial respiration. Glucose uptake, glycolysis, and pentose phosphate pathway were elevated in Sirt5-/- ADMSCs. Inhibition of succinylation by glycine or re-expression of Sirt5 reversed the metabolic alterations in Sirt5-/- ADMSCs, thus abolishing their enhanced proliferative capacities. In the hind limb ischemia mouse model, SIRT5-/- ADMSCs transplantation enhanced blood flow recovery and angiogenesis compared with WT ADMSCs.
    CONCLUSIONS: Our results indicate that SIRT5 deficiency during ADMSC culture expansion leads to reversed metabolic pattern, enhanced proliferative capacities, and improved therapeutic outcomes. These data suggest SIRT5 as a potential target to enhance the functional properties of MSCs for clinical application.
    Keywords:  SIRT5; adipose-derived mesenchymal stem cells; cell proliferation; hind limb ischemia; metabolic switching
    DOI:  https://doi.org/10.1002/ctm2.172
  14. Cancers (Basel). 2020 Sep 30. pii: E2819. [Epub ahead of print]12(10):
    Pascale RM, Calvisi DF, Simile MM, Feo CF, Feo F.
      The deregulation of the oxidative metabolism in cancer, as shown by the increased aerobic glycolysis and impaired oxidative phosphorylation (Warburg effect), is coordinated by genetic changes leading to the activation of oncogenes and the loss of oncosuppressor genes. The understanding of the metabolic deregulation of cancer cells is necessary to prevent and cure cancer. In this review, we illustrate and comment the principal metabolic and molecular variations of cancer cells, involved in their anomalous behavior, that include modifications of oxidative metabolism, the activation of oncogenes that promote glycolysis and a decrease of oxygen consumption in cancer cells, the genetic susceptibility to cancer, the molecular correlations involved in the metabolic deregulation in cancer, the defective cancer mitochondria, the relationships between the Warburg effect and tumor therapy, and recent studies that reevaluate the Warburg effect. Taken together, these observations indicate that the Warburg effect is an epiphenomenon of the transformation process essential for the development of malignancy.
    Keywords:  Warburg effect; glycolysis; oncogenes; oxidative metabolism; tumor therapy
    DOI:  https://doi.org/10.3390/cancers12102819
  15. BMC Cancer. 2020 Sep 29. 20(1): 929
    Fujita M, Imadome K, Somasundaram V, Kawanishi M, Karasawa K, Wink DA.
      BACKGROUND: Metabolic reprogramming is being recognized as a fundamental hallmark of cancer, and efforts to identify drugs that can target cancer metabolism are underway. In this study, we used human breast cancer (BC) cell lines and established their invading phenotype (INV) collected from transwell inserts to compare metabolome differences and evaluate prognostic significance of the metabolome in aggressive BC invasiveness.METHODS: The invasiveness of seven human BC cell lines were compared using the transwell invasion assay. Among these, INV was collected from SUM149, which exhibited the highest invasiveness. Levels of metabolites in INV were compared with those of whole cultured SUM149 cells (WCC) using CE-TOFMS. The impact of glycolysis in INV was determined by glucose uptake assay using fluorescent derivative of glucose (2-NBDG), and significance of glycolysis, or tricarboxylic acid cycle (TCA) and electron transport chain (ETC) in the invasive process were further determined in aggressive BC cell lines, SUM149, MDA-MB-231, HCC1937, using invasion assays in the presence or absence of inhibitors of glycolysis, TCA cycle or ETC.
    RESULTS: SUM149 INV sub-population exhibited a persistent hyperinvasive phenotype. INV were hyper-glycolytic with increased glucose (2-NBDG) uptake; diminished glucose-6-phosphate (G6P) levels but elevated pyruvate and lactate, along with higher expression of phosphorylated-pyruvate dehydrogenase (pPDH) compared to WCC. Notably, inhibiting of glycolysis with lower doses of 2-DG (1 mM), non-cytotoxic to MDA-MB-231 and HCC1937, was effective in diminishing invasiveness of aggressive BC cell lines. In contrast, 3-Nitropropionic acid (3-NA), an inhibitor of succinate dehydrogenase, the enzyme that oxidizes succinate to fumarate in TCA cycle, and functions as complex II of ETC, had no significant effect on their invasiveness, although levels of TCA metabolites or detection of mitochondrial membrane potential with JC-1 staining, indicated that INV cells originally had functional TCA cycles and membrane potential.
    CONCLUSIONS: Hyper-glycolytic phenotype of invading cells caters to rapid energy production required for invasion while TCA cycle/ETC cater to cellular energy needs for sustenance in aggressive BC. Lower, non-cytotoxic doses of 2-DG can hamper invasion and can potentially be used as an adjuvant with other anti-cancer therapies without the usual side-effects associated with cytotoxic doses.
    Keywords:  2-DG; Breast cancer; ETC; Glycolysis; Invasion; Metabolism; TCA cycle
    DOI:  https://doi.org/10.1186/s12885-020-07414-y
  16. Cells. 2020 Sep 29. pii: E2197. [Epub ahead of print]9(10):
    Saldana-Caboverde A, Nissanka N, Garcia S, Lombès A, Diaz F.
      Murine fibroblasts deficient in mitochondria respiratory complexes III (CIII) and IV (CIV) produced by either the ablation of Uqcrfs1 (encoding for Rieske iron sulfur protein, RISP) or Cox10 (encoding for protoheme IX farnesyltransferase, COX10) genes, respectively, showed a pleiotropic effect in complex I (CI). Exposure to 1-5% oxygen increased the levels of CI in both RISP and COX10 KO fibroblasts. De novo assembly of the respiratory complexes occurred at a faster rate and to higher levels in 1% oxygen compared to normoxia in both RISP and COX10 KO fibroblasts. Hypoxia did not affect the levels of assembly of CIII in the COX10 KO fibroblasts nor abrogated the genetic defect impairing CIV assembly. Mitochondrial signaling involving reactive oxygen species (ROS) has been implicated as necessary for HIF-1α stabilization in hypoxia. We did not observe increased ROS production in hypoxia. Exposure to low oxygen levels stabilized HIF-1α and increased CI levels in RISP and COX10 KO fibroblasts. Knockdown of HIF-1α during hypoxic conditions abrogated the beneficial effect of hypoxia on the stability/assembly of CI. These findings demonstrate that oxygen and HIF-1α regulate the assembly of respiratory complexes.
    Keywords:  COX10; HIF-1α; Rieske iron sulfur protein; complex I; complex III; complex IV; hypoxia; mitochondrial respiratory supercomplexes; oxidative phosphorylation
    DOI:  https://doi.org/10.3390/cells9102197
  17. Mol Metab. 2020 Sep 30. pii: S2212-8778(20)30167-8. [Epub ahead of print] 101093
    Maeda Y, Kikuchi R, Kawagoe J, Tsuji T, Koyama N, Yamaguchi K, Nakamura H, Aoshiba K.
      OBJECTIVE: Tumor cells experience hypoxia, acidosis, and hypoglycemia. Metabolic adaptation to a glucose shortage is essential to maintain tumor cell survival because of their high glucose requirement. This study aimed to study the hypothesis that acidosis might promote tumor survival during a glucose shortage and if so, to explore a novel drug targeting metabolic vulnerability to glucose shortage.METHODS: Cell survival and bioenergetics metabolism were assessed in lung cancer cell lines. Our in-house small-molecule compounds were screened to identify those that kill cancer cells under low-glucose conditions. Cytotoxicity against non-cancerous cells was also assessed. Tumor growth was evaluated in vivo using a mouse engraft model.
    RESULTS: Acidosis limited the cellular consumption of glucose and ATP, causing tumor cells to enter a metabolically dormant but energetically economic state, which promoted tumor cell survival during glucose deficiency. We identified ESI-09, a previously known exchange protein directly activated by cAMP (EAPC) inhibitor, as an anticancer compound that inhibited cancer cells under low-glucose conditions, even when associated with acidosis. Bioenergetic studies showed that independent of EPAC inhibition, ESI-09 was a safer mitochondrial uncoupler than a classical uncoupler and created a futile cycling of mitochondrial respiration leading to decreased ATP production, increased ATP dissipation, and fuel scavenging. Accordingly, ESI-09 exhibited more cytotoxic effects under low-glucose conditions than under normal glucose conditions. ESI-09 was also more effective than actively proliferating cells on quiescent glucose-restricted cells. Cisplatin showed opposite effects. ESI-09 inhibited tumor growth in lung-cancer engraft mice.
    CONCLUSIONS: This study highlights the acidosis-induced promotion of tumor survival during glucose shortage and demonstrates that ESI-09 is a novel potent anticancer mitochondrial uncoupler that targets a metabolic vulnerability to glucose shortage even when associated with acidosis. The higher cytotoxicity under lower than normal glucose conditions suggests that ESI-09 is safer than conventional chemotherapy, can target the metabolic vulnerability of tumor cells to low-glucose stress, and is applicable to many cancer cell types.
    Keywords:  acidosis; glucose; lung cancer; mitochondrion; uncoupler
    DOI:  https://doi.org/10.1016/j.molmet.2020.101093
  18. Diabetes. 2020 Sep 29. pii: db200384. [Epub ahead of print]
    Cox AR, Chernis N, Bader DA, Saha PK, Masschelin PM, Felix JB, Sharp R, Lian Z, Putluri V, Rajapakshe K, Kim KH, Villareal DT, Armamento-Villareal R, Wu H, Coarfa C, Putluri N, Hartig SM.
      Obesity fosters low-grade inflammation in white adipose tissue (WAT) that may contribute to the insulin resistance that characterizes type 2 diabetes mellitus (T2DM). However, the causal relationship of these events remains unclear. The established dominance of signal transducer and activator of transcription 1 (STAT1) function in the immune response suggests an obligate link between inflammation and the co-morbidities of obesity. To this end, we sought to determine how STAT1 activity in white adipocytes affects insulin sensitivity. STAT1 expression in WAT inversely correlated with fasting plasma glucose in both obese mice and humans. Metabolomic and gene expression profiling established STAT1 deletion in adipocytes (STAT1 a-KO ) enhanced mitochondrial function and accelerated TCA cycle flux coupled with reduced fat cell size in subcutaneous WAT depots. STAT1 a-KO reduced WAT inflammation, but insulin resistance persisted in obese mice. Rather, elimination of type I cytokine interferon gamma (IFNγ) activity enhanced insulin sensitivity in diet-induced obesity. Our findings reveal a permissive mechanism that bridges WAT inflammation to whole-body insulin sensitivity.
    DOI:  https://doi.org/10.2337/db20-0384
  19. Biology (Basel). 2020 Sep 24. pii: E309. [Epub ahead of print]9(10):
    Belosludtseva NV, Starinets VS, Pavlik LL, Mikheeva IB, Dubinin MV, Belosludtsev KN.
      S-15176, a potent derivative of the anti-ischemic agent trimetazidine, was reported to have multiple effects on the metabolism of mitochondria. In the present work, the effect of S-15176 (1.5 mg/kg/day i.p.) on the ultrastructure and functions of liver mitochondria of C57BL/6 mice with type 2 diabetes mellitus (T2DM) induced by a high-fat diet combined with a low-dose streptozotocin injection was examined. An electron microscopy study showed that T2DM induced mitochondrial swelling and a reduction in the number of liver mitochondria. The number of mtDNA copies in the liver in T2DM decreased. The expression of Drp1 slightly increased, and that of Mfn2 and Opa1 somewhat decreased. The treatment of diabetic animals with S-15176 prevented the mitochondrial swelling, normalized the average mitochondrial size, and significantly decreased the content of the key marker of lipid peroxidation malondialdehyde in liver mitochondria. In S-15176-treated T2DM mice, a two-fold increase in the expression of the PGC-1α and a slight decrease in Drp 1 expression in the liver were observed. The respiratory control ratio, the level of mtDNA, and the number of liver mitochondria of S-15176-treated diabetic mice tended to restore. S-15176 did not affect the decrease in expression of Parkin and Opa1 in the liver of diabetic animals, but slightly suppressed the expression of these proteins in the control. The modulatory effect of S-15176 on dysfunction of liver mitochondria in T2DM can be related to the stimulation of mitochondrial biogenesis and the inhibition of lipid peroxidation in the organelles.
    Keywords:  S-15176 difumarate salt; mitochondrial biogenesis; mitochondrial dynamics; mitochondrial dysfunction; type 2 diabetes mellitus
    DOI:  https://doi.org/10.3390/biology9100309
  20. Cell. 2020 Oct 01. pii: S0092-8674(20)31225-3. [Epub ahead of print]183(1): 11-13
    Yan L, Chen YG.
      Circular RNAs (circRNAs) have emerged as key regulators of a wide variety of biological processes, but the roles of mitochondrial circRNAs are largely unknown. In this issue of Cell, Zhao et al. (2020) reveal that mitochondrial DNA-encoded circRNAs interact with ATP synthase subunit β (ATP5B) to inhibit the output of mitochondrial reactive oxygen species and the activation of liver fibroblasts, which regulate the pathogenesis of liver disease.
    DOI:  https://doi.org/10.1016/j.cell.2020.09.028
  21. Mol Cell Biochem. 2020 Sep 29.
    Sarkar A, Gandhi V.
      Ataxia telangiectasia mutated (ATM), a critical DNA damage sensor, also possesses non-nuclear functions owing to its presence in extra-nuclear compartments, including peroxisomes, lysosomes, and mitochondria. ATM is frequently altered in several human cancers. Recently, we and others have shown that loss of ATM is associated with defective mitochondrial autophagy (mitophagy) in ataxia-telangiectasia (A-T) fibroblasts and B-cell lymphomas. Further, we reported that ATM protein but not ATM kinase activity is required for mitophagy. However, the mechanism of ATM kinase activation during ionophore-induced mitophagy is unknown. In the work reported here, using several ionophores in A-T and multiple T-cell and B-cell lymphoma cell lines, we show that ionophore-induced mitophagy triggers oxidative stress-induced ATMSer1981 phosphorylation through ROS activation, which is different from neocarzinostatin-induced activation of ATMSer1981, Smc1Ser966, and Kap1Ser824. We used A-T cells overexpressed with WT or S1981A (auto-phosphorylation dead) ATM plasmids and show that ATM is activated by ROS-induced oxidative stress emanating from ionophore-induced mitochondrial damage and mitophagy. The antioxidants N-acetylcysteine and glutathione significantly inhibited ROS production and ATMSer1981 phosphorylation but failed to inhibit mitophagy as determined by retroviral infection with mt-mKeima construct followed by lysosomal dual-excitation ratiometric pH measurements. Our data suggest that while ATM kinase does not participate in mitophagy, it is activated via elevated ROS.
    Keywords:  ATM kinase; Leukemia; Lymphoma; Mitophagy; ROS
    DOI:  https://doi.org/10.1007/s11010-020-03917-1
  22. Aging (Albany NY). 2020 Sep 25. 12
    Sun L, Xu H, Wang Y, Ma X, Xu Y, Sun F.
      Chronic renal failure (CRF) is the final outcome of the development of chronic kidney disease with different causes. Although CRF is a common clinical disease, its pathogenesis remains to be improved. SBT-20 belongs to a class of cell-permeable peptides that target the inner mitochondrial membrane, reduce reactive oxygen species (ROS), normalize electron transport chain function, and ATP generation. Our experiment was to evaluate whether SBT-20 affected the oxidative stress and inflammatory process of CRF. The levels of ROS production, mitochondrial membrane potential, NF- κB-p65, TNF-α, Drp1, and mfn2 were measured before and after SBT-20 treatment. We observed that SBT-20 treatment inhibited H2O2-induced mitochondrial ROS production. SBT-20 could also restore the mitochondrial membrane potential and reduce the elevated levels of NF-κB-p65 and TNF-α in HK-2 cells. In vivo, the renal function of CRF mice recovered after treating with SBT-20, the levels of necrotic cells and inflammation decreased, and the morphology of mitochondria recovered. The results showed that SBT-20 had a protective effect on CRF by reducing oxidative stress, inflammation progression via down-regulating of NF-κB-p65, TNF-α, and Drp1 and upregulating of Mfn2. These data support SBT-20 could be used as a potential preparation for CRF.
    Keywords:  SBT-20; chronic renal failure; inflammation; mitochondrial-targeted peptide; oxidative stress
    DOI:  https://doi.org/10.18632/aging.103681
  23. J Biol Chem. 2020 Oct 01. pii: jbc.RA120.014884. [Epub ahead of print]
    Kunz HE, Dorschner JM, Berent TE, Meyer T, Wang X, Jatoi A, Kumar R, Lanza IR.
      Cancer cachexia is characterized by reductions in peripheral lean muscle mass. Prior studies have primarily focused on increased protein breakdown as the driver of cancer-associated muscle wasting. Therapeutic interventions targeting catabolic pathways have, however, largely failed to preserve muscle mass in cachexia, suggesting that other mechanisms might be involved. In pursuit of novel pathways, we used untargeted metabolomics to search for metabolite signatures that may be linked with muscle atrophy. We injected seven-week old C57/BL6 mice with LLC1 tumor cells or vehicle. After 21 days, tumor-bearing mice exhibited reduced body and muscle mass and impaired grip strength compared to controls, which was accompanied by lower synthesis rates of mixed muscle protein and the myofibrillar and sarcoplasmic muscle fractions. Reductions in protein synthesis were accompanied by mitochondrial enlargement and reduced coupling efficiency in tumor-bearing mice. To generate mechanistic insights into impaired protein synthesis, we performed untargeted metabolomic analyses of plasma and muscle and found increased concentrations of two methylarginines, asymmetric dimethylarginine (ADMA) and NG-monomethyl-L-arginine, in tumor-bearing mice compared to control mice. Compared to healthy controls, human cancer patients were also found to have higher levels of ADMA in the skeletal muscle. Treatment of C2C12 myotubes with ADMA impaired protein synthesis and reduced mitochondrial protein quality. These results suggest that increased levels of ADMA and mitochondrial changes may contribute to impaired muscle protein synthesis in cancer cachexia and could point to novel therapeutic targets by which to mitigate cancer cachexia.
    Keywords:  cachexia; cancer; metabolomics; methylarginines; mitochondria; protein synthesis; skeletal muscle
    DOI:  https://doi.org/10.1074/jbc.RA120.014884
  24. Cancer Metab. 2020 ;8 22
    Gaglio D, Bonanomi M, Valtorta S, Bharat R, Ripamonti M, Conte F, Fiscon G, Righi N, Napodano E, Papa F, Raccagni I, Parker SJ, Cifola I, Camboni T, Paci P, Colangelo AM, Vanoni M, Metallo CM, Moresco RM, Alberghina L.
      Abstract: Background: Rewiring of metabolism induced by oncogenic K-Ras in cancer cells involves both glucose and glutamine utilization sustaining enhanced, unrestricted growth. The development of effective anti-cancer treatments targeting metabolism may be facilitated by the identification and rational combinatorial targeting of metabolic pathways.
    Methods: We performed mass spectrometric metabolomics analysis in vitro and in vivo experiments to evaluate the efficacy of drugs and identify metabolic connectivity.
    Results: We show that K-Ras-mutant lung and colon cancer cells exhibit a distinct metabolic rewiring, the latter being more dependent on respiration. Combined treatment with the glutaminase inhibitor CB-839 and the PI3K/aldolase inhibitor NVP-BKM120 more consistently reduces cell growth of tumor xenografts. Maximal growth inhibition correlates with the disruption of redox homeostasis, involving loss of reduced glutathione regeneration, redox cofactors, and a decreased connectivity among metabolites primarily involved in nucleic acid metabolism.
    Conclusions: Our findings open the way to develop metabolic connectivity profiling as a tool for a selective strategy of combined drug repositioning in precision oncology.
    Keywords:  Combinatorial drug treatment; Glutamine; Glycolysis; Metabolic cancer therapy; Metabolic connectivity; Metabolic rewiring; Metabolic signature; Precision oncology
    DOI:  https://doi.org/10.1186/s40170-020-00227-4
  25. STAR Protoc. 2020 Sep 18. pii: 100089. [Epub ahead of print]1(2):
    Timón-Gómez A, Pérez-Pérez R, Nyvltova E, Ugalde C, Fontanesi F, Barrientos A.
      By using negatively charged Coomassie brilliant blue G-250 dye to induce a charge shift on proteins, blue native polyacrylamide gel electrophoresis (BN-PAGE) allows resolution of enzymatically active multiprotein complexes extracted from cellular or subcellular lysates while retaining their native conformation. BN-PAGE was first developed to analyze the size, composition, and relative abundance of the complexes and supercomplexes that form the mitochondrial respiratory chain and OXPHOS system. Here, we present a detailed protocol of BN-PAGE to obtain robust and reproducible results. For complete details on the use and execution of this protocol, please refer to Lobo-Jarne et al. (2018) and Timón-Gómez et al. (2020).
    DOI:  https://doi.org/10.1016/j.xpro.2020.100089
  26. Mol Genet Metab. 2020 Sep 18. pii: S1096-7192(20)30191-8. [Epub ahead of print]
    Kerr M, Hume S, Omar F, Koo D, Barnes H, Khan M, Aman S, Wei XC, Alfuhaid H, McDonald R, McDonald L, Newell C, Sparkes R, Hittel D, Khan A.
      Mitochondrial diseases, due to nuclear or mitochondrial genome mutations causing mitochondrial dysfunction, have a wide range of clinical features involving neurologic, muscular, cardiac, hepatic, visual, and auditory symptoms. Making a diagnosis of a mitochondrial disease is often challenging since there is no gold standard and traditional testing methods have required tissue biopsy which presents technical challenges and most patients prefer a non-invasive approach. Since a diagnosis invariably involves finding a disease-causing DNA variant, new approaches such as next generation sequencing (NGS) have the potential to make it easier to make a diagnosis. We evaluated the ability of our traditional diagnostic pathway (metabolite analysis, tissue neuropathology and respiratory chain enzyme activity) in 390 patients. The traditional diagnostic pathway provided a diagnosis of mitochondrial disease in 115 patients (29.50%). Analysis of mtDNA, tissue neuropathology, skin electron microscopy, respiratory chain enzyme analysis using inhibitor assays, blue native polyacrylamide gel electrophoresis were all statistically significant in distinguishing patients between a mitochondrial and non-mitochondrial diagnosis. From these 390 patients who underwent traditional analysis, we recruited 116 patients for the NGS part of the study (36 patients who had a mitochondrial diagnosis (MITO) and 80 patients who had no diagnosis (No-Dx)). In the group of 36 MITO patients, nuclear whole exome sequencing (nWES) provided a second diagnosis in 2 cases who already had a pathogenic variant in mtDNA, and a revised diagnosis (GLUL) in one case that had abnormal pathology but no pathogenic mtDNA variant. In the 80 NO-Dx patients, nWES found non-mitochondrial diagnosis in 26 patients and a mitochondrial diagnosis in 1 patient. A genetic diagnosis was obtained in 53/116 (45.70%) cases that were recruited for NGS, but not in 11/116 (9.48%) of cases with abnormal mitochondrial neuropathology. Our results show that a non-invasive, bigenomic sequencing (BGS) approach (using both a nWES and optimized mtDNA analysis to include large deletions) should be the first step in investigating for mitochondrial diseases. There may still be a role for tissue biopsy in unsolved cases or when the diagnosis is still not clear after NGS studies.
    DOI:  https://doi.org/10.1016/j.ymgme.2020.08.009
  27. J Immunol. 2020 Sep 30. pii: ji2000459. [Epub ahead of print]
    Sun Y, Preiss NK, Valenteros KB, Kamal Y, Usherwood YK, Frost HR, Usherwood EJ.
      CD8 T cell differentiation is orchestrated by dynamic metabolic changes that direct activation, proliferation, cytotoxic function, and epigenetic changes. We report that the BTB-ZF family transcriptional repressor Zbtb20 negatively regulates CD8 T cell metabolism and memory differentiation in mice. Effector and memory CD8 T cells with conditional Zbtb20 deficiency displayed enhanced mitochondrial and glycolytic metabolism, and memory CD8 T cells had enhanced spare respiratory capacity. Furthermore, Zbtb20-deficient CD8 T cells displayed increased flexibility in the use of mitochondrial fuel sources. Phenotypic and transcriptional skewing toward the memory fate was observed during the CD8 T cell response to Listeria monocytogenes Memory cells mounted larger secondary responses and conferred better protection following tumor challenge. These data suggest that inactivation of Zbtb20 may offer an approach to enhance metabolic activity and flexibility and improve memory CD8 T cell differentiation, useful attributes for T cells used in adoptive immunotherapy.
    DOI:  https://doi.org/10.4049/jimmunol.2000459
  28. Bioeng Transl Med. 2020 Sep;5(3): e10184
    Moon SJ, Dong W, Stephanopoulos GN, Sikes HD.
      Mitochondrial NADPH protects cells against mitochondrial oxidative stress by serving as an electron donor to antioxidant defense systems. However, due to technical challenges, it still remains unknown as to the pool size of mitochondrial NADPH, its dynamics, and NADPH/NADP+ ratio. Here, we have systemically modulated production rates of H2O2 in mitochondria and assessed mitochondrial NADPH metabolism using iNap sensors, 13C glucose isotopic tracers, and a mathematical model. Using sensors, we observed decreases in mitochondrial NADPH caused by excessive generation of mitochondrial H2O2, whereas the cytosolic NADPH was maintained upon perturbation. We further quantified the extent of mitochondrial NADPH/NADP+ based on the mathematical analysis. Utilizing 13C glucose isotopic tracers, we found increased activity in the pentose phosphate pathway (PPP) accompanied small decreases in the mitochondrial NADPH pool, whereas larger decreases induced both PPP activity and glucose anaplerosis. Thus, our integrative and quantitative approach provides insight into mitochondrial NADPH metabolism during mitochondrial oxidative stress.
    Keywords:  NADPH; NADPH metabolism; NADPH sensor; hydrogen peroxide; mitochondria; oxidative stress; redox kinetic model
    DOI:  https://doi.org/10.1002/btm2.10184
  29. Int J Mol Sci. 2020 Sep 28. pii: E7149. [Epub ahead of print]21(19):
    Nakashima C, Kirita T, Yamamoto K, Mori S, Luo Y, Sasaki T, Fujii K, Ohmori H, Kawahara I, Mori T, Goto K, Kishi S, Fujiwara-Tani R, Kuniyasu H.
      Budding at the tumor invasive front has been correlated with the malignant properties of many cancers. Malic enzyme 1 (ME1) promotes the Warburg effect in cancer cells and induces epithelial-mesenchymal transition (EMT) in oral squamous cell carcinoma (OSCC). Therefore, we investigated the role of ME1 in tumor budding in OSCC. Tumor budding was measured in 96 human OSCCs by immunostaining for an epithelial marker (AE1/AE3), and its expression was compared with that of ME1. A significant correlation was observed between tumor budding and ME1 expression. The correlation increased with the progression of cancer. In human OSCC cells, lactate secretion decreased when lactate fermentation was suppressed by knockdown of ME1 and lactate dehydrogenase A or inhibition of pyruvate dehydrogenase (PDH) kinase. Furthermore, the extracellular pH increased, and the EMT phenotype was suppressed. In contrast, when oxidative phosphorylation was suppressed by PDH knockdown, lactate secretion increased, extracellular pH decreased, and the EMT phenotype was promoted. Induction of chemical hypoxia in OSCC cells by CoCl2 treatment resulted in increased ME1 expression along with HIF1α expression and promotion of the EMT phenotype. Hypoxic conditions also increased matrix metalloproteinases expression and decreased mitochondrial membrane potential, mitochondrial oxidative stress, and extracellular pH. Furthermore, the hypoxic treatment resulted in the activation of Yes-associated protein (YAP), which was abolished by ME1 knockdown. These findings suggest that cancer cells at the tumor front in hypoxic environments increase their lactate secretion by switching their energy metabolism from oxidative phosphorylation to glycolysis owing to ME1 overexpression, decrease in extracellular pH, and YAP activation. These alterations enhance EMT and the subsequent tumor budding. Tumor budding and ME1 expression are thus considered useful markers of OSCC malignancy, and ME1 is expected to be a relevant target for molecular therapy.
    Keywords:  EMT; ME1; extracellular pH; hypoxia; tumor budding
    DOI:  https://doi.org/10.3390/ijms21197149
  30. Exp Oncol. 2020 09;42(3): 192-196
    Pyaskovskaya ON, Kolesnik DL, Prokhorova IV, Burlaka АP, Gorbach OI, Solyanik GI.
      BACKGROUND: Taking into account differences in the bioenergetics between malignant and normal cells a search of antitumor drugs among the modifiers of tumor metabolism has a reasonable excuse. Earlier it was found that the cytotoxic/cytostatic action of sodium dichloroacetate (DCA) against Lewis lung carcinoma (LLC) cells in vitro was enhanced in the case of its combination with metformin (MTF).AIM: To study the antitumor action of DCA in combination with MTF against LLC in vivo.
    MATERIALS AND METHODS: LLC/R9, a low metastatic variant of LLC cells, was used. LLC/R9 bearing mice were treated with MTF (at a total dose 0.15 g/kg b.w.) alone or in combination with DCA (at a total dose of 0.75 g/kg b.w.). LLC/R9 growth kinetics and the primary tumor growth and metastasis indices on the 23rd day after tumor cell inoculation were evaluated by routine procedures. The state of the electron transport chain of mitochondria in tumor cells was studied using electron paramagnetic resonance. The content of lactate and glucose in blood plasma from mice was measured by enzymatic methods using biochemical analyzer. The number of tumor-associated macrophages (TAMs) and their distribution by M1/M2 phenotype were estimated by flow cytometry using antibodies against CD68 and CD206.
    RESULTS: In LLC/R9-bearing mice treated with DCA in combination with MTF, tumor growth and metastasis indices, as well as circulating glucose and lactate levels were not significantly different from those in the control group. The level of nitrosylation of non-heme and heme proteins and the content of iron-sulfur centers in the mitochondria of tumor cells in LLC/R9-bearing mice administered with DCA in combination with MTF did not also differ from the corresponding indices in control. Instead, in tumors treated with MTF alone and in combination with DCA the total CD68+ TAMs count was almost 27% (p < 0.05) and 43% lower (p < 0.05) correspondingly than that in control, but this decrease was not accompanied by redistribution of CD68+/CD206+ and CD68+/D206- subsets.
    CONCLUSION: DCA in combination with MTF, at least in doses applied, did not affect LLC/R9 growth and metastasis in vivo. The complete absence of an antitumor effect of DCA in combination with MTF was simultaneously associated with the absence of significant changes in the functional state of electron transport chain of mitochondria in tumor cells, circulating glucose and lactate levels, and the decrease of the TAMs amount in tumors. It suggests that the antitumor activity of DCA and MTF could be determined by both their local effects within a tumor and their multiple systemic impacts.
  31. EMBO J. 2020 Oct 01. 39(19): e103530
    Beretta M, Santos CX, Molenaar C, Hafstad AD, Miller CC, Revazian A, Betteridge K, Schröder K, Streckfuß-Bömeke K, Doroshow JH, Fleck RA, Su TP, Belousov VV, Parsons M, Shah AM.
      Cells subjected to environmental stresses undergo regulated cell death (RCD) when homeostatic programs fail to maintain viability. A major mechanism of RCD is the excessive calcium loading of mitochondria and consequent triggering of the mitochondrial permeability transition (mPT), which is especially important in post-mitotic cells such as cardiomyocytes and neurons. Here, we show that stress-induced upregulation of the ROS-generating protein Nox4 at the ER-mitochondria contact sites (MAMs) is a pro-survival mechanism that inhibits calcium transfer through InsP3 receptors (InsP3 R). Nox4 mediates redox signaling at the MAM of stressed cells to augment Akt-dependent phosphorylation of InsP3 R, thereby inhibiting calcium flux and mPT-dependent necrosis. In hearts subjected to ischemia-reperfusion, Nox4 limits infarct size through this mechanism. These results uncover a hitherto unrecognized stress pathway, whereby a ROS-generating protein mediates pro-survival effects through spatially confined signaling at the MAM to regulate ER to mitochondria calcium flux and triggering of the mPT.
    Keywords:  InsP3 receptor; NADPH oxidase-4; calcium signaling; cell death; mitochondria-associated membrane
    DOI:  https://doi.org/10.15252/embj.2019103530
  32. Nat Commun. 2020 10 01. 11(1): 4913
    Guo L, Cui C, Wang J, Yuan J, Yang Q, Zhang P, Su W, Bao R, Ran J, Wu C.
      Reprograming of proline metabolism is critical for tumor growth. Here we show that PINCH-1 is highly expressed in lung adenocarcinoma and promotes proline synthesis through regulation of mitochondrial dynamics. Knockout (KO) of PINCH-1 increases dynamin-related protein 1 (DRP1) expression and mitochondrial fragmentation, which suppresses kindlin-2 mitochondrial translocation and interaction with pyrroline-5-carboxylate reductase 1 (PYCR1), resulting in inhibition of proline synthesis and cell proliferation. Depletion of DRP1 reverses PINCH-1 deficiency-induced defects on mitochondrial dynamics, proline synthesis and cell proliferation. Furthermore, overexpression of PYCR1 in PINCH-1 KO cells restores proline synthesis and cell proliferation, and suppresses DRP1 expression and mitochondrial fragmentation. Finally, ablation of PINCH-1 from lung adenocarcinoma in mouse increases DRP1 expression and inhibits PYCR1 expression, proline synthesis, fibrosis and tumor growth. Our results identify a signaling axis consisting of PINCH-1, DRP1 and PYCR1 that regulates mitochondrial dynamics and proline synthesis, and suggest an attractive strategy for alleviation of tumor growth.
    DOI:  https://doi.org/10.1038/s41467-020-18753-6
  33. Cell Rep. 2020 Sep 29. pii: S2211-1247(20)31185-2. [Epub ahead of print]32(13): 108196
    Xie S, Ni J, McFaline-Figueroa JR, Wang Y, Bronson RT, Ligon KL, Wen PY, Roberts TM, Zhao JJ.
      Loss of PTEN, the negative regulator of PI3K activity, is frequent in glioblastomas (GBMs). However, the role of the two major PI3K isoforms, p110α and p110β, in PTEN-deficient gliomagenesis remains unknown. We show that PTEN-deficient GBM largely depends on p110α for proliferation and p110β for migration. Genetic ablation of either isoform delays tumor progression in mice, but only ablating both isoforms completely blocks GBM driven by the concurrent ablation of Pten and p53. BKM120 (buparlisib) treatment only modestly prolongs survival in mice bearing intracranial Pten/p53 null tumors due to partial pathway inhibition. BKM120 extends the survival of mice bearing intracranial tumors in which p110β, but not p110α, has been genetically ablated in the Pten/p53 null glioma, indicating that BKM120 fails to inhibit p110β effectively. Our study suggests that the failure of PI3K inhibitors in GBM may be due to insufficient inhibition of p110β and indicates a need to develop brain-penetrant p110α/β inhibitors.
    Keywords:  BKM120; BYL719; GEMM; PDX; PI3K isoform; PTEN; PTEN-deficient; glioblastoma; isoform-selective inhibitor; migration
    DOI:  https://doi.org/10.1016/j.celrep.2020.108196
  34. Biochim Biophys Acta Biomembr. 2020 Sep 28. pii: S0005-2736(20)30326-6. [Epub ahead of print] 183483
    Rokitskaya TI, Kotova EA, Luzhkov VB, Kirsanov RS, Aleksandrova EV, Korshunova GA, Tashlitsky VN, Antonenko YN.
      To clarify the contribution of charge delocalization in a lipophilic ion to the efficacy of its permeation through a lipid membrane, we compared the behavior of alkyl derivatives of triphenylphosphonium, tricyclohexylphosphonium and trihexylphosphonium both in natural and artificial membranes. Exploring accumulation of the lipophilic cations in response to inside-negative membrane potential generation in mitochondria by using an ion-selective electrode revealed similar mitochondrial uptake of butyltricyclohexylphosphonium (C4TCHP) and butyltriphenylphosphonium (C4TPP). Fluorescence correlation spectroscopy also demonstrated similar membrane potential-dependent accumulation of fluorescein derivatives of tricyclohexyldecylphosphonium and decyltriphenylphosphonium in mitochondria. The rate constant of lipophilic cation translocation across the bilayer lipid membrane (BLM), measured by the current relaxation method, moderately increased in the following sequence: trihexyltetradecylphosphonium ([P6,6,6,14]) < triphenyltetradecylphosphonium (C14TPP) < tricyclohexyldodecylphosphonium (C12TCHP). In line with these results, measurements of the BLM stationary conductance indicated that membrane permeability for C4TCHP is 2.5 times higher than that for C4TPP. Values of the difference in the free energy of ion solvation in water and octane calculated using the density functional theory and the polarizable continuum solvent model were similar for methyltriphenylphosphonium, tricyclohexylmethylphosphonium and trihexylmethylphosphonium. Our results prove that both cyclic and aromatic moieties are not necessary for lipophilic ions to effectively permeate through lipid membranes.
    Keywords:  Bilayer lipid membrane; Charge delocalization; Ionic liquids; Lipophilic ions; Permeation
    DOI:  https://doi.org/10.1016/j.bbamem.2020.183483
  35. Redox Rep. 2020 Dec;25(1): 87-94
    Gill R, Mallay S, Young A, Mailloux RJ.
      Our group recently documented that male mice containing a deletion for one copy of the glutaredoxin-2 (Grx2) gene were completely protected from developing diet-induced obesity (DIO). Objectives: Here, we conducted a similar investigation but with female littermates. Results: In comparison to our recent publication using male mice, exposure of WT and GRX2+/- female mice to a HFD from 3-to-10 weeks of age did not induce any changes in body mass, circulating blood glucose, food intake, hepatic glycogen levels, or abdominal fat pad mass. Examination of the bioenergetics of muscle mitochondria revealed no changes in the rate of superoxide ( O 2 ∙ - )/hydrogen peroxide (H2O2) or O2 consumption under different states of respiration or alterations in lipid peroxidation adduct levels regardless of mouse strain or diet. Additionally, we measured the bioenergetics of mitochondria isolated from liver tissue and found that partial loss of GRX2 augmented respiration but did not alter ROS production. Discussion: Overall, our findings demonstrate there are sex differences in the protection of female GRX2+/- mice from DIO, fat accretion, intrahepatic lipid accumulation, and the bioenergetics of mitochondria from muscle and liver tissue.
    Keywords:  Mitochondria; ROS; bioenergetics; glutaredoxin-2; glutathionylation; obesity; redox buffering‌; sex dimorphisms
    DOI:  https://doi.org/10.1080/13510002.2020.1826750
  36. Nat Cell Biol. 2020 Oct;22(10): 1276-1285
    Chen Y, Jiang T, Zhang H, Gou X, Han C, Wang J, Chen AT, Ma J, Liu J, Chen Z, Jing X, Lei H, Wang Z, Bao Y, Baqri M, Zhu Y, Bindra RS, Hansen JE, Dou J, Huang C, Zhou J.
      Breast cancer brain metastasis (BCBM) is a devastating disease. Radiation therapy remains the mainstay for treatment of this disease. Unfortunately, its efficacy is limited by the dose that can be safely applied. One promising approach to overcoming this limitation is to sensitize BCBMs to radiation by inhibiting their ability to repair DNA damage. Here, we report a DNA repair suppressor, leucine-rich repeat-containing protein 31 (LRRC31), that was identified through a genome-wide CRISPR screen. We found that overexpression of LRRC31 suppresses DNA repair and sensitizes BCBMs to radiation. Mechanistically, LRRC31 interacts with Ku70/Ku80 and the ataxia telangiectasia mutated and RAD3-related (ATR) at the protein level, resulting in inhibition of DNA-dependent protein kinase, catalytic subunit (DNA-PKcs) recruitment and activation, and disruption of the MutS homologue 2 (MSH2)-ATR module. We demonstrate that targeted delivery of the LRRC31 gene via nanoparticles improves the survival of tumour-bearing mice after irradiation. Collectively, our study suggests LRRC31 as a major DNA repair suppressor that can be targeted for cancer radiosensitizing therapy.
    DOI:  https://doi.org/10.1038/s41556-020-00586-6
  37. Acta Virol. 2020 Sep 28.
    Szulc-Dąbrowska L, Wyżewski Z, Gregorczyk-Zboroch KP, Toka FN, Szczepanowska J, Struzik J, Nowak-Życzyńska Z, Gieryńska M, Niemiałtowski M.
      Mitochondria are multitasking organelles that play a central role in energy production, survival and primary host defense against viral infections. Therefore, viruses target mitochondria dynamics and functions to benefit their replication and morphogenetic processes. We endeavor to understand the role of mitochondria during infection of ectromelia virus (ECTV), hence our investigations on mitochondria-related genes in non-immune (L929 fibroblasts) and immune (RAW 264.7 macrophages) cells. Our results show that during later stages of infection, ECTV significantly decreases the expression of mitochondria-related genes regulating many aspects of mitochondrial physiology and functions, including mitochondrial transport, small molecule transport, membrane polarization and potential, targeting proteins to mitochondria, inner membrane translocation, and apoptosis. Such down-regulation is cell-specific, since macrophages exhibited a more profound down-regulation of mitochondria-related genes compared to infected L929 fibroblasts. Only L929 cells exhibited up-regulation of two important genes responsible for oxidative phosphorylation and subsequent ATP production: Slc25a23 and Slc25a31. Changes in the expression of mitochondria-related genes are accompanied by altered mitochondria morphology and distribution in both types of cells. In depth Ingenuity Pathway Analysis (IPA) identified the "Sirtuin Signaling Pathway" as the most significant top canonical pathway associated with ECTV infection in both analyzed cell types. Taken together, down-regulation of mitochondria-related genes observed especially in macrophages indicates dysfunctional mitochondria, possibly contributing to energy collapse and induction of intrinsic pathway of apoptosis. Meanwhile, alteration of the expression of several mitochondria-related genes in fibroblasts without apoptosis induction may represent poxviral strategy to control cellular energy metabolism for efficient replication. Keywords: Ectromelia virus; mitochondria; fibroblasts; macrophages.
    DOI:  https://doi.org/10.4149/av_2020_305
  38. Protein Cell. 2020 Oct 01.
    Koppula P, Zhuang L, Gan B.
      The cystine/glutamate antiporter SLC7A11 (also commonly known as xCT) functions to import cystine for glutathione biosynthesis and antioxidant defense and is overexpressed in multiple human cancers. Recent studies revealed that SLC7A11 overexpression promotes tumor growth partly through suppressing ferroptosis, a form of regulated cell death induced by excessive lipid peroxidation. However, cancer cells with high expression of SLC7A11 (SLC7A11high) also have to endure the significant cost associated with SLC7A11-mediated metabolic reprogramming, leading to glucose- and glutamine-dependency in SLC7A11high cancer cells, which presents potential metabolic vulnerabilities for therapeutic targeting in SLC7A11high cancer. In this review, we summarize diverse regulatory mechanisms of SLC7A11 in cancer, discuss ferroptosis-dependent and -independent functions of SLC7A11 in promoting tumor development, explore the mechanistic basis of SLC7A11-induced nutrient dependency in cancer cells, and conceptualize therapeutic strategies to target SLC7A11 in cancer treatment. This review will provide the foundation for further understanding SLC7A11 in ferroptosis, nutrient dependency, and tumor biology and for developing novel effective cancer therapies.
    Keywords:  SLC7A11; cancer therapy; cysteine; cystine; ferroptosis; nutrient dependency; xCT
    DOI:  https://doi.org/10.1007/s13238-020-00789-5
  39. World J Gastroenterol. 2020 Sep 14. 26(34): 5074-5089
    Zhao Y, Feng F, Guo QH, Wang YP, Zhao R.
      Gastrointestinal stromal tumors (GISTs) are the most common mesenchymal tumors of the gastrointestinal tract. At the molecular level, GISTs can be categorized into two groups based on the causative oncogenic mutations. Approximately 85% of GISTs are caused by gain-of-function mutations in the tyrosine kinase receptor KIT or platelet-derived growth factor receptor alpha (PDGFRA). The remaining GISTs, referred to as wild-type (WT) GISTs, are often deficient in succinate dehydrogenase complex (SDH), a key metabolic enzyme complex in the tricarboxylic acid (TCA) cycle and electron transport chain. SDH deficiency leads to the accumulation of succinate, a metabolite produced by the TCA cycle. Succinate inhibits α-ketoglutarate-dependent dioxygenase family enzymes, which comprise approximately 60 members and regulate key aspects of tumorigenesis such as DNA and histone demethylation, hypoxia responses, and m6A mRNA modification. For this reason, succinate and metabolites with similar structures, such as D-2-hydroxyglutarate and fumarate, are considered oncometabolites. In this article, we review recent advances in the understanding of how metabolic enzyme mutations and oncometabolites drive human cancer with an emphasis on SDH mutations and succinate in WT GISTs.
    Keywords:  Epigenetics; Gastrointestinal stromal tumors; Oncometabolite; Succinate; Succinate dehydrogenase; α-ketoglutarate-dependent dioxygenase
    DOI:  https://doi.org/10.3748/wjg.v26.i34.5074
  40. Sci Adv. 2020 Sep;pii: eabc4149. [Epub ahead of print]6(40):
    Popgeorgiev N, Sa JD, Jabbour L, Banjara S, Nguyen TTM, Akhavan-E-Sabet A, Gadet R, Ralchev N, Manon S, Hinds MG, Osigus HJ, Schierwater B, Humbert PO, Rimokh R, Gillet G, Kvansakul M.
      In metazoans, Bcl-2 family proteins are major regulators of mitochondrially mediated apoptosis; however, their evolution remains poorly understood. Here, we describe the molecular characterization of the four members of the Bcl-2 family in the most primitive metazoan, Trichoplax adhaerens All four trBcl-2 homologs are multimotif Bcl-2 group, with trBcl-2L1 and trBcl-2L2 being highly divergent antiapoptotic Bcl-2 members, whereas trBcl-2L3 and trBcl-2L4 are homologs of proapoptotic Bax and Bak, respectively. trBax expression permeabilizes the mitochondrial outer membrane, while trBak operates as a BH3-only sensitizer repressing antiapoptotic activities of trBcl-2L1 and trBcl-2L2. The crystal structure of a trBcl-2L2:trBak BH3 complex reveals that trBcl-2L2 uses the canonical Bcl-2 ligand binding groove to sequester trBak BH3, indicating that the structural basis for apoptosis control is conserved from T. adhaerens to mammals. Finally, we demonstrate that both trBax and trBak BH3 peptides bind selectively to human Bcl-2 homologs to sensitize cancer cells to chemotherapy treatment.
    DOI:  https://doi.org/10.1126/sciadv.abc4149
  41. Cancer Res. 2020 Sep 30. pii: canres.1094.2020. [Epub ahead of print]
    Li M, Kirtane AR, Kiyokawa J, Nagashima H, Lopes A, Tirmizi ZA, Lee CK, Traverso G, Cahill DP, Wakimoto H.
      The aggressive primary brain tumor glioblastoma (GBM) is characterized by aberrant metabolism that fuels its malignant phenotype. Diverse genetic subtypes of malignant glioma are sensitive to selective inhibition of the NAD+ salvage pathway enzyme nicotinamide phosphoribosyltransferase (NAMPT). However, the potential impact of NAD+ depletion on the brain tumor microenvironment has not been elaborated. In addition, systemic toxicity of NAMPT inhibition remains a significant concern. Here we show that microparticle-mediated intratumoral delivery of NAMPT inhibitor GMX1778 induces specific immunological changes in the tumor microenvironment of murine GBM, characterized by upregulation of immune checkpoint PD-L1, recruitment of CD3+, CD4+, and CD8+ T cells, and reduction of M2-polarized immunosuppressive macrophages. NAD+ depletion and autophagy induced by NAMPT inhibitors mediated the upregulation of PD-L1 transcripts and cell surface protein levels in GBM cells. NAMPT inhibitor modulation of the tumor immune microenvironment was therefore combined with PD-1 checkpoint blockade in vivo, significantly increasing the survival of GBM bearing animals. Thus, the therapeutic impacts of NAMPT inhibition extended beyond neoplastic cells, shaping surrounding immune effectors. Microparticle delivery and release of NAMPT inhibitor at the tumor site offers a safe and robust means to alter an immune tumor microenvironment that could potentiate checkpoint immunotherapy for glioblastoma.
    DOI:  https://doi.org/10.1158/0008-5472.CAN-20-1094
  42. Nat Cell Biol. 2020 Oct;22(10): 1180-1186
    Rey T, Zaganelli S, Cuillery E, Vartholomaiou E, Croisier M, Martinou JC, Manley S.
      Mitochondria contain the genetic information and expression machinery to produce essential respiratory chain proteins. Within the mitochondrial matrix, newly synthesized RNA, RNA processing proteins and mitoribosome assembly factors form punctate sub-compartments referred to as mitochondrial RNA granules (MRGs)1-3. Despite their proposed importance in regulating gene expression, the structural and dynamic properties of MRGs remain largely unknown. We investigated the internal architecture of MRGs using fluorescence super-resolution localization microscopy and correlative electron microscopy, and found that the MRG ultrastructure consists of compacted RNA embedded within a protein cloud. Using live-cell super-resolution structured illumination microscopy and fluorescence recovery after photobleaching, we reveal that MRGs rapidly exchange components and can undergo fusion, characteristic properties of fluid condensates4. Furthermore, MRGs associate with the inner mitochondrial membrane and their fusion coincides with mitochondrial remodelling. Inhibition of mitochondrial fission or fusion leads to an aberrant accumulation of MRGs into concentrated pockets, where they remain as distinct individual units despite their close apposition. Together, our findings reveal that MRGs are nanoscale fluid compartments, which are dispersed along mitochondria via membrane dynamics.
    DOI:  https://doi.org/10.1038/s41556-020-00584-8
  43. Nat Commun. 2020 09 30. 11(1): 4907
    Zheng DW, Hong S, Zhang QL, Dong X, Pan P, Song WF, Song W, Cheng SX, Zhang XZ.
      Global alterations in the metabolic network provide substances and energy to support tumor progression. To fuel these metabolic processes, extracellular matrix (ECM) plays a dominant role in supporting the mass transport and providing essential nutrients. Here, we report a fibrinogen and thrombin based coagulation system to construct an artificial ECM (aECM) for selectively cutting-off the tumor metabolic flux. Once a micro-wound is induced, a cascaded gelation of aECM can be triggered to besiege the tumor. Studies on cell behaviors and metabolomics reveal that aECM cuts off the mass transport and leads to a tumor specific starvation to inhibit tumor growth. In orthotopic and spontaneous murine tumor models, this physical barrier also hinders cancer cells from distant metastasis. The in vivo gelation provides an efficient approach to selectively alter the tumor mass transport. This strategy results in a 77% suppression of tumor growth. Most importantly, the gelation of aECM can be induced by clinical operations such as ultrasonic treatment, surgery or radiotherapy, implying this strategy is potential to be translated into a clinical combination regimen.
    DOI:  https://doi.org/10.1038/s41467-020-18493-7
  44. Front Immunol. 2020 ;11 1834
    Li W, Zhang L.
      Memory T cells persist for long term to mediate robust recall response upon rechallenging with previous encountered pathogens. The memory T cell pool is highly heterogeneous based on distinct phenotypic, functional, and locational properties, and contains discrete subsets, which contribute to diverse immune responses. In this mini-review, we will briefly discuss the distinct subsets of memory T cells and then focus on mitochondria-related metabolic and epigenetic regulations of CD8+ T cell memory formation. In particular, we discuss many aspects of mitochondrial quality control systems (biogenesis, dynamics, etc.) in regulating CD8+ T cell fate decision and antitumor immunity. Importantly, targeting mitochondrial metabolism to boost T cell memory formation and metabolic fitness might represent an attractive strategy to improve cancer immunotherapy including CAR-T therapy.
    Keywords:  CD8 T cell; cancer; immunotherapy; memory; mitochondria
    DOI:  https://doi.org/10.3389/fimmu.2020.01834
  45. Nat Commun. 2020 Oct 02. 11(1): 4946
    Jing Y, Liu J, Ye Y, Pan L, Deng H, Wang Y, Yang Y, Diao L, Lin SH, Mills GB, Zhuang G, Xue X, Han L.
      Immune-related adverse events (irAEs), caused by anti-PD-1/PD-L1 antibodies, can lead to fulminant and even fatal consequences and thus require early detection and aggressive management. However, a comprehensive approach to identify biomarkers of irAE is lacking. Here, we utilize a strategy that combines pharmacovigilance data and omics data, and evaluate associations between multi-omics factors and irAE reporting odds ratio across different cancer types. We identify a bivariate regression model of LCP1 and ADPGK that can accurately predict irAE. We further validate LCP1 and ADPGK as biomarkers in an independent patient-level cohort. Our approach provides a method for identifying potential biomarkers of irAE in cancer immunotherapy using both pharmacovigilance data and multi-omics data.
    DOI:  https://doi.org/10.1038/s41467-020-18742-9