bims-mibica Biomed News
on Mitochondrial bioenergetics in cancer
Issue of 2024–11–17
24 papers selected by
Kelsey Fisher-Wellman, Wake Forest University
  1. Targeting mitochondrial metabolism by the mitotoxin bromoxib in leukemia and lymphoma cells.
  2. Targeting mitochondrial metabolism with CPI-613 in chemoresistant ovarian tumors.
  3. Significantly reduced, but balanced, rates of mitochondrial fission and fusion are sufficient to maintain the integrity of yeast mitochondrial DNA.
  4. eIF5A controls mitoprotein import by relieving ribosome stalling at TIM50 translocase mRNA.
  5. Parallel phosphoproteomics and metabolomics map the global metabolic tyrosine phosphoproteome.
  6. A metabolic switch to the pentose-phosphate pathway induces radiation resistance in pancreatic cancer.
  7. GPR68 supports AML cells through the calcium/calcineurin pro-survival pathway and confers chemoresistance by mediating glucose metabolic symbiosis.
  8. Desuccinylation of Inosine-5´-monophosphate Dehydrogenase 1 by SIRT5 Promotes Tumor Cell Proliferation.
  9. PINK1-deficiency facilitates mitochondrial iron accumulation and colon tumorigenesis.
  10. Itaconate promotes an unexpected tumor immune escape mechanism.
  11. Spatial metabolomics highlights metabolic reprogramming in acute myeloid leukemia mice through creatine pathway.
  12. Mitochondrial Membrane Potential (ΔΨ) Fluctuations Associated with the Metabolic States of Mitochondria.
  13. PRMT1-mediated methylation of ME2 promotes hepatocellular carcinoma growth by inhibiting ubiquitination.
  14. CRISPR-Cas9 mediated knockout of NDUFS4 in human iPSCs: A model for mitochondrial complex I deficiency.
  15. Opposing roles for AMPK in regulating distinct mitophagy pathways.
  16. Isocitrate dehydrogenase 2 mutation promotes cytarabine resistance in acute myeloid leukemia by Warburg effect.
  17. PRDX6 contributes to selenocysteine metabolism and ferroptosis resistance.
  18. Deciphering the Metabolic Basis and Molecular Circuitry of the Warburg Paradox in Lymphoma.
  19. Targeting ketone body metabolism in mitigating gemcitabine resistance.
  20. Venetoclax plus low intensity chemotherapy for adults with acute lymphoblastic leukemia.
  21. Osteosarcoma stem cells resist chemotherapy by maintaining mitochondrial dynamic stability via DRP1.
  22. Succinate dehydrogenase deficiency-driven succinate accumulation induces drug resistance in acute myeloid leukemia via ubiquitin-cullin regulation.
  23. Modulatory Effect of Nicotinamide Adenine Dinucleotide Phosphate (NADPH) on the 2-Oxoglutarate Mitochondrial Carrier.
  24. Effect and mechanisms of shikonin on breast cancer cells in vitro and in vivo.
  1. Cell Commun Signal. 2024 Nov 12. 22(1): 541
    Targeting mitochondrial metabolism by the mitotoxin bromoxib in leukemia and lymphoma cells.
    Laura Schmitt, Karina S Krings, Andre Wolsing, Xabier Buque, Marcel Zimmermann, Hector Flores-Romero, Thomas Lenz, Ilka Lechtenberg, Christoph Peter, Björn Stork, Nicole Teusch, Peter Proksch, Kai Stühler, Ana J García-Sáez, Andreas S Reichert, Patricia Aspichueta, Sanil Bhatia, Sebastian Wesselborg.
      Targeting mitochondrial metabolism represents a promising approach for cancer treatment. Here, we investigated the mitotoxic potential of the polybrominated diphenyl ether bromoxib, a natural compound isolated from the marine sponge Dysidea family. We could show that bromoxib comprised strong cytotoxicity in different leukemia and lymphoma cell lines (such as HL60, HPBALL, Jurkat, K562, KOPTK1, MOLT4, SUPB15 and Ramos), but also in solid tumor cell lines (such as glioblastoma cell lines SJ-GBM2 and TP365MG). Bromoxib activated the mitochondrial death pathway as evidenced by the rapid translocation of Bax to the mitochondria and the subsequent mitochondrial release of Smac. Accordingly, bromoxib-induced apoptosis was blocked in caspase 9 deficient Jurkat cells and Jurkat cells overexpressing the antiapoptotic protein Bcl-2. In addition, we could show that bromoxib functioned as an uncoupler of the electron transport chain with similar rapid kinetics as CCCP in terms of dissipation of the mitochondrial membrane potential (ΔΨm), processing of the dynamin-like GTPase OPA1 and subsequent fragmentation of mitochondria. Beyond that, bromoxib strongly abrogated ATP production via glycolysis as well as oxidative phosphorylation (OXPHOS) by targeting electron transport chain complexes II, III, and V (ATP-synthase) in Ramos lymphoma cells. Thus, bromoxib's potential to act on both cytosolic glycolysis and mitochondrial respiration renders it a promising agent for the treatment of leukemia and lymphoma.
    Keywords:  Leukemia; Metabolism; OXPHOS; Protonophore
    DOI:  https://doi.org/10.1186/s12964-024-01913-2
  2. J Ovarian Res. 2024 Nov 14. 17(1): 226
    Targeting mitochondrial metabolism with CPI-613 in chemoresistant ovarian tumors.
    Mary P Udumula, Faraz Rashid, Harshit Singh, Tim Pardee, Sanjeev Luther, Tanya Bhardwaj, Km Anjaly, Sofia Piloni, Miriana Hijaz, Radhika Gogoi, Philip A Philip, Adnan R Munkarah, Shailendra Giri, Ramandeep Rattan.
       BACKGROUND: There is evidence indicating that chemoresistance in tumor cells is mediated by the reconfiguration of the tricarboxylic acid cycle, leading to heightened mitochondrial activity and oxidative phosphorylation (OXPHOS). Previously, we have shown that ovarian cancer cells that are resistant to chemotherapy display increased OXPHOS, mitochondrial function, and metabolic flexibility. To exploit this weakness in chemoresistant ovarian cancer cells, we examined the effectiveness of the mitochondrial inhibitor CPI-613 in treating preclinical ovarian cancer.
    METHODS: Chemosensitive OVCAR3, and chemoresistant CAOV3 and F2 ovarian cancer cells lines and their xenografts in nude mice were used. Functional metabolic studies were performed using Seahorse instrument. Metabolite quantification was performed using LC/MS/MS.
    RESULTS: Mice treated with CPI-613 exhibited a notable increase in overall survival and a reduction in tumor development and burden in OVCAR3, F2, and CAOV3 xenografts. CPI-613 suppressed the activity of pyruvate dehydrogenase and alpha-ketoglutarate dehydrogenase complex, which are two of its targets. This led to a reduction in OXPHOS and tricarboxylic acid cycle activity in all 3 xenografts. The addition of CPI-613 enhanced the responsiveness of chemotherapy in the chemoresistant F2 and CAOV3 tumors, resulting in a notable improvement in survival rates and a reduction in tumor size as compared to using chemotherapy alone. CPI-613 reduced the chemotherapy-induced OXPHOS in chemoresistant tumors. The study revealed that the mechanism by which CPI-613 inhibits tumor growth is through mitochondrial collapse. This is evidenced by an increase in superoxide production within the mitochondria, a decrease in ATP generation, and the release of cytochrome C, which triggers mitochondria-induced apoptosis.
    CONCLUSION: Our study demonstrates the translational potential of CPI-613 against chemoresistant ovarian tumors.
    Keywords:  Apoptosis; CPI-613; Mitochondria; Ovarian cancer
    DOI:  https://doi.org/10.1186/s13048-024-01546-6
  3. Mol Biol Cell. 2024 Nov 13. mbcE24070306
    Significantly reduced, but balanced, rates of mitochondrial fission and fusion are sufficient to maintain the integrity of yeast mitochondrial DNA.
    Brett T Wisniewski, Jason C Casler, Laura L Lackner.
      Mitochondria exist as dynamic tubular networks and the morphology of these networks impacts organelle function and cell health. Mitochondrial morphology is maintained in part by the opposing activities of mitochondrial fission and fusion. Mitochondrial fission and fusion are also required to maintain mitochondrial DNA (mtDNA) integrity. In Saccharomyces cerevisiae, the simultaneous inhibition of mitochondrial fission and fusion results in increased mtDNA mutation and the consequent loss of respiratory competence. The mechanism by which fission and fusion maintain mtDNA integrity is not fully understood. Previous work demonstrates that mtDNA is spatially linked to mitochondrial fission sites. Here, we extend this finding using live-cell imaging to localize mtDNA to mitochondrial fusion sites. While mtDNA is present at sites of mitochondrial fission and fusion, mitochondrial fission and fusion rates are not altered in cells lacking mtDNA. Using alleles that alter mitochondrial fission and fusion rates, we find that mtDNA integrity can be maintained in cells with significantly reduced, but balanced, rates of fission and fusion. In addition, we find that increasing mtDNA copy number reduces the loss of respiratory competence in double mitochondrial fission-fusion mutants. Our findings add novel insights into the relationship between mitochondrial dynamics and mtDNA integrity.
    DOI:  https://doi.org/10.1091/mbc.E24-07-0306
  4. J Cell Biol. 2024 Dec 02. pii: e202404094. [Epub ahead of print]223(12):
    eIF5A controls mitoprotein import by relieving ribosome stalling at TIM50 translocase mRNA.
    Marina Barba-Aliaga, Vanessa Bernal, Cynthia Rong, Madeleine E Volfbeyn, Keguang Zhang, Brian M Zid, Paula Alepuz.
      Efficient import of nuclear-encoded proteins into mitochondria is crucial for proper mitochondrial function. The conserved translation factor eIF5A binds ribosomes, alleviating stalling at polyproline-encoding sequences. eIF5A impacts mitochondrial function across species, though the precise molecular mechanism is unclear. We found that eIF5A depletion in yeast reduces the translation and levels of the TCA cycle and oxidative phosphorylation proteins. Loss of eIF5A causes mitoprotein precursors to accumulate in the cytosol and triggers a mitochondrial import stress response. We identify an essential polyproline protein as a direct target of eIF5A: the mitochondrial inner membrane protein and translocase component Tim50. Thus, eIF5A controls mitochondrial protein import by alleviating ribosome stalling along Tim50 mRNA at the mitochondrial surface. Removal of polyprolines from Tim50 partially rescues the mitochondrial import stress response and translation of oxidative phosphorylation genes. Overall, our findings elucidate how eIF5A impacts the mitochondrial function by promoting efficient translation and reducing ribosome stalling of co-translationally imported proteins, thereby positively impacting the mitochondrial import process.
    DOI:  https://doi.org/10.1083/jcb.202404094
  5. Proc Natl Acad Sci U S A. 2024 Nov 19. 121(47): e2413837121
    Parallel phosphoproteomics and metabolomics map the global metabolic tyrosine phosphoproteome.
    Alissandra L Hillis, Tigist Tamir, Grace E Perry, John M Asara, Jared L Johnson, Tomer M Yaron, Lewis C Cantley, Forest M White, Alex Toker.
      Tyrosine phosphorylation of metabolic enzymes is an evolutionarily conserved posttranslational modification that facilitates rapid and reversible modulation of enzyme activity, localization, or function. Despite the high abundance of tyrosine phosphorylation events detected on metabolic enzymes in high-throughput mass spectrometry-based studies, functional characterization of tyrosine phosphorylation sites has been limited to a subset of enzymes. Since tyrosine phosphorylation is dysregulated across human diseases, including cancer, understanding the consequences of metabolic enzyme tyrosine phosphorylation events is critical for informing disease biology and therapeutic interventions. To globally identify metabolic enzyme tyrosine phosphorylation events and simultaneously assign functional significance to these sites, we performed parallel phosphoproteomics and polar metabolomics in nontumorigenic mammary epithelial cells (MCF10A) stimulated with epidermal growth factor (EGF) in the absence or presence of the EGF receptor inhibitor erlotinib. We performed an integrated analysis of the phosphoproteomic and metabolomic datasets to identify tyrosine phosphorylation sites on metabolic enzymes with functional consequences. We identified two previously characterized (pyruvate kinase muscle isozyme, phosphoglycerate mutase 1) and two uncharacterized (glutathione S-transferase Pi 1, glutamate dehydrogenase 1) tyrosine phosphorylation sites on metabolic enzymes with purported functions based on metabolomic analyses. We validated these hits using a doxycycline-inducible CRISPR interference system in MCF10A cells, in which target metabolic enzymes were depleted with simultaneous reexpression of wild-type, phosphomutant, or phosphomimetic isoforms. Together, these data provide a framework for identification, prioritization, and characterization of tyrosine phosphorylation sites on metabolic enzymes with functional significance.
    Keywords:  EGFR; cancer metabolism; metabolomics; phosphotyrosine; proteomics
    DOI:  https://doi.org/10.1073/pnas.2413837121
  6. Radiother Oncol. 2024 Nov 07. pii: S0167-8140(24)04268-3. [Epub ahead of print] 110606
    A metabolic switch to the pentose-phosphate pathway induces radiation resistance in pancreatic cancer.
    Ariel Shimoni-Sebag, Ifat Abramovich, Bella Agranovich, Rami Massri, Chani Stossel, Dikla Atias, Maria Raites-Gurevich, Keren Yizhak, Talia Golan, Eyal Gottlieb, Yaacov Richard Lawrence.
       PURPOSE: Pancreatic ductal adenocarcinoma (PDAC) is remarkably resistant to standard modalities, including radiotherapy. We hypothesized that metabolic reprogramming may underlie PDAC radioresistance, and moreover, that it would be possible to exploit these metabolic changes for therapeutic intent.
    METHODS AND MATERIALS: We established two matched models of radioresistant PDAC cells by exposing the AsPC-1 and MIAPaCa-2 human pancreatic cancer cells to incremental doses of radiation. The metabolic profile of parental and radioresistant cells was investigated using Nanostring technology, labeled-glucose tracing by liquid chromatography-mass spectrometry, Seahorse analysis and exposure to metabolic inhibitors. The synergistic effect of radiation combined with a pentose-phosphate pathway inhibitor, 6-aminonicotinamide (6-AN) was evaluated in a xenograft model established by subcutaneous injection of radioresistant-AsPC-1 cells into nude mice.
    RESULTS: The radioresistant cells overexpressed pyruvate dehydrogenase kinase (PDK) and consistently, displayed increased glycolysis and downregulated the tricarboxylic acid (TCA) cycle and oxidative phosphorylation. Metabolic flux through the pentose-phosphate pathway (PPP) was increased, as were levels of reduced glutathione; pharmacological inhibition of the PPP dramatically potentiated radiation-induced cell death. Furthermore, the combined treatment of radiation with the PPP inhibitor 6-AN synergistically inhibited tumor growth in-vivo.
    CONCLUSIONS: We provide a mechanistic understanding of the metabolic changes that underlie radioresistance in PDAC. Furthermore, we demonstrate that pancreatic cancer cells can be re-sensitized to radiation via metabolic manipulation, in particular, inhibition of the PPP. Exploitation of the metabolic vulnerabilities of radioresistant pancreatic cancer cells constitutes a new approach to pancreatic cancer, with a potential to improve clinical outcomes.
    Keywords:  DNA damage; Metabolic reprogramming; Pancreatic cancer; Pentose-phosphate pathway; Radiation resistance; Radiation therapy
    DOI:  https://doi.org/10.1016/j.radonc.2024.110606
  7. Biochim Biophys Acta Mol Basis Dis. 2024 Nov 08. pii: S0925-4439(24)00559-3. [Epub ahead of print] 167565
    GPR68 supports AML cells through the calcium/calcineurin pro-survival pathway and confers chemoresistance by mediating glucose metabolic symbiosis.
    Xiaofei He, Caleb Hawkins, Lauren Lawley, Tra Mi Phan, Isaac Park, Nicole Joven, Jiajia Zhang, Mark Wunderlich, Benjamin Mizukawa, Shanshan Pei, Amisha Patel, Jennifer VanOudenhove, Stephanie Halene, Jing Fang.
      Accumulating evidence demonstrates that the "Warburg effect" that glycolysis is enhanced even in the presence of oxygen existed in hematopoietic malignancies, contributing to extracellular acidosis. G-protein coupled receptor 68 (GPR68), as a proton sensing GPCR responding to extracellular acidosis, is expected to play a critical role in hematopoietic malignancies. In the present study, we found that GPR68 was overexpressed in acute myeloid leukemia (AML) cells, and GPR68 deficiency impaired AML cell survival in vitro and cell engraftment in vivo. Mechanistic studies revealed that unlike GPR68 regulates Calpain1 in myelodysplastic syndromes (MDS) cells, GPR68 deficiency reduced cytosolic Ca2+ levels and calcineurin (CaN) activity in AML cells through an NFAT-independent mechanism. Moreover, the decreased Ca2+ levels disturbed cellular respiration (i.e., oxidative phosphorylation, OxPhos) by inhibiting isocitrate dehydrogenase (IDH) activity; this was more pronounced when BCL2 was inhibited simultaneously. Interestingly, GPR68 inhibition also decreased aerobic glycolysis in AML cells in a Ca2+-independent manner, suggesting that GPR68 mediated glucose metabolic symbiosis. As glucose metabolic symbiosis and the heterogeneous dependencies on aerobic glycolysis and cellular respiration tremendously impact chemosensitivity, the inhibition of GPR68 potentiated the tumoricidal effect of first-line chemotherapeutic agents, including BCL-2 inhibitors targeting OxPhos and cytarabine (AraC) targeting glycolysis. Consistent with these in vitro observations, higher levels of GPR68 were associated with inferior clinical outcomes in AML patients who received chemotherapies. In short, GPR68 drives the Ca2+/CaN pro-survival pathway and mediates glucose metabolic pathways in AML cells. Targeting GPR68 eradicates AML cells and alleviates chemoresistance, which could be exploited as a therapeutic target. The overexpression of GPR68 drives a Ca2+/CaN pro-survival pathway and mediates glucose metabolic symbiosis in AML cells, suggesting the diagnostic and therapeutic potential of GPR68 in AML. (GPR68, G proton-coupled receptor 68; PLCβ, phospholipase C beta; CaN, Calcineurin; IDH, isocitrate dehydrogenase; HIF-1α, Hypoxia-inducible factor alpha subunit; GLUT1, Glucose transporter type 1; HK-1, Hexokinase 1).
    Keywords:  Calcineurin; Calcium; Chemoresistance; Glucose metabolic symbiosis; Proton-sensing GPR68
    DOI:  https://doi.org/10.1016/j.bbadis.2024.167565
  8. J Biol Chem. 2024 Nov 08. pii: S0021-9258(24)02478-5. [Epub ahead of print] 107976
    Desuccinylation of Inosine-5´-monophosphate Dehydrogenase 1 by SIRT5 Promotes Tumor Cell Proliferation.
    Chang Xu, Pengbo Yao, Jie Cheng, Peng Jiang.
      Inosine-5´-monophosphate dehydrogenase (IMPDH) catalyzes the rate limiting step of de novo purine synthesis. Currently, it remains still largely unknown how this metabolic event is regulated in tumor cells. Here, we report that a deacetylase sirtuin 5 (SIRT5) may possess a regulatory effect on GMP anabolism by desuccinylating IMPDH1. We found that SIRT5 can directly interacts with IMPDH1 and promotes desuccinylation on the N terminal of IMPDH1, thereby leading to increased IMPDH enzymatic activity, enhanced purine biosynthesis and promoted cell proliferation. Consistently, down-regulation of SIRT5 expression results in decreased IMPDH1 activity and impaired tumor cell proliferation. Therefore, our results reveal that SIRT5-mediated IMPDH1 desuccinylation adapts purine metabolism for rapid cell growth, and could be a potential therapeutic target for tumor cell proliferation inhibition.
    Keywords:  Cell proliferation; Desuccinylation; IMPDH; SIRT5
    DOI:  https://doi.org/10.1016/j.jbc.2024.107976
  9. Autophagy. 2024 Nov 08.
    PINK1-deficiency facilitates mitochondrial iron accumulation and colon tumorigenesis.
    Mariella Arcos, Lavanya Goodla, Hyeoncheol Kim, Sharina P Desai, Rui Liu, Kunlun Yin, Zhaoli Liu, David R Martin, Xiang Xue.
      Mitophagy, the process by which cells eliminate damaged mitochondria, is mediated by PINK1 (PTEN induced kinase 1). Our recent research indicates that PINK1 functions as a tumor suppressor in colorectal cancer by regulating cellular metabolism. Interestingly, PINK1 ablation activated the NLRP3 (NLR family pyrin domain containing 3) inflammasome, releasing IL1B (interleukin 1 beta). However, inhibiting the NLRP3-IL1B signaling pathway with an IL1R (interleukin 1 receptor) antagonist or NLRP3 inhibitor did not hinder colon tumor growth after PINK1 loss. To identify druggable targets in PINK1-deficient tumors, ribonucleic acid sequencing analysis was performed on colon tumors from pink1 knockout and wild-type mice. Gene Set Enrichment Analysis highlighted the enrichment of iron ion transmembrane transporter activity. Subsequent qualitative polymerase chain reaction and western blot analysis revealed an increase in mitochondrial iron transporters, including mitochondrial calcium uniporter, in PINK1-deficient colon tumor cells and tissues. Live-cell iron staining demonstrated elevated cellular and mitochondrial iron levels in PINK1-deficient cells. Clinically used drugs deferiprone and minocycline reduced mitochondrial iron and superoxide levels, resulting in decreased colon tumor cell growth in vitro and in vivo. Manipulating the mitochondrial iron uptake protein MCU (mitochondrial calcium uniporter) also affected cell and xenograft tumor growth. This study suggests that therapies aimed at reducing mitochondrial iron levels may effectively inhibit colon tumor growth, particularly in patients with low PINK1 expression.
    Keywords:  Colorectal cancer; deferiprone; iron chelation; minocycline; mitochondrial iron; mitophagy
    DOI:  https://doi.org/10.1080/15548627.2024.2425594
  10. Cancer Cell. 2024 Oct 30. pii: S1535-6108(24)00399-4. [Epub ahead of print]
    Itaconate promotes an unexpected tumor immune escape mechanism.
    Lara Haase, Christian Frezza.
      Itaconate is a metabolite produced by macrophages upon infection and acts as an antimicrobial molecule. In this issue of Cancer Cell, Lin et al. found that itaconate produced by tumor-associated macrophages is taken up by cancer cells via the transporter solute carrier family 13 member 3 (SLC13A3), promoting resistance to immune checkpoint inhibitors.
    DOI:  https://doi.org/10.1016/j.ccell.2024.10.011
  11. Acta Pharm Sin B. 2024 Oct;14(10): 4461-4477
    Spatial metabolomics highlights metabolic reprogramming in acute myeloid leukemia mice through creatine pathway.
    Yucheng Bao, Jing Qiao, Wenjie Gong, Ruihong Zhang, Yanting Zhou, Yinyin Xie, Yuan Xie, Jiuming He, Tong Yin.
      Acute myeloid leukemia (AML) is recognized as an aggressive cancer that is characterized by significant metabolic reprogramming. Here, we applied spatial metabolomics to achieve high-throughput, in situ identification of metabolites within the liver metastases of AML mice. Alterations at metabolite and protein levels were further mapped out and validated by integrating untargeted metabolomics and proteomics. This study showed a downregulation in arginine's contribution to polyamine biosynthesis and urea cycle, coupled with an upregulation of the creatine metabolism. The upregulation of creatine synthetases Gatm and Gamt, as well as the creatine transporter Slc6a8, resulted in a marked accumulation of creatine within tumor foci. This process further enhances oxidative phosphorylation and glycolysis of leukemia cells, thereby boosting ATP production to foster proliferation and infiltration. Importantly, we discovered that inhibiting Slc6a8 can counter these detrimental effects, offering a new strategy for treating AML by targeting metabolic pathways.
    Keywords:  Acute myeloid leukemia; Creatine; Glycolysis; Metabolic reprogramming; Metastasis; Oxidative phosphorylation; Slc6a8; Spatial metabolomics
    DOI:  https://doi.org/10.1016/j.apsb.2024.07.004
  12. Methods Mol Biol. 2025 ;2878 117-131
    Mitochondrial Membrane Potential (ΔΨ) Fluctuations Associated with the Metabolic States of Mitochondria.
    Ivo F Machado, Raul G Miranda, João S Teodoro, Daniel J Dorta, Anabela P Rolo, Carlos M Palmeira.
      The proton electrochemical gradient generated by the respiratory chain activity accounts for over 90% of the available respiratory energy and, as such, its evaluation and accurate measurement regarding total values and fluctuations are an invaluable component of the understanding of mitochondrial function. Consequently, alterations in electric potential across the inner mitochondrial membrane generated by differential protonic accumulation and transport are known as the mitochondrial membrane potential, or Δψ, and are reflective of the functional metabolic status of mitochondria. There are several experimental approaches to measure Δψ, ranging from fluorometric evaluations to electrochemical probes. In this chapter, we describe how Δψ may be evaluated in isolated mitochondria and live cells using electrochemical and fluorescent methods, such as tetraphenylphosphonium (TPP+) and tetramethylrhodamine methyl ester (TMRM), respectively. These methods are dependent on the accumulation of cationic probes within mitochondria, which are assessed by using a TPP+-selective electrode or instruments that measure fluorescence (microplate reader and flow cytometer).
    Keywords:  Flow cytometry; Membrane potential; Metabolic states; Mitochondria; TMRM; TPP+-selective electrode
    DOI:  https://doi.org/10.1007/978-1-0716-4264-1_7
  13. Cell Death Dis. 2024 Nov 11. 15(11): 814
    PRMT1-mediated methylation of ME2 promotes hepatocellular carcinoma growth by inhibiting ubiquitination.
    Shuai Zhang, Shuling Zhang, Baijuan Xia, Xueying Li, Hongyu Jiang, Su Feng, Yang Xiang, Ya Qiu, Shi Zhou, Peng Luo.
      The mitochondrial malic enzyme 2 (ME2), which is frequently elevated during carcinogenesis and may be a target for cancer therapy, catalyzes the conversion of malate to pyruvate. The processes controlling ME2 activity, however, remain largely unclear. In this work, we show that human hepatocellular carcinoma (HCC) tissues contain high levels of ME2 and that the methylation of ME2 stimulates the growth and migration of HCC cells. Furthermore, we observed that ME2 interacts with protein arginine methyltransferase 1 (PRMT1) and that ME2 enzymatic activity is activated by mutation of ME2 at lysine 67. Mitochondrial respiration was markedly increased by activated ME2, which promoted cell division and carcinogenesis. Furthermore, a negative prognosis for patients was strongly linked with the expression levels of PRMT1 and ME2 R67K in HCC tissues. These findings imply that hepatocellular carcinoma growth is aided by PRMT1-mediated ME2 methylation, that is an essential signaling event that cancer cells need to continue mitochondrial respiration.
    DOI:  https://doi.org/10.1038/s41419-024-07219-y
  14. Biochim Biophys Acta Mol Basis Dis. 2024 Nov 13. pii: S0925-4439(24)00563-5. [Epub ahead of print] 167569
    CRISPR-Cas9 mediated knockout of NDUFS4 in human iPSCs: A model for mitochondrial complex I deficiency.
    Shivani Goolab, Karin Terburgh, Charl du Plessis, Janine Scholefield, Roan Louw.
      Mitochondrial diseases, often caused by defects in complex I (CI) of the oxidative phosphorylation system, currently lack curative treatments. Human-relevant, high-throughput drug screening platforms are crucial for the discovery of effective therapeutics, with induced pluripotent stem cells (iPSCs) emerging as a valuable technology for this purpose. Here, we present a novel iPSC model of NDUFS4-related CI deficiency that displays a strong metabolic phenotype in the pluripotent state. Human iPSCs were edited using CRISPR-Cas9 to target the NDUFS4 gene, generating isogenic NDUFS4 knockout (KO) cell lines. Sanger sequencing detected heterozygous biallelic deletions, whereas no indel mutations were found in isogenic control cells. Western blotting confirmed the absence of NDUFS4 protein in KO iPSCs and CI enzyme kinetics showed a ~56 % reduction in activity compared to isogenic controls. Comprehensive metabolomic profiling revealed a distinct metabolic phenotype in NDUFS4 KO iPSCs, predominantly associated with an elevated NADH/NAD+ ratio, consistent with alterations observed in other models of mitochondrial dysfunction. Additionally, β-lapachone, a recognized NAD+ modulator, alleviated reductive stress in KO iPSCs by modifying the redox state in both the cytosol and mitochondria. Although undifferentiated iPSCs cannot fully replicate the complex cellular dynamics of the disease seen in vivo, these findings highlight the utility of iPSCs in providing a relevant metabolic milieu that can facilitate early-stage, high-throughput exploration of therapeutic strategies for mitochondrial dysfunction.
    Keywords:  CI deficiency; CRISPR-Cas9; Mitochondrial disease; iPSC
    DOI:  https://doi.org/10.1016/j.bbadis.2024.167569
  15. Mol Cell. 2024 Nov 05. pii: S1097-2765(24)00865-7. [Epub ahead of print]
    Opposing roles for AMPK in regulating distinct mitophagy pathways.
    Marianna Longo, Aniketh Bishnu, Pierpaolo Risiglione, Lambert Montava-Garriga, Joyceline Cuenco, Kei Sakamoto, Carol MacKintosh, Ian G Ganley.
      Mitophagy degrades damaged mitochondria, but we show here that it can also target functional mitochondria. This latter scenario occurs during programmed mitophagy and involves the mitophagy receptors NIX and BNIP3. Although AMP-activated protein kinase (AMPK), the energy-sensing protein kinase, can influence damaged-induced mitophagy, its role in programmed mitophagy is unclear. We found that AMPK directly inhibits NIX-dependent mitophagy by triggering 14-3-3-mediated sequestration of ULK1, via ULK1 phosphorylation at two sites: Ser556 and an additional identified site, Ser694. By contrast, AMPK activation increases Parkin phosphorylation and enhances the rate of depolarization-induced mitophagy, independently of ULK1. We show that this happens both in cultured cells and tissues in vivo, using the mito-QC mouse model. Our work unveils a mechanism whereby AMPK activation downregulates mitophagy of functional mitochondria but enhances that of dysfunctional/damaged ones.
    Keywords:  14-3-3; AMPK; NIX; Parkin; ULK1; autophagy; liver; mito-QC; mitophagy; skeletal muscle
    DOI:  https://doi.org/10.1016/j.molcel.2024.10.025
  16. Hematol Oncol. 2024 Nov;42(6): e3316
    Isocitrate dehydrogenase 2 mutation promotes cytarabine resistance in acute myeloid leukemia by Warburg effect.
    Jinrong Yang, Zixu Wang, Kun Wu, Bo Nie, Liyin Li, Jingyan Ruan, Qiang Zhou, Yun Zeng, Mingxia Shi.
      Mutation of isocitrate dehydrogenase 2 (IDH2) is a key factor in promoting cytarabine (Ara-C) resistance in acute myeloid leukemia (AML), however the underly mechanism remains unclear. Acute myeloid leukemia cells, were cultured with either IDH2 knockdown (KD-IDH2) or overexpression (OE-IDH2) to elucidate the role of IDH2 in these leukemic cell lines. Additionally, mutant cell lines were engineered to replicate clinically relevant IDH2 mutations. To investigate cellular responses, the glycolytic inhibitor 2-deoxy-D-glucose (2-DG) was administered to the cells. Cell proliferation was quantified using a Cell Counting Kit-8 (CCK-8), while apoptosis was evaluated through propidium iodide staining followed by flow cytometry. Glycolytic metabolism levels were measured using a specific reagent kit, and Western blotting was employed to determine the expression levels of glycolysis-related proteins. Transcriptome sequencing was conducted to elucidate the mechanisms by which IDH2 mutations influence glycolysis. Furthermore, both in vitro cell experiments and in vivo subcutaneous transplantation tumor models in nude mice were utilized to validate these mechanisms. OE-IDH2 in AML cells, enhances resistance to the Ara-C, promotes cell proliferation and glycolysis, and inhibits apoptosis. KD-IDH2 exhibits opposite effects. Both IDH2 mutations and OE-IDH2 produce similar effects on these cellular processes. The increase in glycolysis levels following IDH2 mutation may contribute to the reduced efficacy of Enasidenib in inhibiting the proliferation of IDH-mutant AML cells. Transcriptome sequencing results indicate an enrichment of the PI3K/Akt signaling pathway in IDH2-mutant AML cells. BEZ235 significantly inhibits the expression of phosphorylated PI3K (p-PI3K), phosphorylated Akt (p-Akt), mTOR, glycolytic metabolism, and Ara-C resistance both in vitro and in vivo. Overexpression and mutation of IDH2 coordinate with the Warburg effect through the PI3K/Akt/mTOR pathway to promote Ara-C resistance in AML.
    Keywords:  AML cell; Ara‐C resistant; IDH2 mutation; Warburg effect; acute myeloid leukemia
    DOI:  https://doi.org/10.1002/hon.3316
  17. Mol Cell. 2024 Nov 07. pii: S1097-2765(24)00867-0. [Epub ahead of print]
    PRDX6 contributes to selenocysteine metabolism and ferroptosis resistance.
    Zhiyi Chen, Alex Inague, Kamini Kaushal, Gholamreza Fazeli, Danny Schilling, Thamara N Xavier da Silva, Ancely Ferreira Dos Santos, Tasneem Cheytan, Florencio Porto Freitas, Umut Yildiz, Lucas Gasparello Viviani, Rodrigo Santiago Lima, Mikaela Peglow Pinz, Isadora Medeiros, Thais Satie Iijima, Thiago Geronimo Pires Alegria, Railmara Pereira da Silva, Larissa Regina Diniz, Simon Weinzweig, Judith Klein-Seetharaman, Andreas Trumpp, Adriana Mañas, Robert Hondal, Christoph Bartenhagen, Matthias Fischer, Briana K Shimada, Lucia A Seale, Thilo Samson Chillon, Marietta Fabiano, Lutz Schomburg, Ulrich Schweizer, Luis E Netto, Flavia C Meotti, Tobias P Dick, Hamed Alborzinia, Sayuri Miyamoto, José Pedro Friedmann Angeli.
      Selenocysteine (Sec) metabolism is crucial for cellular function and ferroptosis prevention and begins with the uptake of the Sec carrier, selenoprotein P (SELENOP). Following uptake, Sec released from SELENOP is metabolized via selenocysteine lyase (SCLY), producing selenide, a substrate for selenophosphate synthetase 2 (SEPHS2), which provides the essential selenium donor, selenophosphate (H2SePO3-), for the biosynthesis of the Sec-tRNA. Here, we discovered an alternative pathway in Sec metabolism mediated by peroxiredoxin 6 (PRDX6), independent of SCLY. Mechanistically, we demonstrate that PRDX6 can readily react with selenide and interact with SEPHS2, potentially acting as a selenium delivery system. Moreover, we demonstrate the functional significance of this alternative route in human cancer cells, revealing a notable association between elevated expression of PRDX6 and human MYCN-amplified neuroblastoma subtype. Our study sheds light on a previously unrecognized aspect of Sec metabolism and its implications in ferroptosis, offering further possibilities for therapeutic exploitation.
    Keywords:  cancer metabolism; cell death; ferroptosis; neuroblastoma; selenium; selenocysteine metabolism
    DOI:  https://doi.org/10.1016/j.molcel.2024.10.027
  18. Cancers (Basel). 2024 Oct 25. pii: 3606. [Epub ahead of print]16(21):
    Deciphering the Metabolic Basis and Molecular Circuitry of the Warburg Paradox in Lymphoma.
    Dashnamoorthy Ravi, Athena Kritharis, Andrew M Evens.
      Background/Objectives: Warburg's metabolic paradox illustrates that malignant cells require both glucose and oxygen to survive, even after converting glucose into lactate. It remains unclear whether sparing glucose from oxidation intersects with TCA cycle continuity and if this confers any metabolic advantage in proliferating cancers. This study seeks to understand the mechanistic basis of Warburg's paradox and its overall implications for lymphomagenesis. Methods: Using metabolomics, we first examined the metabolomic profiles, glucose, and glutamine carbon labeling patterns in the metabolism during the cell cycle. We then investigated proliferation-specific metabolic features of malignant and nonmalignant cells. Finally, through bioinformatics and the identification of appropriate pharmacological targets, we established malignant-specific proliferative implications for the Warburg paradox associated with metabolic features in this study. Results: Our results indicate that pyruvate, lactate, and alanine levels surge during the S phase and are correlated with nucleotide synthesis. By using 13C1,2-Glucose and 13C6,15N2-Glutamine isotope tracers, we observed that the transamination of pyruvate to alanine is elevated in lymphoma and coincides with the entry of glutamine carbon into the TCA cycle. Finally, by using fludarabine as a strong inhibitor of lymphoma, we demonstrate that disrupting the transamination of pyruvate to alanine correlates with the simultaneous suppression of glucose-derived nucleotide biosynthesis and glutamine carbon entry into the TCA cycle. Conclusions: We conclude that the transamination of pyruvate to alanine intersects with reduced glucose oxidation and maintains the TCA cycle as a critical metabolic feature of Warburg's paradox and lymphomagenesis.
    Keywords:  Warburg effect; glucose; glutamine; lactate; lymphoma; nucleotides; targeted inhibitors; transaminase
    DOI:  https://doi.org/10.3390/cancers16213606
  19. JCI Insight. 2024 Nov 07. pii: e177840. [Epub ahead of print]
    Targeting ketone body metabolism in mitigating gemcitabine resistance.
    Krizia Rohena-Rivera, Sungyong You, Minhyung Kim, Sandrine Billet, Johanna Ten Hoeve, Gabrielle Gonzales, Chengqun Huang, Ashley Heard, Keith Syson Chan, Neil A Bhowmick.
      Chemotherapy is often combined with surgery for muscle invasive and non-muscle invasive bladder cancer. However, 70% of the patients recur within 5 years. Metabolic reprogramming is an emerging hallmark in cancer chemoresistance. Here, we report a gemcitabine resistance mechanism which promotes cancer reprogramming via the metabolic enzyme, OXCT1. This mitochondrial enzyme, responsible for the rate-limiting step in β-hydroxybutyrate (βHB) catabolism, was elevated in muscle invasive disease and in chemo-resistant bladder cancer patients. Resistant orthotopic tumors presented an OXCT1-dependent rise in mitochondrial oxygen consumption rate, ATP, and nucleotide biosynthesis. In resistant bladder cancer, knocking out OXCT1 restored gemcitabine sensitivity, and administering the non-metabolizable βHB, enantiomer (S-βHB) only partially restored gemcitabine sensitivity. Suggesting an extra-metabolic role for OXCT1, multi-omics analysis of gemcitabine sensitive and resistant cells revealed an OXCT1-dependent signature with the transcriptional repressor, OVOL1, as a master regulator of epithelial differentiation. The elevation of OVOL1 target genes was associated with its cytoplasmic translocation and poor prognosis in a chemotherapy-treated BCa patient cohort. The knockout of OXCT1 restored OVOL1 transcriptional repressive activity by its nuclear translocation. Orthotopic mouse models of bladder cancer supported OXCT1 as a mediator of gemcitabine sensitivity through ketone metabolism and regulating cancer stem cell differentiation.
    Keywords:  Cancer; Drug therapy; Oncology; Urology
    DOI:  https://doi.org/10.1172/jci.insight.177840
  20. Blood Adv. 2024 Nov 15. pii: bloodadvances.2024014405. [Epub ahead of print]
    Venetoclax plus low intensity chemotherapy for adults with acute lymphoblastic leukemia.
    Marlise R Luskin, Shai Shimony, Julia H Keating, Eric S Winer, Jacqueline S Garcia, Richard M Stone, Elias J Jabbour, Yael Flamand, Kristen Stevenson, Jeremy Adam Ryan, Zhihong Zeng, Anthony Letai, Marina Y Konopleva, Nitin Jain, Daniel J DeAngelo.
      In acute lymphoblastic leukemia (ALL), the BCL2 inhibitor venetoclax may enhance the efficacy of chemotherapy allowing dose reductions that reduce toxicity. We designed a phase 1b study of venetoclax plus attenuated chemotherapy to determine the recommended phase 2 dose (RP2D) of venetoclax. The study enrolled 19 patients with ALL either newly diagnosed (≥60 years, n=11 [B-cell, n=8; T-cell, n=3]) or R/R (≥18 years, n=8 [B-cell, n=3; T-cell, n=5]). Venetoclax was given for 21 days with each cycle of mini-hyper-CVD (cyclophosphamide, vincristine, dexamethasone alternating with methotrexate and cytarabine). There were no dose limiting toxicities at dose level (DL) 1 (n=3, 400 mg/day) or DL2 (n=6, 600 mg/day); DL2 was the RP2D and explored further (n=10). The most common non-hematologic adverse events were grade 3+ infections. There were no deaths within 60 days and no patients discontinued therapy for toxicity. There was no tumor lysis syndrome, hepatotoxicity, or prolonged cytopenias. Among patients with newly diagnosed ALL, 10/11 (90.9%) achieved a measurable residual disease-negative (<0.01% sensitivity) complete remission (CR) including 6 patients with hypodiploid TP53 mutated ALL. All patients in CR bridged to hematopoietic stem cell transplant (n=9) or completed protocol (n=1). With a median follow-up of 60.0 months, median disease-free survival (DFS) for patients with newly diagnosed ALL was 54.6 months (95% CI: 35.5 months - NA) with a 2-year DFS rate of 90% (95% CI, 71% - 100%). Among patients with R/R ALL, 3/8 (37.5%) achieved CR. In summary, for patients with newly diagnosed ALL, venetoclax plus mini-HCVD is well-tolerated with promising efficacy. NCT03319901.
    DOI:  https://doi.org/10.1182/bloodadvances.2024014405
  21. Int J Mol Med. 2025 Jan;pii: 10. [Epub ahead of print]55(1):
    Osteosarcoma stem cells resist chemotherapy by maintaining mitochondrial dynamic stability via DRP1.
    Boren Tian, Yaxuan Wu, Xiaoyun Du, Yan Zhang.
      Osteosarcoma malignancy exhibits significant heterogeneity, comprising both osteosarcoma stem cells (OSCs) and non‑OSCs. OSCs demonstrate increased resistance to chemotherapy due to their distinctive cellular and molecular characteristics. Alterations in mitochondrial morphology and homeostasis may enhance chemoresistance by modulating metabolic and regulatory processes. However, the relationship between mitochondrial homeostasis and chemoresistance in OSCs remains to be elucidated. The present study employed high‑resolution microscopy to perform multi‑layered image reconstructions for a quantitative analysis of mitochondrial morphology. The results indicated that OSCs exhibited larger mitochondria in comparison with non‑OSCs. Furthermore, treatment of OSCs with cisplatin (CIS) or doxorubicin (DOX) resulted in preserved mitochondrial morphological stability, which was not observed in non‑OSCs. This finding suggested a potential association between mitochondrial homeostasis and chemoresistance. Further analysis indicated that dynamin‑related protein 1 (DRP1) might play a pivotal role in maintaining the stability of mitochondrial homeostasis in OSCs. Depletion of DRP1 resulted in the disruption of mitochondrial stability when OSCs were treated with CIS or DOX. Additionally, knocking out DRP1 in OSCs led to a reduction in chemoresistance. These findings unveil a novel mechanism underlying chemoresistance in osteosarcoma and suggest that targeting DRP1 could be a promising therapeutic strategy to overcome chemoresistance in OSCs. This provided valuable insights for enhancing treatment outcomes among patients with osteosarcoma.
    Keywords:  cancer stem cells; chemoresistance; mitochondrial dynamics; osteosarcoma
    DOI:  https://doi.org/10.3892/ijmm.2024.5451
  22. Nat Commun. 2024 Nov 13. 15(1): 9820
    Succinate dehydrogenase deficiency-driven succinate accumulation induces drug resistance in acute myeloid leukemia via ubiquitin-cullin regulation.
    Yifan Chen, Miao Xian, Wenwen Ying, Jiayi Liu, Shaowei Bing, Xiaomin Wang, Jiayi Yu, Xiaojun Xu, Senfeng Xiang, Xuejing Shao, Ji Cao, Qiaojun He, Bo Yang, Meidan Ying.
      Drug resistance is vital for the poor prognosis of acute myeloid leukemia (AML) patients, but the underlying mechanism remains poorly understood. Given the unique microenvironment of bone marrow, we reasoned that drug resistance of AML might rely on distinct metabolic processes. Here, we identify succinate dehydrogenase (SDH) deficiency and over-cumulative succinate as typical features in AML, with a marked function in causing the resistance of AML cells to various anti-cancer therapies. Mechanistically, succinate promotes the accumulation of oncogenic proteins in a manner that precedes transcriptional activation. This function is mediated by succinate-triggered upregulation of ubiquitin-conjugating enzyme E2M (UBC12) phosphorylation, which impairs its E2 function in cullins neddylation. Notably, decreasing succinate by fludarabine can restore the sensitivity of anti-cancer drugs in SDH-deficient AML. Together, we uncover the function of succinate in driving drug resistance by regulating p-UBC12/cullin activity, and indicate reshaping succinate metabolism as a promising treatment for SDH-deficient AML.
    DOI:  https://doi.org/10.1038/s41467-024-53398-9
  23. Molecules. 2024 Oct 31. pii: 5154. [Epub ahead of print]29(21):
    Modulatory Effect of Nicotinamide Adenine Dinucleotide Phosphate (NADPH) on the 2-Oxoglutarate Mitochondrial Carrier.
    Anna Spagnoletta, Daniela Valeria Miniero, Nicola Gambacorta, Francesca Oppedisano, Anna De Grassi, Orazio Nicolotti, Ciro Leonardo Pierri, Annalisa De Palma.
      The 2-oxoglutarate carrier (OGC), pivotal in cellular metabolism, facilitates the exchange of key metabolites between mitochondria and cytosol. This study explores the influence of NADPH on OGC transport activity using proteoliposomes. Experimental data revealed the ability of NADPH to modulate the OGC activity, with a significant increase of 60% at 0.010 mM. Kinetic analysis showed increased Vmax and a reduction in Km for 2-oxoglutarate, suggesting a direct regulatory role. Molecular docking pointed to a specific interaction between NADPH and cytosolic loops of OGC, involving key residues such as K206 and K122. This modulation was unique in mammalian OGC, as no similar effect was observed in a plant OGC structurally/functionally related mitochondrial carrier. These findings propose OGC as a responsive sensor for the mitochondrial redox state, coordinating with the malate/aspartate and isocitrate/oxoglutarate shuttles to maintain redox balance. The results underscore the potential role of OGC in redox homeostasis and its broader implications in cellular metabolism and oxidative stress responses.
    Keywords:  NADPH regulation; isocitrate/oxoglutarate shuttle; kinetic analysis; malate/aspartate shuttle; mitochondrial function; mitochondrial transport; molecular docking; oxoglutarate carrier
    DOI:  https://doi.org/10.3390/molecules29215154
  24. BMC Complement Med Ther. 2024 Nov 08. 24(1): 389
    Effect and mechanisms of shikonin on breast cancer cells in vitro and in vivo.
    Chuyi Yu, Haoyu Xing, Xiaguo Fu, Yingying Zhang, Xiufang Yan, Jianjia Feng, Zhouqin He, Li Ru, Chunlong Huang, Jianming Liang.
       BACKGROUND: Breast cancer seriously affects physical and mental health of women. Despite advances in the clinical use of different treatments, breast cancer remains a major cause of mortality. Therefore, it is imperative to identify promising treatment options. In the present study, we investigated the effects of shikonin on 4T1 breast cancer cells and its potential mechanisms of action.
    METHODS: BALB/c-derived mouse breast cancer 4T1 is very close to human breast cancer in growth characteristics and systemic response, so 4T1 cells were selected for further experiments. Cell viability, apoptosis, intracellular reactive oxygen species (ROS), mitochondrial activity, and cellular calreticulin (CRT) exposure were assessed to evaluate the antitumor effects and mechanisms of shikonin in vitro. Orthotopic tumor growth inhibition and splenic immune cell regulation by shikonin were evaluated in 4T1 breast cancer orthotopic mice in vivo.
    RESULTS: In vitro, shikonin could inhibit cell proliferation, cause apoptosis, disrupt mitochondrial activity, and induce ROS production and CRT exposure. In vivo, shikonin inhibited tumor growth, increased the proportion of CD8+ T cells, and reduced the proportion of regulatory cells (CD25+ Foxp3+ T cells) in the spleen.
    CONCLUSIONS: Shikonin inhibits the growth of 4T1 breast cancer cells by disrupting mitochondrial activity, promoting oxidative stress, and regulating immune function.
    Keywords:  Breast cancer; Immunomodulation; Intracellular reactive oxygen species; Mitochondrial activity; Shikonin
    DOI:  https://doi.org/10.1186/s12906-024-04671-3
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