Abnormalities in cancer metabolism represent potential targets for cancer therapy. We have recently identified a natural compound Quambalarine B (QB), which inhibits proliferation of several leukemic cell lines followed by cell death. We have predicted ubiquinone binding sites of mitochondrial respiratory complexes as potential molecular targets of QB in leukemia cells. Hence, we tracked the effect of QB on leukemia metabolism by applying several omics and biochemical techniques. We have confirmed the inhibition of respiratory complexes by QB and found an increase in the intracellular AMP levels together with respiratory substrates. Inhibition of mitochondrial respiration by QB triggered reprogramming of leukemic cell metabolism involving disproportions in glycolytic flux, inhibition of proteins O-glycosylation, stimulation of glycine synthesis pathway, and pyruvate kinase activity, followed by an increase in pyruvate and a decrease in lactate levels. Inhibition of mitochondrial complex I by QB suppressed folate metabolism as determined by a decrease in formate production. We have also observed an increase in cellular levels of several amino acids except for aspartate, indicating the dependence of Jurkat (T-ALL) cells on aspartate synthesis. These results indicate blockade of mitochondrial complex I and II activity by QB and reduction in aspartate and folate metabolism as therapeutic targets in T-ALL cells. Anti-cancer activity of QB was also confirmed during in vivo studies, suggesting the therapeutic potential of this natural compound.

BIOCEV Institute of Biotechnology v v i The Czech Academy of Sciences Vestec Czech Republic

BIOCEV Institute of Microbiology v v i The Czech Academy of Sciences Vestec Czech Republic

Department of Biochemistry Faculty of Science Charles University Prague Czech Republic

Department of Cell Biology Faculty of Science Charles University Prague Czech Republic

Faculty of Chemical Technology University of Chemistry and Technology Prague Czech Republic

Institute of Microbiology v v i The Czech Academy of Sciences Prague Czech Republic

Zobrazit více v PubMed

Warburg O, Wind F, Negelein E. The Metabolism of Tumors in the Body. J Gen Physiol. 1927;8:519–30. PubMed PMC

Warburg O. On respiratory impairment in cancer cells. Science. 1956;124:269–70. PubMed

Vander Heiden MG, Cantley LC, Thompson CB. Understanding the Warburg effect: the metabolic requirements of cell proliferation. Science. 2009;324:1029–33. https://doi.org/10.1126/science.1160809. PubMed DOI PMC

David CJ, Chen M, Assanah M, Canoll P, Manley JL. HnRNP proteins controlled by c-Myc deregulate pyruvate kinase mRNA splicing in cancer. Nature. 2010;463:364–8. https://doi.org/10.1038/nature08697. PubMed DOI PMC

Papandreou I, Cairns RA, Fontana L, Lim AL, Denko NC. HIF-1 mediates adaptation to hypoxia by actively downregulating mitochondrial oxygen consumption. Cell Metab. 2006;3:187–97. https://doi.org/10.1016/j.cmet.2006.01.012. PubMed DOI

Wong N, Ojo D, Yan J, Tang D. PKM2 contributes to cancer metabolism. Cancer Lett. 2015;356:184–91. https://doi.org/10.1016/j.canlet.2014.01.031. PubMed DOI

Meiser J, Tumanov S, Maddocks O, Labuschagne CF, Athineos D, Van Den Broek N, Mackay GM, Gottlieb E, Blyth K, Vousden K, Kamphorst JJ, Vazquez A. Serine one-carbon catabolism with formate overflow. Sci Adv. 2016;2:e1601273. https://doi.org/10.1126/sciadv.1601273. PubMed DOI PMC

Anastasiou D, Yu Y, Israelsen WJ, Jiang JK, Boxer MB, Hong BS, Tempel W, Dimov S, Shen M, Jha A, Yang H, Mattaini KR, Metallo CM, et al. Pyruvate kinase M2 activators promote tetramer formation and suppress tumorigenesis. Nat Chem Biol. 2012;8:839–47. https://doi.org/10.1038/nchembio.1060. PubMed DOI PMC

Birsoy K, Wang T, Chen WW, Freinkman E, Abu-Remaileh M, Sabatini DM. An Essential Role of the Mitochondrial Electron Transport Chain in Cell Proliferation Is to Enable Aspartate Synthesis. Cell. 2015;162:540–51. https://doi.org/10.1016/j.cell.2015.07.016. PubMed DOI PMC

Sullivan LB, Gui DY, Hosios AM, Bush LN, Freinkman E, Vander Heiden MG. Supporting Aspartate Biosynthesis Is an Essential Function of Respiration in Proliferating Cells. Cell. 2015;162:552–63. https://doi.org/10.1016/j.cell.2015.07.017. PubMed DOI PMC

Wise DR, DeBerardinis RJ, Mancuso A, Sayed N, Zhang XY, Pfeiffer HK, Nissim I, Daikhin E, Yudkoff M, McMahon SB, Thompson CB. Myc regulates a transcriptional program that stimulates mitochondrial glutaminolysis and leads to glutamine addiction. Proc Natl Acad Sci U S A. 2008;105:18782–7. https://doi.org/10.1073/pnas.0810199105. PubMed DOI PMC

Valis K, Talacko P, Grobarova V, Cerny J, Novak P. Shikonin regulates C-MYC and GLUT1 expression through the MST1-YAP1-TEAD1 axis. Exp Cell Res. 2016;349:273–81. https://doi.org/10.1016/j.yexcr.2016.10.018. PubMed DOI

Valis K, Prochazka L, Boura E, Chladova J, Obsil T, Rohlena J, Truksa J, Dong LF, Ralph SJ, Neuzil J. Hippo/Mst1 stimulates transcription of the proapoptotic mediator NOXA in a FoxO1-dependent manner. Cancer Res. 2011;71:946–54. https://doi.org/10.1158/0008-5472.CAN-10-2203. PubMed DOI

Farber S, Diamond LK. Temporary remissions in acute leukemia in children produced by folic acid antagonist, 4-aminopteroyl-glutamic acid. N Engl J Med. 1948;238:787–93. https://doi.org/10.1056/NEJM194806032382301. PubMed DOI

Hermanova I, Arruabarrena-Aristorena A, Valis K, Nuskova H, Alberich-Jorda M, Fiser K, Fernandez-Ruiz S, Kavan D, Pecinova A, Niso-Santano M, Zaliova M, Novak P, Houstek J, et al. Pharmacological inhibition of fatty-acid oxidation synergistically enhances the effect of l-asparaginase in childhood ALL cells. Leukemia. 2016;30:209–18. https://doi.org/10.1038/leu.2015.213. PubMed DOI

Vander Heiden MG. Targeting cancer metabolism: a therapeutic window opens. Nat Rev Drug Discov. 2011;10:671–84. https://doi.org/10.1038/nrd3504. PubMed DOI

Klotz LO, Hou X, Jacob C. 1,4-naphthoquinones: from oxidative damage to cellular and inter-cellular signaling. Molecules. 2014;19:14902–18. https://doi.org/10.3390/molecules190914902. PubMed DOI PMC

Ramirez O, Motta-Mena LB, Cordova A, Estrada A, Li Q, Martinez L, Garza KM. A small library of synthetic di-substituted 1, 4-naphthoquinones induces ROS-mediated cell death in murine fibroblasts. PLoS One. 2014;9:e106828. https://doi.org/10.1371/journal.pone.0106828. PubMed DOI PMC

Yang JT, Li ZL, Wu JY, Lu FJ, Chen CH. An oxidative stress mechanism of shikonin in human glioma cells. PLoS One. 2014;9:e94180. https://doi.org/10.1371/journal.pone.0094180. PubMed DOI PMC

Singh PK, Sarwar M, Maklashina E, Kotlyar V, Rajagukguk S, Tomasiak TM, Cecchini G, Iverson TM. Plasticity of the quinone-binding site of the complex II homolog quinol:fumarate reductase. J Biol Chem. 2013;288:24293–301. https://doi.org/10.1074/jbc.M113.487082. PubMed DOI PMC

Baran I, Ganea C, Privitera S, Scordino A, Barresi V, Musumeci F, Mocanu MM, Condorelli DF, Ursu I, Grasso R, Gulino M, Garaiman A, Musso N, et al. Detailed analysis of apoptosis and delayed luminescence of human leukemia Jurkat T cells after proton irradiation and treatments with oxidant agents and flavonoids. Oxid Med Cell Longev. 2012;2012:498914. https://doi.org/10.1155/2012/498914. PubMed DOI PMC

Abdelmohsen K, Gerber PA, von Montfort C, Sies H, Klotz LO. Epidermal growth factor receptor is a common mediator of quinone-induced signaling leading to phosphorylation of connexin-43: role of glutathione and tyrosine phosphatases. J Biol Chem. 2003;278:38360–7. https://doi.org/10.1074/jbc.M306785200. PubMed DOI

Stodulkova E, Cisarova I, Kolarik M, Chudickova M, Novak P, Man P, Kuzma M, Pavlu B, Cerny J, Flieger M. Biologically active metabolites produced by the basidiomycete Quambalaria cyanescens. PLoS One. 2015;10:e0118913. https://doi.org/10.1371/journal.pone.0118913. PubMed DOI PMC

Grobarova V, Valis K, Talacko P, Pavlu B, Hernychova L, Novakova J, Stodulkova E, Flieger M, Novak P, Cerny J. Quambalarine B, a Secondary Metabolite from Quambalaria cyanescens with Potential Anticancer Properties. J Nat Prod. 2016;79:2304–14. https://doi.org/10.1021/acs.jnatprod.6b00362. PubMed DOI

Guo L, Shestov AA, Worth AJ, Nath K, Nelson DS, Leeper DB, Glickson JD, Blair IA. Inhibition of Mitochondrial Complex II by the Anticancer Agent Lonidamine. J Biol Chem. 2016;291:42–57. https://doi.org/10.1074/jbc.M115.697516. PubMed DOI PMC

Banerjee PS, Hart GW, Cho JW. Chemical approaches to study O-GlcNAcylation. Chem Soc Rev. 2013;42:4345–57. https://doi.org/10.1039/c2cs35412h. PubMed DOI PMC

Warmoes MO, Locasale JW. Heterogeneity of glycolysis in cancers and therapeutic opportunities. Biochem Pharmacol. 2014;92:12–21. https://doi.org/10.1016/j.bcp.2014.07.019. PubMed DOI PMC

Bond MR, Hanover JA. A little sugar goes a long way: the cell biology of O-GlcNAc. J Cell Biol. 2015;208:869–80. https://doi.org/10.1083/jcb.201501101. PubMed DOI PMC

Hart GW, Slawson C, Ramirez-Correa G, Lagerlof O. Cross talk between O-GlcNAcylation and phosphorylation: roles in signaling, transcription, and chronic disease. Annu Rev Biochem. 2011;80:825–58. https://doi.org/10.1146/annurev-biochem-060608-102511. PubMed DOI PMC

Chou TY, Dang CV, Hart GW. Glycosylation of the c-Myc transactivation domain. Proc Natl Acad Sci U S A. 1995;92:4417–21. PubMed PMC

Kim D, Fiske BP, Birsoy K, Freinkman E, Kami K, Possemato RL, Chudnovsky Y, Pacold ME, Chen WW, Cantor JR, Shelton LM, Gui DY, Kwon M, et al. SHMT2 drives glioma cell survival in ischaemia but imposes a dependence on glycine clearance. Nature. 2015;520:363–7. https://doi.org/10.1038/nature14363. PubMed DOI PMC

Gravel SP, Hulea L, Toban N, Birman E, Blouin MJ, Zakikhani M, Zhao Y, Topisirovic I, St-Pierre J, Pollak M. Serine deprivation enhances antineoplastic activity of biguanides. Cancer Res. 2014;74:7521–33. https://doi.org/10.1158/0008-5472.CAN-14-2643-T. PubMed DOI

Israelsen WJ, Vander Heiden MG. Pyruvate kinase: Function, regulation and role in cancer. Semin Cell Dev Biol. 2015;43:43–51. https://doi.org/10.1016/j.semcdb.2015.08.004. PubMed DOI PMC

Chaneton B, Hillmann P, Zheng L, Martin AC, Maddocks OD, Chokkathukalam A, Coyle JE, Jankevics A, Holding FP, Vousden KH, Frezza C, O’Reilly M, Gottlieb E. Serine is a natural ligand and allosteric activator of pyruvate kinase M2. Nature. 2012;491:458–62. https://doi.org/10.1038/nature11540. PubMed DOI PMC

Walsh MJ, Brimacombe KR, Anastasiou D, Yu Y, Israelsen WJ, Hong BS, Tempel W, Dimov S, Veith H, Yang H, Kung C, Yen KE, Dang L, et al. ML265: A potent PKM2 activator induces tetramerization and reduces tumor formation and size in a mouse xenograft model. Probe Reports from the NIH Molecular Libraries Program. (Bethesda (MD) National Center for Biotechnology Information (US) 2010 PubMed

Mattaini KR, Sullivan MR, Vander Heiden MG. The importance of serine metabolism in cancer. J Cell Biol. 2016;214:249–57. https://doi.org/10.1083/jcb.201604085. PubMed DOI PMC

Pavlova NN, Thompson CB. The Emerging Hallmarks of Cancer Metabolism. Cell Metab. 2016;23:27–47. https://doi.org/10.1016/j.cmet.2015.12.006. PubMed DOI PMC

Dong LF, Low P, Dyason JC, Wang XF, Prochazka L, Witting PK, Freeman R, Swettenham E, Valis K, Liu J, Zobalova R, Turanek J, Spitz DR, et al. Alpha-tocopheryl succinate induces apoptosis by targeting ubiquinone-binding sites in mitochondrial respiratory complex II. Oncogene. 2008;27:4324–35. https://doi.org/10.1038/onc.2008.69. PubMed DOI PMC

Dong LF, Neuzil J. Mitochondria in cancer: why mitochondria are a good target for cancer therapy. Prog Mol Biol Transl Sci. 2014;127:211–27. https://doi.org/10.1016/B978-0-12-394625-6.00008-8. PubMed DOI

Jara JA, Lopez-Munoz R. Metformin and cancer: Between the bioenergetic disturbances and the antifolate activity. Pharmacol Res. 2015;101:102–8. https://doi.org/10.1016/j.phrs.2015.06.014. PubMed DOI

Munkley J, Elliott DJ. Hallmarks of glycosylation in cancer. Oncotarget. 2016;7:35478–89. https://doi.org/10.18632/oncotarget.8155. PubMed DOI PMC

Chen J, Xie J, Jiang Z, Wang B, Wang Y, Hu X. Shikonin and its analogs inhibit cancer cell glycolysis by targeting tumor pyruvate kinase-M2. Oncogene. 2011;30:4297–306. https://doi.org/10.1038/onc.2011.137. PubMed DOI

Anastasiou D, Poulogiannis G, Asara JM, Boxer MB, Jiang JK, Shen M, Bellinger G, Sasaki AT, Locasale JW, Auld DS, Thomas CJ, Vander Heiden MG, Cantley LC. Inhibition of pyruvate kinase M2 by reactive oxygen species contributes to cellular antioxidant responses. Science. 2011;334:1278–83. https://doi.org/10.1126/science.1211485. PubMed DOI PMC

Tiwari P. Recent Trends in Therapeutic Approaches for Diabetes Management: A Comprehensive Update. J Diabetes Res. 2015;2015:340838. https://doi.org/10.1155/2015/340838. PubMed DOI PMC

Livak KJ, Schmittgen TD. Analysis of relative gene expression data using real-time quantitative PCR and the 2(-Delta Delta C(T)) Method. Methods. 2001;25:402–8. https://doi.org/10.1006/meth.2001.1262. PubMed DOI

Gottschalk M, Ivanova G, Collins DM, Eustace A, O’Connor R, Brougham DF. Metabolomic studies of human lung carcinoma cell lines using in vitro (1)H NMR of whole cells and cellular extracts. NMR Biomed. 2008;21:809–19. https://doi.org/10.1002/nbm.1258. PubMed DOI

Wishart DS, Tzur D, Knox C, Eisner R, Guo AC, Young N, Cheng D, Jewell K, Arndt D, Sawhney S, Fung C, Nikolai L, Lewis M, et al. HMDB: the Human Metabolome Database. Nucleic Acids Res. 2007;35:D521–6. https://doi.org/10.1093/nar/gkl923. PubMed DOI PMC

Masuda T, Tomita M, Ishihama Y. Phase transfer surfactant-aided trypsin digestion for membrane proteome analysis. J Proteome Res. 2008;7:731–40. https://doi.org/10.1021/pr700658q. PubMed DOI

Cox J, Hein MY, Luber CA, Paron I, Nagaraj N, Mann M. Accurate proteome-wide label-free quantification by delayed normalization and maximal peptide ratio extraction, termed MaxLFQ. Mol Cell Proteomics. 2014;13:2513–26. https://doi.org/10.1074/mcp.M113.031591. PubMed DOI PMC

Mezera V, Endlicher R, Kucera O, Sobotka O, Drahota Z, Cervinkova Z. Effects of Epigallocatechin Gallate on Tert-Butyl Hydroperoxide-Induced Mitochondrial Dysfunction in Rat Liver Mitochondria and Hepatocytes. Oxid Med Cell Longev. 2016;2016:7573131. https://doi.org/10.1155/2016/7573131. PubMed DOI PMC

Voss NR, Gerstein M. 3V: cavity, channel and cleft volume calculator and extractor. Nucleic Acids Res. 2010;38:W555–62. https://doi.org/10.1093/nar/gkq395. PubMed DOI PMC

Cerny J, Jurecka P, Hobza P, Valdes H. Resolution of identity density functional theory augmented with an empirical dispersion term (RI-DFT-D): a promising tool for studying isolated small peptides. J Phys Chem A. 2007;111:1146–54. https://doi.org/10.1021/jp066504m. PubMed DOI

Tao J, Perdew JP, Staroverov VN, Scuseria GE. Climbing the density functional ladder: nonempirical meta-generalized gradient approximation designed for molecules and solids. Phys Rev Lett. 2003;91:146401. https://doi.org/10.1103/PhysRevLett.91.146401. PubMed DOI

Sanner MF, Duncan BS, Carrillo CJ, Olson AJ. Integrating computation and visualization for biomolecular analysis: an example using python and AVS. Pac Symp Biocomput. 1999:401–12. PubMed

Trott O, Olson AJ. AutoDock Vina: improving the speed and accuracy of docking with a new scoring function, efficient optimization, and multithreading. J Comput Chem. 2010;31:455–61. https://doi.org/10.1002/jcc.21334. PubMed DOI PMC

Targeting ERK-Hippo Interplay in Cancer Therapy

The MEK-ERK-MST1 Axis Potentiates the Activation of the Extrinsic Apoptotic Pathway during GDC-0941 Treatment in Jurkat T Cells