Abstract
Cancer cells engage in a metabolic program to enhance biosynthesis and support cell proliferation. The regulatory properties of pyruvate kinase M2 (PKM2) influence altered glucose metabolism in cancer. The interaction of PKM2 with phosphotyrosine-containing proteins inhibits enzyme activity and increases the availability of glycolytic metabolites to support cell proliferation. This suggests that high pyruvate kinase activity may suppress tumor growth. We show that expression of PKM1, the pyruvate kinase isoform with high constitutive activity, or exposure to published small-molecule PKM2 activators inhibits the growth of xenograft tumors. Structural studies reveal that small-molecule activators bind PKM2 at the subunit interaction interface, a site that is distinct from that of the endogenous activator fructose-1,6-bisphosphate (FBP). However, unlike FBP, binding of activators to PKM2 promotes a constitutively active enzyme state that is resistant to inhibition by tyrosine-phosphorylated proteins. These data support the notion that small-molecule activation of PKM2 can interfere with anabolic metabolism.
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Change history
11 October 2012
In the version of this article initially published, the accession codes to the Protein Data Bank were omitted. The error has been corrected in the HTML and PDF versions of the article.
References
Tennant, D.A., Duran, R.V. & Gottlieb, E. Targeting metabolic transformation for cancer therapy. Nat. Rev. Cancer 10, 267–277 (2010).
Vander Heiden, M.G. Targeting cancer metabolism: a therapeutic window opens. Nat. Rev. Drug Discov. 10, 671–684 (2011).
Vander Heiden, M.G., Cantley, L.C. & Thompson, C.B. Understanding the Warburg effect: the metabolic requirements of cell proliferation. Science 324, 1029–1033 (2009).
Cairns, R.A., Harris, I.S. & Mak, T.W. Regulation of cancer cell metabolism. Nat. Rev. Cancer 11, 85–95 (2011).
Levine, A.J. & Puzio-Kuter, A.M. The control of the metabolic switch in cancers by oncogenes and tumor suppressor genes. Science 330, 1340–1344 (2010).
Trachootham, D., Alexandre, J. & Huang, P. Targeting cancer cells by ROS-mediated mechanisms: a radical therapeutic approach? Nat. Rev. Drug Discov. 8, 579–591 (2009).
Weissleder, R. Molecular imaging in cancer. Science 312, 1168–1171 (2006).
Christofk, H.R. et al. The M2 splice isoform of pyruvate kinase is important for cancer metabolism and tumour growth. Nature 452, 230–233 (2008).
Mazurek, S. Pyruvate kinase type M2: a key regulator of the metabolic budget system in tumor cells. Int. J. Biochem. Cell Biol. 43, 969–980 (2011).
Noguchi, T., Inoue, H. & Tanaka, T. The M1- and M2-type isozymes of rat pyruvate kinase are produced from the same gene by alternative RNA splicing. J. Biol. Chem. 261, 13807–13812 (1986).
Clower, C.V. et al. The alternative splicing repressors hnRNP A1/A2 and PTB influence pyruvate kinase isoform expression and cell metabolism. Proc. Natl. Acad. Sci. USA 107, 1894–1899 (2010).
Yamada, K. & Noguchi, T. Regulation of pyruvate kinase M gene expression. Biochem. Biophys. Res. Commun. 256, 257–262 (1999).
Ikeda, Y., Tanaka, T. & Noguchi, T. Conversion of non-allosteric pyruvate kinase isozyme into an allosteric enzyme by a single amino acid substitution. J. Biol. Chem. 272, 20495–20501 (1997).
Ikeda, Y. & Noguchi, T. Allosteric regulation of pyruvate kinase M2 isozyme involves a cysteine residue in the intersubunit contact. J. Biol. Chem. 273, 12227–12233 (1998).
Ashizawa, K., Willingham, M.C., Liang, C.M. & Cheng, S.Y. In vivo regulation of monomer-tetramer conversion of pyruvate kinase subtype M2 by glucose is mediated via fructose 1,6-bisphosphate. J. Biol. Chem. 266, 16842–16846 (1991).
Ashizawa, K., McPhie, P., Lin, K.H. & Cheng, S.Y. An in vitro novel mechanism of regulating the activity of pyruvate kinase M2 by thyroid hormone and fructose 1, 6-bisphosphate. Biochemistry 30, 7105–7111 (1991).
Christofk, H.R., Vander Heiden, M.G., Wu, N., Asara, J.M. & Cantley, L.C. Pyruvate kinase M2 is a phosphotyrosine-binding protein. Nature 452, 181–186 (2008).
Eigenbrodt, E., Reinacher, M., Scheefers-Borchel, U., Scheefers, H. & Friis, R. Double role for pyruvate kinase type M2 in the expansion of phosphometabolite pools found in tumor cells. Crit. Rev. Oncog. 3, 91–115 (1992).
Anastasiou, D. et al. Inhibition of pyruvate kinase M2 by reactive oxygen species contributes to cellular antioxidant responses. Science 334, 1278–1283 (2011).
Vander Heiden, M.G. et al. Evidence for an alternative glycolytic pathway in rapidly proliferating cells. Science 329, 1492–1499 (2010).
Locasale, J.W., Vander Heiden, M.G. & Cantley, L.C. Rewiring of glycolysis in cancer cell metabolism. Cell Cycle 9, 4253 (2010).
Jiang, J.K. et al. Evaluation of thieno[3,2-b]pyrrole[3,2-d]pyridazinones as activators of the tumor cell specific M2 isoform of pyruvate kinase. Bioorg. Med. Chem. Lett. 20, 3387–3393 (2010).
Boxer, M.B. et al. Evaluation of substituted N,N′-diarylsulfonamides as activators of the tumor cell specific M2 isoform of pyruvate kinase. J. Med. Chem. 53, 1048–1055 (2010).
Ikeda, Y., Taniguchi, N. & Noguchi, T. Dominant negative role of the glutamic acid residue conserved in the pyruvate kinase M(1) isozyme in the heterotropic allosteric effect involving fructose-1,6-bisphosphate. J. Biol. Chem. 275, 9150–9156 (2000).
Kato, H., Fukuda, T., Parkison, C., McPhie, P. & Cheng, S.Y. Cytosolic thyroid hormone-binding protein is a monomer of pyruvate kinase. Proc. Natl. Acad. Sci. USA 86, 7861–7865 (1989).
Hitosugi, T. et al. Tyrosine phosphorylation inhibits PKM2 to promote the Warburg effect and tumor growth. Sci. Signal. 2, ra73 (2009).
Lv, L. et al. Acetylation targets the M2 isoform of pyruvate kinase for degradation through chaperone-mediated autophagy and promotes tumor growth. Mol. Cell 42, 719–730 (2011).
Luo, W. et al. Pyruvate kinase M2 is a PHD3-stimulated coactivator for hypoxia-inducible factor 1. Cell 145, 732–744 (2011).
Hoshino, A., Hirst, J.A. & Fujii, H. Regulation of cell proliferation by interleukin-3–induced nuclear translocation of pyruvate kinase. J. Biol. Chem. 282, 17706–17711 (2007).
Steták, A. et al. Nuclear translocation of the tumor marker pyruvate kinase M2 induces programmed cell death. Cancer Res. 67, 1602–1608 (2007).
Hatzivassiliou, G. et al. ATP citrate lyase inhibition can suppress tumor cell growth. Cancer Cell 8, 311–321 (2005).
Vander Heiden, M.G. et al. Identification of small molecule inhibitors of pyruvate kinase M2. Biochem. Pharmacol. 79, 1118–1124 (2010).
Otwinowski, Z. & Minor, W. Processing of X-ray diffraction data collected in oscillation mode. in Methods in Enzymology Vol. 276, 307–326 (Academic Press, 1997).
Minor, W., Cymborowski, M., Otwinowski, Z. & Chruszcz, M. HKL-3000: the integration of data reduction and structure solution—from diffraction images to an initial model in minutes. Acta Crystallogr. D Biol. Crystallogr. 62, 859–866 (2006).
Berman, H.M. et al. The Protein Data Bank. Nucleic Acids Res. 28, 235–242 (2000).
Schüttelkopf, A.W. & van Aalten, D.M. PRODRG: a tool for high-throughput crystallography of protein-ligand complexes. Acta Crystallogr. D Biol. Crystallogr. 60, 1355–1363 (2004).
Emsley, P., Lohkamp, B., Scott, W.G. & Cowtan, K. Features and development of Coot. Acta Crystallogr. D Biol. Crystallogr. 66, 486–501 (2010).
Murshudov, G.N., Vagin, A.A. & Dodson, E.J. Refinement of macromolecular structures by the maximum-likelihood method. Acta Crystallogr. D Biol. Crystallogr. 53, 240–255 (1997).
Davis, I.W., Murray, L.W., Richardson, J.S. & Richardson, D.C. MOLPROBITY: structure validation and all-atom contact analysis for nucleic acids and their complexes. Nucleic Acids Res. 32, W615–W619 (2004).
Schumacker, P.T., Chandel, N. & Agusti, A.G. Oxygen conformance of cellular respiration in hepatocytes. Am. J. Physiol. 265, L395–L402 (1993).
Metallo, C.M. et al. Reductive glutamine metabolism by IDH1 mediates lipogenesis under hypoxia. Nature 481, 380–384 (2011).
Yuan, M., Breitkopf, S.B., Yang, X. & Asara, J.M. A positive/negative ion-switching, targeted mass spectrometry-based metabolomics platform for bodily fluids, cells, and fresh and fixed tissue. Nat. Protoc. 7, 872–881 (2012).
Xia, J., Mandal, R., Sinelnikov, I.V., Broadhurst, D. & Wishart, D.S. MetaboAnalyst 2.0–a comprehensive server for metabolomic data analysis. Nucleic Acids Res. W127–W133 (2012).
Acknowledgements
The Structural Genomics Consortium is a registered charity (1097737) and receives funds from the Canadian Institutes for Health Research, the Canadian Foundation for Innovation, Genome Canada through the Ontario Genomics Institute, GlaxoSmithKline, Karolinska Institutet, the Knut and Alice Wallenberg Foundation, the Ontario Innovation Trust, the Ontario Ministry for Research and Innovation, Merck and Co., Inc., the Novartis Research Foundation, the Swedish Agency for Innovation Systems, the Swedish Foundation for Strategic Research and the Wellcome Trust. The crystallography results shown in this report are derived from work performed at Argonne National Laboratory, Structural Biology Center at the Advanced Photon Source. Argonne is operated by UChicago Argonne, LLC for the US Department of Energy, Office of Biological and Environmental Research under contract DE-AC02-06CH11357. We thank P. Chang for experimental advice related to sucrose gradient ultracentrifugation and SAI Advantium Pharma Ltd. for help with pharmacokinetics studies. We also thank M. Kini for experimental help and acknowledge M. Yuan and S. Breitkopf for help with MS experiments. This work was supported by the Molecular Libraries Initiative of the NIH Roadmap for Medical Research and the Intramural Research Program of the National Human Genome Research Institute, NIH and by NIH grant R03MH085679. This work was also funded by NIH grant R01 GM56203 (L.C.C.). J.M.A. acknowledges funding from NIH 5P01CA120964 and Dana-Farber/Harvard Cancer Center support grant NIH 5P30CA006516. M.G.V.H. acknowledges additional funding support from the Smith Family Foundation, the Burroughs Wellcome Fund, the Damon Runyon Cancer Research Foundation, the Stern family and the National Cancer Institute, including NIH 5P30CA1405141.
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D.A., Y.Y., W.J.I. and M.G.V.H. designed and coordinated the study. M.B.B., C.J.T., L.C.C., H.-W.P. and L.D. advised on various aspects of the study. J.-K.J., M.B.B., M.S., A.P.S., H.V., N.S., M.J.W., K.R.B., W.L., C.P.A., J.I., D.S.A. and C.J.T. designed and provided compounds. B.S.H., W.T., S.D. and H.-W.P. performed all structural studies. A.J. did additional structural analysis. H.Y., C.K., K.E.Y., K.K., F.G.S., S.J. and L.D. performed in vivo pharmacology and ADME studies. C.M.M., J.M.A. and G.S. did MS. M.H.H. reviewed pathology. D.A., Y.Y., W.J.I., K.R.M., B.P.F., K.D.C., S.M., T.M.K., C.K., S.Y.L., Z.R.J., S.M.D., H.R.C. and M.G.V.H. all performed experiments. D.A. and M.G.V.H. wrote the paper with substantial input from Y.Y. and W.J.I.
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L.C.C. is a founder, M.G.V.H. is a consultant and A.J., H.Y., C.K., K.E.Y., K.K., F.G.S., S.J. and L.D are employed by Agios Pharmaceuticals, a company seeking to target metabolic enzymes for cancer therapy.
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Anastasiou, D., Yu, Y., Israelsen, W. et al. Pyruvate kinase M2 activators promote tetramer formation and suppress tumorigenesis. Nat Chem Biol 8, 839–847 (2012). https://doi.org/10.1038/nchembio.1060
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DOI: https://doi.org/10.1038/nchembio.1060
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