Abstract
Hypoxia can promote invasive behavior in cancer cells and alters the response to therapeutic intervention as a result of changes in the expression many genes, including genes involved in intermediary metabolism. Although metabolomics technologies are capable of simultaneously measuring a wide range of metabolites in an untargeted manner, these methods have been relatively under utilized in the study of cancer cell responses to hypoxia. Thus, 1H NMR metabolomics was used to examine the effects of hypoxia in the MDA-MB-231 human breast cancer cell line, both in vitro and in vivo. Cell cultures were compared with respect to their metabolic responses during growth under either hypoxic (1% O2) or normoxic conditions. Orthogonal partial least squares discriminant analysis (OPLS-DA) was used to identify a set of metabolites that were responsive to hypoxia. Via intracardiac administration, MDA-MB-231 cells were also used to generate widespread metastatic disease in immuno-compromised mice. Serum metabolite analysis was conducted to compare animals with and without a large tumor burden. Intriguingly, using a cross-plot of the OPLS loadings, both the in vitro and in vivo samples yielded a subset of metabolites that were significantly altered by hypoxia. These included primarily energy metabolites and amino acids, indicative of known alterations in energy metabolism, and possibly protein synthesis or catabolism. The results suggest that the metabolite pattern identified might prove useful as a marker for intra-tumoral hypoxia.
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Abbreviations
- HIF:
-
Hypoxia inducible factor
- MCT:
-
Monocarboxylate transporter
- MDA-MB-231-Luc2:
-
MDA-MB-231 Luciferase
- OPLS-DA:
-
Orthogonal partial least squares discriminant analysis
- PCA:
-
Principal component analysis
- VIP:
-
Variable influence on projection
References
Allen J, Davey H, Broadhurst D, Heald J, Rowland J, Oliver S, Kell D (2003) High-throughput classification of yeast mutants for functional genomics using metabolic footprinting. Nat Biotechnol 21:692–696
Bondareva A, Downey CM, Ayres F, Liu W, Boyd SK, Hallgrimsson B, Jirik FR (2009) The lysyl oxidase inhibitor, beta-aminopropionitrile, diminishes the metastatic colonization potential of circulating breast cancer cells. PLoS ONE 4:e5620
Chang D, Weljie AM, Newton J (2007) Leveraging latent information in NMR spectra for robust predictive models. Pac Symp Biocomput 12:115–126
Claudino WM, Quattrone A, Biganzoli L, Pestrin M, Bertini I, Di Leo A (2007) Metabolomics: available results, current research projects in breast cancer, and future applications. J Clin Oncol 25:2840–2846
DeBerardinis RJ, Lum JJ, Hatzivassiliou G, Thompson CB (2008) The biology of cancer: metabolic reprogramming fuels cell growth and proliferation. Cell Metab 7:11–20
Dowlatabadi R, Weljie AM, Thorpe TA, Yeung EC, Vogel HJ (2009) Metabolic footprinting study of white spruce somatic embryogenesis using NMR spectroscopy. Plant Physiol Biochem 47:343–350
Eisner R, Stretch C, Eastman T, Xia J, Hau D (2010) Learning to predict cancer-associated skeletal muscle wasting from 1H-NMR profiles of urinary metabolites. Metabolomics (in press)
Erler JT, Bennewith KL, Nicolau M, Dornhöfer N, Kong C, Le Q, Chi JA, Jeffrey SS, Giaccia AJ (2006) Lysyl oxidase is essential for hypoxia-induced metastasis. Nature 440:1222–1226
Gao H, Lu Q, Liu X, Cong H, Zhao L, Wang H, Lin D (2009) Application of 1H NMR-based metabonomics in the study of metabolic profiling of human hepatocellular carcinoma and liver cirrhosis. Cancer Sci 100:782–785
Griffiths JR, McSheehy PM, Robinson SP, Troy H, Chung YL, Leek RD, Williams KJ, Stratford IJ, Harris AL, Stubbs M (2002) Metabolic changes detected by in vivo magnetic resonance studies of HEPA-1 wild-type tumors and tumors deficient in hypoxia-inducible factor-1beta (HIF-1beta): evidence of an anabolic role for the HIF-1 pathway. Cancer Res 62:688–695
Landaas S (1975) The formation of 2-hydroxybutyric acid in experimental animals. Clin Chim Acta 58:23–32
Lindon JC, Nicholson JK, Holmes E, Antti H et al (2003) Contemporary issues in toxicology the role of metabonomics in toxicology and its evaluation by the COMET project. Toxicol Appl Pharmacol 187:137–146
Perrin A, Roudier E, Duborjal H, Bachelet C, Riva-Lavieille C, Leverve X, Massarelli R (2002) Pyruvate reverses metabolic effects produced by hypoxia in glioma and hepatoma cell cultures. Biochimie 84:1003–1011
Schofield CJ, Ratcliffe PJ (2004) Oxygen sensing by HIF hydroxylases. Nat Rev Mol Cell Biol 5:343–354
Sonveaux P, Végran F, Schroeder T, Wergin MC et al (2008) Targeting lactate-fueled respiration selectively kills hypoxic tumor cells in mice. J Clin Invest 118:3930–3942
Ullah MS, Davies AJ, Halestrap AP (2006) The plasma membrane lactate transporter MCT4, but not MCT1, is up-regulated by hypoxia through a HIF-1alpha-dependent mechanism. J Biol Chem 281:9030–9037
Walenta S, Snyder S, Haroon ZA, Braun RD, Amin K, Brizel D, Mueller-Klieser W, Chance B, Dewhirst MW (2001) Tissue gradients of energy metabolites mirror oxygen tension gradients in a rat mammary carcinoma model. Int J Radiat Oncol Biol Phys 51:840–848
Warburg O (1956) On the origin of cancer cells. Science 123:309–314
Webb JD, Coleman ML, Pugh CW (2009) Hypoxia, hypoxia-inducible factors (HIF), HIF hydroxylases and oxygen sensing. Cell Mol Life Sci 66:3539–3554
Weljie A, Dowlatabadi R, Miller B, Vogel H, Jirik F (2007) An inflammatory arthritis-associated metabolite biomarker pattern revealed by 1H NMR spectroscopy. J Proteome Res 6:3456–3464
Weljie AM, Jirik FR (2010) Hypoxia-induced metabolic shifts in cancer cells: moving beyond the Warburg effect. Int J Biochem cell biol
Weljie AM, Newton J, Mercier P, Carlson E, Slupsky CM (2006) Targeted profiling: quantitative analysis of 1H NMR metabolomics data. Anal Chem 78:4430–4442
Weljie AM, Newton J, Jirik FR, Vogel HJ (2008) Evaluating low-intensity unknown signals in quantitative proton NMR mixture analysis. Anal Chem 80:8956–8965
Wigfield SM, Winter SC, Giatromanolaki A, Taylor J, Koukourakis ML, Harris AL (2008) PDK-1 regulates lactate production in hypoxia and is associated with poor prognosis in head and neck squamous cancer. Br J Cancer 98:1975–1984
Wiklund S, Johansson E, Sjöström L, Mellerowicz EJ, Edlund U, Shockcor JP, Gottfries J, Moritz T, Trygg J (2008) Visualization of GC/TOF-MS-based metabolomics data for identification of biochemically interesting compounds using OPLS class models. Anal Chem 80:115–122
Wishart DS, Tzur D, Knox C, Eisner R et al (2007) HMDB: the Human Metabolome Database. Nucleic Acids Res 35:D521–D526
Wouters B, Koritzinsky M (2008) Hypoxia signalling through mTOR and the unfolded protein response in cancer. Nat Rev Cancer 8:851–864
Zhang H, Bosch-Marce M, Shimoda LA, Tan YS, Baek JH, Wesley JB, Gonzalez FJ, Semenza GL (2008) Mitochondrial autophagy is an HIF-1-dependent adaptive metabolic response to hypoxia. J Biol Chem 283:10892–10903
Acknowledgments
The Metabolomics Research Centre at the University of Calgary is supported by funding from Alberta Health Services (AHS)/The Alberta Cancer Foundation (ACF). The studies were also supported in part by a translational grant from the AHS/ACF (to F.R.J.); F.R.J was the recipient of a Canada Research Chairs award.
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Weljie, A.M., Bondareva, A., Zang, P. et al. 1H NMR metabolomics identification of markers of hypoxia-induced metabolic shifts in a breast cancer model system. J Biomol NMR 49, 185–193 (2011). https://doi.org/10.1007/s10858-011-9486-4
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DOI: https://doi.org/10.1007/s10858-011-9486-4