Skip to main content

Thank you for visiting nature.com. You are using a browser version with limited support for CSS. To obtain the best experience, we recommend you use a more up to date browser (or turn off compatibility mode in Internet Explorer). In the meantime, to ensure continued support, we are displaying the site without styles and JavaScript.

  • Article
  • Published:

Metabolic plasticity underpins innate and acquired resistance to LDHA inhibition

Nature Chemical Biology volume 12, pages 779–786 (2016)Cite this article

Subjects

Abstract

Metabolic reprogramming in tumors represents a potential therapeutic target. Herein we used shRNA depletion and a novel lactate dehydrogenase (LDHA) inhibitor, GNE-140, to probe the role of LDHA in tumor growth in vitro and in vivo. In MIA PaCa-2 human pancreatic cells, LDHA inhibition rapidly affected global metabolism, although cell death only occurred after 2 d of continuous LDHA inhibition. Pancreatic cell lines that utilize oxidative phosphorylation (OXPHOS) rather than glycolysis were inherently resistant to GNE-140, but could be resensitized to GNE-140 with the OXPHOS inhibitor phenformin. Acquired resistance to GNE-140 was driven by activation of the AMPK–mTOR–S6K signaling pathway, which led to increased OXPHOS, and inhibitors targeting this pathway could prevent resistance. Thus, combining an LDHA inhibitor with compounds targeting the mitochondrial or AMPK–S6K signaling axis may not only broaden the clinical utility of LDHA inhibitors beyond glycolytically dependent tumors but also reduce the emergence of resistance to LDHA inhibition.

This is a preview of subscription content, access via your institution

Access options

Buy this article

  • Purchase on Springer Link
  • Instant access to full article PDF
Buy now

Prices may be subject to local taxes which are calculated during checkout

Figure 1: Inhibition of LDHA in MIA PaCa-2 cells perturbs glycolysis and inhibits proliferation.
Figure 2: Inhibition of LDHA in MIA PaCa-2 cells with GNE-140 results in modulation of metabolites associated with glycolysis and the pentose phosphate pathway.
Figure 3: Pancreatic cell lines that are sensitive to GNE-140 rely on glycolysis for energy generation.
Figure 4: Acquired resistance to GNE-140 results in increased dependency on OXPHOS.
Figure 5: Acquired resistance to GNE-140 is mediated by AMPK–S6K signaling.
Figure 6: Characterization of in vivo activity of GNE-140.

Similar content being viewed by others

Accession codes

Primary accessions

Protein Data Bank

Referenced accessions

Protein Data Bank

References

  1. Brand, K.A. & Hermfisse, U. Aerobic glycolysis by proliferating cells: a protective strategy against reactive oxygen species. FASEB J. 11, 388–395 (1997).

    Article  CAS  Google Scholar 

  2. Koukourakis, M.I. et al. Lactate dehydrogenase 5 isoenzyme overexpression defines resistance of prostate cancer to radiotherapy. Br. J. Cancer 110, 2217–2223 (2014).

    Article  CAS  Google Scholar 

  3. Koukourakis, M.I., Giatromanolaki, A. & Sivridis, E. Lactate dehydrogenase isoenzymes 1 and 5: differential expression by neoplastic and stromal cells in non-small cell lung cancer and other epithelial malignant tumors. Tumour Biol. 24, 199–202 (2003).

    Article  CAS  Google Scholar 

  4. Koukourakis, M.I., Giatromanolaki, A., Sivridis, E., Gatter, K.C. & Harris, A.L. Tumour Angiogenesis Research Group. Lactate dehydrogenase 5 expression in operable colorectal cancer: strong association with survival and activated vascular endothelial growth factor pathway—a report of the Tumour Angiogenesis Research Group. J. Clin. Oncol. 24, 4301–4308 (2006).

    Article  CAS  Google Scholar 

  5. Girgis, H. et al. Lactate dehydrogenase A is a potential prognostic marker in clear cell renal cell carcinoma. Mol. Cancer 13, 101 (2014).

    Article  Google Scholar 

  6. Shim, H. et al. c-Myc transactivation of LDH-A: implications for tumor metabolism and growth. Proc. Natl. Acad. Sci. USA 94, 6658–6663 (1997).

    Article  CAS  Google Scholar 

  7. Semenza, G.L. et al. Hypoxia response elements in the aldolase A, enolase 1, and lactate dehydrogenase A gene promoters contain essential binding sites for hypoxia-inducible factor 1. J. Biol. Chem. 271, 32529–32537 (1996).

    Article  CAS  Google Scholar 

  8. Fantin, V.R., St-Pierre, J. & Leder, P. Attenuation of LDH-A expression uncovers a link between glycolysis, mitochondrial physiology, and tumor maintenance. Cancer Cell 9, 425–434 (2006).

    Article  CAS  Google Scholar 

  9. Qing, G. et al. Combinatorial regulation of neuroblastoma tumor progression by N-Myc and hypoxia inducible factor HIF-1alpha. Cancer Res. 70, 10351–10361 (2010).

    Article  CAS  Google Scholar 

  10. Seth, P. et al. On-target inhibition of tumor fermentative glycolysis as visualized by hyperpolarized pyruvate. Neoplasia 13, 60–71 (2011).

    Article  CAS  Google Scholar 

  11. Xie, H. et al. Targeting lactate dehydrogenase-A inhibits tumorigenesis and tumor progression in mouse models of lung cancer and impacts tumor-initiating cells. Cell Metab. 19, 795–809 (2014).

    Article  CAS  Google Scholar 

  12. Novoa, W.B., Winer, A.D., Glaid, A.J. & Schwert, G.W. Lactic dehydrogenase. V. Inhibition by oxamate and by oxalate. J. Biol. Chem. 234, 1143–1148 (1959).

    CAS  PubMed  Google Scholar 

  13. Busch, H. & Nair, P.V. Inhibition of lactic acid dehydrogenase by fluoropyruvic acid. J. Biol. Chem. 229, 377–387 (1957).

    CAS  PubMed  Google Scholar 

  14. Billiard, J. et al. Quinoline 3-sulfonamides inhibit lactate dehydrogenase A and reverse aerobic glycolysis in cancer cells. Cancer Metab. 1, 19 (2013).

    Article  Google Scholar 

  15. Dragovich, P.S. et al. Identification of substituted 2-thio-6-oxo-1,6-dihydropyrimidines as inhibitors of human lactate dehydrogenase. Bioorg. Med. Chem. Lett. 23, 3186–3194 (2013).

    Article  CAS  Google Scholar 

  16. Fauber, B.P. et al. Identification of 2-amino-5-aryl-pyrazines as inhibitors of human lactate dehydrogenase. Bioorg. Med. Chem. Lett. 23, 5533–5539 (2013).

    Article  CAS  Google Scholar 

  17. Granchi, C. et al. N-Hydroxyindole-based inhibitors of lactate dehydrogenase against cancer cell proliferation. Eur. J. Med. Chem. 46, 5398–5407 (2011).

    Article  CAS  Google Scholar 

  18. Le, A. et al. Inhibition of lactate dehydrogenase A induces oxidative stress and inhibits tumor progression. Proc. Natl. Acad. Sci. USA 107, 2037–2042 (2010).

    Article  CAS  Google Scholar 

  19. Ward, R.A. et al. Design and synthesis of novel lactate dehydrogenase A inhibitors by fragment-based lead generation. J. Med. Chem. 55, 3285–3306 (2012).

    Article  CAS  Google Scholar 

  20. Vanderporten, E. et al. Label-free high-throughput assays to screen and characterize novel lactate dehydrogenase inhibitors. Anal. Biochem. 441, 115–122 (2013).

    Article  CAS  Google Scholar 

  21. Dragovich, P.S. et al. Identification of substituted 3-hydroxy-2-mercaptocyclohex-2-enones as potent inhibitors of human lactate dehydrogenase. Bioorg. Med. Chem. Lett. 24, 3764–3771 (2014).

    Article  CAS  Google Scholar 

  22. Fauber, B.P. et al. Identification of 3,6-disubstituted dihydropyrones as inhibitors of human lactate dehydrogenase. Bioorg. Med. Chem. Lett. 24, 5683–5687 (2014).

    Article  CAS  Google Scholar 

  23. Aithal, H.N., Walsh-Reitz, M.M. & Toback, F.G. Regulation of glyceraldehyde-3-phosphate dehydrogenase by a cytosolic protein. Am. J. Physiol. 249, C111–C116 (1985).

    Article  CAS  Google Scholar 

  24. Copeland, L. & Zammit, A. Kinetic properties of NAD-dependent glyceraldehyde-3-phosphate dehydrogenase from the host fraction of soybean root nodules. Arch. Biochem. Biophys. 312, 107–113 (1994).

    Article  CAS  Google Scholar 

  25. Owen, M.R., Doran, E. & Halestrap, A.P. Evidence that metformin exerts its anti-diabetic effects through inhibition of complex 1 of the mitochondrial respiratory chain. Biochem. J. 348, 607–614 (2000).

    Article  CAS  Google Scholar 

  26. Shitara, Y. et al. Role of organic cation/carnitine transporter 1 in uptake of phenformin and inhibitory effect on complex I respiration in mitochondria. Toxicol. Sci. 132, 32–42 (2013).

    Article  CAS  Google Scholar 

  27. Borisy, A.A. et al. Systematic discovery of multicomponent therapeutics. Proc. Natl. Acad. Sci. USA 100, 7977–7982 (2003).

    Article  CAS  Google Scholar 

  28. Hardie, D.G. AMPK: positive and negative regulation, and its role in whole-body energy homeostasis. Curr. Opin. Cell Biol. 33, 1–7 (2015).

    Article  CAS  Google Scholar 

  29. Shirwany, N.A. & Zou, M.H. AMPK: a cellular metabolic and redox sensor. A minireview. Front. Biosci. (Landmark Ed.) 19, 447–474 (2014).

    Article  CAS  Google Scholar 

  30. Inoki, K., Zhu, T. & Guan, K.L. TSC2 mediates cellular energy response to control cell growth and survival. Cell 115, 577–590 (2003).

    Article  CAS  Google Scholar 

  31. Gwinn, D.M. et al. AMPK phosphorylation of raptor mediates a metabolic checkpoint. Mol. Cell 30, 214–226 (2008).

    Article  CAS  Google Scholar 

  32. Pearce, L.R. et al. Characterization of PF-4708671, a novel and highly specific inhibitor of p70 ribosomal S6 kinase (S6K1). Biochem. J. 431, 245–255 (2010).

    Article  CAS  Google Scholar 

  33. Ma, X.M. & Blenis, J. Molecular mechanisms of mTOR-mediated translational control. Nat. Rev. Mol. Cell Biol. 10, 307–318 (2009).

    Article  Google Scholar 

  34. Magnuson, B., Ekim, B. & Fingar, D.C. Regulation and function of ribosomal protein S6 kinase (S6K) within mTOR signalling networks. Biochem. J. 441, 1–21 (2012).

    Article  CAS  Google Scholar 

  35. Feng, Z. & Levine, A.J. The regulation of energy metabolism and the IGF-1/mTOR pathways by the p53 protein. Trends Cell Biol. 20, 427–434 (2010).

    Article  CAS  Google Scholar 

  36. Viale, A. et al. Oncogene ablation-resistant pancreatic cancer cells depend on mitochondrial function. Nature 514, 628–632 (2014).

    Article  CAS  Google Scholar 

  37. Birsoy, K. et al. An essential role of the mitochondrial electron transport chain in cell proliferation is to enable aspartate synthesis. Cell 162, 540–551 (2015).

    Article  CAS  Google Scholar 

  38. Chen, J. et al. Piperidine-dione derivatives. World Intellectual Property Organization patent WO 2015/140133 A1 (2015).

  39. Reitman, Z.J. et al. Profiling the effects of isocitrate dehydrogenase 1 and 2 mutations on the cellular metabolome. Proc. Natl. Acad. Sci. USA 108, 3270–3275 (2011).

    Article  CAS  Google Scholar 

  40. Gray, D.C. et al. pHUSH: a single vector system for conditional gene expression. BMC Biotechnol. 7, 61 (2007).

    Article  Google Scholar 

Download references

Acknowledgements

We would like to thank the Discovery Chemistry Small Molecule analytical group for their support; the in vivo cell culture and dosing groups for their support of in vivo studies; R. Bourgon for bioinformatic assistance; the Genentech gCell group (R. Neve, M. Yu and M. Liang-Chu) for cell line banking and maintenance; and members of Genentech gCSI group (E. Lin, B. Lam, Y. Yu and J. Tan) for assistance on cell-based drug screens; S. Sriraman for assistance with the 18FDG-uptake assays; and A. Bruce for assistance with generating figures.

Author information

Author notes

  1. Peter K Jackson

    Present address: Present address: Department of Microbiology and Immunology, Stanford University, Stanford, California, USA.,

  2. Aaron Boudreau and Hans E Purkey: These authors contributed equally to this work.

Authors and Affiliations

  1. Discovery Oncology, Genentech, South San Francisco, California, USA

    Aaron Boudreau, David Peterson, Raju Pusapati & Marie Evangelista

  2. Discovery Chemistry, Genentech, South San Francisco, California, USA

    Hans E Purkey, Kirk Robarge, Sharada Labadie, Aihe Zhou & Binqing Wei

  3. Translational Oncology, Genentech, South San Francisco, California, USA

    Anna Hitz, Mandy Kwong, Rebecca Hong, Min Gao, Christopher Del Nagro, Laura Corson, Marcia Belvin, Peter K Jackson, Georgia Hatzivassiliou, Deepak Sampath & Thomas O'Brien

  4. Drug Metabolism and Pharmacokinetics, Genentech, South San Francisco, California, USA

    Shuguang Ma, Laurent Salphati & Jodie Pang

  5. Chemistry, WuXi AppTec Co., Ltd., Shanghai, China

    Tommy Lai

  6. Structural Biology, WuXi AppTec Co., Ltd., Shanghai, China

    Yingjie Li & Zhongguo Chen

  7. Biochemical and Cellular Pharmacology, Genentech, South San Francisco, California, USA

    Ivana Yen, Steve Sideris & Shiva Malek

  8. Department of Pathology, Genentech, South San Francisco, California, USA.,

    Mark McCleland & Ron Firestein

  9. Biomedical Imaging, Genentech, South San Francisco, California, USA

    Alex Vanderbilt & Simon Williams

  10. Bioinformatics, Genentech, South San Francisco, California, USA

    Anneleen Daemen

  11. Structural Biology, Genentech, South San Francisco, California, USA

    Charles Eigenbrot

Authors
  1. Aaron Boudreau
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. Hans E Purkey
    View author publications

    You can also search for this author in PubMed Google Scholar

  3. Anna Hitz
    View author publications

    You can also search for this author in PubMed Google Scholar

  4. Kirk Robarge
    View author publications

    You can also search for this author in PubMed Google Scholar

  5. David Peterson
    View author publications

    You can also search for this author in PubMed Google Scholar

  6. Sharada Labadie
    View author publications

    You can also search for this author in PubMed Google Scholar

  7. Mandy Kwong
    View author publications

    You can also search for this author in PubMed Google Scholar

  8. Rebecca Hong
    View author publications

    You can also search for this author in PubMed Google Scholar

  9. Min Gao
    View author publications

    You can also search for this author in PubMed Google Scholar

  10. Christopher Del Nagro
    View author publications

    You can also search for this author in PubMed Google Scholar

  11. Raju Pusapati
    View author publications

    You can also search for this author in PubMed Google Scholar

  12. Shuguang Ma
    View author publications

    You can also search for this author in PubMed Google Scholar

  13. Laurent Salphati
    View author publications

    You can also search for this author in PubMed Google Scholar

  14. Jodie Pang
    View author publications

    You can also search for this author in PubMed Google Scholar

  15. Aihe Zhou
    View author publications

    You can also search for this author in PubMed Google Scholar

  16. Tommy Lai
    View author publications

    You can also search for this author in PubMed Google Scholar

  17. Yingjie Li
    View author publications

    You can also search for this author in PubMed Google Scholar

  18. Zhongguo Chen
    View author publications

    You can also search for this author in PubMed Google Scholar

  19. Binqing Wei
    View author publications

    You can also search for this author in PubMed Google Scholar

  20. Ivana Yen
    View author publications

    You can also search for this author in PubMed Google Scholar

  21. Steve Sideris
    View author publications

    You can also search for this author in PubMed Google Scholar

  22. Mark McCleland
    View author publications

    You can also search for this author in PubMed Google Scholar

  23. Ron Firestein
    View author publications

    You can also search for this author in PubMed Google Scholar

  24. Laura Corson
    View author publications

    You can also search for this author in PubMed Google Scholar

  25. Alex Vanderbilt
    View author publications

    You can also search for this author in PubMed Google Scholar

  26. Simon Williams
    View author publications

    You can also search for this author in PubMed Google Scholar

  27. Anneleen Daemen
    View author publications

    You can also search for this author in PubMed Google Scholar

  28. Marcia Belvin
    View author publications

    You can also search for this author in PubMed Google Scholar

  29. Charles Eigenbrot
    View author publications

    You can also search for this author in PubMed Google Scholar

  30. Peter K Jackson
    View author publications

    You can also search for this author in PubMed Google Scholar

  31. Shiva Malek
    View author publications

    You can also search for this author in PubMed Google Scholar

  32. Georgia Hatzivassiliou
    View author publications

    You can also search for this author in PubMed Google Scholar

  33. Deepak Sampath
    View author publications

    You can also search for this author in PubMed Google Scholar

  34. Marie Evangelista
    View author publications

    You can also search for this author in PubMed Google Scholar

  35. Thomas O'Brien
    View author publications

    You can also search for this author in PubMed Google Scholar

Contributions

A.B. contributed in the generation of, and characterized, resistant cell lines and to manuscript writing; H.E.P. and K.R. designed and directed compound generation and characterization of lead molecules and assisted in generating co-crystal structure; A.H. contributed through the generation and characterization shLDHA cell lines and by performing metabolic profiling experiments; D.P. characterized the induction of ROS, performed cell line assays and characterized lactate levels in cell lines; S.L. designed and assisted in compound generation and characterization of lead molecules; M.K. and M.G. performed all MS-based assays; R.H. designed and carried out in vivo studies; C.D.N. performed cell cycle analysis of cells; R.P. assisted in understanding the mechanism of resistance; M.M. and R.F. assisted with knockdown construct design and characterization; S. Ma, L.S. and J.P. designed and carried out pharmacokinetic studies in mice; A.Z. and T.L. designed and assisted in compound generation and characterization of lead molecules; Y.L. and Z.C. obtained a co-crystal structure; B.W. designed molecules and assisted in obtaining a co-crystal structure; I.Y., S.S. and S. Malek designed and performed biochemical and selectivity assays; L.C. designed and generated shLDHA cells; A.V. and S.W. performed cellular glucose uptake assays; A.D. assisted in analysis of metabolite profiling in cells; M.B. performed characterization of lead molecule in cells; C.E. directed protein production and generation of a co-crystal structure; P.K.J. directed the design and characterization of shLDHA cells; G.H. contributed intellectually to the design of all metabolic assays and helped draft the manuscript; D.S. directed in vivo studies and helped draft the manuscript; M.E. directed the generation and characterization of resistant clones, as well as cell line screening and in vivo studies; T.O'B. directed the characterization of the role of LDHA in cells and helped design and direct metabolic and cell cycle and in vivo studies; M.E. and T.O'B. drafted and revised the manuscript for intellectual content and final approval.

Corresponding authors

Correspondence to Marie Evangelista or Thomas O'Brien.

Ethics declarations

Competing interests

All authors were full-time employees of either Genentech or WuXi at the time these studies were conducted.

Supplementary information

Supplementary Text and Figures

Supplementary Results, Supplementary Tables 1–3 and Supplementary Figures 1–9. (PDF 15669 kb)

Supplementary Dataset 1

Complete list of the fold change of all metabolites detected in MIA PaCa-2 cells following treatment with 2 μM GNE-140. (XLSX 61 kb)

Rights and permissions

Reprints and permissions

About this article

Check for updates. Verify currency and authenticity via CrossMark

Cite this article

Boudreau, A., Purkey, H., Hitz, A. et al. Metabolic plasticity underpins innate and acquired resistance to LDHA inhibition. Nat Chem Biol 12, 779–786 (2016). https://doi.org/10.1038/nchembio.2143

Download citation

  • Received:

  • Accepted:

  • Published:

  • Issue Date:

  • DOI: https://doi.org/10.1038/nchembio.2143

This article is cited by

Search

Advanced search

Quick links

Nature Briefing: Cancer

Sign up for the Nature Briefing: Cancer newsletter — what matters in cancer research, free to your inbox weekly.

Get what matters in cancer research, free to your inbox weekly. Sign up for Nature Briefing: Cancer