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:

Inhibition of human copper trafficking by a small molecule significantly attenuates cancer cell proliferation

Nature Chemistry volume 7, pages 968–979 (2015)Cite this article

Subjects

Abstract

Copper is a transition metal that plays critical roles in many life processes. Controlling the cellular concentration and trafficking of copper offers a route to disrupt these processes. Here we report small molecules that inhibit the human copper-trafficking proteins Atox1 and CCS, and so provide a selective approach to disrupt cellular copper transport. The knockdown of Atox1 and CCS or their inhibition leads to a significantly reduced proliferation of cancer cells, but not of normal cells, as well as to attenuated tumour growth in mouse models. We show that blocking copper trafficking induces cellular oxidative stress and reduces levels of cellular ATP. The reduced level of ATP results in activation of the AMP-activated protein kinase that leads to reduced lipogenesis. Both effects contribute to the inhibition of cancer cell proliferation. Our results establish copper chaperones as new targets for future developments in anticancer therapies.

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: Developing small molecules that specifically inhibit human copper-trafficking proteins and an overview of the screening process.
Figure 2: Docking model and binding of DC_AC50 to Atox1 and full-length CCS by FRET measurements (from Tyr/Trp to DC_AC50).
Figure 3: DC_AC50 reduces the proliferation of cancer cells and attenuates tumour growth in xenograft nude mice.
Figure 4: DC_AC50 induces copper accumulation, increases ROS level and decreases the NADPH/NADP+ ratio.
Figure 5: Treatment with DC_AC50 or Atox1/CCS knockdown decreases the cellular ATP level, COX activities and the rate of oxygen consumption in cancer cells.
Figure 6: DC_AC50-induced mitochondria defects and decreased lipid biosynthesis through AMPK activation.

Similar content being viewed by others

Accession codes

Accessions

Protein Data Bank

References

  1. Labbe, S. & Thiele, D. J. Pipes and wiring: the regulation of copper uptake and distribution in yeast. Trends Microbiol. 7, 500–505 (1999).

    Article  CAS  Google Scholar 

  2. Kim, B. E., Nevitt, T. & Thiele, D. J. Mechanisms for copper acquisition, distribution and regulation. Nature Chem. Biol. 4, 176–185 (2008).

    Article  CAS  Google Scholar 

  3. O'Halloran, T. V. & Culotta, V. C. Metallochaperones, an intracellular shuttle service for metal ions. J. Biol. Chem. 275, 25057–25060 (2000).

    Article  CAS  Google Scholar 

  4. Rosenzweig, A. C. Copper delivery by metallochaperone proteins. Acc. Chem. Res. 34, 119–128 (2001).

    Article  CAS  Google Scholar 

  5. Lutsenko, S., Gupta, A., Burkhead, J. L. & Zuzel, V. Cellular multitasking: the dual role of human Cu-ATPases in cofactor delivery and intracellular copper balance. Arch. Biochem. Biophys. 476, 22–32 (2008).

    Article  CAS  Google Scholar 

  6. La Fontaine, S. & Mercer, J. F. Trafficking of the copper-ATPases, ATP7A and ATP7B: role in copper homeostasis. Arch. Biochem. Biophys. 463, 149–167 (2007).

    Article  CAS  Google Scholar 

  7. Rae, T. D., Schmidt, P. J., Pufahl, R. A., Culotta, V. C. & O'Halloran, T. V. Undetectable intracellular free copper: the requirement of a copper chaperone for superoxide dismutase. Science 284, 805–808 (1999).

    Article  CAS  Google Scholar 

  8. Banci, L. et al. Human superoxide dismutase 1 (hSOD1) maturation through interaction with human copper chaperone for SOD1 (hCCS). Proc. Natl Acad. Sci. USA 109, 13555–13560 (2012).

    Article  CAS  Google Scholar 

  9. Caruano-Yzermans, A. L. et al. Mechanisms of the copper-dependent turnover of the copper chaperone for superoxide dismutase. J. Biol. Chem. 281, 13581–13587 (2006).

    Article  CAS  Google Scholar 

  10. Rae, T. D. et al. Mechanism of Cu,Zn-superoxide dismutase activation by the human metallochaperone hCCS. J. Biol. Chem. 276, 5166–5176 (2001).

    Article  CAS  Google Scholar 

  11. Lamb, A. L. et al. Crystal structure of the copper chaperone for superoxide dismutase. Nature Struct. Biol. 6, 724–729 (1999).

    Article  CAS  Google Scholar 

  12. Wernimont, A. K., Huffman, D. L., Lamb, A. L., O'Halloran, T. V. & Rosenzweig, A. C. Structural basis for copper transfer by the metallochaperone for the Menkes/Wilson disease proteins. Nature Struct. Biol. 7, 766–771 (2000).

    Article  CAS  Google Scholar 

  13. Anastassopoulou, I. et al. Solution structure of the apo and copper(I)-loaded human metallochaperone HAH1. Biochemistry 43, 13046–13053 (2004).

    Article  CAS  Google Scholar 

  14. Lamb, A. L., Torres, A. S., O'Halloran, T. V. & Rosenzweig, A. C. Heterodimeric structure of superoxide dismutase in complex with its metallochaperone. Nature Struct. Biol. 8, 751–755 (2001).

    Article  CAS  Google Scholar 

  15. Kawamata, H. & Manfredi, G. Different regulation of wild-type and mutant Cu,Zn superoxide dismutase localization in mammalian mitochondria. Hum. Mol. Genet. 17, 3303–3317 (2008).

    Article  CAS  Google Scholar 

  16. Valentine, J. S. & Hart, P. J. Misfolded CuZnSOD and amyotrophic lateral sclerosis. Proc. Natl Acad. Sci. USA 100, 3617–3622 (2003).

    Article  CAS  Google Scholar 

  17. Mandinov, L. et al. Copper chelation represses the vascular response to injury. Proc. Natl Acad. Sci. USA 100, 6700–6705 (2003).

    Article  CAS  Google Scholar 

  18. Eatock, M. M., Schatzlein, A. & Kaye, S. B. Tumour vasculature as a target for anticancer therapy. Cancer Treat. Rev. 26, 191–204 (2000).

    Article  CAS  Google Scholar 

  19. Lowndes, S. A. & Harris, A. L. The role of copper in tumour angiogenesis. J. Mammary Gland Biol. Neoplasia 10, 299–310 (2005).

    Article  Google Scholar 

  20. Folkman, J. Angiogenesis in cancer, vascular, rheumatoid and other disease. Nature Med. 1, 27–31 (1995).

    Article  CAS  Google Scholar 

  21. Gupte, A. & Mumper, R. J. Elevated copper and oxidative stress in cancer cells as a target for cancer treatment. Cancer Treat. Rev. 35, 32–46 (2009).

    Article  CAS  Google Scholar 

  22. Cai, H. & Peng, F. Knockdown of copper chaperone antioxidant-1 by RNA interference inhibits copper-stimulated proliferation of non-small cell lung carcinoma cells. Oncol. Rep. 30, 269–275 (2013).

    Article  CAS  Google Scholar 

  23. Boal, A. K. & Rosenzweig, A. C. Structural biology of copper trafficking. Chem. Rev. 109, 4760–4779 (2009).

    Article  CAS  Google Scholar 

  24. Lutsenko, S. Human copper homeostasis: a network of interconnected pathways. Curr. Opin. Chem. Biol. 14, 211–217 (2010).

    Article  CAS  Google Scholar 

  25. Wing, T. J., Makino, S., Skillman, A. G. & Kuntz, I. D. DOCK 4.0: search strategies for automated molecular docking of flexible molecule databases. J. Comput. Aided Mol. Des. 15, 411–428 (2001).

    Article  Google Scholar 

  26. Vinkenborg, J. L. et al. Genetically encoded FRET sensors to monitor intracellular Zn2+ homeostasis. Nature Methods 6, 737–740 (2009).

    Article  CAS  Google Scholar 

  27. Hamza, I. et al. The metallochaperone Atox1 plays a critical role in perinatal copper homeostasis. Proc. Natl Acad. Sci. USA 98, 6848–6852 (2001).

    Article  CAS  Google Scholar 

  28. Itoh, S. et al. Novel role of antioxidant-1 (Atox1) as a copper-dependent transcription factor involved in cell proliferation. J. Biol. Chem. 283, 9157–9167 (2008).

    Article  CAS  Google Scholar 

  29. Martinez Molina, D. et al. Monitoring drug target engagement in cells and tissues using the cellular thermal shift assay. Science 341, 84–87 (2013).

    Article  Google Scholar 

  30. Gad, H. et al. MTH1 inhibition eradicates cancer by preventing sanitation of the dNTP pool. Nature 508, 215–221 (2014).

    Article  CAS  Google Scholar 

  31. Huber, K. V. et al. Stereospecific targeting of MTH1 by (S)-crizotinib as an anticancer strategy. Nature 508, 222–227 (2014).

    Article  CAS  Google Scholar 

  32. Sena, L. A. & Chandel, N. S. Physiological roles of mitochondrial reactive oxygen species. Mol. Cell 48, 158–167 (2012).

    Article  CAS  Google Scholar 

  33. Kroemer, G. & Pouyssegur, J. Tumor cell metabolism: cancer's Achilles’ heel. Cancer Cell 13, 472–482 (2008).

    Article  CAS  Google Scholar 

  34. 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).

    Article  CAS  Google Scholar 

  35. Cairns, R. A., Harris, I. S. & Mak, T. W. Regulation of cancer cell metabolism. Nature Rev. Cancer 11, 85–95 (2011).

    Article  CAS  Google Scholar 

  36. Leary, S. C. et al. ‘Pulling the plug’ on cellular copper: the role of mitochondria in copper export. Biochim. Biophys. Acta 1793, 146–153 (2009).

    Article  CAS  Google Scholar 

  37. Sauer, S. W. et al. Severe dysfunction of respiratory chain and cholesterol metabolism in Atp7b(–/–) mice as a model for Wilson disease. Biochim. Biophys. Acta 1812, 1607–1615 (2011).

    Article  CAS  Google Scholar 

  38. Mihaylova, M. M. & Shaw, R. J. The AMPK signalling pathway coordinates cell growth, autophagy and metabolism. Nature Cell Biol. 13, 1016–1023 (2011).

    Article  CAS  Google Scholar 

  39. Hardie, D. G., Ross, F. A. & Hawley, S. A. AMPK: a nutrient and energy sensor that maintains energy homeostasis. Nature Rev. Mol. Cell Biol. 13, 251–262 (2012).

    Article  CAS  Google Scholar 

  40. Carracedo, A., Cantley, L. C. & Pandolfi, P. P. Cancer metabolism: fatty acid oxidation in the limelight. Nature Rev. Cancer 13, 227–232 (2013).

    Article  CAS  Google Scholar 

  41. Schulze, A. & Harris, A. L. How cancer metabolism is tuned for proliferation and vulnerable to disruption. Nature 491, 364–373 (2012).

    Article  CAS  Google Scholar 

  42. Trachootham, D., Alexandre, J. & Huang, P. Targeting cancer cells by ROS-mediated mechanisms: a radical therapeutic approach? Nature Rev. Drug Discov. 8, 579–591 (2009).

    Article  CAS  Google Scholar 

  43. Raj, L. et al. Selective killing of cancer cells by a small molecule targeting the stress response to ROS. Nature 475, 231–234 (2011).

    Article  CAS  Google Scholar 

  44. Gorrini, C., Harris, I. S. & Mak, T. W. Modulation of oxidative stress as an anticancer strategy. Nature Rev. Drug Discov. 12, 931–947, (2013).

    Article  CAS  Google Scholar 

  45. Brady, D. C. et al. Copper is required for oncogenic BRAF signalling and tumorigenesis. Nature 509, 492–496 (2014).

    Article  CAS  Google Scholar 

  46. Ishida, S. et al. Bioavailable copper modulates oxidative phosphorylation and growth of tumors. Proc. Natl Acad. Sci. USA 110, 19507–19512 (2013).

    Article  CAS  Google Scholar 

  47. Santini, C. et al. Advances in copper complexes as anticancer agents. Chem. Rev. 114, 815–862 (2013).

    Article  Google Scholar 

  48. Maryon, E. B. et al. Cellular glutathione plays a key role in copper uptake mediated by human copper transporter 1. Am. J. Physiol. Cell Physiol. 304, C768–C779 (2013).

    Article  CAS  Google Scholar 

  49. Morris, G. M. et al. AutoDock4 and AutoDockTools4: automated docking with selective receptor flexibility. J. Comput. Chem. 30, 2785–2791 (2009).

    Article  CAS  Google Scholar 

  50. Gong, J. et al. ChemMapper: a versatile web server for exploring pharmacology and chemical structure association based on molecular 3D similarity method. Bioinformatics 29, 1827–1829 (2013).

    Article  CAS  Google Scholar 

Download references

Acknowledgements

This work was supported by grants from the National Natural Science Foundation of China (21210003 to H.J. and C.H., and 81230076, 91313000 to H.J.), the Hi-Tech Research and Development Program of China (2012AA020302 and 2012AA01A305 to C.L.), Chinese Academy of Sciences (XDA01040305 to C.L.), National Science Foundation (CHE-1213598 to C.H.) and National Institutes of Health (CA140515 to J.C.). C.H. is supported by the Howard Hughes Medical Institute as an investigator. We thank S. F. Reichard for help with editing the manuscript.

Author information

Author notes

  1. Jing Wang, Cheng Luo, Changliang Shan and Qiancheng You: These authors contributed equally to this work

Authors and Affiliations

  1. Department of Chemistry, Department of Biochemistry and Molecule Biology, Institute for Biophysical Dynamics, The University of Chicago, 929 East 57th Street, Chicago, 60637, Illinois, USA

    Jing Wang, Qiancheng You, Dali Han, Yuxin Xie, Jason Karpus & Chuan He

  2. Drug Discovery and Design Center, State Key Laboratory of Drug Research, Shanghai Institute of Materia Medica, Chinese Academy of Sciences, Shanghai 201203, China

    Cheng Luo, Junyan Lu, Yu Zhou, Yi Wen, Shijie Chen, Shisheng Ouyang, Naixia Zhang, Hong Ding, Hong Liu & Hualiang Jiang

  3. Department of Hematology and Medical Oncology, Winship Cancer Institute of Emory, Emory University School of Medicine, Atlanta, 30322, Georgia, USA

    Changliang Shan, Shannon Elf, Jun Fan, Heebum Kang, Ruiting Lin & Jing Chen

  4. Laboratory of Chemical Biology, Eindhoven University of Technology, MB Eindhoven, 5600, Netherlands

    Jan L. Vinkenborg & Maarten Merkx

  5. Department of Molecular BioSciences, Northwestern University, Evanston, 60208, Illinois, USA

    Chihao Luan

  6. Howard Hughes Medical Institute, The University of Chicago, 929 East 57th Street, Chicago, 60637, Illinois, USA

    Jing Wang, Qiancheng You, Dali Han, Yuxin Xie, Jason Karpus & Chuan He

Authors
  1. Jing Wang
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. Cheng Luo
    View author publications

    You can also search for this author in PubMed Google Scholar

  3. Changliang Shan
    View author publications

    You can also search for this author in PubMed Google Scholar

  4. Qiancheng You
    View author publications

    You can also search for this author in PubMed Google Scholar

  5. Junyan Lu
    View author publications

    You can also search for this author in PubMed Google Scholar

  6. Shannon Elf
    View author publications

    You can also search for this author in PubMed Google Scholar

  7. Yu Zhou
    View author publications

    You can also search for this author in PubMed Google Scholar

  8. Yi Wen
    View author publications

    You can also search for this author in PubMed Google Scholar

  9. Jan L. Vinkenborg
    View author publications

    You can also search for this author in PubMed Google Scholar

  10. Jun Fan
    View author publications

    You can also search for this author in PubMed Google Scholar

  11. Heebum Kang
    View author publications

    You can also search for this author in PubMed Google Scholar

  12. Ruiting Lin
    View author publications

    You can also search for this author in PubMed Google Scholar

  13. Dali Han
    View author publications

    You can also search for this author in PubMed Google Scholar

  14. Yuxin Xie
    View author publications

    You can also search for this author in PubMed Google Scholar

  15. Jason Karpus
    View author publications

    You can also search for this author in PubMed Google Scholar

  16. Shijie Chen
    View author publications

    You can also search for this author in PubMed Google Scholar

  17. Shisheng Ouyang
    View author publications

    You can also search for this author in PubMed Google Scholar

  18. Chihao Luan
    View author publications

    You can also search for this author in PubMed Google Scholar

  19. Naixia Zhang
    View author publications

    You can also search for this author in PubMed Google Scholar

  20. Hong Ding
    View author publications

    You can also search for this author in PubMed Google Scholar

  21. Maarten Merkx
    View author publications

    You can also search for this author in PubMed Google Scholar

  22. Hong Liu
    View author publications

    You can also search for this author in PubMed Google Scholar

  23. Jing Chen
    View author publications

    You can also search for this author in PubMed Google Scholar

  24. Hualiang Jiang
    View author publications

    You can also search for this author in PubMed Google Scholar

  25. Chuan He
    View author publications

    You can also search for this author in PubMed Google Scholar

Contributions

C.H. conceived the project with H.J. and J.C., and J.W., C.Luo, C.S. and Q.Y. designed and performed most of the experiments. J.L. and S.O. performed the virtual screening and bioinformatics analysis. S.E., J.F. and H.K. assisted with the cell and mice experiments. Y.Z. and H.L. assisted in the synthesis of the compounds. J.L.V. and M.M. assisted in setting up the FRET-based compound screening, Y.W. and N.Z. conducted the NMR experimental and data analysis of DC_AC2 with Atox1. C.Luan, H.D. and S.C. performed the SPR experiments for DC_AC50 with Atox1 and CCS. C.H. and J.W. wrote the manuscript with input from H.J. and J.C.

Corresponding authors

Correspondence to Jing Chen, Hualiang Jiang or Chuan He.

Ethics declarations

Competing interests

A patent application on DC_AC50 has been filed by the University of Chicago and the Shanghai Institute of Materia Medica.

Supplementary information

Supplementary information

Supplementary information (PDF 3391 kb)

Rights and permissions

Reprints and permissions

About this article

Check for updates. Verify currency and authenticity via CrossMark

Cite this article

Wang, J., Luo, C., Shan, C. et al. Inhibition of human copper trafficking by a small molecule significantly attenuates cancer cell proliferation. Nature Chem 7, 968–979 (2015). https://doi.org/10.1038/nchem.2381

Download citation

  • Received:

  • Accepted:

  • Published:

  • Issue Date:

  • DOI: https://doi.org/10.1038/nchem.2381

This article is cited by

Search

Advanced search

Quick links

Nature Briefing: Translational Research

Sign up for the Nature Briefing: Translational Research newsletter — top stories in biotechnology, drug discovery and pharma.

Get what matters in translational research, free to your inbox weekly. Sign up for Nature Briefing: Translational Research