Abstract

SIRT6, a member of the SIRT deacetylase family, is responsible for deacetylation of histone H3 Nε-acetyl-lysines 9 (H3K9ac) and 56 (H3K56ac). As a tumor suppressor, SIRT6 has frequently been found to have low expression in various cancers. Here, we report the identification of MDL-800, a selective SIRT6 activator. MDL-800 increased the deacetylase activity of SIRT6 by up to 22-fold via binding to an allosteric site; this interaction led to a global decrease in H3K9ac and H3K56ac levels in human hepatocellular carcinoma (HCC) cells. Consequently, MDL-800 inhibited the proliferation of HCC cells via SIRT6-driven cell-cycle arrest and was effective in a tumor xenograft model. Together, these data demonstrate that pharmacological activation of SIRT6 is a potential therapeutic approach for the treatment of HCC. MDL-800 is a first-in-class small-molecule cellular SIRT6 activator that can be used to physiologically and pathologically investigate the roles of SIRT6 deacetylation.

Access optionsAccess options

Rent or Buy article

Get time limited or full article access on ReadCube.

from$8.99

All prices are NET prices.

Data availability

Crystal structures and diffraction data have been deposited in the Protein Data Bank under accession codes PDB 5X16 (SIRT6-ADPR) and PDB 5Y2F (SIRT6–ADPR–MDL-801). All other data generated or analyzed during the study in this published article (and its supplementary information files) are available from the corresponding author on reasonable request.

Additional information

Publisher’s note: Springer Nature remains neutral with regard to jurisdictional claims in published maps and institutional affiliations.

References

  1. 1.

    Chalkiadaki, A. & Guarente, L. The multifaceted functions of sirtuins in cancer. Nat. Rev. Cancer 15, 608–624 (2015).

  2. 2.

    Feldman, J. L., Dittenhafer-Reed, K. E. & Denu, J. M. Sirtuin catalysis and regulation. J. Biol. Chem. 287, 42419–42427 (2012).

  3. 3.

    Kugel, S. & Mostoslavsky, R. Chromatin and beyond: the multitasking roles for SIRT6. Trends Biochem. Sci. 39, 72–81 (2014).

  4. 4.

    Tasselli, L., Zheng, W. & Chua, K. F. SIRT6: novel mechanisms and links to aging and disease. Trends Endocrinol. Metab. 28, 168–185 (2017).

  5. 5.

    Irminger-Finger, I. Science of cancer and aging. J. Clin. Oncol. 25, 1844–1851 (2007).

  6. 6.

    Sebastián, C. et al. The histone deacetylase SIRT6 is a tumor suppressor that controls cancer metabolism. Cell 151, 1185–1199 (2012).

  7. 7.

    Lin, Z. et al. USP10 antagonizes c-Myc transcriptional activation through SIRT6 stabilization to suppress tumor formation. Cell Rep. 5, 1639–1649 (2013).

  8. 8.

    Marquardt, J. U. et al. Sirtuin-6-dependent genetic and epigenetic alterations are associated with poor clinical outcome in hepatocellular carcinoma patients. Hepatology 58, 1054–1064 (2013).

  9. 9.

    Kugel, S. et al. SIRT6 suppresses pancreatic cancer through control of Lin28b. Cell 165, 1401–1415 (2016).

  10. 10.

    Van Meter, M., Gorbunova, V. & Seluanov, A. SIRT6: a promising target for cancer prevention and therapy. Adv. Exp. Med. Biol. 818, 181–196 (2014).

  11. 11.

    Pan, P. W. et al. Structure and biochemical functions of SIRT6. J. Biol. Chem. 286, 14575–14587 (2011).

  12. 12.

    Jiang, H. et al. SIRT6 regulates TNF-α secretion through hydrolysis of long-chain fatty acyl lysine. Nature 496, 110–113 (2013).

  13. 13.

    Kawahara, T. L. et al. SIRT6 links histone H3 lysine 9 deacetylation to NF-kappaB-dependent gene expression and organismal life span. Cell 136, 62–74 (2009).

  14. 14.

    Michishita, E. et al. Cell cycle-dependent deacetylation of telomeric histone H3 lysine K56 by human SIRT6. Cell Cycle 8, 2664–2666 (2009).

  15. 15.

    Gil, R., Barth, S., Kanfi, Y. & Cohen, H. Y. SIRT6 exhibits nucleosome-dependent deacetylase activity. Nucleic Acids Res. 41, 8537–8545 (2013).

  16. 16.

    Tasselli, L. et al. SIRT6 deacetylates H3K18ac at pericentric chromatin to prevent mitotic errors and cellular senescence. Nat. Struct. Mol. Biol. 23, 434–440 (2016).

  17. 17.

    Ghosh, S., Liu, B., Wang, Y., Hao, Q. & Zhou, Z. Lamin A is an endogenous SIRT6 activator and promotes SIRT6-mediated DNA repair. Cell Rep. 13, 1396–1406 (2015).

  18. 18.

    Feldman, J. L., Baeza, J. & Denu, J. M. Activation of the protein deacetylase SIRT6 by long-chain fatty acids and widespread deacylation by mammalian sirtuins. J. Biol. Chem. 288, 31350–31356 (2013).

  19. 19.

    You, W. et al. Structural basis of sirtuin 6 activation by synthetic small molecules. Angew. Chem. Int. Edn Engl. 56, 1007–1011 (2017).

  20. 20.

    Parenti, M. D. et al. Discovery of novel and selective SIRT6 inhibitors. J. Med. Chem. 57, 4796–4804 (2014).

  21. 21.

    He, B., Hu, J., Zhang, X. & Lin, H. Thiomyristoyl peptides as cell-permeable Sirt6 inhibitors. Org. Biomol. Chem. 12, 7498–7502 (2014).

  22. 22.

    Huang, W. et al. Allosite: a method for predicting allosteric sites. Bioinformatics 29, 2357–2359 (2013).

  23. 23.

    Kokkonen, P. et al. Studying SIRT6 regulation using H3K56 based substrate and small molecules. Eur. J. Pharm. Sci. 63, 71–76 (2014).

  24. 24.

    Pacholec, M. et al. SRT1720, SRT2183, SRT1460, and resveratrol are not direct activators of SIRT1. J. Biol. Chem. 285, 8340–8351 (2010).

  25. 25.

    Sinclair, D. A. & Guarente, L. Small-molecule allosteric activators of sirtuins. Annu. Rev. Pharmacol. Toxicol. 54, 363–380 (2014).

  26. 26.

    Borra, M. T., Smith, B. C. & Denu, J. M. Mechanism of human SIRT1 activation by resveratrol. J. Biol. Chem. 280, 17187–17195 (2005).

  27. 27.

    Kaeberlein, M. et al. Substrate-specific activation of sirtuins by resveratrol. J. Biol. Chem. 280, 17038–17045 (2005).

  28. 28.

    Mao, Z. et al. SIRT6 promotes DNA repair under stress by activating PARP1. Science 332, 1443–1446 (2011).

  29. 29.

    Hu, J., He, B., Bhargava, S. & Lin, H. A fluorogenic assay for screening Sirt6 modulators. Org. Biomol. Chem. 11, 5213–5216 (2013).

  30. 30.

    Tan, Y. et al. A SIRT4-like auto ADP-ribosyltransferase is essential for the environmental growth of Mycobacterium smegmatis. Acta Biochim. Biophys. Sin. (Shanghai) 48, 145–152 (2016).

  31. 31.

    Dai, H., Ellis, J. L., Sinclair, D. A. & Hubbard, B. P. Synthesis and assay of SIRT1-activating compounds. Methods Enzymol. 574, 213–244 (2016).

  32. 32.

    Davenport, A. M., Huber, F. M. & Hoelz, A. Structural and functional analysis of human SIRT1. J. Mol. Biol. 426, 526–541 (2014).

  33. 33.

    Moniot, S., Schutkowski, M. & Steegborn, C. Crystal structure analysis of human Sirt2 and its ADP-ribose complex. J. Struct. Biol. 182, 136–143 (2013).

  34. 34.

    Nguyen, G. T., Schaefer, S., Gertz, M., Weyand, M. & Steegborn, C. Structures of human sirtuin 3 complexes with ADP-ribose and with carba-NAD+ and SRT1720: binding details and inhibition mechanism. Acta Crystallogr. D Biol. Crystallogr 69, 1423–1432 (2013).

  35. 35.

    Du, J. et al. Sirt5 is a NAD-dependent protein lysine demalonylase and desuccinylase. Science 334, 806–809 (2011).

  36. 36.

    Hubatsch, I., Ragnarsson, E. G. & Artursson, P. Determination of drug permeability and prediction of drug absorption in Caco-2 monolayers. Nat. Protoc. 2, 2111–2119 (2007).

  37. 37.

    Min, L. et al. Liver cancer initiation is controlled by AP-1 through SIRT6-dependent inhibition of survivin. Nat. Cell Biol. 14, 1203–1211 (2012).

  38. 38.

    Elhanati, S. et al. Reciprocal regulation between SIRT6 and miR-122 controls liver metabolism and predicts hepatocarcinoma prognosis. Cell Rep. 14, 234–242 (2016).

  39. 39.

    Suter, M. A. et al. A maternal high-fat diet modulates fetal SIRT1 histone and protein deacetylase activity in nonhuman primates. FASEB J. 26, 5106–5114 (2012).

  40. 40.

    Gu, Y., Turck, C. W. & Morgan, D. O. Inhibition of CDK2 activity in vivo by an associated 20K regulatory subunit. Nature 366, 707–710 (1993).

  41. 41.

    el-Deiry, W. S. et al. WAF1, a potential mediator of p53 tumor suppression. Cell 75, 817–825 (1993).

  42. 42.

    Harper, J. W., Adami, G. R., Wei, N., Keyomarsi, K. & Elledge, S. J. The p21 Cdk-interacting protein Cip1 is a potent inhibitor of G1 cyclin-dependent kinases. Cell 75, 805–816 (1993).

  43. 43.

    Hengst, L., Dulic, V., Slingerland, J. M., Lees, E. & Reed, S. I. A cell cycle-regulated inhibitor of cyclin-dependent kinases. Proc. Natl Acad. Sci. USA 91, 5291–5295 (1994).

  44. 44.

    Polyak, K. et al. Cloning of p27Kip1, a cyclin-dependent kinase inhibitor and a potential mediator of extracellular antimitogenic signals. Cell 78, 59–66 (1994).

  45. 45.

    Toyoshima, H. & Hunter, T. p27, a novel inhibitor of G1 cyclin-Cdk protein kinase activity, is related to p21. Cell 78, 67–74 (1994).

  46. 46.

    Morgan, D. O. Principles of CDK regulation. Nature 374, 131–134 (1995).

  47. 47.

    Hubbard, B. P. et al. Evidence for a common mechanism of SIRT1 regulation by allosteric activators. Science 339, 1216–1219 (2013).

  48. 48.

    Dai, H. et al. Crystallographic structure of a small molecule SIRT1 activator-enzyme complex. Nat. Commun. 6, 7645 (2015).

  49. 49.

    Huang, M. et al. Screening and biological evaluation of a novel STAT3 signaling pathway inhibitor against cancer. Bioorg. Med. Chem. Lett. 26, 5172–5176 (2016).

  50. 50.

    Zhao, Y. et al. Crystal structures of PI3Kα complexed with PI103 and its derivatives: new directions for inhibitors design. ACS Med. Chem. Lett. 5, 138–142 (2013).

  51. 51.

    Jiang, H. et al. Peptidomimetic inhibitors of APC-Asef interaction block colorectal cancer migration. Nat. Chem. Biol. 13, 994–1001 (2017).

  52. 52.

    Ran, F. A. et al. Genome engineering using the CRISPR-Cas9 system. Nat. Protoc. 8, 2281–2308 (2013).

Download references

Acknowledgements

We sincerely thank D. Sinclair for discussion on activator application, and C. Steegborn and A. Mai for discussion on crystallization. We thank Z.-G. Han (Key Laboratory of Systems Biomedicine, Ministry of Education, and Collaborative Innovation Center of Systems Biomedicine of Rui-Jin Hospital, Shanghai Jiao Tong University School of Medicine) for providing cell lines. This work was supported in part by grants from the National Basic Research Program of China (973 Program) (2015CB910403 to J.Z. and G.C.), the National Natural Science Foundation of China (81721004 to G.C. and J.Z.; 91753117 to J.Z.; 81322046 to J.Z.; 81302698 to S.L.; 31671459 to Y.X.; U1605221 to H.L.; and 2181001006 to H.J.), the Program for Changjiang Scholars and the Innovative Research Team of the University of the Ministry Education of China (2017 to J.Z.), the CAS Interdisciplinary Innovation Team (2017 to J.Z.), the Innovation Program of the Shanghai Municipal Education Commission (2019 to J.Z.), the State Key Laboratory of Luminescence and application (SKLA-2016-12 to Y.X.), and the Strategic Priority Research Program of the Chinese Academy of Sciences, ‘Personalized Medicines—Molecular Signature-based Drug Discovery and Development’ (XDA12040100 to H.J.).

Author information

Author notes

  1. These authors contributed equally: Zhimin Huang, Junxing Zhao, Wei Deng, Yingyi Chen, Jialin Shang.

Affiliations

  1. Key Laboratory of Cell Differentiation and Apoptosis, Ministry of Education, Department of Pathophysiology, Ruijin Hospital, Shanghai Jiao-Tong University School of Medicine, Shanghai, China

    • Zhimin Huang
    • , Junxing Zhao
    • , Yingyi Chen
    • , Jialin Shang
    • , Kun Song
    • , Lu Zhang
    • , Chengxiang Wang
    • , Shaoyong Lu
    • , Xiuyan Yang
    • , Jianrong Xu
    • , Qiufen Zhang
    • , Jie Zhong
    • , Ying Xu
    • , Guoqiang Chen
    •  & Jian Zhang
  2. Shanghai Institutes for Biological Sciences, Chinese Academy of Sciences, Shanghai, China

    • Wei Deng
    • , Mingzhe Xiao
    •  & Y Eugene Chin
  3. Engineering Research Center for the Development and Application of Ethnic Medicine and TCM, Ministry of Education, Guizhou Medical University, Guiyang, China

    • Bin He
  4. Structural Genomics Consortium, University of Toronto, Toronto, Ontario, Canada

    • Jinrong Min
  5. State Key Laboratory of Drug Research, Shanghai Institute of Materia Medica, Chinese Academy of Sciences, Shanghai, China

    • Hao Hu
    • , Minjia Tan
    •  & Hualiang Jiang
  6. School of Life Sciences and Technology, Tongji University, Shanghai, China

    • Xiaoxiang Sun
    •  & Zhiyong Mao
  7. Basic Clinical Research Center, Renji Hospital, Shanghai Jiao Tong University School of Medicine, Shanghai, China

    • Houwen Lin
    •  & Jian Zhang
  8. Medicinal Bioinformatics Center, Shanghai JiaoTong University School of Medicine, Shanghai, China

    • Jian Zhang

Authors

  1. Search for Zhimin Huang in:

  2. Search for Junxing Zhao in:

  3. Search for Wei Deng in:

  4. Search for Yingyi Chen in:

  5. Search for Jialin Shang in:

  6. Search for Kun Song in:

  7. Search for Lu Zhang in:

  8. Search for Chengxiang Wang in:

  9. Search for Shaoyong Lu in:

  10. Search for Xiuyan Yang in:

  11. Search for Bin He in:

  12. Search for Jinrong Min in:

  13. Search for Hao Hu in:

  14. Search for Minjia Tan in:

  15. Search for Jianrong Xu in:

  16. Search for Qiufen Zhang in:

  17. Search for Jie Zhong in:

  18. Search for Xiaoxiang Sun in:

  19. Search for Zhiyong Mao in:

  20. Search for Houwen Lin in:

  21. Search for Mingzhe Xiao in:

  22. Search for Y Eugene Chin in:

  23. Search for Hualiang Jiang in:

  24. Search for Ying Xu in:

  25. Search for Guoqiang Chen in:

  26. Search for Jian Zhang in:

Contributions

J. Zhang conceived and supervised the project. J. Zhang and Z.H. designed the experiments. Z.H., J. Zhao,Y.C., J.S., L.Z., C.W., H.H., J.X., J. Zhong, Q.Z., X.S., and Y.X. performed the biological experiments. J. Zhang, G.C.,Y.X., Z.H., S.L., and H.L. analyzed data. Z.H. and K.S. performed the crystallography. Y.C. and X.Y. carried out synthesis, purification, and characterization of compounds. W.D., S.L., B.H., J.M., H.H., M.T., X.S., Z.M., M.X., Y.E.C., and H.J. generated key protein reagents. K.S. solved the crystal structures. J. Zhang wrote the manuscript, and all other authors contributed specific parts of the manuscript; J. Zhang, G.C., and Y.X. assume responsibility for the manuscript in its entirety.

Competing interests

The authors declare no competing interests.

Corresponding authors

Correspondence to Ying Xu or Guoqiang Chen or Jian Zhang.

Supplementary information

  1. Supplementary Text and Figures

    Supplementary Figures 1–69 and Supplementary Tables 1–20

  2. Reporting Summary

  3. Supplementary Note

    Synthetic Procedures

About this article

Publication history

Received

Accepted

Published

DOI

https://doi.org/10.1038/s41589-018-0150-0