Interfering with bromodomain epigenome readers as therapeutic option in mucoepidermoid carcinoma
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Emerging evidence indicates that bromodomains comprise a conserved class of epigenome readers involved in cancer development and inflammation. Bromodomains are associated with epigenetic modifications of gene transcription through interactions with lysine residues of histone tails. Particularly, the bromodomain and extra-terminal domain (BET) family member BRD4 has been found to be involved in the control over oncogenes, including c-MYC, and in the maintenance of downstream inflammatory processes. The objective of this study was to evaluate the effect of pharmacologically displacing BRD4 in mucoepidermoid carcinoma (MEC) cells.
We assessed the presence of BRD4 levels in a panel of human MEC tissue samples in conjunction with histological grading and clinical information. In vitro studies were carried out using human MEC-derived cell lines. The BET inhibitor iBET762 was administered to MEC cells to assess the impact of disrupted BRD4 signaling on colony forming capacities and cell cycle status. The activation of cellular senescence induced by iBET762 was determined by immunohistochemical staining for p16ink4. Flow cytometry was used to identify populations of cancer stem cells in MEC-derived cell lines.
We found that primary human MECs and MEC-derived cell lines are endowed with high BRD4 expression levels compared to those in normal salivary glands. We also found that, by displacing BRD4 from chromatin using the BET inhibitor iBET762, MEC cells lose their colony forming capacities and undergo G1 cell cycle arrest and senescence. Finally, we found that targeted displacement of BRD4 from chromatin results in depletion of cancer stem cells from the overall MEC cell populations.
Our findings indicate that bromodomain-mediated gene regulation constitutes an epigenetic mechanism that is deregulated in MEC cells and that the use of BET inhibitors may serve as a feasible therapeutic strategy to manage MECs.
KeywordsMucoepidermoid carcinoma BRD4 Epigenetic Cancer stem cells iBET762 Epi-drug
This work was conducted during a visiting scholar period at the University of Michigan, sponsored by the Capes Foundation within the Ministry of Education, Brazil (grant n. BEX / 88881.132606/2016-01 PDSE). This grant was funded by a University of Michigan School of Dentistry faculty grant, and a Cancer Center Support Grant (P30 CA046592). The authors declare no potential conflicts of interest with respect to authorships and/or publication of this article.
RLM performed most of the cell culture-based assays, IHC, IF, and participated in the organization of the figures. CHVNF performed the flow cytometry assays, LAR helped with the IHC and IF assays, and LPW helped with the immunofluorescence assays and its quantifications. VZ and MDM contributed to the collection of MEC tissue samples and their clinical data and helped with their re-evaluation. PAV, MAL, CHS and RMC contributed to the conception, design, data organization and writing of the manuscript. All authors gave their final approval and agreed to be accountable for all aspects of the work.
Compliance with ethical standards
Conflict of interest
All procedures performed in studies involving human participants were in accordance with the ethical standards of the institutional and/or national research committee and with the 1964 Helsinki declaration and its later amendments or comparable ethical standards.
- 4.E.S. Choi, S. Oh, B. Jang, H.J. Yu, J.A. Shin, N.P. Cho, I.H. Yang, D.H. Won, H.J. Kwon, S.D. Hong, S.D. Cho, Silymarin and its active component silibinin act as novel therapeutic alternatives for salivary gland cancer by targeting the ERK1/2-Bim signaling cascade. Cell Oncol. 40, 235–246 (2017)Google Scholar
- 6.J.H. Mikesch, W. Hartmann, L. Angenendt, O. Huber, C. Schliemann, M.F. Arteaga, E. Wardelmann, C. Rudack, W.E. Berdel, M. Stenner and I. Grunewald, AAA+ ATPases Reptin and Pontin as potential diagnostic and prognostic biomarkers in salivary gland cancer - a short report. Cell Oncol. 41, 455–462 (2018)Google Scholar
- 7.L. Barnes, Universitäts-Spital Zurich. Dept. Pathologie., International Academy of Pathology., World Health Organization. and International Agency for Research on Cancer., Pathology and genetics of head and neck tumours, (IARC Press, Lyon, 2007)Google Scholar
- 10.T. Cerda, X.S. Sun, S. Vignot, P.Y. Marcy, B. Baujat, A.C. Baglin, A.M. Ali, S. Testelin, E. Reyt, F. Janot, J. Thariat, A rationale for chemoradiation (vs radiotherapy) in salivary gland cancers? On behalf of the REFCOR (French rare head and neck cancer network). Crit. Rev. Oncol. Hematol. 91, 142–158 (2014)CrossRefGoogle Scholar
- 12.R.M. Castilho, C.H. Squarize, L.O. Almeida, Epigenetic modifications and head and neck Cancer: Implications for tumor progression and resistance to therapy. Int J Mol Sci. 18 E15063 (2017)Google Scholar
- 13.M.S. Gilardini Montani, M. Granato, C. Santoni, P. Del Porto, N. Merendino, G. D'Orazi, A. Faggioni, M. Cirone, Histone deacetylase inhibitors VPA and TSA induce apoptosis and autophagy in pancreatic cancer cells. Cell Oncol. 40, 167–180 (2017)Google Scholar
- 14.M. Staberg, S.R. Michaelsen, R.D. Rasmussen, M. Villingshoj, H.S. Poulsen, P. Hamerlik, Inhibition of histone deacetylases sensitizes glioblastoma cells to lomustine. Cell Oncol. 40, 21–32 (2017)Google Scholar
- 15.M.F. Segura, B. Fontanals-Cirera, A. Gaziel-Sovran, M.V. Guijarro, D. Hanniford, G. Zhang, P. Gonzalez-Gomez, M. Morante, L. Jubierre, W. Zhang, F. Darvishian, M. Ohlmeyer, I. Osman, M.M. Zhou, E. Hernando, BRD4 sustains melanoma proliferation and represents a new target for epigenetic therapy. Cancer Res. 73, 6264–6276 (2013)CrossRefGoogle Scholar
- 23.E. Nicodeme, K.L. Jeffrey, U. Schaefer, S. Beinke, S. Dewell, C.W. Chung, R. Chandwani, I. Marazzi, P. Wilson, H. Coste, J. White, J. Kirilovsky, C.M. Rice, J.M. Lora, R.K. Prinjha, K. Lee, A. Tarakhovsky, Suppression of inflammation by a synthetic histone mimic. Nature 468, 1119–1123 (2010)CrossRefGoogle Scholar
- 24.O. Mirguet, R. Gosmini, J. Toum, C.A. Clement, M. Barnathan, J.M. Brusq, J.E. Mordaunt, R.M. Grimes, M. Crowe, O. Pineau, M. Ajakane, A. Daugan, P. Jeffrey, L. Cutler, A.C. Haynes, N.N. Smithers, C.W. Chung, P. Bamborough, I.J. Uings, A. Lewis, J. Witherington, N. Parr, R.K. Prinjha, E. Nicodeme, Discovery of epigenetic regulator I-BET762: Lead optimization to afford a clinical candidate inhibitor of the BET bromodomains. J. Med. Chem. 56, 7501–7515 (2013)CrossRefGoogle Scholar
- 26.L.O. Almeida, D.M. Guimaraes, M.D. Martins, M.A.T. Martins, K.A. Warner, J.E. Nor, R.M. Castilho, C.H. Squarize, Unlocking the chromatin of adenoid cystic carcinomas using HDAC inhibitors sensitize cancer stem cells to cisplatin and induces tumor senescence. Stem Cell Res. 21, 94–105 (2017)CrossRefGoogle Scholar
- 28.D.M. Guimaraes, L.O. Almeida, M.D. Martins, K.A. Warner, A.R. Silva, P.A. Vargas, F.D. Nunes, C.H. Squarize, J.E. Nor, R.M. Castilho, Sensitizing mucoepidermoid carcinomas to chemotherapy by targeted disruption of cancer stem cells. Oncotarget 7, 42447–42460 (2016)Google Scholar
- 29.A. Adams, K. Warner, A.T. Pearson, Z. Zhang, H.S. Kim, D. Mochizuki, G. Basura, J. Helman, A. Mantesso, R.M. Castilho, M.S. Wicha, J.E. Nor, ALDH/CD44 identifies uniquely tumorigenic cancer stem cells in salivary gland mucoepidermoid carcinomas. Oncotarget 6, 26633–26650 (2015)Google Scholar
- 31.M. Granic, P. Suton, D. Mueller, I. Cvrljevic, I. Luksic, Prognostic factors in head and neck mucoepidermoid carcinoma: experience at a single institution based on 64 consecutive patients over a 28-year period. Int J Oral Maxillofac Surg. 47, 283-288 (2018)Google Scholar
- 32.A.C. Birkeland, S.K. Foltin, N.L. Michmerhuizen, R.C. Hoesli, A.J. Rosko, S. Byrd, M. Yanik, J.E. Nor, C.R. Bradford, M.E. Prince, T.E. Carey, J.B. McHugh, M.E. Spector, J.C. Brenner, Correlation of Crtc1/3-Maml2 fusion status, grade and survival in mucoepidermoid carcinoma. Oral Oncol. 68, 5–8 (2017)CrossRefGoogle Scholar
- 35.J. Zuber, J. Shi, E. Wang, A.R. Rappaport, H. Herrmann, E.A. Sison, D. Magoon, J. Qi, K. Blatt, M. Wunderlich, M.J. Taylor, C. Johns, A. Chicas, J.C. Mulloy, S.C. Kogan, P. Brown, P. Valent, J.E. Bradner, S.W. Lowe, C.R. Vakoc, RNAi screen identifies Brd4 as a therapeutic target in acute myeloid leukaemia. Nature 478, 524–528 (2011)CrossRefGoogle Scholar
- 36.A. Chaidos, V. Caputo, K. Gouvedenou, B. Liu, I. Marigo, M.S. Chaudhry, A. Rotolo, D.F. Tough, N.N. Smithers, A.K. Bassil, T.D. Chapman, N.R. Harker, O. Barbash, P. Tummino, N. Al-Mahdi, A.C. Haynes, L. Cutler, B. Le, A. Rahemtulla, I. Roberts, M. Kleijnen, J.J. Witherington, N.J. Parr, R.K. Prinjha, A. Karadimitris, Potent antimyeloma activity of the novel bromodomain inhibitors I-BET151 and I-BET762. Blood 123, 697–705 (2014)CrossRefGoogle Scholar
- 37.J.E. Delmore, G.C. Issa, M.E. Lemieux, P.B. Rahl, J. Shi, H.M. Jacobs, E. Kastritis, T. Gilpatrick, R.M. Paranal, J. Qi, M. Chesi, A.C. Schinzel, M.R. McKeown, T.P. Heffernan, C.R. Vakoc, P.L. Bergsagel, I.M. Ghobrial, P.G. Richardson, R.A. Young, W.C. Hahn, K.C. Anderson, A.L. Kung, J.E. Bradner, C.S. Mitsiades, BET bromodomain inhibition as a therapeutic strategy to target c-Myc. Cell 146, 904–917 (2011)CrossRefGoogle Scholar
- 38.S.B. Baylin, P.A. Jones, Epigenetic determinants of Cancer. Cold Spring Harb Perspect Biol. 8(9) (2016). https://doi.org/10.1101/cshperspect.a019505
- 39.Y. Yokoyama, H. Zhu, J.H. Lee, A.V. Kossenkov, S.Y. Wu, J.M. Wickramasinghe, X. Yin, K.C. Palozola, A. Gardini, L.C. Showe, K.S. Zaret, Q. Liu, D. Speicher, J.R. Conejo-Garcia, J.E. Bradner, Z. Zhang, A.K. Sood, T. Ordog, B.G. Bitler, R. Zhang, BET inhibitors suppress ALDH activity by targeting ALDH1A1 super-enhancer in ovarian Cancer. Cancer Res. 76, 6320–6330 (2016)CrossRefGoogle Scholar
- 40.N. Tasdemir, A. Banito, J.S. Roe, D. Alonso-Curbelo, M. Camiolo, D.F. Tschaharganeh, C.H. Huang, O. Aksoy, J.E. Bolden, C.C. Chen, M. Fennell, V. Thapar, A. Chicas, C.R. Vakoc, S.W. Lowe, BRD4 connects enhancer remodeling to senescence immune surveillance. Cancer Discov. 6, 612–629 (2016)CrossRefGoogle Scholar