Advertisement

Endocrine

pp 1–9 | Cite as

Human telomerase reverse transcriptase in papillary thyroid cancer: gene expression, effects of silencing and regulation by BET inhibitors in thyroid cancer cells

  • Valentina Maggisano
  • Marilena Celano
  • Saverio Massimo Lepore
  • Marialuisa Sponziello
  • Francesca Rosignolo
  • Valeria Pecce
  • Antonella Verrienti
  • Federica Baldan
  • Catia Mio
  • Lorenzo Allegri
  • Marianna Maranghi
  • Rosa Falcone
  • Giuseppe Damante
  • Diego RussoEmail author
  • Stefania Bulotta
Original Article
  • 76 Downloads

Abstract

Purpose

Mutations in TERT promoter have been detected in the more aggressive papillary thyroid cancers (PTCs). To elucidate the role of TERT as an eligible molecular target in these tumors, the expression of hTERT was analyzed in a series of PTCs and the effects of both pharmacological and RNA-interference-induced hTERT silencing were investigated in two human PTC cell lines (K1 and BCPAP).

Methods

The expression levels of hTERT mRNA and protein were evaluated by real-time PCR and western blot assays, respectively. Effects of hTERT silencing on PTC cell lines were analyzed by MTT, migration and western blot assays. Pharmacological inhibition of hTERT was performed using two bromodomain and extra-terminal (BET) inhibitors, JQ1 and I-BET762.

Results

hTERT expression results increased in 20 out of 48 PTCs, including tumors either positive or negative for the presence of hTERT promoter and/or BRAF mutations. In K1 and BCPAP cells, hTERT silencing determined a reduction in cell viability (~50% for K1 and ~70%, for BCPAP, vs control) and migration properties that were associated with a decrease of AKT phosphorylation and β-Catenin expression. Moreover, hTERT mRNA levels were down-regulated by two BET inhibitors, JQ1 and I-BET762, which at the same dosage (0.5 and 5 µM) reduced the growth of these thyroid cancer cells.

Conclusions

These findings demonstrate that hTERT may represent an excellent therapeutic target in subgroups of aggressive PTCs.

Keywords

Papillary thyroid cancer siRNA anti-hTERT phospho-AKT BET inhibitors 

Notes

Funding

The study was supported by the Fondazione Umberto Di Mario ONLUS and Banca d’Italia, and by a Grant to G.D. from Italian Minister of Foreigner Affairs and International Cooperation (MAECI) (Progetti Grande Rilevanza Italia-Serbia No. PGR00223). F. Baldan is a recipient of a fellowship from AIRC.

Compliance with ethical standards

Conflict of interest

The authors declare that they have no conflict of interest.

Ethics approval

All procedures performed in the studies involving human participants were in accordance with the ethical standards of the institutional research committee and with the 1964 Helsinki declaration and its later amendments or comparable ethical standards.

Informed consent

Informed consent was obtained from all individual participants included in the study.

Supplementary material

12020_2018_1836_MOESM1_ESM.doc (60 kb)
Supplementary Table 1

References

  1. 1.
    C.M. Kitahara, J.A. Sosa, The changing incidence of thyroid cancer. Nat. Rev. Endocrinol. 12, 646–653 (2016)CrossRefGoogle Scholar
  2. 2.
    M.E. Cabanillas, D.G. McFadden, C. Durante, Thyroid cancer. Lancet 388, 2783–2795 (2016)CrossRefGoogle Scholar
  3. 3.
    S. Bulotta, M. Celano, G. Costante, D. Russo, Emerging strategies for managing differentiated thyroid cancers refractory to radioiodine. Endocrine 52, 214–221 (2016)CrossRefGoogle Scholar
  4. 4.
    Y. Jin, D. van Nostrand, L. Cheng, M. Liu, L. Chen, Radioiodine refractory differentiated thyroid cancer. Crit. Rev. Oncol. Hematol. 125, 111–120 (2018)CrossRefGoogle Scholar
  5. 5.
    E.N. Klein Hesselink, D. Steenvoorden, E. Kapiteijn, E.P. Corssmit, A.N. van der Horst-Schrivers, J.D. Lefrandt, T.P. Links, O.M. Dekkers, Therapy of endocrine disease: response and toxicity of small-molecule tyrosine kinase inhibitors in patients with thyroid carcinoma: a systematic review and meta-analysis. Eur. J. Endocrinol. 172, R215–R225 (2015)CrossRefGoogle Scholar
  6. 6.
    W. Yimaer, A. Abudouyimu, Y. Tian, S. Magaoweiya, D. Bagedati, H. Wen, Efficacy and safety of vascular endothelial growth factor receptor tyrosine kinase inhibitors in the treatment of advanced thyroid cancer: a meta-analysis of randomized controlled trials. Onco Targets Ther. 9, 1167–1173 (2016)Google Scholar
  7. 7.
    R. Leão, J.D. Apolónio, D. Lee, A. Figueiredo, U. Tabori, P. Castelo-Branco, Mechanisms of human telomerase reverse transcriptase (hTERT) regulation: clinical impacts in cancer. J. Biomed. Sci. 25(1), 22 (2018)CrossRefGoogle Scholar
  8. 8.
    A. Pestana, J. Vinagre, M. Sobrinho-Simões, P. Soares, TERT biology and function in cancer: beyond immortalisation. J. Mol. Endocrinol. 58(2), R129–R146 (2017)CrossRefGoogle Scholar
  9. 9.
    A. Alzahrani, R. Alsaadi, A.K. Murugan, B.B. Sadiq, TERT promoter mutations in thyroid cancer. Horm. Cancer 7, 165–177 (2016)CrossRefGoogle Scholar
  10. 10.
    X. Liu, J. Bishop, Y. Shan, S. Pai, D. Liu, A.K. Murugan, H. Sun, A.K. El-Naggar, M. Xing, Highly prevalent TERT promoter mutations in aggressive thyroid cancers. Endocr. Rel. Cancer 20, 603–610 (2013)CrossRefGoogle Scholar
  11. 11.
    B. Xu, R. Ghossein, Genomic landscape of poorly differentiated and anaplastic thyroid carcinoma. Endocr. Pathol. 27, 205–212 (2016)CrossRefGoogle Scholar
  12. 12.
    R. Liu, M. Xing, TERT promoter mutations in thyroid cancer. Endocr. Rel. Cancer 23, R143–R155 (2016)CrossRefGoogle Scholar
  13. 13.
    H.G. Vuong, A.M. Altibi, U.N. Duong, H.T. Ngo, T.Q. Pham, H.M. Tran, N. Oishi, K. Mochizuki, T. Nakazawa, L. Hassell, R. Katoh, T. Kondo, Role of molecular markers to predict distant metastasis in papillary thyroid carcinoma: promising value of TERT promoter mutations and insignificant role of BRAF mutations—a meta-analysis. Tumour Biol. 39(10), 1010428317713913 (2017)CrossRefGoogle Scholar
  14. 14.
    G.C. Penna, A. Pestana, J.M. Cameselle, D. Momesso, F.A. de Andrade, A.P.A. Vidal, M.L. Araujo Junior, M. Melo, P.V. Fernandes, R. Corbo, M. Vaisman, M. Sobrinho-Simões, P. Soares et al. TERTp mutation is associated with a shorter progression free survival in patients with aggressive histology subtypes of follicular-cell derived thyroid carcinoma. Endocrine 61(3), 489–498 (2018)CrossRefGoogle Scholar
  15. 15.
    M. Muzza, C. Colombo, S. Rossi, D. Tosi, V. Cirello, M. Perrino, S. De Leo, E. Magnani, E. Pignatti, B. Vigo, M. Simoni, G. Bulfamante, L. Vicentini et al. Telomerase in differentiated thyroid cancer: promoter mutations, expression and localization. Mol. Cell. Endocrinol. 399, 288–295 (2015)CrossRefGoogle Scholar
  16. 16.
    V. Maggisano, M. Celano, S. Lepore, G.E. Lombardo, M. Sponziello, F. Rosignolo, A. Verrienti, F. Baldan, E. Puxeddu, C. Durante, S. Filetti, G. Damante, D. Russo et al. Silencing of hTERT blocks growth and migration of anaplastic thyroid cancer cells. Mol. Cell. Endocrinol. 448, 34–40 (2017)CrossRefGoogle Scholar
  17. 17.
    D. Cheng, Y. Zhao, S. Wang, F. Zhang, M. Russo, S.B. McMahon, J. Zhu, Repression of telomerase gene promoter requires human-specific genomic context and is mediated by multiple HDAC1-containing corepressor complexes. FASEB J. 31, 1165–1178 (2017)CrossRefGoogle Scholar
  18. 18.
    M. Celano, C. Mio, M. Sponziello, A. Verrienti, S. Bulotta, C. Durante, G. Damante, D. Russo, Targeting post-translational histone modifications for the treatment of non-medullary thyroid cancer. Mol. Cell. Endocrinol. 469, 38–47 (2018)CrossRefGoogle Scholar
  19. 19.
    X. Zhu, S.Y. Cheng, Epigenetic modifications: novel therapeutic approach for thyroid cancer. Endocrinol. Metab. 32, 326–331 (2017)CrossRefGoogle Scholar
  20. 20.
    C. Mio, E. Lavarone, F. Baldan, B. Toffoletto, C. Puppin, S. Filetti, C. Durante, D. Russo, A. Orlacchio, A. Di Cristofano, C. Di Loreto, G. Damante, MCM5 as a target of BET inhibitors in thyroid cancer cells. Endocr. Relat. Cancer 23(4), 335–347 (2016)CrossRefGoogle Scholar
  21. 21.
    X. Gao, X. Wu, X. Zhang, W. Hua, Y. Zhang, Y. Maimaiti, Z. Gao, Y. Zhang, Inhibition of BRD4 suppresses tumor growth and enhances iodine uptake in thyroid cancer. Biochem. Biophys. Res. Commun. 469(3), 679–685 (2016)CrossRefGoogle Scholar
  22. 22.
    M. Pérez-Salvia, M. Esteller, Bromodomain inhibitors and cancer therapy: from structures to applications. Epigenetics 12(5), 323–339 (2017)CrossRefGoogle Scholar
  23. 23.
    E. Wadhwa, T. Nicolaides, Bromodomain inhibitor review: bromodomain and extra-terminal family protein inhibitors as a potential new therapy in central nervous system tumors. Cureus 8(5), e620 (2016)Google Scholar
  24. 24.
    S. Natarajan, Z. Chen, E.V. Wancewicz, B.P. Monia, D.R. Corey, Telomerase reverse transcriptase (hTERT) mRNA and telomerase RNA (hTR) as targets for downregulation of telomerase activity. Oligonucleotides 14, 263–273 (2004)CrossRefGoogle Scholar
  25. 25.
    W. Zhang, L. Xing, RNAi gene therapy of SiHa cells via targeting human TERT induces growth inhibition and enhances radiosensitivity. Int. J. Oncol. 43, 1228–1234 (2013)CrossRefGoogle Scholar
  26. 26.
    A.Q. Liu, L.Y. Ge, X. Lu, X.L. Luo, Y. Cai, X.Q. Ye, F.F. Geng, Silencing of the hTERT gene by shRNA inhibits colon cancer SW480 cell growth in vitro and in vivo. PLoS ONE 9, e107019 2014).CrossRefGoogle Scholar
  27. 27.
    P. Chen, W.L. Gu, M.Z. Gong, J. Wang, D.Q. Li, shRNA-mediated silencing of hTERT suppresses proliferation and promotes apoptosis in osteosarcoma cells. Cancer Gene Ther. 24, 325–332 (2017)CrossRefGoogle Scholar
  28. 28.
    L. Teng, M.C. Specht, C.B. Barden, T.J.Fahey III, Antisense hTERT inhibits thyroid cancer cell growth. J. Clin. Endocrinol. Metab. 88, 1362–1366 (2003).CrossRefGoogle Scholar
  29. 29.
    G.E. Lombardo, V. Maggisano, M. Celano, D. Cosco, C. Mignogna, F. Baldan, S.M. Lepore, L. Allegri, S. Moretti, C. Durante, G. Damante, M. Fresta, D. Russo et al. Anti-hTERT siRNA-loaded nanoparticles block the growth of anaplastic thyroid cancer xenograft. Mol. Cancer Ther. 17(6), 1187–1195 (2018)CrossRefGoogle Scholar
  30. 30.
    B.R. Haugen, E.K. Alexander, K.C. Bible, G. Doherty, S.J. Mandel, Y.E. Nikiforov, F. Pacini, G. Randolph, A. Sawka, M. Schlumberger, K.G. Schuff, S.I. Sherman, J.A. Sosa et al. 2015 American Thyroid Association Management Guidelines for Adult Patients with Thyroid Nodules and Differentiated Thyroid Cancer. Thyroid 26, 1–133 (2016)CrossRefGoogle Scholar
  31. 31.
    M. Celano, C. Mignogna, F. Rosignolo, M. Sponziello, M. Iannone, S.M. Lepore, G.E. Lombardo, V. Maggisano, A. Verrienti, S. Bulotta, C. Durante, C. Di Loreto, G. Damante et al. Expression of YAP1 in aggressive thyroid cancer. Endocrine 59, 209–212 (2018)CrossRefGoogle Scholar
  32. 32.
    M. Sponziello, F. Rosignolo, M. Celano, V. Maggisano, V. Pecce, R.F. De Rose, G.E. Lombardo, C. Durante, S. Filetti, G. Damante, D. Russo, S. Bulotta, Fibronectin-1 expression is increased in aggressive thyroid cancer and favors the migration and invasion of cancer cells. Mol. Cell. Endocrinol. 431, 123–132 (2016)CrossRefGoogle Scholar
  33. 33.
    R.E. Schweppe, J.P. Klopper, C. Korch, U. Puqazhenthi, M. Benezra, J.A. Knauf, J.A. Fagin, L.A. Marlow, J.A. Copland, R.C. Smallridge, B.R. Haugen, Deoxyribonucleic acid profiling analysis of 40 human thyroid cancer cell lines reveals cross-contamination resulting in cell line redundancy and misidentification. J. Clin. Endocrinol. Metab. 93(11), 4331–4341 (2008)CrossRefGoogle Scholar
  34. 34.
    M.J. Jeon, W.G. Kim, S. Sim, S. Lim, H. Kwon, T.Y. Kim, Y.K. Shong, W.B. Kim, Low prevalence of somatic TERT promoter mutations in classic papillary thyroid carcinoma. Endocrinol. Metab. 31, 100–104 (2016)CrossRefGoogle Scholar
  35. 35.
    M. D’Agostino, M. Sponziello, C. Puppin, M. Celano, V. Maggisano, F. Baldan, M. Biffoni, S. Bulotta, C. Durante, S. Filetti, G. Damante, D. Russo, Different expression of TSH receptor and NIS genes in thyroid cancer: role of epigenetics. J. Mol. Endocrinol. 52, 121–131 (2014)CrossRefGoogle Scholar
  36. 36.
    M. Celano, V. Maggisano, R.F. De Rose, S. Bulotta, J. Maiuolo, M. Navarra, D. Russo, Flavonoid fraction of Citrus reticulata juice reduces proliferation and migration of anaplastic thyroid carcinoma cells. Nutr. Cancer 67(7), 1183–1190 (2015)CrossRefGoogle Scholar
  37. 37.
    S.C. Akıncılar, E. Khattar, P.L. Boon, B. Unal, M.J. Fullwood, V. Tergaonkar, Long-range chromatin interactions drive mutant TERT promoter activation. Cancer Discov. 6(11), 1276–1291 (2016)CrossRefGoogle Scholar
  38. 38.
    A. Berdelou, L. Lamartina, M. Klain, S. Leboulleux, M. Schlumberger, Treatment of refractory thyroid cancer. Endocr. Rel. Cancer 25, R209–R223 (2018)CrossRefGoogle Scholar
  39. 39.
    J.A. Fagin, S.A. Wells Jr., Biologic and clinical perspectives on thyroid cancer. N. Engl. J. Med. 375, 1054–1067 (2016).CrossRefGoogle Scholar
  40. 40.
    T.J. Giordano, Genomic hallmarks of thyroid neoplasia. Annu. Rev. Pathol. 13, 141–162 (2018)CrossRefGoogle Scholar
  41. 41.
    M. Molina-Vega, J. García-Alemán, A. Sebastián-Ochoa, I. Mancha-Doblas, M. Trigo-Pérez, F. Tinahones-Madueño, Tyrosine kinase inhibitors in iodine-refractory differentiated thyroid cancer: experience in clinical practice. Endocrine 59(2), 395–401 (2018)CrossRefGoogle Scholar
  42. 42.
    D. Rusinek, A. Pfeifer, J. Krajewska, M. Oczko-Wojciechowska, D. Handkiewicz-Junak, A. Pawlaczek, J. Zebracka-Gala, M. Kowalska, R. Cyplinska, E. Zembala-Nozynska, M. Chekan, E. Chmielik, A. Kropinska et al. Coexistence of TERT promoter mutations and the BRAF V600E alteration and its impact on histopathological features of papillary thyroid carcinoma in a selected series of Polish patients. Int. J. Mol. Sci. 19(9), E2647 (2018)CrossRefGoogle Scholar
  43. 43.
    M. Xing, R. Liu, X. Liu, A.K. Murugan, G. Zhu, M.A. Zeiger, S. Pai, J. Bishop, BRAF V600E and TERT promoter mutations cooperatively identify the most aggressive papillary thyroid cancer with highest recurrence. J. Clin. Oncol. 32, 2718–2726 (2014)CrossRefGoogle Scholar
  44. 44.
    L. Jin, E. Chen, S. Dong, Y. Cai, X. Zhang, Y. Zhou, R. Zeng, F. Yang, C. Pan, Y. Liu, W. Wu, M. Xing, X. Zhang et al. BRAF and TERT promoter mutations in the aggressiveness of papillary thyroid carcinoma: a study of 653 patients. Oncotarget 7, 18346–18355 (2016)Google Scholar
  45. 45.
    S. Moon, Y.S. Song, Y.A. Kim, J.A. Lim, S.W. Cho, J.H. Moon, S. Hahn, D.J. Park, Y.J. Park, Effects of coexistent BRAFV600E and TERT promoter mutations on poor clinical outcomes in papillary thyroid cancer: a meta-analysis. Thyroid 27(5), 651–660 (2017)CrossRefGoogle Scholar
  46. 46.
    Z. Kordestani, M. Sanjari, M. Safavi, M. Mashrouteh, G. Asadikaram, M.F.S. Abadi, A. Mirzazadeh, Enhanced beta-catenin expression is associated with recurrence of papillary thyroid carcinoma. Endocr. Pract. 24(5), 411–418 (2018)CrossRefGoogle Scholar
  47. 47.
    X. Liu, T. Zhang, G. Zhu, M. Xing, Regulation of mutant TERT by BRAF V600E/MAP kinase pathway through FOS/GABP in human cancer. Nat. Commun. 9, 579 (2018)CrossRefGoogle Scholar
  48. 48.
    S.L. Asa, S. Ezzat, The epigenetic landscape of differentiated thyroid cancer. Mol. Cell. Endocrinol. 469, 3–10 (2018)CrossRefGoogle Scholar
  49. 49.
    D.B. Doroshow, J.P. Eder, P.M. LoRusso, BET inhibitors: a novel epigenetic approach. Ann. Oncol. 28(8), 1776–1787 (2017)CrossRefGoogle Scholar
  50. 50.
    I. Ali, G. Choi, K. Lee, BET inhibitors as anticancer agents: a patent review. Recent Pat Anticancer Drug Discov. 12(4), 340–364 (2017)CrossRefGoogle Scholar
  51. 51.
    C. Mio, K. Conzatti, F. Baldan, L. Allegri, M. Sponziello, F. Rosignolo, D. Russo, S. Filetti, G. Damante, BET bromodomain inhibitor JQ1 modulates microRNA expression in thyroid cancer cells. Oncol. Rep. 39, 582–588 (2018)Google Scholar

Copyright information

© Springer Science+Business Media, LLC, part of Springer Nature 2019

Authors and Affiliations

  • Valentina Maggisano
    • 1
  • Marilena Celano
    • 1
  • Saverio Massimo Lepore
    • 1
  • Marialuisa Sponziello
    • 2
  • Francesca Rosignolo
    • 2
  • Valeria Pecce
    • 2
  • Antonella Verrienti
    • 2
  • Federica Baldan
    • 2
  • Catia Mio
    • 3
  • Lorenzo Allegri
    • 3
  • Marianna Maranghi
    • 2
  • Rosa Falcone
    • 2
  • Giuseppe Damante
    • 3
  • Diego Russo
    • 1
    Email author
  • Stefania Bulotta
    • 1
  1. 1.Department of Health Sciences“Magna Graecia” University of CatanzaroCatanzaroItaly
  2. 2.Department of Translational and Precision Medicine“Sapienza” University of RomeRomeItaly
  3. 3.Department of Medical AreaUniversity of UdineUdineItaly

Personalised recommendations