Cellular Oncology

, Volume 39, Issue 6, pp 573–582 | Cite as

WT1, MSH6, GATA5 and PAX5 as epigenetic oral squamous cell carcinoma biomarkers - a short report

  • Ilda Patrícia Ribeiro
  • Francisco Caramelo
  • Francisco Marques
  • Ana Domingues
  • Margarida Mesquita
  • Leonor Barroso
  • Hugo Prazeres
  • Maria José Julião
  • Isabel Poiares Baptista
  • Artur Ferreira
  • Joana Barbosa Melo
  • Isabel Marques Carreira



Oral squamous cell carcinoma (OSCC) is a frequently occurring aggressive malignancy with a heterogeneous clinical behavior. Based on the paucity of specific early diagnostic and prognostic biomarkers, which hampers the appropriate treatment and, ultimately the development of novel targeted therapies, we aimed at identifying such biomarkers through a genetic and epigenetic analysis of these tumors.


93 primary OSCCs were subjected to DNA copy number alteration (CNA) and methylation status analyses using methylation-specific multiplex ligation-dependent probe amplification (MS-MPLA). The genetic and epigenetic OSCC profiles obtained were associated with the patients’ clinic-pathological features.


We found that WT1 gene promoter methylation is a predictor of a better prognosis and that MSH6 and GATA5 gene promoter methylation serve as predictors of a worse prognosis. GATA5 gene promoter methylation was found to be significantly associated with a shorter survival rate. In addition, we found that PAX5 gene promoter methylation was significantly associated with tongue tumors. To the best of our knowledge, this is the first study that highlights this specific set of genes as epigenetic diagnostic and prognostic biomarkers in OSCC.


Our data highlight the importance of epigenetically assessing OSCCs to identify key genes that may serve as diagnostic and prognostic biomarkers and, potentially, as candidate therapeutic targets.


DNA methylation Copy number gains and losses Predictors of prognosis Oral squamous cell carcinoma 



Ribeiro I.P. is a recipient of a PhD fellowship (SFRH/BD/52290/2013) from the Portuguese Foundation for Science and Technology. This work was in part supported by CIMAGO (Center of Investigation on Environment Genetics and Oncobiology - Faculty of Medicine, University of Coimbra) and by the Portuguese Foundation for Science and Technology (grant: UID/NEU/04539/2013).

Compliance with ethical standards

Conflict of interest

The authors have no conflict of interest to disclose.

Supplementary material

13402_2016_293_MOESM1_ESM.doc (77 kb)
ESM 1 Supplementary Table 1 (DOC 77 kb)


  1. 1.
    L. M. Arantes, A. C. de Carvalho, M. E. Melendez, C. C. Centrone, J. F. Gois-Filho, T. N. Toporcov, D. N. Caly, E. H. Tajara, E. M. Goloni-Bertollo, A. L. Carvalho, And Gencapo, validation of methylation markers for diagnosis of oral cavity cancer. Eur. J. Cancer 51, 632–641 (2015). doi: 10.1016/j.ejca.2015.01.060 CrossRefPubMedGoogle Scholar
  2. 2.
    A.K. Markopoulos, Current aspects on oral squamous cell carcinoma. Open Dent. J. 6, 126–130 (2012) doi: 10.2174/1874210601206010126
  3. 3.
    C. Salazar, R. Nagadia, P. Pandit, J. Cooper-White, N. Banerjee, N. Dimitrova, W. B. Coman, C. Punyadeera, A novel saliva-based microRNA biomarker panel to detect head and neck cancers. Cell. Oncol. 37, 331–338 (2014). doi: 10.1007/s13402-014-0188-2 CrossRefGoogle Scholar
  4. 4.
    K. Mohankumar, S. Pajaniradje, S. Sridharan, V. K. Singh, L. Ronsard, A. C. Banerjea, B. C. Selvanesan, M. S. Coumar, L. Periyasamy, R. Rajagopalan, Apoptosis induction by an analog of curcumin (BDMC-A) in human laryngeal carcinoma cells through intrinsic and extrinsic pathways. Cell. Oncol. 37, 439–454 (2014). doi: 10.1007/s13402-014-0207-3 CrossRefGoogle Scholar
  5. 5.
    D. Pulte, H. Brenner, Changes in survival in head and neck cancers in the late 20th and early twenty-first century: a period analysis. Oncologist 15, 994–1001 (2010). doi: 10.1634/theoncologist.2009-0289 CrossRefPubMedPubMedCentralGoogle Scholar
  6. 6.
    T. Nakaoka, A. Ota, T. Ono, S. Karnan, H. Konishi, A. Furuhashi, Y. Ohmura, Y. Yamada, Y. Hosokawa, Y. Kazaoka, Combined arsenic trioxide-cisplatin treatment enhances apoptosis in oral squamous cell carcinoma cells. Cell. Oncol. 37, 119–129 (2014). doi: 10.1007/s13402-014-0167-7 CrossRefGoogle Scholar
  7. 7.
    Y. Tokumaru, K. Yamashita, M. Osada, S. Nomoto, D. I. Sun, Y. Xiao, M. O. Hoque, W. H. Westra, J. A. Califano, D. Sidransky, Inverse correlation between cyclin A1 hypermethylation and p53 mutation in head and neck cancer identified by reversal of epigenetic silencing. Cancer Res. 64, 5982–5987 (2004). doi: 10.1158/0008-5472.CAN-04-0993 CrossRefPubMedGoogle Scholar
  8. 8.
    C. R. Leemans, B. J. Braakhuis, R. H. Brakenhoff, The molecular biology of head and neck cancer. Nat. Rev. Cancer 11, 9–22 (2011). doi: 10.1038/nrc2982 CrossRefPubMedGoogle Scholar
  9. 9.
    T. Bezabeh, O. Odlum, R. Nason, P. Kerr, D. Sutherland, R. Patel, I. C. Smith, Prediction of treatment response in head and neck cancer by magnetic resonance spectroscopy. AJNR Am. J. Neuroradiol. 26, 2108–2113 (2005)PubMedGoogle Scholar
  10. 10.
    R. J. Nobre, E. Cruz, O. Real, L. P. de Almeida, T. C. Martins, Characterization of common and rare human papillomaviruses in Portuguese women by the polymerase chain reaction, restriction fragment length polymorphism and sequencing. J. Med. Virol. 82, 1024–1032 (2010). doi: 10.1002/jmv.21756 CrossRefPubMedGoogle Scholar
  11. 11.
    A. O. Nygren, N. Ameziane, H. M. Duarte, R. N. Vijzelaar, Q. Waisfisz, C. J. Hess, J. P. Schouten, A. Errami, Methylation-specific MLPA (MS-MLPA): simultaneous detection of CpG methylation and copy number changes of up to 40 sequences. Nucleic Acids Res. 33, e128 (2005). doi: 10.1093/nar/gni127 CrossRefPubMedPubMedCentralGoogle Scholar
  12. 12.
    I. P. Ribeiro, F. Marques, F. Caramelo, J. Ferrao, H. Prazeres, M. J. Juliao, W. Rifi, S. Savola, J. B. de Melo, I. P. Baptista, I. M. Carreira, Genetic imbalances detected by multiplex ligation-dependent probe amplification in a cohort of patients with oral squamous cell carcinoma-the first step towards clinical personalized medicine. Tumour Biol. 35, 4687–4695 (2014). doi: 10.1007/s13277-014-1614-9 PubMedGoogle Scholar
  13. 13.
    I. P. Ribeiro, F. Marques, F. Caramelo, J. Pereira, M. Patricio, H. Prazeres, J. Ferrao, M. J. Juliao, M. Castelo-Branco, J. B. de Melo, I. P. Baptista, I. M. Carreira, Genetic gains and losses in oral squamous cell carcinoma: impact on clinical management. Cell. Oncol. 37, 29–39 (2014). doi: 10.1007/s13402-013-0161-5 CrossRefGoogle Scholar
  14. 14.
    M. Chmelarova, E. Dvorakova, J. Spacek, J. Laco, M. Mzik, V. Palicka, Promoter methylation of GATA4, WIF1, NTRK1 and other selected tumour suppressor genes in ovarian cancer. Folia Biol. 59, 87–92 (2013)Google Scholar
  15. 15.
    S. Maruya, J. P. Issa, R. S. Weber, D. I. Rosenthal, J. C. Haviland, R. Lotan, A. K. El-Naggar, Differential methylation status of tumor-associated genes in head and neck squamous carcinoma: incidence and potential implications. Clin. Cancer Res. 10, 3825–3830 (2004). doi: 10.1158/1078-0432.CCR-03-0370 CrossRefPubMedGoogle Scholar
  16. 16.
    S. Sharma, T. K. Kelly, P. A. Jones, Epigenetics in cancer. Carcinogenesis 31, 27–36 (2010). doi: 10.1093/carcin/bgp220 CrossRefPubMedGoogle Scholar
  17. 17.
    A. S. Ho, S. Turcan, T. A. Chan, Epigenetic therapy: use of agents targeting deacetylation and methylation in cancer management. Onco Targets Ther. 6, 223–232 (2013). doi: 10.2147/OTT.S34680 PubMedPubMedCentralGoogle Scholar
  18. 18.
    M. J. Worsham, J. K. Stephen, K. M. Chen, S. Havard, V. Shah, G. Gardner, V. G. Schweitzer, Delineating an epigenetic continuum in head and neck cancer. Cancer Lett. 342, 178–184 (2014). doi: 10.1016/j.canlet.2012.02.018 CrossRefPubMedGoogle Scholar
  19. 19.
    M. Federico and L. Bagella, Histone deacetylase inhibitors in the treatment of hematological malignancies and solid tumors. J. Biomed. Biotechnol. 2011, 475641 (2011) doi: 10.1155/2011/475641
  20. 20.
    Y. Oji, H. Yamamoto, M. Nomura, Y. Nakano, A. Ikeba, S. Nakatsuka, S. Abeno, E. Kiyotoh, T. Jomgeow, M. Sekimoto, R. Nezu, Y. Yoshikawa, Y. Inoue, N. Hosen, M. Kawakami, A. Tsuboi, Y. Oka, H. Ogawa, S. Souda, K. Aozasa, M. Monden, H. Sugiyama, Overexpression of the Wilms’ tumor gene WT1 in colorectal adenocarcinoma. Cancer Sci. 94, 712–717 (2003)CrossRefPubMedGoogle Scholar
  21. 21.
    H. Sugiyama, WT1 (Wilms’ tumor gene 1): biology and cancer immunotherapy. Jpn. J. Clin. Oncol. 40, 377–387 (2010) doi: 10.1093/jjco/hyp194
  22. 22.
    L. Yang, Y. Han, F. Suarez Saiz, M. D. Minden, A tumor suppressor and oncogene: the WT1 story. Leukemia 21, 868–876 (2007). doi: 10.1038/sj.leu.2404624 PubMedGoogle Scholar
  23. 23.
    X. W. Qi, F. Zhang, H. Wu, J. L. Liu, B. G. Zong, C. Xu, J. Jiang, Wilms’ tumor 1 (WT1) expression and prognosis in solid cancer patients: a systematic review and meta-analysis. Sci. Rep. 5, 8924 (2015). doi: 10.1038/srep08924 CrossRefPubMedPubMedCentralGoogle Scholar
  24. 24.
    S. Yip, J. Miao, D. P. Cahill, A. J. Iafrate, K. Aldape, C. L. Nutt, D. N. Louis, MSH6 mutations arise in glioblastomas during temozolomide therapy and mediate temozolomide resistance. Clin. Cancer Res. 15, 4622–4629 (2009). doi: 10.1158/1078-0432.CCR-08-3012 CrossRefPubMedPubMedCentralGoogle Scholar
  25. 25.
    I. Peters, N. Dubrowinskaja, M. Kogosov, M. Abbas, J. Hennenlotter, C. von Klot, A. S. Merseburger, A. Stenzl, R. Scherer, M. A. Kuczyk, J. Serth, Decreased GATA5 mRNA expression associates with CpG island methylation and shortened recurrence-free survival in clear cell renal cell carcinoma. BMC Cancer 14, 101 (2014). doi: 10.1186/1471-2407-14-101 CrossRefPubMedPubMedCentralGoogle Scholar
  26. 26.
    I. Peters, K. Gebauer, N. Dubrowinskaja, F. Atschekzei, M. W. Kramer, J. Hennenlotter, H. Tezval, M. Abbas, R. Scherer, A. S. Merseburger, A. Stenzl, M. A. Kuczyk, J. Serth, GATA5 CpG island hypermethylation is an independent predictor for poor clinical outcome in renal cell carcinoma. Oncol. Rep. 31, 1523–1530 (2014). doi: 10.3892/or.2014.3030 PubMedPubMedCentralGoogle Scholar
  27. 27.
    R. Zheng, G. A. Blobel, G. A. T. A. Transcription Factors, And cancer. Genes Cancer 1, 1178–1188 (2010). doi: 10.1177/1947601911404223 CrossRefPubMedPubMedCentralGoogle Scholar
  28. 28.
    R. Guerrero-Preston, C. Michailidi, L. Marchionni, C. R. Pickering, M. J. Frederick, J. N. Myers, S. Yegnasubramanian, T. Hadar, M. G. Noordhuis, V. Zizkova, E. Fertig, N. Agrawal, W. Westra, W. Koch, J. Califano, V. E. Velculescu, D. Sidransky, Key tumor suppressor genes inactivated by "greater promoter" methylation and somatic mutations in head and neck cancer. Epigenetics 9, 1031–1046 (2014). doi: 10.4161/epi.29025 CrossRefPubMedPubMedCentralGoogle Scholar
  29. 29.
    M. J. Worsham, H. Ali, J. Dragovic, V. P. Schweitzer, Molecular characterization of head and neck cancer: how close to personalized targeted therapy? Mol. Diagn. Ther. 16, 209–222 (2012). doi: 10.2165/11635330-000000000-00000 CrossRefPubMedPubMedCentralGoogle Scholar
  30. 30.
    S. I. Pai, W. H. Westra, Molecular pathology of head and neck cancer: implications for diagnosis, prognosis, and treatment. Annu. Rev. Pathol. 4, 49–70 (2009)CrossRefPubMedPubMedCentralGoogle Scholar
  31. 31.
    K. T. Robbins, G. Clayman, P. A. Levine, J. Medina, R. Sessions, A. Shaha, P. Som, G. T. Wolf, H. American, S. Neck, O.-H. American Academy, Of and S. Neck, neck dissection classification update: revisions proposed by the American head and neck society and the American Academy of Otolaryngology-Head and Neck Surgery. Arch. Otolaryngol. Head Neck Surg. 128, 751–758 (2002)CrossRefGoogle Scholar
  32. 32.
    Cancer Genome Atlas Network, comprehensive genomic characterization of head and neck squamous cell carcinomas. Nature 517, 576–582 (2015) doi: 10.1038/nature14129
  33. 33.
    S. B. Chinn, J. N. Myers, Oral cavity carcinoma: current management, controversies, and future directions. J. Clin. Oncol. 33, 3269–3276 (2015). doi: 10.1200/JCO.2015.61.2929 CrossRefPubMedGoogle Scholar

Copyright information

© International Society for Cellular Oncology 2016

Authors and Affiliations

  • Ilda Patrícia Ribeiro
    • 1
    • 2
  • Francisco Caramelo
    • 3
  • Francisco Marques
    • 2
    • 4
    • 5
  • Ana Domingues
    • 1
  • Margarida Mesquita
    • 6
  • Leonor Barroso
    • 6
  • Hugo Prazeres
    • 7
  • Maria José Julião
    • 8
  • Isabel Poiares Baptista
    • 2
    • 4
  • Artur Ferreira
    • 6
  • Joana Barbosa Melo
    • 1
    • 2
  • Isabel Marques Carreira
    • 1
    • 2
  1. 1.Cytogenetics and Genomics Laboratory, Faculty of MedicineUniversity of CoimbraCoimbraPortugal
  2. 2.CIMAGO - Center of Investigation on Environment Genetics and Oncobiology - Faculty of MedicineUniversity of CoimbraCoimbraPortugal
  3. 3.Laboratory of Biostatistics and Medical Informatics, IBILI - Faculty of MedicineUniversity of CoimbraCoimbraPortugal
  4. 4.Department of Dentistry, Faculty of MedicineUniversity of CoimbraCoimbraPortugal
  5. 5.Stomatology UnitCoimbra Hospital and University Centre (CHUC)CoimbraPortugal
  6. 6.Maxillofacial Surgery Department, Coimbra Hospital and University Centre (CHUC)CoimbraPortugal
  7. 7.Molecular Pathology LaboratoryPortuguese Institute of Oncology of Coimbra FGCoimbraPortugal
  8. 8.Department of PathologyCoimbra Hospital and University Centre (CHUC)CoimbraPortugal

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