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BMC Cancer
Published in: BMC Cancer 1/2016

Open Access 01-12-2016 | Research article

Salivary DNA methylation panel to diagnose HPV-positive and HPV-negative head and neck cancers

Authors: Yenkai Lim, Yunxia Wan, Dimitrios Vagenas, Dmitry A. Ovchinnikov, Chris F. L. Perry, Melissa J. Davis, Chamindie Punyadeera

Published in: BMC Cancer | Issue 1/2016

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Abstract

Background

Head and neck squamous cell carcinoma (HNSCC) is a heterogeneous group of tumours with a typical 5 year survival rate of <40 %. DNA methylation in tumour-suppressor genes often occurs at an early stage of tumorigenesis, hence DNA methylation can be used as an early tumour biomarker. Saliva is an ideal diagnostic medium to detect early HNSCC tumour activities due to its proximity to tumour site, non-invasiveness and ease of sampling. We test the hypothesis that the surveillance of DNA methylation in five tumour-suppressor genes (RASSF1α, p16 INK4a , TIMP3, PCQAP/MED15) will allow us to diagnose HNSCC patients from a normal healthy control group as well as to discriminate between Human Papillomavirus (HPV)-positive and HPV-negative patients.

Methods

Methylation-specific PCR (MSP) was used to determine the methylation levels of RASSF1α, p16 INK4a , TIMP3 and PCQAP/MED15 in DNA isolated from saliva. Statistical analysis was carried out using non-parametric Mann-Whitney’s U-test for individually methylated genes. A logistic regression analysis was carried out to determine the assay sensitivity when combing the five genes. Further, a five-fold cross-validation with a bootstrap procedure was carried out to determine how well the panel will perform in a real clinical scenario.

Results

Salivary DNA methylation levels were not affected by age. Salivary DNA methylation levels for RASSF1α, p16 INK4a , TIMP3 and PCQAP/MED15 were higher in HPV-negative HNSCC patients (n = 88) compared with a normal healthy control group (n = 122) (sensitivity of 71 % and specificity of 80 %). Conversely, DNA methylation levels for these genes were lower in HPV-positive HNSCC patients (n = 45) compared with a normal healthy control group (sensitivity of 80 % and specificity of 74 %), consistent with the proposed aetiology of HPV-positive HNSCCs.

Conclusions

Salivary DNA tumour-suppressor methylation gene panel has the potential to detect early-stage tumours in HPV-negative HNSCC patients. HPV infection was found to deregulate the methylation levels in HPV-positive HNSCC patients. Large-scale double-blinded clinical trials are crucial before this panel can potentially be integrated into a clinical setting.
Appendix
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Literature
1.
2.
Lim Y, et al. Salivary epigenetic biomarkers in head and neck squamous cell carcinomas. Biomark Med. 2016;10(3):301–13.CrossRefPubMed
3.
Chang MC, et al. Cell-mediated immunity and head and neck cancer: with special emphasis on betel quid chewing habit. Oral Oncol. 2005;41(8):757–75.CrossRefPubMed
4.
5.
Pfister DG, Fury MG. New chapter in our understanding of human papillomavirus-related head and neck cancer. J Clin Oncol. 2014;32(30):3349–52.CrossRefPubMed
6.
7.
Syrjanen S. The role of human papillomavirus infection in head and neck cancers. Ann Oncol. 2010;21 Suppl 7:vii243–5.PubMed
8.
9.
Gillison ML, et al. Distinct risk factor profiles for human papillomavirus type 16-positive and human papillomavirus type 16-negative head and neck cancers. J Natl Cancer Inst. 2008;100(6):407–20.CrossRefPubMed
10.
Ragin CC, Modugno F, Gollin SM. The epidemiology and risk factors of head and neck cancer: a focus on human papillomavirus. J Dent Res. 2007;86(2):104–14.CrossRefPubMed
11.
12.
13.
Warnakulasuriya S. Global epidemiology of oral and oropharyngeal cancer. Oral Oncol. 2009;45(4–5):309–16.CrossRefPubMed
14.
Hall SF, et al. TNM-based stage groupings in head and neck cancer: application in cancer of the hypopharynx. Head Neck. 2009;31(1):1–8.CrossRefPubMed
15.
Price KA, Cohen EE. Current treatment options for metastatic head and neck cancer. Curr Treat Options Oncol. 2012;13(1):35–46.CrossRefPubMed
16.
Villa A, Villa C, Abati S. Oral cancer and oral erythroplakia: an update and implication for clinicians. Aust Dent J. 2011;56(3):253–6.CrossRefPubMed
17.
Mehanna H, et al. Head and neck cancer—Part 1: epidemiology, presentation, and prevention. BMJ. 2010;341:c4684.CrossRefPubMed
18.
Iorgulescu G. Saliva between normal and pathological. Important factors in determining systemic and oral health. J Med Life. 2009;2(3):303–7.PubMedPubMedCentral
19.
Pfaffe T, et al. Diagnostic potential of saliva: current state and future applications. Clin Chem. 2011;57(5):675–87.CrossRefPubMed
20.
21.
na. Head and neck cancer biomarkers detected in saliva. Cancer Biol Ther. 2004. 4(1):6–12.
22.
23.
Topkas E, et al. Evaluation of saliva collection devices for the analysis of proteins. Clin Chim Acta. 2012;413(13–14):1066–70.CrossRefPubMed
24.
Zhang X, Dimeski G, Punyadeera C. Validation of an immunoassay to measure plasminogen-activator inhibitor-1 concentrations in human saliva. Biochem Med. 2014;24(2):258–65.CrossRef
25.
Ovchinnikov DA, et al. DNA methylation at the novel CpG sites in the promoter of MED15/PCQAP gene as a biomarker for head and neck cancers. Biomark Insights. 2014;9:53–60.PubMedPubMedCentral
26.
27.
Worsham MJ, et al. Delineating an epigenetic continuum in head and neck cancer. Cancer Lett. 2014;342(2):178–84.CrossRefPubMed
28.
Carvalho AL, et al. Detection of promoter hypermethylation in salivary rinses as a biomarker for head and neck squamous cell carcinoma surveillance. Clin Cancer Res. 2011;17(14):4782–9.CrossRefPubMedPubMedCentral
29.
30.
Tollefsbol TO, et al. Epigenetics protocols. New York: Humana Press; 2011.
31.
Huang Z, Bassil C, Murphy S. Methylation-specific PCR. In: Malek A, Tchernitsa O, editors. Ovarian cancer. New York: Humana Press; 2013. p. 75–82.
32.
Chai RC, et al. A pilot study to compare the detection of HPV-16 biomarkers in salivary oral rinses with tumour p16INK4a expression in head and neck squamous cell carcinoma patients. BMC Cancer. 2016;16(1):1–8.CrossRef
33.
Divine KK, et al. Nested multigene MSP/DHPLC method for analyzing promoter hypermethylation status in clinical samples. BioTechniques. 2006. 40(1):40, 42, 44 passim.
34.
Righini CA, et al. Tumor-specific methylation in saliva: a promising biomarker for early detection of head and neck cancer recurrence. Clin Cancer Res. 2007;13(4):1179–85.CrossRefPubMed
35.
Eads CA, et al. Epigenetic patterns in the progression of esophageal adenocarcinoma. Cancer Res. 2001;61(8):3410–8.PubMed
36.
Jiang L, et al. CAL 27 is an oral adenosquamous carcinoma cell line. Oral Oncol. 2009;45(11):e204–7.CrossRefPubMed
37.
38.
Kohavi R. A study of cross-validation and bootstrap for accuracy estimation and model selection. In: Proceedings of the 14th international joint conference on Artificial intelligence, vol. 2. Montreal: Morgan Kaufmann Publishers Inc; 1995. p. 1137–43.
39.
Geisser S. Predictive inference. New York: Chapman and Hall; 1993.
40.
Devijver PA. Pattern recognition: a statistical approach. London: Prentice-Hall; 1982.
41.
Tang K-W, et al. The landscape of viral expression and host gene fusion and adaptation in human cancer. Nat Commun. 2013;4:2513.PubMedPubMedCentral
42.
43.
44.
Arantes LM, et al. Methylation as a biomarker for head and neck cancer. Oral Oncol. 2014;50(6):587–92.CrossRefPubMed
45.
Richards KL, et al. Genome-wide hypomethylation in head and neck cancer is more pronounced in HPV-negative tumors and is associated with genomic instability. PLoS One. 2009;4(3):e4941.CrossRefPubMedPubMedCentral
46.
47.
Dong SM, et al. Epigenetic inactivation of RASSF1A in head and neck cancer. Clin Cancer Res. 2003;9(10):3635–40.PubMed
48.
Demokan S, et al. Promoter methylation and loss of p16(INK4a) gene expression in head and neck cancer. Head Neck. 2012;34(10):1470–5.CrossRefPubMed
49.
Guan Z, et al. Promoter methylation and expression of TIMP3 gene in gastric cancer. Diagn Pathol. 2013;8(1):1–6.
50.
51.
Sandhu HK, et al. An association study of PCQAP polymorphisms and schizophrenia. Psychiatr Genet. 2004;14(3):169–72.CrossRefPubMed
52.
53.
Zhao M, et al. Mediator MED15 modulates transforming growth factor beta (TGFbeta)/Smad signaling and breast cancer cell metastasis. J Mol Cell Biol. 2013;5(1):57–60.CrossRefPubMed
54.
Guo SX, et al. Hypermethylation of p16 and p15 genes and RB protein expression in acute leukemia. Leuk Res. 2000;24(1):39–46.CrossRefPubMed
55.
Calmon MF, et al. Methylation profile of genes CDKN2A (p14 and p16), DAPK1, CDH1, and ADAM23 in head and neck cancer. Cancer Genet Cytogenet. 2007;173(1):31–7.CrossRefPubMed
56.
Sanchez-Cespedes M, et al. Gene promoter hypermethylation in tumors and serum of head and neck cancer patients. Cancer Res. 2000;60(4):892–5.PubMed
57.
58.
Meng W, et al. Combined RASSF1A and RASSF2A promoter methylation analysis as diagnostic biomarker for bladder cancer. Mol Biol Int. 2012;2012:8.CrossRef
59.
60.
Xu J, et al. Methylation of HIN-1, RASSF1A, RIL and CDH13 in breast cancer is associated with clinical characteristics, but only RASSF1A methylation is associated with outcome. BMC Cancer. 2012;12:243.CrossRefPubMedPubMedCentral
61.
Shi DT, et al. Association of RASSF1A promoter methylation with gastric cancer risk: a meta-analysis. Tumour Biol. 2014;35(2):943–8.CrossRefPubMed
62.
Wang HL, et al. Aberrant promoter methylation of RASSF1A gene may be correlated with colorectal carcinogenesis: a meta-analysis. Mol Biol Rep. 2014;41(6):3991–9.CrossRefPubMed
63.
Gao T, et al. The association of RAS Association Domain Family Protein1A (RASSF1A) methylation states and bladder cancer risk: a systematic review and meta-analysis. PLoS One. 2012;7(11):e48300.CrossRefPubMedPubMedCentral
64.
65.
Koutsimpelas D, et al. Promoter methylation of MGMT, MLH1 and RASSF1A tumor suppressor genes in head and neck squamous cell carcinoma: pharmacological genome demethylation reduces proliferation of head and neck squamous carcinoma cells. Oncol Rep. 2012;27(4):1135–41.PubMedPubMedCentral
66.
67.
Konig F, et al. Relation between human papillomavirus positivity and p16 expression in head and neck carcinomas--a tissue microarray study. Anticancer Res. 2007;27(1a):283–8.PubMed
68.
69.
Boscolo-Rizzo P, et al. New insights into human papillomavirus-associated head and neck squamous cell carcinoma. Acta Otorhinolaryngol Ital. 2013;33(2):77–87.PubMedPubMedCentral
70.
De Schutter H, et al. Promoter methylation of TIMP3 and CDH1 predicts better outcome in head and neck squamous cell carcinoma treated by radiotherapy only. Oncol Rep. 2009;21(2):507–13.PubMed
71.
Rettori MM, et al. Prognostic significance of TIMP3 hypermethylation in post-treatment salivary rinse from head and neck squamous cell carcinoma patients. Carcinogenesis. 2013;34(1):20–7.CrossRefPubMed
72.
Rettori MM, et al. TIMP3 and CCNA1 hypermethylation in HNSCC is associated with an increased incidence of second primary tumors. J Transl Med. 2013;11:316.CrossRefPubMedPubMedCentral
73.
Sun W, et al. Detection of TIMP3 promoter hypermethylation in salivary rinse as an independent predictor of local recurrence-free survival in head and neck cancer. Clin Cancer Res. 2012;18(4):1082–91.CrossRefPubMedPubMedCentral
74.
Swift-Scanlan T, et al. Two-color quantitative multiplex methylation-specific PCR. Biotechniques. 2006;40(2):210–9.CrossRefPubMed
Metadata
Title
Salivary DNA methylation panel to diagnose HPV-positive and HPV-negative head and neck cancers
Authors
Yenkai Lim
Yunxia Wan
Dimitrios Vagenas
Dmitry A. Ovchinnikov
Chris F. L. Perry
Melissa J. Davis
Chamindie Punyadeera
Publication date
01-12-2016
Publisher
BioMed Central
Published in
BMC Cancer / Issue 1/2016
Electronic ISSN: 1471-2407
DOI
https://doi.org/10.1186/s12885-016-2785-0

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