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. 2017 Aug 30;17(1):601.
doi: 10.1186/s12885-017-3582-0.

LINE-1 hypermethylation in white blood cell DNA is associated with high-grade cervical intraepithelial neoplasia

Martina Barchitta  1 , Annalisa Quattrocchi  1 , Andrea Maugeri  1 , Carolina Canto  2 , Nadia La Rosa  3 , Maria Antonietta Cantarella  3 , Giuseppa Spampinato  3 , Aurora Scalisi  3 , Antonella Agodi  4
Affiliations

LINE-1 hypermethylation in white blood cell DNA is associated with high-grade cervical intraepithelial neoplasia

Martina Barchitta et al. BMC Cancer. .

Abstract

Background: Long Interspersed Nuclear Elements-1 (LINEs-1) methylation from white blood cells (WBCs) DNA has been proposed as biomarker associated with different types of cancer. The aim of the present study was to investigate the degree of WBCs LINE-1 methylation, according to high-risk Human Papilloma Virus (hrHPV) status in a healthy population, and the association with high-grade Cervical Intraepithelial Neoplasia (CIN2+) in hrHPV positive women.

Methods: Women with abnormal cervical cells were enrolled and classified by histological diagnosis and hrHPV infection. A structured questionnaire was used to obtain information on socio-demographic variables and lifestyle factors. LINE-1 methylation level in WBCs was measured by pyrosequencing-based methylation analysis after bisulfite conversion.

Results: Among 252 women diagnosed with normal cervical epithelium, with regard to LINE-1 methylation level no significant difference was observed between hrHPV positive and hrHPV negative women, also adjusting for known risk factors of infection. The association between WBCs LINE-1 methylation and CIN2+ status was analyzed in hrHPV positive women. The median value of LINE-1 methylation levels was higher in cases (CIN2+) than in controls (75.00% versus 73.17%; p = 0.002). For a one-unit increase in LINE-1 methylation level, the odds of being diagnosed with CIN2+ increased by 10%, adjusting for known factors related to LINE-1 methylation (adjOR: 1.10; 95% CI:1.01-1.20; p = 0.032). The Receiver-Operating Characteristic (ROC) curve analysis identified the cut-off value of 73.8% as the best threshold to separate cases from controls (sensitivity: 63.4% and specificity: 61.8%).

Conclusions: LINE-1 methylation status in WBCs DNA may represent a cost-effective and tissue-accessible biomarker for high-grade CIN in hrHPV positive women. However, LINE-1 hypermethylation cannot be considered specific for cervical cancer (CC) and a model based solely on LINE-1 methylation levels has limited performance. Further investigations are necessary to propose and validate a novel methylation biomarker panel, based on LINE-1 methylation and other differentially methylated regions, for the screening of women at risk of CC.

Keywords: Cervical Intraepitelial Neoplasia; Cervical cancer; Global DNA methylation; Hypermethylation; LINE-1 methylation; Prevention; Pyrosequencing-based methylation analysis; ROC curve analysis.

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Conflict of interest statement

Ethics approval and consent to participate

The study protocol was approved by the ethics committee of the involved Institution (CE Catania 2; Prot. N. 227/BE and 275/BE) and performed according to the Declaration of Helsinki. Participants were fully informed of the purpose and procedures of the study, and a signed written consent was obtained.

Consent for publication

Not Applicable.

Competing interests

Carolina Canto is an employee of Oncopath s.r.l.; the other authors declare that they have no competing interests.

Publisher’s Note

Springer Nature remains neutral with regard to jurisdictional claims in published maps and institutional affiliations.

Figures

Fig. 1
Fig. 1
Methylation levels of LINE-1 in cases (CIN2+) and controls (≤CIN1). Mean methylation levels of LINE-1 sequences (mean percentage of methylation of the three evaluated CpG sites) in CIN2+ patients (cases) and in CIN 1 or normal cervical epithelium patients (controls) obtained using pyrosequencing of bisulfite converted DNA from WBCs (p-value = 0.002, based on the Mann-Whitney U test)
Fig. 2
Fig. 2
ROC curve analysis of LINE-1 methylation and CIN2+ detection. ROC (Receiver Operator Characteristics) curve of LINE-1 methylation levels for the detection of CIN2+. LINE-1 methylation level was suitable for detecting CIN2+ with an AUC of 0.652 (95% CI = 0.560–0.744). The cut-off value of 73.83% is the best threshold to separate cases from controls
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