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Prediction of Human Papillomavirus 16 E6 Gene Expression and Cervical Intraepithelial Neoplasia Progression by Methylation Status
  1. Pavla Hublarova, MSc*,
  2. Roman Hrstka, PhD*,
  3. Pavla Rotterova, PhD,§,
  4. Leopold Rotter, MD,§,
  5. Marie Coupkova, MD,
  6. Vinay Badal, PhD,
  7. Rudolf Nenutil, PhD* and
  8. Borivoj Vojtesek, DrSci*
  1. * Department of Oncological and Experimental Pathology, Masaryk Memorial Cancer Institute;
  2. 1st Pathological-Anatomical Institute, St Ann's Faculty Hospital Brno;
  3. Gynecological-Obstetric Clinic, Faculty Hospital Brno;
  4. § Sanatorium Helios;
  5. Gynecological Ambulance, Brno, Czech Republic; and
  6. Laboratory of DNA Repair and DNA Methylation in Chemical Carcinogenesis, Institute of Molecular and Cell Biology, Proteos, Singapore.
  1. Address correspondence and reprint requests to Borivoj Vojtesek, DrSci, Department of Oncological and Experimental Pathology, Masaryk Memorial Cancer Institute, Zluty kopec 7, 656 53 Brno, Czech Republic. E-mail: vojtesek{at}


Introduction: Human papillomavirus (HPV) infection represents the most important risk factor for the development of cervical intraepithelial neoplasia (CIN) and cervical cancer. We aimed to analyze the consequences of methylation of the E6 gene promoter in distinct stages of HPV-16-induced cellular transformation to assess its importance for disease progression.

Methods: Human papillomavirus 16 was detected by sensitive polymerase chain reaction (PCR). Determination of E6 gene promoter methylation was analyzed by digestion with specific restriction endonuclease McrBC followed by PCR amplification. Expression of the E6 gene was determined by quantitative real-time PCR.

Results: Of 103 cervical smears from asymptomatic women with no cytological and colposcopic abnormalities, 20.4% were HPV-16-positive. Human papillomavirus 16 was present in 44.4% of 18 patients with CIN I, in 62.2% of 143 patients with CIN II/III, and in 74.2% of 31 cervix carcinoma specimens. The incidence of HPV-16 in all lesions compared with asymptomatic women was statistically significant (P < 0.001, Pearson χ2 test). Methylation was detected in 81% (n = 21) of HPV-16-positive asymptomatic smears compared with 62.5% in CIN I (n = 8), 31.5% (n = 89) in CIN II/III, and 43.4% (n = 23) in carcinomas; a statistical significance between lesions and healthy women was found (P < 0.001, Pearson χ2 test). Expression of E6 mRNA correlated with methylation status (P = 0.010, Mann-Whitney U test).

Conclusions: We conclude that methylation of the E6 gene promoter in HPV-16 genome is a predictive biomarker for cervical cancer progression by regulating the expression of the E6 oncogene.

  • Cervix
  • Cervical intraepithelial neoplasia
  • HPV-16
  • E6 gene
  • Promoter methylation

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