Prediction of human papillomavirus 16 e6 gene expression and cervical intraepithelial neoplasia progression by methylation status.
BACKGROUND
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 chi 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 chi 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.
Hublarova
Pavla
P
Department of Oncological and Experimental Pathology, Masaryk Memorial Cancer Institute, Brno, Czech Republic.
Hrstka
Roman
R
Rotterova
Pavla
P
Rotter
Leopold
L
Coupkova
Marie
M
Badal
Vinay
V
Nenutil
Rudolf
R
Vojtesek
Borivoj
B
D000293:Adolescent; D000328:Adult; D000368:Aged; D002471:Cell Transformation, Neoplastic; D019175:DNA Methylation; D004279:DNA, Viral / Q000032:analysis; D018450:Disease Progression; D005260:Female; D052162:Human papillomavirus 16 / Q000235:genetics / Q000378:metabolism; D006801:Humans; D008875:Middle Aged; D009856:Oncogene Proteins, Viral / Q000235:genetics*; D030361:Papillomavirus Infections / Q000235:genetics* / Q000473:pathology / Q000821:virology; D011379:Prognosis; D011401:Promoter Regions, Genetic / Q000235:genetics; D012333:RNA, Messenger / Q000235:genetics / Q000378:metabolism; D012097:Repressor Proteins / Q000235:genetics*; D020133:Reverse Transcriptase Polymerase Chain Reaction; D002583:Uterine Cervical Neoplasms / Q000235:genetics* / Q000473:pathology / Q000821:virology; D055815:Young Adult; D002578:Uterine Cervical Dysplasia / Q000235:genetics* / Q000473:pathology / Q000821:virology
D004279:DNA, Viral; C068651:E6 protein, Human papillomavirus type 16; D009856:Oncogene Proteins, Viral; D012333:RNA, Messenger; D012097:Repressor Proteins