Methylation of CpG 5962 in L1 of the human papillomavirus 16 genome as a potential predictive marker for viral persistence: A prospective large cohort study using cervical swab samples

Abstract

Several studies have demonstrated that the viral genome can be methylated by the host cell during progression from persistent infection to cervical cancer. The aim of this study was to investigate whether methylation at a specific site could predict the development of viral persistence and whether viral load shows a correlation with specific methylation patterns. HPV16-positive samples from women aged 20-29 years (n = 99) with a follow-up time of 13 years, were included from a Danish cohort comprising 11 088 women. Viral load was measured by real-time PCR and methylation status was determined for 39 CpG sites in the upstream regulatory region (URR), E6/E7, and L1 region of HPV16 by next-generation sequencing. Participants were divided into two groups according to whether they were persistently (≥ 24 months) or transiently HPV16 infected. The general methylation status was significantly different between women with a persistent and women with a transient infection outcome (P = .025). One site located in L1 (nt. 5962) was statistically significantly (P = .00048) different in the methylation status after correction using the Holm-Sidak method (alpha = 0.05). Correlation analyses of samples from HPV16 persistently infected women suggest that methylation is higher although viral load is lower.This study indicates that methylation at position 5962 of the HPV16 genome within the L1 gene might be a predictive marker for the development of a persistent HPV16 infection. K E Y W O R D SBiomarker, DNA Methylation, HPV16, Persistence

Risk stratification of HPV 16 DNA methylation combined with E6 oncoprotein in cervical cancer screening: a 10-year prospective cohort study

“…With regard to specific CpG sites of HPV 16 with good predictive utility in our study, CpG 5602 in L1 gene had won the top AUC value in the detection of CIN3/AIS in Clarke et al's nested case-control study reaching 0.84 [18]; CpG 6650 and 7034 in L1 gene in two studies of Chinese population also showed strong 16 positive women in our current study [31]. Albeit with the consensus on the role of LCR region in regulating viral gene expression [29,30], it remains undetermined which methylation patterns, hypomethylation or hypermethylation, in this region are associated with the risk of cervical cancer [25,27,31,32]. Our current study showed that high methylations in CpG 31, 37, or 43 were associated with CIN3+ risk, which were consistent with Sun's finding [27], but different from Fertey's finding s [32].…”

“…Albeit with the consensus on the role of LCR region in regulating viral gene expression [29,30], it remains undetermined which methylation patterns, hypomethylation or hypermethylation, in this region are associated with the risk of cervical cancer [25,27,31,32]. Our current study showed that high methylations in CpG 31, 37, or 43 were associated with CIN3+ risk, which were consistent with Sun's finding [27], but different from Fertey's finding s [32]. These discrepancies among these observations may partly result from study design (e.g., prospective cohort, nested case-control, or cross-sectional study), methylation assays (e.g., pyrosequencing or next-generation bisulfite sequencing), study population, and/or disease endpoints (e.g., CIN2+ versus CIN3 in our study).…”

The role of HPV‑induced epigenetic changes in cervical carcinogenesis (Review)

“…It was revealed that the methylation status of the viral genome at position 5,962, corresponding to the L1 gene was significantly higher in samples of the virus isolated from women with persistent infection compared with those isolated from women with transient infection. This suggests that hypermethylation of the L1 gene of HPV16 is associated with viral persistence ( 62 ). A similar result has also been reported in another study, in which a high level of methylation was found in several CpG islands of the L1 gene of HPV16 and this condition was shown to be associated with an increased risk of persistent infection by HPV16( 63 ).…”

A paradigm for post-embryonic Oct4 re-expression: E7-induced hydroxymethylation regulates Oct4 expression in cervical cancer

“…Upon HPV infection, the viral and host DNA methylation facilitates viral persistence and carcinogenesis. The grade of HPV L1 methylation is indicative of disease severity and persistent HPV infection in the cervix [27], while persistent HPV infection is due the hypermethylation of the E2 binding site on the Long Control region (LCR) [28].…”