Abstract 1991: Methylation of the HPV16 L1 gene is associated with disease progression in a prospective population-based cohort
Authors: Mirabello Lisa , Sun Chang , Ghosh Arpita , Rodriguez Ana C. , Schiffman Mark , Wentzensen Nicolas , Wacholder Sholom , Lorincz Attila , Burk Robert D. 
Published: August 2014

Persistent infection with one of the carcinogenic types of human papillomavirus (HPV) is the primary cause of cervical cancer, the 3rd most commonly diagnosed cancer worldwide. HPV type 16 is the most prevalent HPV type, the most persistent, and is found in about half of all cancers. Only a small minority of women infected with HPV16 progress to cervical precancer, with a substantial fraction of those eventually developing invasive cancer. The viral and host determinants of transitions to HPV 16 infection clearance, persistence and progression to precancer are still largely unknown.

CpG methylation can be used as a defense mechanism against expression from foreign, e.g. viral, DNA. Methylation of HPV DNA by infected cells may alter viral expression patterns relevant for viral infection and transformation. The goal of our study was to evaluate whether DNA methylation in the late structural gene, L1, or the upstream regulatory region (URR) of HPV16 are associated with clearance of infection, persistence without progression to precancer, or progression to precancer among women in the prospective Guanacaste, Costa Rica cohort.

HPV16 DNA was extracted from cervical cells collected prior to diagnosis from 36 women randomly selected from each of the 3 main infection outcomes that can be distinguished within the cohort: 1) women who cleared their HPV16 infection, 2) women with persistent HPV16 infection without progression to precancer (defined as CIN3), and 3) women with HPV16 infection who progressed to precancer (CIN3). DNA was bisulfite modified and methylation was quantified using pyrosequencing assays at 16 CpG sites in L1 and 16 CpGs in URR. The Kruskal-Wallis test was used to determine if methylation at each individual CpG site was associated with disease outcome. To account for multiple testing, the Benjamini-Hochberg method was used to ensure the expected proportion of false positives was less than 0.05. To determine whether methylation of L1 or URR was associated with disease outcome, incorporating information from all sites in each region, a global test based on the minimum of the P-values was performed. The P-value corresponding to the global test was obtained via permutation.

Methylation at six different CpG sites in the L1 region were found to be associated with outcome status (P = 0.01-0.0008). The L1 region was significantly associated with the disease outcome (Global P = 0.008). At each differentially methylated CpG site in the L1 region, we noted increased methylation with each transition towards precancer. No significant associations were detected with the URR region or the individual CpG sites in this region.

Viral CpG methylation of L1 might alter gene expression and thereby influence infection outcome. This is the first study to prospectively analyze outcomes related to baseline HPV16 methylation of L1 and URR and to suggest that methylation of L1 may be driving viral progression to precancer.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 1991. doi:10.1158/1538-7445.AM2011-1991