Cervical Cancer Preventive Measures Based on HPV 16 Epigenome

描述（由申请人提供）：HPV 16发挥致癌作用的主要机制是通过两种病毒早期基因E6和E7的表达。因此，调节E6和E7表达的因素可能会影响HPV 16感染向更高级别的宫颈上皮内瘤变（CIN 2+）或浸润性宫颈癌（ICC）的转化。HPV 16 DNA甲基化（DNAm）的程度已被证明影响其基因表达、病毒生命周期、持久性和复制行为。我们最近的研究表明：1）在宫颈脱落细胞（EC）中检测的HPV 16 DNAm的较高程度与被诊断为CIN 2+的可能性降低79%相关。2）HPV 16 DNAm的程度显示出识别CIN 2+女性的最佳阳性预测值（PPV），与任何致癌HPV类型检测阳性或异常pap检测阳性的女性相比，卫生部门常用的两种检测方法来识别高危女性3）HPV 16-叶酸循环浓度较低的HPV阳性女性被诊断为CIN 2+的可能性是叶酸水平较高的HPV 16阴性女性的9倍。由于HPV使用人类甲基化机制用于其DNAm，因此影响人类细胞中DNAm的相同因素，即叶酸代谢途径产生的甲基的可用性，也可能具有改变HPV 16 DNAm程度的能力。在此背景下，本研究的目的如下：目的1。使用从我们已发表的研究中确定的部位的EC和宫颈活检配对样本中提取的DNA，评估HPV 16 DNAm程度与CIN之间的相关性（HPV 16 E6增强子的7862位和HPV 16 E6启动子的31、37、43、52和58位）和CIN 2+，以便在单独的，更大的女性群体。这一目标将在已经从300名诊断为CIN 2+（病例）的妇女和300名诊断为d CIN 1（非病例）的妇女中收集的样本中进行测试。评价目标1相同部位的HPV 16 DNAm程度与叶酸循环浓度之间的相关性，以评估美国叶酸强化计划导致的不同水平叶酸暴露的影响。这一目标将在已经从300名诊断为d CIN 1的妇女中收集的样本中进行测试。 这项研究将提供一个独特的机会，以确定检测HPV 16 DNA m的程度，以确定发展HPV 16相关CIN 2+的高风险妇女的诊断价值。此外，该研究整合了一个重要的人群范围内的环境暴露（叶酸）和分子数据，通过测试的假设，暴露于叶酸可能有能力改变HPV 16 DNAm的程度，并修改HPV 16相关的风险发展CIN在美国后叶酸强化时代。 公共卫生相关性：该研究的目的是评估HPV 16的特异性甲基化变化是否可用作晚期宫颈上皮内瘤变风险增加的生物标志物，并确定叶酸水平是否可预测HPV 16甲基化程度。

DESCRIPTION (provided by applicant): A main mechanism by which HPV 16 exerts oncogenic effects is through the expression of two viral early genes, E6 and E7. Therefore, factors that regulate the expression of E6 and E7 are likely to influence transformation of HPV 16 infections to higher grades of cervical intraepithelial neoplasia (CIN 2+) or invasive cervical cancer (ICC). The degree of HPV 16 DNA methylation (DNAm) has been shown to influence its gene expression, viral life cycle, persistence and replicative behavior. Our recent studies demonstrated the following: 1) A higher degree of HPV 16 DNAm assayed in exfoliated cells (ECs) from the cervix was associated with 79% reduced likelihood of being diagnosed with CIN 2+ 2) The degree of HPV 16 DNAm showed the best positive predictive value (PPV) for identifying women with CIN 2+, compared to women tested positive for any carcinogenic HPV type or tested positive for abnormal pap, two tests commonly used by Health Departments to identify high risk women 3) HPV 16-positive women with lower circulating concentrations of folate were nine times more likely to be diagnosed with CIN 2+ compared to HPV 16-negative women with higher folate. Because HPV uses the human methylation machinery for its DNAm, the same factors which influence DNAm in human cells, namely the availability of methyl groups generated by the folate metabolic pathway, may also have the ability to alter the degree of HPV 16 DNAm. With this background, the aims of the study are as follows: Aim 1. To evaluate the association between the degree of HPV 16 DNAm and CIN using DNA extracted from paired samples of ECs and cervical biopsies at sites identified in our published study (position 7862 of the HPV 16 E6 enhancer and positions 31, 37, 43, 52 and 58 of the HPV 16 E6 promoter) and CIN 2+, in order to replicate our findings in a separate, larger group of women. This aim will be tested among samples already collected from 300 women diagnosed with CIN 2+ (cases) and 300 women diagnosed with d CIN 1 (non-cases) Aim 2. To evaluate the association between the degree of HPV 16 DNAm at the same sites as in aim 1 and circulating concentrations of folate, in order to assess the influence of exposure to different levels of folate as a result of the US folic acid fortification program. This aim will be tested among samples already collected from 300 women diagnosed with d CIN 1. The study will provide a unique opportunity to determine the diagnostic value of testing the degree of HPV 16 DNAm for identifying women at high risk for developing HPV 16 associated CIN 2+. Further, the study integrates an important population-wide environmental exposure (folate) and molecular data by testing the hypothesis that exposure to folate may have the ability to alter the degree of HPV 16 DNAm and modify the HPV 16 associated risk of developing CIN in the US post-folic acid fortification era. PUBLIC HEALTH RELEVANCE: The purpose of the study is to assess whether specific methylation changes in HPV 16 can be used as a biomarker for enhanced risk of advanced cervical intraepithelial neoplasia and to determine whether folic acid levels predict the degree of HPV 16 methylation.