Prognostic Significance of DNA & Histone Methylation

描述(由申请人提供)： 目前，还没有经过验证的诊断或预后标准来识别低级别宫颈病变(CIN 1)，这些病变最终会发展为CIN 2和CIN 3或宫颈癌。我们认为，CIN 1病变的常规组织病理学检查或常规护理就诊中的高危(HR)-HPV检测不足以提供预测高级别病变的信息。根据我们最近的研究结果，叶酸水平降低与HR-HPV持续存在和高度宫颈发育不良有关，我们推测这些病变中的叶酸和叶酸相关的表观遗传学改变(全局、CpG岛和DNA和组蛋白的基因特异性甲基化)可能为识别CIN 1病变提供有价值的信息，这些病变是CIN 2或CIN 3的目标。从2003年1月开始，我们机构的所有CIN 1和HR-HPV女性前瞻性跟踪并每年在阿拉巴马大学伯明翰分校妇女健康研究中心(CRWH)进行HPV检测，作为常规护理的一部分。这为我们提供了一个独特且经济实惠的机会，可以招募这些妇女参加前瞻性的后续研究，以评估叶酸和叶酸相关的表观遗传学标志物在识别高危低级别宫颈病变中的重要性。我们推测，在HR-HPV阳性的CIN 1患者中，在12-24个月后发生CIN 2或3的患者的循环和宫颈细胞叶酸浓度以及DNA和组蛋白的甲基化程度将不同于在类似时间段未发生CIN 2或3的类似病变患者。我们建议招募600名HR-HPV阳性的CIN 1妇女进行为期24个月的随访研究。为了评估叶酸和甲基化是否是CIN 2/3的独立预测因子，我们将把结果与宫颈癌和其他癌症保护性维生素的已知流行病学和HPV风险因素相关联。新开发和测试的免疫组织化学技术，测量完整和特定类型的宫颈细胞中DNA和病史(lys4和9)的全局甲基化程度，将在拟议的研究中用于评估全局甲基化。胞嘧啶延伸试验和实时定量聚合酶链式反应将分别用于评估CpG岛甲基化和基因特异性甲基化。这将是第一个全面的前瞻性随访研究，旨在评估维生素和相关表观遗传生物标记物对宫颈发育不良的预后意义。由于HR-HPV感染和低级别宫颈病变是常见的，对需要干预的宫颈病变具有更高特异性的新标记物将改善未来的宫颈癌筛查。这项研究旨在验证表观遗传生物标记物是可行的和具有成本效益的大规模执行。UAB的研究人员人才、设备、设施和接触研究对象的机会都是进行这项研究的理想选择。

DESCRIPTION (provided by applicant): Currently, there are no validated diagnostic or prognostic criteria that will identify low-grade cervical lesions (CIN 1) that are destined toward CIN 2 & 3 or cervical cancer. We believe that the conventional histopathological examination of CIN 1 lesions or testing for high-risk (HR)-HPV at a routine care visit provides insufficient information to predict high-grade lesions. Based on our recent results which demonstrated that lower circulating levels of folate are associated with HR-HPV persistence and development of high-grade cervical dysplasia, we hypothesize that folate and folate-related epigenetic alterations (global, CpG island and gene specific methylation of DNA and histones) in these lesions may provide valuable information for identifying CIN 1 lesions that are destined toward CIN 2 or 3. From January 2003, all women with CIN 1 and HR-HPV at our institution are followed prospectively and tested annually for HPV at the University of Alabama at Birmingham (UAB) Center for Research in Women's Health (CRWH) as part of their routine care. This gives us a unique and cost-effective opportunity to recruit these women into a prospective follow-up study to evaluate the significance of folate and folate-related epigenetic markers in the identification high-risk low-grade cervical lesions. We hypothesize that the circulating and cervical cell concentrations of folate and the degree of methylation of DNA and histones in HR-HPV-positive CIN 1 subjects who develop CIN 2 or 3 after a 12-24 month period will be different than that of similar lesions ill subjects who do not develop CIN 2 or 3 lesions during a similar time period. We propose to recruit 600 women with HR-HPV positive CIN 1 to a 24-month follow-up study. To evaluate whether folate and methylation are independent predictors of CIN 2/3, we will correlate results with known epidemiological and HPV risk factors for cervical cancer and other cancer-protective vitamins. Newly developed and tested immunohistochemical techniques, which measure the degree of global methylation of DNA and histories (lys4 & 9) in intact and specific types of cervical ceils, will be used to evaluate global methylation in the proposed study. Cytosine extension assay and real-time PCR assays will be used to evaluate CpG island methylation and gene-specific methylation respectively. This will be the first comprehensive prospective follow-up study designed to evaluate the prognostic significance of vitamins and related epigenetic biomarkers for cervical dysplasia. Since HR-HPV infections and low-grade cervical lesions are common, novel markers with higher specificity for cervical lesions requiring intervention will improve cervical cancer screening in the future. The study intends to validate epigenetic biomarkers that are feasible and cost-effective to perform on a large scale. Investigator talent, equipment, facilities and access to study subjects at UAB are ideal for conducting this study.