DNA Repair Capacity Assays for Lung Disease Risk Assessment

项目摘要 肺癌(LC)是与癌症相关的主要死亡原因，COPD在所有原因中排名第三 死亡率。在美国，大约有7000万现在和以前吸烟的人(SM)，尽管 分别只有15%-20%和~17%的人会被诊断为LC或COPD。因此，敏感度的差异 已被提出在LC和COPD疾病的病因中发挥重要作用，这种疾病的病因也不同于 西班牙裔白人(Nhw)和西班牙裔族裔。据估计，吸烟是85%的LC死亡的原因， 与慢性阻塞性肺疾病密切相关。暴露在其他环境呼吸道致癌物质中，包括氡， 木材烟雾和颗粒物2.5微米(PM2.5)可能与吸烟相互作用，或单独 致癌物质。在美国，从不吸烟者(NS)的LC病例每年增加15-20,000例。风险因素 包括环境烟草烟雾(侧流烟雾是主要成分)、氡、木材烟雾和 PM2.5。木材烟雾和PM2.5也与慢性阻塞性肺疾病和最近的电子产品有关 香烟已被证明会影响肺功能。所有这些环境暴露都有 它们诱导DNA损伤的能力具有共性。DNA修复能力降低(DRC)已被证明 主要通过以医院为基础的病例对照研究与LC有关，但尚未得到严格的研究 用于慢性阻塞性肺病。这些研究使用宿主细胞重新激活或诱变剂敏感性试验来响应 特定的DNA损伤剂。RFA的一个主要目标是将基因组完整性分析扩展到人群 研究“是开发和/或评估高通量、功能性DNA修复分析在 在人群环境中评估疾病表型和发病机制的背景。我们对这一指控的处理方法 目的是评估三种新兴的高通量DRC检测方法--Litron体外流式细胞术MN检测， Trevigen彗星试验和Trevigen hOGG1 FLARETM彗星试验。此外，通过与 Nagel博士，他的流式细胞仪宿主细胞再激活分析(FM-HCR)的表现衡量了 人类细胞修复不同类型损伤的能力将被导入细胞内的质粒 评估过了。我们将利用两个已建立的队列，LSC和新墨西哥州LC队列来评估 这些DRC分析通过暴露预测SM和NS(AIM 1)的LC风险以及SM(AIM 2)的COPD风险 从主流和侧流香烟烟雾、木材烟雾、 电子烟气雾剂和过氧化氢(模拟氡)。目标3将解决将私营军事公司用作 通过比较三种DRC检测和暴露在初级人群中预测LC和COPD风险的替代物 支气管镜下获得的支气管上皮细胞与PMC相匹配。此外，FM-HCR检测将 来自目标1和目标2的研究对照，在对一个或多个 以确定这一测试是否可以复制这一结果。无论是一个或多个特定的DNA 修复途径(例如，碱基切除)是导致DRC差异的原因也将得到确定。

Project Summary Lung cancer (LC) is the leading cancer-related cause of death and COPD is third among all cause mortality. There are approximately 70 million current and former cigarette smokers (SM) in the US, although only 15-20% and ~17% will be diagnosed with LC or COPD, respectively. Thus, differences in susceptibility have been proposed to play an important role in LC and COPD disease etiology that also differs between non- Hispanic white (NHW) and Hispanic ethnicity. Smoking is estimated to be the cause of 85% of LC deaths and strongly associated with COPD. Exposure to other environmental respiratory carcinogens that include radon, wood smoke, and particulate matter 2.5 µ (PM2.5) may interact with cigarette smoking or individually be carcinogenic. LC in never smokers (NS) is increasing with 15-20,000 cases annually in the US. Risk factors include environmental tobacco smoke (sidestream smoke is the major component), radon, wood smoke and PM2.5. Wood smoke and PM2.5 are also associated with risk for COPD and most recently electronic cigarettes have been shown to affect pulmonary function. All of these environmental exposures have commonality in their ability to induce DNA damage. Reduced DNA repair capacity (DRC) has been shown to be associated with LC largely through hospital-based case-control studies, but has not been rigorously studied for COPD. These studies used the host cell reactivation or the mutagen sensitivity assay in response to specific DNA damaging agents. A major goal for this RFA “Expanding Genome Integrity Assays to Population Studies” is to develop and/or evaluate the performance of high throughput, functional DNA repair assays in the context of assessing disease phenotype and pathogenesis in a population setting. Our approach to this charge is to evaluate three emerging high throughput DRC assays – Litron in vitro flow cytometry-based MN assay, Trevigen comet assay, and the Trevigen hOGG1 FLARETM comet assay. In addition, through collaboration with Dr. Nagel, the performance of his flow cytometric host cell reactivation assay (FM-HCR) that measures the ability of human cells to repair different types of damage within plasmids transfected into the cells will be assessed. We will take advantage of two established cohorts, the LSC and New Mexico LC cohort to evaluate these DRC assays for predicting LC risk in SM and NS (Aim 1) and COPD in SM (Aim 2) through the exposure of their PMCs to total particulate matter (TPM) from mainstream and sidestream cigarette smoke, wood smoke, electronic cigarette aerosol, and H2O2 (mimic for radon). Aim 3 will address the ability to use the PMCs as a surrogate for risk prediction for LC and COPD by comparing the three DRC assays and exposures in primary bronchial epithelial cells obtained by bronchoscopy and matched PMCs. In addition, the FM-HCR assay will study controls from aims 1 and 2 that show the largest difference in DRC in response to one or more of the exposures to determine if this assay can replicate that outcome. Whether one or more of the specific DNA repair pathways (e.g., base excision) are driving the differences in DRC will also be determined.