Identification of Free Radical Induced Biomarkers of Exposure to Electronic Cigarette Aerosol

电子烟（e-cig）的使用正在上升，特别是在年轻人中; 然而，它们的潜在危害还不为人所知，使制定知情的监管战略变得复杂。 缺乏电子烟相关危害的数据，部分原因是缺乏暴露于电子烟的特定生物标志物。 气溶胶我们发现，电子烟气雾剂含有高活性的自由基，可导致氧化损伤， 用户.自由基可以破坏许多细胞途径，可能有助于疾病的进展。 癌症和其他疾病。电子烟自由基的检测可以通过自旋阱捕获来实现 (e.g., DMPO）并通过电子顺磁共振（EPR）光谱进行分析。初步研究， EPR表明，电子烟气溶胶中由电子液体溶剂丙二醇（PG）和 甘油（GLY），所有电子烟都有，显示出独特的结构特征。我们的目标是找出这些 自由基结构，并利用其独特的结构来开发暴露的电子烟特异性生物标志物。的 本研究的具体目标是：（目的1）确定所产生的自由基的结构 目的2）确定自由基的主要作用靶点及其形成的加合物 在啮齿动物模型中暴露于电子烟的组织中的攻击以及这些自由基形成的可能代谢物 暴露于电子烟的啮齿动物血清中的加合物。该项目代表了一个重要的研究方向， 化学/生物方法可以为烟草监管科学提供信息。因此，这方面的一个重要方面是 应用它来扩展我在翻译科学相关领域的背景，除了提供具体的 在新的生物标志物相关研究领域的培训，包括代谢组学和自由基结构 识别.为此，我的培训将通过一系列课程、相关指导和实践来进行。 经验，每一个面向，以确保我的过渡到一个独立的研究人员在生物标志物领域 发展和监管科学。课程作业，指导，会议参与，和实践 培训/体验将在K99阶段完成。在这一阶段，研究计划的目标1将 完成并启动目标2（在授予的R 00阶段完成）。完成 研究目标，先进的脉冲EPR和质谱方法将用于产生的自由基 被PG和GLY吸入电子烟气雾剂中。在小鼠暴露模型中，自由基暴露和攻击目标在小鼠体内的浓度是相同的。 将使用新的体内DMPO/抗DMPO抗体方法测定肺。这将允许 通过TOF-MS鉴定特异性蛋白质和DNA加合物。DMPO暴露后研究将包括 非靶向成对代谢组学方法，以观察电子烟前后代谢产物谱的变化 exposure.通过利用电子烟的独特结构产生的自由基和他们的攻击目标， 肺，暴露于特定于电子烟气溶胶的生物标志物可以被识别并用于开发监管 旨在减少电子烟危害的战略。

Project Summary/Abstract: Electronic cigarette (e-cig) usage is on the rise, particularly among youths; however, their potential for harm is not understood, complicating development of informed regulatory strategies. The lack of data on e-cig related harm is, in part, due to the lack of specific biomarkers for exposure to e-cig aerosols. We found that e-cig aerosol contains highly reactive free radicals that can cause oxidative damage to the user. Free radicals can damage numerous cellular pathways possibly contributing to the progression of cancers and other diseases. Detection of e-cig free radicals can be accomplished by trapping with spin traps (e.g., DMPO) and analysis by electron paramagnetic resonance (EPR) spectroscopy. Preliminary research with EPR shows that free radicals produced in the e-cig aerosol by e-liquid solvents, propylene glycol (PG) and glycerin (GLY), common to all e-cigs, display unique structural characteristics. Our objective is to identify these free radical structures and utilize their unique structure to develop an e-cig specific biomarker of exposure. The specific aims of the proposed research are: (Aim 1) To determine the structures of the free radicals produced by PG and GLY in e-cigs; (Aim 2) To determine the primary targets of and adducts formed from free radical assault in the tissue of e-cig exposures in rodent models and possible metabolites formed from the these radical adducts in the serum of e-cig exposed rodent. This project represents an important research direction where a chemical/biological approach can inform tobacco regulatory science. As such, an important aspect of this application it to extend my background in areas relevant to translational science in addition to providing specific training in new biomarker-relevant research areas including metabolomics and free radical structural identification. To this end, my training will occur through a series of courses, relevant mentorship, and practical experience, each geared to ensure my transition to an independent researcher in the fields of biomarker development and regulatory science. Coursework, mentorship, conference participation, and practical training/experience will be completed during the K99 phase. During this phase, Aim 1 of the research plan will be completed and Aim 2 will be initiated (for completion during the R00 phase of the award). To accomplish the research aims, advanced pulsed EPR and mass spectroscopy approaches will be utilized for radicals produced by PG and GLY in e-cig aerosols. In a mouse exposure model, free radical exposure and targets of attack in the lung will be determined using a novel in vivo DMPO/anti-DMPO antibody approach. This will allow for the identification of specific protein and DNA adducts by TOF-MS. A post-DMPO exposure study will consist of an untargeted pairwise metabolomics approach to look at changes in metabolite profiles before and after e-cig exposure. By leveraging the unique structures of e-cig produced free radicals and their targets of attack in the lung, biomarkers of exposure specific to e-cig aerosols can be identified and used to develop regulatory strategies aimed at reducing harm from e-cig exposure.