Project 2: Attenuating Pulmonary Toxicity of Cutaneous Exposure to Arsenicals

皮肤暴露于高活性和毒性的砷，如路易氏剂、二苯氯砷、二乙基 氯化砷和二苯基氰化砷可能是偶然发生的，也可能是故意释放的。急性皮肤 暴露会导致严重的局部起泡和远端器官(如肺部)的炎症，导致损伤和 呼吸衰竭。砷致肺损伤的发病机制尚不完全清楚 寻找有效的治疗方案一直是一个挑战。利用经皮肺组织的RNA-Seq 暴露于砷后，我们鉴定了几个被改变的溴域4(BRD4)靶基因。BRD4是一种 与乙酰化组蛋白结合的溴末端和末端外(BET)结构域蛋白质家族的成员 引发炎症级联反应。这项建议的重点是了解BET蛋白和 皮肤砷中毒所致急性肺损伤和迟发性肺损伤的相关炎症途径 (DLI)。我们的初步数据显示，皮肤损伤后肺组织中BRD4蛋白和组蛋白乙酰化水平增加。 接触过砷。我们还证明，BRD4抑制剂JQ1可以减轻肺损伤并减少 皮肤暴露于苯基精氨酸(PAO；替代物)后的炎性细胞因子，如IL-6 阿森纳)。因此，我们假设有毒剂量的砷会导致蛋白质的乙酰化和 随后与BET蛋白结合，导致损伤通路的激活，并阻断BET信号转导 或者它的下游效应物可以减轻砷化合物引起的肺损伤。建议的研究分为 三个具体目标。目的1将描述皮肤暴露后砷致小鼠肺损伤的特征 敬军火库。这些曝光将在MRIGlobal进行，这是一个专门从事 这样的曝光率。对于机制研究和治疗测试，Aim 1还将开发内部PAO 皮肤砷致急性肺损伤模型和三氧化二砷吸入模型。AIM 2中的研究 将确定阿森纳引起急性肺毒性的机制。这些体外和体内研究 将在原代肺细胞中使用与战争有关的军火库和军火药的替代品 并将确定下游治疗靶点和分子，以减轻由 这些化学物质。这些研究将利用药物开发核心来获得新的合成或FDA- 批准的分子用于测试治疗效果。目标3的研究将确定小分子 溴域信号转导的抑制剂可以减轻砷中毒所致的肺损伤和相关的发病率。这些 在小鼠体内的研究最初将使用ATO或PAO进行。成功的候选分子，获得 通过我们的药物开发核心，然后将用更强大的武器库进行测试。建议的结果 研究将有助于确定皮肤砷暴露所致肺损伤的治疗方案，以及 其他与起泡和发炎有关的潜在有毒化学物质。

Cutaneous exposure to highly reactive and toxic arsenicals such as lewisite, diphenyl chloroarsine, diethyl chloroarsine and diphenyl cyanoarsine may occur accidently or by deliberate release. Acute cutaneous exposures cause severe local blistering and inflammation in distal organs, such as the lung, causing injury and respiratory failure. The pathogenesis of arsenical-induced lung injury is incompletely understood and a search for effective treatment regimens has been a challenge. Using RNA-Seq of lungs obtained from cutaneous exposed arsenical we identified several bromodomain 4 (BRD4) target genes that were altered. BRD4 is a member of the bromo- and extra-terminal (BET) domain family of proteins that on binding to acetylated histones initiates the inflammatory cascade. This proposal focuses on understanding the role of BET proteins and associated inflammatory pathways in cutaneous arsenical-induced acute lung injury (ALI) and delayed lung injury (DLI). Our preliminary data indicates increased BRD4 protein and histone acetylation in lung following cutaneous arsenical exposures. We also demonstrate that JQ1, a BRD4 inhibitor, mitigates lung injury and decreases inflammatory cytokines such as IL-6, following cutaneous exposure to phenyl arsineoxide (PAO; a surrogate arsenical). Therefore, we hypothesize that toxic doses of arsenicals cause acetylation of proteins and subsequent binding to BET proteins, resulting in activation of injury pathways, and that blocking BET signaling or its downstream effectors can mitigate arsenicals-induced lung injury. The proposed studies are divided into three specific aims. Aim 1 will characterize arsenical-induced lung injury in mice following cutaneous exposures to arsenicals. These exposures will be carried out at MRIGlobal, a contract organization specializing in carrying out such exposures. For mechanistic studies and testing of therapies, aim 1 will also develop an in-house PAO model of cutaneous arsenical-induced ALI and an inhalation model of arsenic trioxide (ATO). Studies in Aim 2 will determine mechanisms by which arsenicals cause acute pulmonary toxicity. These in vitro and in vivo studies will be carried out using both warfare-related arsenicals and surrogates of arsenicals in primary lung cells and mice and will identify downstream therapeutic targets and molecules that can mitigate the toxicity induced by these chemicals. These studies will utilize the Drug Development Core in acquiring novel synthetic or FDA- approved molecules to test for therapeutic efficacy. Studies in Aim 3 will determine whether small molecule inhibitors of bromodomain signaling can mitigate arsenical-induced lung injury and associated morbidity. These in vivo mouse studies will initially be carried out using ATO or PAO. Successful candidate molecules, obtained through our Drug Development Core, will then be tested with more potent arsenicals. Results of the proposed research will help identify treatment options for cutaneous arsenical exposure-induced lung injury as well as for other potentially toxic chemicals associated with blistering and inflammation.