Biochemistry connecting glutathione and isocyanate asthma

连接谷胱甘肽和异氰酸酯哮喘的生物化学

• 批准号: 8738279
• 负责人: ADAM WISNEWSKI
• 金额: $ 20.87万
• 依托单位: YALE UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2013
• 资助国家: 美国
• 起止时间: 2013-09-01 至 2016-08-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8738279
• 关键词: Biochemistry; connecting; glutathione; isocyanate; asthma

DESCRIPTION (provided by applicant): Isocyanate, the essential cross-linking chemical for making polyurethane, are a major cause of occupational asthma world-wide. However, the mechanisms by which they cause disease remain unclear. We theorize that isocyanate asthma pathogenesis depends upon dynamic biochemical reactivity of isocyanate with multiple/different self-proteins/peptides, through metabolic pathways previously unappreciated for their connection to isocyanate allergenicity. Our specific hypothesis is that inhaled isocyanate reacts first, and preferentially, with the free thiol of the self-peptide, glutathione (GSH), which is preent at high concentrations (100-1000 ¿M) in the airway fluid. However, thiol-linked isocyanate-GSH reaction products are "quasi-stable" at physiologic pH, and if not rapidly metabolized and excreted, can "transfer" the isocyanate group to another self-protein/peptide, especially albumin, one of the most abundant proteins in the body. Isocyanate conjugation of albumin is known to cause antigenic changes that trigger adaptive immunologic responses (specific IgE, T-cell responses) known to promote airway inflammation and asthma. The novelty of the present hypothesis vs. previously proposed mechanisms of isocyanate asthma is the formation of immunogenically distinct isocyanate antigens (isocyanate-albumin conjugates) via GSH-mediated transcarbomylation, rather than direct reactivity with isocyanate. If correct, this paradigm would explain many of the unique features of "isocyanate" vs. typical "environmental" asthma, and provide critical new insights into pathogenesis, risk factors, and diagnostic indicators of disease. In this grant application, we present unpublished Preliminary data that strongly support our novel hypotheses, and outline a Research Plan to better understand GSH-isocyanate reactivity and its connection with pathogenic responses to occupational exposure. Given the toxicity of isocyanates, the study design involves in vitro experiments followed by in vivo experiments in an animal model of isocyanate asthma, to directly test our hypotheses in a manner not possible in human subjects. The highly ambitious proposal will be facilitated through unique infrastructure (W.M. Keck Biotechnology Resource Laboratory) at the applicant's institution and collaboration with ongoing clinical isocyanate asthma research. The experimental data from the grant is expected to define key steps in isocyanate asthma pathogenesis, and biomarkers, upon which to base better strategies for disease surveillance and diagnosis. Successful completion of the proposed studies should also provide compelling evidence for further clinical studies to translate these initial findings into practical methods of disease prevention. The two specific aims are to Aim 1. Characterize the reaction products between glutathione (GSH) and isocyanate, and their carbamoylating capacity, under physiologic conditions in vitro. Aim 2. Determine the role of GSH-isocyanate interactions in the development of asthma in vivo in an animal model.

描述（由申请人提供）：异氰酸酯是制造聚氨酯的重要交联化学物质，是全世界职业性哮喘的主要原因。然而，它们引起疾病的机制仍不清楚。 我们推测，异氰酸酯哮喘发病机制取决于异氰酸酯与多种/不同自身蛋白/肽的动态生化反应性，通过以前未被认识到与异氰酸酯过敏性相关的代谢途径。我们的具体假设是，吸入的异氰酸酯首先且优先与自肽谷胱甘肽 (GSH) 的游离硫醇发生反应，谷胱甘肽 (GSH) 在气道液体中以高浓度 (100-1000 µM) 存在。然而，硫醇连接的异氰酸酯-GSH反应产物在生理pH下是“准稳定”的，如果不快速代谢和排泄，可以将异氰酸酯基团“转移”到另一种自身蛋白质/肽上，尤其是白蛋白，这是体内最丰富的蛋白质之一。众所周知，白蛋白的异氰酸酯缀合会引起抗原变化，从而引发适应性免疫反应（特异性 IgE、T 细胞反应），从而促进气道炎症和哮喘。 与之前提出的异氰酸酯哮喘机制相比，本假设的新颖之处在于通过 GSH 介导的转羧基化形成免疫原性不同的异氰酸酯抗原（异氰酸酯-白蛋白缀合物），而不是与异氰酸酯直接反应。如果正确的话，这一范式将解释“异氰酸酯”与典型“环境”哮喘的许多独特特征，并为疾病的发病机制、危险因素和诊断指标提供重要的新见解。在本次拨款申请中，我们提供了未发表的初步数据，这些数据强烈支持我们的新假设，并概述了一项研究计划，以更好地了解 GSH-异氰酸酯反应性及其与职业暴露致病反应的关系。考虑到异氰酸酯的毒性，研究设计涉及体外实验，然后在异氰酸酯哮喘动物模型中进行体内实验，以在人类受试者中不可能的方式直接检验我们的假设。这项雄心勃勃的提案将通过申请人所在机构独特的基础设施（W.M. Keck 生物技术资源实验室）以及与正在进行的临床异氰酸酯哮喘研究的合作来促进。这笔资助的实验数据预计将定义异氰酸酯哮喘发病机制和生物标志物的关键步骤，以此为疾病监测和诊断的更好策略奠定基础。拟议研究的成功完成还应该为进一步的临床研究提供令人信服的证据，以将这些初步发现转化为疾病预防的实际方法。这两个具体目标是： 目标 1：在体外生理条件下表征谷胱甘肽 (GSH) 和异氰酸酯之间的反应产物及其氨基甲酰化能力。目标 2. 确定 GSH-异氰酸酯相互作用在动物模型体内哮喘发展中的作用。

项目成果

Reaction products of hexamethylene diisocyanate vapors with "self" molecules in the airways of rabbits exposed via tracheostomy.

• DOI: 10.1080/00498254.2017.1329569
• 发表时间: 2018-05
• 期刊: Xenobiotica; the fate of foreign compounds in biological systems
• 作者: Wisnewski AV;Kanyo J;Asher J;Goodrich JA;Barnett G;Patrylak L;Liu J;Redlich CA;Nassar AF
• 通讯作者: Nassar AF

UPLC-MS for metabolomics: a giant step forward in support of pharmaceutical research.

• DOI: 10.1016/j.drudis.2016.11.020
• 发表时间: 2017-03
• 期刊: Drug discovery today
• 影响因子: 7.4
• 作者: Nassar AF;Wu T;Nassar SF;Wisnewski AV
• 通讯作者: Wisnewski AV

Biotransformation and Rearrangement of Laromustine.

拉莫司汀的生物转化和重排。

• DOI: 10.1124/dmd.116.069823
• 发表时间: 2016
• 期刊: Drug metabolism and disposition: the biological fate of chemicals
• 作者: Nassar,Alaa-EldinF;Wisnewski,AdamV;King,Ivan
• 通讯作者: King,Ivan

Identification of novel reaction products of methylene-bis-phenylisocyanate ("MDI") with oxidized glutathione in aqueous solution and also during incubation of MDI with a murine hepatic S9 fraction.

鉴定亚甲基双苯基异氰酸酯（“MDI”）与氧化型谷胱甘肽在水溶液中以及在 MDI 与鼠肝 S9 级分孵育期间的新反应产物。

• DOI: 10.1016/j.tiv.2016.07.011
• 发表时间: 2016
• 期刊: Toxicology in vitro : an international journal published in association with BIBRA
• 作者: Wisnewski,AV;Liu,J;Nassar,AF
• 通讯作者: Nassar,AF