Adduct Formation in the Toxicity of Dithiocarbamates

描述（由申请人提供）：人类接触二硫代氨基甲酸酯发生在农业，工业和医药中。尽管对降解和代谢途径的理解正在不断发展，但目前对二硫代氨基甲酸酯生物学作用的分子靶点和机制知之甚少。该项目的长期目标是描述二硫代氨基甲酸酯及其代谢物在生物系统中的相互作用，并确定这些相互作用作为毒性机制和生物标志物的相关性。先前的研究揭示了某些二硫代氨基甲酸酯产生选择性雪旺细胞毒性的能力，并伴随着神经中铜和脂质过氧化产物水平的增加。该应用的研究基于以下假设：这些二硫代氨基甲酸酯结合内源性铜，形成亲脂复合物，并在髓鞘内积聚，导致脂质过氧化、氧化损伤和脱髓鞘增加。这一假设将通过两个特定目的进行验证：1)确定铜积累和氧化应激是否需要二硫代氨基甲酸盐介导的节段性脱髓鞘；2)确定二硫代氨基甲酸盐介导的周围神经脱髓鞘的分子靶点和细胞反应。目标1将通过以下方式实现：A)合成对铜的亲和力不同的二硫代氨基甲酸酯及其铜络合物的溶解度，并评估其在体内脂质过氧化和脱髓鞘的相对效力，b)确定非二硫代氨基甲酸铜螯合剂铜酮提高神经中铜和脂质过氧化的能力。C)确定脂质过氧化和铜积累是否是二硫代氨基甲酸酯的直接影响，通过确定这些影响与髓鞘损伤发生的剂量反应和时间关系，d)确定氧化损伤和铜毒性受损的转基因模型是否对二硫代氨基甲酸酯介导的脱髓鞘更敏感。目标2将通过a)确定蛋白质表达的变化，b)识别受损蛋白质，c)测量铜酶活性以及d)表征二硫代氨基甲酸盐在神经中产生的过量铜的位置和化学种类来实现。这些研究的意义在于建立结构-活性关系，有助于预测可能通过类似机制起作用的药物，确定易感人群，制定基于机制的暴露建议，并预测长期低水平暴露的影响。

DESCRIPTION (provided by applicant): Human exposure to dithiocarbamates occurs in agriculture, industry and medicine. Although an understanding of degradative and metabolic pathways is evolving, there is currently little knowledge on the molecular targets and mechanisms underlying the biological affects of dithiocarbamates. The long-range objectives of this project are to delineate the interactions of dithiocarbamates and their metabolites within biological systems and to determine the relevance of these interactions as mechanisms of toxicity and biomarkers. Previous studies revealed the ability of certain dithiocarbamates to produce selective Schwann cell toxicity accompanied by increased levels of copper and lipid peroxidation products in nerve. Investigations in this application are guided by the working hypothesis that these dithiocarbamates bind endogenous copper, form lipophilic complexes and accumulate within myelin resulting in increased lipid peroxidation, oxidative injury and demyelination. This hypothesis will be tested through two specific aims: 1) To determine if copper accumulation and oxidative stress are required for dithiocarbamate-mediated segmental demyelination and 2) To determine the molecular targets and cellular responses in dithiocarbamate-mediated peripheral nerve demyelination. Aim 1 will be achieved through: a) synthesis of dithiocarbamates differing in their affinity for copper and their copper complex solubility and evaluating their relative potency for lipid peroxidation and demyelination in vivo, b) determining the ability of the non-dithiocarbamate copper chelating agent, cuprizone, to elevate copper and lipid peroxidation in nerve, c) determining if lipid peroxidation and copper accumulation are direct affects of dithiocarbamates through defining the dose response and temporal relationship of these affects to the onset of myelin injury and d) determining if transgenic models with compromised defense to oxidative injury and copper toxicity are more sensitive to dithiocarbamate-mediated demyelination. Aim 2 will be achieved through a) determining changes in protein expression, b) identifying damaged proteins, c) measuring cuproenzyme activities and d) characterizing the location and chemical species of excess copper produced in nerve by dithiocarbamates. The significance of these studies lies in establishing structure-activity relationships useful for predicting agents that may act through similar mechanisms, identifying susceptible populations, developing mechanistically based exposure recommendations and predicting the affects of long term low level exposures.