Identification of novel brain-penetrating oxime antidotes for phorate toxicity

新型脑穿透性肟解毒剂甲拌磷毒性的鉴定

• 批准号: 9633107
• 负责人: Janice Elaine Chambers
• 金额: $ 21.56万
• 依托单位: MISSISSIPPI STATE UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2018
• 资助国家: 美国
• 起止时间: 2018-09-30 至 2020-08-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9633107
• 关键词: Acetylcholinesterase; Acetylcholinesterase Inhibitors; Antidotes; Atropine; Attenuated; Behavior; Binding; Biological; Blood - brain barrier anatomy; Brain; Brain Injuries; Butyrylcholinesterase; Cause of Death; Cessation of life; Chemicals; Chemistry; Cholinesterase Inhibitors; Cholinesterases; Computer Simulation; Cytochrome P450; Data; Development; Distal; Ethers; FDA approved; Female; Future; Height; Hepatic; In Vitro; Insecticides; Kinetics; Laboratories; Legal patent; Lethal Dose 50; Libraries; Life; Mediating; Metabolic Activation; Mixed Function Oxygenases; Modeling; Muscarinic Acetylcholine Receptor; Neuraxis; Neurotoxicity Syndromes; Oral; Organophosphates; Oxides; Oximes; Paraoxon; Parathion; Parents; Pattern; Peptides; Peripheral; Phorate; Poisoning; Property; Rattus; Sarin; Savings; Seizures; Series; Sulfones; Sulfoxide; Sulfur; Survivors; Testing; Therapeutic; Time; Toxic effect; Treatment Efficacy; Violence; attenuation; combat; comparative efficacy; dosage; experimental study; improved; in vivo; inorganic phosphate; lipophilicity; liquid chromatography mass spectrometry; male; nerve agent; novel; phosphorodithioic acid; respiratory; response; toxic organophosphate insecticide exposure

7. Project Summary/Abstract Many of the organophosphate (OP) insecticides, such as phorate (O,O-diethyl S-ethylthiomethyl phosphorodithioate), are highly toxic with rat oral LD50's in the low mg/kg range, and they or their active metabolites are potent inhibitors of acetylcholinesterase (AChE). Phorate is consistently more toxic to females than males; for example, rat oral LD50's for males and females are 3.7 and 1.4 mg/kg, respectively. Phorate requires monooxygenase-mediated bioactivation to active anticholinesterase metabolites, similar to a number of other OP insecticides, such as parathion (O,O-diethyl O-nitrophenyl phosphorothionate). Our preliminary studies with a phorate metabolite phorate-oxon (PHO) have indicated a longer time delay and more violent signs of poisoning than with paraoxon (PXN), the active metabolite of parathion. Additionally, patterns of oxime-mediated cholinesterase reactivation differ between PHO and PXN, which is unexpected because both are diethyl phosphates, and would be expected to phosphylate cholinesterase with the same diethyl moiety and therefore display similar reactivation patterns. PHO undergoes additional bioactivation of its terminal sulfur ether to a sulfoxide, then to a sulfone. Estimates of binding energies and the unusual preliminary results observed thus far have suggested that there may be an ethoxy leaving group instead of the expected ethylthiomethyl group and that the slow bioactivation of PHO in the brain to PHO-sulfoxide and then to PHO- sulfone might be responsible for the unexpected preliminary observations. 2-PAM is the currently FDA- approved oxime AChE reactivator. However the need for a different oxime reactivator that is more effective with an unconventional phosphylating moiety as well as an oxime that can penetrate into the brain will be needed for effective phorate therapy. Our laboratories have invented and patented novel substituted phenoxyalkyl pyridinium oximes that show preliminary evidence of survival efficacy with PHO as well as convincing evidence of entry into the brain with other OP's in our rat model. Therefore the following Specific Aims are proposed: Aim 1. To confirm the leaving group of PHO or PHO metabolites through a mass spectral analysis of AChE-phosphylated peptides and through computational modeling to determine barrier height for the potential leaving groups. Aim 2. To determine bioactivation efficiency through analysis of brain and hepatic bioactivation kinetics for phorate to its several metabolites as quantified by LC/MS/MS. Aim 3. To identify more effective oxime reactivators from our novel oxime library of AChE inhibited by the three phorate metabolites through in vitro reactivation studies and limited in vivo phorate survival studies. The results of this R21 project will be the identification of a few down-selected novel oximes that can be further developed in a subsequent U01 project into effective therapeutics for phorate poisoning.

7.项目总结/摘要 许多有机磷杀虫剂，如甲拌磷（O，O-二乙基S-乙硫基甲基 二硫代磷酸盐）具有高毒性，大鼠口服LD 50在低mg/kg范围内，它们或其活性成分 代谢物是乙酰胆碱酯酶（AChE）的有效抑制剂。甲拌磷对女性的毒性更大 例如，雄性和雌性大鼠的口服半数致死剂量分别为3.7和1.4毫克/千克。拌磷 需要单加氧酶介导的生物活化活性抗胆碱酯酶代谢物，类似于一些 其他OP杀虫剂，如敌百虫（O，O-二乙基O-硝基苯基硫代磷酸酯）。我们的初步 对甲拌磷代谢物甲拌磷的研究表明， 中毒的迹象比对氧磷（PXN），活性代谢产物的磷。此外， 肟介导的胆碱酯酶再活化在PHO和PXN之间不同，这是出乎意料的，因为两者都 是磷酸二乙酯，并且预期将磷酸化具有相同二乙基部分的胆碱酯酶 因此显示出相似的再激活模式。PHO经历其末端硫的额外生物活化 醚变成亚砜，然后变成砜。结合能的估计和不寻常的初步结果 迄今为止观察到的结果表明，可能存在乙氧基离去基团，而不是预期的乙氧基离去基团。 乙基硫甲基和缓慢的生物激活的PHO在脑中的PHO亚砜，然后PHO- 砜可能是导致非预期初步观察结果的原因。2-目前，FDA- 批准的肟AChE再活化剂。然而，需要一种更有效的不同肟活化剂， 带有非常规磷酸化部分以及可以渗透到大脑中的肟， 有效的甲拌磷治疗所需的。我们的实验室已经发明并获得专利的新型替代品 苯氧基烷基吡啶鎓肟，其显示出与PHO的存活功效的初步证据，以及 在我们的大鼠模型中与其他OP一起进入大脑的令人信服的证据。因此，以下具体 提出的目标：目标1。通过质谱确认PHO或PHO代谢物的离去基团 分析乙酰胆碱酯酶磷酸化肽，并通过计算建模来确定 潜在的离去基团。目标2.为了通过分析脑和脑组织来确定生物活化效率， 用LC/MS/MS定量甲拌磷几种代谢物肝生物活化动力学目的3.到 从三种甲拌磷抑制的AChE的新肟库中鉴定出更有效的肟重活化剂 通过体外再活化研究和有限的体内甲拌磷存活研究，对代谢物进行了研究。的结果 R21项目将鉴定一些向下选择的新肟，这些肟可以在一个 随后U 01计划成为甲拌磷中毒有效治疗剂。