Eating worms: an unusual nicotinic acetylcholine receptor from the nematode pharynx as a potential drug target

 描述(申请人提供)：被忽视的热带疾病(NTDS)包括土壤传播的蠕虫硫酶(STH)，它是由不同的肠道线虫群引起的。这些寄生虫包括蛔虫、鞭虫和钩虫。这些感染很常见。例如，蛔虫病影响着全世界14亿人，最常见的是3至8岁的儿童。这些线虫寄生虫的控制依赖于有效的驱虫剂供应。以兽医为例，我们知道继续使用驱虫药进行大规模化疗会导致耐药性。有必要确定驱虫药开发的新靶点。这样的位置曾经是线虫咽部的烟碱型乙酰胆碱受体。我们发现，这种受体对目前使用的抗虫药物不起作用。方法：本应用的具体目的是：1.优化线虫EAT-2和EAT-18的表达，并对所产生的受体进行药理学鉴定。我们将研究受体表达的最佳条件，并使用一系列激动剂、拮抗剂和潜在的变构调节剂来表征所产生的受体反应。2.克隆和表达猪链霉菌Eat-2和Eat-18。咽感受器还需要其他亚单位吗？我们将从猪蛔虫中鉴定EAT-2和EAT-18的同源基因，克隆并在非洲爪哇卵母细胞中表达，用于鉴定。此外，我们还将检验形成受体需要其他亚单位基因的假设。3.Eat-2和Eat 18结合形成成熟受体吗？我们将检验EAT-2和EAT-18结合形成功能性nAChR的新假设。在这个项目完成后，我们将在一个异源系统中表达一个来自寄生线虫的重要的潜在药物靶点。我们将描述一种非常新颖的nAChR(缺乏任何阿尔法亚基)。我们将在猪蛔虫寄生虫中识别编码这些受体的基因。最后，我们将确定小(71A.A.)成熟的离子通道受体需要蛋白EAT-18。

 DESCRIPTION (provided by applicant): The Neglected Tropical Diseases (NTDs) include the soil-transmitted helminthiases (STHs) which are caused by diverse groups of intestinal nematodes. The parasites include Ascaris, Trichuris and hookworms. These infections are common. Ascariasis for example, affects 1.4 billion people worldwide and is most common in children between the ages of 3 and 8. Control of these nematode parasites relies on an effective supply of anthelmintics. Taking veterinary medicine as an example, we know that continued use of anthelmintic compounds for mass chemotherapy will lead to drug resistance. There is a need to identify novel target sites for anthelmintic development. Once such site is the nicotinic acetylcholine receptor on the nematode pharynx. We have discovered this receptor does not respond to currently used anrthelmintic drugs. Approach: The specific aims of this application are: 1. Optimize expression of C. elegans eat-2 and eat 18 and pharmacologically characterize the resulting receptor. We will investigate optimal conditions for receptor expression and characterize the resulting receptor responses using a range of agonists, antagonists and potential allosteric modulators. 2. Clone and express A. suum eat-2 and eat-18. Are other subunits required for the pharyngeal receptor? We will identify orthologs of eat-2 and eat-18 from Ascaris suum, clone, and express them in Xenopus oocytes for characterization. Additionally we will test the hypothesis that other subunit genes are required to form the receptor. 3. Do eat-2 and eat 18 combine to form the mature receptor? We will test the novel hypothesis that eat-2 and eat-18 combine to form a functional nAChR. On completion of this project we will have expressed in a heterologous system an important new potential drug target from a parasitic nematode. We will have characterized an extremely novel nAChR (one that lacks any alpha subunits). We will have identified the genes that encode these receptors in the parasitic nematode Ascaris suum. Finally, we will have determined whether the small (71 a.a.) protein eat-18 is required in the mature ion channel receptor.