Cholinergic receptors on the nematode pharynx: an unexploited drug target

描述（由申请人提供）：被忽视的热带病（NTD）包括由不同肠道线虫引起的土壤传播蠕虫病（STH）。寄生虫包括蛔虫、鞭虫和钩虫。这些感染很常见。例如，蛔虫病影响全世界14亿人，在3至8岁的儿童中最常见。控制这些线虫寄生虫依赖于有效供应驱虫剂。以兽药为例，我们知道继续使用驱虫化合物进行大规模化疗会导致耐药性。需要鉴定用于驱虫剂开发的新靶位点。一旦这样的网站是烟碱乙酰胆碱受体的线虫咽。我们已经发现这种受体对目前使用的驱虫药没有反应。方法：本申请的具体目的是：1。表征A中的nAChR反应。咽部：采用电流钳、电压钳和膜片钳技术。我们将检验A.猪咽nAChR与胆碱能驱虫剂激活的肌肉nAChR截然不同。2.咽部特异性烟碱乙酰胆碱受体亚单位的鉴定与克隆。我们将确定nAChR亚基基因存在于咽部产生功能性受体所需的。3.应用电压钳和膜片钳技术研究非洲爪蟾卵母细胞咽部胆碱能受体。我们将测试的假设，咽nAChR亚单位的表达结果在功能受体，这是在体内发现的类似。在这个项目完成后，我们将在寄生线虫中描述一个重要的新的潜在药物靶点的药理学特征。我们将已经确定了编码这些受体的基因。最后，我们将在一个可访问的平台上重建受体，适合筛选潜在的新驱虫剂。本研究的长期目标是在适合药物发现筛选的平台（非洲爪蟾卵母细胞）中提供来自一系列重要寄生虫物种的离子通道药物靶点。 公共卫生相关性：被忽视的热带病包括土壤传播的线虫寄生虫，如蛔虫、鞭虫和钩虫。例如，蛔虫病是人类最常见的寄生虫感染之一。1.5全世界有10亿人患有蛔虫病，这种疾病在3至8岁的儿童中最常见。蠕虫病的治疗包括使用烟碱类驱虫剂，如噻嘧啶和oxantel，通过激活肌肉上的胆碱能离子通道（nAChR）选择性麻痹线虫。然而，对驱虫药的耐药性是一个真实的问题。我们已经确定了一个新的潜在的新的驱虫剂，nAChR的线虫咽，这是目前使用的药物不敏感的目标网站。我们建议，该受体的药理学特性，克隆的组件亚基基因和表达的功能性受体在非洲爪蟾卵母细胞。在项目完成后，我们将表征潜在的新药物靶点（咽nAChR），并将其重建在适合筛选化合物文库的平台（非洲爪蟾卵母细胞）中，以鉴定新的驱虫化合物。

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. Characterize nAChR responses in A. suum pharynx: using current-, voltage- & patch- clamp techniques. We will test the hypothesis that the A. suum pharynx nAChRs are pharmacologically distinct from the muscle nAChRs activated by cholinergic anthelmintics. 2. Identify & clone pharyngeal specific nicotinic acetylcholine receptor subunits. We will identify the nAChR subunit genes present in the pharynx required to produce functional receptors. 3. Characterize the pharyngeal cholinergic receptors in Xenopus laevis oocytes using voltage- & patch-clamp techniques. We will test the hypothesis that expression of pharyngeal nAChR subunits results in functioning receptors that are pharmacologically similar to those found in vivo. On completion of this project we will have characterized the pharmacology of an important new potential drug target in a parasitic nematode. We will have identified the genes that encode these receptors. Finally, we will have reconstituted the receptors in an accessible platform suitable for screening potential new anthelmintics. The longer term goal of this research is provide ion- channel drug targets from a range of important parasite species in a platform (Xenopus oocytes) suitable for drug discovery screening. PUBLIC HEALTH RELEVANCE: The Neglected Tropical Diseases (NTDs) include soil transmitted nematode parasites like ascariasis, trichuriasis and hookworm. Ascariasis, for example, is one of the most common human parasitic infections. 1.5 billion people worldwide have ascariasis, and the disease is most common in children between the ages of 3 and 8. Treatment of helminthiasis includes use of nicotinic anthelmintics like pyrantel and oxantel which selectively paralyze nematodes by activating cholinergic ion-channels (nAChRs) on their muscle. However, resistance to anthelmintics drugs is a real concern. We have identified a novel potential target site for new anthelmintics, the nAChR on the nematode pharynx, which is not sensitive to currently used drugs. We propose to characterize the pharmacological properties of this receptor, clone the component subunit genes and express the functional receptor in Xenopus oocytes. On completion of the project we will have characterized a potential new drug target (the pharynx nAChR) and reconstituted it in a platform (Xenopus laevis oocytes) suitable for screening compound libraries to identify new anthelmintic compounds.