Diethylcarbamazine, Emodepside and SLO-1 K Channels of Filaria

项目摘要 丝虫病是一组被忽视的热带疾病，由克雷德微丝虫病感染引起。 由叮咬的昆虫传播的寄生线虫。由以下原因造成的河流失明 螺旋体丝虫病和由马来丝虫引起的淋巴丝虫病就是这样的例子 疾病。河盲症是由寄生虫引起的，这种寄生虫会导致角膜和 严重的瘙痒和皮炎；它感染了西非和中非的1700万人。淋巴 丝虫病是一种使人衰弱和毁容的疾病，全世界有1.2亿人患有丝虫病。 这些线虫寄生虫的控制依赖于少量的驱虫药，这些药物具有 有限的行动范围。目前还没有实用的大杀丝虫剂，可以杀死成虫。 也有人担心大量的微丝杀菌剂化疗会导致 抗药性的发展。 乙基卡马津是治疗淋巴丝虫病的主要药物，在美国大部分地区 世界上，除了在存在盘尾丝虫病的地区，因为它是相反的风险 失明。它能迅速清除微丝虫病，并导致约40%的成虫死亡 (大杀丝剂)。然而，令人震惊的是，在它推出68年后，我们还没有证明 了解其作用的分子机制。在这里，我们建议重新激活这一点 研究其对丝虫离子通道的影响，包括对SLO-1 K通道的影响。 依莫地平是一种新兴的重要的环八肽类驱虫药，也是一种 对微丝虫和成虫有影响。单一的情绪化治疗可能会让主要的 比现有的大规模药物管理(MDA)计划更先进，这些计划需要定期 杀死成虫的治疗方法。其中一个可能的作用部位是线虫。 SLO-1 K通道，开放通道可抑制运动，但对ALL无效 丝虫。在这里，我们建议研究丝状SLO-1 K通道作为情绪肽的作用部位。 我们的方法将侧重于马来丝虫，但我们也将使用包括对丝虫的研究 和人性化的频道。我们将使用膜片钳，dsRNA击倒，蠕虫运动分析 和Xenopus表达来表征天然SLO-1 K通道的功能特性 来自马来布鲁氏菌。在……里面 目标1， 我们将检验假设马来丝虫SLO-1K 通道是乙基卡马津和情绪苷的唯一靶点。我们将使用 膜片钳记录丝虫肌肉细胞SLO-1钾通道及其作用 推定的目标被击倒。我们建议，在中，表达丝虫，人类和 布鲁氏菌SLO-1 K通道在非洲爪哇卵母细胞中拼接变异体以检验假设 不同种类的丝虫和人类SLO-1钾通道在药物上是可分离的 使用情绪化和K通道激动剂。 目标2 这项提议是创新的，使用多种技术组合来测试 二乙基乙胺嘧啶和情绪化作用于其推测的靶位SLO-1丝虫K通道。 使用这种混合技术的总体影响将是发现 乙基卡马津和莫地平对丝虫性SLO-1 K通道剪接变异体的影响 乙基卡马津和情绪苷作用模式的表征。了解以下内容 这些药物的分子作用部位需要：a)耐药的分子检测； B)设计新药和联合疗法；c)预测和了解敏感性 不同线虫寄生虫种类；以及d)预测寄主毒性

Project Summary Filariasis is a group of neglected tropical diseases produced by infection with microfilaria of Clade III parasitic nematodes that ae transmitted by biting insects. River Blindness caused by Onchocerca volvulus, and Lymphatic Filariasis produced by Brugia malayi are examples of these diseases. River Blindness is caused by parasites that produce scaring of the cornea as well as severe itching and dermatitis; it infects 17 million people in West and Central Africa. Lymphatic filariasis is a debilitating and disfiguring disease, which occurs in 120 million people worldwide. Control of these nematode parasites relies on a small number of anthelmintic drugs, which have a limited spectrum of action. There are no practical macrofilaricides, which kill the adult parasites in the host; and there are concerns that mass microfilaricide chemotherapy will lead to the development of resistance. Diethylcarbamazine is a mainstay for the treatment of lymphatic filariasis in most parts of the world, except in areas where onchocerciasis is present because it is contra-indicated by risks of blindness. It produces rapid clearance of microfilaria and causes ~40% mortality of adult parasites (macrofilaricide). It is striking however, that 68 years after its introduction, we have no proven understanding of the molecular mechanism of its action. Here, we propose to re-invigorate this investigation by studying its effects on filarial ion-channels, including effects on SLO-1 K channels. Emodepside is an emerging and important cyclooctadepsipeptide class of anthelmintic that also has effects on microfilaria and adult filaria. Single emodepside treatments could allow a major advance over existing mass drug administration (MDA) programs which require regular treatments to kill adult parasites. One of the putative sites of action of emodepside is on nematode SLO-1 K channels where opening of the channels inhibits motility, but it is not effective against all filaria. Here we propose to examine filarial SLO-1 K channels as sites of action of emodepside. Our approach will focus on Brugia malayi but we will also use include studies on Onchocerca and human channels. We will use patch-clamp, dsRNA knock down, Worminator motility assays and Xenopus expression to characterize the functional properties of innate SLO-1 K channels from Brugia malayi. In Aim #1, we will test the hypothesis that Brugia malayi SLO-1 K channels are the only target sites of diethylcarbamazine and emodepside. We will use patch-clamp recordings of SLO-1 K channels from Brugia muscle cells and examine effects of knock down of putative targets. We propose, in , to express Onchocerca, human and Brugia SLO-1 K channels splice variants in Xenopus oocytes to test the hypothesis that different species of filaria and human SLO-1 K channels are pharmacologically separable using emodepside and K channel agonists. Aim #2 The proposal is innovative, using a combination of techniques to test the effects of diethylcarbamazine and emodepside on their putative target sites, SLO-1 K channels of filarial. The overall impact of using this mixture of techniques, will be the discovery of effects of diethylcarbamazine and emodepside on filarial SLO-1 K channel splice variants, and an improved characterization of the modes of action of diethylcarbamazine and emodepside. Knowledge of the molecular sites of action of these drugs is required for: a) molecular detection of resistance; b) designing new drugs and combination therapies; c) predicting and understanding sensitivities of different nematode parasite species; and d) predicting host toxicity