A high throughput screen for inhibitors of nematode detoxification genes

线虫解毒基因抑制剂的高通量筛选

• 批准号: 8000247
• 负责人: KEVIN STRANGE
• 金额: $ 11.78万
• 依托单位: MOUNT DESERT ISLAND BIOLOGICAL LAB
• 依托单位国家: 美国
• 财政年份: 2009
• 资助国家: 美国
• 起止时间: 2009-09-25 至 2012-03-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8000247
• 关键词: Drug Metabolic Detoxication; Genes; Nematoda; high throughput screening; inhibitor/antagonist

DESCRIPTION (provided by applicant): Nematodes parasitize ~25% of the human population. Helminth targeting drugs, or anthelmintics, have been used to control parasitic nematodes for decades and many species are evolving multidrug resistance. In diverse organisms, multidrug resistance is mediated by increased expression and activity of enzymes that detoxify xenobiotics. Pharmacological compounds that target xenobiotic detoxification pathways would provide much needed tools for studying multidrug resistance and could greatly increase the useful life of current and future anthelmintics. Few pharmacological compounds are available for studying and targeting multidrug resistance and those that are available are not specific for nematodes and only target a single enzyme or class of enzymes. The transcription factor SKN-1 activates the expression of xenobiotic detoxification genes in the nematode Caenorhabditis elegans. Genetic inhibition of SKN-1 sensitizes C. elegans to diverse xenobiotics, and C. elegans can acquire resistance to anthelmintics by increasing the expression of genes that are regulated by SKN-1. SKN-1 is also essential for the development of embryos. Our recent studies have identified a principal pathway regulating SKN-1 that is highly divergent from pathways that regulate xenobiotic detoxification in mammals. Therefore, SKN-1 is a promising target for the development of drugs that disrupt embryonic development, decrease stress resistance, and inhibit xenobiotic detoxification in nematodes without affecting analogous pathways in humans. The small size, simple culturing characteristics, and genetic tractability of C. elegans make it an ideal system to screen for inhibitors of xenobiotic detoxification genes. The first goal of this project is to develop and optimize a fluorescence-based assay of SKN-1 activity in C. elegans. The second goal of this project is to develop four secondary and counter screens to rapidly prioritize hit compounds and to test the assay in a pilot screen of 2000 compounds. After completion of these goals, a detailed description of the optimized assay will be submitted for entry into the Molecular Libraries Probe Production Centers Network (MLPCN). PUBLIC HEALTH RELEVANCE: Nematodes parasitize ~25% of humans and many strains are resistant to all currently used antiparasitic drugs. In many organisms, drug resistance is caused by over active drug detoxification pathways. The studies proposed in this application will develop, optimize, and validate a screen for pharmacological inhibitors of drug detoxification in nematodes.

描述（由申请人提供）：线虫寄生于约 25% 的人口。蠕虫靶向药物或驱虫药几十年来一直用于控制寄生线虫，许多物种正在进化出多重耐药性。在不同的生物体中，多药耐药性是由异生物质解毒酶的表达和活性增加介导的。针对外源性解毒途径的药理学化合物将为研究多药耐药性提供急需的工具，并可以大大延长当前和未来驱虫药的使用寿命。很少有药理学化合物可用于研究和靶向多药耐药性，并且可用的药理学化合物对线虫不具有特异性，并且仅针对单一酶或一类酶。 转录因子 SKN-1 激活线虫秀丽隐杆线虫中异生物质解毒基因的表达。 SKN-1 的基因抑制使秀丽隐杆线虫对多种外源物质敏感，并且秀丽隐杆线虫可以通过增加 SKN-1 调节的基因的表达来获得对驱虫药的抗性。 SKN-1 对于胚胎发育也至关重要。我们最近的研究发现了调节 SKN-1 的主要途径，该途径与调节哺乳动物异生物质解毒的途径高度不同。因此，SKN-1 是开发药物的一个有前途的靶点，这些药物可以破坏线虫的胚胎发育、降低应激抵抗力和抑制外源性解毒，而不影响人类的类似途径。秀丽隐杆线虫体积小、培养简单、遗传易驯化，使其成为筛选外源解毒基因抑制剂的理想系统。 该项目的首要目标是开发和优化线虫中基于荧光的 SKN-1 活性检测方法。该项目的第二个目标是开发四种二级筛选和反筛选，以快速确定命中化合物的优先级，并在 2000 种化合物的试点筛选中测试检测方法。完成这些目标后，将提交优化测定的详细描述以进入分子库探针生产中心网络 (MLPCN)。 公共卫生相关性：线虫寄生于约 25% 的人类，并且许多菌株对目前使用的所有抗寄生虫药物都具有抗药性。在许多生物体中，耐药性是由药物解毒途径过度活跃引起的。本申请中提出的研究将开发、优化和验证线虫药物解毒的药理学抑制剂的筛选。