Compound library screening using Giardia labmblia high throughput viability assay

描述（由申请人提供）：贾第虫是一种困扰贫困国家的人类病原体，也是美国腹泻暴发的最常见原因。贾第虫已被疾病预防控制中心列为B类生物恐怖主义生物。在美国，最受欢迎的抗贾第虫病药物是甲硝唑或替硝唑，这些化合物具有不良副作用。此外，对药物制度的抗药性和复发的增加也是一个令人关切的问题。因此，显然需要替代药物治疗。我们已经开发了一种基于滋养体ATP含量的生物发光Giarida活力测定法，该测定法适合于高通量筛选，并使该测定法小型化以用于1536孔板中的化合物筛选。我们建议使用最初的CHO细胞和后来的CACO-2细胞筛选NIH化合物库并研究命中化合物的选择性。通过再生长实验确定表现出低IC 50的选择性化合物的最小致死浓度。对于一些最高的命中，将进行贾第虫目标的鉴定，如果成功，将推进到基于结构的化合物优化。 公共卫生相关性：贾第虫是一种人类病原体，每年影响数十亿人，是B类生物恐怖主义生物。贾第虫病治疗具有不良副作用、复发和增加的耐药性。为了促进更好的药物的发现，我们已经开发了贾第虫活力测定法，并将其小型化用于高通量化合物筛选。该试验将用于筛选NIH化合物库并鉴定杀死贾第虫滋养体的化合物。

DESCRIPTION (provided by applicant): Giardia lamblia is a human pathogen afflicting impoverished nations, and the most common cause of outbreaks of diarrhea in the United States. Giardia has been classified by the CDC as a category B bioterrorism organism. The favored anti giardiasis drugs in the US are metronidazole or tinidazole, compounds that have undesirable side effects. Moreover, increasing resistance to drug regimes and recurrence are a concern. It is thus clear that alternative drug treatments are needed. We have developed a bioluminescence Giarida viability assay based on the trophozoites ATP content that is suitable for high throughput screening and miniaturized the assay for compound screening in 1536-well plates. We propose to screen the NIH compound library and investigate the selectivity of hit compounds using initially CHO cells and later CACO-2 cells. The minimum lethal concentration of selective compounds exhibiting low IC50 will be determined by re- growth experiments. For a few top hits, identification of the Giardia target will be undertaken and if successful, advanced to structure-based compound optimization. PUBLIC HEALTH RELEVANCE: Giardia lamblia is a human pathogen afflicting billions of people annually and a category B bioterrorism organism. Giardiasis treatments have undesirable side effects, recurrence, and increasing drug resistance. To facilitate discovery of better drugs, we have developed a Giardia viability assay and miniaturized it for high throughput compound screening. This assay will be used to screen the NIH compound library and identify compounds that kill Giardia trophozoites.