Compound library screening using Giardia labmblia high throughput viability assay

描述（由申请人提供）：贾第鞭毛虫是一种折磨贫困国家的人类病原体，也是美国腹泻爆发的最常见原因。贾第鞭毛虫已被疾病控制与预防中心列为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，这些化合物有不良的副作用。此外，对药物的耐药性增加和复发也是一个问题。因此，很明显，需要替代药物治疗。我们开发了一种基于滋养体ATP含量的贾鞭毛虫生物发光活力测定方法，适用于高通量筛选，并将该方法小型化，适用于1536孔板的化合物筛选。我们建议筛选NIH化合物文库，并首先使用CHO细胞和后来的CACO-2细胞研究命中化合物的选择性。具有低IC50的选择性化合物的最小致死浓度将通过再生长实验确定。对于一些最热门的目标，贾第鞭毛虫靶点的鉴定将进行，如果成功，将推进到基于结构的化合物优化。

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.