I-Corps: A C. elegans model using human genes for high-throughput screening

I-Corps：使用人类基因进行高通量筛选的秀丽隐杆线虫模型

• 批准号: 1664261
• 负责人: Federico Sesti
• 金额: $ 5万
• 依托单位: Rutgers, The State University of New Jersey-RBHS-Robert Wood
• 依托单位国家: 美国
• 项目类别: Standard Grant
• 财政年份: 2017
• 资助国家: 美国
• 起止时间: 2017-01-01 至 2018-06-30
• 项目状态: 已结题
• 来源: https://www.nsf.gov/awardsearch/showAward?AWD_ID=1664261&HistoricalAwards=false
• 关键词: Corps; C.; elegans; model; using

The broader impact/commercial potential of this I-Corps project is in the improvement of drug discovery. Pharmaceutical companies screen millions of compounds before a single agent is selected to undergo human clinical trials. Initial screening of potential therapeutics is conducted in cell culture assays that allow for low-cost evaluation, but are limited in determining therapeutic relevance. This limitation is overcome by additional screening in multiple mammalian models where potential compounds are further assessed for their therapeutic effects, toxicology etc. Yet, the expenses and time associated with mammalian model testing means that only a small percentage of compounds that pass cellculture screening will make it to this stage, effectively overlooking thousands of compounds that may be potent therapeutics. This project?s technology aims at bypassing this bottleneck. It is labor friendly and cost-effective but at the same time will enable companies to test compounds efficacy on an animal model rather than cells in culture thus allowing for further informed decision making in regards to the development of their compounds.This I-Corps project offers an innovative method to screen compounds targeting the actin cytoskeleton and its associated genes. C. elegans worms are engineered to replace a cytoskeletal gene, OSG-1, with its human homolog, (ARHGEF10), and worms lacking OSG-1, (OSG-1 KO), are maintained in 96-well plates each containing a different compound. Worms are subjected to a 4-hr heat shock and scored for survival. Because the mortality of OSG-1 KO worms is significantly higher than ARHGEF10, only compounds that target the actin cytoskeleton pathway can differentially change the mortality rates of the KO compared to the ARHGEF10 worm. Compounds that are efficacious but affect the mortality rates of the KO and the ARHGEF10 worm to the same extent do not impinge on the actin cytoskeleton pathway (they represent a second cut because they might act on pathways and mechanisms that are not conserved in humans). Worms are engineered to express GFP through most of the body. Only living worms express GFP whereas dead ones do not. Survival assessment only requires scoring fluorescent worms. With costs only accruing from agar and plate costs, early compounds can be screened for potency at a fraction of the cost of traditional early phase drug discovery methods.

I-Corps项目的更广泛影响/商业潜力在于改进药物发现。制药公司在选择一种药物进行人体临床试验之前，要筛选数百万种化合物。潜在治疗方法的初步筛选是在细胞培养试验中进行的，这允许低成本的评估，但在确定治疗相关性方面受到限制。通过在多种哺乳动物模型中进行额外筛选，进一步评估潜在化合物的治疗效果、毒理学等，可以克服这一限制。然而，与哺乳动物模型测试相关的费用和时间意味着只有一小部分化合物通过细胞培养筛选才能进入这一阶段，从而有效地忽略了数千种可能成为有效治疗药物的化合物。这个项目吗?美国的技术旨在绕过这一瓶颈。这是劳动友好和成本效益高的，但同时将使公司能够在动物模型上测试化合物的功效，而不是在培养细胞中，从而允许进一步明智的决策，有关他们的化合物的发展。这个I-Corps项目提供了一种创新的方法来筛选针对肌动蛋白细胞骨架及其相关基因的化合物。线虫被改造成用其人类同源基因（ARHGEF10）替换OSG-1细胞骨架基因，缺乏OSG-1的线虫（OSG-1 KO）被维持在96孔板中，每个孔板含有不同的化合物。蠕虫受到4小时的热休克，并为生存评分。由于OSG-1型KO蠕虫的死亡率明显高于ARHGEF10，因此只有靶向肌动蛋白细胞骨架途径的化合物才能区别改变与ARHGEF10型相比的KO蠕虫的死亡率。有效但影响KO和ARHGEF10蠕虫死亡率的化合物在相同程度上不会影响肌动蛋白细胞骨架途径（它们代表了第二次切割，因为它们可能作用于人类不保守的途径和机制）。蠕虫被设计成在身体的大部分表达绿色荧光蛋白。只有活的蠕虫表达GFP，而死的蠕虫则不表达。生存评估只需要给荧光虫打分。由于成本仅来自琼脂和平板成本，早期化合物可以筛选效力，而成本只是传统早期药物发现方法的一小部分。