In Vitro Screening of New Compounds and Analogs in Cell-Based Assays

这个UW NCDDG的目标将是确定新的化合物，证明有希望作为新的 在临床前测定中的抗癌治疗剂。NCDDG核心B将负责体外试验 项目1和项目2中生产的化合物，以及这些化合物的效力和机制的评价 化合物. PI和指导委员会将使用岩心B的数据选择：1）类似物， 2）需要进一步体外生物测定的类似物，和3） 核心C和项目3应使用的类似物，用于在本地小鼠中进行体内有效性试验 人类癌症的模型。为了每年以低成本有效筛选数百种新化合物， 我们建议以高效的方式在UWCCC小分子筛选中心开展核心B服务。 设施（hts.wisc.edu）。筛选设施是华盛顿大学现有的核心服务设施 综合癌症中心。该设施具有现有的基础设施（细胞培养设施，自动化 液体处理机器人和读板机）、经验丰富的员工和快速筛选的良好记录 成千上万的化学化合物在一个基于细胞和生化分析的多样化阵列。使用 现有的设施将允许更快和更便宜的分析NCDDG候选化合物相比， 在单个学术实验室中运行这些分析或与NCDDG建立新的分析设施 资金核心B的目标是提供一种过滤器，其中化合物移动到体内测试，而不移动到体内测试。 详细建立了大量化合物类似物的作用机制。核心将使用四个 96孔格式的基于细胞的测定，每个都利用机器人技术和实验室的经验。 筛选设施。化合物类似物将在一组八个人癌细胞中测试细胞毒性。 从NCI组的60个细胞系获得的细胞系。为化合物提供更多的机理性见解 功能，我们将集中在两个关键的细胞表型，细胞存活和细胞迁移。化合物将 测定它们触发细胞凋亡的能力和它们改变关键细胞存活介质Akt的能力。一 还将测定化合物子集对体外肿瘤细胞迁移的影响。所有数据将 进行统计分析，以确保获得的新化合物信息的质量， 以促进对任何化合物类似物的未来测试做出最明智的决定。

The goal of this UW NCDDG will be to identify novel compounds that demonstrate promise as leads for new anti-cancer therapeutics in preclinical assays. The NCDDG Core B will be responsible for in vitro assays of the compounds produced in Projects 1 and 2 and evaluation of the potency and mechanism of those compounds. Data from Core B will be used by the PI and the steering committee to select: 1)analogs that should be dropped from further analysis, 2) analogs that require further in vitro biological assays and 3) analogs that should be used by Core C and Project 3 for in vivo efficacy testing in autochthonous mouse models of human cancer. In order to efficiently screen many hundreds of new compounds per year in a cost efficient manner, we propose to perform the Core B services at the Keck-UWCCC Small Molecule Screening Facility (http://hts.wisc.edu). The Screening Facility is an existing core service facility for the UW Comprehensive Cancer Center. This facility has the existing infrastructure (cell culture facility, automated liquid handling robots and plate readers), experienced staff and a track record of rapidly screening thousands of chemical compounds in a diverse array of cell-based and biochemical assays. Use of the existing facility will permit faster and cheaper analysis of the NCDDG candidate compounds compared to running those assays in an individual academic laboratory or establishing a new assay facility with NCDDG funds. The goal of Core B is to provide a filter for which compounds move to in vivo testing and not to establish in detail the mechanism of action of large numbers of compound analogs. The Core will use four cell-based assays in 96-well formats that each take advantage of the robotics and experience of the screening facility. Compound analogs will be tested for cytotoxicity in a panel of eight human cancer cell lines obtained from the NCI panel of 60 cell lines. To provide more mechanistic insights into compound function we will focus on two key cellular phenotypes, cell survival and cell migration. Compounds will be assayed for their ability to trigger apoptosis and for their ability to alter the key cell survival mediator Akt. A subset of compounds will also be assayed for their effect on tumor cell migration in vitro. All data will be subject to statistical analysis to ensure the quality of information that is obtained on the new compounds and to facilitate the most informed decisions about the future testing of any compound analog.