期刊
COMBINATORIAL CHEMISTRY & HIGH THROUGHPUT SCREENING
卷 17, 期 7, 页码 596-609出版社
BENTHAM SCIENCE PUBL LTD
DOI: 10.2174/1386207317666140609122714
关键词
Cancer; fluorescence polarization; HTS; RNA-binding protein; MUSASHI; small-molecule inhibitors
资金
- Commonwealth Foundation for Cancer Research
- Experimental Therapeutics Center of the Memorial Sloan Kettering Cancer Center
- William Randolph Hearst Fund in Experimental Therapeutics
- Lillian S. Wells Foundation
- NIH/NCI Cancer Center [5 P30 CA008748-44]
- US National Institutes of Health(NIH)
- National Institute of Diabetes and Digestive and Kidney Diseases [K01DK084261]
- V-foundation
- Sidney Kimmel Award
- Louis V Gerstner Young Investigator Award
RNA-binding proteins (RBPs) can act as stem cell modulators and oncogenic drivers, but have been largely ignored by the pharmaceutical industry as potential therapeutic targets for cancer. The MUSASHI (MSI) family has recently been demonstrated to be an attractive clinical target in the most aggressive cancers. Therefore, the discovery and development of small molecule inhibitors could provide a novel therapeutic strategy. In order to find novel compounds with MSI RNA binding inhibitory activity, we have developed a fluorescence polarization (FP) assay and optimized it for high throughput screening (HTS) in a 1536-well microtiter plate format. Using a chemical library of 6,208 compounds, we performed pilot screens, against both MSI1 and MSI2, leading to the identification of 7 molecules for MSI1, 15 for MSI2 and 5 that inhibited both. A secondary FP dose-response screen validated 3 MSI inhibitors with IC50 below 10 mu M. Out of the 25 compounds retested in the secondary screen only 8 demonstrated optical interference due to high fluorescence. Utilizing a SYBR-based RNA electrophoresis mobility shift assay (EMSA), we further verified MSI inhibition of the top 3 compounds. Surprisingly, even though several aminoglycosides were present in the library, they failed to demonstrate MSI inhibitor activity challenging the concept that these compounds are pan-active against RBPs. In summary, we have developed an in vitro strategy to identify MSI specific inhibitors using an FP HTS platform, which will facilitate novel drug discovery for this class of RBPs.
作者
我是这篇论文的作者
点击您的名字以认领此论文并将其添加到您的个人资料中。
推荐
暂无数据