1. Development of a high-throughput fluorescence polarization assay for the discovery of EZH2-EED interaction inhibitors.
- Author
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Zhu MR, Du DH, Hu JC, Li LC, Liu JQ, Ding H, Kong XQ, Jiang HL, Chen KX, and Luo C
- Subjects
- Apomorphine pharmacology, Enhancer of Zeste Homolog 2 Protein chemical synthesis, Ergonovine pharmacology, Fluorescence Polarization, Humans, Hydrogen-Ion Concentration, Limit of Detection, Nifedipine pharmacology, Oxyphenbutazone pharmacology, Peptide Fragments antagonists & inhibitors, Peptide Fragments chemical synthesis, Peptide Fragments metabolism, Protein Binding drug effects, Temperature, Enhancer of Zeste Homolog 2 Protein antagonists & inhibitors, Enhancer of Zeste Homolog 2 Protein metabolism, High-Throughput Screening Assays methods, Polycomb Repressive Complex 2 antagonists & inhibitors, Polycomb Repressive Complex 2 metabolism, Protein Multimerization drug effects
- Abstract
Aberrant activity of enhancer of zeste homolog 2 (EZH2) is associated with a wide range of human cancers. The interaction of EZH2 with embryonic ectoderm development (EED) is required for EZH2's catalytic activity. Inhibition of the EZH2-EED complex thus represents a novel strategy for interfering with the oncogenic potentials of EZH2 by targeting both its catalytic and non-catalytic functions. To date, there have been no reported high-throughput screening (HTS) assays for inhibitors acting at the EZH2-EED interface. In this study, we developed a fluorescence polarization (FP)-based HTS system for the discovery of EZH2-EED interaction inhibitors. The tracer peptide sequences, positions of fluorescein labeling, and a variety of physicochemical conditions were optimized. The high Z' factors (>0.9) at a variety of DMSO concentrations suggested that this system is robust and suitable for HTS. The minimal sequence requirement for the EZH2-EED interaction was determined by using this system. A pilot screening of an in-house compound library containing 1600 FDA-approved drugs identified four compounds (apomorphine hydrochloride, oxyphenbutazone, nifedipine and ergonovine maleate) as potential EZH2-EED interaction inhibitors.
- Published
- 2018
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