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A Fluorescence-based Lymphocyte Assay Suitable for High-throughput Screening of Small Molecules
Published on: March 10, 2017
Development and Characterization of a Fluorescent Probe for GLS1 and the Application for High-Throughput Screening of
Xi Xu1,2, Zijian Kuang1, Jie Han2
1Jiangsu Key Laboratory of Drug Design and Optimization, Department of Medicinal Chemistry, School of Pharmacy , China Pharmaceutical University , Tongjiaxiang 24 , Nanjing 210009 , P.R. China.
Abstract:
Glutaminase (GLS1) is a cancer energy metabolism protein which plays a predominant role in cell growth and proliferation. Because of its major involvement in malignant tumor, small-molecule GLS1 inhibitors are urgently needed to assess its therapeutic potential and for probing their underlying biology function. Recent studies showed that targeting the allosteric binding site represented a promising strategy for identifying potent and selective GLS1 inhibitors. Herein, we present the synthesis of two fluorescent probes targeting the allosteric binding site of GLS1 and their usage as mechanistic tools in multiple applicable assay platform. The fluorescence polarization (FP)-based binding assay enables easy, fast, and reliable screen of allosteric inhibitors from our in-house compound library obtained through click chemistry method. The obtained compound C147 (named as CPU-L1) has been proved to be more potent and with greater solubility than the control compound CB839, which could serve as promising leads for further optimization as novel GLS1 inhibitors.
Insights
Researchers developed novel fluorescent probes to discover glutaminase (GLS1) inhibitors, crucial for cancer metabolism. A new compound, CPU-L1, shows promise as a potent and soluble GLS1 inhibitor for cancer therapy.
Area of Science:
- Biochemistry
- Medicinal Chemistry
- Cancer Biology
Background:
- Glutaminase (GLS1) is a key protein in cancer energy metabolism, vital for tumor cell growth and proliferation.
- Targeting the allosteric binding site of GLS1 is a promising strategy for developing potent and selective inhibitors.
- Small-molecule GLS1 inhibitors are needed for therapeutic assessment and biological function studies.
Purpose of the Study:
- To synthesize novel fluorescent probes targeting the allosteric binding site of GLS1.
- To utilize these probes as mechanistic tools in various assay platforms.
- To identify and characterize novel GLS1 inhibitors.
Main Methods:
- Synthesis of two fluorescent probes utilizing click chemistry.
- Development of a fluorescence polarization (FP)-based binding assay.
- Screening of an in-house compound library for allosteric GLS1 inhibitors.
Main Results:
- Successful synthesis of two fluorescent probes targeting GLS1's allosteric site.
- The FP assay provided a fast and reliable method for screening inhibitors.
- Compound C147 (CPU-L1) demonstrated higher potency and solubility compared to the control CB839.
Conclusions:
- The developed fluorescent probes are effective mechanistic tools for studying GLS1.
- CPU-L1 is a promising lead compound for further optimization as a novel GLS1 inhibitor.
- Targeting GLS1 allosterically offers a viable strategy for cancer therapeutic development.
