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Using the Protein Chip to Screen Agonists and Antagonists of the Androgen Receptor

National Center of Pharmaceutical Screening, Institute of Materia Medica, Chinese Academy of Medical Sciences and Peking Union Medical College, Beijing, China, Ottawa Health Research Institute, Faculty of Medicine, University of Ottawa, Ottawa, Canada

Ailin Liu

National Center of Pharmaceutical Screening, Institute of Materia Medica, Chinese Academy of Medical Sciences and Peking Union Medical College, Beijing, China

Wei Wang

The Key Laboratory of Biosynthesis, Ministry of Medicine, China, Institute of Materia Medica, Chinese Academy of Medical Sciences and Peking Union Medical College, Beijing, China

Guanhua Du

National Center of Pharmaceutical Screening, Institute of Materia Medica, Chinese Academy of Medical Sciences and Peking Union Medical College, Beijing, China, dugh{at}imm.ac.cn, Institute of Materia Medica Chinese Academy of Medical Sciences and Peking Union Medical College 1 Xian Nong Tan Street Beijing, China, 100050

Based on its important physical and pathological function, the androgen receptor (AR) is regarded as a significant drug target. In this report, the authors describe a novel strategy of protein chip technology to screen agonists and antagonists of AR. First, the AR ligand binding domain (AR-LBD) was expressed in Escherichia coli, purified, and then immobilized on a silane-polysaccharide surface of a protein chip. Second, the affinities of methyltestosterone (MT) and fluorescent-labeled testosterone for the AR-LBD protein chip were determined. Third, a converse strategy of the protein chip was tested to evaluate its reliability as a drug screening method. Fourth, a 10,067-compound library was screened to find new ligands of AR. From the results, the K_d of testosterone and the IC₅₀ of MT are consistent with the literature (0.61 vs. 0.49 nM 2.88 vs. 3.90 nM, respectively). The Z' factor of the high-throughput screening (HTS) method was 0.76, which meets the requirement of drug screening (>0.4). Finally, 3 active ligands of AR were identified with their IC ₅₀ values of 3.63, 2.19, and 1.71 µM, respectively. In summary, the novel strategy of the AR-LBD protein chip was suitable for HTS at the molecular level. (Journal of Biomolecular Screening 2008:276-284)

Key Words: protein chip • androgen receptor • high-throughput screen • E. coli • ligand binding assay

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