Biotechnology Bulletin ›› 2014, Vol. 0 ›› Issue (4): 187-192.

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The Establishment and Application of a High Throughput Screening Model for Inhibitors of STAT3

Pan Li, Zhang Ning, Niu Guojun, Meng Jing, Liu Xiang, Yang Cheng   

  1. (1. College of Biotechnology,Tianjin University of Science and Technology,Tianjin 300457;
    2. College of Pharmacy,Nankai University,Tianjin 300071;
    3. Tianjin International Joint Academy of Biotechnology and Medicine,Tianjin 300457; 4. College of Life Sciences,Nankai UniversityTianjin 300071)
  • Received:2014-01-13 Online:2014-04-29 Published:2014-04-29

Abstract: It was to establish a viable high-throughput drug screening model for discovering inhibitors of Signal Transducer and Activator of Transcription3, and applied the model to screen potential anti-cancer drugs. The STAT3 gene was amplified with PCR and cloned into the expression vector pET-28a. The recombinant STAT3 was over-expressed and purified, and then was used to bind with specific phosphotyrosine peptides. We established a high-throughput drug screening model based on ELISA to obtain some effective compounds. Further, we tested the effect of them on the growth ability and proliferation of the Hepatoma cells using MTT assay. Results showed that it was not only successfully constructed the expression vector pET-28a-STAT3, but also established a reliable and stable model for drug screening. Besides, 8 248 samples were screened, and a positive sample MDC6 was finally obtained. When the drug concentration was less than 500 μmol/L, the inhibition rate of MDC6 reached 92%. The minimum value of IC50 was 3.37 μmol/L, while at the cellular level it was 15.92 μmol/L. In this work, we developed a repeatable and reliable assay for screening of STAT3 inhibitors.

Key words: STAT3, Target, High-throughput drug screening model, ELISA