2. Academic Validation
  3. A HTRF based competitive binding assay for screening specific inhibitors of HIV-1 capsid assembly targeting the C-Terminal domain of capsid

A HTRF based competitive binding assay for screening specific inhibitors of HIV-1 capsid assembly targeting the C-Terminal domain of capsid

  • Antiviral Res. 2019 Sep;169:104544. doi: 10.1016/j.antiviral.2019.104544.
Da-Wei Zhang 1,  Rong-Hua Luo 2,  Lei Xu 3,  Liu-Meng Yang 2,  Xiao-Shuang Xu 3,  Gregory J Bedwell 4,  Alan N Engelman 4,  Yong-Tang Zheng 5,  Shan Chang 6
Affiliations

Affiliations

  • 1 Institute of Bioinformatics and Medical Engineering, School of Electrical and Information Engineering, Jiangsu University of Technology, Changzhou, 213001, China; Department of Cancer Immunology and Virology, Dana-Farber Cancer Institute and Department of Medicine, Harvard Medical School, 450 Brookline Avenue, Boston, MA, 02215, USA.
  • 2 Key Laboratory of Animal Models and Human Disease Mechanisms of the Chinese Academy of Sciences, KIZ-CUHK Joint Laboratory of Bioresources and Molecular Research in Common Diseases, The National Kunming High Level Biosafety Research Center for Nonhuman Primate, Kunming Institute of Zoology, Chinese Academy of Sciences, Kunming, Yunnan, 650223, China.
  • 3 Institute of Bioinformatics and Medical Engineering, School of Electrical and Information Engineering, Jiangsu University of Technology, Changzhou, 213001, China.
  • 4 Department of Cancer Immunology and Virology, Dana-Farber Cancer Institute and Department of Medicine, Harvard Medical School, 450 Brookline Avenue, Boston, MA, 02215, USA.
  • 5 Key Laboratory of Animal Models and Human Disease Mechanisms of the Chinese Academy of Sciences, KIZ-CUHK Joint Laboratory of Bioresources and Molecular Research in Common Diseases, The National Kunming High Level Biosafety Research Center for Nonhuman Primate, Kunming Institute of Zoology, Chinese Academy of Sciences, Kunming, Yunnan, 650223, China. Electronic address: [email protected].
  • 6 Institute of Bioinformatics and Medical Engineering, School of Electrical and Information Engineering, Jiangsu University of Technology, Changzhou, 213001, China. Electronic address: [email protected].
PMID: 31254557 DOI: 10.1016/j.antiviral.2019.104544
Abstract

Due to its multifaceted essential roles in virus replication and extreme genetic fragility, the human immunodeficiency virus type 1 (HIV-1) capsid (CA) protein is a valued therapeutic target. However, CA is as yet unexploited clinically, as there are no Antiviral agents that target it currently on the market. To facilitate the identification of potential HIV-1 CA inhibitors, we established a homogeneous time-resolved fluorescence (HTRF) assay to screen for small molecules that target a biologically active and specific binding pocket in the C-terminal domain of HIV-1 CA (CA CTD). The assay, which is based on competition of small molecules for the binding of a known CA inhibitor (CAI) to the CA CTD, exhibited a signal-to-background ratio (S/B) > 10 and a Z' value > 0.9. In a pilot screen of three kinase inhibitor libraries containing 464 compounds, we identified one compound, TX-1918, as a low micromolecular inhibitor of the HIV-1 CA CTD-CAI interaction (IC50 = 3.81 μM) that also inhibited viral replication at moderate micromolar concentration (EC50 = 15.16 μM) and inhibited CA assembly in vitro. Based on the structure of TX-1918, an additional compound with an Antiviral EC50 of 6.57 μM and cellular cytotoxicity CC50 of 102.55 μM was obtained from a compound similarity search. Thus, the HTRF-based assay has properties that are suitable for screening large compound libraries to identify novel anti-HIV-1 inhibitors targeting the CA CTD.

Keywords

Capsid C-Terminal domain; Drug discovery; HIV-1 capsid; HTRF; TX-1918.

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