Yeast-two-hybrid based high-throughput screening to discover SARS-CoV-2 fusion inhibitors by targeting the HR1/HR2 interaction.
10.1016/j.apsb.2025.06.029
- Author:
Jing ZHANG
1
;
Dongsheng LI
1
;
Wenwen ZHOU
1
;
Chao LIU
1
;
Peirong WANG
2
;
Baoqing YOU
1
;
Bingjie SU
1
;
Keyu GUO
1
;
Wenjing SHI
1
;
Tin Mong TIMOTHY YUNG
2
;
Richard Yi TSUN KAO
2
;
Peng GAO
3
;
Yan LI
1
;
Shuyi SI
1
Author Information
1. State Key Laboratory of Bioactive Substances and Functions of Natural Medicines, Institute of Medicinal Biotechnology, Chinese Academy of Medical Sciences & Peking Union Medical College, Beijing 100050, China.
2. Department of Microbiology, Li Ka Shing Faculty of Medicine, the University of Hong Kong, Pok Fu Lam, Hong Kong SAR, China.
3. Applied Oral Sciences & Community Dental Care, Faculty of Dentistry, the University of Hong Kong, Pok Fu Lam, Hong Kong SAR, China.
- Publication Type:Journal Article
- Keywords:
COVID-19;
Fusion inhibitor;
HR1/HR2 interaction;
High-throughput screening;
IMB-9C;
SARS-CoV-2;
Variants;
Yeast-two-hybrid
- From:
Acta Pharmaceutica Sinica B
2025;15(9):4829-4843
- CountryChina
- Language:English
-
Abstract:
The continuous emergence of SARS-CoV-2 variants as well as other potential future coronavirus has challenged the effectiveness of current COVID-19 vaccines. Therefore, there remains a need for alternative antivirals that target processes less susceptible to mutations, such as the formation of six-helix bundle (6-HB) during the viral fusion step of host cell entry. In this study, a novel high-throughput screening (HTS) assay employing a yeast-two-hybrid (Y2H) system was established to identify inhibitors of HR1/HR2 interaction. The compound IMB-9C, which achieved single-digit micromolar inhibition of SARS-CoV-2 and its Omicron variants with low cytotoxicity, was selected. IMB-9C effectively blocks the HR1/HR2 interaction in vitro and inhibits SARS-CoV-2-S-mediated cell-cell fusion. It binds to both HR1 and HR2 through non-covalent interaction and influences the secondary structure of HR1/HR2 complex. In addition, virtual docking and site-mutagenesis results suggest that amino acid residues A930, I931, K933, T941, and L945 are critical for IMB-9C binding to HR1. Collectively, in this study, we have developed a novel screening method for HR1/HR2 interaction inhibitors and identified IMB-9C as a potential antiviral small molecule against COVID-19 and its variants.
