A research team led by Prof. ZHENG Yongtang from Kunming Institute of Zoology of the Chinese Academy of Sciences, collaborating with Prof. YANG Shengyong and Prof. LEI Jian from West China Hospital of Sichuan University, developed anti-SARS-CoV-2 inhibitors. The finding was published in Science. 

The COVID-19 pandemic caused by the SARS-CoV-2 virus continually poses serious threats to global public health. To date, there is still a lack of specific drugs against SARS-CoV-2. 

The main protease (M^pro) plays a critical role in SARS-CoV-2 replication, and it is conservative in all coronaviruses. Human doesn’t express an enzyme as the M^pro of SARS-CoV-2, making M^pro to be a superior target to develop anti-SARS-CoV-2 inhibitors. Although a few inhibitors of the M^pro have been reported, their anti-viral activities are only determined in vitro.  

Based on the bicycloproline fragment of approved anti-HCV drugs, telaprevir and boceprevir, the researchers first designed and synthesized 32 new bicycloproline-containing small molecules using rational drug design strategy. All compounds displayed potential in vitro anti-SARS-CoV-2 M^pro activities with the most potential inhibitor’s half-maximal inhibitory concentration (IC50) of 7.6 nM. 

Then, they determined the co-crystal structure of M^pro with MI-23, one inhibitor which showed the strongest anti-SARS-CoV-2 activity, and found that MI-23 covalently interacted with the active site of M^pro. The interaction occupied three sites, P1, P2, and P3, and formed hydrogen bonds with many critical amino acids. The structure data suggests that the interaction pattern is consistent with their design concept. 

By analyzing the cellular anti-SARS-CoV-2 activity as well as pharmacokinetic properties and safety, the researchers evaluated the designed compounds in rats. 20 molecules selected from 32 inhibitors displayed good antiviral activity in multiple cell lines. Six molecules with potential cellular antiviral activity were evaluated in rats, and two molecules, MI-09 and MI-30, displayed good pharmacokinetic properties and safety. 

Furthermore, they evaluated in vivo anti-SARS-CoV-2 activity of MI-09 and MI-30 using an hACE2 transgenic mouse model. Oral or intraperitoneal treatment with MI-09 or MI-30 significantly reduced lung viral loads and lung lesions. The mice receiving MI-09 or MI-30 displayed mild alveolar septal thickening and inflammatory cell infiltration, compared with the control mice showing moderate symptoms. Fewer neutrophils and macrophages were observed in the lung of mice treated with MI-09 or MI-30, compared with the control mice, suggesting that MI-09 and MI-30 suppress immune cell infiltration. 

This study reported a series of new potential M^pro inhibitors and evaluated their mechanism and antiviral activity in molecular, cellular and animal level. The data on pharmacokinetic properties and safety demonstrated that these compounds could be potentially developed to be anti-SARS-CoV-2 drugs.