Inhibition of Cysteine Proteases via Thiol-Michael Addition Explains the Anti-SARS-CoV-2 and Bioactive Properties of Arteannuin B

Artemisia annua is the plant that producesartemisinin,an endoperoxide-containing sesquiterpenoid used for the treatmentof malaria. A. annua extracts, which contain otherbioactive compounds, have been used to treat other diseases, includingcancer and COVID-19, the disease caused by the virus SARS-CoV-2. Inthis study, a methyl ester derivative of arteannuin B was isolatedwhen A. annua leaves were extracted with a 1:1 mixtureof methanol and dichloromethane. This methyl ester was thought tobe formed from the reaction between arteannuin B and the extractingsolvent, which was supported by the fact that arteannuin B underwent 1,2-addition when it was dissolved in deuteromethanol. In contrast,in the presence of N-acetylcysteine methyl ester,a 1,4-addition (thiol-Michael reaction) occurred. Arteannuin B hinderedthe activity of the SARS CoV-2 main protease (nonstructural protein5, NSP5), a cysteine protease, through time-dependent inhibition.The active site cysteine residue of NSP5 (cysteine-145) formed a covalentbond with arteannuin B as determined by mass spectrometry. In orderto determine whether cysteine adduction by arteannuin B can inhibitthe development of cancer cells, similar experiments were performedwith caspase-8, the cysteine protease enzyme overexpressed in glioblastoma.Time-dependent inhibition and cysteine adduction assays suggestedarteannuin B inhibits caspase-8 and adducts to the active site cysteineresidue (cysteine-360), respectively. Overall, these results enhanceour understanding of how A. annua possesses antiviraland cytotoxic activities.

Automated summary

Artemisia annua produces artemisinin, a compound used to treat malaria. A. annua extracts contain other bioactive compounds and have potential therapeutic effects on cancer and COVID-19.