A Comparative Study of the Inhibitory Effects of Rheum palmatum L. And Isatis Indigotica Root, on 3CLpro from SARS-CoV-2 Using Both Cell-based and Cell-free Assay Models

Lan, Y.

Purpose: A recent outbreak of a type of coronavirus - SARS-CoV-2 has caused over 200,000 deaths and limited drugs are available. SARS-CoV-2 shows similarity with SARS-CoV. Previous studies have proven that RH121 and indigo are effective in inhibiting 3CLpro from SARS-CoV, but no research has been done to test the two inhibitors on 3CLpro from SARS-CoV-2. This study investigates if RH121 and indigo are still effective in inhibiting 3CLpro from SARS-CoV-2, and compares the two inhibitors in terms of their inhibitory efficiency. Method: For the cell-free assay, substrate Thr-Ser-Ala-Val-Leu-Gln-pNA will be used, and pNA will be released. The absorbance of pNA will be measured using a microplate reader. The cell-based assay will use vero cell lines, vector, plasmid pcDNA3.1-3CLpro-S-Luc, luciferin, and luminometer. The luciferase activity will be tested by Luciferase Reporter Assay System. Possible results: There are seven possible results: (1) RH121 failed to inhibit 3CLpro from SARS-CoV2, but Indigo successfully inhibits 3CLpro from SARS-CoV2. (2) Indigo failed to inhibit 3CLpro from SARS-CoV2, but RH121 successfully inhibits 3CLpro from SARS-CoV-2. (3) Both Indigo and RH121 failed to inhibit 3CLpro from SARS-CoV-2. (4) Indigo has a higher IC50 value than RH121 in cell-based assay, but has a lesser IC50 value than RH121 in cell-free assay. (5) RH121 has a higher IC50 value than Indigo in cell-based assay, but has a lesser IC50 value than Indigo in cell-free assay. (6) Indigo has a higher IC50 value than RH121 in both cell-based assay and cell-free assay. (7) RH121 has a higher IC50 value than Indigo in both cell-based assay and cell-free assay. Conclusion: The result of this study would provide valuable information regarding which inhibitor best inhibits the 3CLpro of SARS-CoV-2 for future trials. Further studies should be done regarding the bioavailability of the two inhibitors and their competitive/uncompetitive nature. The ED50 and TD50 of the two inhibitors should also be investigated in future trials to obtain the therapeutic index and the therapeutic window of RH121 and indigo. © 2021 ACM.