Chloropyridinyl Esters of Nonsteroidal Anti-Inflammatory Agents and Related Derivatives as Potent SARS-CoV-2 3CL Protease Inhibitors.

Arun K Ghosh,Monika Yadav,Carlos A Brito-Sierra,Andrew D Mesecar,Dana Shahabi,Shin-Ichiro Hattori,Brandon J Anson,Devika Sirohi,Satish Kovela,Connie Bonham,Richard Kuhn,Emma K Lendy,Hiroaki Mitsuya

doi:10.3390/molecules26195782

Abstract

We report the design and synthesis of a series of new 5-chloropyridinyl esters of salicylic acid, ibuprofen, indomethacin, and related aromatic carboxylic acids for evaluation against SARS-CoV-2 3CL protease enzyme. These ester derivatives were synthesized using EDC in the presence of DMAP to provide various esters in good to excellent yields. Compounds are stable and purified by silica gel chromatography and characterized using 1H-NMR, 13C-NMR, and mass spectral analysis. These synthetic derivatives were evaluated in our in vitro SARS-CoV-2 3CLpro inhibition assay using authentic SARS-CoV-2 3CLpro enzyme. Compounds were also evaluated in our in vitro antiviral assay using quantitative VeroE6 cell-based assay with RNAqPCR. A number of compounds exhibited potent SARS-CoV-2 3CLpro inhibitory activity and antiviral activity. Compound 9a was the most potent inhibitor, with an enzyme IC50 value of 160 nM. Compound 13b exhibited an enzyme IC50 value of 4.9 µM. However, it exhibited a potent antiviral EC50 value of 24 µM in VeroE6 cells. Remdesivir, an RdRp inhibitor, exhibited an antiviral EC50 value of 2.4 µM in the same assay. We assessed the mode of inhibition using mass spectral analysis which suggested the formation of a covalent bond with the enzyme. To obtain molecular insight, we have created a model of compound 9a bound to SARS-CoV-2 3CLpro in the active site.

Highlights

Novel Coronavirus disease 2019 (COVID-19) caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is responsible for the ongoing COVID-19 pandemic [1,2]
SARS-CoV-2 belongs to a family of beta-coronaviruses, including SARS-CoV and MERS-CoV, which were responsible for earlier outbreaks of SARS and MERS in 2003 and 2012, respectively [10,11,12]
SARS-CoV-2 encodes two proteases, a 3-chymotrypsinlike cysteine protease (3CLpro) known as main protease (Mpro) and a papain-like protease (PLpro) for proteolytic processing of viral replication and maturation. [13,14] Both of these proteases are essential for replication of SARS-CoV-2 and other coronaviruses

Summary

Introduction

Novel Coronavirus disease 2019 (COVID-19) caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is responsible for the ongoing COVID-19 pandemic [1,2]. Based upon our X-ray structural studies and mode of inhibition, we have further investigated other aromatic and heteroaromatic scaffolds and their ability to block SARS-CoV-2 3CL protease activity as well as antiviral activity in VeroE6 cells [29]. We plan to synthesize 5-chloropyridin-3-yl esters of widely used nonsteroidal anti-inflammatory agents (NSAIDs) [30,31] and evaluate their potential as irreversible inhibitors of SARS-CoV-2 3CLpro enzyme. Such acylated thioesters of these NSAIDs would hydrolyze slowly over time and release parent NSAIDs in the cell [32].

Biological Evaluation

Chemistry

IC50 Value Determination

Mass Analysis of Enzyme-Inhibitor Complex

Findings

Conclusions