GRL-0920, an Indole Chloropyridinyl Ester, Completely Blocks SARS-CoV-2 Infection

• Published in: mBio Vol. 11; no. 4
• Main Authors: Hattori, Shin-ichiro, Higshi-Kuwata, Nobuyo, Raghavaiah, Jakka, Das, Debananda, Bulut, Haydar, Davis, David A., Takamatsu, Yuki, Matsuda, Kouki, Takamune, Nobutoki, Kishimoto, Naoki, Okamura, Tadashi, Misumi, Shogo, Yarchoan, Robert, Maeda, Kenji, Ghosh, Arun K., Mitsuya, Hiroaki
• Format: Journal Article
• Language: English
• Published: United States American Society for Microbiology 20.08.2020
• Subjects: Amino Acid Sequence; Animals; antiviral agents; Antiviral Agents - pharmacology; Betacoronavirus - drug effects; Betacoronavirus - enzymology; Chlorocebus aethiops; Chloroquine - pharmacology; Coronavirus 3C Proteases; Coronavirus Infections - drug therapy; Coronavirus Infections - virology; COVID-19; Cysteine Endopeptidases - chemistry; Cysteine Endopeptidases - genetics; Cysteine Endopeptidases - metabolism; Indoles - chemistry; Indoles - pharmacology; Indoles - therapeutic use; main protease; Models, Molecular; Molecular Biology and Physiology; Pandemics; Pneumonia, Viral - drug therapy; Pneumonia, Viral - virology; SARS-CoV-2; Vero Cells; Viral Nonstructural Proteins - antagonists & inhibitors; Viral Nonstructural Proteins - chemistry; Viral Nonstructural Proteins - genetics; Viral Nonstructural Proteins - metabolism; COVID-19; SARS-CoV-2; antiviral agents; main protease
• ISSN: 2161-2129; 2150-7511; 2150-7511
• DOI: 10.1128/mBio.01833-20

Targeting the main protease (M pro ) of SARS-CoV-2, we identified two indole-chloropyridinyl-ester derivatives, GRL-0820 and GRL-0920, active against SARS-CoV-2, employing RNA-qPCR and immunocytochemistry and show that the two compounds exerted potent activity against SARS-CoV-2. While GRL-0820 and remdesivir blocked SARS-CoV-2 infection, viral breakthrough occurred as examined with immunocytochemistry. In contrast, GRL-0920 completely blocked the infectivity and cytopathic effect of SARS-CoV-2 without significant toxicity. Structural modeling showed that indole and chloropyridinyl of the derivatives interacted with two catalytic dyad residues of M pro , Cys145 and His41, resulting in covalent bonding, which was verified using HPLC/MS. The present data should shed light on the development of therapeutics for COVID-19, and optimization of GRL-0920 based on the present data is essential to develop more-potent anti-SARS-CoV-2 compounds for treating COVID-19. We assessed various newly generated compounds that target the main protease (M pro ) of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) and various previously known compounds reportedly active against SARS-CoV-2, employing RNA quantitative PCR (RNA-qPCR), cytopathicity assays, and immunocytochemistry. Here, we show that two indole-chloropyridinyl-ester derivatives, GRL-0820 and GRL-0920, exerted potent activity against SARS-CoV-2 in cell-based assays performed using VeroE6 cells and TMPRSS2-overexpressing VeroE6 cells. While GRL-0820 and the nucleotide analog remdesivir blocked SARS-CoV-2 infection, viral breakthrough occurred. No significant anti-SARS-CoV-2 activity was found for several compounds reportedly active against SARS-CoV-2 such as lopinavir, nelfinavir, nitazoxanide, favipiravir, and hydroxychroloquine. In contrast, GRL-0920 exerted potent activity against SARS-CoV-2 (50% effective concentration [EC 50 ] = 2.8 μM) and dramatically reduced the infectivity, replication, and cytopathic effect of SARS-CoV-2 without significant toxicity as examined with immunocytochemistry. Structural modeling shows that indole and chloropyridinyl of the derivatives interact with two catalytic dyad residues of M pro , Cys145 and His41, resulting in covalent bonding, which was verified using high-performance liquid chromatography–mass spectrometry (HPLC/MS), suggesting that the indole moiety is critical for the anti-SARS-CoV-2 activity of the derivatives. GRL-0920 might serve as a potential therapeutic for coronavirus disease 2019 (COVID-19) and might be optimized to generate more-potent anti-SARS-CoV-2 compounds. IMPORTANCE Targeting the main protease (M pro ) of SARS-CoV-2, we identified two indole-chloropyridinyl-ester derivatives, GRL-0820 and GRL-0920, active against SARS-CoV-2, employing RNA-qPCR and immunocytochemistry and show that the two compounds exerted potent activity against SARS-CoV-2. While GRL-0820 and remdesivir blocked SARS-CoV-2 infection, viral breakthrough occurred as examined with immunocytochemistry. In contrast, GRL-0920 completely blocked the infectivity and cytopathic effect of SARS-CoV-2 without significant toxicity. Structural modeling showed that indole and chloropyridinyl of the derivatives interacted with two catalytic dyad residues of M pro , Cys145 and His41, resulting in covalent bonding, which was verified using HPLC/MS. The present data should shed light on the development of therapeutics for COVID-19, and optimization of GRL-0920 based on the present data is essential to develop more-potent anti-SARS-CoV-2 compounds for treating COVID-19.