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1.
Steroids ; 188: 109120, 2022 Dec.
Article in English | MEDLINE | ID: covidwho-2113189

ABSTRACT

The present work reports simple and effective protocol for preparing 6α-nitro-5α-cholestano[7α,5-cd] pyrazolines (4-7) by the reaction of 7α-bromo-6-nitrocholest-5-enes (1-3) with hydrazine hydrate under reflux [the substrate (2) gave products (5) and (6) and the later on acetylation with AC2O/Py gave (7)]. In the case of reaction of 3ß-hydroxy analogue (3) with hydrazine, however, 6α-nitro-5α-cholestano [3α,5-cd] pyrazoline (8) and 6α-nitro-3ß, 5-oxido-5ß-cholestane (9) were obtained. The probable mechanism of the formation of pyrazolines has also been outlined. In the current pandemic coronavirus disease 2019 scenario, the in-silico study was performed with reactants (1-3), their products (4-9) against SARS-CoV-2 omicron protease (PDB ID:7T9L) for knowing significant interactions between them. Docking results give information that both reactants and products have binding energies ranges from -5.7 to 7.7 kcal/mol and strong interactions with various hydrophilic and hydrophobic amino acids such as ASP, PRO, PHE, SER and LEU which are significant residues playing important role in SARS-CoV-2 Omicron main protease (Mpro).


Subject(s)
COVID-19 , Coronavirus 3C Proteases , SARS-CoV-2 , Humans , COVID-19/drug therapy , Hydrazines , Molecular Docking Simulation , Molecular Dynamics Simulation , Peptide Hydrolases , SARS-CoV-2/enzymology , SARS-CoV-2/metabolism , Coronavirus 3C Proteases/antagonists & inhibitors
2.
Biochemistry ; 61(22): 2495-2505, 2022 Nov 15.
Article in English | MEDLINE | ID: covidwho-2096611

ABSTRACT

The main protease (Mpro) of SARS-CoV-2 is essential for viral replication and has been the focus of many drug discovery efforts since the start of the COVID-19 pandemic. Nirmatrelvir (NTV) is an inhibitor of SARS-CoV-2 Mpro that is used in the combination drug Paxlovid for the treatment of mild to moderate COVID-19. However, with increased use of NTV across the globe, there is a possibility that future SARS-CoV-2 lineages will evolve resistance to NTV. Early prediction and monitoring of resistance mutations could allow for measures to slow the spread of resistance and for the development of new compounds with activity against resistant strains. In this work, we have used in silico mutational scanning and inhibitor docking of Mpro to identify potential resistance mutations. Subsequent in vitro experiments revealed five mutations (N142L, E166M, Q189E, Q189I, and Q192T) that reduce the potency of NTV and of a previously identified non-covalent cyclic peptide inhibitor of Mpro. The E166M mutation reduced the half-maximal inhibitory concentration (IC50) of NTV 24-fold and 118-fold for the non-covalent peptide inhibitor. Our findings inform the ongoing genomic surveillance of emerging SARS-CoV-2 lineages.


Subject(s)
Antiviral Agents , COVID-19 , Coronavirus 3C Proteases , Drug Resistance, Viral , Protease Inhibitors , SARS-CoV-2 , Humans , Antiviral Agents/pharmacology , Antiviral Agents/chemistry , COVID-19/drug therapy , COVID-19/virology , Molecular Docking Simulation , Mutation , Pandemics , Protease Inhibitors/pharmacology , Protease Inhibitors/chemistry , SARS-CoV-2/drug effects , SARS-CoV-2/genetics , Drug Resistance, Viral/genetics , Coronavirus 3C Proteases/antagonists & inhibitors
3.
Bioorg Chem ; 129: 106185, 2022 12.
Article in English | MEDLINE | ID: covidwho-2060449

ABSTRACT

The evolving SARS-CoV-2 epidemic buffets the world, and the concerted efforts are needed to explore effective drugs. Mpro is an intriguing antiviral target for interfering with viral RNA replication and transcription. In order to get potential anti-SARS-CoV-2 agents, we established an enzymatic assay using a fluorogenic substrate to screen the inhibitors of Mpro. Fortunately, Acriflavine (ACF) and Proflavine Hemisulfate (PRF) with the same acridine scaffold were picked out for their good inhibitory activity against Mpro with IC50 of 5.60 ± 0.29 µM and 2.07 ± 0.01 µM, respectively. Further evaluation of MST assay and enzymatic kinetics experiment in vitro showed that they had a certain affinity to SARS-CoV-2 Mpro and were both non-competitive inhibitors. In addition, they inhibited about 90 % HCoV-OC43 replication in BHK-21 cells at 1 µM. Both compounds showed nano-molar activities against SARS-CoV-2 virus, which were superior to GC376 for anti-HCoV-43, and equivalent to the standard molecule remdesivir. Our study demonstrated that ACF and PRF were inhibitors of Mpro, and ACF has been previously reported as a PLpro inhibitor. Taken together, ACF and PRF might be dual-targeted inhibitors to provide protection against infections of coronaviruses.


Subject(s)
Acriflavine , COVID-19 , Coronavirus 3C Proteases , Cysteine Proteinase Inhibitors , Proflavine , SARS-CoV-2 , Viral Protease Inhibitors , Acriflavine/pharmacology , COVID-19/drug therapy , Proflavine/pharmacology , SARS-CoV-2/drug effects , SARS-CoV-2/enzymology , Coronavirus 3C Proteases/antagonists & inhibitors , Cysteine Proteinase Inhibitors/pharmacology , Viral Protease Inhibitors/pharmacology , Mesocricetus , Animals , Cricetinae , Cell Line , Virus Replication/drug effects
4.
Phys Chem Chem Phys ; 24(37): 22898-22904, 2022 Sep 28.
Article in English | MEDLINE | ID: covidwho-2036937

ABSTRACT

Coronavirus 3C-like protease (3CLpro) is found in SARS-CoV-2 virus, which causes COVID-19. 3CLpro controls virus replication and is a major target for target-based antiviral discovery. As reported by Pfizer, Nirmatrelvir (PF-07321332) is a competitive protein inhibitor and a clinical candidate for orally delivered medication. However, the binding mechanisms between Nirmatrelvir and 3CLpro complex structures remain unknown. This study incorporated ligand Gaussian accelerated molecular dynamics, the one-dimensional and two-dimensional potential of mean force, normal molecular dynamics, and Kramers' rate theory to determine the binding and dissociation rate constants (koff and kon) associated with the binding of the 3CLpro protein to the Nirmatrelvir inhibitor. The proposed approach addresses the challenges in designing small-molecule antiviral drugs.


Subject(s)
Antiviral Agents , Coronavirus 3C Proteases , SARS-CoV-2 , Antiviral Agents/chemistry , Antiviral Agents/pharmacology , Coronavirus 3C Proteases/antagonists & inhibitors , Cysteine Endopeptidases/metabolism , Lactams , Leucine , Ligands , Molecular Docking Simulation , Molecular Dynamics Simulation , Nitriles , Peptide Hydrolases/metabolism , Proline , SARS-CoV-2/drug effects
5.
J Mol Model ; 28(9): 279, 2022 Aug 29.
Article in English | MEDLINE | ID: covidwho-2014173

ABSTRACT

Main protease (Mpro) plays a key role in replication of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). This study was designed for finding natural inhibitors of SARS-CoV-2 Mpro by in silico methods. To this end, the co-crystal structure of Mpro with telaprevir was explored and receptor-ligand pharmacophore models were developed and validated using pharmit. The database of "ZINC Natural Products" was screened, and 288 compounds were filtered according to pharmacophore features. In the next step, Lipinski's rule of five was applied and absorption, distribution, metabolism, excretion, and toxicity (ADMET) of the filtered compounds were calculated using in silico methods. The resulted 15 compounds were docked into the active site of Mpro and those with the highest binding scores and better interaction including ZINC61991204, ZINC67910260, ZINC61991203, and ZINC08790293 were selected. Further analysis by molecular dynamic simulation studies showed that ZINC61991203 and ZINC08790293 dissociated from Mpro active site, while ZINC426421106 and ZINC5481346 were stable. Root mean square deviation (RMSD), radius of gyration (Rg), number of hydrogen bonds between ligand and protein during the time of simulation, and root mean square fluctuations (RMSF) of protein and ligands were calculated, and components of binding free energy were calculated using the molecular mechanic/Poisson-Boltzmann surface area (MM/PBSA) method. The result of all the analysis indicated that ZINC61991204 and ZINC67910260 are drug-like and nontoxic and have a high potential for inhibiting Mpro.


Subject(s)
Coronavirus 3C Proteases , Protease Inhibitors , SARS-CoV-2 , COVID-19 , Coronavirus 3C Proteases/antagonists & inhibitors , Humans , Ligands , Molecular Docking Simulation , Molecular Dynamics Simulation , Protease Inhibitors/pharmacology , SARS-CoV-2/drug effects
6.
J Virol ; 96(17): e0090722, 2022 09 14.
Article in English | MEDLINE | ID: covidwho-2001774

ABSTRACT

The rapid global emergence of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) has caused serious health problems, highlighting the urgent need for antiviral drugs. The viral main protease (Mpro) plays an important role in viral replication and thus remains the target of choice for the prevention or treatment of several viral diseases due to high sequence and structural conservation. Prolonged use of viral protease inhibitors can lead to the development of mutants resistant to those inhibitors and to many of the available antiviral drugs. Here, we used feline infectious peritonitis virus (FIPV) as a model to investigate its development of resistance under pressure from the Mpro inhibitor GC376. Passage of wild-type (WT) FIPV in the presence of GC376 selected for a mutation in the nsp12 region where Mpro cleaves the substrate between nsp12 and nsp13. This mutation confers up to 3-fold resistance to GC376 and nirmatrelvir, as determined by EC50 assay. In vitro biochemical and cellular experiments confirmed that FIPV adapts to the stress of GC376 by mutating the nsp12 and nsp13 hydrolysis site to facilitate cleavage by Mpro and release to mediate replication and transcription. Finally, we demonstrate that GC376 cannot treat FIP-resistant mutants that cause FIP in animals. Taken together, these results suggest that Mpro affects the replication of coronaviruses (CoVs) and the drug resistance to GC376 by regulating the amount of RdRp from a distant site. These findings provide further support for the use of an antiviral drug combination as a broad-spectrum therapy to protect against contemporary and emerging CoVs. IMPORTANCE CoVs cause serious human infections, and antiviral drugs are currently approved to treat these infections. The development of protease-targeting therapeutics for CoV infection is hindered by resistance mutations. Therefore, we should pay attention to its resistance to antiviral drugs. Here, we identified possible mutations that lead to relapse after clinical treatment of FIP. One amino acid substitution in the nsp12 polymerase at the Mpro cleavage site provided low-level resistance to GC376 after selection exposure to the GC376 parental nucleoside. Resistance mutations enhanced FIPV viral fitness in vitro and attenuated the therapeutic effect of GC376 in an animal model of FIPV infection. Our research explains the evolutionary characteristics of coronaviruses under antiviral drugs, which is helpful for a more comprehensive understanding of the molecular basis of virus resistance and provides important basic data for the effective prevention and control of CoVs.


Subject(s)
Antiviral Agents , Coronavirus 3C Proteases , Coronavirus, Feline , Drug Resistance, Viral , Mutation , Protease Inhibitors , Animals , Antiviral Agents/pharmacology , Cats/virology , Coronavirus 3C Proteases/antagonists & inhibitors , Coronavirus 3C Proteases/genetics , Coronavirus 3C Proteases/metabolism , Coronavirus, Feline/drug effects , Coronavirus, Feline/enzymology , Coronavirus, Feline/genetics , Drug Resistance, Viral/genetics , Protease Inhibitors/pharmacology
7.
Drug Dev Res ; 83(7): 1623-1640, 2022 Nov.
Article in English | MEDLINE | ID: covidwho-1999851

ABSTRACT

The global emergence of coronavirus disease 2019 (COVID-19) has caused substantial human casualties. Clinical manifestations of this disease vary from asymptomatic to lethal, and the symptomatic form can be associated with cytokine storm and hyperinflammation. In face of the urgent demand for effective drugs to treat COVID-19, we have searched for candidate compounds using in silico approach followed by experimental validation. Here we identified celastrol, a pentacyclic triterpene isolated from Tripterygium wilfordii Hook F, as one of the best compounds out of 39 drug candidates. Celastrol reverted the gene expression signature from severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2)-infected cells and irreversibly inhibited the recombinant forms of the viral and human cysteine proteases involved in virus invasion, such as Mpro (main protease), PLpro (papain-like protease), and recombinant human cathepsin L. Celastrol suppressed SARS-CoV-2 replication in human and monkey cell lines and decreased interleukin-6 (IL-6) secretion in the SARS-CoV-2-infected human cell line. Celastrol acted in a concentration-dependent manner, with undetectable signs of cytotoxicity, and inhibited in vitro replication of the parental and SARS-CoV-2 variant. Therefore, celastrol is a promising lead compound to develop new drug candidates to face COVID-19 due to its ability to suppress SARS-CoV-2 replication and IL-6 production in infected cells.


Subject(s)
Antiviral Agents , COVID-19 , Coronavirus 3C Proteases , Pentacyclic Triterpenes , Humans , Antiviral Agents/pharmacology , Coronavirus 3C Proteases/antagonists & inhibitors , COVID-19/drug therapy , Interleukin-6 , Molecular Docking Simulation , Pentacyclic Triterpenes/pharmacology , Protease Inhibitors/pharmacology , SARS-CoV-2/drug effects , Viral Nonstructural Proteins/genetics , Viral Nonstructural Proteins/metabolism
8.
Eur J Med Chem ; 238: 114508, 2022 Aug 05.
Article in English | MEDLINE | ID: covidwho-1982957

ABSTRACT

The COVID-19 posed a serious threat to human life and health, and SARS-CoV-2 Mpro has been considered as an attractive drug target for the treatment of COVID-19. Herein, we report 2-(furan-2-ylmethylene)hydrazine-1-carbothioamide derivatives as novel inhibitors of SARS-CoV-2 Mpro developed by in-house library screening and biological evaluation. Similarity search led to the identification of compound F8-S43 with the enzymatic IC50 value of 10.76 µM. Further structure-based drug design and synthetic optimization uncovered compounds F8-B6 and F8-B22 as novel non-peptidomimetic inhibitors of Mpro with IC50 values of 1.57 µM and 1.55 µM, respectively. Moreover, enzymatic kinetic assay and mass spectrometry demonstrated that F8-B6 was a reversible covalent inhibitor of Mpro. Besides, F8-B6 showed low cytotoxicity with CC50 values of more than 100 µM in Vero and MDCK cells. Overall, these novel SARS-CoV-2 Mpro non-peptidomimetic inhibitors provide a useful starting point for further structural optimization.


Subject(s)
COVID-19 , Coronavirus 3C Proteases , Furans , SARS-CoV-2 , Antiviral Agents/chemistry , Antiviral Agents/pharmacology , COVID-19/drug therapy , Coronavirus 3C Proteases/antagonists & inhibitors , Coronavirus 3C Proteases/chemistry , Coronavirus 3C Proteases/metabolism , Drug Discovery/methods , Furans/chemistry , Furans/pharmacology , Humans , Hydrazines/pharmacology , Molecular Docking Simulation , Protease Inhibitors/chemistry , Protease Inhibitors/pharmacology , SARS-CoV-2/drug effects , SARS-CoV-2/enzymology
9.
Biochem Biophys Res Commun ; 616: 8-13, 2022 08 06.
Article in English | MEDLINE | ID: covidwho-1982607

ABSTRACT

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) provoked a pandemic of acute respiratory disease, namely coronavirus disease 2019 (COVID-19). Currently, effective drugs for this disease are urgently warranted. Anisodamine is a traditional Chinese medicine that is predicted as a potential therapeutic drug for the treatment of COVID-19. Therefore, this study aimed to investigate its antiviral activity and crucial targets in SARS-CoV-2 infection. SARS-CoV-2 and anisodamine were co-cultured in Vero E6 cells, and the antiviral activity of anisodamine was assessed by immunofluorescence assay. The antiviral activity of anisodamine was further measured by pseudovirus entry assay in HEK293/hACE2 cells. Finally, the predictions of crucial targets of anisodamine on SARS-CoV-2 were analyzed by molecular docking studies. We discovered that anisodamine suppressed SARS-CoV-2 infection in Vero E6 cells, and reduced the SARS-CoV-2 pseudovirus entry to HEK293/hACE2 cells. Furthermore, molecular docking studies indicated that anisodamine may target SARS-CoV-2 main protease (Mpro) with the docking score of -6.63 kcal/mol and formed three H-bonds with Gly143, Cys145, and Cys44 amino acid residues at the predicted active site of Mpro. This study suggests that anisodamine is a potent antiviral agent for treating COVID-19.


Subject(s)
COVID-19 , Coronavirus 3C Proteases , SARS-CoV-2 , Solanaceous Alkaloids , Antiviral Agents/chemistry , Antiviral Agents/pharmacology , COVID-19/drug therapy , COVID-19/virology , Coronavirus 3C Proteases/antagonists & inhibitors , Coronavirus 3C Proteases/drug effects , Coronavirus 3C Proteases/metabolism , HEK293 Cells , Humans , Molecular Docking Simulation , Peptide Hydrolases , Protease Inhibitors/pharmacology , Solanaceous Alkaloids/pharmacology , Viral Nonstructural Proteins/chemistry
10.
Antiviral Res ; 204: 105350, 2022 08.
Article in English | MEDLINE | ID: covidwho-1944193

ABSTRACT

Two years after its emergence, SARS-CoV-2 still represents a serious and global threat to human health. Antiviral drug development usually takes a long time and, to increase the chances of success, chemical variability of hit compounds represents a valuable source for the discovery of new antivirals. In this work, we applied a platform of variably oriented virtual screening campaigns to seek for novel chemical scaffolds for SARS-CoV-2 main protease (Mpro) inhibitors. The study on the resulting 30 best hits led to the identification of a series of structurally unrelated Mpro inhibitors. Some of them exhibited antiviral activity in the low micromolar range against SARS-CoV-2 and other human coronaviruses (HCoVs) in different cell lines. Time-of-addition experiments demonstrated an antiviral effect during the viral replication cycle at a time frame consistent with the inhibition of SARS-CoV-2 Mpro activity. As a proof-of-concept, to validate the pharmaceutical potential of the selected hits against SARS-CoV-2, we rationally optimized one of the hit compounds and obtained two potent SARS-CoV-2 inhibitors with increased activity against Mpro both in vitro and in a cellular context, as well as against SARS-CoV-2 replication in infected cells. This study significantly contributes to the expansion of the chemical variability of SARS-CoV-2 Mpro inhibitors and provides new scaffolds to be exploited for pan-coronavirus antiviral drug development.


Subject(s)
Antiviral Agents , Coronavirus 3C Proteases , Protease Inhibitors , SARS-CoV-2 , Antiviral Agents/chemistry , Antiviral Agents/pharmacology , Coronavirus 3C Proteases/antagonists & inhibitors , Molecular Docking Simulation , Protease Inhibitors/chemistry , SARS-CoV-2/drug effects , SARS-CoV-2/enzymology
11.
PLoS One ; 17(2): e0263251, 2022.
Article in English | MEDLINE | ID: covidwho-1938414

ABSTRACT

The main protease (3CLpro) is one of the essential components of the SARS-CoVs viral life cycle, which makes it an interesting target for overpowering these viruses. Although many covalent and noncovalent inhibitors have been designed to inhibit this molecular target, none have gained FDA approval as a drug. Because of the high rate of COVID-19 pandemic development, in addition to laboratory research, we require in silico methods to accelerate rational drug design. The unbinding pathways of two SARS-CoV and SARS-CoV-2 3CLpro noncovalent inhibitors with the PDB IDs: 3V3M, 4MDS, 6W63, 5RF7 were explored from a comparative perspective using unbiased molecular dynamics (UMD) simulations. We uncovered common weak points for selected inhibitors that could not interact significantly with a binding pocket at specific residues by all their fragments. So water molecules entered the free binding S regions and weakened protein-inhibitor fundamental interactions gradually. N142, G143, and H163 are the essential residues, which cause key protein-ligand interactions in the binding pocket. We believe that these results will help design new potent inhibitors against SARS-CoV-2.


Subject(s)
Antiviral Agents/pharmacology , COVID-19/drug therapy , Coronavirus 3C Proteases/antagonists & inhibitors , Protease Inhibitors/pharmacology , SARS-CoV-2/drug effects , Antiviral Agents/chemistry , COVID-19/virology , Coronavirus 3C Proteases/chemistry , Coronavirus 3C Proteases/metabolism , Drug Design , Humans , Molecular Docking Simulation , Molecular Dynamics Simulation , Protease Inhibitors/chemistry , SARS-CoV-2/chemistry , SARS-CoV-2/metabolism
12.
J Mol Biol ; 434(16): 167706, 2022 08 30.
Article in English | MEDLINE | ID: covidwho-1914637

ABSTRACT

New variants of the severe acute respiratory syndrome Coronavirus 2 (SARS-CoV-2) emerged and spread rapidly all over the world, which strongly supports the need for pharmacological options to complement vaccine strategies. Main protease (Mpro or 3CLpro) is a critical enzyme in the life cycle of SARS-CoV-2 and appears to be highly conserved among different genera of coronaviruses, making it an ideal target for the development of drugs with broad-spectrum property. PF-07304814 developed by Pfizer is an intravenously administered inhibitor targeting SARS-CoV-2 Mpro. Here we showed that PF-07304814 displays broad-spectrum inhibitory activity against Mpros from multiple coronaviruses. Crystal structures of Mpros of SARS-CoV-2, SARS-CoV, MERS-CoV, and HCoV-NL63 bound to the inhibitor PF-07304814 revealed a conserved ligand-binding site, providing new insights into the mechanism of inhibition of viral replication. A detailed analysis of these crystal structures complemented by comprehensive comparison defined the key structural determinants essential for inhibition and illustrated the binding mode of action of Mpros from different coronaviruses. In view of the importance of Mpro for the medications of SARS-CoV-2 infection, insights derived from the present study should accelerate the design of pan-coronaviral main protease inhibitors that are safer and more effective.


Subject(s)
Coronavirus 3C Proteases , Coronavirus Protease Inhibitors , Indoles , Leucine , Pyrrolidinones , SARS-CoV-2 , Coronavirus 3C Proteases/antagonists & inhibitors , Coronavirus 3C Proteases/chemistry , Coronavirus Protease Inhibitors/chemistry , Coronavirus Protease Inhibitors/pharmacology , Drug Design , Humans , Indoles/chemistry , Indoles/pharmacology , Leucine/chemistry , Leucine/pharmacology , Ligands , Protein Binding , Pyrrolidinones/chemistry , Pyrrolidinones/pharmacology , SARS-CoV-2/drug effects , SARS-CoV-2/enzymology
13.
Viruses ; 14(5)2022 05 04.
Article in English | MEDLINE | ID: covidwho-1820420

ABSTRACT

The coronavirus disease 2019 (COVID-19) pandemic is caused by the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), a recently emerged human coronavirus. COVID-19 vaccines have proven to be successful in protecting the vaccinated from infection, reducing the severity of disease, and deterring the transmission of infection. However, COVID-19 vaccination faces many challenges, such as the decline in vaccine-induced immunity over time, and the decrease in potency against some SARS-CoV-2 variants including the recently emerged Omicron variant, resulting in breakthrough infections. The challenges that COVID-19 vaccination is facing highlight the importance of the discovery of antivirals to serve as another means to tackle the pandemic. To date, neutralizing antibodies that block viral entry by targeting the viral spike protein make up the largest class of antivirals that has received US FDA emergency use authorization (EUA) for COVID-19 treatment. In addition to the spike protein, other key targets for the discovery of direct-acting antivirals include viral enzymes that are essential for SARS-CoV-2 replication, such as RNA-dependent RNA polymerase and proteases, as judged by US FDA approval for remdesivir, and EUA for Paxlovid (nirmatrelvir + ritonavir) for treating COVID-19 infections. This review presents an overview of the current status and future direction of antiviral drug discovery for treating SARS-CoV-2 infections, covering important antiviral targets such as the viral spike protein, non-structural protein (nsp) 3 papain-like protease, nsp5 main protease, and the nsp12/nsp7/nsp8 RNA-dependent RNA polymerase complex.


Subject(s)
Antiviral Agents , COVID-19 , Drug Discovery , Antiviral Agents/pharmacology , COVID-19/drug therapy , COVID-19 Vaccines , Coronavirus 3C Proteases/antagonists & inhibitors , Humans , RNA-Dependent RNA Polymerase/antagonists & inhibitors , SARS-CoV-2 , Spike Glycoprotein, Coronavirus/antagonists & inhibitors , Viral Proteins/metabolism
14.
Bioorg Med Chem Lett ; 58: 128526, 2022 02 15.
Article in English | MEDLINE | ID: covidwho-1814173

ABSTRACT

The COVID-19 pandemic has drastically impacted global economies and public health. Although vaccine development has been successful, it was not sufficient against more infectious mutant strains including the Delta variant indicating a need for alternative treatment strategies such as small molecular compound development. In this work, a series of SARS-CoV-2 main protease (Mpro) inhibitors were designed and tested based on the active compound from high-throughput diverse compound library screens. The most efficacious compound (16b-3) displayed potent SARS-CoV-2 Mpro inhibition with an IC50 value of 116 nM and selectivity against SARS-CoV-2 Mpro when compared to PLpro and RdRp. This new class of compounds could be used as potential leads for further optimization in anti COVID-19 drug discovery.


Subject(s)
Antiviral Agents/pharmacology , Coronavirus 3C Proteases/antagonists & inhibitors , Drug Discovery , Protease Inhibitors/pharmacology , SARS-CoV-2/drug effects , Thiazoles/pharmacology , Antiviral Agents/chemical synthesis , Antiviral Agents/chemistry , COVID-19/drug therapy , Coronavirus 3C Proteases/metabolism , Humans , Microbial Sensitivity Tests , Molecular Structure , Protease Inhibitors/chemical synthesis , Protease Inhibitors/chemistry , SARS-CoV-2/enzymology , Thiazoles/chemical synthesis , Thiazoles/chemistry
15.
Drug Des Devel Ther ; 16: 1067-1082, 2022.
Article in English | MEDLINE | ID: covidwho-1808738

ABSTRACT

Coronavirus disease 2019 (COVID-19) caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) currently poses a threat to human health. 3C-like proteinase (3CLpro) plays an important role in the viral life cycle. Hence, it is considered an attractive antiviral target protein. Whole-genome sequencing showed that the sequence homology between SARS-CoV-2 3CLpro and SARS-CoV 3CLpro is 96.08%, with high similarity in the substrate-binding region. Thus, assessing peptidomimetic inhibitors of SARS-CoV 3CLpro could accelerate the development of peptidomimetic inhibitors for SARS-CoV-2 3CLpro. Accordingly, we herein discuss progress on SARS-CoV-2 3CLpro peptidomimetic inhibitors. Inflammation plays a major role in the pathophysiological process of COVID-19. Small-molecule compounds targeting 3CLpro with both antiviral and anti-inflammatory effects are also briefly discussed in this paper.


Subject(s)
Antiviral Agents , COVID-19 , Coronavirus 3C Proteases , Peptidomimetics , Protease Inhibitors , Anti-Inflammatory Agents/pharmacology , Antiviral Agents/pharmacology , COVID-19/drug therapy , Coronavirus 3C Proteases/antagonists & inhibitors , Humans , Peptidomimetics/pharmacology , Protease Inhibitors/chemistry , Protease Inhibitors/pharmacology , SARS-CoV-2
16.
mBio ; 13(3): e0078422, 2022 06 28.
Article in English | MEDLINE | ID: covidwho-1807327

ABSTRACT

The main protease, Mpro, of SARS-CoV-2 is required to cleave the viral polyprotein into precise functional units for virus replication and pathogenesis. Here, we report quantitative reporters for Mpro function in living cells in which protease inhibition by genetic or chemical methods results in robust signal readouts by fluorescence (enhanced green fluorescent protein [eGFP]) or bioluminescence (firefly luciferase). These gain-of-signal systems are scalable to high-throughput platforms for quantitative discrimination between Mpro mutants and/or inhibitor potencies as evidenced by validation of several reported inhibitors. Additional utility is shown by single Mpro amino acid variants and structural information combining to demonstrate that both inhibitor conformational dynamics and amino acid differences are able to influence inhibitor potency. We further show that a recent variant of concern (Omicron) has an unchanged response to a clinically approved drug, nirmatrelvir, whereas proteases from divergent coronavirus species show differential susceptibility. Together, we demonstrate that these gain-of-signal systems serve as robust, facile, and scalable assays for live cell quantification of Mpro inhibition, which will help expedite the development of next-generation antivirals and enable the rapid testing of emerging variants. IMPORTANCE The main protease, Mpro, of SARS-CoV-2 is an essential viral protein required for the earliest steps of infection. It is therefore an attractive target for antiviral drug development. Here, we report the development and implementation of two complementary cell-based systems for quantification of Mpro inhibition by genetic or chemical approaches. The first is fluorescence based (eGFP), and the second is luminescence based (firefly luciferase). Importantly, both systems rely upon gain-of-signal readouts such that stronger inhibitors yield higher fluorescent or luminescent signal. The high versatility and utility of these systems are demonstrated by characterizing Mpro mutants and natural variants, including Omicron, as well as a panel of existing inhibitors. These systems rapidly, safely, and sensitively identify Mpro variants with altered susceptibilities to inhibition, triage-nonspecific, or off-target molecules and validate bona fide inhibitors, with the most potent thus far being the first-in-class drug nirmatrelvir.


Subject(s)
Antiviral Agents , Coronavirus 3C Proteases , Protease Inhibitors , SARS-CoV-2 , Amino Acids , Antiviral Agents/pharmacology , Coronavirus 3C Proteases/antagonists & inhibitors , Luciferases, Firefly , Protease Inhibitors/pharmacology , SARS-CoV-2/drug effects , SARS-CoV-2/genetics
17.
Biochem Biophys Res Commun ; 604: 76-82, 2022 05 14.
Article in English | MEDLINE | ID: covidwho-1797136

ABSTRACT

The coronavirus disease 2019 (COVID-19) pandemic, caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), has resulted in millions of deaths and seriously threatened public health and safety. Despite COVID-19 vaccines being readily popularized worldwide, targeted therapeutic agents for the treatment of this disease remain very limited. Here, we studied the inhibitory activity of the scutellarein and its methylated derivatives against SARS-CoV-2 main protease (Mpro) by the fluorescence resonance energy transfer (FRET) assay. Among all the methylated derivatives we studied, 4'-O-methylscutellarein exhibited the most promising enzyme inhibitory activity in vitro, with the half-maximal inhibitory concentration value (IC50) of 0.40 ± 0.03 µM. Additionally, the mechanism of action of the hits was further characterized through enzyme kinetic studies and molecular docking. Overall, our results implied that 4'-O-methylscutellarein could be a primary lead compound with clinical potential for the development of inhibitors against the SARS-CoV-2 Mpro.


Subject(s)
Alkaloids , Coronavirus 3C Proteases , Indoles , SARS-CoV-2 , Viral Protease Inhibitors , Alkaloids/pharmacology , Coronavirus 3C Proteases/antagonists & inhibitors , Humans , Indoles/pharmacology , Kinetics , Molecular Docking Simulation , SARS-CoV-2/drug effects , SARS-CoV-2/enzymology , Viral Protease Inhibitors/pharmacology
18.
Nat Commun ; 13(1): 1891, 2022 04 07.
Article in English | MEDLINE | ID: covidwho-1783979

ABSTRACT

The SARS-CoV-2 3CL protease is a critical drug target for small molecule COVID-19 therapy, given its likely druggability and essentiality in the viral maturation and replication cycle. Based on the conservation of 3CL protease substrate binding pockets across coronaviruses and using screening, we identified four structurally distinct lead compounds that inhibit SARS-CoV-2 3CL protease. After evaluation of their binding specificity, cellular antiviral potency, metabolic stability, and water solubility, we prioritized the GC376 scaffold as being optimal for optimization. We identified multiple drug-like compounds with <10 nM potency for inhibiting SARS-CoV-2 3CL and the ability to block SARS-CoV-2 replication in human cells, obtained co-crystal structures of the 3CL protease in complex with these compounds, and determined that they have pan-coronavirus activity. We selected one compound, termed coronastat, as an optimized lead and characterized it in pharmacokinetic and safety studies in vivo. Coronastat represents a new candidate for a small molecule protease inhibitor for the treatment of SARS-CoV-2 infection for eliminating pandemics involving coronaviruses.


Subject(s)
Antiviral Agents , COVID-19 , Coronavirus 3C Proteases , Protease Inhibitors , Antiviral Agents/chemistry , Antiviral Agents/therapeutic use , COVID-19/drug therapy , Coronavirus 3C Proteases/antagonists & inhibitors , Humans , Molecular Docking Simulation , Pandemics , Protease Inhibitors/chemistry , Protease Inhibitors/pharmacology , Protease Inhibitors/therapeutic use , SARS-CoV-2
19.
Virology ; 571: 21-33, 2022 06.
Article in English | MEDLINE | ID: covidwho-1783830

ABSTRACT

Since December 2019, the deadly novel severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) has caused the current COVID-19 pandemic. To date, vaccines are available in the developed countries to prevent the infection of this virus; however, medicines are necessary to help control COVID-19. Human coronavirus 229E (HCoV-229E) causes the common cold. The main protease (Mpro) is an essential enzyme required for the multiplication of these two viruses in the host cells, and thus is an appropriate candidate to screen potential medicinal compounds. Flavonols and dihydroflavonols are two groups of plant flavonoids. In this study, we report docking simulation with two Mpro enzymes and five flavonols and three dihydroflavonols, in vitro inhibition of the SARS-CoV-2 Mpro, and in vitro inhibition of the HCoV 229E replication. The docking simulation results predicted that (+)-dihydrokaempferol, (+)- dihydroquercetin, (+)-dihydromyricetin, kaempferol, quercetin, myricentin, isoquercitrin, and rutin could bind to at least two subsites (S1, S1', S2, and S4) in the binding pocket and inhibit the activity of SARS-CoV-2 Mpro. Their affinity scores ranged from -8.8 to -7.4 (kcal/mol). Likewise, these compounds were predicted to bind and inhibit the HCoV-229E Mpro activity with affinity scores ranging from -7.1 to -7.8 (kcal/mol). In vitro inhibition assays showed that seven available compounds effectively inhibited the SARS-CoV-2 Mpro activity and their IC50 values ranged from 0.125 to 12.9 µM. Five compounds inhibited the replication of HCoV-229E in Huh-7 cells. These findings indicate that these antioxidative flavonols and dihydroflavonols are promising candidates for curbing the two viruses.


Subject(s)
Coronavirus 229E, Human , Coronavirus 3C Proteases , Flavonols , SARS-CoV-2 , COVID-19 , Coronavirus 229E, Human/drug effects , Coronavirus 229E, Human/physiology , Coronavirus 3C Proteases/antagonists & inhibitors , Flavonols/pharmacology , Humans , Molecular Docking Simulation , SARS-CoV-2/drug effects , Virus Replication/drug effects
20.
Molecules ; 27(7)2022 Mar 31.
Article in English | MEDLINE | ID: covidwho-1776288

ABSTRACT

A new dicoumarin, jusan coumarin, (1), has been isolated from Artemisia glauca aerial parts. The chemical structure of jusan coumarin was estimated, by 1D, 2D NMR as well as HR-Ms spectroscopic methods, to be 7-hydroxy-6-methoxy-3-[(2-oxo-2H-chromen-6-yl)oxy]-2H-chromen-2-one. As the first time to be introduced in nature, its potential against SARS-CoV-2 has been estimated using various in silico methods. Molecular similarity and fingerprints experiments have been utilized for 1 against nine co-crystallized ligands of COVID-19 vital proteins. The results declared a great similarity between Jusan Coumarin and X77, the ligand of COVID-19 main protease (PDB ID: 6W63), Mpro. To authenticate the obtained outputs, a DFT experiment was achieved to confirm the similarity of X77 and 1. Consequently, 1 was docked against Mpro. The results clarified that 1 bonded in a correct way inside Mpro active site, with a binding energy of -18.45 kcal/mol. Furthermore, the ADMET and toxicity profiles of 1 were evaluated and showed the safety of 1 and its likeness to be a drug. Finally, to confirm the binding and understand the thermodynamic characters between 1 and Mpro, several molecular dynamics (MD) simulations studies have been administered. Additionally, the known coumarin derivative, 7-isopentenyloxycoumarin (2), has been isolated as well as ß-sitosterol (3).


Subject(s)
Artemisia , Coronavirus 3C Proteases , Coumarins , Protease Inhibitors , SARS-CoV-2 , Artemisia/chemistry , Coronavirus 3C Proteases/antagonists & inhibitors , Coumarins/chemistry , Coumarins/pharmacology , Dicumarol/chemistry , Dicumarol/pharmacology , Ligands , Molecular Docking Simulation , Molecular Dynamics Simulation , Protease Inhibitors/chemistry , Protease Inhibitors/pharmacology , SARS-CoV-2/drug effects , SARS-CoV-2/enzymology
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