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1.
Science ; 374(6575): 1586-1593, 2021 Dec 24.
Article in English | MEDLINE | ID: covidwho-1666355

ABSTRACT

The worldwide outbreak of COVID-19 caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) has become a global pandemic. Alongside vaccines, antiviral therapeutics are an important part of the healthcare response to countering the ongoing threat presented by COVID-19. Here, we report the discovery and characterization of PF-07321332, an orally bioavailable SARS-CoV-2 main protease inhibitor with in vitro pan-human coronavirus antiviral activity and excellent off-target selectivity and in vivo safety profiles. PF-07321332 has demonstrated oral activity in a mouse-adapted SARS-CoV-2 model and has achieved oral plasma concentrations exceeding the in vitro antiviral cell potency in a phase 1 clinical trial in healthy human participants.


Subject(s)
COVID-19/drug therapy , Lactams/pharmacology , Lactams/therapeutic use , Leucine/pharmacology , Leucine/therapeutic use , Nitriles/pharmacology , Nitriles/therapeutic use , Proline/pharmacology , Proline/therapeutic use , SARS-CoV-2/drug effects , Viral Protease Inhibitors/pharmacology , Viral Protease Inhibitors/therapeutic use , Administration, Oral , Animals , COVID-19/virology , Clinical Trials, Phase I as Topic , Coronavirus/drug effects , Disease Models, Animal , Drug Therapy, Combination , Humans , Lactams/administration & dosage , Lactams/pharmacokinetics , Leucine/administration & dosage , Leucine/pharmacokinetics , Mice , Mice, Inbred BALB C , Microbial Sensitivity Tests , Nitriles/administration & dosage , Nitriles/pharmacokinetics , Proline/administration & dosage , Proline/pharmacokinetics , Randomized Controlled Trials as Topic , Ritonavir/administration & dosage , Ritonavir/therapeutic use , SARS-CoV-2/physiology , Viral Protease Inhibitors/administration & dosage , Viral Protease Inhibitors/pharmacokinetics , Virus Replication/drug effects
2.
Nature ; 601(7894): 496, 2022 01.
Article in English | MEDLINE | ID: covidwho-1641925

Subject(s)
Antiviral Agents/therapeutic use , COVID-19/drug therapy , COVID-19/virology , Drug Development/trends , Drug Resistance, Viral , Research Personnel , SARS-CoV-2/drug effects , Adenosine Monophosphate/administration & dosage , Adenosine Monophosphate/analogs & derivatives , Adenosine Monophosphate/pharmacology , Adenosine Monophosphate/therapeutic use , Administration, Oral , Alanine/administration & dosage , Alanine/analogs & derivatives , Alanine/pharmacology , Alanine/therapeutic use , Antiviral Agents/administration & dosage , Antiviral Agents/pharmacology , Antiviral Agents/supply & distribution , COVID-19/mortality , COVID-19/prevention & control , COVID-19 Vaccines/supply & distribution , Cytidine/administration & dosage , Cytidine/analogs & derivatives , Cytidine/pharmacology , Cytidine/therapeutic use , Drug Approval , Drug Combinations , Drug Resistance, Viral/drug effects , Drug Resistance, Viral/genetics , Drug Therapy, Combination , Hospitalization/statistics & numerical data , Humans , Hydroxylamines/administration & dosage , Hydroxylamines/pharmacology , Hydroxylamines/therapeutic use , Lactams/administration & dosage , Lactams/pharmacology , Lactams/therapeutic use , Leucine/administration & dosage , Leucine/pharmacology , Leucine/therapeutic use , Medication Adherence , Molecular Targeted Therapy , Mutagenesis , Nitriles/administration & dosage , Nitriles/pharmacology , Nitriles/therapeutic use , Proline/administration & dosage , Proline/pharmacology , Proline/therapeutic use , Public-Private Sector Partnerships/economics , Ritonavir/administration & dosage , Ritonavir/pharmacology , Ritonavir/therapeutic use , SARS-CoV-2/enzymology , SARS-CoV-2/genetics
5.
J Phys Chem Lett ; 12(20): 4814-4822, 2021 May 27.
Article in English | MEDLINE | ID: covidwho-1387121

ABSTRACT

Angiotensin converting enzyme 2 (ACE2) plays a key role in renin-angiotensin system regulation and amino acid homeostasis. Human ACE2 acts as the receptor for severe acute respiratory syndrome coronaviruses SARS-CoV and SARS-CoV-2. ACE2 is also widely expressed in epithelial cells of the lungs, heart, kidney, and pancreas. It is considered an important drug target for treating SARS-CoV-2 as well as pulmonary diseases, heart failure, hypertension, renal diseases, and diabetes. Despite the critical importance, the mechanism of ligand binding to the human ACE2 receptor remains unknown. Here, we have addressed this challenge through all-atom simulations using a novel ligand Gaussian accelerated molecular dynamics (LiGaMD) method. Microsecond time scale LiGaMD simulations have unprecedentedly captured multiple times of spontaneous binding and unbinding of a potent inhibitor MLN-4760 in the ACE2 receptor. With ligand far away in the unbound state, the ACE2 receptor samples distinct Open, Partially Open, Closed, and Fully Closed conformations. Upon ligand binding to the active site, conformational ensemble of the ACE2 receptor is biased toward the Closed state as observed in the X-ray experimental structure. The LiGaMD simulations thus suggest a conformational selection mechanism for ligand recognition by the highly flexible ACE2 receptor, which is expected to facilitate rational drug design targeting human ACE2 against coronaviruses and other related human diseases.


Subject(s)
Angiotensin-Converting Enzyme 2/antagonists & inhibitors , Antiviral Agents/chemistry , COVID-19/drug therapy , Imidazoles/chemistry , Leucine/analogs & derivatives , Protease Inhibitors/chemistry , SARS-CoV-2/drug effects , Antiviral Agents/pharmacology , COVID-19/metabolism , Catalytic Domain , Drug Design , Humans , Imidazoles/pharmacology , Leucine/chemistry , Leucine/pharmacology , Ligands , Molecular Dynamics Simulation , Protease Inhibitors/pharmacology , Protein Binding , Protein Conformation , SARS-CoV-2/metabolism
6.
Int J Mol Sci ; 22(17)2021 Aug 24.
Article in English | MEDLINE | ID: covidwho-1374423

ABSTRACT

The novel coronavirus disease, caused by severe acute respiratory coronavirus 2 (SARS-CoV-2), rapidly spreading around the world, poses a major threat to the global public health. Herein, we demonstrated the binding mechanism of PF-07321332, α-ketoamide, lopinavir, and ritonavir to the coronavirus 3-chymotrypsin-like-protease (3CLpro) by means of docking and molecular dynamic (MD) simulations. The analysis of MD trajectories of 3CLpro with PF-07321332, α-ketoamide, lopinavir, and ritonavir revealed that 3CLpro-PF-07321332 and 3CLpro-α-ketoamide complexes remained stable compared with 3CLpro-ritonavir and 3CLpro-lopinavir. Investigating the dynamic behavior of ligand-protein interaction, ligands PF-07321332 and α-ketoamide showed stronger bonding via making interactions with catalytic dyad residues His41-Cys145 of 3CLpro. Lopinavir and ritonavir were unable to disrupt the catalytic dyad, as illustrated by increased bond length during the MD simulation. To decipher the ligand binding mode and affinity, ligand interactions with SARS-CoV-2 proteases and binding energy were calculated. The binding energy of the bespoke antiviral PF-07321332 clinical candidate was two times higher than that of α-ketoamide and three times than that of lopinavir and ritonavir. Our study elucidated in detail the binding mechanism of the potent PF-07321332 to 3CLpro along with the low potency of lopinavir and ritonavir due to weak binding affinity demonstrated by the binding energy data. This study will be helpful for the development and optimization of more specific compounds to combat coronavirus disease.


Subject(s)
Antiviral Agents/pharmacology , COVID-19/drug therapy , Coronavirus 3C Proteases/antagonists & inhibitors , Coronavirus Protease Inhibitors/pharmacology , Lactams/pharmacology , Leucine/pharmacology , Nitriles/pharmacology , Proline/pharmacology , Antiviral Agents/therapeutic use , Catalytic Domain/drug effects , Coronavirus 3C Proteases/metabolism , Coronavirus Protease Inhibitors/therapeutic use , Humans , Lactams/therapeutic use , Leucine/therapeutic use , Lopinavir/pharmacology , Molecular Docking Simulation , Molecular Dynamics Simulation , Nitriles/therapeutic use , Proline/therapeutic use , Ritonavir/pharmacology , SARS-CoV-2/drug effects , SARS-CoV-2/enzymology
7.
Biomed Res Int ; 2021: 6614000, 2021.
Article in English | MEDLINE | ID: covidwho-1327769

ABSTRACT

Chloroquine (CQ) and hydroxychloroquine (HCQ) have shown the ability to inhibit in vitro viral replications of coronaviridae viruses such as SARS-CoV and SARS-CoV-2. However, clinical trial outcomes have been disparate, suggesting that CQ and HCQ antiviral mechanisms are not fully understood. Based on three-dimensional structural similarities between HCQ and the known ACE2 specific inhibitor MLN-4760, we compared their modulation on ACE2 activity. Here we describe, for the first time, in a cell-free in vitro system that HCQ directly and dose-dependently inhibits the activity of recombinant human ACE2, with a potency similar to the MLN-4760. Further analysis suggests that HCQ binds to a noncompetitive site other than the one occupied by MLN-4760. We also determined that the viral spike glycoprotein segment that comprises the RBD segment has no effect on ACE2 activity but unexpectedly was able to partially reverse the inhibition induced by HCQ but not that by MLN-4760. In summary, here we demonstrate the direct inhibitory action of HCQ over the activity of the enzyme ACE2. Then, by determining the activity of ACE2, we reveal that the interaction with the spike protein of SARS-CoV-2 leads to structural changes that at least partially displace the interaction of the said enzyme with HCQ. These results may help to explain why the effectiveness of HCQ in clinical trials has been so variable. Additionally, this knowledge could be used for to develop techniques for the detection of SARS-CoV-2.


Subject(s)
Angiotensin-Converting Enzyme 2 , Antiviral Agents , COVID-19/drug therapy , Hydroxychloroquine , Angiotensin-Converting Enzyme 2/antagonists & inhibitors , Angiotensin-Converting Enzyme 2/chemistry , Angiotensin-Converting Enzyme 2/metabolism , Antiviral Agents/chemistry , Antiviral Agents/metabolism , Antiviral Agents/pharmacology , Humans , Hydroxychloroquine/chemistry , Hydroxychloroquine/metabolism , Hydroxychloroquine/pharmacology , Imidazoles/chemistry , Imidazoles/metabolism , Imidazoles/pharmacology , Leucine/analogs & derivatives , Leucine/chemistry , Leucine/metabolism , Leucine/pharmacology , Recombinant Proteins/chemistry , Recombinant Proteins/metabolism , SARS-CoV-2 , Spike Glycoprotein, Coronavirus/chemistry , Spike Glycoprotein, Coronavirus/metabolism
8.
J Enzyme Inhib Med Chem ; 36(1): 1646-1650, 2021 Dec.
Article in English | MEDLINE | ID: covidwho-1320278

ABSTRACT

The chemical structure of PF-07321332, the first orally available Covid-19 clinical candidate, has recently been revealed by Pfizer. No information has been provided about the interaction pattern between PF-07321332 and its biomolecular counterpart, the SARS-CoV-2 main protease (Mpro). In the present work, we exploited Supervised Molecular Dynamics (SuMD) simulations to elucidate the key features that characterise the interaction between this drug candidate and the protease, emphasising similarities and differences with other structurally related inhibitors such as Boceprevir and PF-07304814. The structural insights provided by SuMD will hopefully be able to inspire the rational discovery of other potent and selective protease inhibitors.


Subject(s)
Antiviral Agents/chemistry , Lactams/chemistry , Leucine/chemistry , Molecular Dynamics Simulation , Nitriles/chemistry , Proline/chemistry , Protease Inhibitors/chemistry , Antiviral Agents/pharmacology , Humans , Lactams/pharmacology , Leucine/pharmacology , Ligands , Nitriles/pharmacology , Peptide Hydrolases/metabolism , Proline/pharmacology , Protease Inhibitors/pharmacology , SARS-CoV-2/drug effects , SARS-CoV-2/enzymology , Software
9.
Sci Rep ; 11(1): 9609, 2021 05 05.
Article in English | MEDLINE | ID: covidwho-1217711

ABSTRACT

The COVID-19 pandemic caused by the Severe Acute Respiratory Syndrome Coronavirus-2 (SARS-CoV-2) is the defining global health emergency of this century. GC-376 is a Mpro inhibitor with antiviral activity against SARS-CoV-2 in vitro. Using the K18-hACE2 mouse model, the in vivo antiviral efficacy of GC-376 against SARS-CoV-2 was evaluated. GC-376 treatment was not toxic in K18-hACE2 mice. Overall outcome of clinical symptoms and survival upon SARS-CoV-2 challenge were not improved in mice treated with GC-376 compared to controls. The treatment with GC-376 slightly improved survival from 0 to 20% in mice challenged with a high virus dose at 105 TCID50/mouse. Most notably, GC-376 treatment led to milder tissue lesions, reduced viral loads, fewer presence of viral antigen, and reduced inflammation in comparison to vehicle-treated controls in mice challenged with a low virus dose at 103 TCID50/mouse. This was particularly the case in the brain where a 5-log reduction in viral titers was observed in GC-376 treated mice compared to vehicle controls. This study supports the notion that GC-376 represents a promising lead candidate for further development to treat SARS-CoV-2 infection and that the K18-hACE2 mouse model is suitable to study antiviral therapies against SARS-CoV-2.


Subject(s)
Angiotensin-Converting Enzyme 2/genetics , Antiviral Agents/pharmacology , COVID-19/drug therapy , Carbonates/pharmacology , Leucine/pharmacology , Sulfonic Acids/pharmacology , Animals , Brain/drug effects , Brain/pathology , COVID-19/pathology , COVID-19/virology , Chlorocebus aethiops , Disease Models, Animal , Female , Keratin-18/genetics , Lung/drug effects , Lung/pathology , Lung/virology , Mice, Transgenic , Vero Cells , Viral Load
10.
Molecules ; 26(6)2021 Mar 17.
Article in English | MEDLINE | ID: covidwho-1138745

ABSTRACT

The COVID-19 outbreak continues to spread worldwide at a rapid rate. Currently, the absence of any effective antiviral treatment is the major concern for the global population. The reports of the occurrence of various point mutations within the important therapeutic target protein of SARS-CoV-2 has elevated the problem. The SARS-CoV-2 main protease (Mpro) is a major therapeutic target for new antiviral designs. In this study, the efficacy of PF-00835231 was investigated (a Mpro inhibitor under clinical trials) against the Mpro and their reported mutants. Various in silico approaches were used to investigate and compare the efficacy of PF-00835231 and five drugs previously documented to inhibit the Mpro. Our study shows that PF-00835231 is not only effective against the wild type but demonstrates a high affinity against the studied mutants as well.


Subject(s)
Antiviral Agents/chemistry , Antiviral Agents/pharmacology , Coronavirus 3C Proteases/antagonists & inhibitors , Indoles/chemistry , Indoles/pharmacology , Leucine/chemistry , Leucine/pharmacology , Protease Inhibitors/chemistry , Protease Inhibitors/pharmacology , Pyrrolidinones/chemistry , Pyrrolidinones/pharmacology , Binding Sites , COVID-19/drug therapy , Computer Simulation , Coronavirus 3C Proteases/chemistry , Coronavirus 3C Proteases/genetics , Databases, Protein , Diarylquinolines/chemistry , Diarylquinolines/pharmacology , Dihydropyridines/chemistry , Dihydropyridines/pharmacology , Humans , Ligands , Molecular Docking Simulation , Molecular Dynamics Simulation , Nitrobenzenes/chemistry , Nitrobenzenes/pharmacology , Nitrophenols/chemistry , Nitrophenols/pharmacology , Organophosphorus Compounds/chemistry , Organophosphorus Compounds/pharmacology , Piperazines/chemistry , Piperazines/pharmacology , Proline/analogs & derivatives , Proline/chemistry , Proline/pharmacology , SARS-CoV-2/drug effects , SARS-CoV-2/genetics
11.
ACS Infect Dis ; 7(3): 586-597, 2021 03 12.
Article in English | MEDLINE | ID: covidwho-1108883

ABSTRACT

As the COVID-19 pandemic continues to unfold, the morbidity and mortality are increasing daily. Effective treatment for SARS-CoV-2 is urgently needed. We recently discovered four SARS-CoV-2 main protease (Mpro) inhibitors including boceprevir, calpain inhibitors II and XII, and GC-376 with potent antiviral activity against infectious SARS-CoV-2 in cell culture. In this study, we further characterized the mechanism of action of these four compounds using the SARS-CoV-2 pseudovirus neutralization assay. It was found that GC-376 and calpain inhibitors II and XII have a dual mechanism of action by inhibiting both viral Mpro and host cathepsin L in Vero cells. To rule out the cell-type dependent effect, the antiviral activity of these four compounds against SARS-CoV-2 was also confirmed in type 2 transmembrane serine protease-expressing Caco-2 cells using the viral yield reduction assay. In addition, we found that these four compounds have broad-spectrum antiviral activity in inhibiting not only SARS-CoV-2 but also SARS-CoV, and MERS-CoV, as well as human coronaviruses (CoVs) 229E, OC43, and NL63. The mechanism of action is through targeting the viral Mpro, which was supported by the thermal shift-binding assay and enzymatic fluorescence resonance energy transfer assay. We further showed that these four compounds have additive antiviral effect when combined with remdesivir. Altogether, these results suggest that boceprevir, calpain inhibitors II and XII, and GC-376 might be promising starting points for further development against existing human coronaviruses as well as future emerging CoVs.


Subject(s)
Antiviral Agents/pharmacology , Carbonates/pharmacology , Glycoproteins/pharmacology , Leucine/pharmacology , Oligopeptides/pharmacology , Proline/analogs & derivatives , SARS-CoV-2/drug effects , Sulfonic Acids/pharmacology , Adenosine Monophosphate/analogs & derivatives , Adenosine Monophosphate/pharmacology , Alanine/analogs & derivatives , Alanine/pharmacology , Animals , COVID-19/drug therapy , Caco-2 Cells , Cathepsin L/antagonists & inhibitors , Cell Line , Chlorocebus aethiops , Coronavirus 229E, Human/drug effects , Coronavirus 3C Proteases/antagonists & inhibitors , Coronavirus NL63, Human/drug effects , Coronavirus OC43, Human/drug effects , Drug Combinations , HEK293 Cells , Humans , Middle East Respiratory Syndrome Coronavirus/drug effects , Proline/pharmacology , Serine Endopeptidases/metabolism , Vero Cells
12.
Cell ; 181(2): 271-280.e8, 2020 04 16.
Article in English | MEDLINE | ID: covidwho-4561

ABSTRACT

The recent emergence of the novel, pathogenic SARS-coronavirus 2 (SARS-CoV-2) in China and its rapid national and international spread pose a global health emergency. Cell entry of coronaviruses depends on binding of the viral spike (S) proteins to cellular receptors and on S protein priming by host cell proteases. Unravelling which cellular factors are used by SARS-CoV-2 for entry might provide insights into viral transmission and reveal therapeutic targets. Here, we demonstrate that SARS-CoV-2 uses the SARS-CoV receptor ACE2 for entry and the serine protease TMPRSS2 for S protein priming. A TMPRSS2 inhibitor approved for clinical use blocked entry and might constitute a treatment option. Finally, we show that the sera from convalescent SARS patients cross-neutralized SARS-2-S-driven entry. Our results reveal important commonalities between SARS-CoV-2 and SARS-CoV infection and identify a potential target for antiviral intervention.


Subject(s)
Betacoronavirus/metabolism , Coronavirus Infections/drug therapy , Peptidyl-Dipeptidase A/metabolism , Pneumonia, Viral/drug therapy , Protease Inhibitors/pharmacology , Serine Endopeptidases/metabolism , Spike Glycoprotein, Coronavirus/metabolism , Virus Internalization/drug effects , Ammonium Chloride/pharmacology , Angiotensin-Converting Enzyme 2 , Animals , Antibodies, Neutralizing/immunology , Antibodies, Viral/immunology , Betacoronavirus/chemistry , Betacoronavirus/genetics , COVID-19 , Cell Line , Coronavirus/chemistry , Coronavirus/genetics , Coronavirus/physiology , Coronavirus Infections/immunology , Coronavirus Infections/therapy , Drug Development , Esters , Gabexate/analogs & derivatives , Gabexate/pharmacology , Guanidines , Humans , Immunization, Passive , Leucine/analogs & derivatives , Leucine/pharmacology , Pandemics , Peptidyl-Dipeptidase A/chemistry , Receptors, Virus/chemistry , Receptors, Virus/metabolism , SARS Virus/physiology , SARS-CoV-2 , Spike Glycoprotein, Coronavirus/chemistry , Spike Glycoprotein, Coronavirus/genetics , Vesiculovirus/genetics
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