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1.
Sci Rep ; 12(1): 5320, 2022 03 29.
Article in English | MEDLINE | ID: covidwho-1768843

ABSTRACT

The ongoing pandemic caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) requires treatments with rapid clinical translatability. Here we develop a multi-target and multi-ligand virtual screening method to identify FDA-approved drugs with potential activity against SARS-CoV-2 at traditional and understudied viral targets. 1,268 FDA-approved small molecule drugs were docked to 47 putative binding sites across 23 SARS-CoV-2 proteins. We compared drugs between binding sites and filtered out compounds that had no reported activity in an in vitro screen against SARS-CoV-2 infection of human liver (Huh-7) cells. This identified 17 "high-confidence", and 97 "medium-confidence" drug-site pairs. The "high-confidence" group was subjected to molecular dynamics simulations to yield six compounds with stable binding poses at their optimal target proteins. Three drugs-amprenavir, levomefolic acid, and calcipotriol-were predicted to bind to 3 different sites on the spike protein, domperidone to the Mac1 domain of the non-structural protein (Nsp) 3, avanafil to Nsp15, and nintedanib to the nucleocapsid protein involved in packaging the viral RNA. Our "two-way" virtual docking screen also provides a framework to prioritize drugs for testing in future emergencies requiring rapidly available clinical drugs and/or treating diseases where a moderate number of targets are known.


Subject(s)
COVID-19 , Coronavirus Papain-Like Proteases , Nucleocapsid Proteins , SARS-CoV-2 , Spike Glycoprotein, Coronavirus , Binding Sites , COVID-19/drug therapy , Coronavirus Papain-Like Proteases/antagonists & inhibitors , Humans , Nucleocapsid Proteins/antagonists & inhibitors , RNA, Viral , SARS-CoV-2/drug effects , Spike Glycoprotein, Coronavirus/antagonists & inhibitors
2.
Commun Biol ; 5(1): 169, 2022 02 25.
Article in English | MEDLINE | ID: covidwho-1713217

ABSTRACT

SARS-CoV-2 proteases Mpro and PLpro are promising targets for antiviral drug development. In this study, we present an antiviral screening strategy involving a novel in-cell protease assay, antiviral and biochemical activity assessments, as well as structural determinations for rapid identification of protease inhibitors with low cytotoxicity. We identified eight compounds with anti-SARS-CoV-2 activity from a library of 64 repurposed drugs and modeled at protease active sites by in silico docking. We demonstrate that Sitagliptin and Daclatasvir inhibit PLpro, and MG-101, Lycorine HCl, and Nelfinavir mesylate inhibit Mpro of SARS-CoV-2. The X-ray crystal structure of Mpro in complex with MG-101 shows a covalent bond formation between the inhibitor and the active site Cys145 residue indicating its mechanism of inhibition is by blocking the substrate binding at the active site. Thus, we provide methods for rapid and effective screening and development of inhibitors for blocking virus polyprotein processing as SARS-CoV-2 antivirals. Additionally, we show that the combined inhibition of Mpro and PLpro is more effective in inhibiting SARS-CoV-2 and the delta variant.


Subject(s)
Coronavirus 3C Proteases/antagonists & inhibitors , Coronavirus Papain-Like Proteases/antagonists & inhibitors , Drug Evaluation, Preclinical/methods , SARS-CoV-2/enzymology , Viral Protease Inhibitors/analysis , COVID-19/drug therapy , Drug Repositioning , HEK293 Cells , Humans , Molecular Docking Simulation , Molecular Targeted Therapy
3.
Sci Rep ; 12(1): 2145, 2022 02 09.
Article in English | MEDLINE | ID: covidwho-1692555

ABSTRACT

The most common host entry point of human adapted coronaviruses (CoV) including SARS-CoV-2 is through the initial colonization in the nostril and mouth region which is responsible for spread of the infection. Most recent studies suggest that the commercially available oral and nasal rinse products are effective in inhibiting the viral replication. However, the anti-viral mechanism of the active ingredients present in the oral rinses have not been studied. In the present study, we have assessed in vitro enzymatic inhibitory activity of active ingredients in the oral mouth rinse products: aloin A and B, chlorhexidine, eucalyptol, hexetidine, menthol, triclosan, methyl salicylate, sodium fluoride and povidone, against two important proteases of SARS-CoV-2 PLpro and 3CLpro. Our results indicate only aloin A and B effectively inhibited proteolytic activity of PLpro with an IC50 of 13.16 and 16.08 µM. Interestingly, neither of the aloin isoforms inhibited 3CLpro enzymatic activity. Computational structural modelling of aloin A and B interaction with PLpro revealed that, both aloin isoforms form hydrogen bond with Tyr268 of PLpro, which is critical for their proteolytic activity. Furthermore, 100 ns molecular dynamics (MD) simulation studies predicted that both aloin isoforms have strong interaction with Glu167, which is required for PLpro deubiquitination activity. Our results from the in vitro deubiquitinase inhibition assay show that aloin A and B isomers exhibit deubiquitination inhibitory activity with an IC50 value of 15.68 and 17.51 µM, respectively. In conclusion, the isoforms of aloin inhibit both proteolytic and the deubiquitinating activity of SARS-CoV-2 PLpro, suggesting potential in inhibiting the replication of SARS-CoV-2 virus.


Subject(s)
Coronavirus Papain-Like Proteases/metabolism , Emodin/analogs & derivatives , SARS-CoV-2/enzymology , Animals , Binding Sites , COVID-19/pathology , COVID-19/virology , Cell Survival/drug effects , Chlorocebus aethiops , Coronavirus 3C Proteases/antagonists & inhibitors , Coronavirus 3C Proteases/metabolism , Coronavirus Papain-Like Proteases/antagonists & inhibitors , Emodin/chemistry , Emodin/metabolism , Emodin/pharmacology , Humans , Molecular Dynamics Simulation , Protein Isoforms/chemistry , Protein Isoforms/metabolism , Protein Isoforms/pharmacology , SARS-CoV-2/isolation & purification , Vero Cells
4.
Molecules ; 27(4)2022 Feb 11.
Article in English | MEDLINE | ID: covidwho-1686903

ABSTRACT

Two rare 2-phenoxychromone derivatives, 6-demethoxy-4`-O-capillarsine (1) and tenuflorin C (2), were isolated from the areal parts of Artemisia commutata and A. glauca, respectively, for the first time. Being rare in nature, the inhibition potentialities of 1 and 2 against SARS-CoV-2 was investigated using multistage in silico techniques. At first, molecular similarity and fingerprint studies were conducted for 1 and 2 against co-crystallized ligands of eight different COVID-19 enzymes. The carried-out studies indicated the similarity of 1 and 2 with TTT, the co-crystallized ligand of COVID-19 Papain-Like Protease (PLP), (PDB ID: 3E9S). Therefore, molecular docking studies of 1 and 2 against the PLP were carried out and revealed correct binding inside the active site exhibiting binding energies of -18.86 and -18.37 Kcal/mol, respectively. Further, in silico ADMET in addition to toxicity evaluation of 1 and 2 against seven models indicated the general safety and the likeness of 1 and 2 to be drugs. Lastly, to authenticate the binding and to investigate the thermodynamic characters, molecular dynamics (MD) simulation studies were conducted on 1 and PLP.


Subject(s)
Artemisia/chemistry , COVID-19/enzymology , Chromones/chemistry , Coronavirus Papain-Like Proteases , Protease Inhibitors/chemistry , SARS-CoV-2/enzymology , COVID-19/drug therapy , Coronavirus Papain-Like Proteases/antagonists & inhibitors , Coronavirus Papain-Like Proteases/chemistry , Humans
5.
Molecules ; 26(24)2021 Dec 09.
Article in English | MEDLINE | ID: covidwho-1572566

ABSTRACT

This study demonstrates the inhibitory effect of 42 pyrimidonic pharmaceuticals (PPs) on the 3-chymotrypsin-like protease of SARS-CoV-2 (3CLpro) through molecular docking, molecular dynamics simulations, and free binding energies by means of molecular mechanics-Poisson Boltzmann surface area (MM-PBSA) and molecular mechanics-generalized Born surface area (MM-GBSA). Of these tested PPs, 11 drugs approved by the US Food and Drug Administration showed an excellent binding affinity to the catalytic residues of 3CLpro of His41 and Cys145: uracil mustard, cytarabine, floxuridine, trifluridine, stavudine, lamivudine, zalcitabine, telbivudine, tipiracil, citicoline, and uridine triacetate. Their percentage of residues involved in binding at the active sites ranged from 56 to 100, and their binding affinities were in the range from -4.6 ± 0.14 to -7.0 ± 0.19 kcal/mol. The molecular dynamics as determined by a 200 ns simulation run of solvated docked complexes confirmed the stability of PP conformations that bound to the catalytic dyad and the active sites of 3CLpro. The free energy of binding also demonstrates the stability of the PP-3CLpro complexes. Citicoline and uridine triacetate showed free binding energies of -25.53 and -7.07 kcal/mol, respectively. Therefore, I recommend that they be repurposed for the fight against COVID-19, following proper experimental and clinical validation.


Subject(s)
COVID-19/drug therapy , Coronavirus 3C Proteases/antagonists & inhibitors , Coronavirus Papain-Like Proteases/antagonists & inhibitors , Drug Repositioning/methods , Protease Inhibitors/pharmacology , SARS-CoV-2/drug effects , Acetates/chemistry , Acetates/pharmacology , Antiviral Agents/chemistry , Antiviral Agents/pharmacology , Cytidine Diphosphate Choline/chemistry , Cytidine Diphosphate Choline/pharmacology , Drug Evaluation, Preclinical , Humans , Molecular Docking Simulation , Molecular Dynamics Simulation , Protease Inhibitors/chemistry , Uridine/analogs & derivatives , Uridine/chemistry , Uridine/pharmacology
6.
Angew Chem Int Ed Engl ; 61(9): e202113617, 2022 02 21.
Article in English | MEDLINE | ID: covidwho-1565164

ABSTRACT

The main protease (Mpro ) and papain-like protease (PLpro ) play critical roles in SARS-CoV-2 replication and are promising targets for antiviral inhibitors. The simultaneous visualization of Mpro and PLpro is extremely valuable for SARS-CoV-2 detection and rapid inhibitor screening. However, such a crucial investigation has remained challenging because of the lack of suitable probes. We have now developed a dual-color probe (3MBP5) for the simultaneous detection of Mpro and PLpro by fluorescence (or Förster) resonance energy transfer (FRET). This probe produces fluorescence from both the Cy3 and Cy5 fluorophores that are cleaved by Mpro and PLpro . 3MBP5-activatable specificity was demonstrated with recombinant proteins, inhibitors, plasmid-transfected HEK 293T cells, and SARS-CoV-2-infected TMPRSS2-Vero cells. Results from the dual-color probe first verified the simultaneous detection and intracellular distribution of SARS-CoV-2 Mpro and PLpro . This is a powerful tool for the simultaneous detection of different proteases with value for the rapid screening of inhibitors.


Subject(s)
Color , Coronavirus 3C Proteases/metabolism , Coronavirus Papain-Like Proteases/metabolism , Fluorescent Dyes/chemistry , Protease Inhibitors/pharmacology , SARS-CoV-2/enzymology , Coronavirus 3C Proteases/antagonists & inhibitors , Coronavirus Papain-Like Proteases/antagonists & inhibitors , Fluorescence Resonance Energy Transfer , HEK293 Cells , Humans
7.
Int J Mol Sci ; 22(22)2021 Nov 17.
Article in English | MEDLINE | ID: covidwho-1524024

ABSTRACT

The worldwide outbreak of COVID-19 was caused by a pathogenic virus called Severe Acute Respiratory Syndrome Coronavirus-2 (SARS-CoV-2). Therapies against SARS-CoV-2 target the virus or human cells or the immune system. However, therapies based on specific antibodies, such as vaccines and monoclonal antibodies, may become inefficient enough when the virus changes its antigenicity due to mutations. Polyphenols are the major class of bioactive compounds in nature, exerting diverse health effects based on their direct antioxidant activity and their effects in the modulation of intracellular signaling. There are currently numerous clinical trials investigating the effects of polyphenols in prophylaxis and the treatment of COVID-19, from symptomatic, via moderate and severe COVID-19 treatment, to anti-fibrotic treatment in discharged COVID-19 patients. Antiviral activities of polyphenols and their impact on immune system modulation could serve as a solid basis for developing polyphenol-based natural approaches for preventing and treating COVID-19.


Subject(s)
Antiviral Agents/therapeutic use , COVID-19/prevention & control , Polyphenols/therapeutic use , Antiviral Agents/chemistry , Antiviral Agents/metabolism , COVID-19/virology , Coronavirus 3C Proteases/antagonists & inhibitors , Coronavirus 3C Proteases/metabolism , Coronavirus Papain-Like Proteases/antagonists & inhibitors , Coronavirus Papain-Like Proteases/metabolism , Humans , Plants, Medicinal/chemistry , Plants, Medicinal/metabolism , Polyphenols/chemistry , Polyphenols/metabolism , SARS-CoV-2/isolation & purification , SARS-CoV-2/metabolism , Spike Glycoprotein, Coronavirus/antagonists & inhibitors , Spike Glycoprotein, Coronavirus/metabolism
8.
Chemistry ; 27(71): 17928-17940, 2021 Dec 20.
Article in English | MEDLINE | ID: covidwho-1487453

ABSTRACT

The global spread of the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) has called for an urgent need for dedicated antiviral therapeutics. Metal complexes are commonly underrepresented in compound libraries that are used for screening in drug discovery campaigns, however, there is growing evidence for their role in medicinal chemistry. Based on previous results, we have selected more than 100 structurally diverse metal complexes for profiling as inhibitors of two relevant SARS-CoV-2 replication mechanisms, namely the interaction of the spike (S) protein with the ACE2 receptor and the papain-like protease PLpro . In addition to many well-established types of mononuclear experimental metallodrugs, the pool of compounds tested was extended to approved metal-based therapeutics such as silver sulfadiazine and thiomersal, as well as polyoxometalates (POMs). Among the mononuclear metal complexes, only a small number of active inhibitors of the S/ACE2 interaction was identified, with titanocene dichloride as the only strong inhibitor. However, among the gold and silver containing complexes many turned out to be very potent inhibitors of PLpro activity. Highly promising activity against both targets was noted for many POMs. Selected complexes were evaluated in antiviral SARS-CoV-2 assays confirming activity for gold complexes with N-heterocyclic carbene (NHC) or dithiocarbamato ligands, a silver NHC complex, titanocene dichloride as well as a POM compound. These studies might provide starting points for the design of metal-based SARS-CoV-2 antiviral agents.


Subject(s)
Antiviral Agents/pharmacology , Coronavirus Papain-Like Proteases/antagonists & inhibitors , SARS-CoV-2 , Spike Glycoprotein, Coronavirus/antagonists & inhibitors , Angiotensin-Converting Enzyme 2 , SARS-CoV-2/drug effects
9.
J Med Chem ; 65(4): 2940-2955, 2022 02 24.
Article in English | MEDLINE | ID: covidwho-1475245

ABSTRACT

Antiviral agents that complement vaccination are urgently needed to end the COVID-19 pandemic. The SARS-CoV-2 papain-like protease (PLpro), one of only two essential cysteine proteases that regulate viral replication, also dysregulates host immune sensing by binding and deubiquitination of host protein substrates. PLpro is a promising therapeutic target, albeit challenging owing to featureless P1 and P2 sites recognizing glycine. To overcome this challenge, we leveraged the cooperativity of multiple shallow binding sites on the PLpro surface, yielding novel 2-phenylthiophenes with nanomolar inhibitory potency. New cocrystal structures confirmed that ligand binding induces new interactions with PLpro: by closing of the BL2 loop of PLpro forming a novel "BL2 groove" and by mimicking the binding interaction of ubiquitin with Glu167 of PLpro. Together, this binding cooperativity translates to the most potent PLpro inhibitors reported to date, with slow off-rates, improved binding affinities, and low micromolar antiviral potency in SARS-CoV-2-infected human cells.


Subject(s)
Antiviral Agents/pharmacology , COVID-19/drug therapy , Coronavirus Papain-Like Proteases/antagonists & inhibitors , Cysteine Proteinase Inhibitors/pharmacology , Antiviral Agents/chemical synthesis , Antiviral Agents/chemistry , Binding Sites/drug effects , COVID-19/metabolism , Coronavirus Papain-Like Proteases/isolation & purification , Coronavirus Papain-Like Proteases/metabolism , Crystallography, X-Ray , Cysteine Proteinase Inhibitors/chemical synthesis , Cysteine Proteinase Inhibitors/chemistry , Humans , Microbial Sensitivity Tests , Microsomes, Liver/chemistry , Microsomes, Liver/metabolism , Models, Molecular , Pandemics , Surface Plasmon Resonance , Tumor Cells, Cultured
10.
Int J Mol Sci ; 22(19)2021 Oct 02.
Article in English | MEDLINE | ID: covidwho-1444233

ABSTRACT

Considering the current dramatic and fatal situation due to the high spreading of SARS-CoV-2 infection, there is an urgent unmet medical need to identify novel and effective approaches for prevention and treatment of Coronavirus disease (COVID 19) by re-evaluating and repurposing of known drugs. For this, tomatidine and patchouli alcohol have been selected as potential drugs for combating the virus. The hit compounds were subsequently docked into the active site and molecular docking analyses revealed that both drugs can bind the active site of SARS-CoV-2 3CLpro, PLpro, NSP15, COX-2 and PLA2 targets with a number of important binding interactions. To further validate the interactions of promising compound tomatidine, Molecular dynamics study of 100 ns was carried out towards 3CLpro, NSP15 and COX-2. This indicated that the protein-ligand complex was stable throughout the simulation period, and minimal backbone fluctuations have ensued in the system. Post dynamic MM-GBSA analysis of molecular dynamics data showed promising mean binding free energy 47.4633 ± 9.28, 51.8064 ± 8.91 and 54.8918 ± 7.55 kcal/mol, respectively. Likewise, in silico ADMET studies of the selected ligands showed excellent pharmacokinetic properties with good absorption, bioavailability and devoid of toxicity. Therefore, patchouli alcohol and especially, tomatidine may provide prospect treatment options against SARS-CoV-2 infection by potentially inhibiting virus duplication though more research is guaranteed and secured.


Subject(s)
Coronavirus 3C Proteases/antagonists & inhibitors , Coronavirus Papain-Like Proteases/antagonists & inhibitors , Endoribonucleases/antagonists & inhibitors , SARS-CoV-2/enzymology , Sesquiterpenes/pharmacology , Tomatine/analogs & derivatives , Viral Nonstructural Proteins/antagonists & inhibitors , Antiviral Agents/pharmacology , COVID-19/drug therapy , COVID-19/virology , Coronavirus 3C Proteases/metabolism , Coronavirus Papain-Like Proteases/metabolism , Endoribonucleases/metabolism , Enzyme Inhibitors/pharmacology , Humans , Molecular Docking Simulation , Molecular Dynamics Simulation , SARS-CoV-2/drug effects , Tomatine/pharmacology , Viral Nonstructural Proteins/metabolism
11.
Molecules ; 26(17)2021 Aug 24.
Article in English | MEDLINE | ID: covidwho-1374471

ABSTRACT

The emergence of COVID-19 continues to pose severe threats to global public health. The pandemic has infected over 171 million people and claimed more than 3.5 million lives to date. We investigated the binding potential of antiviral cyanobacterial proteins including cyanovirin-N, scytovirin and phycocyanin with fundamental proteins involved in attachment and replication of SARS-CoV-2. Cyanovirin-N displayed the highest binding energy scores (-16.8 ± 0.02 kcal/mol, -12.3 ± 0.03 kcal/mol and -13.4 ± 0.02 kcal/mol, respectively) with the spike protein, the main protease (Mpro) and the papainlike protease (PLpro) of SARS-CoV-2. Cyanovirin-N was observed to interact with the crucial residues involved in the attachment of the human ACE2 receptor. Analysis of the binding affinities calculated employing the molecular mechanics-Poisson-Boltzmann surface area (MM-PBSA) approach revealed that all forms of energy, except the polar solvation energy, favourably contributed to the interactions of cyanovirin-N with the viral proteins. With particular emphasis on cyanovirin-N, the current work presents evidence for the potential inhibition of SARS-CoV-2 by cyanobacterial proteins, and offers the opportunity for in vitro and in vivo experiments to deploy the cyanobacterial proteins as valuable therapeutics against COVID-19.


Subject(s)
Antiviral Agents/pharmacology , Bacterial Proteins/pharmacology , COVID-19/drug therapy , Coronavirus Protease Inhibitors/pharmacology , Antiviral Agents/therapeutic use , Bacterial Proteins/therapeutic use , Bacterial Proteins/ultrastructure , COVID-19/virology , Coronavirus 3C Proteases/antagonists & inhibitors , Coronavirus 3C Proteases/metabolism , Coronavirus 3C Proteases/ultrastructure , Coronavirus Papain-Like Proteases/antagonists & inhibitors , Coronavirus Papain-Like Proteases/metabolism , Coronavirus Papain-Like Proteases/ultrastructure , Coronavirus Protease Inhibitors/therapeutic use , Coronavirus Protease Inhibitors/ultrastructure , Humans , Molecular Docking Simulation , Molecular Dynamics Simulation , Protein Binding , Protein Interaction Mapping , Spike Glycoprotein, Coronavirus/antagonists & inhibitors , Spike Glycoprotein, Coronavirus/metabolism , Spike Glycoprotein, Coronavirus/ultrastructure , X-Ray Diffraction
12.
Mol Biotechnol ; 64(1): 1-8, 2022 Jan.
Article in English | MEDLINE | ID: covidwho-1366408

ABSTRACT

Because of the essential roles of SARS-CoV-2 papain-like protease (PLpro) in the viral polyprotein processing and suppression of host immune responses, it is a crucial target for drug discovery against COVID-19. To develop robust biochemical methodologies for inhibitor screening against PLpro, extensive characterization of recombinant protein is important. Here we report cloning, expression, and purification of the recombinant SARS-CoV-2 PLpro, and explore various parameters affecting its stability and the catalytic activity. We also report the optimum conditions which should be used for high-throughput inhibitor screening using a fluorogenic tetrapeptide substrate.


Subject(s)
Coronavirus Papain-Like Proteases/chemistry , Coronavirus Papain-Like Proteases/metabolism , High-Throughput Screening Assays/methods , Antiviral Agents/pharmacology , Coronavirus Papain-Like Proteases/antagonists & inhibitors , Coronavirus Papain-Like Proteases/isolation & purification , Coumarins/chemistry , Coumarins/metabolism , Cysteine Proteinase Inhibitors/pharmacology , Dimethyl Sulfoxide/chemistry , Dynamic Light Scattering , Edetic Acid/chemistry , Enzyme Stability , Fluorometry/methods , Hydrogen-Ion Concentration , Osmolar Concentration , Peptides/chemistry , Peptides/metabolism , Temperature
13.
ChemMedChem ; 17(1): e202100455, 2022 01 05.
Article in English | MEDLINE | ID: covidwho-1366225

ABSTRACT

As the pathogen of COVID-19, SARS-CoV-2 encodes two essential cysteine proteases that process the pathogen's two large polypeptide products pp1a and pp1ab in the human cell host to form 15 functionally important, mature nonstructural proteins. One of the two enzymes is papain-like protease or PLPro . It possesses deubiquitination and deISGylation activities that suppress host innate immune responses toward SARS-CoV-2 infection. To repurpose drugs for PLPro , we experimentally screened libraries of 33 deubiquitinase and 37 cysteine protease inhibitors on their inhibition of PLPro . Our results showed that 15 deubiquitinase and 1 cysteine protease inhibitors exhibit strong inhibition of PLPro at 200 µM. More comprehensive characterizations revealed seven inhibitors GRL0617, SJB2-043, TCID, DUB-IN-1, DUB-IN-3, PR-619, and S130 with an IC50 value below 40 µM and four inhibitors GRL0617, SJB2-043, TCID, and PR-619 with an IC50 value below 10 µM. Among four inhibitors with an IC50 value below 10 µM, SJB2-043 is the most unique in that it does not fully inhibit PLPro but has a noteworthy IC50 value of 0.56 µM. SJB2-043 likely binds to an allosteric site of PLPro to convene its inhibition effect, which needs to be further investigated. As a pilot study, the current work indicates that COVID-19 drug repurposing by targeting PLPro holds promise, but in-depth analysis of repurposed drugs is necessary to avoid omitting critical allosteric inhibitors.


Subject(s)
Antiviral Agents/pharmacology , Coronavirus Papain-Like Proteases/antagonists & inhibitors , Cysteine Proteinase Inhibitors/pharmacology , Drug Repositioning , SARS-CoV-2/drug effects , Antiviral Agents/chemistry , Cysteine Proteinase Inhibitors/chemistry , Humans , Inhibitory Concentration 50 , Structure-Activity Relationship
14.
SAR QSAR Environ Res ; 32(9): 699-718, 2021 Sep.
Article in English | MEDLINE | ID: covidwho-1358390

ABSTRACT

SARS-CoV-2 PLpro was investigated as a therapeutic target for potent antiviral drugs due to its essential role in not only viral replication but also in regulating the inborn immune response. Several computational approaches, including homology modelling, molecular docking, and molecular dynamics (MD) studies, were employed to search for promising drugs in treating SARS-CoV-2. Eighty-one compounds, sub-structurally similar to the antiviral drug, were used as potential inhibitors of PLpro. From our results, three complexes containing the ligands with Pubchem IDs: 153012995, 12149203, and 123608715 showed lower binding energies than the control (Ritonavir), indicating that they may become promising inhibitors for PLpro. MD was performed in a water solvent to validate the stability of the three complexes. All complexes achieved stable structure during the simulation as no significant fluctuations were observed in the validation parameters. Moreover, the binding energy for each complex was estimated using the MM-GBSA method. Complex 1 was the most stable structure based on the lowest binding energy score and its structure remained in a similar cavity with the docket snapshot. Based on our studies, three ligands were assumed to be potential inhibitors. The ligand of complex 1 may become the most promising antiviral drug against SARS-CoV-2 targeting PLpro.


Subject(s)
Antiviral Agents/pharmacology , COVID-19/drug therapy , Coronavirus Papain-Like Proteases/antagonists & inhibitors , Coronavirus Papain-Like Proteases/chemistry , Molecular Docking Simulation , Molecular Dynamics Simulation , Protease Inhibitors/pharmacology , SARS-CoV-2/drug effects , Humans
15.
IEEE/ACM Trans Comput Biol Bioinform ; 18(4): 1262-1270, 2021.
Article in English | MEDLINE | ID: covidwho-1349900

ABSTRACT

SARS-CoV-2 encodes the Mac1 domain within the large nonstructural protein 3 (Nsp3), which has an ADP-ribosylhydrolase activity conserved in other coronaviruses. The enzymatic activity of Mac1 makes it an essential virulence factor for the pathogenicity of coronavirus (CoV). They have a regulatory role in counteracting host-mediated antiviral ADP-ribosylation, which is unique part of host response towards viral infections. Mac1 shows highly conserved residues in the binding pocket for the mono and poly ADP-ribose. Therefore, SARS-CoV-2 Mac1 enzyme is considered as an ideal drug target and inhibitors developed against them can possess a broad antiviral activity against CoV. ADP-ribose-1 phosphate bound closed form of Mac1 domain is considered for screening with large database of ZINC. XP docking and QPLD provides strong potential lead compounds, that perfectly fits inside the binding pocket. Quantum mechanical studies expose that, substrate and leads have similar electron donor ability in the head regions, that allocates tight binding inside the substrate-binding pocket. Molecular dynamics study confirms the substrate and new lead molecules presence of electron donor and acceptor makes the interactions tight inside the binding pocket. Overall binding phenomenon shows both substrate and lead molecules are well-adopt to bind with similar binding mode inside the closed form of Mac1.


Subject(s)
COVID-19/drug therapy , COVID-19/virology , Coronavirus Papain-Like Proteases/antagonists & inhibitors , Coronavirus Papain-Like Proteases/chemistry , SARS-CoV-2/drug effects , Adenosine Diphosphate Ribose/metabolism , Amino Acid Sequence , Antiviral Agents/pharmacology , Computational Biology , Coronavirus Papain-Like Proteases/genetics , High-Throughput Screening Assays/methods , High-Throughput Screening Assays/statistics & numerical data , Humans , Models, Molecular , Molecular Docking Simulation , Molecular Dynamics Simulation , Protein Domains , Quantum Theory , SARS-CoV-2/genetics , SARS-CoV-2/physiology , User-Computer Interface
16.
Int J Biol Macromol ; 188: 137-146, 2021 Oct 01.
Article in English | MEDLINE | ID: covidwho-1345340

ABSTRACT

COVID-19 is a disease caused by SARS-CoV-2, which has led to more than 4 million deaths worldwide. As a result, there is a worldwide effort to develop specific drugs for targeting COVID-19. Papain-like protease (PLpro) is an attractive drug target because it has multiple essential functions involved in processing viral proteins, including viral genome replication and removal of post-translational ubiquitination modifications. Here, we established two assays for screening PLpro inhibitors according to protease and anti-ISGylation activities, respectively. Application of the two screening techniques to the library of clinically approved drugs led to the discovery of tanshinone IIA sulfonate sodium and chloroxine with their IC50 values of lower than 10 µM. These two compounds were found to directly interact with PLpro and their molecular mechanisms of binding were illustrated by docking and molecular dynamics simulations. The results highlight the usefulness of the two developed screening techniques for locating PLpro inhibitors.


Subject(s)
Antiviral Agents/pharmacology , COVID-19/drug therapy , Coronavirus Papain-Like Proteases/antagonists & inhibitors , Coronavirus Protease Inhibitors/pharmacology , Drug Repositioning , SARS-CoV-2/enzymology , Antiviral Agents/chemistry , Binding Sites , Chloroquinolinols/chemistry , Chloroquinolinols/pharmacology , Coronavirus Papain-Like Proteases/genetics , Coronavirus Papain-Like Proteases/isolation & purification , Coronavirus Protease Inhibitors/chemistry , High-Throughput Screening Assays/methods , Humans , Molecular Docking Simulation , Molecular Dynamics Simulation , Phenanthrenes/chemistry , Phenanthrenes/pharmacology , SARS-CoV-2/drug effects
17.
Mol Inform ; 40(8): e2100028, 2021 08.
Article in English | MEDLINE | ID: covidwho-1345038

ABSTRACT

The COVID-19 pandemic caused by the SARS-CoV-2 has mobilized scientific attention in search of a treatment. The cysteine-proteases, main protease (Mpro) and papain-like protease (PLpro) are important targets for antiviral drugs. In this work, we simulate the interactions between the Mpro and PLpro with Ebselen, its metabolites and derivatives with the aim of finding molecules that can potentially inhibit these enzymes. The docking data demonstrate that there are two main interactions between the thiol (-SH) group of Cys (from the protease active sites) and the electrophilic centers of the organoselenium molecules, i. e. the interaction with the carbonyl group (O=C… SH) and the interaction with the Se moiety (Se… SH). Both interactions may lead to an adduct formation and enzyme inhibition. Density Functional Theory (DFT) calculations with Ebselen indicate that the energetics of the thiol nucleophilic attack is more favorable on Se than on the carbonyl group, which is in accordance with experimental data (Jin et al. Nature, 2020, 582, 289-293). Therefore, organoselenium molecules should be further explored as inhibitors of the SARS-CoV-2 proteases. Furthermore, we suggest that some metabolites of Ebselen (e. g. Ebselen diselenide and methylebselenoxide) and derivatives ethaselen and ebsulfur should be tested in vitro as inhibitors of virus replication and its proteases.


Subject(s)
Azoles/pharmacology , COVID-19/drug therapy , Coronavirus Papain-Like Proteases/metabolism , Organoselenium Compounds/pharmacology , Protease Inhibitors/pharmacology , SARS-CoV-2/drug effects , Viral Matrix Proteins/metabolism , Antiviral Agents/chemistry , Antiviral Agents/metabolism , Antiviral Agents/pharmacology , Azoles/chemistry , Azoles/metabolism , COVID-19/metabolism , Catalytic Domain/drug effects , Coronavirus Papain-Like Proteases/antagonists & inhibitors , Drug Discovery , Humans , Isoindoles , Molecular Docking Simulation , Organoselenium Compounds/chemistry , Organoselenium Compounds/metabolism , Protease Inhibitors/chemistry , Protease Inhibitors/metabolism , Viral Matrix Proteins/antagonists & inhibitors
18.
J Pharm Pharm Sci ; 24: 390-399, 2021.
Article in English | MEDLINE | ID: covidwho-1329250

ABSTRACT

PURPOSE: SARS-CoV-2 infection is associated with substantial mortality and high morbidity. This study tested the effect of angiotensin II type I receptor blocker, losartan, on SARS-CoV-2 replication and inhibition of the papain-like protease of the virus. METHODS: The dose-dependent inhibitory effect of losartan, in concentrations from 1µM to 100µM as determined by quantitative cell analysis combining fluorescence microscopy, image processing, and cellular measurements (Cellomics analysis) on SARS-CoV-2 replication was investigated in Vero E6 cells. The impact of losartan on deubiquitination and deISGylation of SARS-CoV-2 papain-like protease (PLpro) were also evaluated.  Results: Losartan reduced PLpro cleavage of tetraUbiquitin to diUbiquitin.  It was less effective in inhibiting PLpro's cleavage of ISG15-AMC than Ubiquitin-AMC.  To determine if losartan inhibited SARS-CoV-2 replication, losartan treatment of SARS-CoV-2 infected Vero E6 was examined. Losartan treatment one hour prior to SARS-CoV-2 infection reduced levels of SARS-CoV-2 nuclear protein, an indicator of virus replication, by 80% and treatment one-hour post-infection decreased viral replication by 70%. CONCLUSION: Losartan was not an effective inhibitor of deubiquitinase or deISGylase activity of the PLpro but affected the SARS-CoV-2 replication of Vero E6 cells in vitro.  As losartan has a favorable safety profile and is currently available it has features necessary for efficacious drug repurposing and treatment of COVID-19.


Subject(s)
Angiotensin II Type 1 Receptor Blockers/pharmacology , Antiviral Agents/pharmacology , Losartan/pharmacology , SARS-CoV-2/drug effects , Animals , COVID-19/drug therapy , Chlorocebus aethiops , Computational Biology , Coronavirus Papain-Like Proteases/antagonists & inhibitors , Coronavirus Papain-Like Proteases/metabolism , Deubiquitinating Enzymes/antagonists & inhibitors , Deubiquitinating Enzymes/metabolism , SARS-CoV-2/metabolism , SARS-CoV-2/physiology , Ubiquitin/metabolism , Vero Cells , Virus Replication/drug effects
19.
Proc Natl Acad Sci U S A ; 118(29)2021 07 20.
Article in English | MEDLINE | ID: covidwho-1294550

ABSTRACT

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection continues to be a serious global public health threat. The 3C-like protease (3CLpro) is a virus protease encoded by SARS-CoV-2, which is essential for virus replication. We have previously reported a series of small-molecule 3CLpro inhibitors effective for inhibiting replication of human coronaviruses including SARS-CoV-2 in cell culture and in animal models. Here we generated a series of deuterated variants of a 3CLpro inhibitor, GC376, and evaluated the antiviral effect against SARS-CoV-2. The deuterated GC376 displayed potent inhibitory activity against SARS-CoV-2 in the enzyme- and the cell-based assays. The K18-hACE2 mice develop mild to lethal infection commensurate with SARS-CoV-2 challenge doses and were proposed as a model for efficacy testing of antiviral agents. We treated lethally infected mice with a deuterated derivative of GC376. Treatment of K18-hACE2 mice at 24 h postinfection with a derivative (compound 2) resulted in increased survival of mice compared to vehicle-treated mice. Lung virus titers were decreased, and histopathological changes were ameliorated in compound 2-treated mice compared to vehicle-treated mice. Structural investigation using high-resolution crystallography illuminated binding interactions of 3CLpro of SARS-CoV-2 and SARS-CoV with deuterated variants of GC376. Taken together, deuterated GC376 variants have excellent potential as antiviral agents against SARS-CoV-2.


Subject(s)
Antiviral Agents/therapeutic use , COVID-19/drug therapy , Coronavirus 3C Proteases/antagonists & inhibitors , Coronavirus Papain-Like Proteases/antagonists & inhibitors , Protease Inhibitors/therapeutic use , Pyrrolidines/therapeutic use , SARS-CoV-2/drug effects , Angiotensin-Converting Enzyme 2/genetics , Animals , Antiviral Agents/chemical synthesis , Antiviral Agents/chemistry , Antiviral Agents/pharmacology , COVID-19/pathology , Coronavirus 3C Proteases/chemistry , Coronavirus Papain-Like Proteases/chemistry , Crystallography, X-Ray , Deuterium , Disease Models, Animal , Drug Evaluation, Preclinical , Female , Lung/pathology , Mice , Mice, Transgenic , Models, Molecular , Molecular Structure , Protease Inhibitors/chemical synthesis , Protease Inhibitors/chemistry , Protease Inhibitors/pharmacology , Protein Conformation , Pyrrolidines/chemistry , SARS-CoV-2/enzymology , Sulfonic Acids , Transgenes
20.
Biochem J ; 478(13): 2517-2531, 2021 07 16.
Article in English | MEDLINE | ID: covidwho-1290988

ABSTRACT

The COVID-19 pandemic has emerged as the biggest life-threatening disease of this century. Whilst vaccination should provide a long-term solution, this is pitted against the constant threat of mutations in the virus rendering the current vaccines less effective. Consequently, small molecule antiviral agents would be extremely useful to complement the vaccination program. The causative agent of COVID-19 is a novel coronavirus, SARS-CoV-2, which encodes at least nine enzymatic activities that all have drug targeting potential. The papain-like protease (PLpro) contained in the nsp3 protein generates viral non-structural proteins from a polyprotein precursor, and cleaves ubiquitin and ISG protein conjugates. Here we describe the expression and purification of PLpro. We developed a protease assay that was used to screen a custom compound library from which we identified dihydrotanshinone I and Ro 08-2750 as compounds that inhibit PLpro in protease and isopeptidase assays and also inhibit viral replication in cell culture-based assays.


Subject(s)
Antiviral Agents/chemistry , Antiviral Agents/pharmacology , Coronavirus Papain-Like Proteases/antagonists & inhibitors , Drug Evaluation, Preclinical , SARS-CoV-2/enzymology , Small Molecule Libraries/pharmacology , Adenosine Monophosphate/analogs & derivatives , Adenosine Monophosphate/pharmacology , Alanine/analogs & derivatives , Alanine/pharmacology , Aniline Compounds/pharmacology , Animals , Benzamides/pharmacology , Chlorocebus aethiops , Coronavirus Papain-Like Proteases/genetics , Coronavirus Papain-Like Proteases/isolation & purification , Coronavirus Papain-Like Proteases/metabolism , Drug Synergism , Enzyme Assays , Flavins/pharmacology , Fluorescence Resonance Energy Transfer , Furans/pharmacology , High-Throughput Screening Assays , Inhibitory Concentration 50 , Naphthalenes/pharmacology , Phenanthrenes/pharmacology , Quinones/pharmacology , Reproducibility of Results , SARS-CoV-2/drug effects , SARS-CoV-2/growth & development , Small Molecule Libraries/chemistry , Vero Cells , Virus Replication/drug effects
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