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1.
Nat Commun ; 12(1): 4396, 2021 07 20.
Article in English | MEDLINE | ID: covidwho-1387353

ABSTRACT

Rapid development of antisense therapies can enable on-demand responses to new viral pathogens and make personalized medicine for genetic diseases practical. Antisense phosphorodiamidate morpholino oligomers (PMOs) are promising candidates to fill such a role, but their challenging synthesis limits their widespread application. To rapidly prototype potential PMO drug candidates, we report a fully automated flow-based oligonucleotide synthesizer. Our optimized synthesis platform reduces coupling times by up to 22-fold compared to previously reported methods. We demonstrate the power of our automated technology with the synthesis of milligram quantities of three candidate therapeutic PMO sequences for an unserved class of Duchenne muscular dystrophy (DMD). To further test our platform, we synthesize a PMO that targets the genomic mRNA of SARS-CoV-2 and demonstrate its antiviral effects. This platform could find broad application not only in designing new SARS-CoV-2 and DMD antisense therapeutics, but also for rapid development of PMO candidates to treat new and emerging diseases.


Subject(s)
Chemistry Techniques, Synthetic/instrumentation , Chemistry, Pharmaceutical/instrumentation , High-Throughput Screening Assays/instrumentation , Morpholinos/chemical synthesis , Oligonucleotides, Antisense/chemical synthesis , Animals , COVID-19/drug therapy , COVID-19/virology , Chlorocebus aethiops , Communicable Diseases, Emerging/drug therapy , Communicable Diseases, Emerging/microbiology , Disease Models, Animal , High-Throughput Screening Assays/methods , Humans , Morpholinos/pharmacology , Morpholinos/therapeutic use , Muscular Dystrophy, Duchenne/drug therapy , Muscular Dystrophy, Duchenne/genetics , Oligonucleotides, Antisense/pharmacology , Oligonucleotides, Antisense/therapeutic use , Precision Medicine/methods , RNA, Messenger/antagonists & inhibitors , RNA, Viral/antagonists & inhibitors , SARS-CoV-2/genetics , Time Factors , Vero Cells
2.
Eur J Med Chem ; 223: 113622, 2021 Nov 05.
Article in English | MEDLINE | ID: covidwho-1263253

ABSTRACT

The emerging severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is responsible for the global pandemic coronavirus disease (COVID-19), but no specific antiviral drug has been proven effective for controlling this pandemic to date. In this study, several 2-((indol-3-yl)thio)-N-benzyl-acetamides were identified as SARS-CoV-2 RNA-dependent RNA polymerase (RdRp) inhibitors. After a two-round optimization, a new series of 2-((indol-3-yl)thio)-N-benzyl-acetamides was designed, synthesized, and evaluated for SARS-CoV-2 RdRp inhibitory effect. Compounds 6b2, 6b5, 6c9, 6d2, and 6d5 were identified as potent inhibitors with IC50 values of 3.35 ± 0.21 µM, 4.55 ± 0.2 µM, 1.65 ± 0.05 µM, 3.76 ± 0.79 µM, and 1.11 ± 0.05 µM, respectively; the IC50 of remdesivir (control) was measured as 1.19 ± 0.36 µM. All of the compounds inhibited RNA synthesis by SARS-CoV-2 RdRp. The most potent compound 6d5, which showed a stronger inhibitory activity against the human coronavirus HCoV-OC43 than remdesivir, is a promising candidate for further investigation.


Subject(s)
Acetamides/chemical synthesis , Antiviral Agents/chemical synthesis , COVID-19/drug therapy , Enzyme Inhibitors/chemical synthesis , RNA, Viral/antagonists & inhibitors , RNA-Dependent RNA Polymerase/antagonists & inhibitors , SARS-CoV-2/drug effects , Acetamides/pharmacology , Adenosine Monophosphate/analogs & derivatives , Adenosine Monophosphate/pharmacology , Adenosine Monophosphate/standards , Alanine/analogs & derivatives , Alanine/pharmacology , Alanine/standards , Antiviral Agents/pharmacology , Drug Evaluation, Preclinical , Enzyme Inhibitors/pharmacology , Humans , Inhibitory Concentration 50 , Molecular Docking Simulation , Molecular Structure , Protein Binding , SARS-CoV-2/genetics , Structure-Activity Relationship
3.
Bioorg Med Chem Lett ; 43: 128052, 2021 07 01.
Article in English | MEDLINE | ID: covidwho-1196690

ABSTRACT

Ciclesonide is an inhaled corticosteroid used to treat asthma and is currently undergoing clinical trials for treatment of coronavirus disease 2019 (COVID-19). An active metabolite of ciclesonide, Cic2, was recently reported to repress severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2) genomic RNA replication. Herein, we designed and synthesized a few types of ciclesonide analogues. Cic4 (bearing an azide group) and Cic6 (bearing a chloro group) potently decreased SARS-CoV-2 viral replication and had low cytotoxicity compared with Cic2 (bearing a hydroxy group). These compounds are promising as novel therapeutic agents for COVID-19 that show significant antiviral activity.


Subject(s)
COVID-19/drug therapy , Pregnenediones/pharmacology , RNA, Viral/antagonists & inhibitors , SARS-CoV-2/drug effects , COVID-19/virology , Glucocorticoids/pharmacology , Humans , RNA, Viral/genetics , SARS-CoV-2/genetics , Virus Replication/genetics
4.
SLAS Discov ; 26(6): 757-765, 2021 07.
Article in English | MEDLINE | ID: covidwho-1194439

ABSTRACT

Frequent outbreaks of novel coronaviruses (CoVs), highlighted by the current severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) pandemic, necessitate the development of therapeutics that could be easily and effectively administered worldwide. The conserved mRNA-capping process enables CoVs to evade their host immune system and is a target for antiviral development. Nonstructural protein (nsp) 16 in complex with nsp10 catalyzes the final step of coronaviral mRNA capping through its 2'-O-methylation activity. Like other methyltransferases, the SARS-CoV-2 nsp10-nsp16 complex is druggable. However, the availability of an optimized assay for high-throughput screening (HTS) is an unmet need. Here, we report the development of a radioactivity-based assay for the methyltransferase activity of the nsp10-nsp16 complex in a 384-well format, kinetic characterization, and optimization of the assay for HTS (Z' factor = 0.83). Considering the high conservation of nsp16 across known CoV species, the potential inhibitors targeting the SARS-CoV-2 nsp10-nsp16 complex may also be effective against other emerging pathogenic CoVs.


Subject(s)
Adenosine/analogs & derivatives , High-Throughput Screening Assays , RNA Caps/antagonists & inhibitors , RNA, Viral/antagonists & inhibitors , Viral Nonstructural Proteins/antagonists & inhibitors , Viral Regulatory and Accessory Proteins/antagonists & inhibitors , Adenosine/chemistry , Adenosine/pharmacology , COVID-19/virology , Cloning, Molecular , Enzyme Assays , Enzyme Inhibitors/chemistry , Enzyme Inhibitors/pharmacology , Escherichia coli/genetics , Escherichia coli/metabolism , Gene Expression , Genetic Vectors/chemistry , Genetic Vectors/metabolism , Humans , Kinetics , Methylation , Methyltransferases , Models, Molecular , RNA Caps/genetics , RNA Caps/metabolism , RNA, Viral/genetics , RNA, Viral/metabolism , Recombinant Proteins/chemistry , Recombinant Proteins/genetics , Recombinant Proteins/metabolism , SARS-CoV-2/drug effects , SARS-CoV-2/enzymology , SARS-CoV-2/genetics , Tritium , Viral Nonstructural Proteins/chemistry , Viral Nonstructural Proteins/genetics , Viral Nonstructural Proteins/metabolism , Viral Regulatory and Accessory Proteins/chemistry , Viral Regulatory and Accessory Proteins/genetics , Viral Regulatory and Accessory Proteins/metabolism
5.
SLAS Discov ; 26(6): 766-774, 2021 07.
Article in English | MEDLINE | ID: covidwho-1192708

ABSTRACT

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is the virus responsible for the global COVID-19 pandemic. Nonstructural protein 14 (NSP14), which features exonuclease (ExoN) and guanine N7 methyltransferase activity, is a critical player in SARS-CoV-2 replication and fidelity and represents an attractive antiviral target. Initiating drug discovery efforts for nucleases such as NSP14 remains a challenge due to a lack of suitable high-throughput assay methodologies. This report describes the combination of self-assembled monolayers and matrix-assisted laser desorption ionization mass spectrometry to enable the first label-free and high-throughput assay for NSP14 ExoN activity. The assay was used to measure NSP14 activity and gain insight into substrate specificity and the reaction mechanism. Next, the assay was optimized for kinetically balanced conditions and miniaturized, while achieving a robust assay (Z factor > 0.8) and a significant assay window (signal-to-background ratio > 200). Screening 10,240 small molecules from a diverse library revealed candidate inhibitors, which were counterscreened for NSP14 selectivity and RNA intercalation. The assay methodology described here will enable, for the first time, a label-free and high-throughput assay for NSP14 ExoN activity to accelerate drug discovery efforts and, due to the assay flexibility, can be more broadly applicable for measuring other enzyme activities from other viruses or implicated in various pathologies.


Subject(s)
Antiviral Agents/pharmacology , Enzyme Inhibitors/pharmacology , Exonucleases/antagonists & inhibitors , Exoribonucleases/antagonists & inhibitors , High-Throughput Screening Assays , RNA, Viral/antagonists & inhibitors , SARS-CoV-2/drug effects , Viral Nonstructural Proteins/antagonists & inhibitors , Antiviral Agents/chemistry , COVID-19/virology , Cloning, Molecular , Enzyme Assays , Enzyme Inhibitors/chemistry , Escherichia coli/genetics , Escherichia coli/metabolism , Exonucleases/genetics , Exonucleases/metabolism , Exoribonucleases/genetics , Exoribonucleases/metabolism , Gene Expression , Genetic Vectors/chemistry , Genetic Vectors/metabolism , Humans , Kinetics , RNA, Viral/genetics , RNA, Viral/metabolism , Recombinant Proteins/genetics , Recombinant Proteins/metabolism , SARS-CoV-2/enzymology , SARS-CoV-2/genetics , Small Molecule Libraries/chemistry , Small Molecule Libraries/pharmacology , Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization/methods , Substrate Specificity , Viral Nonstructural Proteins/genetics , Viral Nonstructural Proteins/metabolism , Virus Replication/drug effects
6.
FEBS Open Bio ; 11(5): 1452-1464, 2021 05.
Article in English | MEDLINE | ID: covidwho-1168813

ABSTRACT

Human pathogenic RNA viruses are threats to public health because they are prone to escaping the human immune system through mutations of genomic RNA, thereby causing local outbreaks and global pandemics of emerging or re-emerging viral diseases. While specific therapeutics and vaccines are being developed, a broad-spectrum therapeutic agent for RNA viruses would be beneficial for targeting newly emerging and mutated RNA viruses. In this study, we conducted a screen of repurposed drugs using Sendai virus (an RNA virus of the family Paramyxoviridae), with human-induced pluripotent stem cells (iPSCs) to explore existing drugs that may present anti-RNA viral activity. Selected hit compounds were evaluated for their efficacy against two important human pathogens: Ebola virus (EBOV) using Huh7 cells and severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) using Vero E6 cells. Selective estrogen receptor modulators (SERMs), including raloxifene, exhibited antiviral activities against EBOV and SARS-CoV-2. Pioglitazone, a PPARγ agonist, also exhibited antiviral activities against SARS-CoV-2, and both raloxifene and pioglitazone presented a synergistic antiviral effect. Finally, we demonstrated that SERMs blocked entry steps of SARS-CoV-2 into host cells. These findings suggest that the identified FDA-approved drugs can modulate host cell susceptibility against RNA viruses.


Subject(s)
Antiviral Agents/pharmacology , Drug Repositioning , RNA Viruses/drug effects , RNA, Viral/antagonists & inhibitors , SARS-CoV-2/drug effects , Animals , COVID-19/drug therapy , Cell Line , Chlorocebus aethiops , Drug Repositioning/methods , Ebolavirus/drug effects , Ebolavirus/physiology , Humans , Induced Pluripotent Stem Cells/virology , Microbial Sensitivity Tests/methods , Pioglitazone/pharmacology , RNA Viruses/physiology , Raloxifene Hydrochloride/pharmacology , SARS-CoV-2/physiology , Selective Estrogen Receptor Modulators/pharmacology , Sendai virus/drug effects , Sendai virus/physiology , Vero Cells
7.
SLAS Discov ; 26(6): 749-756, 2021 07.
Article in English | MEDLINE | ID: covidwho-1136206

ABSTRACT

Severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2) represents a significant threat to human health. Despite its similarity to related coronaviruses, there are currently no specific treatments for COVID-19 infection, and therefore there is an urgent need to develop therapies for this and future coronavirus outbreaks. Formation of the cap at the 5' end of viral RNA has been shown to help coronaviruses evade host defenses. Nonstructural protein 14 (nsp14) is responsible for N7-methylation of the cap guanosine in coronaviruses. This enzyme is highly conserved among coronaviruses and is a bifunctional protein with both N7-methyltransferase and 3'-5' exonuclease activities that distinguish nsp14 from its human equivalent. Mutational analysis of SARS-CoV nsp14 highlighted its role in viral replication and translation efficiency of the viral genome. In this paper, we describe the characterization and development of a high-throughput assay for nsp14 utilizing RapidFire technology. The assay has been used to screen a library of 1771 Food and Drug Administration (FDA)-approved drugs. From this, we have validated nitazoxanide as a selective inhibitor of the methyltransferase activity of nsp14. Although modestly active, this compound could serve as a starting point for further optimization.


Subject(s)
Antiviral Agents/pharmacology , Exoribonucleases/antagonists & inhibitors , High-Throughput Screening Assays , Nitro Compounds/pharmacology , RNA Caps/antagonists & inhibitors , RNA, Viral/antagonists & inhibitors , SARS-CoV-2/drug effects , Thiazoles/pharmacology , Viral Nonstructural Proteins/antagonists & inhibitors , Antiparasitic Agents/chemistry , Antiparasitic Agents/pharmacology , Antiviral Agents/chemistry , COVID-19/virology , Cloning, Molecular , Drug Repositioning , Enzyme Assays , Enzyme Inhibitors/chemistry , Enzyme Inhibitors/pharmacology , Escherichia coli/genetics , Escherichia coli/metabolism , Exoribonucleases/genetics , Exoribonucleases/metabolism , Gene Expression , Genetic Vectors/chemistry , Genetic Vectors/metabolism , Humans , Kinetics , Mass Spectrometry/methods , Methylation , Nitro Compounds/chemistry , Prescription Drugs/chemistry , Prescription Drugs/pharmacology , RNA Caps/genetics , RNA Caps/metabolism , RNA, Viral/genetics , RNA, Viral/metabolism , Recombinant Fusion Proteins/chemistry , Recombinant Fusion Proteins/genetics , Recombinant Fusion Proteins/metabolism , SARS-CoV-2/enzymology , SARS-CoV-2/genetics , Small Molecule Libraries/chemistry , Small Molecule Libraries/pharmacology , Thiazoles/chemistry , Viral Nonstructural Proteins/genetics , Viral Nonstructural Proteins/metabolism , Virus Replication/drug effects
8.
Artif Cells Nanomed Biotechnol ; 49(1): 204-218, 2021 Dec.
Article in English | MEDLINE | ID: covidwho-1109121

ABSTRACT

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is a zoo tonic, highly pathogenic virus. The new type of coronavirus with contagious nature spread from Wuhan (China) to the whole world in a very short time and caused the new coronavirus disease (COVID-19). COVID-19 has turned into a global public health crisis due to spreading by close person-to-person contact with high transmission capacity. Thus, research about the treatment of the damages caused by the virus or prevention from infection increases everyday. Besides, there is still no approved and definitive, standardized treatment for COVID-19. However, this disaster experienced by human beings has made us realize the significance of having a system ready for use to prevent humanity from viral attacks without wasting time. As is known, nanocarriers can be targeted to the desired cells in vitro and in vivo. The nano-carrier system targeting a specific protein, containing the enzyme inhibiting the action of the virus can be developed. The system can be used by simple modifications when we encounter another virus epidemic in the future. In this review, we present a potential treatment method consisting of a nanoparticle-ribozyme conjugate, targeting ACE-2 receptors by reviewing the virus-associated ribozymes, their structures, types and working mechanisms.


Subject(s)
COVID-19/drug therapy , Nanoparticles/administration & dosage , RNA, Catalytic/therapeutic use , RNA, Viral/antagonists & inhibitors , SARS-CoV-2/drug effects , Angiotensin-Converting Enzyme 2/antagonists & inhibitors , Clinical Trials as Topic , Drug Carriers , Drug Compounding , Drug Design , HIV Infections/drug therapy , HIV-1/drug effects , HIV-1/genetics , Humans , Middle East Respiratory Syndrome Coronavirus/drug effects , Middle East Respiratory Syndrome Coronavirus/genetics , Models, Molecular , Nucleic Acid Conformation , RNA Interference , RNA, Catalytic/administration & dosage , RNA, Catalytic/chemistry , RNA, Catalytic/classification , RNA, Untranslated/classification , RNA, Untranslated/genetics , RNA, Untranslated/therapeutic use , Receptors, Coronavirus/antagonists & inhibitors , SARS Virus/drug effects , SARS Virus/genetics , SARS-CoV-2/genetics , SARS-CoV-2/physiology , Spike Glycoprotein, Coronavirus/physiology , Virus Replication/drug effects
9.
Rev Med Virol ; 31(6): e2225, 2021 11.
Article in English | MEDLINE | ID: covidwho-1095682

ABSTRACT

Convalescent plasma therapy (CPT) has been investigated as a treatment for COVID-19. This review evaluates CPT in COVID-19 and other viral respiratory diseases, including severe acute respiratory syndrome (SARS), Middle East respiratory syndrome (MERS) and influenza. PubMed and Google scholar databases were used to collect eligible publications until 8 December 2020. Meta-analysis used Mantel-Haenszel risk ratio (RR) with 95% confidence interval (CI) and pooled analysis for individual patient data with inverse variance weighted average. The study is registered at PROSPERO with the number of CRD4200270579. Forty-four studies with 36,716 participants were included in the pooled analysis and 20 studies in the meta-analysis. Meta-analysis showed reduction of mortality (RR 0.57, 95% CI [0.43, 0.76], z = 3.86 [p < 0.001], I2  = 44% [p = 0.03]) and higher number of discharged patients (RR 2.53, 95% CI [1.72, 3.72], z = 4.70 [p < 0.001], I2  = 3% [p = 0.39]) in patients receiving CPT compared to standard care alone. A possible mechanism of action is prompt reduction in viral titre. Serious transfusion-related adverse events were reported to be less than 1% of cases, suggesting the overall safety of CPT; nevertheless, the number of patients participating in the studies was still limited. It is also important to notice that in all the studies, the majority of patients were also given other medications, such as antivirals, antibiotics and corticosteroid; furthermore, randomized controlled studies involving more patients and in combination with other treatment modalities are urgently needed.


Subject(s)
COVID-19/therapy , Coronavirus Infections/therapy , Influenza, Human/therapy , Severe Acute Respiratory Syndrome/therapy , Adrenal Cortex Hormones/therapeutic use , Anti-Bacterial Agents/therapeutic use , Antiviral Agents/therapeutic use , COVID-19/immunology , COVID-19/mortality , COVID-19/virology , Combined Modality Therapy/methods , Coronavirus Infections/immunology , Coronavirus Infections/mortality , Coronavirus Infections/virology , Humans , Immunization, Passive , Influenza A Virus, H1N1 Subtype/genetics , Influenza A Virus, H1N1 Subtype/immunology , Influenza, Human/immunology , Influenza, Human/mortality , Influenza, Human/virology , Middle East Respiratory Syndrome Coronavirus/drug effects , Middle East Respiratory Syndrome Coronavirus/immunology , Middle East Respiratory Syndrome Coronavirus/pathogenicity , RNA, Viral/antagonists & inhibitors , RNA, Viral/genetics , RNA, Viral/immunology , SARS Virus/drug effects , SARS Virus/immunology , SARS Virus/pathogenicity , SARS-CoV-2/drug effects , SARS-CoV-2/immunology , SARS-CoV-2/pathogenicity , Severe Acute Respiratory Syndrome/immunology , Severe Acute Respiratory Syndrome/mortality , Severe Acute Respiratory Syndrome/virology , Survival Analysis , Treatment Outcome
13.
Chem Soc Rev ; 50(6): 3647-3655, 2021 Mar 21.
Article in English | MEDLINE | ID: covidwho-1057718

ABSTRACT

Clinically approved antiviral drugs are currently available for only 10 of the more than 220 viruses known to infect humans. The SARS-CoV-2 outbreak has exposed the critical need for compounds that can be rapidly mobilised for the treatment of re-emerging or emerging viral diseases, while vaccine development is underway. We review the current status of antiviral therapies focusing on RNA viruses, highlighting strategies for antiviral drug discovery and discuss the challenges, solutions and options to accelerate drug discovery efforts.


Subject(s)
Antiviral Agents/pharmacology , COVID-19/drug therapy , Drug Discovery/methods , Molecular Targeted Therapy/methods , Pandemics/prevention & control , RNA, Viral/antagonists & inhibitors , Antiviral Agents/chemistry , Biological Products/chemistry , Biological Products/pharmacology , COVID-19/prevention & control , COVID-19/virology , Coronavirus Protease Inhibitors/chemistry , Coronavirus Protease Inhibitors/pharmacology , Humans , Molecular Docking Simulation , Nucleic Acid Synthesis Inhibitors/chemistry , Nucleic Acid Synthesis Inhibitors/pharmacology , RNA, Viral/chemistry , RNA, Viral/genetics , RNA, Viral/metabolism , SARS-CoV-2/chemistry , SARS-CoV-2/drug effects , SARS-CoV-2/enzymology , SARS-CoV-2/genetics , Small Molecule Libraries/chemistry , Small Molecule Libraries/pharmacology
14.
SLAS Discov ; 26(5): 620-627, 2021 06.
Article in English | MEDLINE | ID: covidwho-1021348

ABSTRACT

SARS-CoV-2, the coronavirus that causes COVID-19, evades the human immune system by capping its RNA. This process protects the viral RNA and is essential for its replication. Multiple viral proteins are involved in this RNA capping process, including the nonstructural protein 16 (nsp16), which is an S-adenosyl-l-methionine (SAM)-dependent 2'-O-methyltransferase. Nsp16 is significantly active when in complex with another nonstructural protein, nsp10, which plays a key role in its stability and activity. Here we report the development of a fluorescence polarization (FP)-based RNA displacement assay for nsp10-nsp16 complex in a 384-well format with a Z' factor of 0.6, suitable for high-throughput screening. In this process, we purified the nsp10-nsp16 complex to higher than 95% purity and confirmed its binding to the methyl donor SAM, the product of the reaction, S-adenosyl-l-homocysteine (SAH), and a common methyltransferase inhibitor, sinefungin, using isothermal titration calorimetry (ITC). The assay was further validated by screening a library of 1124 drug-like compounds. This assay provides a cost-effective high-throughput method for screening the nsp10-nsp16 complex for RNA competitive inhibitors toward developing COVID-19 therapeutics.


Subject(s)
Antiviral Agents/pharmacology , High-Throughput Screening Assays , RNA, Viral/antagonists & inhibitors , SARS-CoV-2/drug effects , Small Molecule Libraries/pharmacology , Viral Nonstructural Proteins/antagonists & inhibitors , Viral Regulatory and Accessory Proteins/antagonists & inhibitors , Adenosine/analogs & derivatives , Adenosine/pharmacology , Binding, Competitive , COVID-19/drug therapy , COVID-19/virology , Enzyme Inhibitors/pharmacology , Fluorescence Polarization , Gene Expression Regulation , Host-Pathogen Interactions/drug effects , Humans , Methyltransferases , Protein Binding , RNA Caps/antagonists & inhibitors , RNA Caps/genetics , RNA Caps/metabolism , RNA, Viral/genetics , RNA, Viral/metabolism , SARS-CoV-2/genetics , SARS-CoV-2/metabolism , Signal Transduction , Viral Nonstructural Proteins/genetics , Viral Nonstructural Proteins/metabolism , Viral Regulatory and Accessory Proteins/genetics , Viral Regulatory and Accessory Proteins/metabolism , Virus Replication
18.
Genomics ; 113(1 Pt 1): 331-343, 2021 01.
Article in English | MEDLINE | ID: covidwho-972544

ABSTRACT

An outbreak, caused by an RNA virus, SARS-CoV-2 named COVID-19 has become pandemic with a magnitude which is daunting to all public health institutions in the absence of specific antiviral treatment. Surface glycoprotein and nucleocapsid phosphoprotein are two important proteins of this virus facilitating its entry into host cell and genome replication. Small interfering RNA (siRNA) is a prospective tool of the RNA interference (RNAi) pathway for the control of human viral infections by suppressing viral gene expression through hybridization and neutralization of target complementary mRNA. So, in this study, the power of RNA interference technology was harnessed to develop siRNA molecules against specific target genes namely, nucleocapsid phosphoprotein gene and surface glycoprotein gene. Conserved sequence from 139 SARS-CoV-2 strains from around the globe was collected to construct 78 siRNA that can inactivate nucleocapsid phosphoprotein and surface glycoprotein genes. Finally, based on GC content, free energy of folding, free energy of binding, melting temperature, efficacy prediction and molecular docking analysis, 8 siRNA molecules were selected which are proposed to exert the best action. These predicted siRNAs should effectively silence the genes of SARS-CoV-2 during siRNA mediated treatment assisting in the response against SARS-CoV-2.


Subject(s)
COVID-19/therapy , Computational Chemistry , Coronavirus Nucleocapsid Proteins/genetics , Drug Design , Genetic Therapy/methods , Molecular Docking Simulation , RNA Interference , RNA, Messenger/antagonists & inhibitors , RNA, Small Interfering/chemistry , RNA, Viral/antagonists & inhibitors , SARS-CoV-2/genetics , Spike Glycoprotein, Coronavirus/genetics , Argonaute Proteins/chemistry , Argonaute Proteins/genetics , Base Composition , COVID-19/drug therapy , COVID-19/virology , Evolution, Molecular , Gene Expression Regulation, Viral/drug effects , Humans , Pandemics , Phosphoproteins/genetics , Phylogeny , RNA Folding , RNA, Guide/chemistry , RNA, Guide/genetics , RNA, Messenger/genetics , RNA, Small Interfering/pharmacology , RNA, Small Interfering/therapeutic use , RNA, Viral/genetics , SARS-CoV-2/drug effects , Sequence Alignment , Thermodynamics
19.
Nat Commun ; 11(1): 6041, 2020 11 27.
Article in English | MEDLINE | ID: covidwho-947535

ABSTRACT

The etiologic agent of the Covid-19 pandemic is the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). The viral membrane of SARS-CoV-2 surrounds a helical nucleocapsid in which the viral genome is encapsulated by the nucleocapsid protein. The nucleocapsid protein of SARS-CoV-2 is produced at high levels within infected cells, enhances the efficiency of viral RNA transcription, and is essential for viral replication. Here, we show that RNA induces cooperative liquid-liquid phase separation of the SARS-CoV-2 nucleocapsid protein. In agreement with its ability to phase separate in vitro, we show that the protein associates in cells with stress granules, cytoplasmic RNA/protein granules that form through liquid-liquid phase separation and are modulated by viruses to maximize replication efficiency. Liquid-liquid phase separation generates high-density protein/RNA condensates that recruit the RNA-dependent RNA polymerase complex of SARS-CoV-2 providing a mechanism for efficient transcription of viral RNA. Inhibition of RNA-induced phase separation of the nucleocapsid protein by small molecules or biologics thus can interfere with a key step in the SARS-CoV-2 replication cycle.


Subject(s)
COVID-19/virology , Coronavirus Nucleocapsid Proteins/metabolism , RNA, Viral/metabolism , SARS-CoV-2/physiology , Animals , Antiviral Agents/pharmacology , Antiviral Agents/therapeutic use , COVID-19/drug therapy , COVID-19/epidemiology , Coronavirus Nucleocapsid Proteins/antagonists & inhibitors , Coronavirus RNA-Dependent RNA Polymerase/metabolism , HeLa Cells , Humans , Insecta , Intravital Microscopy , Microscopy, Fluorescence , Molecular Dynamics Simulation , Pandemics/prevention & control , Phosphoproteins/antagonists & inhibitors , Phosphoproteins/metabolism , RNA, Viral/antagonists & inhibitors , SARS-CoV-2/drug effects , Viral Transcription/drug effects , Viral Transcription/physiology , Virus Replication/drug effects , Virus Replication/genetics
20.
Biomed Pharmacother ; 131: 110738, 2020 Nov.
Article in English | MEDLINE | ID: covidwho-773856

ABSTRACT

The novel coronavirus called severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) can be considered as the most important current global issue, as it has caused the novel coronavirus disease (COVID-19) pandemic, which has resulted in high mortality and morbidity rates all around the world. Although scientists are trying to discover novel therapies and develop and evaluate various previous treatments, at the time of writing this paper, there was no definite therapy and vaccine for COVID-19. So, as COVID-19 has called ideas for treatment, controlling, and diagnosis, we discussed the application of Clustered Regularly Interspaced Short Palindromic Repeats/Cas13 (CRISPR/Cas13) as a treatment of COVID-19, which received less attention compared with other potential therapeutic options.


Subject(s)
Betacoronavirus/genetics , CRISPR-Cas Systems , Coronavirus Infections/therapy , Gene Editing , Genetic Therapy/methods , Pneumonia, Viral/therapy , RNA, Viral/genetics , Betacoronavirus/drug effects , COVID-19 , CRISPR-Associated Proteins/pharmacology , Conserved Sequence , Coronavirus Infections/drug therapy , Coronavirus Infections/genetics , Genome, Viral , Humans , Pandemics , Pneumonia, Viral/genetics , RNA, Guide/genetics , RNA, Viral/antagonists & inhibitors , SARS-CoV-2
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