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1.
Molecules ; 27(3)2022 Jan 21.
Article in English | MEDLINE | ID: covidwho-1649980

ABSTRACT

COVID-19 has spread around the world and caused serious public health and social problems. Although several vaccines have been authorized for emergency use, new effective antiviral drugs are still needed. Some repurposed drugs including Chloroquine, Hydroxychloroquine and Remdesivir were immediately used to treat COVID-19 after the pandemic. However, the therapeutic effects of these drugs have not been fully demonstrated in clinical studies. In this paper, we found an antimalarial drug, Naphthoquine, showed good broad-spectrum anti-coronavirus activity. Naphthoquineinhibited HCoV-229E, HCoV-OC43 and SARS-CoV-2 replication in vitro, with IC50 = 2.05 ± 1.44 µM, 5.83 ± 0.74 µM, and 2.01 ± 0.38 µM, respectively. Time-of-addition assay was also performed to explore at which stage Naphthoquine functions during SARS-CoV-2 replication. The results suggested that Naphthoquine may influence virus entry and post-entry replication. Considering the safety of Naphthoquine was even better than that of Chloroquine, we think Naphthoquine has the potential to be used as a broad-spectrum drug for coronavirus infection.


Subject(s)
1-Naphthylamine/analogs & derivatives , Aminoquinolines/pharmacology , Antiviral Agents/pharmacology , Coronavirus/drug effects , SARS-CoV-2/drug effects , 1-Naphthylamine/pharmacology , Animals , Cell Line , Chlorocebus aethiops , Coronavirus 229E, Human/drug effects , Coronavirus NL63, Human/drug effects , Coronavirus OC43, Human/drug effects , Humans , In Vitro Techniques , Vero Cells , Virus Replication/drug effects
2.
J Ethnopharmacol ; 287: 114965, 2022 Apr 06.
Article in English | MEDLINE | ID: covidwho-1587284

ABSTRACT

ETHNOPHARMACOLOGICAL RELEVANCE: Coronavirus and influenza virus infection seriously threaten human health. Cangma Huadu Granules (CMHD) is an in-hospital preparation composed of eight traditional Chinese medicines (TCM), which has been clinically used against COVID-19 in China and may be a promising candidate for the treatment of influenza. However, the role of its treatment urgently needs to be studied. AIM OF THE STUDY: To evaluate the therapeutic effects of CMHD on pneumonia induced by coronavirus (HCoV-229E) and influenza A virus (H1N1/FM1) in mice and explore its mechanism of anti-infection. MATERIALS AND METHODS: Mice were infected with HCoV-229E or H1N1/FM1 virus through the nasal cavity. CMHD (12.1, 6.05 and 3.03 g/kg/d) or the positive control drugs were administered intragastrically. The lung index and histopathological changes were used to evaluate the therapeutic effect of CMHD. The expression of TNF-α, IL-1ß, IL-6 and IL-4 in Serum and the proportion of CD4+ and CD8+ T lymphocytes in peripheral blood were detected to evaluate the anti-inflammatory and immune regulation effects of CMHD, respectively. Furthermore, the levels of p-NF-κBp65/ NF-κB p65, which was the key targets of the NF-κB pathway was analyzed. RESULTS: In HCoV-229E-induced pneumonia, the lung index was markedly reduced, and lung pathology was improved in mice that treated with CMHD (12.1, 6.05 g/kg/d). Meanwhile, the expression of TNF-α, IL-6 were obviously inhibited, but the expression of IL-4 was significantly increased in CMHD groups. Compared with the model group, CMHD could also markedly upregulate the level of CD4+ and CD8+. Furthermore, CMHD has a markedly effect on inhibit the expression of p-NF-κB p65/NF-κB p65 in the lung. In H1N1-induced pneumonia, the lung index of mice in the CMHD (12.1 g/kg/d) treatment group was lower than that in the model group, and less inflammatory infiltration could be seen in the lung pathological. Moreover, CMHD could also obviously decrease the expression of TNF-α, IL-1ß, IL-6, but significantly increase the expression of IL-4. Except for that, CMHD could also markedly downregulate the level of CD4+ and upregulate the level of CD8+ compared with the model group. In addition, CMHD has a markedly effect on inhibit the expression of p-NF-κB p65/NF-κB p65 in the lung. CONCLUSION: CMHD can significantly combats viral infections caused by HCoV-229E and H1N1, and the mechanism may be related to its multiple functions of anti-inflammatory, immunity regulating and inhibiting NF-κB signal transduction pathway.


Subject(s)
Anti-Inflammatory Agents/pharmacology , Coronavirus Infections/drug therapy , Drugs, Chinese Herbal/pharmacology , Influenza A Virus, H1N1 Subtype/drug effects , Medicine, Chinese Traditional/methods , Orthomyxoviridae Infections/drug therapy , Animals , Anti-Inflammatory Agents/therapeutic use , Coronavirus 229E, Human/drug effects , Cytokines/metabolism , Disease Models, Animal , Drugs, Chinese Herbal/therapeutic use , Female , Immunity/drug effects , Male , Mice, Inbred BALB C , Mice, Inbred ICR , Pneumonia/drug therapy , Pneumonia/pathology , T-Lymphocytes/metabolism , Transcription Factor RelA/metabolism
3.
Phytomedicine ; 95: 153874, 2022 Jan.
Article in English | MEDLINE | ID: covidwho-1560696

ABSTRACT

BACKGROUND: Although severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) and human coronavirus 229E (HCoV-229E) pose a huge threat to human public health, no specific treatment is available. Jinzhen granule (JZ) is a traditional eight ingredients-Chinese medicine with prominent efficacy for treating viral-induced diseases. However, little is known about the antiviral effect and mechanism of JZ against SARS-CoV-2 and HCoV-229E. PURPOSE: This study aimed to reveal the antiviral effects of JZ against SARS-CoV-2 and HCoV-229E, and to further explore the underlying mechanisms regulating the host immune response. METHODS: The chromatographic separation of JZ was performed using a Shimadzu analytical high-performance liquid chromatograph with UV detection and Alltech ELSD 2000ES. We conducted cytopathic effect (CPE) and plaque reduction assays to evaluate the antiviral effect of JZ. A lethal human angiotensin converting enzyme 2 (hACE2) transgenic mouse model of SARS-CoV-2 was established to determine the protective effect of JZ on mortality and lung virus titers. Real-time quantitative PCR assays were used to analyze the expression of proinflammatory cytokines in vitro and in vivo. Western blotting was further performed to determine the activities on regulating the nuclear factor kappa B (NF-κB)/MAPK pathway. Finally, mitochondrial membrane potential assays, flow cytometry analysis and western blotting were used to assess the anti-apoptotic potency toward HCoV-229E infection. RESULTS: The results showed that 13 chemical components were identified and five peaks were determined and quantitated (gallic acid 1.97 mg/g, baicalin 20.69 mg/g, glycyrrhizic acid 4.92 mg/g, hyodeoxycholic acid 4.86 mg/g, cholic acid 4.07 mg/g). We found that JZ exerted inhibitory potency against SARS-CoV-2 and HCoV-229E in vitro by using CPE and plaque reduction assays, and it was further found that JZ protected mice infected by SARS-CoV-2 from death and inhibited lung virus titers. JZ also significantly decreased the induction of inflammatory cytokines (IL-1α, IL-6, CCL-5 and MIP-1ß), similar to the observed in vitro effect. Moreover, JZ suppressed the release of inflammatory cytokines in vitro and it decreased the protein expression of p-p38 MAPK, p-JNK, p-NF-κB p65 and p-IκBα induced by HCoV-229E and increased the expression of IκBα. Notably, JZ significantly protected HCoV-229E-infected Huh-7 cells from mitochondrial damage and decreased apoptotic cells. The activation of the mitochondria-mediated apoptotic pathway was inhibited by JZ, as shown by the reduced expression of cleaved caspase-9, caspase-3 and p-PARP. CONCLUSIONS: In conclusion, JZ (gallic acid 1.97 mg/g, baicalin 20.69 mg/g, glycyrrhizic acid 4.92 mg/g, hyodeoxycholic acid 4.86 mg/g, cholic acid 4.07 mg/g) exhibited antiviral activities against SARS-CoV-2 and HCoV-229E by regulating the NF-κB/MAPK pathway and the mitochondria-mediated apoptotic pathway. These findings demonstrated the efficacy of JZ against CoVs and suggested JZ treatment as a novel clinical therapeutic strategy for COVID-19.


Subject(s)
Antiviral Agents , Coronavirus 229E, Human , Drugs, Chinese Herbal/pharmacology , SARS-CoV-2/drug effects , Animals , Antiviral Agents/pharmacology , COVID-19 , Coronavirus 229E, Human/drug effects , Humans , MAP Kinase Signaling System , Mice , NF-kappa B
4.
Viruses ; 13(11)2021 11 04.
Article in English | MEDLINE | ID: covidwho-1502533

ABSTRACT

The COVID-19 pandemic has deeply influenced sanitization procedures, and high-level disinfection has been massively used to prevent SARS-CoV-2 spread, with potential negative impact on the environment and on the threat of antimicrobial resistance (AMR). Aiming to overcome these concerns, yet preserving the effectiveness of sanitization against enveloped viruses, we assessed the antiviral properties of the Probiotic Cleaning Hygiene System (PCHS), an eco-sustainable probiotic-based detergent previously proven to stably abate pathogen contamination and AMR. PCHS (diluted 1:10, 1:50 and 1:100) was tested in comparison to common disinfectants (70% ethanol and 0.5% sodium hypochlorite), in suspension and carrier tests, according with the European UNI EN 14476:2019 and UNI EN 16777:2019 standards. Human alpha- and beta-coronaviruses hCoV-229E and SARS-CoV-2, human herpesvirus type 1, human and animal influenza viruses, and vaccinia virus were included in the study. The results showed that PCHS was able to inactivate 99.99% of all tested viruses within 1-2 h of contact, both in suspension and on surface. Notably, while control disinfectants became inactive within 2 h after application, the PCHS antiviral action persisted up to 24 h post-application, suggesting that its use may effectively allow a continuous prevention of virus spread via contaminated environment, without worsening environmental pollution and AMR concern.


Subject(s)
Disinfection/methods , Probiotics/pharmacology , Sanitation/methods , Virus Diseases/prevention & control , Viruses/drug effects , Animals , Antiviral Agents/pharmacology , COVID-19/prevention & control , COVID-19/virology , Coronavirus 229E, Human/drug effects , Disinfectants/pharmacology , Environmental Microbiology , Herpesvirus 1, Human/drug effects , Humans , Orthomyxoviridae/drug effects , SARS-CoV-2/drug effects , Vaccinia virus/drug effects , Virus Diseases/virology
5.
Int J Mol Sci ; 22(21)2021 Nov 01.
Article in English | MEDLINE | ID: covidwho-1488619

ABSTRACT

The COVID-19 pandemic caused by SARS-CoV-2 infection poses a serious threat to global public health and the economy. The enzymatic product of cholesterol 25-hydroxylase (CH25H), 25-Hydroxycholesterol (25-HC), was reported to have potent anti-SARS-CoV-2 activity. Here, we found that the combination of 25-HC with EK1 peptide, a pan-coronavirus (CoV) fusion inhibitor, showed a synergistic antiviral activity. We then used the method of 25-HC modification to design and synthesize a series of 25-HC-modified peptides and found that a 25-HC-modified EK1 peptide (EK1P4HC) was highly effective against infections caused by SARS-CoV-2, its variants of concern (VOCs), and other human CoVs, such as HCoV-OC43 and HCoV-229E. EK1P4HC could protect newborn mice from lethal HCoV-OC43 infection, suggesting that conjugation of 25-HC with a peptide-based viral inhibitor was a feasible and universal strategy to improve its antiviral activity.


Subject(s)
Antiviral Agents/pharmacology , Hydroxycholesterols/chemistry , Lipopeptides/chemistry , SARS-CoV-2/drug effects , Amino Acid Sequence , Animals , Antiviral Agents/chemistry , Antiviral Agents/therapeutic use , Body Weight/drug effects , COVID-19/drug therapy , COVID-19/virology , Coronavirus 229E, Human/drug effects , Coronavirus 229E, Human/pathogenicity , Coronavirus Infections/drug therapy , Coronavirus Infections/mortality , Coronavirus Infections/virology , Coronavirus OC43, Human/drug effects , Coronavirus OC43, Human/pathogenicity , Disease Models, Animal , Drug Synergism , Humans , Hydroxycholesterols/pharmacology , Hydroxycholesterols/therapeutic use , Lipopeptides/pharmacology , Lipopeptides/therapeutic use , Mice , Mice, Inbred BALB C , Polyethylene Glycols/chemistry , SARS-CoV-2/isolation & purification , SARS-CoV-2/physiology , Survival Rate , Virus Internalization/drug effects
6.
Cells ; 10(11)2021 10 29.
Article in English | MEDLINE | ID: covidwho-1488496

ABSTRACT

Human coronavirus (HCoV) similar to other viruses rely on host cell machinery for both replication and to spread. The p97/VCP ATPase is associated with diverse pathways that may favor HCoV replication. In this study, we assessed the role of p97 and associated host responses in human lung cell line H1299 after HCoV-229E or HCoV-OC43 infection. Inhibition of p97 function by small molecule inhibitors shows antiviral activity, particularly at early stages of the virus life cycle, during virus uncoating and viral RNA replication. Importantly, p97 activity inhibition protects human cells against HCoV-induced cytopathic effects. The p97 knockdown also inhibits viral production in infected cells. Unbiased quantitative proteomics analyses reveal that HCoV-OC43 infection resulted in proteome changes enriched in cellular senescence and DNA repair during virus replication. Further analysis of protein changes between infected cells with control and p97 shRNA identifies cell cycle pathways for both HCoV-229E and HCoV-OC43 infection. Together, our data indicate a role for the essential host protein p97 in supporting HCoV replication, suggesting that p97 is a therapeutic target to treat HCoV infection.


Subject(s)
Coronavirus 229E, Human/physiology , Coronavirus OC43, Human/physiology , Valosin Containing Protein/metabolism , Virus Replication/physiology , Antiviral Agents/pharmacology , Cell Cycle/drug effects , Cell Line , Coronavirus 229E, Human/drug effects , Coronavirus OC43, Human/drug effects , Cytopathogenic Effect, Viral/drug effects , Humans , Proteome/drug effects , Proteome/metabolism , RNA, Small Interfering/genetics , RNA, Viral/biosynthesis , Valosin Containing Protein/antagonists & inhibitors , Valosin Containing Protein/genetics , Virus Replication/drug effects , Virus Uncoating/drug effects
7.
Nat Commun ; 12(1): 6055, 2021 10 18.
Article in English | MEDLINE | ID: covidwho-1475294

ABSTRACT

COVID-19 caused by the SARS-CoV-2 virus has become a global pandemic. 3CL protease is a virally encoded protein that is essential across a broad spectrum of coronaviruses with no close human analogs. PF-00835231, a 3CL protease inhibitor, has exhibited potent in vitro antiviral activity against SARS-CoV-2 as a single agent. Here we report, the design and characterization of a phosphate prodrug PF-07304814 to enable the delivery and projected sustained systemic exposure in human of PF-00835231 to inhibit coronavirus family 3CL protease activity with selectivity over human host protease targets. Furthermore, we show that PF-00835231 has additive/synergistic activity in combination with remdesivir. We present the ADME, safety, in vitro, and in vivo antiviral activity data that supports the clinical evaluation of PF-07304814 as a potential COVID-19 treatment.


Subject(s)
COVID-19/drug therapy , Coronavirus 3C Proteases/antagonists & inhibitors , Coronavirus Protease Inhibitors/administration & dosage , Indoles/administration & dosage , Leucine/administration & dosage , Pyrrolidinones/administration & dosage , Adenosine Monophosphate/administration & dosage , Adenosine Monophosphate/adverse effects , Adenosine Monophosphate/analogs & derivatives , Adenosine Monophosphate/pharmacokinetics , Alanine/administration & dosage , Alanine/adverse effects , Alanine/analogs & derivatives , Alanine/pharmacokinetics , Animals , COVID-19/virology , Chlorocebus aethiops , Coronavirus 229E, Human/drug effects , Coronavirus 229E, Human/enzymology , Coronavirus Protease Inhibitors/adverse effects , Coronavirus Protease Inhibitors/pharmacokinetics , Disease Models, Animal , Drug Design , Drug Synergism , Drug Therapy, Combination , HeLa Cells , Humans , Indoles/adverse effects , Indoles/pharmacokinetics , Infusions, Intravenous , Leucine/adverse effects , Leucine/pharmacokinetics , Mice , Pyrrolidinones/adverse effects , Pyrrolidinones/pharmacokinetics , SARS Virus/drug effects , SARS Virus/enzymology , SARS-CoV-2/drug effects , SARS-CoV-2/enzymology , Vero Cells
8.
Virology ; 564: 33-38, 2021 12.
Article in English | MEDLINE | ID: covidwho-1447220

ABSTRACT

Endemic seasonal coronaviruses cause morbidity and mortality in a subset of patients, but no specific treatment is available. Molnupiravir is a promising pipeline antiviral drug for treating SARS-CoV-2 infection potentially by targeting RNA-dependent RNA polymerase (RdRp). This study aims to evaluate the potential of repurposing molnupiravir for treating seasonal human coronavirus (HCoV) infections. Molecular docking revealed that the active form of molnupiravir, ß-D-N4-hydroxycytidine (NHC), has similar binding affinity to RdRp of SARS-CoV-2 and seasonal HCoV-NL63, HCoV-OC43 and HCoV-229E. In cell culture models, treatment of molnupiravir effectively inhibited viral replication and production of infectious viruses of the three seasonal coronaviruses. A time-of-drug-addition experiment indicates the specificity of molnupiravir in inhibiting viral components. Furthermore, combining molnupiravir with the protease inhibitor GC376 resulted in enhanced antiviral activity. Our findings highlight that the great potential of repurposing molnupiravir for treating seasonal coronavirus infected patients.


Subject(s)
Coronavirus 229E, Human/genetics , Coronavirus Infections/drug therapy , Coronavirus NL63, Human/genetics , Coronavirus OC43, Human/genetics , Cytidine/analogs & derivatives , Hydroxylamines/pharmacology , Antiviral Agents/chemistry , Antiviral Agents/metabolism , Antiviral Agents/pharmacology , Common Cold/drug therapy , Coronavirus 229E, Human/drug effects , Coronavirus 229E, Human/physiology , Coronavirus NL63, Human/drug effects , Coronavirus NL63, Human/physiology , Coronavirus OC43, Human/drug effects , Coronavirus OC43, Human/physiology , Cytidine/pharmacology , Humans , Molecular Docking Simulation , Protein Binding/drug effects , Pyrrolidines/pharmacology , RNA-Dependent RNA Polymerase/chemistry , RNA-Dependent RNA Polymerase/genetics , RNA-Dependent RNA Polymerase/metabolism , Seasons , Sulfonic Acids/pharmacology , Virus Replication/drug effects , Virus Replication/genetics
9.
Virol J ; 17(1): 136, 2020 09 09.
Article in English | MEDLINE | ID: covidwho-1435256

ABSTRACT

BACKGROUND: Coronaviruses (CoVs) were long thought to only cause mild respiratory and gastrointestinal symptoms in humans but outbreaks of Middle East Respiratory Syndrome (MERS)-CoV, Severe Acute Respiratory Syndrome (SARS)-CoV-1, and the recently identified SARS-CoV-2 have cemented their zoonotic potential and their capacity to cause serious morbidity and mortality, with case fatality rates ranging from 4 to 35%. Currently, no specific prophylaxis or treatment is available for CoV infections. Therefore we investigated the virucidal and antiviral potential of Echinacea purpurea (Echinaforce®) against human coronavirus (HCoV) 229E, highly pathogenic MERS- and SARS-CoVs, as well as the newly identified SARS-CoV-2, in vitro. METHODS: To evaluate the antiviral potential of the extract, we pre-treated virus particles and cells and evaluated remaining infectivity by limited dilution. Furthermore, we exposed cells to the extract after infection to further evaluate its potential as a prophylaxis and treatment against coronaviruses. We also determined the protective effect of Echinaforce® in re-constituted nasal epithelium. RESULTS: In the current study, we found that HCoV-229E was irreversibly inactivated when exposed to Echinaforce® at 3.2 µg/ml IC50. Pre-treatment of cell lines, however, did not inhibit infection with HCoV-229E and post-infection treatment had only a marginal effect on virus propagation at 50 µg/ml. However, we did observe a protective effect in an organotypic respiratory cell culture system by exposing pre-treated respiratory epithelium to droplets of HCoV-229E, imitating a natural infection. The observed virucidal activity of Echinaforce® was not restricted to common cold coronaviruses, as both SARS-CoV-1 and MERS-CoVs were inactivated at comparable concentrations. Finally, the causative agent of COVID-19, SARS-CoV-2 was also inactivated upon treatment with 50µg/ml Echinaforce®. CONCLUSIONS: These results show that Echinaforce® is virucidal against HCoV-229E, upon direct contact and in an organotypic cell culture model. Furthermore, MERS-CoV and both SARS-CoV-1 and SARS-CoV-2 were inactivated at similar concentrations of the extract. Therefore we hypothesize that Echinacea purpurea preparations, such as Echinaforce®, could be effective as prophylactic treatment for all CoVs due to their structural similarities.


Subject(s)
Antiviral Agents/pharmacology , Betacoronavirus/drug effects , Coronavirus 229E, Human/drug effects , Coronavirus Infections/drug therapy , Coronavirus/drug effects , Plant Extracts/pharmacology , Plant Extracts/therapeutic use , Animals , COVID-19 , Cell Line , Chlorocebus aethiops , Common Cold/drug therapy , Common Cold/virology , Coronavirus Infections/virology , Humans , Middle East Respiratory Syndrome Coronavirus/drug effects , Pandemics , Pneumonia, Viral/drug therapy , Pneumonia, Viral/virology , RNA Viruses/drug effects , Randomized Controlled Trials as Topic , SARS-CoV-2 , Severe Acute Respiratory Syndrome/drug therapy , Severe Acute Respiratory Syndrome/virology , Vero Cells
10.
Appl Environ Microbiol ; 87(19): e0109821, 2021 09 10.
Article in English | MEDLINE | ID: covidwho-1403023

ABSTRACT

A novel and robust approach to evaluate the antiviral activity of coatings was developed, assessing three commercially available leave-on surface coating products for efficacy against human coronaviruses (HCoVs) HCoV-229E and severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). The assessment is based on three criteria that reflect real-life settings, namely, (i) immediate antiviral effect, (ii) effect after repeated cleaning of the coated surface, and (iii) antiviral activity in the presence of organic material. The results showed that only a copper compound-based coating successfully met all three criteria. A quaternary ammonium compound-based coating did not meet the second criterion, and a coating based on reactive oxygen species showed no antiviral effect. Moreover, the study demonstrated that HCoV-229E is a relevant SARS-CoV-2 surrogate for such experiments. This new approach allows benchmarking of currently available antiviral coatings and future coating developments to avoid unjustified claims. The deployment of efficient antiviral coatings can offer an additional measure to mitigate the risk of transmission of respiratory viruses like SARS-CoV-2 or influenza viruses from high-touch surfaces. IMPORTANCE SARS-CoV-2, the virus responsible for the coronavirus disease 2019 (COVID-19) pandemic, is transmitted mainly person-to-person through respiratory droplets, while the contribution of fomite transmission is less important than suspected at the beginning of the pandemic. Nevertheless, antiviral-coating solutions can offer an additional measure to mitigate the risk of SARS-CoV-2 transmission from high-touch surfaces. The deployment of antiviral coatings is not new, but what is currently lacking is solid scientific evidence of the efficacy of commercially available self-disinfecting surfaces under real-life conditions. Therefore, we developed a novel, robust approach to evaluate the antiviral activity of such coatings, applying strict quality criteria to three commercially available products to test their efficacies against SARS-CoV-2. We also showed that HCoV-229E is a relevant surrogate for such experiments. Our approach will also bring significant benefit to the evaluation of the effects of coatings on the survival of nonenveloped viruses, which are known to be more tolerant to desiccation and disinfectants and for which high-touch surfaces play an important role.


Subject(s)
Antiviral Agents/pharmacology , Coronavirus 229E, Human/drug effects , Disinfectants/pharmacology , SARS-CoV-2/drug effects
11.
Viruses ; 13(8)2021 08 23.
Article in English | MEDLINE | ID: covidwho-1367925

ABSTRACT

An escalating pandemic of the novel SARS-CoV-2 virus is impacting global health, and effective antivirals are needed. Umifenovir (Arbidol) is an indole-derivative molecule, licensed in Russia and China for prophylaxis and treatment of influenza and other respiratory viral infections. It has been shown that umifenovir has broad spectrum activity against different viruses. We evaluated the sensitivity of different coronaviruses, including the novel SARS-CoV-2 virus, to umifenovir using in vitro assays. Using a plaque assay, we revealed an antiviral effect of umifenovir against seasonal HCoV-229E and HCoV-OC43 coronaviruses in Vero E6 cells, with estimated 50% effective concentrations (EC50) of 10.0 ± 0.5 µM and 9.0 ± 0.4 µM, respectively. Umifenovir at 90 µM significantly suppressed plaque formation in CMK-AH-1 cells infected with SARS-CoV. Umifenovir also inhibited the replication of SARS-CoV-2 virus, with EC50 values ranging from 15.37 ± 3.6 to 28.0 ± 1.0 µM. In addition, 21-36 µM of umifenovir significantly suppressed SARS-CoV-2 virus titers (≥2 log TCID50/mL) in the first 24 h after infection. Repurposing of antiviral drugs is very helpful in fighting COVID-19. A safe, pan-antiviral drug such as umifenovir could be extremely beneficial in combating the early stages of a viral pandemic.


Subject(s)
Antiviral Agents/pharmacology , Coronavirus 229E, Human/drug effects , Coronavirus OC43, Human/drug effects , Indoles/pharmacology , SARS Virus/drug effects , SARS-CoV-2/drug effects , Animals , Antiviral Agents/administration & dosage , Cell Survival/drug effects , Chlorocebus aethiops , Coronavirus 229E, Human/physiology , Coronavirus OC43, Human/physiology , Cytopathogenic Effect, Viral/drug effects , Humans , Indoles/administration & dosage , Microbial Sensitivity Tests , SARS Virus/physiology , SARS-CoV-2/physiology , Vero Cells , Viral Load/drug effects , Viral Plaque Assay , Virus Replication/drug effects
12.
Int J Mol Sci ; 22(15)2021 Jul 27.
Article in English | MEDLINE | ID: covidwho-1332900

ABSTRACT

A novel series of N-substituted cis- and trans-3-aryl-4-(diethoxyphosphoryl)azetidin-2-ones were synthesized by the Kinugasa reaction of N-methyl- or N-benzyl-(diethyoxyphosphoryl)nitrone and selected aryl alkynes. Stereochemistry of diastereoisomeric adducts was established based on vicinal H3-H4 coupling constants in azetidin-2-one ring. All the obtained azetidin-2-ones were evaluated for the antiviral activity against a broad range of DNA and RNA viruses. Azetidin-2-one trans-11f showed moderate inhibitory activity against human coronavirus (229E) with EC50 = 45 µM. The other isomer cis-11f was active against influenza A virus H1N1 subtype (EC50 = 12 µM by visual CPE score; EC50 = 8.3 µM by TMS score; MCC > 100 µM, CC50 = 39.9 µM). Several azetidin-2-ones 10 and 11 were tested for their cytostatic activity toward nine cancerous cell lines and several of them appeared slightly active for Capan-1, Hap1 and HCT-116 cells values of IC50 in the range 14.5-97.9 µM. Compound trans-11f was identified as adjuvant of oxacillin with significant ability to enhance the efficacy of this antibiotic toward the highly resistant S. aureus strain HEMSA 5. Docking and molecular dynamics simulations showed that enantiomer (3R,4S)-11f can be responsible for the promising activity due to the potency in displacing oxacillin at ß-lactamase, thus protecting the antibiotic from undesirable biotransformation.


Subject(s)
Adjuvants, Pharmaceutic/chemistry , Adjuvants, Pharmaceutic/pharmacology , Antiviral Agents/chemistry , Antiviral Agents/pharmacology , Azetidines/pharmacology , Infections/drug therapy , Anti-Bacterial Agents/chemistry , Anti-Bacterial Agents/pharmacology , Azetidines/chemistry , Bacterial Proteins/chemistry , Cell Line, Tumor , Cell Proliferation/drug effects , Coronavirus 229E, Human/drug effects , Cytostatic Agents/chemistry , Cytostatic Agents/pharmacology , Humans , Influenza A Virus, H1N1 Subtype/drug effects , Molecular Dynamics Simulation , Oxacillin/chemistry , Penicillin-Binding Proteins/chemistry , Staphylococcus aureus/drug effects , Stereoisomerism , beta-Lactamases/chemistry
13.
Biosensors (Basel) ; 11(8)2021 Jul 26.
Article in English | MEDLINE | ID: covidwho-1325599

ABSTRACT

Cell-based assays are a valuable tool for examination of virus-host cell interactions and drug discovery processes, allowing for a more physiological setting compared to biochemical assays. Despite the fact that cell-based SPR assays are label-free and thus provide all the associated benefits, they have never been used to study viral growth kinetics and to predict drug antiviral response in cells. In this study, we prove the concept that the cell-based SPR assay can be applied in the kinetic analysis of the early stages of viral infection of cells and the antiviral drug activity in the infected cells. For this purpose, cells immobilized on the SPR slides were infected with human coronavirus HCov-229E and treated with hydroxychloroquine. The SPR response was measured at different time intervals within the early stages of infection. Methyl Thiazolyl Tetrazolium (MTT) assay was used to provide the reference data. We found that the results of the SPR and MTT assays were consistent, and SPR is a reliable tool in investigating virus-host cell interaction and the mechanism of action of viral inhibitors. SPR assay was more sensitive and accurate in the first hours of infection within the first replication cycle, whereas the MTT assay was not so effective. After the second replication cycle, noise was generated by the destruction of the cell layer and by the remnants of dead cells, and masks useful SPR signals.


Subject(s)
Antiviral Agents/therapeutic use , Coronavirus 229E, Human/physiology , Coronavirus Infections/drug therapy , Hydroxychloroquine/therapeutic use , Surface Plasmon Resonance/methods , Animals , Antiviral Agents/pharmacology , Chlorocebus aethiops , Coronavirus 229E, Human/drug effects , Coronavirus 229E, Human/isolation & purification , Coronavirus Infections/pathology , Coronavirus Infections/virology , Humans , Hydroxychloroquine/pharmacology , Kinetics , Severity of Illness Index , Vero Cells
14.
Antiviral Res ; 193: 105127, 2021 09.
Article in English | MEDLINE | ID: covidwho-1293551

ABSTRACT

In this study, a series of 10 quinoline analogues was evaluated for their in vitro antiviral activity against a panel of alpha- and beta-coronaviruses, including the severe acute respiratory syndrome coronaviruses 1 and 2 (SARS-CoV-1 and SARS-CoV-2), as well as the human coronaviruses (HCoV) 229E and OC43. Chloroquine and hydroxychloroquine were the most potent with antiviral EC50 values in the range of 0.12-12 µM. Chloroquine displayed the most favorable selectivity index (i.e. ratio cytotoxic versus antiviral concentration), being 165 for HCoV-OC43 in HEL cells. Potent anti-coronavirus activity was also observed with amodiaquine, ferroquine and mefloquine, although this was associated with substantial cytotoxicity for mefloquine. Primaquine, quinidine, quinine and tafenoquine only blocked coronavirus replication at higher concentrations, while piperaquine completely lacked antiviral and cytotoxic effects. A time-of-addition experiment in HCoV-229E-infected HEL cells revealed that chloroquine interferes with viral entry at a post-attachment stage. Using confocal microscopy, no viral RNA synthesis could be detected upon treatment of SARS-CoV-2-infected cells with chloroquine. The inhibition of SARS-CoV-2 replication by chloroquine and hydroxychloroquine coincided with an inhibitory effect on the autophagy pathway as visualized by a dose-dependent increase in LC3-positive puncta. The latter effect was less pronounced or even absent with the other quinolines. In summary, we showed that several quinoline analogues, including chloroquine, hydroxychloroquine, amodiaquine, ferroquine and mefloquine, exhibit broad anti-coronavirus activity in vitro.


Subject(s)
Antimalarials/pharmacology , Antiviral Agents/pharmacology , Coronavirus Infections/drug therapy , Coronavirus/drug effects , Quinolines/pharmacology , Animals , COVID-19/drug therapy , Chlorocebus aethiops , Chloroquine/pharmacology , Coronavirus 229E, Human/drug effects , Coronavirus OC43, Human/drug effects , Humans , Hydroxychloroquine/pharmacology , SARS-CoV-2/drug effects , Vero Cells , Virus Internalization/drug effects , Virus Replication/drug effects
15.
mSphere ; 6(2)2021 04 28.
Article in English | MEDLINE | ID: covidwho-1207482

ABSTRACT

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) persists on stainless steel and plastic for up to 7 days, suggesting that coronavirus disease 2019 (COVID-19) could be spread by fomite transmission. There is limited research on the stability of SARS-CoV-2 on textiles, with the risk of textiles acting as fomites not being well understood. To date, there does not appear to be any published research on the stability of coronaviruses during laundering, which is required to determine the efficacy of current laundering policies in the decontamination of health care textiles. The aim of this study was to investigate the environmental stability of human coronaviruses HCoV-OC43 and HCoV-229E on different textile fiber types and the persistence of HCoV-OC43 on textiles during domestic and industrial laundering. This study demonstrated that human coronaviruses (5 log10 50% tissue culture infective doses [TCID50]) remain infectious on polyester for ≥72 h, cotton for ≥24 h, and polycotton for ≥6 h; HCoV-OC43 was also able to transfer from polyester to PVC or polyester after 72 h. Under clean conditions, HCoV-OC43 was not detectable on cotton swatches laundered with industrial and domestic wash cycles without temperature and detergent (≥4.57-log10-TCID50 reduction), suggesting that the dilution and agitation of wash cycles are sufficient to remove human coronaviruses from textiles. In the presence of interfering substances (artificial saliva), ≤1.78 log10 TCID50 HCoV-OC43 was detected after washing domestically without temperature and detergent, unlike industrial laundering, where the virus was completely removed. However, no infectious HCoV-OC43 was detected when washed domestically with detergent.IMPORTANCE Synthetic textiles such as polyester could potentially act as fomites of human coronaviruses, indicating the importance of infection control procedures during handling of contaminated textiles prior to laundering. This study provides novel evidence that human coronaviruses can persist on textiles for up to 3 days and are readily transferred from polyester textile to other surfaces after 72 h of incubation. This is of particular importance for the domestic laundering of contaminated textiles such as health care uniforms in the United Kingdom and United States, where there may be a risk of cross-contaminating the domestic environment. It was demonstrated that human coronaviruses are removed from contaminated textiles by typical domestic and commercial wash cycles, even at low temperatures without detergent, indicating that current health care laundering policies are likely sufficient in the decontamination of SARS-CoV-2 from textiles.


Subject(s)
COVID-19/transmission , Common Cold/transmission , Coronavirus 229E, Human/drug effects , Coronavirus OC43, Human/drug effects , Detergents/pharmacology , Textiles/virology , Cell Line , Cotton Fiber/virology , Fomites/virology , Humans , Laundering , Polyesters , SARS-CoV-2/drug effects
16.
J Med Virol ; 93(3): 1792-1795, 2021 03.
Article in English | MEDLINE | ID: covidwho-1196499

ABSTRACT

BACKGROUND: The coronavirus disease 2019 (COVID-19) pandemic calls for effective and safe treatments. Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) causing COVID-19 actively replicates in the throat, unlike SARS-CoV, and shows high pharyngeal viral shedding even in patients with mild symptoms of the disease. HCoV-229E is one of four coronaviruses causing the common cold. In this study, the efficacy of ColdZyme® (CZ-MD), a medical device mouth spray, was tested against SARS-CoV-2 and HCoV-229E in vitro. The CZ-MD provides a protective glycerol barrier containing cod trypsin as an ancillary component. Combined, these ingredients can inactivate common cold viruses in the throat and mouth. The CZ-MD is believed to act on the viral surface proteins that would perturb their entry pathway into cells. The efficacy and safety of the CZ-MD have been demonstrated in clinical trials on the common cold. METHOD OF STUDY: The ability of the CZ-MD to inactivate SARS-CoV-2 and HCoV-229E was tested using an in vitro virucidal suspension test (ASTM E1052). RESULTS: CZ-MD inactivated SARS-CoV-2 by 98.3% and HCoV-229E by 99.9%. CONCLUSION: CZ-MD mouth spray can inactivate the respiratory coronaviruses SARS-CoV-2 and HCoV-229E in vitro. Although the in vitro results presented cannot be directly translated into clinical efficacy, the study indicates that CZ-MD might offer a protective barrier against SARS-CoV-2 and a decreased risk of COVID-19 transmission.


Subject(s)
Antiviral Agents/pharmacology , Coronavirus 229E, Human/drug effects , Glycerol/pharmacology , SARS-CoV-2/drug effects , Trypsin/pharmacology , Virus Inactivation/drug effects , COVID-19/drug therapy , COVID-19/prevention & control , COVID-19/transmission , Common Cold/drug therapy , Common Cold/prevention & control , Common Cold/transmission , Disinfectants/pharmacology , Humans , Viral Proteins/drug effects
17.
J Med Chem ; 64(9): 5632-5644, 2021 05 13.
Article in English | MEDLINE | ID: covidwho-1193564

ABSTRACT

To develop antiviral therapeutics against human coronavirus (HCoV) infections, suitable coronavirus drug targets and corresponding lead molecules must be urgently identified. Here, we describe the discovery of a class of HCoV inhibitors acting on nsp15, a hexameric protein component of the viral replication-transcription complexes, endowed with immune evasion-associated endoribonuclease activity. Structure-activity relationship exploration of these 1,2,3-triazolo-fused betulonic acid derivatives yielded lead molecule 5h as a strong inhibitor (antiviral EC50: 0.6 µM) of HCoV-229E replication. An nsp15 endoribonuclease active site mutant virus was markedly less sensitive to 5h, and selected resistance to the compound mapped to mutations in the N-terminal part of HCoV-229E nsp15, at an interface between two nsp15 monomers. The biological findings were substantiated by the nsp15 binding mode for 5h, predicted by docking. Hence, besides delivering a distinct class of inhibitors, our study revealed a druggable pocket in the nsp15 hexamer with relevance for anti-coronavirus drug development.


Subject(s)
Antiviral Agents/pharmacology , Coronavirus 229E, Human/drug effects , Coronavirus 229E, Human/enzymology , Endoribonucleases/antagonists & inhibitors , Enzyme Inhibitors/pharmacology , Oleanolic Acid/analogs & derivatives , Viral Nonstructural Proteins/antagonists & inhibitors , Virus Replication/drug effects , Antiviral Agents/chemical synthesis , Antiviral Agents/chemistry , Cell Line , Dose-Response Relationship, Drug , Endoribonucleases/metabolism , Enzyme Inhibitors/chemical synthesis , Enzyme Inhibitors/chemistry , Humans , Microbial Sensitivity Tests , Models, Molecular , Oleanolic Acid/chemical synthesis , Oleanolic Acid/chemistry , Oleanolic Acid/pharmacology , Viral Nonstructural Proteins/metabolism
18.
Viruses ; 13(3)2021 03 23.
Article in English | MEDLINE | ID: covidwho-1154525

ABSTRACT

The emergent human coronavirus SARS-CoV-2 and its high infectivity rate has highlighted the strong need for new disinfection systems. Evidence has proven that airborne transmission is an important route of spreading for this virus. Therefore, this short communication introduces CLODOS Technology®, a novel strategy to disinfect contaminated surfaces. It is a product based on stable and 99% pure chlorine dioxide, already certified as a bactericide, fungicide and virucide against different pathogens. In this study, CLODOS Technology®, by direct contact or thermonebulization, showed virucidal activity against the human coronavirus HCoV-229E at non-cytotoxic doses. Different conditions such as nebulization, exposure time and product concentration have been tested to standardize and optimize this new feasible method for disinfection.


Subject(s)
Coronavirus 229E, Human/drug effects , Disinfectants/pharmacology , Disinfection/methods , Cell Line , Chlorine Compounds/analysis , Chlorine Compounds/pharmacology , Coronavirus Infections/prevention & control , Coronavirus Infections/virology , Disinfectants/analysis , Disinfection/instrumentation , Humans , Nebulizers and Vaporizers , Oxides/analysis , Oxides/pharmacology
19.
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
20.
Viruses ; 13(2)2021 02 23.
Article in English | MEDLINE | ID: covidwho-1100154

ABSTRACT

A novel coronavirus, severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), emerged in China at the end of 2019 causing a large global outbreak. As treatments are of the utmost importance, drug repurposing embodies a rich and rapid drug discovery landscape, where candidate drug compounds could be identified and optimized. To this end, we tested seven compounds for their ability to reduce replication of human coronavirus (HCoV)-229E, another member of the coronavirus family. Among these seven drugs tested, four of them, namely rapamycin, disulfiram, loperamide and valproic acid, were highly cytotoxic and did not warrant further testing. In contrast, we observed a reduction of the viral titer by 80% with resveratrol (50% effective concentration (EC50) = 4.6 µM) and lopinavir/ritonavir (EC50 = 8.8 µM) and by 60% with chloroquine (EC50 = 5 µM) with very limited cytotoxicity. Among these three drugs, resveratrol was less cytotoxic (cytotoxic concentration 50 (CC50) = 210 µM) than lopinavir/ritonavir (CC50 = 102 µM) and chloroquine (CC50 = 67 µM). Thus, among the seven drugs tested against HCoV-229E, resveratrol demonstrated the optimal antiviral response with low cytotoxicity with a selectivity index (SI) of 45.65. Similarly, among the three drugs with an anti-HCoV-229E activity, namely lopinavir/ritonavir, chloroquine and resveratrol, only the latter showed a reduction of the viral titer on SARS-CoV-2 with reduced cytotoxicity. This opens the door to further evaluation to fight Covid-19.


Subject(s)
Antiviral Agents/pharmacology , Coronavirus 229E, Human/drug effects , Resveratrol/pharmacology , Ritonavir/pharmacology , SARS-CoV-2/drug effects , Virus Replication/drug effects , Cell Line , Chloroquine/pharmacology , Coronavirus 229E, Human/physiology , Drug Repositioning , Humans , Lopinavir/pharmacology , Male , SARS-CoV-2/physiology , Viral Load
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