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1.
EBioMedicine ; 82: 104148, 2022 Aug.
Article in English | MEDLINE | ID: covidwho-1991004

ABSTRACT

BACKGROUND: To address the emergence of SARS-CoV-2, multiple clinical trials in humans were rapidly started, including those involving an oral treatment by nitazoxanide, despite no or limited pre-clinical evidence of antiviral efficacy. METHODS: In this work, we present a complete pre-clinical evaluation of the antiviral activity of nitazoxanide against SARS-CoV-2. FINDINGS: First, we confirmed the in vitro efficacy of nitazoxanide and tizoxanide (its active metabolite) against SARS-CoV-2. Then, we demonstrated nitazoxanide activity in a reconstructed bronchial human airway epithelium model. In a SARS-CoV-2 virus challenge model in hamsters, oral and intranasal treatment with nitazoxanide failed to impair viral replication in commonly affected organs. We hypothesized that this could be due to insufficient diffusion of the drug into organs of interest. Indeed, our pharmacokinetic study confirmed that concentrations of tizoxanide in organs of interest were always below the in vitro EC50. INTERPRETATION: These preclinical results suggest, if directly applicable to humans, that the standard formulation and dosage of nitazoxanide is not effective in providing antiviral therapy for Covid-19. FUNDING: This work was supported by the Fondation de France "call FLASH COVID-19", project TAMAC, by "Institut national de la santé et de la recherche médicale" through the REACTing (REsearch and ACTion targeting emerging infectious diseases), by REACTING/ANRS MIE under the agreement No. 21180 ('Activité des molécules antivirales dans le modèle hamster'), by European Virus Archive Global (EVA 213 GLOBAL) funded by the European Union's Horizon 2020 research and innovation program under grant agreement No. 871029 and DNDi under support by the Wellcome Trust Grant ref: 222489/Z/21/Z through the COVID-19 Therapeutics Accelerator".


Subject(s)
COVID-19 , SARS-CoV-2 , Animals , Antiviral Agents/pharmacology , Antiviral Agents/therapeutic use , COVID-19/drug therapy , Cricetinae , Humans , Nitro Compounds , Thiazoles
2.
Microorganisms ; 10(8):1639, 2022.
Article in English | MDPI | ID: covidwho-1987897

ABSTRACT

In the absence of drugs to treat or prevent COVID-19, drug repurposing can be a valuable strategy. Despite a substantial number of clinical trials, drug repurposing did not deliver on its promise. While success was observed with some repurposed drugs (e.g., remdesivir, dexamethasone, tocilizumab, baricitinib), others failed to show clinical efficacy. One reason is the lack of clear translational processes based on adequate preclinical profiling before clinical evaluation. Combined with limitations of existing in vitro and in vivo models, there is a need for a systematic approach to urgent antiviral drug development in the context of a global pandemic. We implemented a methodology to test repurposed and experimental drugs to generate robust preclinical evidence for further clinical development. This translational drug development platform comprises in vitro, ex vivo, and in vivo models of SARS-CoV-2, along with pharmacokinetic modeling and simulation approaches to evaluate exposure levels in plasma and target organs. Here, we provide examples of identified repurposed antiviral drugs tested within our multidisciplinary collaboration to highlight lessons learned in urgent antiviral drug development during the COVID-19 pandemic. Our data confirm the importance of assessing in vitro and in vivo potency in multiple assays to boost the translatability of pre-clinical data. The value of pharmacokinetic modeling and simulations for compound prioritization is also discussed. We advocate the need for a standardized translational drug development platform for mild-to-moderate COVID-19 to generate preclinical evidence in support of clinical trials. We propose clear prerequisites for progression of drug candidates for repurposing into clinical trials. Further research is needed to gain a deeper understanding of the scope and limitations of the presented translational drug development platform.

3.
Sci Rep ; 12(1): 12609, 2022 Jul 23.
Article in English | MEDLINE | ID: covidwho-1956425

ABSTRACT

The replacement of the Omicron BA.1 variant of SARS-CoV-2 by the BA.2 and the rapid growth of the BA.5 sub lineage, which have both different sets of mutations in the spike glycoprotein, alters the spectrum of activity of therapeutic antibodies currently licensed in the European Union. Using clinical strains of the Omicron BA.2 and BA.5 variants, we compared the neutralising power of monoclonal antibodies against the Omicron BA.1, BA.2 and BA.5 variants, using an ancestral strain (lineage B.1, D614G) and a Delta variant strain as reference. Sotrovimab/Vir-7831 is less active against BA.2 than against BA.1 (fold change reduction ~ 1,4) and even less active against BA.5 (fold change reduction ~ 2.7). Within the Evusheld /AZD7442 cocktail, Cilgavimab/AZD1061 is more active against BA.2 and BA.5 than against BA.1 (fold change increase ~ 32), whilst the very low activity of Tixagevimab/AZD8895 against BA.1 is not enhanced against BA.2 nor BA.5. In total, compared to BA.1, the activity of the Evusheld/AZD7442 is significantly improved against BA.2 while BA.5 is intermediate but closer to BA.2.


Subject(s)
COVID-19 , Spike Glycoprotein, Coronavirus , Antibodies, Monoclonal , Antibodies, Monoclonal, Humanized , Antibodies, Neutralizing , Antibodies, Viral , COVID-19/drug therapy , Drug Combinations , Humans , SARS-CoV-2/genetics , Spike Glycoprotein, Coronavirus/genetics
4.
EuropePMC; 2022.
Preprint in English | EuropePMC | ID: ppcovidwho-329966

ABSTRACT

The replacement of the Omicron BA.1 variant of SARS-CoV-2 by the BA.2 sub lineage, which has a different set of mutations in the spike glycoprotein, alters the spectrum of activity of therapeutic antibodies. In this study, we compared the neutralising power of monoclonal antibodies against the Omicron BA.1 and BA.2 variants, with an ancestral strain (D614G) and a Delta variant as reference. Sotrovimab is less active against BA.2 than against BA.1 (fold change reduction ~1,5). Within the Evusheld/ AZD744 cocktail, Cilgavimab is more active against BA.2 than against BA.1 (fold change increase ~32), whilst activity of Tixagevimab remains very low. In total, the activity of Evusheld is significantly improved (fold change increase ~10 compared to BA.1).

5.
Commun Biol ; 5(1): 225, 2022 03 10.
Article in English | MEDLINE | ID: covidwho-1740485

ABSTRACT

Late 2020, SARS-CoV-2 Alpha variant emerged in United Kingdom and gradually replaced G614 strains initially involved in the global spread of the pandemic. In this study, we use a Syrian hamster model to compare a clinical strain of Alpha variant with an ancestral G614 strain. The Alpha variant succeed to infect animals and to induce a pathology that mimics COVID-19. However, both strains replicate to almost the same level and induced a comparable disease and immune response. A slight fitness advantage is noted for the G614 strain during competition and transmission experiments. These data do not corroborate the epidemiological situation observed during the first half of 2021 in humans nor reports that showed a more rapid replication of Alpha variant in human reconstituted bronchial epithelium. This study highlights the need to combine data from different laboratories using various animal models to decipher the biological properties of newly emerging SARS-CoV-2 variants.


Subject(s)
COVID-19 , Disease Models, Animal , Mesocricetus , SARS-CoV-2/physiology , Animals , Antibodies, Neutralizing/blood , COVID-19/blood , COVID-19/immunology , COVID-19/virology , Cytokines/genetics , Female , Gastrointestinal Tract/virology , Genome, Viral , Lung/virology , Nasal Lavage Fluid/virology , SARS-CoV-2/genetics , Virus Replication
6.
Viruses ; 12(6)2020 06 25.
Article in English | MEDLINE | ID: covidwho-1726024

ABSTRACT

The recent emergence of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) worldwide has highlighted the importance of reliable and rapid diagnostic testing to prevent and control virus circulation. Dozens of monoplex in-house RT-qPCR assays are already available; however, the development of dual-target assays is suited to avoid false-negative results caused by polymorphisms or point mutations, that can compromise the accuracy of diagnostic and screening tests. In this study, two mono-target assays recommended by WHO (E-Sarbeco (enveloppe gene, Charite University, Berlin, Germany) and RdRp-IP4 (RdRp, Institut Pasteur, Paris, France)) were selected and combined in a unique robust test; the resulting duo SARS-CoV-2 RT-qPCR assay was compared to the two parental monoplex tests. The duo SARS-CoV-2 assay performed equally, or better, in terms of sensitivity, specificity, linearity and signal intensity. We demonstrated that combining two single systems into a dual-target assay (with or without an MS2-based internal control) did not impair performances, providing a potent tool adapted for routine molecular diagnosis in clinical microbiology laboratories.


Subject(s)
Betacoronavirus/isolation & purification , Coronavirus Infections/diagnosis , Pneumonia, Viral/diagnosis , RNA-Dependent RNA Polymerase/genetics , Real-Time Polymerase Chain Reaction/methods , Viral Envelope Proteins/genetics , Viral Nonstructural Proteins/genetics , Betacoronavirus/genetics , COVID-19 , Coronavirus Envelope Proteins , Coronavirus Infections/virology , Coronavirus RNA-Dependent RNA Polymerase , Humans , Pandemics , Pneumonia, Viral/virology , RNA, Viral/analysis , SARS-CoV-2 , Sensitivity and Specificity , World Health Organization
7.
EMBO Rep ; 23(5): e53820, 2022 05 04.
Article in English | MEDLINE | ID: covidwho-1726972

ABSTRACT

Engineering recombinant viruses is a pre-eminent tool for deciphering the biology of emerging viral pathogens such as the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). However, the large size of coronavirus genomes renders the current reverse genetics methods challenging. Here, we describe a simple method based on "infectious subgenomic amplicons" (ISA) technology to generate recombinant infectious coronaviruses with no need for reconstruction of the complete genomic cDNA and apply this method to SARS-CoV-2 and also to the feline enteric coronavirus. In both cases we rescue wild-type viruses with biological characteristics similar to original strains. Specific mutations and fluorescent red reporter genes can be readily incorporated into the SARS-CoV-2 genome enabling the generation of a genomic variants and fluorescent reporter strains for in vivo experiments, serological diagnosis, and antiviral assays. The swiftness and simplicity of the ISA method has the potential to facilitate the advance of coronavirus reverse genetics studies, to explore the molecular biological properties of the SARS-CoV-2 variants, and to accelerate the development of effective therapeutic reagents.


Subject(s)
COVID-19 , SARS-CoV-2 , Animals , Antiviral Agents , COVID-19/genetics , Cats , Reverse Genetics , SARS-CoV-2/genetics
8.
EuropePMC; 2022.
Preprint in English | EuropePMC | ID: ppcovidwho-329017

ABSTRACT

The efficacy of pre-exposure prophylaxis by the Tixagevimab/Cilgavimab cocktail (AZD7442) was evaluated in hamsters against a clinical BA.1 strain of SARS-CoV-2 variant Omicron. AZD7442 retains inhibitory activity against Omicron despite a substantial loss of efficacy. We estimate that Omicron virus requires about 20-times more antibodies in plasma than the ancestral B.1 strain (G614) virus to achieve a similar drug efficacy in reducing lung infectious titers.

9.
EuropePMC; 2021.
Preprint in English | EuropePMC | ID: ppcovidwho-316871

ABSTRACT

Engineering recombinant viruses is capital for deciphering the biology of emerging viral pathogens such as the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). However, the large size of coronaviruses genome makes reverse genetics methods challenging.Here we describe a simple method based on “infectious subgenomic amplicons” (ISA) technology to generate recombinant infectious coronaviruses with no need for reconstructing a full genomic cDNA. The method was applied to the SARS-CoV-2 and the feline enteric coronavirus, and allowed to rescue wild-type viruses with biological characteristics closely similar to original strains. Mutations and fluorescent red reporter gene were rapidly incorporated into the SARS-CoV-2 genome allowing the generation of a genomic variant and a fluorescent reporter strains which were studied during in vivo experiments, serological diagnosis and antiviral assays. The swiftness and simplicity of the ISA method has the potential to facilitate the advance of coronavirus reverse genetics studies and to explore biological properties of SARS-CoV-2 variants or accelerating the development of therapeutic measures.

10.
PLoS One ; 16(12): e0260958, 2021.
Article in English | MEDLINE | ID: covidwho-1546973

ABSTRACT

SARS-CoV-2 variants are emerging with potential increased transmissibility highlighting the great unmet medical need for new therapies. Niclosamide is a potent anti-SARS-CoV-2 agent that has advanced in clinical development. We validate the potent antiviral efficacy of niclosamide in a SARS-CoV-2 human airway model. Furthermore, niclosamide remains its potency against the D614G, Alpha (B.1.1.7), Beta (B.1.351), and Delta (B.1.617.2) variants. Our data further support the potent anti-SARS-CoV-2 properties of niclosamide and highlights its great potential as a therapeutic agent for COVID-19.


Subject(s)
Antiviral Agents/therapeutic use , COVID-19/drug therapy , Niclosamide/therapeutic use , SARS-CoV-2/drug effects , Animals , Caco-2 Cells , Chlorocebus aethiops , Humans , Inhibitory Concentration 50 , Respiratory Mucosa/virology , Vero Cells
11.
Antiviral Res ; 197: 105212, 2022 01.
Article in English | MEDLINE | ID: covidwho-1530602

ABSTRACT

Drug repositioning has been used extensively since the beginning of the COVID-19 pandemic in an attempt to identify antiviral molecules for use in human therapeutics. Hydroxychloroquine and azithromycin have shown inhibitory activity against SARS-CoV-2 replication in different cell lines. Based on such in vitro data and despite the weakness of preclinical assessment, many clinical trials were set up using these molecules. In the present study, we show that hydroxychloroquine and azithromycin alone or combined does not block SARS-CoV-2 replication in human bronchial airway epithelia. When tested in a Syrian hamster model, hydroxychloroquine and azithromycin administrated alone or combined displayed no significant effect on viral replication, clinical course of the disease and lung impairments, even at high doses. Hydroxychloroquine quantification in lung tissues confirmed strong exposure to the drug, above in vitro inhibitory concentrations. Overall, this study does not support the use of hydroxychloroquine and azithromycin as antiviral drugs for the treatment of SARS-CoV-2 infections.


Subject(s)
Anti-Infective Agents/pharmacology , Azithromycin/pharmacology , COVID-19/drug therapy , Hydroxychloroquine/pharmacology , SARS-CoV-2/drug effects , Animals , Anti-Infective Agents/administration & dosage , Anti-Infective Agents/therapeutic use , Azithromycin/administration & dosage , Azithromycin/pharmacokinetics , Azithromycin/therapeutic use , Bronchi/cytology , Bronchi/virology , Chlorocebus aethiops , Cricetinae , Disease Models, Animal , Drug Therapy, Combination , Female , Humans , Hydroxychloroquine/administration & dosage , Hydroxychloroquine/therapeutic use , Lung/pathology , Mesocricetus , Middle Aged , Plasma/virology , Real-Time Polymerase Chain Reaction , Vero Cells
12.
Viruses ; 13(9)2021 09 12.
Article in English | MEDLINE | ID: covidwho-1411086

ABSTRACT

Our therapeutic arsenal against viruses is very limited and the current pandemic of SARS-CoV-2 highlights the critical need for effective antivirals against emerging coronaviruses. Cellular assays allowing a precise quantification of viral replication in high-throughput experimental settings are essential to the screening of chemical libraries and the selection of best antiviral chemical structures. To develop a reporting system for SARS-CoV-2 infection, we generated cell lines expressing a firefly luciferase maintained in an inactive form by a consensus cleavage site for the viral protease 3CLPro of coronaviruses, so that the luminescent biosensor is turned on upon 3CLPro expression or SARS-CoV-2 infection. This cellular assay was used to screen a metabolism-oriented library of 492 compounds to identify metabolic vulnerabilities of coronaviruses for developing innovative therapeutic strategies. In agreement with recent reports, inhibitors of pyrimidine biosynthesis were found to prevent SARS-CoV-2 replication. Among the top hits, we also identified the NADPH oxidase (NOX) inhibitor Setanaxib. The anti-SARS-CoV-2 activity of Setanaxib was further confirmed using ACE2-expressing human pulmonary cells Beas2B as well as human primary nasal epithelial cells. Altogether, these results validate our cell-based functional assay and the interest of screening libraries of different origins to identify inhibitors of SARS-CoV-2 for drug repurposing or development.


Subject(s)
Antiviral Agents/isolation & purification , Biosensing Techniques/methods , Coronavirus 3C Proteases/metabolism , SARS-CoV-2/physiology , Virus Replication , Animals , Antiviral Agents/pharmacology , Cell Line , Chlorocebus aethiops , Drug Discovery , Drug Evaluation, Preclinical , Enzyme Activation , HEK293 Cells , Humans , Luciferases, Firefly/metabolism , Nasal Mucosa/virology , Pyrazolones/pharmacology , Pyridones/pharmacology , SARS-CoV-2/metabolism , Vero Cells , Virus Internalization/drug effects , Virus Replication/drug effects
13.
Nat Commun ; 12(1): 1735, 2021 03 19.
Article in English | MEDLINE | ID: covidwho-1387332

ABSTRACT

Despite no or limited pre-clinical evidence, repurposed drugs are massively evaluated in clinical trials to palliate the lack of antiviral molecules against SARS-CoV-2. Here we use a Syrian hamster model to assess the antiviral efficacy of favipiravir, understand its mechanism of action and determine its pharmacokinetics. When treatment is initiated before or simultaneously to infection, favipiravir has a strong dose effect, leading to reduction of infectious titers in lungs and clinical alleviation of the disease. Antiviral effect of favipiravir correlates with incorporation of a large number of mutations into viral genomes and decrease of viral infectivity. Antiviral efficacy is achieved with plasma drug exposure comparable with those previously found during human clinical trials. Notably, the highest dose of favipiravir tested is associated with signs of toxicity in animals. Thereby, pharmacokinetic and tolerance studies are required to determine whether similar effects can be safely achieved in humans.


Subject(s)
Amides/pharmacology , Antiviral Agents/pharmacology , COVID-19/drug therapy , Pyrazines/pharmacology , SARS-CoV-2/drug effects , Animals , COVID-19/virology , Chlorocebus aethiops , Cricetinae , Disease Models, Animal , Female , Genome, Viral , Lung/virology , Mesocricetus , SARS-CoV-2/genetics , Vero Cells , Viral Load/drug effects
14.
Antiviral Res ; 193: 105137, 2021 09.
Article in English | MEDLINE | ID: covidwho-1306846

ABSTRACT

Following the emergence of SARS-CoV-2, the search for an effective and rapidly available treatment was initiated worldwide based on repurposing of available drugs. Previous reports described the antiviral activity of certain tyrosine kinase inhibitors (TKIs) targeting the Abelson kinase 2 against pathogenic coronaviruses. Imatinib, one of them, has more than twenty years of safe utilization for the treatment of hematological malignancies. In this context, Imatinib was rapidly evaluated in clinical trials against Covid-19. Here, we present the pre-clinical evaluation of imatinib in multiple models. Our results indicated that imatinib and another TKI, the masitinib, exhibit an antiviral activity in VeroE6 cells. However, imatinib was inactive in a reconstructed bronchial human airway epithelium model. In vivo, imatinib therapy failed to impair SARS-CoV-2 replication in a golden Syrian hamster model despite high concentrations in plasma and in the lung. Overall, these results do not support the use of imatinib and similar TKIs as antivirals in the treatment of Covid-19.


Subject(s)
Antiviral Agents/pharmacology , COVID-19/drug therapy , Imatinib Mesylate/pharmacology , SARS-CoV-2/drug effects , Animals , COVID-19/epidemiology , COVID-19/virology , Cell Line , Chlorocebus aethiops , Drug Evaluation, Preclinical , Drug Repositioning , Enzyme Inhibitors/pharmacology , Epithelium , Female , Humans , Lung/pathology , Male , Mesocricetus , Vero Cells , Virus Replication/drug effects
15.
mBio ; 12(4): e0085021, 2021 08 31.
Article in English | MEDLINE | ID: covidwho-1297961

ABSTRACT

Since its emergence in 2019, circulating populations of the new coronavirus (CoV) continuously acquired genetic diversity. At the end of 2020, a variant named 20I/501Y.V1 (lineage B.1.1.7) emerged and replaced other circulating strains in several regions. This phenomenon has been poorly associated with biological evidence that this variant and the original strain exhibit different phenotypic characteristics. Here, we analyze the replication ability of this new variant in different cellular models using for comparison an ancestral D614G European strain (lineage B1). Results from comparative replication kinetics experiments in vitro and in a human reconstituted bronchial epithelium showed no difference. However, when both viruses were put in competition in human reconstituted bronchial epithelium, the 20I/501Y.V1 variant outcompeted the ancestral strain. All together, these findings demonstrate that this new variant replicates more efficiently and may contribute to a better understanding of the progressive replacement of circulating strains by the severe acute respiratory CoV-2 (SARS-CoV-2) 20I/501Y.V1 variant. IMPORTANCE The emergence of several SARS-CoV-2 variants raised numerous questions concerning the future course of the pandemic. We are currently observing a replacement of the circulating viruses by the variant from the United Kingdom known as 20I/501Y.V1, from the B.1.1.7 lineage, but there is little biological evidence that this new variant exhibits a different phenotype. In the present study, we used different cellular models to assess the replication ability of the 20I/501Y.V1 variant. Our results showed that this variant replicates more efficiently in human reconstituted bronchial epithelium, which may explain why it spreads so rapidly in human populations.


Subject(s)
COVID-19/transmission , Genetic Fitness/genetics , SARS-CoV-2/growth & development , SARS-CoV-2/genetics , Virus Replication/genetics , Animals , COVID-19/pathology , Caco-2 Cells , Cell Line , Chlorocebus aethiops , Humans , Respiratory Mucosa/virology , Vero Cells , Viral Load
17.
Sci Rep ; 10(1): 13093, 2020 08 04.
Article in English | MEDLINE | ID: covidwho-697117

ABSTRACT

A novel coronavirus, named SARS-CoV-2, emerged in 2019 in China and rapidly spread worldwide. As no approved therapeutics exists to treat COVID-19, the disease associated to SARS-Cov-2, there is an urgent need to propose molecules that could quickly enter into clinics. Repurposing of approved drugs is a strategy that can bypass the time-consuming stages of drug development. In this study, we screened the PRESTWICK CHEMICAL LIBRARY composed of 1,520 approved drugs in an infected cell-based assay. The robustness of the screen was assessed by the identification of drugs that already demonstrated in vitro antiviral effect against SARS-CoV-2. Thereby, 90 compounds were identified as positive hits from the screen and were grouped according to their chemical composition and their known therapeutic effect. Then EC50 and CC50 were determined for a subset of 15 compounds from a panel of 23 selected drugs covering the different groups. Eleven compounds such as macrolides antibiotics, proton pump inhibitors, antiarrhythmic agents or CNS drugs emerged showing antiviral potency with 2 < EC50 ≤ 20 µM. By providing new information on molecules inhibiting SARS-CoV-2 replication in vitro, this study provides information for the selection of drugs to be further validated in vivo. Disclaimer: This study corresponds to the early stages of antiviral development and the results do not support by themselves the use of the selected drugs to treat SARS-CoV-2 infection.


Subject(s)
Betacoronavirus/physiology , Small Molecule Libraries/chemistry , Animals , Antiviral Agents/chemistry , Antiviral Agents/metabolism , Antiviral Agents/pharmacology , Betacoronavirus/isolation & purification , COVID-19 , Caco-2 Cells , Cell Survival/drug effects , Chlorocebus aethiops , Coronavirus Infections/pathology , Coronavirus Infections/virology , Drug Approval , Drug Evaluation, Preclinical , Drug Repositioning , Humans , Pandemics , Pneumonia, Viral/pathology , Pneumonia, Viral/virology , SARS-CoV-2 , Small Molecule Libraries/metabolism , Small Molecule Libraries/pharmacology , Vero Cells , Virus Replication/drug effects
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