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2.
J Infect Dis ; 224(3): 415-419, 2021 08 02.
Article in English | MEDLINE | ID: covidwho-1526165

ABSTRACT

Mutagenic ribonucleosides can act as broad-based antiviral agents. They are metabolized to the active ribonucleoside triphosphate form and concentrate in genomes of RNA viruses during viral replication. ß-d-N4-hydroxycytidine (NHC, initial metabolite of molnupiravir) is >100-fold more active than ribavirin or favipiravir against severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), with antiviral activity correlated to the level of mutagenesis in virion RNA. However, NHC also displays host mutational activity in an animal cell culture assay, consistent with RNA and DNA precursors sharing a common intermediate of a ribonucleoside diphosphate. These results indicate highly active mutagenic ribonucleosides may hold risk for the host.


Subject(s)
Antiviral Agents/pharmacology , Cytidine/analogs & derivatives , Mutagens/pharmacology , SARS-CoV-2/drug effects , Animals , Antiviral Agents/adverse effects , CHO Cells/drug effects , Cells, Cultured , Cricetulus , Cytidine/adverse effects , Cytidine/pharmacology , Dose-Response Relationship, Drug , Mutagenesis/drug effects , Mutagens/adverse effects , SARS-CoV-2/genetics , Virus Replication/drug effects
3.
Cell ; 184(21): 5432-5447.e16, 2021 10 14.
Article in English | MEDLINE | ID: covidwho-1454060

ABSTRACT

Understanding vaccine-elicited protection against SARS-CoV-2 variants and other sarbecoviruses is key for guiding public health policies. We show that a clinical stage multivalent SARS-CoV-2 spike receptor-binding domain nanoparticle (RBD-NP) vaccine protects mice from SARS-CoV-2 challenge after a single immunization, indicating a potential dose-sparing strategy. We benchmarked serum neutralizing activity elicited by RBD-NPs in non-human primates against a lead prefusion-stabilized SARS-CoV-2 spike (HexaPro) using a panel of circulating mutants. Polyclonal antibodies elicited by both vaccines are similarly resilient to many RBD residue substitutions tested, although mutations at and surrounding position 484 have negative consequences for neutralization. Mosaic and cocktail nanoparticle immunogens displaying multiple sarbecovirus RBDs elicit broad neutralizing activity in mice and protect mice against SARS-CoV challenge even in the absence of SARS-CoV RBD in the vaccine. This study provides proof of principle that multivalent sarbecovirus RBD-NPs induce heterotypic protection and motivates advancing such broadly protective sarbecovirus vaccines to the clinic.

4.
mBio ; 12(4): e0157221, 2021 08 31.
Article in English | MEDLINE | ID: covidwho-1349194

ABSTRACT

Tissue- and cell-specific expression patterns are highly variable within and across individuals, leading to altered host responses after acute virus infection. Unraveling key tissue-specific response patterns provides novel opportunities for defining fundamental mechanisms of virus-host interaction in disease and the identification of critical tissue-specific networks for disease intervention in the lung. Currently, there are no approved therapeutics for Middle East respiratory syndrome coronavirus (MERS-CoV) patients, and little is understood about how lung cell types contribute to disease outcomes. MERS-CoV replicates equivalently in primary human lung microvascular endothelial cells (MVE) and fibroblasts (FB) and to equivalent peak titers but with slower replication kinetics in human airway epithelial cell cultures (HAE). However, only infected MVE demonstrate observable virus-induced cytopathic effect. To explore mechanisms leading to reduced MVE viability, donor-matched human lung MVE, HAE, and FB were infected, and their transcriptomes, proteomes, and lipidomes were monitored over time. Validated functional enrichment analysis demonstrated that MERS-CoV-infected MVE were dying via an unfolded protein response (UPR)-mediated apoptosis. Pharmacologic manipulation of the UPR in MERS-CoV-infected primary lung cells reduced viral titers and in male mice improved respiratory function with accompanying reductions in weight loss, pathological signatures of acute lung injury, and times to recovery. Systems biology analysis and validation studies of global kinetic transcript, protein, and lipid data sets confirmed that inhibition of host stress pathways that are differentially regulated following MERS-CoV infection of different tissue types can alleviate symptom progression to end-stage lung disease commonly seen following emerging coronavirus outbreaks. IMPORTANCE Middle East respiratory syndrome coronavirus (MERS-CoV) causes severe atypical pneumonia in infected individuals, but the underlying mechanisms of pathogenesis remain unknown. While much has been learned from the few reported autopsy cases, an in-depth understanding of the cells targeted by MERS-CoV in the human lung and their relative contribution to disease outcomes is needed. The host response in MERS-CoV-infected primary human lung microvascular endothelial (MVE) cells and fibroblasts (FB) was evaluated over time by analyzing total RNA, proteins, and lipids to determine the cellular pathways modulated postinfection. Findings revealed that MERS-CoV-infected MVE cells die via apoptotic mechanisms downstream of the unfolded protein response (UPR). Interruption of enzymatic processes within the UPR in MERS-CoV-infected male mice reduced disease symptoms, virus-induced lung injury, and time to recovery. These data suggest that the UPR plays an important role in MERS-CoV infection and may represent a host target for therapeutic intervention.


Subject(s)
Acute Lung Injury/pathology , Apoptosis/physiology , Coronavirus Infections/pathology , Unfolded Protein Response/physiology , Acute Lung Injury/virology , Animals , Cell Line , Endothelial Cells/metabolism , Endothelial Cells/virology , Female , Fibroblasts/metabolism , Fibroblasts/virology , Humans , Male , Mice , Middle East Respiratory Syndrome Coronavirus/immunology
5.
Nature ; 599(7885): 465-470, 2021 11.
Article in English | MEDLINE | ID: covidwho-1428880

ABSTRACT

Monoclonal antibodies with neutralizing activity against SARS-CoV-2 have demonstrated clinical benefits in cases of mild-to-moderate SARS-CoV-2 infection, substantially reducing the risk for hospitalization and severe disease1-4. Treatment generally requires the administration of high doses of these monoclonal antibodies and has limited efficacy in preventing disease complications or mortality among hospitalized patients with COVID-195. Here we report the development and evaluation of anti-SARS-CoV-2 monoclonal antibodies with optimized Fc domains that show superior potency for prevention or treatment of COVID-19. Using several animal disease models of COVID-196,7, we demonstrate that selective engagement of activating Fcγ receptors results in improved efficacy in both preventing and treating disease-induced weight loss and mortality, significantly reducing the dose required to confer full protection against SARS-CoV-2 challenge and for treatment of pre-infected animals. Our results highlight the importance of Fcγ receptor pathways in driving antibody-mediated antiviral immunity and exclude the possibility of pathogenic or disease-enhancing effects of Fcγ receptor engagement of anti-SARS-CoV-2 antibodies upon infection. These findings have important implications for the development of Fc-engineered monoclonal antibodies with optimal Fc-effector function and improved clinical efficacy against COVID-19 disease.

6.
Mol Cell ; 80(6): 1078-1091.e6, 2020 12 17.
Article in English | MEDLINE | ID: covidwho-1386333

ABSTRACT

We report that the SARS-CoV-2 nucleocapsid protein (N-protein) undergoes liquid-liquid phase separation (LLPS) with viral RNA. N-protein condenses with specific RNA genomic elements under physiological buffer conditions and condensation is enhanced at human body temperatures (33°C and 37°C) and reduced at room temperature (22°C). RNA sequence and structure in specific genomic regions regulate N-protein condensation while other genomic regions promote condensate dissolution, potentially preventing aggregation of the large genome. At low concentrations, N-protein preferentially crosslinks to specific regions characterized by single-stranded RNA flanked by structured elements and these features specify the location, number, and strength of N-protein binding sites (valency). Liquid-like N-protein condensates form in mammalian cells in a concentration-dependent manner and can be altered by small molecules. Condensation of N-protein is RNA sequence and structure specific, sensitive to human body temperature, and manipulatable with small molecules, and therefore presents a screenable process for identifying antiviral compounds effective against SARS-CoV-2.


Subject(s)
COVID-19/metabolism , Coronavirus Nucleocapsid Proteins/metabolism , Genome, Viral , Nucleocapsid/metabolism , RNA, Viral/metabolism , SARS-CoV-2/metabolism , Animals , Antiviral Agents/pharmacology , COVID-19/drug therapy , COVID-19/genetics , Chlorocebus aethiops , Coronavirus Nucleocapsid Proteins/genetics , Drug Evaluation, Preclinical , HEK293 Cells , Humans , Nucleocapsid/genetics , Phosphoproteins/genetics , Phosphoproteins/metabolism , SARS-CoV-2/genetics , Vero Cells
7.
Nat Struct Mol Biol ; 28(9): 747-754, 2021 09.
Article in English | MEDLINE | ID: covidwho-1370728

ABSTRACT

Drug discovery campaigns against COVID-19 are beginning to target the SARS-CoV-2 RNA genome. The highly conserved frameshift stimulation element (FSE), required for balanced expression of viral proteins, is a particularly attractive SARS-CoV-2 RNA target. Here we present a 6.9 Å resolution cryo-EM structure of the FSE (88 nucleotides, ~28 kDa), validated through an RNA nanostructure tagging method. The tertiary structure presents a topologically complex fold in which the 5' end is threaded through a ring formed inside a three-stem pseudoknot. Guided by this structure, we develop antisense oligonucleotides that impair FSE function in frameshifting assays and knock down SARS-CoV-2 virus replication in A549-ACE2 cells at 100 nM concentration.


Subject(s)
COVID-19/prevention & control , Cryoelectron Microscopy/methods , Frameshift Mutation/genetics , Oligonucleotides, Antisense/genetics , RNA, Viral/genetics , Response Elements/genetics , SARS-CoV-2/genetics , A549 Cells , Animals , Base Sequence , COVID-19/virology , Cell Line, Tumor , Chlorocebus aethiops , Genome, Viral/genetics , Humans , Models, Molecular , Nucleic Acid Conformation , Oligonucleotides, Antisense/pharmacology , RNA, Viral/chemistry , RNA, Viral/ultrastructure , SARS-CoV-2/physiology , SARS-CoV-2/ultrastructure , Vero Cells , Virus Replication/drug effects , Virus Replication/genetics
8.
Cell Rep ; 36(4): 109450, 2021 07 27.
Article in English | MEDLINE | ID: covidwho-1306890

ABSTRACT

Improving clinical care for individuals infected with SARS-CoV-2 variants is a global health priority. Small-molecule antivirals like remdesivir (RDV) and biologics such as human monoclonal antibodies (mAbs) have demonstrated therapeutic efficacy against SARS-CoV-2, the causative agent of coronavirus disease 2019 (COVID-19). It is not known whether combination RDV/mAb will improve outcomes over single-agent therapies or whether antibody therapies will remain efficacious against variants. Here, we show that a combination of two mAbs in clinical trials, C144 and C135, have potent antiviral effects against even when initiated 48 h after infection and have therapeutic efficacy in vivo against the B.1.351 variant of concern (VOC). Combining RDV and antibodies provided a modest improvement in outcomes compared with single agents. These data support the continued use of RDV to treat SARS-CoV-2 infections and the continued clinical development of the C144 and C135 antibody combination to treat patients infected with SARS-CoV-2 variants.


Subject(s)
Antibodies, Monoclonal/pharmacology , COVID-19/drug therapy , SARS-CoV-2/drug effects , Animals , Antibodies, Monoclonal/immunology , Antiviral Agents/pharmacology , Humans , Mice , SARS-CoV-2/pathogenicity
9.
BMC Bioinformatics ; 22(1): 287, 2021 May 29.
Article in English | MEDLINE | ID: covidwho-1257920

ABSTRACT

BACKGROUND: Representing biological networks as graphs is a powerful approach to reveal underlying patterns, signatures, and critical components from high-throughput biomolecular data. However, graphs do not natively capture the multi-way relationships present among genes and proteins in biological systems. Hypergraphs are generalizations of graphs that naturally model multi-way relationships and have shown promise in modeling systems such as protein complexes and metabolic reactions. In this paper we seek to understand how hypergraphs can more faithfully identify, and potentially predict, important genes based on complex relationships inferred from genomic expression data sets. RESULTS: We compiled a novel data set of transcriptional host response to pathogenic viral infections and formulated relationships between genes as a hypergraph where hyperedges represent significantly perturbed genes, and vertices represent individual biological samples with specific experimental conditions. We find that hypergraph betweenness centrality is a superior method for identification of genes important to viral response when compared with graph centrality. CONCLUSIONS: Our results demonstrate the utility of using hypergraphs to represent complex biological systems and highlight central important responses in common to a variety of highly pathogenic viruses.


Subject(s)
Algorithms , Models, Biological , Genomics , Proteins
10.
J Infect Dis ; 224(Supplement_1): S1-S21, 2021 Jul 15.
Article in English | MEDLINE | ID: covidwho-1263668

ABSTRACT

The NIH Virtual SARS-CoV-2 Antiviral Summit, held on 6 November 2020, was organized to provide an overview on the status and challenges in developing antiviral therapeutics for coronavirus disease 2019 (COVID-19), including combinations of antivirals. Scientific experts from the public and private sectors convened virtually during a live videocast to discuss severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) targets for drug discovery as well as the preclinical tools needed to develop and evaluate effective small-molecule antivirals. The goals of the Summit were to review the current state of the science, identify unmet research needs, share insights and lessons learned from treating other infectious diseases, identify opportunities for public-private partnerships, and assist the research community in designing and developing antiviral therapeutics. This report includes an overview of therapeutic approaches, individual panel summaries, and a summary of the discussions and perspectives on the challenges ahead for antiviral development.


Subject(s)
Antiviral Agents/therapeutic use , COVID-19/drug therapy , SARS-CoV-2/drug effects , Antiviral Agents/pharmacology , COVID-19/virology , Drug Development , Humans , National Institutes of Health (U.S.) , Peptide Hydrolases/metabolism , Protease Inhibitors/pharmacology , Protease Inhibitors/therapeutic use , United States , Virus Replication/drug effects
11.
J Infect Dis ; 224(3): 415-419, 2021 08 02.
Article in English | MEDLINE | ID: covidwho-1221478

ABSTRACT

Mutagenic ribonucleosides can act as broad-based antiviral agents. They are metabolized to the active ribonucleoside triphosphate form and concentrate in genomes of RNA viruses during viral replication. ß-d-N4-hydroxycytidine (NHC, initial metabolite of molnupiravir) is >100-fold more active than ribavirin or favipiravir against severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), with antiviral activity correlated to the level of mutagenesis in virion RNA. However, NHC also displays host mutational activity in an animal cell culture assay, consistent with RNA and DNA precursors sharing a common intermediate of a ribonucleoside diphosphate. These results indicate highly active mutagenic ribonucleosides may hold risk for the host.


Subject(s)
Antiviral Agents/pharmacology , Cytidine/analogs & derivatives , Mutagens/pharmacology , SARS-CoV-2/drug effects , Animals , Antiviral Agents/adverse effects , CHO Cells/drug effects , Cells, Cultured , Cricetulus , Cytidine/adverse effects , Cytidine/pharmacology , Dose-Response Relationship, Drug , Mutagenesis/drug effects , Mutagens/adverse effects , SARS-CoV-2/genetics , Virus Replication/drug effects
12.
Bio Protoc ; 11(5): e3938, 2021 Mar 05.
Article in English | MEDLINE | ID: covidwho-1162408

ABSTRACT

Next generations sequencing (NGS) has become an important tool in biomedical research. The Primer ID approach combined with the MiSeq platform overcomes the limitation of PCR errors and reveals the true sampling depth of population sequencing, making it an ideal tool to study mutagenic effects of potential broad-spectrum antivirals on RNA viruses. In this report we describe a protocol using Primer ID sequencing to study the mutations induced by antivirals in a coronavirus genome from an in vitro cell culture model and an in vivo mouse model. Viral RNA or total lung tissue RNA is tagged with Primer ID-containing cDNA primers during the initial reverse transcription step, followed by two rounds of PCR to amplify viral sequences and incorporate sequencing adaptors. Purified and pooled libraries are sequenced using the MiSeq platform. Sequencing data are processed using the template consensus sequence (TCS) web-app. The Primer ID approach provides an accurate sequencing protocol to measure mutation error rates in viral RNA genomes and host mRNA. Sequencing results suggested that ß-D-N4-hydroxycytidine (NHC) greatly increased the transition substitution rate but not the transversion substitution rate in the viral RNA genomes, and cytosine (C) to uridine (U) was found as the most frequently seen mutation.

13.
mBio ; 12(2)2021 03 16.
Article in English | MEDLINE | ID: covidwho-1138305

ABSTRACT

The angiotensin-converting enzyme 2 (ACE2) receptor is a major severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) host range determinant, and understanding SARS-CoV-2-ACE2 interactions will provide important insights into COVID-19 pathogenesis and animal model development. SARS-CoV-2 cannot infect mice due to incompatibility between its receptor binding domain and the murine ACE2 receptor. Through molecular modeling and empirical in vitro validation, we identified 5 key amino acid differences between murine and human ACE2 that mediate SARS-CoV-2 infection, generating a chimeric humanized murine ACE2. Additionally, we examined the ability of the humanized murine ACE2 receptor to permit infection by an additional preemergent group 2B coronavirus, WIV-1, providing evidence for the potential pan-virus capabilities of this chimeric receptor. Finally, we predicted the ability of these determinants to inform host range identification of preemergent coronaviruses by evaluating hot spot contacts between SARS-CoV-2 and additional potential host receptors. Our results identify residue determinants that mediate coronavirus receptor usage and host range for application in SARS-CoV-2 and emerging coronavirus animal model development.IMPORTANCE SARS-CoV-2 (the causative agent of COVID-19) is a major public health threat and one of two related coronaviruses that have caused epidemics in modern history. A method of screening potential infectible hosts for preemergent and future emergent coronaviruses would aid in mounting rapid response and intervention strategies during future emergence events. Here, we evaluated determinants of SARS-CoV-2 receptor interactions, identifying key changes that enable or prevent infection. The analysis detailed in this study will aid in the development of model systems to screen emergent coronaviruses as well as treatments to counteract infections.


Subject(s)
Angiotensin-Converting Enzyme 2/chemistry , Angiotensin-Converting Enzyme 2/metabolism , Betacoronavirus/physiology , Virus Replication , Amino Acid Sequence , Angiotensin-Converting Enzyme 2/genetics , Animals , Betacoronavirus/metabolism , Binding Sites , COVID-19/virology , Cell Line , Coronavirus Infections/virology , Host Specificity , Humans , Mice , Models, Molecular , Mutation , Protein Binding , Recombinant Proteins/chemistry , Recombinant Proteins/genetics , Recombinant Proteins/metabolism , SARS-CoV-2/metabolism , SARS-CoV-2/physiology , Spike Glycoprotein, Coronavirus/chemistry , Spike Glycoprotein, Coronavirus/metabolism
15.
J Exp Med ; 218(3)2021 03 01.
Article in English | MEDLINE | ID: covidwho-1044017

ABSTRACT

SARS-CoV-2, the causative agent of COVID-19, has been responsible for over 42 million infections and 1 million deaths since its emergence in December 2019. There are few therapeutic options and no approved vaccines. Here, we examine the properties of highly potent human monoclonal antibodies (hu-mAbs) in a Syrian hamster model of SARS-CoV-2 and in a mouse-adapted model of SARS-CoV-2 infection (SARS-CoV-2 MA). Antibody combinations were effective for prevention and in therapy when administered early. However, in vitro antibody neutralization potency did not uniformly correlate with in vivo protection, and some hu-mAbs were more protective in combination in vivo. Analysis of antibody Fc regions revealed that binding to activating Fc receptors contributes to optimal protection against SARS-CoV-2 MA. The data indicate that intact effector function can affect hu-mAb protective activity and that in vivo testing is required to establish optimal hu-mAb combinations for COVID-19 prevention.


Subject(s)
Antibodies, Monoclonal, Murine-Derived , Antibodies, Neutralizing , Antibodies, Viral , Betacoronavirus/immunology , Coronavirus Infections , Pandemics , Pneumonia, Viral , Animals , Antibodies, Monoclonal, Murine-Derived/immunology , Antibodies, Monoclonal, Murine-Derived/pharmacology , Antibodies, Neutralizing/immunology , Antibodies, Neutralizing/pharmacology , Antibodies, Viral/immunology , Antibodies, Viral/pharmacology , COVID-19 , Cell Line , Coronavirus Infections/immunology , Coronavirus Infections/therapy , Female , Humans , Mesocricetus , Mice , Mice, Inbred BALB C , Pneumonia, Viral/immunology , Pneumonia, Viral/therapy , SARS-CoV-2
16.
Journal of Virology ; 93(24):1-14, 2019.
Article | Academic Search Complete | ID: covidwho-1017136

ABSTRACT

Coronaviruses (CoVs) have emerged from animal reservoirs to cause severe and lethal disease in humans, but there are currently no FDA-approved antivirals to treat the infections. One class of antiviral compounds, nucleoside analogues, mimics naturally occurring nucleosides to inhibit viral replication. While these compounds have been successful therapeutics for several viral infections, mutagenic nucleoside analogues, such as ribavirin and 5-fluorouracil, have been ineffective at inhibiting CoVs. This has been attributed to the proofreading activity of the viral 3'-5' exoribonuclease (ExoN). β-d-N4-Hydroxycytidine (NHC) (EIDD-1931;Emory Institute for Drug Development) has recently been reported to inhibit multiple viruses. Here, we demonstrate that NHC inhibits both murine hepatitis virus (MHV) (50% effective concentration [EC50] = 0.17 μM) and Middle East respiratory syndrome CoV (MERS-CoV) (EC50 = 0.56 μM) with minimal cytotoxicity. NHC inhibited MHV lacking ExoN proofreading activity similarly to wild-type (WT) MHV, suggesting an ability to evade or overcome ExoN activity. NHC inhibited MHV only when added early during infection, decreased viral specific infectivity, and increased the number and proportion of G:A and C:U transition mutations present after a single infection. Low-level NHC resistance was difficult to achieve and was associated with multiple transition mutations across the genome in both MHV and MERS-CoV. These results point to a virus-mutagenic mechanism of NHC inhibition in CoVs and indicate a high genetic barrier to NHC resistance. Together, the data support further development of NHC for treatment of CoVs and suggest a novel mechanism of NHC interaction with the CoV replication complex that may shed light on critical aspects of replication. IMPORTANCE The emergence of coronaviruses (CoVs) into human populations from animal reservoirs has demonstrated their epidemic capability, pandemic potential, and ability to cause severe disease. However, no antivirals have been approved to treat these infections. Here, we demonstrate the potent antiviral activity of a broad-spectrum ribonucleoside analogue, β-d-N4-hydroxycytidine (NHC), against two divergent CoVs. Viral proofreading activity does not markedly impact sensitivity to NHC inhibition, suggesting a novel interaction between a nucleoside analogue inhibitor and the CoV replicase. Further, passage in the presence of NHC generates only low-level resistance, likely due to the accumulation of multiple potentially deleterious transition mutations. Together, these data support a mutagenic mechanism of inhibition by NHC and further support the development of NHC for treatment of CoV infections. [ABSTRACT FROM AUTHOR] Copyright of Journal of Virology is the property of American Society for Microbiology and its content may not be copied or emailed to multiple sites or posted to a listserv without the copyright holder's express written permission. However, users may print, download, or email articles for individual use. This abstract may be abridged. No warranty is given about the accuracy of the copy. Users should refer to the original published version of the material for the full abstract. (Copyright applies to all Abstracts.)

17.
CPT Pharmacometrics Syst Pharmacol ; 10(2): 89-99, 2021 02.
Article in English | MEDLINE | ID: covidwho-966959

ABSTRACT

The severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) outbreak initiated the global coronavirus disease 2019 (COVID-19) pandemic resulting in 42.9 million confirmed infections and > 1.1 million deaths worldwide as of October 26, 2020. Remdesivir is a broad-spectrum nucleotide prodrug shown to be effective against enzootic coronaviruses. The pharmacokinetics (PKs) of remdesivir in plasma have recently been described. However, the distribution of its active metabolite nucleoside triphosphate (NTP) to the site of pulmonary infection is unknown in humans. Our objective was to use existing in vivo mouse PK data for remdesivir and its metabolites to develop a mechanism-based model to allometrically scale and simulate the human PK of remdesivir in plasma and NTP in lung homogenate. Remdesivir and GS-441524 concentrations in plasma and total phosphorylated nucleoside concentrations in lung homogenate from Ces1c-/- mice administered 25 or 50 mg/kg of remdesivir subcutaneously were simultaneously fit to estimate PK parameters. The mouse PK model was allometrically scaled to predict human PK parameters to simulate the clinically recommended 200 mg loading dose followed by 100 mg daily maintenance doses administered as 30-minute intravenous infusions. Simulations of unbound remdesivir concentrations in human plasma were below 2.48 µM, the 90% maximal inhibitory concentration for SARS-CoV-2 inhibition in vitro. Simulations of NTP in the lungs were below high efficacy in vitro thresholds. We have identified a need for alternative dosing strategies to achieve more efficacious concentrations of NTP in human lungs, perhaps by reformulating remdesivir for direct pulmonary delivery.


Subject(s)
Adenosine Monophosphate/analogs & derivatives , Alanine/analogs & derivatives , Antiviral Agents/pharmacokinetics , COVID-19/drug therapy , Models, Animal , Adenosine Monophosphate/pharmacokinetics , Adenosine Monophosphate/therapeutic use , Alanine/pharmacokinetics , Alanine/therapeutic use , Animals , Antiviral Agents/therapeutic use , COVID-19/metabolism , Cells, Cultured , Female , Humans , Mice , Mice, Inbred C57BL , Mice, Knockout , Respiratory Mucosa/drug effects , Respiratory Mucosa/metabolism , Species Specificity
18.
Cell ; 183(5): 1367-1382.e17, 2020 11 25.
Article in English | MEDLINE | ID: covidwho-893667

ABSTRACT

A safe, effective, and scalable vaccine is needed to halt the ongoing SARS-CoV-2 pandemic. We describe the structure-based design of self-assembling protein nanoparticle immunogens that elicit potent and protective antibody responses against SARS-CoV-2 in mice. The nanoparticle vaccines display 60 SARS-CoV-2 spike receptor-binding domains (RBDs) in a highly immunogenic array and induce neutralizing antibody titers 10-fold higher than the prefusion-stabilized spike despite a 5-fold lower dose. Antibodies elicited by the RBD nanoparticles target multiple distinct epitopes, suggesting they may not be easily susceptible to escape mutations, and exhibit a lower binding:neutralizing ratio than convalescent human sera, which may minimize the risk of vaccine-associated enhanced respiratory disease. The high yield and stability of the assembled nanoparticles suggest that manufacture of the nanoparticle vaccines will be highly scalable. These results highlight the utility of robust antigen display platforms and have launched cGMP manufacturing efforts to advance the SARS-CoV-2-RBD nanoparticle vaccine into the clinic.


Subject(s)
Antibodies, Neutralizing/immunology , Antibodies, Viral/immunology , COVID-19 Vaccines/immunology , COVID-19/prevention & control , Nanoparticles/chemistry , Protein Domains/immunology , SARS-CoV-2/immunology , Spike Glycoprotein, Coronavirus/chemistry , Vaccination , Adolescent , Adult , Aged , Animals , COVID-19/virology , Chlorocebus aethiops , Cohort Studies , Epitopes/immunology , Female , HEK293 Cells , Humans , Macaca nemestrina , Male , Mice, Inbred BALB C , Middle Aged , SARS-CoV-2/genetics , Spike Glycoprotein, Coronavirus/immunology , Vero Cells , Young Adult
19.
bioRxiv ; 2020 Sep 15.
Article in English | MEDLINE | ID: covidwho-807036

ABSTRACT

SARS-CoV-2, the causative agent of COVID-19, is responsible for over 24 million infections and 800,000 deaths since its emergence in December 2019. There are few therapeutic options and no approved vaccines. Here we examine the properties of highly potent human monoclonal antibodies (hu-mAbs) in a mouse adapted model of SARS-CoV-2 infection (SARS-CoV-2 MA). In vitro antibody neutralization potency did not uniformly correlate with in vivo activity, and some hu-mAbs were more potent in combination in vivo . Analysis of antibody Fc regions revealed that binding to activating Fc receptors is essential for optimal protection against SARS-CoV-2 MA. The data indicate that hu-mAb protective activity is dependent on intact effector function and that in vivo testing is required to establish optimal hu-mAb combinations for COVID-19 prevention.

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