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1.
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
2.
J Virol ; 95(23): e0097421, 2021 11 09.
Article in English | MEDLINE | ID: covidwho-1410203

ABSTRACT

The global COVID-19 pandemic has sparked intense interest in the rapid development of vaccines as well as animal models to evaluate vaccine candidates and to define immune correlates of protection. We recently reported a mouse-adapted SARS-CoV-2 virus strain (MA10) with the potential to infect wild-type laboratory mice, driving high levels of viral replication in respiratory tract tissues as well as severe clinical and respiratory symptoms, aspects of COVID-19 disease in humans that are important to capture in model systems. We evaluated the immunogenicity and protective efficacy of novel rhesus adenovirus serotype 52 (RhAd52) vaccines against MA10 challenge in mice. Baseline seroprevalence is lower for rhesus adenovirus vectors than for human or chimpanzee adenovirus vectors, making these vectors attractive candidates for vaccine development. We observed that RhAd52 vaccines elicited robust binding and neutralizing antibody titers, which inversely correlated with viral replication after challenge. These data support the development of RhAd52 vaccines and the use of the MA10 challenge virus to screen novel vaccine candidates and to study the immunologic mechanisms that underscore protection from SARS-CoV-2 challenge in wild-type mice. IMPORTANCE We have developed a series of SARS-CoV-2 vaccines using rhesus adenovirus serotype 52 (RhAd52) vectors, which exhibit a lower seroprevalence than human and chimpanzee vectors, supporting their development as novel vaccine vectors or as an alternative adenovirus (Ad) vector for boosting. We sought to test these vaccines using a recently reported mouse-adapted SARS-CoV-2 (MA10) virus to (i) evaluate the protective efficacy of RhAd52 vaccines and (ii) further characterize this mouse-adapted challenge model and probe immune correlates of protection. We demonstrate that RhAd52 vaccines elicit robust SARS-CoV-2-specific antibody responses and protect against clinical disease and viral replication in the lungs. Further, binding and neutralizing antibody titers correlated with protective efficacy. These data validate the MA10 mouse model as a useful tool to screen and study novel vaccine candidates, as well as the development of RhAd52 vaccines for COVID-19.


Subject(s)
Adenovirus Vaccines/immunology , Antibodies, Neutralizing/immunology , COVID-19 Vaccines/immunology , COVID-19/immunology , COVID-19/prevention & control , Pandemics/prevention & control , SARS-CoV-2/immunology , Adenoviridae Infections/immunology , Adenoviruses, Simian/immunology , Animals , Antibodies, Viral/immunology , Disease Models, Animal , Female , Humans , Immunogenicity, Vaccine , Macaca mulatta/virology , Mice , Mice, Inbred BALB C , SARS-CoV-2/pathogenicity , Vaccination
3.
J Heart Lung Transplant ; 40(10): 1076-1081, 2021 10.
Article in English | MEDLINE | ID: covidwho-1267682

ABSTRACT

While COVID-19 is best known as a respiratory infection, SARS-CoV-2 causes systemic disease manifestations including coagulopathies. Both dysregulated extracellular matrix remodeling pathways and circulating coagulation proteins are hallmarks of severe COVID-19 and often continue after the resolution of acute infection. Coagulation proteins have proven effective as biomarkers for severe disease and anticoagulants are a mainstay of COVID-19 therapeutics in hospitalized patients. While much knowledge has been gained about the role of clotting pathway activation in COVID-19, much remains to be elucidated in this complex network of signaling pathways.


Subject(s)
Blood Coagulation Disorders/etiology , COVID-19/complications , COVID-19/physiopathology , Wound Healing , Humans
4.
PLoS Pathog ; 17(1): e1009287, 2021 01.
Article in English | MEDLINE | ID: covidwho-1105834

ABSTRACT

The COVID-19 pandemic has revealed that infection with SARS-CoV-2 can result in a wide range of clinical outcomes in humans. An incomplete understanding of immune correlates of protection represents a major barrier to the design of vaccines and therapeutic approaches to prevent infection or limit disease. This deficit is largely due to the lack of prospectively collected, pre-infection samples from individuals that go on to become infected with SARS-CoV-2. Here, we utilized data from genetically diverse Collaborative Cross (CC) mice infected with SARS-CoV to determine whether baseline T cell signatures are associated with a lack of viral control and severe disease upon infection. SARS-CoV infection of CC mice results in a variety of viral load trajectories and disease outcomes. Overall, a dysregulated, pro-inflammatory signature of circulating T cells at baseline was associated with severe disease upon infection. Our study serves as proof of concept that circulating T cell signatures at baseline can predict clinical and virologic outcomes upon SARS-CoV infection. Identification of basal immune predictors in humans could allow for identification of individuals at highest risk of severe clinical and virologic outcomes upon infection, who may thus most benefit from available clinical interventions to restrict infection and disease.


Subject(s)
COVID-19/immunology , COVID-19/virology , SARS-CoV-2/physiology , T-Lymphocytes/immunology , Animals , COVID-19/genetics , Female , Humans , Male , Mice , Mice, Inbred C57BL , Phenotype , Viral Load
5.
Nature ; 591(7850): 451-457, 2021 03.
Article in English | MEDLINE | ID: covidwho-1075231

ABSTRACT

All coronaviruses known to have recently emerged as human pathogens probably originated in bats1. Here we use a single experimental platform based on immunodeficient mice implanted with human lung tissue (hereafter, human lung-only mice (LoM)) to demonstrate the efficient in vivo replication of severe acute respiratory syndrome coronavirus (SARS-CoV), Middle East respiratory syndrome coronavirus (MERS-CoV) and severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), as well as two endogenous SARS-like bat coronaviruses that show potential for emergence as human pathogens. Virus replication in this model occurs in bona fide human lung tissue and does not require any type of adaptation of the virus or the host. Our results indicate that bats contain endogenous coronaviruses that are capable of direct transmission to humans. Our detailed analysis of in vivo infection with SARS-CoV-2 in human lung tissue from LoM showed a predominant infection of human lung epithelial cells, including type-2 pneumocytes that are present in alveoli and ciliated airway cells. Acute infection with SARS-CoV-2 was highly cytopathic and induced a robust and sustained type-I interferon and inflammatory cytokine and chemokine response. Finally, we evaluated a therapeutic and pre-exposure prophylaxis strategy for SARS-CoV-2 infection. Our results show that therapeutic and prophylactic administration of EIDD-2801-an oral broad-spectrum antiviral agent that is currently in phase II/III clinical trials-markedly inhibited SARS-CoV-2 replication in vivo, and thus has considerable potential for the prevention and treatment of COVID-19.


Subject(s)
COVID-19/drug therapy , COVID-19/prevention & control , Cytidine/analogs & derivatives , Hydroxylamines/administration & dosage , Hydroxylamines/therapeutic use , Administration, Oral , Alveolar Epithelial Cells/immunology , Alveolar Epithelial Cells/pathology , Alveolar Epithelial Cells/virology , Animals , COVID-19/immunology , Chemoprevention , Chiroptera/virology , Clinical Trials, Phase II as Topic , Clinical Trials, Phase III as Topic , Cytidine/administration & dosage , Cytidine/therapeutic use , Cytokines/immunology , Epithelial Cells/virology , Female , Heterografts , Humans , Immunity, Innate , Interferon Type I/immunology , Lung/immunology , Lung/pathology , Lung/virology , Lung Transplantation , Male , Mice , Post-Exposure Prophylaxis , Pre-Exposure Prophylaxis , SARS-CoV-2/immunology , SARS-CoV-2/pathogenicity , Virus Replication
7.
Science ; 371(6531): 823-829, 2021 02 19.
Article in English | MEDLINE | ID: covidwho-1048643

ABSTRACT

The recurrent zoonotic spillover of coronaviruses (CoVs) into the human population underscores the need for broadly active countermeasures. We employed a directed evolution approach to engineer three severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) antibodies for enhanced neutralization breadth and potency. One of the affinity-matured variants, ADG-2, displays strong binding activity to a large panel of sarbecovirus receptor binding domains and neutralizes representative epidemic sarbecoviruses with high potency. Structural and biochemical studies demonstrate that ADG-2 employs a distinct angle of approach to recognize a highly conserved epitope that overlaps the receptor binding site. In immunocompetent mouse models of SARS and COVID-19, prophylactic administration of ADG-2 provided complete protection against respiratory burden, viral replication in the lungs, and lung pathology. Altogether, ADG-2 represents a promising broad-spectrum therapeutic candidate against clade 1 sarbecoviruses.


Subject(s)
Antibodies, Monoclonal/immunology , Antibodies, Viral/immunology , Betacoronavirus/immunology , Broadly Neutralizing Antibodies/immunology , SARS-CoV-2/immunology , Spike Glycoprotein, Coronavirus/immunology , Angiotensin-Converting Enzyme 2/metabolism , Animals , Antibodies, Monoclonal/genetics , Antibodies, Monoclonal/metabolism , Antibodies, Viral/genetics , Antibodies, Viral/metabolism , Antibody Affinity , Binding Sites , Binding Sites, Antibody , Broadly Neutralizing Antibodies/genetics , Broadly Neutralizing Antibodies/metabolism , COVID-19/prevention & control , COVID-19/therapy , Cell Surface Display Techniques , Directed Molecular Evolution , Epitopes/immunology , Humans , Immunization, Passive , Immunoglobulin Fc Fragments/immunology , Mice, Inbred BALB C , Protein Domains , Protein Engineering , Receptors, Coronavirus/metabolism , SARS Virus/immunology , Severe Acute Respiratory Syndrome/immunology , Severe Acute Respiratory Syndrome/prevention & control , Severe Acute Respiratory Syndrome/therapy , Spike Glycoprotein, Coronavirus/metabolism
8.
Science ; 370(6523): 1464-1468, 2020 12 18.
Article in English | MEDLINE | ID: covidwho-922513

ABSTRACT

The spike aspartic acid-614 to glycine (D614G) substitution is prevalent in global severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) strains, but its effects on viral pathogenesis and transmissibility remain unclear. We engineered a SARS-CoV-2 variant containing this substitution. The variant exhibits more efficient infection, replication, and competitive fitness in primary human airway epithelial cells but maintains similar morphology and in vitro neutralization properties, compared with the ancestral wild-type virus. Infection of human angiotensin-converting enzyme 2 (ACE2) transgenic mice and Syrian hamsters with both viruses resulted in similar viral titers in respiratory tissues and pulmonary disease. However, the D614G variant transmits significantly faster and displayed increased competitive fitness than the wild-type virus in hamsters. These data show that the D614G substitution enhances SARS-CoV-2 infectivity, competitive fitness, and transmission in primary human cells and animal models.


Subject(s)
COVID-19/transmission , COVID-19/virology , SARS-CoV-2/genetics , SARS-CoV-2/pathogenicity , Amino Acid Substitution , Angiotensin-Converting Enzyme 2/genetics , Animals , Asparagine/genetics , Cricetinae , Genetic Fitness/genetics , Glycine/genetics , Humans , Mesocricetus , Mice , Mice, Transgenic , Respiratory Mucosa/virology , Virulence/genetics , Virus Replication/genetics
9.
Proc Natl Acad Sci U S A ; 117(47): 29832-29838, 2020 11 24.
Article in English | MEDLINE | ID: covidwho-900111

ABSTRACT

Effective therapies are urgently needed for the SARS-CoV-2/COVID-19 pandemic. We identified panels of fully human monoclonal antibodies (mAbs) from large phage-displayed Fab, scFv, and VH libraries by panning against the receptor binding domain (RBD) of the SARS-CoV-2 spike (S) glycoprotein. A high-affinity Fab was selected from one of the libraries and converted to a full-size antibody, IgG1 ab1, which competed with human ACE2 for binding to RBD. It potently neutralized replication-competent SARS-CoV-2 but not SARS-CoV, as measured by two different tissue culture assays, as well as a replication-competent mouse ACE2-adapted SARS-CoV-2 in BALB/c mice and native virus in hACE2-expressing transgenic mice showing activity at the lowest tested dose of 2 mg/kg. IgG1 ab1 also exhibited high prophylactic and therapeutic efficacy in a hamster model of SARS-CoV-2 infection. The mechanism of neutralization is by competition with ACE2 but could involve antibody-dependent cellular cytotoxicity (ADCC) as IgG1 ab1 had ADCC activity in vitro. The ab1 sequence has a relatively low number of somatic mutations, indicating that ab1-like antibodies could be quickly elicited during natural SARS-CoV-2 infection or by RBD-based vaccines. IgG1 ab1 did not aggregate, did not exhibit other developability liabilities, and did not bind to any of the 5,300 human membrane-associated proteins tested. These results suggest that IgG1 ab1 has potential for therapy and prophylaxis of SARS-CoV-2 infections. The rapid identification (within 6 d of availability of antigen for panning) of potent mAbs shows the value of large antibody libraries for response to public health threats from emerging microbes.


Subject(s)
COVID-19 Serological Testing/methods , COVID-19 Vaccines/immunology , COVID-19/therapy , Angiotensin-Converting Enzyme 2/metabolism , Animals , Antibodies, Viral/blood , Antibodies, Viral/immunology , Antibody-Dependent Cell Cytotoxicity , COVID-19 Serological Testing/standards , COVID-19 Vaccines/standards , Chlorocebus aethiops , Cricetinae , Female , Humans , Immunization, Passive/methods , Immunization, Passive/standards , Immunogenicity, Vaccine , Immunoglobulin G/blood , Immunoglobulin G/immunology , Mice , Mice, Inbred BALB C , SARS-CoV-2/immunology , Spike Glycoprotein, Coronavirus/chemistry , Spike Glycoprotein, Coronavirus/immunology , Vero Cells
10.
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
11.
bioRxiv ; 2020 Sep 29.
Article in English | MEDLINE | ID: covidwho-835242

ABSTRACT

The D614G substitution in the S protein is most prevalent SARS-CoV-2 strain circulating globally, but its effects in viral pathogenesis and transmission remain unclear. We engineered SARS-CoV-2 variants harboring the D614G substitution with or without nanoluciferase. The D614G variant replicates more efficiency in primary human proximal airway epithelial cells and is more fit than wildtype (WT) virus in competition studies. With similar morphology to the WT virion, the D614G virus is also more sensitive to SARS-CoV-2 neutralizing antibodies. Infection of human ACE2 transgenic mice and Syrian hamsters with the WT or D614G viruses produced similar titers in respiratory tissue and pulmonary disease. However, the D614G variant exhibited significantly faster droplet transmission between hamsters than the WT virus, early after infection. Our study demonstrated the SARS-CoV2 D614G substitution enhances infectivity, replication fitness, and early transmission.

12.
bioRxiv ; 2020 Sep 21.
Article in English | MEDLINE | ID: covidwho-807105

ABSTRACT

The COVID-19 pandemic has revealed that infection with SARS-CoV-2 can result in a wide range of clinical outcomes in humans, from asymptomatic or mild disease to severe disease that can require mechanical ventilation. An incomplete understanding of immune correlates of protection represents a major barrier to the design of vaccines and therapeutic approaches to prevent infection or limit disease. This deficit is largely due to the lack of prospectively collected, pre-infection samples from indiviuals that go on to become infected with SARS-CoV-2. Here, we utilized data from a screen of genetically diverse mice from the Collaborative Cross (CC) infected with SARS-CoV to determine whether circulating baseline T cell signatures are associated with a lack of viral control and severe disease upon infection. SARS-CoV infection of CC mice results in a variety of viral load trajectories and disease outcomes. Further, early control of virus in the lung correlates with an increased abundance of activated CD4 and CD8 T cells and regulatory T cells prior to infections across strains. A basal propensity of T cells to express IFNg and IL17 over TNFa also correlated with early viral control. Overall, a dysregulated, pro-inflammatory signature of circulating T cells at baseline was associated with severe disease upon infection. While future studies of human samples prior to infection with SARS-CoV-2 are required, our studies in mice with SARS-CoV serve as proof of concept that circulating T cell signatures at baseline can predict clinical and virologic outcomes upon SARS-CoV infection. Identification of basal immune predictors in humans could allow for identification of individuals at highest risk of severe clinical and virologic outcomes upon infection, who may thus most benefit from available clinical interventions to restrict infection and disease. SUMMARY: We used a screen of genetically diverse mice from the Collaborative Cross infected with mouse-adapted SARS-CoV in combination with comprehensive pre-infection immunophenotyping to identify baseline circulating immune correlates of severe virologic and clinical outcomes upon SARS-CoV infection.

13.
Nature ; 586(7830): 509-515, 2020 10.
Article in English | MEDLINE | ID: covidwho-792975

ABSTRACT

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is the aetiological agent of coronavirus disease 2019 (COVID-19), an emerging respiratory infection caused by the introduction of a novel coronavirus into humans late in 2019 (first detected in Hubei province, China). As of 18 September 2020, SARS-CoV-2 has spread to 215 countries, has infected more than 30 million people and has caused more than 950,000 deaths. As humans do not have pre-existing immunity to SARS-CoV-2, there is an urgent need to develop therapeutic agents and vaccines to mitigate the current pandemic and to prevent the re-emergence of COVID-19. In February 2020, the World Health Organization (WHO) assembled an international panel to develop animal models for COVID-19 to accelerate the testing of vaccines and therapeutic agents. Here we summarize the findings to date and provides relevant information for preclinical testing of vaccine candidates and therapeutic agents for COVID-19.


Subject(s)
Coronavirus Infections/drug therapy , Coronavirus Infections/prevention & control , Disease Models, Animal , Pandemics/prevention & control , Pneumonia, Viral/drug therapy , Pneumonia, Viral/prevention & control , Animals , Betacoronavirus/drug effects , Betacoronavirus/immunology , COVID-19 , COVID-19 Vaccines , Coronavirus Infections/immunology , Ferrets/virology , Humans , Mesocricetus/virology , Mice , Pneumonia, Viral/immunology , Primates/virology , SARS-CoV-2 , Viral Vaccines/immunology
14.
Cell ; 183(4): 1070-1085.e12, 2020 11 12.
Article in English | MEDLINE | ID: covidwho-785288

ABSTRACT

The SARS-CoV-2 pandemic has caused extreme human suffering and economic harm. We generated and characterized a new mouse-adapted SARS-CoV-2 virus that captures multiple aspects of severe COVID-19 disease in standard laboratory mice. This SARS-CoV-2 model exhibits the spectrum of morbidity and mortality of COVID-19 disease as well as aspects of host genetics, age, cellular tropisms, elevated Th1 cytokines, and loss of surfactant expression and pulmonary function linked to pathological features of acute lung injury (ALI) and acute respiratory distress syndrome (ARDS). This model can rapidly access existing mouse resources to elucidate the role of host genetics, underlying molecular mechanisms governing SARS-CoV-2 pathogenesis, and the protective or pathogenic immune responses related to disease severity. The model promises to provide a robust platform for studies of ALI and ARDS to evaluate vaccine and antiviral drug performance, including in the most vulnerable populations (i.e., the aged) using standard laboratory mice.


Subject(s)
Acute Lung Injury/pathology , Betacoronavirus/pathogenicity , Coronavirus Infections/pathology , Pneumonia, Viral/pathology , Animals , Betacoronavirus/isolation & purification , Betacoronavirus/physiology , COVID-19 , Cell Line , Chemokines/blood , Coronavirus Infections/mortality , Coronavirus Infections/virology , Cytokines/blood , Disease Models, Animal , Female , Humans , Lung/pathology , Lung/physiology , Lung/virology , Mice , Mice, Inbred BALB C , Mice, Inbred C57BL , Pandemics , Pneumonia, Viral/mortality , Pneumonia, Viral/virology , Respiratory Distress Syndrome/pathology , SARS-CoV-2 , Severity of Illness Index , Survival Rate
15.
Nature ; 586(7830): 560-566, 2020 10.
Article in English | MEDLINE | ID: covidwho-733515

ABSTRACT

Coronaviruses are prone to transmission to new host species, as recently demonstrated by the spread to humans of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), the causative agent of the coronavirus disease 2019 (COVID-19) pandemic1. Small animal models that recapitulate SARS-CoV-2 disease are needed urgently for rapid evaluation of medical countermeasures2,3. SARS-CoV-2 cannot infect wild-type laboratory mice owing to inefficient interactions between the viral spike protein and the mouse orthologue of the human receptor, angiotensin-converting enzyme 2 (ACE2)4. Here we used reverse genetics5 to remodel the interaction between SARS-CoV-2 spike protein and mouse ACE2 and designed mouse-adapted SARS-CoV-2 (SARS-CoV-2 MA), a recombinant virus that can use mouse ACE2 for entry into cells. SARS-CoV-2 MA was able to replicate in the upper and lower airways of both young adult and aged BALB/c mice. SARS-CoV-2 MA caused more severe disease in aged mice, and exhibited more clinically relevant phenotypes than those seen in Hfh4-ACE2 transgenic mice, which express human ACE2 under the control of the Hfh4 (also known as Foxj1) promoter. We demonstrate the utility of this model using vaccine-challenge studies in immune-competent mice with native expression of mouse ACE2. Finally, we show that the clinical candidate interferon-λ1a (IFN-λ1a) potently inhibits SARS-CoV-2 replication in primary human airway epithelial cells in vitro-both prophylactic and therapeutic administration of IFN-λ1a diminished SARS-CoV-2 replication in mice. In summary, the mouse-adapted SARS-CoV-2 MA model demonstrates age-related disease pathogenesis and supports the clinical use of pegylated IFN-λ1a as a treatment for human COVID-196.


Subject(s)
Betacoronavirus , Coronavirus Infections/drug therapy , Coronavirus Infections/prevention & control , Disease Models, Animal , Interferons/pharmacology , Interferons/therapeutic use , Interleukins/pharmacology , Interleukins/therapeutic use , Pandemics/prevention & control , Pneumonia, Viral/drug therapy , Pneumonia, Viral/prevention & control , Viral Vaccines/immunology , Aging/immunology , Angiotensin-Converting Enzyme 2 , Animals , Betacoronavirus/drug effects , Betacoronavirus/immunology , Betacoronavirus/pathogenicity , COVID-19 , COVID-19 Vaccines , Coronavirus Infections/genetics , Coronavirus Infections/immunology , Female , Forkhead Transcription Factors/genetics , Humans , Interferon-alpha/administration & dosage , Interferon-alpha/pharmacology , Interferon-alpha/therapeutic use , Interferons/administration & dosage , Interleukins/administration & dosage , Male , Mice , Mice, Inbred BALB C , Mice, Transgenic , Models, Molecular , Peptidyl-Dipeptidase A/genetics , Peptidyl-Dipeptidase A/metabolism , Pneumonia, Viral/genetics , Pneumonia, Viral/immunology , Receptors, Virus/genetics , Receptors, Virus/metabolism , SARS-CoV-2
16.
Cell ; 183(1): 169-184.e13, 2020 10 01.
Article in English | MEDLINE | ID: covidwho-720448

ABSTRACT

The coronavirus disease 2019 pandemic has made deployment of an effective vaccine a global health priority. We evaluated the protective activity of a chimpanzee adenovirus-vectored vaccine encoding a prefusion stabilized spike protein (ChAd-SARS-CoV-2-S) in challenge studies with severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) and mice expressing the human angiotensin-converting enzyme 2 receptor. Intramuscular dosing of ChAd-SARS-CoV-2-S induces robust systemic humoral and cell-mediated immune responses and protects against lung infection, inflammation, and pathology but does not confer sterilizing immunity, as evidenced by detection of viral RNA and induction of anti-nucleoprotein antibodies after SARS-CoV-2 challenge. In contrast, a single intranasal dose of ChAd-SARS-CoV-2-S induces high levels of neutralizing antibodies, promotes systemic and mucosal immunoglobulin A (IgA) and T cell responses, and almost entirely prevents SARS-CoV-2 infection in both the upper and lower respiratory tracts. Intranasal administration of ChAd-SARS-CoV-2-S is a candidate for preventing SARS-CoV-2 infection and transmission and curtailing pandemic spread.


Subject(s)
Coronavirus Infections/immunology , Immunogenicity, Vaccine , Pneumonia, Viral/immunology , Viral Vaccines/immunology , Adenoviridae/genetics , Administration, Intranasal , Animals , Antibodies, Neutralizing/blood , Antibodies, Viral/blood , COVID-19 , COVID-19 Vaccines , Chlorocebus aethiops , Coronavirus Infections/pathology , Coronavirus Infections/prevention & control , Female , HEK293 Cells , Humans , Injections, Intramuscular , Mice , Mice, Inbred BALB C , Pandemics , Pneumonia, Viral/pathology , Respiratory Mucosa/immunology , Respiratory Mucosa/pathology , Respiratory Mucosa/virology , Spike Glycoprotein, Coronavirus/genetics , Spike Glycoprotein, Coronavirus/immunology , Vero Cells , Viral Vaccines/administration & dosage
18.
Nature ; 584(7821): 443-449, 2020 08.
Article in English | MEDLINE | ID: covidwho-647154

ABSTRACT

The ongoing pandemic of coronavirus disease 2019 (COVID-19), which is caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), is a major threat to global health1 and the medical countermeasures available so far are limited2,3. Moreover, we currently lack a thorough understanding of the mechanisms of humoral immunity to SARS-CoV-24. Here we analyse a large panel of human monoclonal antibodies that target the spike (S) glycoprotein5, and identify several that exhibit potent neutralizing activity and fully block the receptor-binding domain of the S protein (SRBD) from interacting with human angiotensin-converting enzyme 2 (ACE2). Using competition-binding, structural and functional studies, we show that the monoclonal antibodies can be clustered into classes that recognize distinct epitopes on the SRBD, as well as distinct conformational states of the S trimer. Two potently neutralizing monoclonal antibodies, COV2-2196 and COV2-2130, which recognize non-overlapping sites, bound simultaneously to the S protein and neutralized wild-type SARS-CoV-2 virus in a synergistic manner. In two mouse models of SARS-CoV-2 infection, passive transfer of COV2-2196, COV2-2130 or a combination of both of these antibodies protected mice from weight loss and reduced the viral burden and levels of inflammation in the lungs. In addition, passive transfer of either of two of the most potent ACE2-blocking monoclonal antibodies (COV2-2196 or COV2-2381) as monotherapy protected rhesus macaques from SARS-CoV-2 infection. These results identify protective epitopes on the SRBD and provide a structure-based framework for rational vaccine design and the selection of robust immunotherapeutic agents.


Subject(s)
Antibodies, Neutralizing/immunology , Antibodies, Viral/immunology , Betacoronavirus/immunology , Coronavirus Infections/immunology , Coronavirus Infections/prevention & control , Pandemics/prevention & control , Pneumonia, Viral/immunology , Pneumonia, Viral/prevention & control , Angiotensin-Converting Enzyme 2 , Animals , Antibodies, Monoclonal/immunology , Betacoronavirus/chemistry , Binding, Competitive , COVID-19 , Cell Line , Cross Reactions , Disease Models, Animal , Epitopes, B-Lymphocyte/chemistry , Epitopes, B-Lymphocyte/immunology , Female , Humans , Macaca mulatta , Male , Mice , Middle Aged , Neutralization Tests , Peptidyl-Dipeptidase A/genetics , Peptidyl-Dipeptidase A/metabolism , Pre-Exposure Prophylaxis , SARS Virus/chemistry , SARS Virus/immunology , SARS-CoV-2 , Severe Acute Respiratory Syndrome/immunology , Spike Glycoprotein, Coronavirus/chemistry , Spike Glycoprotein, Coronavirus/immunology , Spike Glycoprotein, Coronavirus/metabolism
19.
Cell ; 182(2): 429-446.e14, 2020 07 23.
Article in English | MEDLINE | ID: covidwho-381993

ABSTRACT

The mode of acquisition and causes for the variable clinical spectrum of coronavirus disease 2019 (COVID-19) remain unknown. We utilized a reverse genetics system to generate a GFP reporter virus to explore severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) pathogenesis and a luciferase reporter virus to demonstrate sera collected from SARS and COVID-19 patients exhibited limited cross-CoV neutralization. High-sensitivity RNA in situ mapping revealed the highest angiotensin-converting enzyme 2 (ACE2) expression in the nose with decreasing expression throughout the lower respiratory tract, paralleled by a striking gradient of SARS-CoV-2 infection in proximal (high) versus distal (low) pulmonary epithelial cultures. COVID-19 autopsied lung studies identified focal disease and, congruent with culture data, SARS-CoV-2-infected ciliated and type 2 pneumocyte cells in airway and alveolar regions, respectively. These findings highlight the nasal susceptibility to SARS-CoV-2 with likely subsequent aspiration-mediated virus seeding to the lung in SARS-CoV-2 pathogenesis. These reagents provide a foundation for investigations into virus-host interactions in protective immunity, host susceptibility, and virus pathogenesis.


Subject(s)
Betacoronavirus/genetics , Coronavirus Infections/pathology , Coronavirus Infections/virology , Pneumonia, Viral/pathology , Pneumonia, Viral/virology , Respiratory System/virology , Reverse Genetics/methods , Aged , Angiotensin-Converting Enzyme 2 , Animals , Antibodies, Monoclonal/immunology , Antibodies, Neutralizing/immunology , Betacoronavirus/immunology , Betacoronavirus/pathogenicity , COVID-19 , Cell Line , Cells, Cultured , Chlorocebus aethiops , Coronavirus Infections/immunology , Coronavirus Infections/therapy , Cystic Fibrosis/pathology , DNA, Recombinant , Female , Furin/metabolism , Humans , Immunization, Passive , Lung/metabolism , Lung/pathology , Lung/virology , Male , Middle Aged , Nasal Mucosa/metabolism , Nasal Mucosa/pathology , Nasal Mucosa/virology , Pandemics , Peptidyl-Dipeptidase A/metabolism , Pneumonia, Viral/immunology , Respiratory System/pathology , SARS-CoV-2 , Serine Endopeptidases/metabolism , Vero Cells , Virulence , Virus Replication
20.
J Infect Dis ; 221(6): 882-889, 2020 03 02.
Article in English | MEDLINE | ID: covidwho-27190

ABSTRACT

BACKGROUND: Virus infections result in a range of clinical outcomes for the host, from asymptomatic to severe or even lethal disease. Despite global efforts to prevent and treat virus infections to limit morbidity and mortality, the continued emergence and re-emergence of new outbreaks as well as common infections such as influenza persist as a health threat. Challenges to the prevention of severe disease after virus infection include both a paucity of protective vaccines as well as the early identification of individuals with the highest risk that may require supportive treatment. METHODS: We completed a screen of mice from the Collaborative Cross (CC) that we infected with influenza, severe acute respiratory syndrome-coronavirus, and West Nile virus. RESULTS: The CC mice exhibited a range of disease manifestations upon infections, and we used this natural variation to identify strains with mortality after infection and strains exhibiting no mortality. We then used comprehensive preinfection immunophenotyping to identify global baseline immune correlates of protection from mortality to virus infection. CONCLUSIONS: These data suggest that immune phenotypes might be leveraged to identify humans at highest risk of adverse clinical outcomes upon infection, who may most benefit from intensive clinical interventions, in addition to providing insight for rational vaccine design.


Subject(s)
Mortality , RNA Virus Infections/immunology , RNA Virus Infections/mortality , Animals , Collaborative Cross Mice , Cytokines/metabolism , Disease Models, Animal , Female , Humans , Influenza A virus/immunology , Influenza, Human , Male , Mice , Orthomyxoviridae Infections/immunology , Orthomyxoviridae Infections/mortality , RNA , RNA Virus Infections/virology , SARS Virus/immunology , Severe Acute Respiratory Syndrome/immunology , Severe Acute Respiratory Syndrome/mortality , T-Lymphocytes/immunology , T-Lymphocytes/metabolism , Viral Vaccines/immunology , West Nile Fever/immunology , West Nile Fever/mortality , West Nile virus/immunology
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