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2.
Vaccine ; 40(4): 574-586, 2022 01 28.
Article in English | MEDLINE | ID: covidwho-1586276

ABSTRACT

A series of recombinant human type 5 adenoviruses that express the full-length or membrane-truncated spike protein (S) of SARS-CoV-2 (AdCoV2-S or AdCoV2-SdTM, respectively) was tested the efficacy against SARS-CoV-2 via intranasal (i.n.) or subcutaneous (s.c.) immunization in a rodent model. Mucosal delivery of adenovirus (Ad) vaccines could induce anti-SARS-CoV-2 IgG and IgA in the serum and in the mucosal, respectively as indicated by vaginal wash (vw) and bronchoalveolar lavage fluid (BALF). Serum anti-SARS-CoV-2 IgG but not IgA in the vw and BALF was induced by AdCoV2-S s.c.. Administration of AdCoV2-S i.n. was able to induce higher anti-SARS-CoV-2 binding and neutralizing antibody levels than s.c. injection. AdCoV2-SdTM i.n. induced a lower antibody responses than AdCoV2-S i.n.. Induced anti-S antibody responses by AdCoV2-S via i.n. or s.c. were not influenced by the pre-existing serum anti-Ad antibody. Novelty, S-specific IgG1 which represented Th2-mediated humoral response was dominantly induced in Ad i.n.-immunized serum in contrast to more IgG2a which represented Th1-mediated cellular response found in Ad s.c.-immunized serum. The activation of S-specific IFN-É£ and IL-4 in splenic Th1 and Th2 cells, respectively, was observed in the AdCoV2-S i.n. and s.c. groups, indicating the Th1 and Th2 immunity were activated. AdCoV2-S and AdCoV2-SdTM significantly prevented body weight loss and reduced pulmonary viral loads in hamsters. A reduction in inflammation in the lungs was observed in AdCoV-S via i.n. or s.c.-immunized hamsters following a SARS-CoV-2 challenge. It correlated to Th1 cytokine but no inflammatory cytokines secretions found in AdCoV-S i.n. -immunized BALF. These results indicate that intranasal delivery of AdCoV2-S vaccines is safe and potent at preventing SARS-CoV-2 infections.


Subject(s)
Adenovirus Vaccines , COVID-19 , Animals , Antibodies, Viral , COVID-19 Vaccines , Cricetinae , Female , Humans , SARS-CoV-2 , Spike Glycoprotein, Coronavirus/genetics
3.
MAbs ; 14(1): 2005507, 2022.
Article in English | MEDLINE | ID: covidwho-1585297

ABSTRACT

The emergence of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) has triggered a serious public health crisis worldwide, and considering the novelty of the disease, preventative and therapeutic measures alike are urgently needed. To accelerate such efforts, the development of JS016, a neutralizing monoclonal antibody directed against the SARS-CoV-2 spike protein, was expedited from a typical 12- to 18-month period to a 4-month period. During this process, transient Chinese hamster ovary cell lines are used to support preclinical, investigational new drug-enabling toxicology research, and early Chemistry, Manufacturing and Controls development; mini-pool materials to supply Phase 1 clinical trials; and a single-clone working cell bank for late-stage and pivotal clinical trials were successively adopted. Moreover, key process performance and product quality investigations using a series of orthogonal and state-of-the-art techniques were conducted to demonstrate the comparability of products manufactured using these three processes, and the results indicated that, despite observed variations in process performance, the primary and high-order structures, purity and impurity profiles, biological and immunological functions, and degradation behaviors under stress conditions were largely comparable. The study suggests that, in particular situations, this strategy can be adopted to accelerate the development of therapeutic biopharmaceuticals and their access to patients.


Subject(s)
Antibodies, Monoclonal/immunology , Antibodies, Neutralizing/immunology , Antibodies, Viral/immunology , COVID-19/immunology , SARS-CoV-2/immunology , Spike Glycoprotein, Coronavirus/immunology , Animals , Antibodies, Monoclonal/chemistry , Antibodies, Monoclonal/therapeutic use , Antibody Affinity/immunology , Antibody Specificity/immunology , CHO Cells , COVID-19/prevention & control , COVID-19/virology , Chromatography, High Pressure Liquid/methods , Circular Dichroism , Clone Cells , Cricetinae , Cricetulus , Humans , Immunoglobulin G/chemistry , Immunoglobulin G/immunology , Immunoglobulin G/therapeutic use , Isoelectric Point , SARS-CoV-2/metabolism , SARS-CoV-2/physiology , Spike Glycoprotein, Coronavirus/metabolism
4.
Gut Microbes ; 14(1): 2018900, 2022.
Article in English | MEDLINE | ID: covidwho-1585291

ABSTRACT

Mounting evidence suggests that the gut-to-lung axis is critical during respiratory viral infections. We herein hypothesized that disruption of gut homeostasis during severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection may associate with early disease outcomes. To address this question, we took advantage of the Syrian hamster model. Our data confirmed that this model recapitulates some hallmark features of the human disease in the lungs. We further showed that SARS-CoV-2 infection associated with mild intestinal inflammation, relative alteration in intestinal barrier property and liver inflammation and altered lipid metabolism. These changes occurred concomitantly with an alteration of the gut microbiota composition over the course of infection, notably characterized by a higher relative abundance of deleterious bacterial taxa such as Enterobacteriaceae and Desulfovibrionaceae. Conversely, several members of the Ruminococcaceae and Lachnospiraceae families, including bacteria known to produce the fermentative products short-chain fatty acids (SCFAs), had a reduced relative proportion compared to non-infected controls. Accordingly, infection led to a transient decrease in systemic SCFA amounts. SCFA supplementation during infection had no effect on clinical and inflammatory parameters. Lastly, a strong correlation between some gut microbiota taxa and clinical and inflammation indices of SARS-CoV-2 infection severity was evidenced. Collectively, alteration of the gut microbiota correlates with disease severity in hamsters making this experimental model valuable for the design of interventional, gut microbiota-targeted, approaches for the control of COVID-19.Abbreviations: SARS-CoV-2, severe acute respiratory syndrome coronavirus 2; COVID-19, coronavirus disease 2019; SCFAs, short-chain fatty acids; dpi, day post-infection; RT-PCR, reverse transcription polymerase chain reaction; IL, interleukin. ACE2, angiotensin converting enzyme 2; TMPRSS2, transmembrane serine protease 2.


Subject(s)
COVID-19/microbiology , COVID-19/physiopathology , Disease Models, Animal , Gastrointestinal Microbiome , Mesocricetus , Animals , Bacteria/classification , Bacteria/isolation & purification , Bacteria/metabolism , COVID-19/drug therapy , COVID-19/pathology , Cricetinae , Fatty Acids, Volatile/administration & dosage , Fatty Acids, Volatile/metabolism , Humans , Male , SARS-CoV-2/physiology , Severity of Illness Index
5.
Front Immunol ; 12: 765211, 2021.
Article in English | MEDLINE | ID: covidwho-1581337

ABSTRACT

Saturation suppressor mutagenesis was used to generate thermostable mutants of the SARS-CoV-2 spike receptor-binding domain (RBD). A triple mutant with an increase in thermal melting temperature of ~7°C with respect to the wild-type B.1 RBD and was expressed in high yield in both mammalian cells and the microbial host, Pichia pastoris, was downselected for immunogenicity studies. An additional derivative with three additional mutations from the B.1.351 (beta) isolate was also introduced into this background. Lyophilized proteins were resistant to high-temperature exposure and could be stored for over a month at 37°C. In mice and hamsters, squalene-in-water emulsion (SWE) adjuvanted formulations of the B.1-stabilized RBD were considerably more immunogenic than RBD lacking the stabilizing mutations and elicited antibodies that neutralized all four current variants of concern with similar neutralization titers. However, sera from mice immunized with the stabilized B.1.351 derivative showed significantly decreased neutralization titers exclusively against the B.1.617.2 (delta) VOC. A cocktail comprising stabilized B.1 and B.1.351 RBDs elicited antibodies with qualitatively improved neutralization titers and breadth relative to those immunized solely with either immunogen. Immunized hamsters were protected from high-dose viral challenge. Such vaccine formulations can be rapidly and cheaply produced, lack extraneous tags or additional components, and can be stored at room temperature. They are a useful modality to combat COVID-19, especially in remote and low-resource settings.


Subject(s)
Antibodies, Neutralizing/immunology , COVID-19 Vaccines/immunology , COVID-19/prevention & control , SARS-CoV-2/immunology , Spike Glycoprotein, Coronavirus/immunology , Animals , Antibodies, Viral/immunology , Cricetinae , Immunogenicity, Vaccine/immunology , Mice , Spike Glycoprotein, Coronavirus/genetics
6.
Front Immunol ; 12: 766112, 2021.
Article in English | MEDLINE | ID: covidwho-1581336

ABSTRACT

The coronavirus disease 2019 (COVID-19) pandemic caused by the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) has become a global health concern. The development of vaccines with high immunogenicity and safety is crucial for controlling the global COVID-19 pandemic and preventing further illness and fatalities. Here, we report the development of a SARS-CoV-2 vaccine candidate, Nanocovax, based on recombinant protein production of the extracellular (soluble) portion of the spike (S) protein of SARS-CoV-2. The results showed that Nanocovax induced high levels of S protein-specific IgG and neutralizing antibodies in three animal models: BALB/c mouse, Syrian hamster, and a non-human primate (Macaca leonina). In addition, a viral challenge study using the hamster model showed that Nanocovax protected the upper respiratory tract from SARS-CoV-2 infection. Nanocovax did not induce any adverse effects in mice (Mus musculus var. albino) and rats (Rattus norvegicus). These preclinical results indicate that Nanocovax is safe and effective.


Subject(s)
COVID-19 Vaccines/immunology , COVID-19 Vaccines/toxicity , COVID-19/prevention & control , Immunogenicity, Vaccine/immunology , Spike Glycoprotein, Coronavirus/immunology , Animals , Antibodies, Neutralizing/immunology , Antibodies, Viral/immunology , Cricetinae , Macaca , Mice , Rats , SARS-CoV-2 , Vaccines, Synthetic/immunology , Vaccines, Synthetic/toxicity
7.
Front Immunol ; 12: 719077, 2021.
Article in English | MEDLINE | ID: covidwho-1575525

ABSTRACT

The COVID-19 pandemic caused by the coronavirus SARS-CoV-2 is a major global public threat. Currently, a worldwide effort has been mounted to generate billions of effective SARS-CoV-2 vaccine doses to immunize the world's population at record speeds. However, there is still a demand for alternative effective vaccines that rapidly confer long-term protection and rely upon cost-effective, easily scaled-up manufacturing. Here, we present a Sindbis alphavirus vector (SV), transiently expressing the SARS-CoV-2 spike protein (SV.Spike), combined with the OX40 immunostimulatory antibody (αOX40) as a novel, highly effective vaccine approach. We show that SV.Spike plus αOX40 elicits long-lasting neutralizing antibodies and a vigorous T-cell response in mice. Protein binding, immunohistochemical, and cellular infection assays all show that vaccinated mice sera inhibits spike functions. Immunophenotyping, RNA Seq transcriptome profiles, and metabolic analysis indicate a reprogramming of T cells in vaccinated mice. Activated T cells were found to mobilize to lung tissue. Most importantly, SV.Spike plus αOX40 provided robust immune protection against infection with authentic coronavirus in transgenic mice expressing the human ACE2 receptor (hACE2-Tg). Finally, our immunization strategy induced strong effector memory response, potentiating protective immunity against re-exposure to SARS-CoV-2 spike protein. Our results show the potential of a new Sindbis virus-based vaccine platform to counteract waning immune response, which can be used as a new candidate to combat SARS-CoV-2. Given the T-cell responses elicited, our vaccine is likely to be effective against variants that are proving challenging, as well as serve as a platform to develop a broader spectrum pancoronavirus vaccine. Similarly, the vaccine approach is likely to be applicable to other pathogens.


Subject(s)
Antigens, Differentiation/immunology , COVID-19 Vaccines/immunology , COVID-19/prevention & control , SARS-CoV-2/immunology , Sindbis Virus/immunology , Spike Glycoprotein, Coronavirus/immunology , Angiotensin-Converting Enzyme 2/metabolism , Animals , Antibodies, Neutralizing/immunology , Antibodies, Viral/immunology , COVID-19/immunology , Cricetinae , Female , HEK293 Cells , Humans , Mice , Mice, Inbred C57BL , Mice, Transgenic , Sindbis Virus/genetics , T-Lymphocytes/immunology , Vaccination
8.
Methods Mol Biol ; 2410: 177-192, 2022.
Article in English | MEDLINE | ID: covidwho-1575553

ABSTRACT

The SARS-CoV-2 pandemic has impacted the health of humanity after the outbreak in Hubei, China in late December 2019. Ever since, it has taken unprecedented proportions and rapidity causing over a million fatal cases. Recently, a robust Syrian golden hamster model recapitulating COVID-19 was developed in search for effective therapeutics and vaccine candidates. However, overt clinical disease symptoms were largely absent despite high levels of virus replication and associated pathology in the respiratory tract. Therefore, we used micro-computed tomography (µCT) to longitudinally visualize lung pathology and to preclinically assess candidate vaccines. µCT proved to be crucial to quantify and noninvasively monitor disease progression, to evaluate candidate vaccine efficacy, and to improve screening efforts by allowing longitudinal data without harming live animals. Here, we give a comprehensive guide on how to use low-dose high-resolution µCT to follow-up SARS-CoV-2-induced disease and test the efficacy of COVID-19 vaccine candidates in hamsters. Our approach can likewise be applied for the preclinical assessment of antiviral and anti-inflammatory drug treatments in vivo.


Subject(s)
COVID-19 Vaccines , COVID-19 , Animals , COVID-19/prevention & control , Cricetinae , X-Ray Microtomography
9.
Viruses ; 13(12)2021 12 14.
Article in English | MEDLINE | ID: covidwho-1572667

ABSTRACT

Pre-existing comorbidities such as obesity or metabolic diseases can adversely affect the clinical outcome of COVID-19. Chronic metabolic disorders are globally on the rise and often a consequence of an unhealthy diet, referred to as a Western Diet. For the first time in the Syrian hamster model, we demonstrate the detrimental impact of a continuous high-fat high-sugar diet on COVID-19 outcome. We observed increased weight loss and lung pathology, such as exudate, vasculitis, hemorrhage, fibrin, and edema, delayed viral clearance and functional lung recovery, and prolonged viral shedding. This was accompanied by an altered, but not significantly different, systemic IL-10 and IL-6 profile, as well as a dysregulated serum lipid response dominated by polyunsaturated fatty acid-containing phosphatidylethanolamine, partially recapitulating cytokine and lipid responses associated with severe human COVID-19. Our data support the hamster model for testing restrictive or targeted diets and immunomodulatory therapies to mediate the adverse effects of metabolic disease on COVID-19.


Subject(s)
COVID-19 , Diet, High-Fat/adverse effects , Dietary Carbohydrates/adverse effects , Lipid Metabolism , Severity of Illness Index , Animals , COVID-19/pathology , Cricetinae , Cytokines/blood , Disease Models, Animal , Edema , Fibrin , Hemorrhage , Humans , Interleukin-10 , Interleukin-6 , Lipidomics , Lipids/blood , Liver/pathology , Lung/pathology , Male , Mesocricetus , Obesity , SARS-CoV-2 , Sugars , Vasculitis/pathology , Virus Shedding
11.
Viruses ; 13(12)2021 12 03.
Article in English | MEDLINE | ID: covidwho-1555015

ABSTRACT

We have developed a monoclonal antibody (mAb) cocktail (ZRC-3308) comprising of ZRC3308-A7 and ZRC3308-B10 in the ratio 1:1 for COVID-19 treatment. The mAbs were designed to have reduced immune effector functions and increased circulation half-life. mAbs showed good binding affinities to non-competing epitopes on RBD of SARS-CoV-2 spike protein and were found neutralizing SARS-CoV-2 variants B.1, B.1.1.7, B.1.351, B.1.617.2, and B.1.617.2 AY.1 in vitro. The mAb cocktail demonstrated effective prophylactic and therapeutic activity against SARS-CoV-2 infection in Syrian hamsters. The antibody cocktail appears to be a promising candidate for prophylactic use and for therapy in early COVID-19 cases that have not progressed to severe disease.


Subject(s)
Antibodies, Monoclonal, Humanized/immunology , COVID-19/therapy , SARS-CoV-2/immunology , Animals , Antibodies, Monoclonal, Humanized/therapeutic use , Antibodies, Neutralizing/immunology , Antibodies, Neutralizing/therapeutic use , Antibody Affinity , Binding Sites , COVID-19/prevention & control , Cricetinae , Disease Models, Animal , Epitopes , Humans , Immunization, Passive , Mesocricetus , Mutation , SARS-CoV-2/genetics , Spike Glycoprotein, Coronavirus/genetics , Spike Glycoprotein, Coronavirus/immunology
12.
Viruses ; 13(12)2021 12 04.
Article in English | MEDLINE | ID: covidwho-1554793

ABSTRACT

SARS-CoV-2 pathogenesis, vaccine, and therapeutic studies rely on the use of animals challenged with highly pathogenic virus stocks produced in cell cultures. Ideally, these virus stocks should be genetically and functionally similar to the original clinical isolate, retaining wild-type properties to be reliably used in animal model studies. It is well-established that SARS-CoV-2 isolates serially passaged on Vero cell lines accumulate mutations and deletions in the furin cleavage site; however, these can be eliminated when passaged on Calu-3 lung epithelial cell lines, as presented in this study. As numerous stocks of SARS-CoV-2 variants of concern are being grown in cell cultures with the intent for use in animal models, it is essential that propagation methods generate virus stocks that are pathogenic in vivo. Here, we found that the propagation of a B.1.351 SARS-CoV-2 stock on Calu-3 cells eliminated viruses that previously accumulated mutations in the furin cleavage site. Notably, there were alternative variants that accumulated at the same nucleotide positions in virus populations grown on Calu-3 cells at multiple independent facilities. When a Calu-3-derived B.1.351 virus stock was used to infect hamsters, the virus remained pathogenic and the Calu-3-specific variants persisted in the population. These results suggest that Calu-3-derived virus stocks are pathogenic but care should still be taken to evaluate virus stocks for newly arising mutations during propagation.


Subject(s)
SARS-CoV-2/growth & development , Serial Passage/methods , Spike Glycoprotein, Coronavirus/genetics , Animals , COVID-19/virology , Cell Line, Tumor , Chlorocebus aethiops , Cricetinae , Furin/metabolism , Humans , Mutation , SARS-CoV-2/genetics , SARS-CoV-2/pathogenicity , Vero Cells
13.
Eur Rev Med Pharmacol Sci ; 25(21): 6745-6766, 2021 11.
Article in English | MEDLINE | ID: covidwho-1524863

ABSTRACT

The severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) that has created the current pandemic, has caused a worldwide worry. Different countries have since enforced varying levels of lockdowns and guidelines for their populations to follow in a serious effort to mitigate the spread. Up until recently, the majority of these regulations and policies were established on the assumption that the dominant routes of transmission of this virus are through droplets and fomite contact. However, there is now a substantial amount of research pointing towards the strong possibility that SARS-CoV-2 can spread through airborne means. The World Health Organization (WHO) and the Center for Disease Control and Prevention (CDC) have recently recognized this, which poses the question of whether our collective methods of lessening transmission risk and keeping people safe have been sufficient. This paper is a comprehensive review of the evidence on SARS-CoV-2 being an airborne disease, through different epidemiological, experimental, and animal-model based published research. Studies opposing this evidence have also been discussed. The majority of these studies are favoring the high plausibility of SARS-CoV-2 aerosol transmission, and therefore the many implications of aerosol transmission have been discussed in this paper to suggest effective mitigation and control strategies.


Subject(s)
Aerosols , COVID-19/transmission , SARS-CoV-2/physiology , Animals , COVID-19/epidemiology , COVID-19/prevention & control , COVID-19/virology , Cricetinae , Disease Models, Animal , Equipment and Supplies, Hospital/virology , Feces/virology , Humans , Masks , Pandemics , Particulate Matter , RNA, Viral/metabolism , SARS-CoV-2/genetics , SARS-CoV-2/isolation & purification , Ventilation/standards , Virus Inactivation
14.
Cell Rep ; 37(3): 109838, 2021 10 19.
Article in English | MEDLINE | ID: covidwho-1517083

ABSTRACT

As severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) spreads, variants with enhanced virulence and transmissibility have emerged. Although in vitro systems allow rapid characterization, they do not fully recapitulate the dynamic interaction of virions and neutralizing antibodies in the airway. Here, we demonstrate that the N501Y variant permits respiratory infection in unmodified mice. We utilize N501Y to survey in vivo pseudovirus infection dynamics and susceptibility to reinfection with the L452R (Los Angeles), K417N + E484K (South Africa), and L452R + K417N + E484Q (India) variants. Human coronavirus disease 2019 (COVID-19)+ or vaccinated antibody isotypes, titers, variant receptor binding domain (RBD) binding, and neutralization potential are studied, revealing numerous significant correlations. Immune escape of the K417N + E484K variant is observed because infection can be appreciated in the nasopharynx, but not lungs, of mice transferred with low-antibody-tier plasma. Conversely, near-complete protection is observed in animals receiving high-antibody-tier plasma, a phenomenon that can only be appreciated in vivo.


Subject(s)
Antibodies, Viral/immunology , COVID-19/immunology , COVID-19/therapy , SARS-CoV-2/genetics , SARS-CoV-2/immunology , Animals , Antibodies, Neutralizing/immunology , Cell Line , Cricetinae , Genetic Variation , HEK293 Cells , Humans , Immune System , Immunization, Passive/methods , In Vitro Techniques , Mice , Mutation , Nasopharynx/virology , Protein Binding , Recombinant Proteins/metabolism , Spike Glycoprotein, Coronavirus/genetics
15.
Theranostics ; 12(1): 1-17, 2022.
Article in English | MEDLINE | ID: covidwho-1512993

ABSTRACT

Background: Administration of potent anti-receptor-binding domain (RBD) monoclonal antibodies has been shown to curtail viral shedding and reduce hospitalization in patients with SARS-CoV-2 infection. However, the structure-function analysis of potent human anti-RBD monoclonal antibodies and its links to the formulation of antibody cocktails remains largely elusive. Methods: Previously, we isolated a panel of neutralizing anti-RBD monoclonal antibodies from convalescent patients and showed their neutralization efficacy in vitro. Here, we elucidate the mechanism of action of antibodies and dissect antibodies at the epitope level, which leads to a formation of a potent antibody cocktail. Results: We found that representative antibodies which target non-overlapping epitopes are effective against wild type virus and recently emerging variants of concern, whilst being encoded by antibody genes with few somatic mutations. Neutralization is associated with the inhibition of binding of viral RBD to ACE2 and possibly of the subsequent fusion process. Structural analysis of representative antibodies, by cryo-electron microscopy and crystallography, reveals that they have some unique aspects that are of potential value while sharing some features in common with previously reported neutralizing monoclonal antibodies. For instance, one has a common VH 3-53 public variable region yet is unusually resilient to mutation at residue 501 of the RBD. We evaluate the in vivo efficacy of an antibody cocktail consisting of two potent non-competing anti-RBD antibodies in a Syrian hamster model. We demonstrate that the cocktail prevents weight loss, reduces lung viral load and attenuates pulmonary inflammation in hamsters in both prophylactic and therapeutic settings. Although neutralization of one of these antibodies is abrogated by the mutations of variant B.1.351, it is also possible to produce a bi-valent cocktail of antibodies both of which are resilient to variants B.1.1.7, B.1.351 and B.1.617.2. Conclusions: These findings support the up-to-date and rational design of an anti-RBD antibody cocktail as a therapeutic candidate against COVID-19.


Subject(s)
Antibodies, Monoclonal/chemistry , Antibodies, Monoclonal/pharmacology , COVID-19/drug therapy , SARS-CoV-2/metabolism , Angiotensin-Converting Enzyme 2/genetics , Angiotensin-Converting Enzyme 2/metabolism , Animals , Antibodies, Monoclonal/metabolism , Antibodies, Neutralizing/chemistry , Antibodies, Neutralizing/immunology , Antibodies, Neutralizing/pharmacology , Binding Sites , Binding, Competitive , COVID-19/virology , Cricetinae , Cryoelectron Microscopy , Crystallography, X-Ray , Dogs , Epitopes , Female , Humans , Madin Darby Canine Kidney Cells , Neutralization Tests , Protein Domains , SARS-CoV-2/genetics , SARS-CoV-2/immunology , SARS-CoV-2/pathogenicity , Spike Glycoprotein, Coronavirus/genetics , Spike Glycoprotein, Coronavirus/immunology , Spike Glycoprotein, Coronavirus/metabolism
16.
Sci Rep ; 11(1): 21849, 2021 11 08.
Article in English | MEDLINE | ID: covidwho-1505527

ABSTRACT

The huge worldwide demand for vaccines targeting SARS-CoV-2 has necessitated the continued development of novel improved formulations capable of reducing the burden of the COVID-19 pandemic. Herein, we evaluated novel protein subunit vaccine formulations containing a resistin-trimerized spike antigen, SmT1. When combined with sulfated lactosyl archaeol (SLA) archaeosome adjuvant, formulations induced robust antigen-specific humoral and cellular immune responses in mice. Antibodies had strong neutralizing activity, preventing viral spike binding and viral infection. In addition, the formulations were highly efficacious in a hamster challenge model reducing viral load and body weight loss even after a single vaccination. The antigen-specific antibodies generated by our vaccine formulations had stronger neutralizing activity than human convalescent plasma, neutralizing the spike proteins of the B.1.1.7 and B.1.351 variants of concern. As such, our SmT1 antigen along with SLA archaeosome adjuvant comprise a promising platform for the development of efficacious protein subunit vaccine formulations for SARS-CoV-2.


Subject(s)
Adjuvants, Immunologic/chemistry , Antigens, Archaeal/chemistry , COVID-19 Vaccines/therapeutic use , Lipids/chemistry , Animals , Antibodies, Neutralizing/immunology , Antibodies, Viral/immunology , Body Weight , COVID-19/therapy , Chlorocebus aethiops , Cricetinae , Cytokines/metabolism , Female , Humans , Immunity, Cellular , Immunity, Humoral , Immunization, Passive , Mesocricetus , Mice , Mice, Inbred C57BL , Neutralization Tests , Peptides/chemistry , Protein Domains , SARS-CoV-2 , Toll-Like Receptors/immunology , Vero Cells , Viral Load
17.
Nat Neurosci ; 24(11): 1522-1533, 2021 11.
Article in English | MEDLINE | ID: covidwho-1500484

ABSTRACT

Coronavirus disease 2019 (COVID-19) can damage cerebral small vessels and cause neurological symptoms. Here we describe structural changes in cerebral small vessels of patients with COVID-19 and elucidate potential mechanisms underlying the vascular pathology. In brains of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2)-infected individuals and animal models, we found an increased number of empty basement membrane tubes, so-called string vessels representing remnants of lost capillaries. We obtained evidence that brain endothelial cells are infected and that the main protease of SARS-CoV-2 (Mpro) cleaves NEMO, the essential modulator of nuclear factor-κB. By ablating NEMO, Mpro induces the death of human brain endothelial cells and the occurrence of string vessels in mice. Deletion of receptor-interacting protein kinase (RIPK) 3, a mediator of regulated cell death, blocks the vessel rarefaction and disruption of the blood-brain barrier due to NEMO ablation. Importantly, a pharmacological inhibitor of RIPK signaling prevented the Mpro-induced microvascular pathology. Our data suggest RIPK as a potential therapeutic target to treat the neuropathology of COVID-19.


Subject(s)
Blood-Brain Barrier/metabolism , Brain/metabolism , Coronavirus 3C Proteases/metabolism , Intracellular Signaling Peptides and Proteins/metabolism , Microvessels/metabolism , SARS-CoV-2/metabolism , Animals , Blood-Brain Barrier/pathology , Brain/pathology , Chlorocebus aethiops , Coronavirus 3C Proteases/genetics , Cricetinae , Female , Humans , Intracellular Signaling Peptides and Proteins/genetics , Male , Mesocricetus , Mice , Mice, Inbred C57BL , Mice, Knockout , Mice, Transgenic , Microvessels/pathology , SARS-CoV-2/genetics , Vero Cells
19.
mBio ; 12(5): e0198721, 2021 10 26.
Article in English | MEDLINE | ID: covidwho-1494967

ABSTRACT

The coronavirus disease 2019 (COVID-19) pandemic has raised concerns about the detrimental effects of antibodies. Antibody-dependent enhancement (ADE) of infection is one of the biggest concerns in terms of not only the antibody reaction to severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) upon reinfection with the virus but also the reaction to COVID-19 vaccines. In this study, we evaluated ADE of infection by using COVID-19 convalescent-phase plasma and BHK cells expressing human Fcγ receptors (FcγRs). We found that FcγRIIA and FcγRIIIA mediated modest ADE of infection against SARS-CoV-2. Although ADE of infection was observed in monocyte-derived macrophages infected with SARS-CoV-2, including its variants, proinflammatory cytokine/chemokine expression was not upregulated in macrophages. SARS-CoV-2 infection thus produces antibodies that elicit ADE of infection, but these antibodies do not contribute to excess cytokine production by macrophages. IMPORTANCE Viruses infect cells mainly via specific receptors at the cell surface. Antibody-dependent enhancement (ADE) of infection is an alternative mechanism of infection for viruses to infect immune cells that is mediated by antibodies and IgG receptors (FcγRs). Because ADE of infection contributes to the pathogenesis of some viruses, such as dengue virus and feline coronavirus, it is important to evaluate the precise mechanism of ADE and its contribution to the pathogenesis of SARS-CoV-2. Here, using convalescent-phase plasma from COVID-19 patients, we found that two types of FcγRs, FcγRIIA and FcγRIIIA, mediate ADE of SARS-CoV-2 infection. Although ADE of infection was observed for SARS-CoV-2 and its recent variants, proinflammatory cytokine production in monocyte-derived macrophages was not upregulated. These observations suggest that SARS-CoV-2 infection produces antibodies that elicit ADE of infection, but these antibodies may not be involved in aberrant cytokine release by macrophages during SARS-CoV-2 infection.


Subject(s)
Cytokines/metabolism , Macrophages/metabolism , Receptors, IgG/metabolism , SARS-CoV-2/pathogenicity , Animals , Antibody-Dependent Enhancement/physiology , Cell Line , Cricetinae , Humans , Real-Time Polymerase Chain Reaction , Receptors, IgG/genetics
20.
Emerg Microbes Infect ; 10(1): 2016-2029, 2021 Dec.
Article in English | MEDLINE | ID: covidwho-1493580

ABSTRACT

ABSTRACTA COVID-19 vaccine that can give early protection is needed to eliminate the viral spread efficiently. Here, we demonstrate the development of a nanoparticle vaccine candidate, REVC-128, in which multiple trimeric spike ectodomains with glycine (G) at position 614 were multimerized onto a nanoparticle. In-vitro characterization of this vaccine confirms its structural and antigenic integrity. In-vivo immunogenicity evaluation in mice indicates that a single dose of this vaccine induces potent serum neutralizing antibody titre at two weeks post-immunization. This is significantly higher than titre caused by trimeric spike protein without nanoparticle presentation. The comparison of serum binding to spike subunits between animals immunized by a spike with and without nanoparticle presentation indicates that nanoparticle prefers the display of spike RBD (Receptor-Binding Domain) over S2 subunit, likely resulting in a more neutralizing but less cross-reactive antibody response. Moreover, a Syrian golden hamster in-vivo model for the SARS-CoV-2 virus challenge was implemented two weeks post a single dose of REVC-128 immunization. The results showed that vaccination protects hamsters against the SARS-CoV-2 virus challenge with evidence of steady body weight, suppressed viral loads and alleviation of tissue damage for protected animals, compared with ∼10% weight loss, high viral loads and tissue damage in unprotected animals. Furthermore, the data showed that vaccine REVC-128 is thermostable at up to 37°C for at least 4 weeks. These findings, along with a history of safety for protein vaccines, suggest that the REVC-128 is a safe, stable and efficacious single-shot vaccine to give the earliest protection against SARS-CoV-2 infection.


Subject(s)
COVID-19 Vaccines/immunology , COVID-19/prevention & control , Nanoparticles/chemistry , SARS-CoV-2 , Animals , Antibodies, Neutralizing , Antibodies, Viral/blood , Antibody Formation , COVID-19 Vaccines/administration & dosage , Cricetinae , Humans , Immunization , Immunization Schedule , Immunogenicity, Vaccine , Mesocricetus , Mice , Spike Glycoprotein, Coronavirus , Vaccination , Viral Load
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