ABSTRACT
The SARS-CoV-2 Omicron variant and its sublineages show pronounced viral escape from neutralizing antibodies elicited by vaccination or prior SARS-CoV-2 variant infection owing to over 30 amino acid alterations within the spike (S) glycoprotein. We and others have recently reported that breakthrough infection of vaccinated individuals with Omicron sublineages BA.1 and BA.2 are associated with distinct patterns of cross-neutralizing activity against SARS-CoV-2 variants of concern (VOCs). BA.2 breakthrough infection mediated overall stronger cross-neutralization of BA.2 and its descendants (BA.2.12.1, BA.4, and BA.5) compared to BA.1 breakthrough infection. Here we characterized the effect of Omicron BA.4/BA.5 S glycoprotein exposure on the magnitude and breadth of the neutralizing antibody response upon breakthrough infection in vaccinated individuals and in mice upon booster vaccination. We show that immune sera from triple mRNA-vaccinated individuals with subsequent Omicron BA.4/BA.5 breakthrough infection display broad and robust neutralizing activity against Omicron BA.1, BA.2, BA.2.12.1, and BA.4/BA.5. Administration of a prototypic BA.4/BA.5-adapted mRNA booster vaccine to mice following SARS-CoV-2 wild-type strain-based primary immunization is associated with similarly broad neutralizing activity. Immunization of naive mice with a bivalent mRNA vaccine (wild-type + Omicron BA.4/BA.5) induces strong and broad neutralizing activity against Omicron VOCs and previous variants. These findings suggest that when administered as boosters, mono- and bivalent Omicron BA.4/BA.5-adapted vaccines may enhance neutralization breadth, and in a bivalent format may also have the potential to confer protection to individuals with no pre-existing immunity against SARS-CoV-2.
Subject(s)
Severe Acute Respiratory Syndrome , Breakthrough PainABSTRACT
A safe and effective vaccine against COVID-19 is urgently needed in quantities sufficient to immunise large populations. We report the preclinical development of two BNT162b vaccine candidates, which contain lipid-nanoparticle (LNP) formulated nucleoside-modified mRNA encoding SARS-CoV-2 spike glycoprotein-derived immunogens. BNT162b1 encodes a soluble, secreted, trimerised receptor-binding domain (RBD-foldon). BNT162b2 encodes the full-length transmembrane spike glycoprotein, locked in its prefusion conformation (P2 S). The flexibly tethered RBDs of the RBD-foldon bind ACE2 with high avidity. Approximately 20% of the P 2S trimers are in the two-RBD down, one-RBD up state. In mice, one intramuscular dose of either candidate elicits a dose-dependent antibody response with high virus-entry inhibition titres and strong TH1 CD4+ and IFN{gamma}+ CD8+ T-cell responses. Prime/boost vaccination of rhesus macaques with BNT162b candidates elicits SARS-CoV-2 neutralising geometric mean titres 8.2 to 18.2 times that of a SARS-CoV-2 convalescent human serum panel. The vaccine candidates protect macaques from SARS-CoV-2 challenge, with BNT162b2 protecting the lower respiratory tract from the presence of viral RNA and with no evidence of disease enhancement. Both candidates are being evaluated in phase 1 trials in Germany and the United States. BNT162b2 is being evaluated in an ongoing global, pivotal Phase 2/3 trial (NCT04380701, NCT04368728).
Subject(s)
COVID-19ABSTRACT
To contain the coronavirus disease 2019 (COVID-19) pandemic, a safe and effective vaccine against the new severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2) is urgently needed in quantities sufficient to immunise large populations. In this study, we report the design, preclinical development, immunogenicity and anti-viral protective effect in rhesus macaques of the BNT162b2 vaccine candidate. BNT162b2 contains an LNP-formulated nucleoside-modified mRNA that encodes the spike glycoprotein captured in its prefusion conformation. After expression of the BNT162b2 coding sequence in cells, approximately 20% of the spike molecules are in the one-RBD up, two-RBD down state. Immunisation of mice with a single dose of BNT162b2 induced dose level-dependent increases in pseudovirus neutralisation titers. Prime-boost vaccination of rhesus macaques elicited authentic SARS-CoV-2 neutralising geometric mean titers 10.2 to 18.0 times that of a SARS-CoV-2 convalescent human serum panel. BNT162b2 generated strong TH1 type CD4+ and IFNy+ CD8+ T-cell responses in mice and rhesus macaques. The BNT162b2 vaccine candidate fully protected the lungs of immunised rhesus macaques from infectious SARS-CoV-2 challenge. BNT162b2 is currently being evaluated in a global, pivotal Phase 2/3 trial (NCT04368728).
Subject(s)
Coronavirus Infections , COVID-19ABSTRACT
Objectives: SARS-CoV-2 infection is the cause of a worldwide pandemic, currently with limited therapeutic options. It is characterised by being highly contagious and nasal mucosa appears to be the primary site with subsequent spread to the lungs and elsewhere. BromAc (Bromelain & Acetylcysteine) has been described to disrupt glycoproteins by the synchronous breakage of glycosidic linkages and disulphide bonds. The spike protein of SARS-CoV-2 is an attractive target as it is essential for binding to the ACE2 receptor in host cells and is formed of glycoprotein and disulphide bridges for stabilisation. Hence, we sought to determine whether BromAc has activity on the spike and envelope protein specific to SARS-CoV-2 virus. Design: Gel electrophoresis analysis was carried out on recombinant spike and envelope proteins that were treated with a range of concentrations of single agents and BromAc. For UV analysis of disulfide bonds reduction, both spike and envelope protein were treated with Acetylcysteine with the determination of loss of disulfide bonds. Results: Recombinant spike and envelope SARS-CoV-2 protein were fragmented by BromAc whilst single agents had minimal effect. Spike and envelope proteins disulphide bonds were reduced by Acetylcysteine. Conclusion: BromAc disintegrates the spike and envelope protein from SARS-CoV-2 and may render it non-infective. In vitro tests on live virus have been encouraging and clinical testing through nasal administration in patients with early SARS-CoV-2 infection is imminent.
Subject(s)
COVID-19ABSTRACT
Convalescent plasma from SARS-CoV-2 infected individuals and monoclonal antibodies were shown to potently neutralize viral and pseudoviral particles carrying the S glycoprotein. However, a non-negligent proportion of plasma samples from infected individuals as well as S-specific monoclonal antibodies were reported to be non-neutralizing despite efficient interaction with the S glycoprotein in different biochemical assays using soluble recombinant forms of S or when expressed at the cell surface. How neutralization relates to binding of S glycoprotein in the context of viral particles remains to be established. Here we developed a pseudovirus capture assay (VCA) to measure the capacity of plasma samples or antibodies immobilized on ELISA plates to bind to membrane-bound S glycoproteins from SARS-CoV-2 expressed at the surface of lentiviral particles. By performing VCA and neutralization assays we observed a strong correlation between these two parameters. However, while we found that plasma samples unable to capture viral particles did not neutralize, capture did not guarantee neutralization, indicating that the capacity of antibodies to bind to the S glycoprotein at the surface of viral particles is required but not sufficient to mediate neutralization. Altogether, our results highlights the importance of better understanding the inactivation of S by plasma and neutralizing antibodies.
Subject(s)
Severe Acute Respiratory SyndromeABSTRACT
A novel severe acute respiratory (SARS)-like coronavirus (SARS-CoV-2) is responsible for the current global coronavirus disease 2019 (COVID-19) pandemic, infecting millions of people and causing hundreds of thousands of deaths. The viral entry of SARS-CoV-2 depends on an interaction between the receptor binding domain of its trimeric Spike glycoprotein and the human angiotensin converting enzyme 2 (ACE2) receptor. A better understanding of the Spike/ACE2 interaction is still required to design anti-SARS-CoV-2 therapeutics. Here, we investigated the degree of cooperativity of ACE2 within both the SARS-CoV-2 and the closely related SARS-CoV-1 membrane-bound S glycoproteins. We show that there exist differential inter-protomer conformational transitions between both Spike trimers. Interestingly, the SARS-CoV-2 spike exhibits a positive cooperativity for monomeric soluble ACE2 binding when compared to the SARS-CoV-1 spike, which might have more structural restrains. Our findings can be of importance in the development of therapeutics that block the Spike/ACE2 interaction.