Intranasal or airborne transmission-mediated delivery of an attenuated SARS-CoV-2 protects Syrian hamsters against new variants.

Detection of secretory antibodies in the airway is highly desirable when evaluating mucosal protection by vaccines against a respiratory virus, such as the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). We show that intranasal delivery of an attenuated SARS-CoV-2 (Nsp1-K164A/H165A) induces both mucosal and systemic IgA and IgG in male Syrian hamsters. Interestingly, either direct intranasal immunization or airborne transmission-mediated delivery of Nsp1-K164A/H165A in Syrian hamsters offers protection against heterologous challenge with variants of concern (VOCs) including Delta, Omicron BA.1, BA.2.12.1 and BA.5. Vaccinated animals show significant reduction in both tissue viral loads and lung inflammation. Similarly attenuated viruses bearing BA.1 and BA.5 spike boost variant-specific neutralizing antibodies in male mice that were first vaccinated with modified vaccinia virus Ankara vectors (MVA) expressing full-length WA1/2020 Spike protein. Together, these results demonstrate that our attenuated virus may be a promising nasal vaccine candidate for boosting mucosal immunity against future SARS-CoV-2 VOCs.

Neutralizing and protective murine monoclonal antibodies to the hemagglutinin of influenza H5 clades 2.3.2.1 and 2.3.4.4.

BACKGROUND: Highly pathogenic avian H5 influenza viruses have spread and diversified genetically and antigenically into multiple clades and subclades. Most isolates of currently circulating H5 viruses are in clade 2.3.2.1 or 2.3.4.4. METHODS: Panels of murine monoclonal antibodies (mAbs) were generated to the influenza hemagglutinin (HA) of H5 viruses from the clade 2.3.2.1 H5N1 vaccine virus A/duck/Bangladesh/19097/2013 and the clade 2.3.4.4 H5N8 vaccine virus A/gyrfalcon/Washington/41088-6/2014. Antibodies were selected and characterized for binding, neutralization, epitope recognition, cross-reactivity with other H5 viruses, and the ability to provide protection in passive transfer experiments. RESULTS: All mAbs bound homologous HA in an ELISA format; mAbs 5C2 and 6H6 were broadly binding for other H5 HAs. Potently neutralizing mAbs were identified in each panel, and all neutralizing mAbs provided protection in passive transfer experiments in mice challenged with a homologous clade influenza virus. Cross-reacting mAb 5C2 neutralized a wide variety of clade 2.3.2.1 viruses, as well as H5 viruses from other clades, and also provided protection against heterologous H5 clade influenza virus challenge. Epitope analysis indicated that the majority of mAbs recognized epitopes in the globular head of the HA. The mAb 5C2 appeared to recognize an epitope below the globular head but above the stalk region of HA. CONCLUSIONS: The results suggested that these H5 mAbs would be useful for virus and vaccine characterization. The results confirmed the functional cross-reactivity of mAb 5C2, which appears to bind a novel epitope, and suggest the therapeutic potential for H5 infections in humans with further development.

Distinct in vitro and in vivo neutralization profiles of monoclonal antibodies elicited by the receptor binding domain of the ancestral SARS-CoV-2.

Broadly neutralizing antibodies against severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) variants are sought to curb coronavirus disease 2019 (COVID-19) infections. Here we produced and characterized a set of mouse monoclonal antibodies (mAbs) specific for the ancestral SARS-CoV-2 receptor binding domain (RBD). Two of them, 17A7 and 17B10, were highly potent in microneutralization assay with 50% inhibitory concentration (IC50 ) ≤135 ng/mL against infectious SARS-CoV-2 variants, including G614, Alpha, Beta, Gamma, Delta, Epsilon, Zeta, Kappa, Lambda, B.1.1.298, B.1.222, B.1.5, and R.1. Both mAbs (especially 17A7) also exhibited strong in vivo efficacy in protecting K18-hACE2 transgenic mice from the lethal infection with G614, Alpha, Beta, Gamma, and Delta viruses. Structural analysis indicated that 17A7 and 17B10 target the tip of the receptor binding motif in the RBD-up conformation. A third RBD-reactive mAb (3A6) although escaped by Beta and Gamma, was highly effective in cross-neutralizing Delta and Omicron BA.1 variants in vitro and in vivo. In competition experiments, antibodies targeting epitopes similar to these 3 mAbs were rarely enriched in human COVID-19 convalescent sera or postvaccination sera. These results are helpful to inform new antibody/vaccine design and these mAbs can be useful tools for characterizing SARS-CoV-2 variants and elicited antibody responses.

Intranasal delivery of a rationally attenuated SARS-CoV-2 is immunogenic and protective in Syrian hamsters.

Few live attenuated severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) vaccines are in pre-clinical or clinical development. We seek to attenuate SARS-CoV-2 (isolate WA1/2020) by removing the polybasic insert within the spike protein and the open reading frames (ORFs) 6-8, and by introducing mutations that abolish non-structural protein 1 (Nsp1)-mediated toxicity. The derived virus (WA1-ΔPRRA-ΔORF6-8-Nsp1K164A/H165A) replicates to 100- to 1000-fold-lower titers than the ancestral virus and induces little lung pathology in both K18-human ACE2 (hACE2) transgenic mice and Syrian hamsters. Immunofluorescence and transcriptomic analyses of infected hamsters confirm that three-pronged genetic modifications attenuate the proinflammatory pathways more than the removal of the polybasic cleavage site alone. Finally, intranasal administration of just 100 PFU of the WA1-ΔPRRA-ΔORF6-8-Nsp1K164A/H165A elicits robust antibody responses in Syrian hamsters and protects against SARS-CoV-2-induced weight loss and pneumonia. As a proof-of-concept study, we demonstrate that live but sufficiently attenuated SARS-CoV-2 vaccines may be attainable by rational design.