Omicron subvariant BA.5 efficiently infects lung cells.

The SARS-CoV-2 Omicron subvariants BA.1 and BA.2 exhibit reduced lung cell infection relative to previously circulating SARS-CoV-2 variants, which may account for their reduced pathogenicity. However, it is unclear whether lung cell infection by BA.5, which displaced these variants, remains attenuated. Here, we show that the spike (S) protein of BA.5 exhibits increased cleavage at the S1/S2 site and drives cell-cell fusion and lung cell entry with higher efficiency than its counterparts from BA.1 and BA.2. Increased lung cell entry depends on mutation H69Δ/V70Δ and is associated with efficient replication of BA.5 in cultured lung cells. Further, BA.5 replicates in the lungs of female Balb/c mice and the nasal cavity of female ferrets with much higher efficiency than BA.1. These results suggest that BA.5 has acquired the ability to efficiently infect lung cells, a prerequisite for causing severe disease, suggesting that evolution of Omicron subvariants can result in partial loss of attenuation.

SARS-CoV-2-specific immunoglobulin Y antibodies are protective in infected mice.

Safe, passive immunization methods are required against severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2) and its variants. Immunization of chickens with antigen is known to induce specific IgY antibodies concentrated in the egg yolk and has a good safety profile, high yield of IgY per egg, can be topically applied, not requiring parenteral delivery. Our data provide the first evidence of the prophylactic efficacy of Immunoglobulin Y antibodies against SARS-CoV-2 in mice. Lohmann hens were injected with recombinant SARS-CoV-2 RBD protein; IgY-Abs were extracted from the eggs and characterized using SDS-PAGE. Antiviral activity was evaluated using plaque reduction neutralization tests. In additional experiments, IgY-RBD efficacy was examined in mice sensitized to SARS-CoV-2 infection by transduction with Ad5-hACE2 (mild disease) or by using mouse-adapted virus (severe disease). In both cases, prophylactic intranasal administration of IgY-Abs reduced SARS-CoV-2 replication, and reduced morbidity, inflammatory cell infiltration, hemorrhage, and edema in the lungs and increased survival compared to control groups that received non-specific IgY-Abs. These results indicate that further evaluation of IgY-RBD antibodies in humans is warranted.

A Glycosylated RBD Protein Induces Enhanced Neutralizing Antibodies against Omicron and Other Variants with Improved Protection against SARS-CoV-2 Infection.

SARS-CoV-2 has mutated frequently since its first emergence in 2019. Numerous variants, including the currently emerging Omicron variant, have demonstrated high transmissibility or increased disease severity, posing serious threats to global public health. This study describes the identification of an immunodominant non-neutralizing epitope on SARS-CoV-2 receptor-binding domain (RBD). A subunit vaccine against this mutant RBD, constructed by masking this epitope with a glycan probe, did not significantly affect RBD's receptor-binding affinity or antibody-binding affinity, or its ability to induce antibody production. However, this vaccine enhanced the neutralizing activity of this RBD and its protective efficacy in immunized mice. Specifically, this vaccine elicited significantly higher-titer neutralizing antibodies than the prototypic RBD protein against Alpha (B.1.1.7 lineage), Beta (B.1.351 lineage), Gamma (P.1 lineage), and Epsilon (B.1.427 or B.1.429 lineage) variant pseudoviruses containing single or combined mutations in the spike (S) protein, albeit the neutralizing antibody titers against some variants were slightly lower than against original SARS-CoV-2. This vaccine also significantly improved the neutralizing activity of the prototypic RBD against pseudotyped and authentic Delta (B.1.617.2 lineage) and Omicron (B.1.1.529 lineage) variants, although the neutralizing antibody titers were lower than against original SARS-CoV-2. In contrast to the prototypic RBD, the mutant RBD completely protected human ACE2 (hACE2)-transgenic mice from lethal challenge with a prototype SARS-CoV-2 strain and a Delta variant without weight loss. Overall, these findings indicate that this RBD vaccine has broad-spectrum activity against multiple SARS-CoV-2 variants, as well as the potential to be effective and have improved efficacy against Omicron and other pandemic variants. IMPORTANCE Several SARS-CoV-2 variants have shown increased transmissibility, calling for a need to develop effective vaccines with broadly neutralizing activity against multiple variants. This study identified a non-neutralizing epitope on the receptor-binding domain (RBD) of SARS-CoV-2 spike protein, and further shielded it with a glycan probe. A subunit vaccine based on this mutant RBD significantly enhanced the ability of prototypic RBD against multiple SARS-CoV-2 variants, including the Delta and Omicron strains, although the neutralizing antibody titers against some of these variants were lower than those against original SARS-CoV-2. This mutant vaccine also enhanced the protective efficacy of the prototypic RBD vaccine against SARS-CoV-2 infection in immunized animals. In conclusion, this study identified an engineered RBD vaccine against Omicron and other SARS-CoV-2 variants that induced stronger neutralizing antibodies and protection than the original RBD vaccine. It also highlights the need to improve the effectiveness of current COVID-19 vaccines to prevent pandemic SARS-CoV-2 variants.

RBD-mRNA vaccine induces broadly neutralizing antibodies against Omicron and multiple other variants and protects mice from SARS-CoV-2 challenge.

Multiple SARS-CoV-2 variants are identified with higher rates of transmissibility or greater disease severity. Particularly, recent emergence of Omicron variant with rapid human-to-human transmission posts new challenges to the current prevention strategies. In this study, following vaccination with an mRNA vaccine encoding SARS-CoV-2 receptor-binding domain (RBD-mRNA), we detected serum antibodies that neutralized pseudoviruses expressing spike (S) protein harboring single or multiple mutations, as well as authentic SARS-CoV-2 variants, and evaluated its protection against SARS-CoV-2 infection. The vaccine induced durable antibodies that potently neutralized prototypic strain and B.1.1.7 lineage variant pseudoviruses containing N501Y or D614G mutations alone or in combination with a N439K mutation (B.1.258 lineage), with a L452R mutation (B.1.427 or B.1.429 lineage), or a L452R-E484Q double mutation (B.1.617.1 variant), although neutralizing activity against B.1.1.7 lineage variant containing 10 amino acid changes in the S protein was slightly reduced. The RBD-mRNA-induced antibodies exerted moderate neutralization against authentic B.1.617.2 and B.1.1.529 variants, and pseudotyped B.1.351 and P.1 lineage variants containing K417N/T, E484K, and N501Y mutations, the B.1.617.2 lineage variant harboring L452R, T478K, and P681R mutations, and the B.1.1.529 lineage variant containing 38 mutations in the S protein. Particularly, RBD-mRNA vaccine completely protected mice from challenge with a virulent mouse-adapted SARS-CoV-2 variant. Among these lineages, B.1.1.7, B.1.351, P.1, B.1.617.2, and B.1.1.529 belong to Alpha, Beta, Gamma, Delta, and Omicron variants, respectively. Our observations reveal that RBD-mRNA vaccine is promising and highlights the need to design novel vaccines with improved neutralization against current and future pandemic SARS-CoV-2 variants.

SARS-CoV-2 Omicron virus causes attenuated disease in mice and hamsters.

The recent emergence of B.1.1.529, the Omicron variant1,2, has raised concerns of escape from protection by vaccines and therapeutic antibodies. A key test for potential countermeasures against B.1.1.529 is their activity in preclinical rodent models of respiratory tract disease. Here, using the collaborative network of the SARS-CoV-2 Assessment of Viral Evolution (SAVE) programme of the National Institute of Allergy and Infectious Diseases (NIAID), we evaluated the ability of several B.1.1.529 isolates to cause infection and disease in immunocompetent and human ACE2 (hACE2)-expressing mice and hamsters. Despite modelling data indicating that B.1.1.529 spike can bind more avidly to mouse ACE2 (refs. 3,4), we observed less infection by B.1.1.529 in 129, C57BL/6, BALB/c and K18-hACE2 transgenic mice than by previous SARS-CoV-2 variants, with limited weight loss and lower viral burden in the upper and lower respiratory tracts. In wild-type and hACE2 transgenic hamsters, lung infection, clinical disease and pathology with B.1.1.529 were also milder than with historical isolates or other SARS-CoV-2 variants of concern. Overall, experiments from the SAVE/NIAID network with several B.1.1.529 isolates demonstrate attenuated lung disease in rodents, which parallels preliminary human clinical data.

The development of Nanosota-1 as anti-SARS-CoV-2 nanobody drug candidates.

Combating the COVID-19 pandemic requires potent and low-cost therapeutics. We identified a series of single-domain antibodies (i.e., nanobody), Nanosota-1, from a camelid nanobody phage display library. Structural data showed that Nanosota-1 bound to the oft-hidden receptor-binding domain (RBD) of SARS-CoV-2 spike protein, blocking viral receptor angiotensin-converting enzyme 2 (ACE2). The lead drug candidate possessing an Fc tag (Nanosota-1C-Fc) bound to SARS-CoV-2 RBD ~3000 times more tightly than ACE2 did and inhibited SARS-CoV-2 pseudovirus ~160 times more efficiently than ACE2 did. Administered at a single dose, Nanosota-1C-Fc demonstrated preventive and therapeutic efficacy against live SARS-CoV-2 infection in both hamster and mouse models. Unlike conventional antibodies, Nanosota-1C-Fc was produced at high yields in bacteria and had exceptional thermostability. Pharmacokinetic analysis of Nanosota-1C-Fc documented an excellent in vivo stability and a high tissue bioavailability. As effective and inexpensive drug candidates, Nanosota-1 may contribute to the battle against COVID-19.

Exponential increase in neutralizing and spike specific antibodies following vaccination of COVID-19 convalescent plasma donors.

BACKGROUND: With the recent approval of COVID-19 vaccines, recovered COVID-19 subjects who are vaccinated may be ideal candidates to donate COVID-19 convalescent plasma (CCP). CASE SERIES: Eleven recovered COVID-19 patients were screened to donate CCP. All had molecularly confirmed COVID-19, and all but one were antibody positive by chemiluminescence immunoassay (DiaSorin) prior to vaccination. All were tested again for antibodies 11-21 days after they were vaccinated (Pfizer/Moderna). All showed dramatic increases (~50-fold) in spike-specific antibody levels and had at least a 20-fold increase in the IC50 neutralizing antibody titer based on plaque reduction neutralization testing (PRNT). The spike-specific antibody levels following vaccination were significantly higher than those seen in any non-vaccinated COVID-19 subjects tested to date at our facility. CONCLUSION: Spike-specific and neutralizing antibodies demonstrated dramatic increases following a single vaccination after COVID-19 infection, which significantly exceeded values seen with COVID-19 infection alone. Recovered COVID-19 subjects who are vaccinated may make ideal candidates for CCP donation.