Ipsilateral or contralateral boosting of mice with mRNA vaccines confers equivalent immunity and protection against a SARS-CoV-2 Omicron strain (preprint)

Boosting with mRNA vaccines encoding variant-matched spike proteins has been implemented to mitigate their reduced efficacy against emerging SARS-CoV-2 variants. Nonetheless, in humans, it remains unclear whether boosting in the ipsilateral or contralateral arm with respect to the priming doses impacts immunity and protection. Here, we boosted K18-hACE2 mice with either monovalent mRNA-1273 (Wuhan-1 spike) or bivalent mRNA-1273.214 (Wuhan-1 + BA.1 spike) vaccine in the ipsilateral or contralateral leg relative to a two-dose priming series with mRNA-1273. Boosting in the ipsilateral or contralateral leg elicited equivalent levels of serum IgG and neutralizing antibody responses against Wuhan-1 and BA.1. While contralateral boosting with mRNA vaccines resulted in expansion of spike-specific B and T cells beyond the ipsilateral draining lymph node (DLN) to the contralateral DLN, administration of a third mRNA vaccine dose at either site resulted in similar levels of antigen-specific germinal center B cells, plasmablasts/plasma cells, T follicular helper cells and CD8+ T cells in the DLNs and the spleen. Furthermore, ipsilateral and contralateral boosting with mRNA-1273 or mRNA-1273.214 vaccines conferred similar homologous or heterologous immune protection against SARS-CoV-2 BA.1 virus challenge with equivalent reductions in viral RNA and infectious virus in the nasal turbinates and lungs. Collectively, our data show limited differences in B and T cell immune responses after ipsilateral and contralateral site boosting by mRNA vaccines that do not substantively impact protection against an Omicron strain.

Intestinal helminth infection impairs vaccine-induced T cell responses and protection against SARS-CoV-2 (preprint)

Although vaccines have reduced COVID-19 disease burden, their efficacy in helminth infection endemic areas is not well characterized. We evaluated the impact of infection by Heligmosomoides polygyrus bakeri (Hpb), a murine intestinal hookworm, on the efficacy of an mRNA vaccine targeting the Wuhan-1 spike protein of SARS-CoV-2. Although immunization generated similar B cell responses in Hpb-infected and uninfected mice, polyfunctional CD4+ and CD8+ T cell responses were markedly reduced in Hpb-infected mice. Hpb-infected and mRNA vaccinated mice were protected against the ancestral SARS-CoV-2 strain WA1/2020, but control of lung infection was diminished against an Omicron variant compared to animals immunized without Hpb infection. Helminth mediated suppression of spike-specific CD8+ T cell responses occurred independently of STAT6 signaling, whereas blockade of IL-10 rescued vaccine-induced CD8+ T cell responses. In mice, intestinal helminth infection impairs vaccine induced T cell responses via an IL-10 pathway and compromises protection against antigenically shifted SARS-CoV-2 variants.

Mucosal Adenoviral-vectored Vaccine Boosting Durably Prevents XBB.1.16 Infection in Nonhuman Primates (preprint)

Waning immunity and continued virus evolution have limited the durability of protection from symptomatic infection mediated by intramuscularly (IM)-delivered mRNA vaccines against COVID-19 although protection from severe disease remains high. Mucosal vaccination has been proposed as a strategy to increase protection at the site of SARS-CoV-2 infection by enhancing airway immunity, potentially reducing rates of infection and transmission. Here, we compared protection against XBB.1.16 virus challenge 5 months following IM or mucosal boosting in non-human primates (NHP) that had previously received a two-dose mRNA-1273 primary vaccine regimen. The mucosal boost was composed of a bivalent chimpanzee adenoviral-vectored vaccine encoding for both SARS-CoV-2 WA1 and BA.5 spike proteins (ChAd-SARS-CoV-2-S) and delivered either by an intranasal mist or an inhaled aerosol. An additional group of animals was boosted by the IM route with bivalent WA1/BA.5 spike-matched mRNA (mRNA-1273.222) as a benchmark control. NHP were challenged in the upper and lower airways 18 weeks after boosting with XBB.1.16, a heterologous Omicron lineage strain. Cohorts boosted with ChAd-SARS-CoV-2-S by an aerosolized or intranasal route had low to undetectable virus replication as assessed by levels of subgenomic SARS-CoV-2 RNA in the lungs and nose, respectively. In contrast, animals that received the mRNA-1273.222 boost by the IM route showed minimal protection against virus replication in the upper airway but substantial reduction of virus RNA levels in the lower airway. Immune analysis showed that the mucosal vaccines elicited more durable antibody and T cell responses than the IM vaccine. Protection elicited by the aerosolized vaccine was associated with mucosal IgG and IgA responses, whereas protection elicited by intranasal delivery was mediated primarily by mucosal IgA. Thus, durable immunity and effective protection against a highly transmissible heterologous variant in both the upper and lower airways can be achieved by mucosal delivery of a virus-vectored vaccine. Our study provides a template for the development of mucosal vaccines that limit infection and transmission against respiratory pathogens. Graphical abstract O_FIG O_LINKSMALLFIG WIDTH=166 HEIGHT=200 SRC="FIGDIR/small/565765v1_ufig1.gif" ALT="Figure 1"> View larger version (48K): org.highwire.dtl.DTLVardef@d49b9dorg.highwire.dtl.DTLVardef@347455org.highwire.dtl.DTLVardef@1c1a196org.highwire.dtl.DTLVardef@1579130_HPS_FORMAT_FIGEXP M_FIG C_FIG

A bivalent ChAd nasal vaccine protects against SARS-CoV-2 BQ.1.1 and XBB.1.5 infection and disease in mice and hamsters (preprint)

We previously described a nasally delivered monovalent adenoviral-vectored SARS-CoV-2 vaccine (ChAd-SARS-CoV-2-S, targeting Wuhan-1 spike [S]; iNCOVACC) that is currently used in India as a primary or booster immunization. Here, we updated the mucosal vaccine for Omicron variants by creating ChAd-SARS-CoV-2-BA.5-S, which encodes for a pre-fusion and surface-stabilized S protein of the BA.5 strain, and then tested monovalent and bivalent vaccines for efficacy against circulating variants including BQ.1.1 and XBB.1.5. Whereas monovalent ChAd-vectored vaccines effectively induced systemic and mucosal antibody responses against matched strains, the bivalent ChAd-vectored vaccine elicited greater breadth. However, serum neutralizing antibody responses induced by both monovalent and bivalent vaccines were poor against the antigenically distant XBB.1.5 Omicron strain and did not protect in passive transfer experiments. Nonetheless, nasally delivered bivalent ChAd-vectored vaccines induced robust antibody and spike-specific memory T cell responses in the respiratory mucosa, and conferred protection against WA1/2020 D614G and Omicron variants BQ.1.1 and XBB.1.5 in the upper and lower respiratory tracts of both mice and hamsters. Our data suggest that a nasally delivered bivalent adenoviral-vectored vaccine induces protective mucosal and systemic immunity against historical and emerging SARS-CoV-2 strains without requiring high levels of serum neutralizing antibody.

Development of SARS-CoV-2 mRNA vaccines encoding spike N-terminal and receptor binding domains (preprint)

With the success of mRNA vaccines against coronavirus disease 2019 (COVID-19), strategies can now focus on improving vaccine potency, breadth, and stability. We present the design and preclinical evaluation of domain-based mRNA vaccines encoding the wild-type spike-protein receptor-binding (RBD) and/or N-terminal domains (NTD). An NTD-RBD linked candidate vaccine, mRNA-1283, showed improved antigen expression, antibody responses, and stability at refrigerated temperatures (2-8{degrees}C) compared with the clinically available mRNA-1273, which encodes the full-length spike protein. In mice administered mRNA-1283 as a primary series, booster, or variant-specific booster, similar or greater immune responses and protection from viral challenge were observed against wild-type, beta, delta, or omicron (BA.1) compared with mRNA-1273 immunized mice, especially at lower vaccine dosages. These results support clinical assessment of mRNA-1283 (NCT05137236).

SARS-CoV-2 Omicron boosting induces de novo B cell response in humans (preprint)

The primary two-dose SARS-CoV-2 mRNA vaccine series are strongly immunogenic in humans, but the emergence of highly infectious variants necessitated additional doses of these vaccines and the development of new variant-derived ones. SARS-CoV-2 booster immunizations in humans primarily recruit pre-existing memory B cells (MBCs). It remains unclear, however, whether the additional doses induce germinal centre (GC) reactions where reengaged B cells can further mature and whether variant-derived vaccines can elicit responses to novel epitopes specific to such variants. Here, we show that boosting with the original SARS-CoV-2 spike vaccine (mRNA-1273) or a B.1.351/B.1.617.2 (Beta/Delta) bivalent vaccine (mRNA-1273.213) induces robust spike-specific GC B cell responses in humans. The GC response persisted for at least eight weeks, leading to significantly more mutated antigen-specific MBC and bone marrow plasma cell compartments. Interrogation of MBC-derived spike-binding monoclonal antibodies (mAbs) isolated from individuals boosted with either mRNA-1273, mRNA-1273.213, or a monovalent Omicron BA.1-based vaccine (mRNA-1273.529) revealed a striking imprinting effect by the primary vaccination series, with all mAbs (n=769) recognizing the original SARS-CoV-2 spike protein. Nonetheless, using a more targeted approach, we isolated mAbs that recognized the spike protein of the SARS-CoV-2 Omicron (BA.1) but not the original SARS-CoV-2 spike from the mRNA-1273.529 boosted individuals. The latter mAbs were less mutated and recognized novel epitopes within the spike protein, suggesting a naive B cell origin. Thus, SARS-CoV-2 boosting in humans induce robust GC B cell responses, and immunization with an antigenically distant spike can overcome the antigenic imprinting by the primary vaccination series.

Bivalent SARS-CoV-2 mRNA vaccines increase breadth of neutralization and protect against the BA.5 Omicron variant (preprint)

The emergence of SARS-CoV-2 variants in the Omicron lineage with large number of substitutions in the spike protein that can evade antibody neutralization has resulted in diminished vaccine efficacy and persistent transmission. One strategy to broaden vaccine-induced immunity is to administer bivalent vaccines that encode for spike proteins from both historical and newly-emerged variant strains. Here, we evaluated the immunogenicity and protective efficacy of two bivalent vaccines that recently were authorized for use in Europe and the United States and contain two mRNAs encoding Wuhan-1 and either BA.1 (mRNA-1273.214) or BA.4/5 (mRNA-1273.222) spike proteins. As a primary immunization series in BALB/c mice, both bivalent vaccines induced broader neutralizing antibody responses than the constituent monovalent vaccines (mRNA-1273 [Wuhan-1], mRNA-1273.529 [BA.1], and mRNA-1273-045 [BA.4/5]). When administered to K18-hACE2 transgenic mice as a booster at 7 months after the primary vaccination series with mRNA-1273, the bivalent vaccines induced greater breadth and magnitude of neutralizing antibodies compared to an mRNA-1273 booster. Moreover, the response in bivalent vaccine-boosted mice was associated with increased protection against BA.5 infection and inflammation in the lung. Thus, boosting with bivalent Omicron-based mRNA-1273.214 or mRNA-1273.222 vaccines enhances immunogenicity and protection against currently circulating SARS-CoV-2 strains.

Boosting with Omicron-matched or historical mRNA vaccines increases neutralizing antibody responses and protection against B.1.1.529 infection in mice (preprint)

The B.1.1.529 Omicron variant jeopardizes vaccines designed with early pandemic spike antigens. Here, we evaluated in mice the protective activity of the Moderna mRNA-1273 vaccine against B.1.1.529 before or after boosting with preclinical mRNA-1273 or mRNA-1273.529, an Omicron-matched vaccine. Whereas two doses of mRNA-1273 vaccine induced high levels of serum neutralizing antibodies against historical WA1/2020 strains, levels were lower against B.1.1.529 and associated with infection and inflammation in the lung. A primary vaccination series with mRNA-1273.529 potently neutralized B.1.1.529 but showed limited inhibition of historical or other SARS-CoV-2 variants. However, boosting with mRNA-1273 or mRNA-1273.529 vaccines increased serum neutralizing titers and protection against B.1.1.529 infection. Nonetheless, the levels of inhibitory antibodies were higher and viral burden and cytokines in the lung were slightly lower in mice given the Omicron-matched mRNA booster. Thus, in mice, boosting with mRNA-1273 or mRNA-1273.529 enhances protection against B.1.1.529 infection with limited differences in efficacy measured.

The SARS-CoV-2 B.1.1.529 Omicron virus causes attenuated infection and disease in mice and hamsters (preprint)

Despite the development and deployment of antibody and vaccine countermeasures, rapidly-spreading SARS-CoV-2 variants with mutations at key antigenic sites in the spike protein jeopardize their efficacy. The recent emergence of B.1.1.529, the Omicron variant1,2, which has more than 30 mutations in the spike protein, has raised concerns for escape from protection by vaccines and therapeutic antibodies. A key test for potential countermeasures against B.1.1.529 is their activity in pre-clinical rodent models of respiratory tract disease. Here, using the collaborative network of the SARS-CoV-2 Assessment of Viral Evolution (SAVE) program of the National Institute of Allergy and Infectious Diseases (NIAID), we evaluated the ability of multiple B.1.1.529 Omicron isolates to cause infection and disease in immunocompetent and human ACE2 (hACE2) expressing mice and hamsters. Despite modeling and binding data suggesting that B.1.1.529 spike can bind more avidly to murine ACE2, we observed attenuation of infection in 129, C57BL/6, and BALB/c mice as compared with previous SARS-CoV-2 variants, with limited weight loss and lower viral burden in the upper and lower respiratory tracts. Although K18-hACE2 transgenic mice sustained infection in the lungs, these animals did not lose weight. In wild-type and hACE2 transgenic hamsters, lung infection, clinical disease, and pathology with B.1.1.529 also were milder compared to historical isolates or other SARS-CoV-2 variants of concern. Overall, experiments from multiple independent laboratories of the SAVE/NIAID network with several different B.1.1.529 isolates demonstrate attenuated lung disease in rodents, which parallels preliminary human clinical data.

mRNA-1273 and Ad26.COV2.S vaccines protect against the B.1.621 variant of SARS-CoV-2 (preprint)

ABSTRACT Since the emergence of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) in 2019, viral variants with greater transmissibility or immune evasion properties have arisen, which could jeopardize recently deployed vaccine and antibody-based countermeasures. Here, we evaluated in mice and hamsters the efficacy of preclinical non-GMP Moderna mRNA vaccine (mRNA-1273) and the Johnson & Johnson recombinant adenoviral-vectored vaccine (Ad26.COV2.S) against the B.1.621 (Mu) South American variant of SARS-CoV-2, which contains spike mutations T95I, Y144S, Y145N, R346K, E484K, N501Y, D614G, P681H, and D950N. Immunization of 129S2 and K18-human ACE2 transgenic mice with mRNA-1273 vaccine protected against weight loss, lung infection, and lung pathology after challenge with B.1.621 or WA1/2020 N501Y/D614G SARS-CoV-2 strain. Similarly, immunization of 129S2 mice and Syrian hamsters with a high dose of Ad26.COV2.S reduced lung infection after B.1.621 virus challenge. Thus, immunity induced by mRNA-1273 or Ad26.COV2.S vaccines can protect against the B.1.621 variant of SARS-CoV-2 in multiple animal models.

Protective activity of mRNA vaccines against ancestral and variant SARS-CoV-2 strains (preprint)

SUMMARY Although mRNA vaccines prevent COVID-19, variants jeopardize their efficacy as immunity wanes. Here, we assessed the immunogenicity and protective activity of historical (mRNA-1273, designed for Wuhan-1 spike) or modified (mRNA-1273.351, designed for B.1.351 spike) preclinical Moderna mRNA vaccines in 129S2 and K18-hACE2 mice. Immunization with high or low dose formulations of mRNA vaccines induced neutralizing antibodies in serum against ancestral SARS-CoV-2 and several variants, although levels were lower particularly against the B.1.617.2 (Delta) virus. Protection against weight loss and lung pathology was observed with all high-dose vaccines against all viruses. Nonetheless, low-dose formulations of the vaccines, which produced lower magnitude antibody and T cell responses, and serve as a possible model for waning immunity, showed breakthrough lung infection and pneumonia with B.1.617.2. Thus, as levels of immunity induced by mRNA vaccines decline, breakthrough infection and disease likely will occur with some SARS-CoV-2 variants, suggesting a need for additional booster regimens.

Multivalent designed proteins protect against SARS-CoV-2 variants of concern (preprint)

Escape variants of SARS-CoV-2 are threatening to prolong the COVID-19 pandemic. To address this challenge, we developed multivalent protein-based minibinders as potential prophylactic and therapeutic agents. Homotrimers of single minibinders and fusions of three distinct minibinders were designed to geometrically match the SARS-CoV-2 spike (S) trimer architecture and were optimized by cell-free expression and found to exhibit virtually no measurable dissociation upon binding. Cryo-electron microscopy (cryoEM) showed that these trivalent minibinders engage all three receptor binding domains on a single S trimer. The top candidates neutralize SARS-CoV-2 variants of concern with IC50 values in the low pM range, resist viral escape, and provide protection in highly vulnerable human ACE2-expressing transgenic mice, both prophylactically and therapeutically. Our integrated workflow promises to accelerate the design of mutationally resilient therapeutics for pandemic preparedness.

In vivo monoclonal antibody efficacy against SARS-CoV-2 variant strains (preprint)

Rapidly-emerging variants jeopardize antibody-based countermeasures against SARS-CoV-2. While recent cell culture experiments have demonstrated loss of potency of several anti-spike neutralizing antibodies against SARS-CoV-2 variant strains1-3, the in vivo significance of these results remains uncertain. Here, using a panel of monoclonal antibodies (mAbs) corresponding to many in advanced clinical development by Vir Biotechnology, AbbVie, AstraZeneca, Regeneron, and Lilly we report the impact on protection in animals against authentic SARS-CoV-2 variants including WA1/2020 strains, a B.1.1.7 isolate, and chimeric strains with South African (B.1.351) or Brazilian (B.1.1.28) spike genes. Although some individual mAbs showed reduced or abrogated neutralizing activity against B.1.351 and B.1.1.28 viruses with E484K spike protein mutations in cell culture, low prophylactic doses of mAb combinations protected against infection in K18-hACE2 transgenic mice, 129S2 immunocompetent mice, and hamsters without emergence of resistance. Two exceptions were mAb LY-CoV555 monotherapy which lost all protective activity in vivo, and AbbVie 2B04/47D11, which showed partial loss of activity. When administered after infection as therapy, higher doses of mAb cocktails protected in vivo against viruses displaying a B.1.351 spike gene. Thus, many, but not all, of the antibody products with Emergency Use Authorization should retain substantial efficacy against the prevailing SARS-CoV-2 variant strains.