Simultaneous and sequential multi-species coronavirus vaccination

Although successful COVID-19 vaccines have been developed, multiple pathogenic coronavirus species exist, urging for development of multi-species coronavirus vaccines. Here we developed prototype LNP-mRNA vaccine candidates against SARS-CoV-2 (Delta variant), SARS-CoV and MERS-CoV, and test how multiplexing of these LNP-mRNAs can induce effective immune responses in animal models. A triplex scheme of LNP-mRNA vaccination induced antigen-specific antibody responses against SARS-CoV-2, SARS-CoV and MERS-CoV, with a relatively weaker MERS-CoV response in this setting. Single cell RNA-seq profiled the global systemic immune repertoires and the respective transcriptome signatures of multiplexed vaccinated animals, which revealed a systemic increase in activated B cells, as well as differential gene expression signatures across major adaptive immune cells. Sequential vaccination showed potent antibody responses against all three species, significantly stronger than simultaneous vaccination in mixture. These data demonstrated the feasibility, antibody responses and single cell immune profiles of multi-species coronavirus vaccination. The direct comparison between simultaneous and sequential vaccination offers insights on optimization of vaccination schedules to provide broad and potent antibody immunity against three major pathogenic coronavirus species. One sentence summaryMultiplexed mRNA vaccination in simultaneous and sequential modes provide broad and potent immunity against pathogenic coronavirus species.

Heterotypic vaccination responses against SARS-CoV-2 Omicron BA.2

The Omicron sub-lineage BA.2 of SARS-CoV-2 has recently become dominant across many areas in the world in the on-going waves of COVID-19. Compared to the ancestral/wild-type (WT) virus, Omicron lineage variants, both BA.1 and BA.2, contain high number of mutations, especially in the spike protein, causing significant immune escape that leads to substantial reduction of vaccine and antibody efficacy. Because of this antigenic drift, BA.2 exhibited differential resistance profile to monoclonal antibodies than BA.1. Thus, it is important to understand whether the immunity elicited by currently available vaccines are effective against the BA.2 subvariant. We directly tested the heterotypic vaccination responses against Omicron BA.2, using vaccinated serum from animals receiving WT- and variant-specific mRNA vaccine in lipid nanoparticle (LNP) formulations. Omicron BA.1 and BA.2 antigen showed similar reactivity to serum antibodies elicited by two doses of WT, B.1.351 and B.1.617 LNP-mRNAs. Neutralizing antibody titers of B.1.351 and B.1.617 LNP-mRNA were ~2-fold higher than that of WT LNP-mRNA. Both homologous boosting with WT LNP-mRNA and heterologous boosting with BA.1 LNP-mRNA substantially increased waning immunity of WT vaccinated mice against both BA.1 and BA.2 subvariants. The BA.1 LNP-mRNA booster was ~3-fold more efficient than WT LNP-mRNA at elevating neutralizing antibody titers of BA.2. Together, these data provided a direct preclinical evaluation of WT and variant-specific LNP-mRNAs in standard two-dose and as boosters against BA.1 and BA.2 subvariants.

Potent and specific human monoclonal antibodies against SARS-CoV-2 Omicron variant by rapid mRNA immunization of humanized mice

The Omicron variant (B.1.1.529) of SARS-CoV-2 rapidly becomes dominant globally. Its extensive mutations confer severe efficacy reduction to most of existing antibodies or vaccines. Here, we developed RAMIHM, a highly efficient strategy to generate fully human monoclonal antibodies (mAbs), directly applied it with Omicron-mRNA immunization, and isolated three potent and specific clones against Omicron. Rapid mRNA immunization elicited strong anti-Omicron antibody response in humanized mice, along with broader anti-coronavirus activity. Customized single cell BCR sequencing mapped the clonal repertoires. Top-ranked clones collectively from peripheral blood, plasma B and memory B cell populations showed high rate of Omicron-specificity (93.3%) from RAMIHM-scBCRseq. Clone-screening identified three highly potent neutralizing antibodies that have low nanomolar affinity for Omicron RBD, and low ng/mL level IC50 in neutralization, more potent than majority of currently approved or authorized clinical RBD-targeting mAbs. These lead mAbs are fully human and ready for downstream IND-enabling and/or translational studies.

SARS-CoV-2 Omicron-specific mRNA vaccine induces potent and broad antibody responses in vivo

The Omicron variant of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) has high transmissibility and recently swept the globe. Due to the extensive number of mutations, this variant has high level of immune evasion, which drastically reduced the efficacy of existing antibodies and vaccines. Thus, it is important to test an Omicron-specific vaccine, evaluate its immune response against Omicron and other variants, and compare its immunogenicity as boosters with existing vaccine designed against the reference wildtype virus (WT). Here, we generated an Omicron-specific lipid nanoparticle (LNP) mRNA vaccine candidate, and tested its activity in animals, both alone and as a heterologous booster to existing WT mRNA vaccine. Our Omicron-specific LNP-mRNA vaccine elicited strong and specific antibody response in vaccination-naive mice. Mice that received two-dose WT LNP-mRNA, the one mimicking the commonly used Pfizer/Moderna mRNA vaccine, showed a >40-fold reduction in neutralization potency against Omicron variant than that against WT two weeks post second dose, which further reduced to background level >3 months post second dose. As a booster shot for two-dose WT mRNA vaccinated mice, a single dose of either a homologous booster with WT LNP-mRNA or a heterologous booster with Omicron LNP-mRNA restored the waning antibody response against Omicron, with over 40-fold increase at two weeks post injection as compared to right before booster. Interestingly, the heterologous Omicron LNP-mRNA booster elicited neutralizing titers 10-20 fold higher than the homologous WT booster against the Omicron variant, with comparable titers against the Delta variant. All three types of vaccination, including Omicron mRNA alone, WT mRNA homologous booster, and Omicron heterologous booster, elicited broad binding antibody responses against SARS-CoV-2 WA-1, Beta, and Delta variants, as well as other Betacoronavirus species such as SARS-CoV, but not Middle East respiratory syndrome coronavirus (MERS-CoV). These data provided direct proof-of-concept assessments of an Omicron-specific mRNA vaccination in vivo, both alone and as a heterologous booster to the existing widely-used WT mRNA vaccine form.