mRNA vaccines encoding membrane-anchored receptor-binding domains of SARS-CoV-2 mutants induce strong humoral responses and can overcome immune imprinting (preprint)

To address the limitations of whole-spike COVID vaccines, we explored mRNA vaccines encoding membrane-anchored receptor-binding domain (RBD-TMs), each a fusion of a variant RBD, the transmembrane (TM) and cytoplasmic tail (CT) fragments of the SARS-CoV-2 spike protein. In naive mice, RBD-TM mRNA vaccines against ancestral SARS-CoV-2, Beta, Delta, Delta-plus, Kappa, Omicron BA.1 or BA.5, all induced strong humoral responses against the target RBD. Multiplex surrogate viral neutralization (sVNT) assays indicated broad neutralizing activity against a range of variant RBDs. In the setting of a heterologous boost, against the background of exposure to ancestral whole spike vaccines, sVNT studies suggested that RBD-TM vaccines were able to overcome the detrimental effects of immune imprinting. Omicron BA.1 and BA.5 RBD-TM booster vaccines induced serum antibodies with 12 and 22-fold higher neutralizing activity against the target RBD than their equivalent whole spike variants. Boosting with BA.1 or BA.5 RBD-TM provided good protection against more recent variants including XBB and XBB.1.5. Each RBD-TM mRNA is 28% of the length of its whole-spike equivalent. This advantage will enable tetravalent mRNA vaccines to be developed at well-tolerated doses of formulated mRNA.

Uncovering strain- and age- dependent differences in innate immune response to SARS-CoV-2 infection in nasal epithelia using combined short and long-read scRNA-seq (preprint)

Assessing the impact of SARS-CoV-2 variants on host immune systems is crucial with the continuous arrival of novel variants. However, there is a lack of reported studies utilizing single-cell RNA-sequencing (scRNA-seq) to elucidate the age-dependent host-viral interactions with different SARS-CoV-2 strains. Therefore, we employed Full-Length Transcriptome Sequencing (FLT-Seq) combining Illumina and Oxford Nanopore Technologies (ONT) with 10X 3-prime single cell sequencing to investigate host transcriptomic changes during wild-type (WT) and Alpha-strain SARS-CoV-2 infections within air-liquid-interface (ALI)-cultured human nasal epithelial cells from adults and adolescents. The results revealed increased innate immune responses in cells with lower viral loads which were lacking in cells with higher viral load. Alpha-infections showed heightened expression of genes related to interferon (IFN)-responses and protein-folding compared with WT, implying the increased immune response and endoplasmic reticulum stress with the variant of a higher transmission rate. ACE2 expression was elevated in infected populations of secretory and goblet cells with high IFN-stimulation and a subpopulation of ciliated cells but was lacking in bystander cells and other infected-cell types, despite detectable IFN-stimulation and regardless of strain. We used long-read sequencing to identify differential transcript usage (DTU) of IFN-response and translational genes, including IFI27, RPL4 and RPL15 (padj < 0.05) in infected cells compared with control, and consistent DTU of RPL15 and MX1 between Alpha and WT strain-infected cells in various cell-types. Together, these results reveal the dynamic transcriptional/translational/post-translational activity upon SARS-CoV-2 infection, which appeared to be age and strain-dependent. Overall, this study highlights the complexity of cell-type, age and viral-strain-dependent host epithelial responses to SARS-CoV-2.

Broad immunity to SARS-CoV-2 variants of concern mediated by a SARS-CoV-2 receptor-binding domain protein vaccine (preprint)

The SARS-CoV-2 global pandemic has fuelled the generation of vaccines at an unprecedented pace and scale. However, many challenges remain, including: the emergence of vaccine-resistant mutant viruses, vaccine stability during storage and transport, waning vaccine-induced immunity, and concerns about infrequent adverse events associated with existing vaccines. Here, we report on a protein subunit vaccine comprising the receptor-binding domain (RBD) of the ancestral SARS-CoV-2 spike protein, dimerised with an immunoglobulin IgG1 Fc domain. These were tested in conjunction with three different adjuvants: a TLR2 agonist R4-Pam2Cys, an NKT cell agonist glycolipid alpha-Galactosylceramide, or MF59 squalene oil-in-water adjuvant. Each formulation drove strong neutralising antibody (nAb) responses and provided durable and highly protective immunity against lower and upper airway infection in mouse models of COVID-19. We have also developed an RBD-human IgG1 Fc vaccine with an RBD sequence of the highly immunoevasive beta variant (N501Y, E484K, K417N). This beta variant RBD vaccine, combined with MF59 adjuvant, induced strong protection in mice against the beta strain as well as the ancestral strain. Furthermore, when used as a third dose booster vaccine following priming with whole spike vaccine, anti-sera from beta-RBD-Fc immunised mice increased titres of nAb against other variants including alpha, delta, delta+, gamma, lambda, mu, and omicron BA.1 and BA.2. These results demonstrated that an RBD-Fc protein subunit/MF59 adjuvanted vaccine can induce high levels of broad nAbs, including when used as a booster following prior immunisation of mice with whole ancestral-strain Spike vaccines. This vaccine platform offers a potential approach to augment some of the currently approved vaccines in the face of emerging variants of concern, and it has now entered a phase I clinical trial.