Prime hAd5 Spike + Nucleocapsid Vaccination Induces Ten-Fold Increases in Mean T-Cell Responses in Phase 1 Subjects that are Sustained Against Spike Variants

Abstract

In response to the need for a safe, efficacious vaccine that provides broad immune protection against SARS-CoV-2 infection, we have developed a dual-antigen COVID-19 vaccine. The vaccine delivers both the viral spike (S) protein modified to increase cell-surface expression (S-Fusion) and the viral nucleocapsid (N) protein with an Enhanced T-cell Stimulation Domain (N-ETSD) to enhance MHC class I and II presentation and T-cell responses. The vaccine antigens are delivered using a human adenovirus serotype 5 (hAd5) platform with E1, E2b, and E3 regions deleted that has been shown in previous cancer vaccine studies to be effective in the presence of pre-existing hAd5 immunity. Here, we demonstrate the hAd5 S-Fusion + N-ETSD (hAd5 S + N) vaccine antigens when expressed by dendritic cells (DCs) of previously SARS-CoV-2-infected patients elicit Th1 dominant activation of autologous patient T cells, indicating the vaccine antigens have the potential for generating immune responses in patients previously infected or vaccinated. We further demonstrate that participants in our open-label Phase 1b study of the dual-antigen hAd5 S + N vaccine generate Th1 dominant S- and N-specific T cells after a single prime subcutaneous injection and that the magnitude of these responses were comparable to those seen for T cells from previously infected patients. We further present our in silico prediction of T-cell epitope HLA binding for both the first-wave SARS-CoV-2 ‘A’ strain and the K417N, E484K, and N501Y S as well as the T201I N variants that suggests T-cell responses to the hAd5 S + N vaccine will retain efficacy against these variants. These findings that the dual-antigen hAd5 S + N vaccine elicits SARS-CoV-2-relevant T-cell responses and that such cell-mediated protection is likely to be sustained against emerging variants supports the testing of this vaccine as a universal booster that would enhance and broaden existing immune protection conferred by currently approved S-based vaccines.

Comprehensive Analysis of CD4+ T Cell Response Cross-Reactive to SARS-CoV-2 Antigens at the Single Allele Level of HLA Class II

“…The BNT162b2 COVID-19 spike mRNA vaccine boosted the pre-existing SARS-CoV-2 spike cross-reactive T cells. Moreover, an adenovirus-based COVID-19 vaccine encoding both spike and nucleocapsid proteins elicited Th1 dominant responses after a single prime injection ( 40 ). A vaccine encoding all the spike, nucleocapsid, and membrane protein might re-activate pre-existing cross-reactive T cells and evoke secondary-like immune responses in more individuals than the only spike-encoding vaccine.…”

Peptidome Surveillance Across Evolving SARS-CoV-2 Lineages Reveals HLA Binding Conservation in Nucleocapsid Among Variants With Most Potential for T-Cell Epitope Loss in Spike

“…Motivated by vaccine approaches currently in trials that include both the S and N proteins ( 20 – 22 ), we investigated the relationship between HLA binder loss in S and N across all viral lineages in the data.…”

“…High Fraction of Lineages With the Most Binder Loss in S Show Conserved HLA Binding in N Motivated by vaccine approaches currently in trials that include both the S and N proteins (20)(21)(22), we investigated the relationship between HLA binder loss in S and N across all viral lineages in the data.…”

“…HLA presentation is an essential precursor to T-cell recognition, and recent results have demonstrated that immunodominant CD4+ T-cell epitopes specific to SARS-CoV-2 correlate with HLA binding promiscuity (14). We analyzed both the S and N peptidomes, motivated by next generation vaccines currently in development that include both S and N antigens (20)(21)(22) and findings discussed above indicating broad T-cell response across structural proteins.…”

Heterologous saRNA Prime, DNA Dual-Antigen Boost SARS-CoV-2 Vaccination Elicits Robust Cellular Immunogenicity and Cross-Variant Neutralizing Antibodies

“…To address the need for a vaccine regimen that would be highly efficacious against predominating and emerging variants as well as distributable in currently underserved areas, we previously developed a next-generation human adenovirus serotype 5 (hAd5)-vectored dual-antigen spike (S) plus nucleocapsid (N) vaccine (AdS+N) (7,8) to leverage the resilience of cell-mediated immunity against variants. This vaccine, encoding Wuhan strain or 'wild type' (wt) SARS-CoV-2 S and modified with a fusion sequence (S-Fusion) to enhance cell-surface expression (7,8), as well as N modified with an Enhanced T-cell Stimulation Domain (N-ETSD) (9) for increased MHC class I and II stimulation (10)(11)(12), has been shown to elicit humoral and T-cell responses in mice (8), nonhuman primates (NHP) (7), and participants in Phase 1b trials (9). The AdS+N vaccine given as a subcutaneous (SC) prime with two oral boosts protected NHP from SARS-CoV-2 infection (7), and a single prime vaccination of clinical trial participants generated Tcell responses that were sustained against a series of variant S peptide sequences, including those for the B.1.351, B.1.1.7, P.1, and B.1.426 variants (9).…”