Intranasal delivery of a chimpanzee adenovirus vector expressing a pre-fusion spike (BV-AdCoV-1) protects golden Syrian hamsters against SARS-CoV-2 infection

Abstract

We evaluated the immunogenicity and protective ability of a chimpanzee replication-deficient adenovirus vectored COVID-19 vaccine (BV-AdCoV-1) expressing a stabilized pre-fusion SARS-CoV-2 spike glycoprotein in golden Syrian hamsters. Intranasal administration of BV-AdCoV-1 elicited strong humoral and cellular immunity in the animals. Furthermore, vaccination prevented weight loss, reduced SARS-CoV-2 infectious virus titers in the lungs as well as lung pathology and provided protection against SARS-CoV-2 live challenge. In addition, there was no vaccine-induced enhanced disease nor immunopathological exacerbation in BV-AdCoV-1-vaccinated animals. Furthermore, the vaccine induced cross-neutralizing antibody responses against the ancestral strain and the B.1.617.2, Omicron(BA.1), Omicron(BA.2.75) and Omicron(BA.4/5) variants of concern. These results demonstrate that BV-AdCoV-1 is potentially a promising candidate vaccine to prevent SARS-CoV-2 infection, and to curtail pandemic spread in humans.

Keywords: COVID-19 vaccine; Chimpanzee Adenovirus Serotype 68; challenge study; golden Syrian hamsters; intranasal.

Figure 1

Construction and characterization of BV-AdCoV-1. (A) Schematic diagram of the DNA sequence coding for the trimeric prefusion spike (Pre-S). The extracellular domain of SARS-COV-2 spike protein (1-1201 aa) was fused to the T4 fibritin trimerization motif, and the original spike’s signal peptide was replaced by the Japanese encephalitis virus E protein signal peptide (JEV SP). Mutations in the furin cleavage site and proline substitutions (K986P, V987P) were introduced. (B) Western blot analysis of HEK 293 cell lysate and supernatant. (C) Pre-S protein elution profile on a calibrated Superdex 200 increase 10/300 GL. (D) Cryo-EM picture of purified pre-S protein.

Figure 2

Experimental study scheme and anti-SARS-CoV-2 antibody detection. (A) Schedule of vaccine administration and virus challenge. dpi (days post-infection). (B) Binding anti-spike IgG antibody in golden Syrian hamsters’ sera. The antibody level was determined by Multiplex ELISA. (C) Binding anti-S1 RBD IgG antibody in golden Syrian hamsters’ sera. The antibody levels were determined by Multiplex ELISA. (D) Anti-nucleocapsid IgG antibody in golden Syrian hamsters’ sera. The antibody level was determined by Multiplex ELISA. (E) Live neutralizing antibody titers in golden Syrian hamsters’ sera. The neutralizing antibody level was determined by live SARS-CoV-2 cytopathogenicity-based assay. ***p < 0.001.

Figure 3

Experimental study scheme and cross-neutralizing antibody titers. (A) Schedule of vaccine administration. (B) Binding anti-spike IgG antibody titers in golden Syrian hamsters’ sera. (C–E) Cross-neutralizing antibody titers in golden Syrian hamsters’ sera. *p < 0.05, **p < 0.01.

Figure 4

(A–E) IFN-γ ELISpots in golden Syrian hamsters’ splenocytes re-stimulated with: medium-only (negative control); PMA and ionomycin (positive control); and peptide pools from RBD, nucleocapsid (N) and spike (S). T-cell responses against SARS-CoV-2 antigens were evaluated using a hamster ELISpot IFN-γ kit at dpi 7. **p<0.01.

Figure 5

Body weight changes in golden Syrian hamsters after live challenge. Data presented as mean ± SD of values. *P<0.05 or **P<0.01 between Negative control and Saline. Intergroup statistical analysis was performed using a two-way ANOVA test, using the infected, saline-treated group as a reference.

Figure 6

Intranasal BV-AdCoV-1 vaccination protects golden Syrian hamsters from SARS-CoV-2 infection. (A) SARS-CoV-2 virus infection titers in the lungs of golden Syrian hamsters were defined as the virus 50% tissue culture infectious dose (TCID₅₀). (B) SARS-CoV-2 relative levels of RNA in the lungs of golden Syrian hamsters. (C) Lung histopathology scores in golden Syrian hamsters. Left lung slides were stained with Hematoxylin-Phloxine to visualize histomorphometric changes. Slides were scanned using the NanoZoomer Digital Pathology System C9600-02. **p < 0.01, ***p < 0.001, ****p < 0.0001.

Figure 7

Ratios of cytokine expression levels in the lungs of golden Syrian hamsters at dpi 3. The cytokine expression levels in the lungs of golden Syrian hamsters were determined by RT-qPCR at dpi 3. (A) Ratios of TNF-α cytokine expression levels in the lungs of golden Syrian hamsters at dpi 3, (B) Ratios of IFN-γ cytokine expression levels in the lungs of golden Syrian hamsters at dpi 3, (C) Ratios of IL-2 cytokine expression levels in the lungs of golden Syrian hamsters at dpi 3, (D) Ratios of IL-4 cytokine expression levels in the lungs of golden Syrian hamsters at dpi 3, (E) Ratios of IL-5 cytokine expression levels in the lungs of golden Syrian hamsters at dpi 3, (F) Ratios of IL-6 cytokine expression levels in the lungs of golden Syrian hamsters at dpi 3, (G) Ratios of IL-10 cytokine expression levels in the lungs of golden Syrian hamsters at dpi 3, (H)Ratios of IL-12p40 cytokine expression levels in the lungs of golden Syrian hamsters at dpi 3, (I) Ratios of IL-17 cytokine expression levels in the lungs of golden Syrian hamsters at dpi 3, (J) Ratios of IL-21 cytokine expression levels in the lungs of golden Syrian hamsters at dpi 3. ****p < 0.0001.

Figure 8

Ratios of cytokine expression levels in the lungs of golden Syrian hamsters at dpi 7. The cytokine expression levels in the lungs of golden Syrian hamsters were determined by RT-qPCR at dpi 7. (A) Ratios of TNF-α cytokine expression levels in the lungs of golden Syrian hamsters at dpi 7, (B) Ratios of IFN-γ cytokine expression levels in the lungs of golden Syrian hamsters at dpi 7, (C) Ratios of IL-2 cytokine expression levels in the lungs of golden Syrian hamsters at dpi 7, (D) Ratios of IL-4 cytokine expression levels in the lungs of golden Syrian hamsters at dpi 7, (E) Ratios of IL-5 cytokine expression levels in the lungs of golden Syrian hamsters at dpi 7, (F) Ratios of IL-6 cytokine expression levels in the lungs of golden Syrian hamsters at dpi 7, (G) Ratios of IL-10 cytokine expression levels in the lungs of golden Syrian hamsters at dpi 7, (H) Ratios of IL-12p40 cytokine expression levels in the lungs of golden Syrian hamsters at dpi 7, (I) Ratios of IL-17 cytokine expression levels in the lungs of golden Syrian hamsters at dpi 7, (J) Ratios of IL-21 cytokine expression levels in the lungs of golden Syrian hamsters at dpi 7. *p < 0.05, **p < 0.01, ***p < 0.001, ****p < 0.0001.