A single intranasal or intramuscular immunization with chimpanzee adenovirus-vectored SARS-CoV-2 vaccine protects against pneumonia in hamsters

Abstract

The development of an effective vaccine against severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), the etiologic agent of coronavirus disease 2019 (COVID-19), is a global priority. Here, we compare the protective capacity of intranasal and intramuscular delivery of a chimpanzee adenovirus-vectored vaccine encoding a prefusion stabilized spike protein (chimpanzee adenovirus [ChAd]-SARS-CoV-2-S) in Golden Syrian hamsters. Although immunization with ChAd-SARS-CoV-2-S induces robust spike-protein-specific antibodies capable of neutralizing the virus, antibody levels in serum are higher in hamsters vaccinated by an intranasal compared to intramuscular route. Accordingly, against challenge with SARS-CoV-2, ChAd-SARS-CoV-2-S-immunized hamsters are protected against less weight loss and have reduced viral infection in nasal swabs and lungs, and reduced pathology and inflammatory gene expression in the lungs, compared to ChAd-control immunized hamsters. Intranasal immunization with ChAd-SARS-CoV-2-S provides superior protection against SARS-CoV-2 infection and inflammation in the upper respiratory tract. These findings support intranasal administration of the ChAd-SARS-CoV-2-S candidate vaccine to prevent SARS-CoV-2 infection, disease, and possibly transmission.

Keywords: ChAd vectored vaccine; SARS-CoV-2; Syrian hamster; intranasal immunization.

Graphical abstract

Figure 1

Development of the SARS-CoV-2 hamster model (A) Mean ± standard deviation (SD) weight loss or weight gain of uninfected (n = 3) or SARS-CoV-2 infected (n = 21). (B) Daily food intake of uninfected and infected hamsters. Data points for the uninfected hamsters (n = 3 per day) were recorded for 14 days and plotted. For infected hamsters, food intake 1–10 dpi was recorded (^∗∗∗∗p < 0.0001 and ^∗∗∗p < 0.001 by ANOVA with Dunnett’s multiple comparison against the uninfected hamsters). (C) Infectious virus titer was quantified by FFA from homogenates of the left lung lobe at indicated time points. Each dot is an individual hamster, and bars indicate median values (dotted line is the limit of detection). (D) Lung viral RNA was quantified in the left lung lobe at indicated time points after infection. Each dot is an individual hamster, and bars indicate median values (dotted line is the limit of detection). (E) Pathology score of the lungs from infected hamsters. <10% affected = 1, >10% but <50% = 2, and >50% = 3. Each lobe was scored, and the average score was plotted per animals. The solid line is the average score per day for RNA in situ hybridization (red line and dots) or inflammation (blue line and dots). (F) Representative images at 5× and 20× magnification of H&E staining of SARS-CoV-2-infected hamsters sacrificed at different time points after inoculation (n = 5 for 2 dpi, n = 3 for 3 dpi, n = 3 for 4 dpi, n = 5 for 5 dpi, n = 2 for 8 dpi, n = 3 for 14 dpi, and n = 3 for uninfected). The scale bar represents 1 mm. (G) Serum S-protein-specific IgG(H+L) responses in SARS-CoV-2-infected hamsters. Each color is a different day after infection. (C and D) Bars indicate median values, and dotted lines are the LOD of the assays. See also Figure S1.

Figure 2

Humoral immune response following IN and IM immunization (A–C) Anti-S-protein-specific serum IgG(H+L) titers in hamsters immunized IN with ChAd-control (A) or with ChAd-SARS-CoV-2-S IM (B) or IN (C). Each line is an individual animal. (D–F) Receptor binding domain (RBD)-specific serum IgM titers in hamsters immunized IN with ChAd-control (D) or with ChAd-SARS-CoV-2-S IM (E) or IN (F). Each line is an individual animal. (G and H) IC₅₀ values for S-protein-specific or RBD-specific IgG(H+L) (G) or IgG2/IgG3 (H) serum antibodies in hamsters vaccinated IM (blue symbols) or IN (red symbols) with ChAd-control (open symbols) or ChAd-SARS-CoV-2-S (closed symbols). ^∗∗∗∗p < 0.0001 and ^∗∗∗p < 0.001 by Mann-Whitney test with a Bonferroni correction for multiple comparisons. (I) SARS-CoV-2 serum neutralizing titer, measured by FRNT, in hamsters vaccinated IM or IN with ChAd-control or ChAd-SARS-CoV-2-S. ^∗∗∗∗p < 0.0001 and ^∗p < 0.05 by Mann-Whitney test with a Bonferroni correction for multiple comparisons. (G–I) Bars indicate median values, and dotted lines are the LOD of the assays. See also Figures S2, S3, and S7.

Figure 3

IN immunization offers superior protection against challenge with SARS-CoV-2 (A–D) 28 days after a single IM (blue symbols) or IN (red symbols) vaccination with ChAd-control (open symbols) or ChAd-SARS-CoV-2-S (closed symbols), hamsters were challenged with 2.5 × 10⁵ PFUs of SARS-CoV-2, and nasal swabs (A and B) and lungs (C and D) were collected for analysis of viral RNA levels by qPCR (A and C) and infectious virus by plaque assay (B and D). ^∗∗∗∗p < 0.0001, ^∗∗∗p < 0.001, ^∗∗p < 0.01, ^∗p < 0.05, and ns, not significant by Mann-Whitney test with a Bonferroni correction for multiple comparisons. (E) Mean ± SD of weight loss or gain in SARS-CoV-2-challenged hamsters (n = 4 per group). (F) SARS-CoV-2 N protein serum titer, measured by ELISA, in hamsters vaccinated IM or IN with ChAd-control or ChAd-SARS-CoV-2-S. (A–D and F) Bars indicate median values, and dotted lines are the limit of detection of the assays. See also Figures S4 and S7.

Figure 4

IN immunization offers superior protection against SARS-CoV-2 infection of the nasal epithelium (A) RNA in situ hybridization (ISH) for SARS-CoV-2 viral RNA in hamster lung sections. Representative images at 5× and 20× magnification of the ChAd-control (IM), ChAd-SARS-CoV-2-S (IM), and ChAd-SARS-CoV-2-S (IN) sections. The scale bar represents 1 mm. (B) Comparison of RNA-ISH staining between groups of hamsters. Each lobe was scored according to the following system, <10% RNA-positive = 1, >10% but <50% RNA-positive = 2, and >50% RNA-positive = 3, and the average score was plotted per animals. ^∗∗p < 0.01 and ns by Mann-Whitney U test with a Bonferroni correction for multiple comparisons. (C) Representative images of sagittal sections of hamster heads infected with SARS-CoV-2 for 2 days or uninfected control. RNA-ISH was performed on the sections, and SARS-CoV-2 viral RNA was detected in the nasal turbinate. (D) Detection of SARS-CoV-2 viral by RNA-ISH in sagittal sections of hamster heads from the immunized and SARS-CoV-2-challenged animals. See also Figures S5 and S6.

Figure 5

ChAd-SARS-CoV-2 immunization ameliorates inflammatory gene expression following SARS-CoV-2 challenge Inflammatory gene-expression (n = 22) was quantified by RT-PCR in RNA extracted from lung homogenates 3 dpi (primer and probe sequences are in Table S1). (A) Fold increase in gene expression for ChAd-control-immunized (IN in red and IM in black) and SARS-CoV-2-challenged hamsters. (B) ΔΔCt values for Ifit3, Irf7, Ccl5, Cxcl10, Ddx58, and Ccl3 in ChAd-control- (open symbols) and ChAd-SARS-CoV-2-S (closed symbols)-immunized and SARS-CoV-2-challenged animals 3 dpi. ns, ^∗∗∗∗p < 0.0001, ^∗∗∗p < 0.001, and ^∗∗p < 0.01 by one-way ANOVA with a Šidák correction for multiple comparisons. Each dot is an individual animal from two experiments. Bars indicate average values. See also Figure S7.