Protection from SARS-CoV-2 Variants by MVAs expressing matched or mismatched S administered intranasally to mice

Figures

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  Fig. 1. Protective immunity to SARS-CoV-2 variants following IM vaccination with rMVA-W. a Timeline of K18-hACE2 mice vaccinated IM on days 0 and 21 with 2 × 107 PFU of MVA control (n = 24) or rMVA-W (n = 24). On day 35, the mice vaccinated with MVA and rMVA-W were each divided into 3 groups of n = 8 and each group was challenged with 104 TCID50 of CoV-W, -B, or -D and 5 mice from each group were humanely killed 5 days later. The remaining 3 mice from each group were followed for weight loss and morbidity. b Serum antibody binding to the Wuhan S RBD and c neutralization of pseudoviruses rVSV-W, -B and -D at 3 and 5 weeks after vaccination were carried out with all MVA vaccinated mice (n = 24) and all rMVA-W vaccinated mice (n = 24). d Time course of weight loss and e survival on days after challenge with CoV-W, CoV-B or CoV-D were determined for 3 mice from each challenge group. Recovery of SARS-CoV-2 from f lungs and g nasal turbinates on day 5 after challenge of 5 mice from each challenge group. Copies of sgRNAs N and S relative to 18 S RNA from h lungs and i nasal turbinates on day 5 of the same mice analyzed in f and g. D day, HK humanely killed. Statistics: for c one way ANOVA with…

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  Fig. 2. Duration of cross-protective immunity following IM vaccination with rMVA-W. a Timeline of K18-hACE2 mice vaccinated IM twice with 2 × 107 PFU of MVA control (n = 8) divided into groups of 2–3 or rMVA-W (n = 15) divided into groups of 5 and challenged with 104 TCID50 of CoV-W, -B, or -D approximately 9 months later. Mice were H.K. 5 days after challenge. b Binding of serum antibodies elicited by vaccination with rMVA-W to Wuhan S protein determined by ELISA (n = 15). c Neutralization of pseudoviruses rVSV-W, -B and -D by serum obtained at weeks 3, 5, and 41 (n = 15). d Weights of mice on day 5 after challenge relative to starting weights (MVA n = 2 or 3; rMVAs N = 5 per group). Recovery of SARS-CoV-2 from e lungs and f nasal turbinates on day 5 after challenge (n as in d). Copies of sgRNAs N and S relative to 18 S rRNA from g lungs and h nasal turbinates on day 5 after challenge (n as in e, f). Statistics: for D, F, H Welch’s t-test. *p ≤ 0.03; **p ≤ 0.002. Dotted line, limit of detection.

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  Fig. 3. Inhibition of early stages of variant SARS-CoV-2 infections in mice immunized IM with rMVA-W. a Timeline of K18-hACE2 mice vaccinated IM twice with 2 × 107 PFU of MVA control (n = 40) or rMVA-W (n = 40), divided into groups of 10 and challenged 2 weeks later with 104 TCID50 of CoV-W, -B, -D or 5 × 104 TCID50 of CoV-O. Half the mice were H.K. on day 2 and remainder on day 4. b Binding of serum antibodies to Wuhan RBD after first and second immunizations determined by ELISA (n = 40 per group). c Neutralization of pseudoviruses rVSV-W, -B, -D and -O by sera obtained after the first and second immunizations (n = 9 per group). Recovery of SARS-CoV-2 from d lungs and e nasal turbinates on days 2 (n = 5 per group) and 4 (n = 5 per group) after challenge. Copies of sgRNAs N and S relative to 18 S RNA from f lungs and g nasal turbinates on days 2 and 4 after challenge. (n = same as in d and e). Statistics: for c ANOVA with Tukey post-hoc; for e–g Mann-Whitney. *p ≤ 0.03; **p ≤ 0.002; ****p < 0.0001. Dotted line, limit of detection.

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  Fig. 4. Inhibition of variant SARS-CoV-2 infections in mice immunized IN with rMVA-W. a Timeline of K18-hACE2 mice vaccinated IN twice with 2 × 107 PFU of MVA control (n = 31) or rMVA-W (n = 32), divided into groups of 7 or 8 and 2 weeks later challenged with 104 TCID50 of CoV-W, -B, or -D or 5 × 104 TCID50 of CoV-O. Half the mice were H.K. on day 2 and remainder on day 4. b Serum antibody binding to Wuhan RBD determined by ELISA after the first and second immunizations. (n= MVA 31, rMVA 32). c Neutralization of pseudoviruses rVSV-W, -B, -D and -O by serum obtained after the first and second immunizations. (n = 8 per group). Recovery of SARS-CoV-2 from d lungs and e nasal turbinates on days 2 and 4 and f brain on day 4 after challenge. n = 3 or 4 per group. Copies of sgRNAs N and S relative to 18 S RNA from g lungs and h nasal turbinates on days 2 and 4 after challenge. Fold-reduction of sgRNAs in nasal turbinates on day 2 for mice immunized i IN and j IM. Data replotted from Figs. 3g and 4h. Statistics: for c ANOVA with Tukey post-hoc; for g, h Mann-Whitney. *p ≤ 0.03; **p ≤ 0.002; ****p < 0.0001. Dotted line, limit of detection.

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  Fig. 5. Neutralizing antibody responses following boosts with rMVAs expressing matched or mismatched S. a Timeline of matched prime and boost IM vaccinations of C57BL/6 mice with 2 × 107 PFU of MVA or rMVA-W, -D, and -O. (n = 10 per group). b Neutralization of pseudoviruses rVSV-W, -D and -O by serum obtained after the prime with rMVA-W, -D and -O and after the matched boost. n = 10 per group. c Neutralization of pseudoviruses by sera from mice sublethally infected with CoV-W, CoV-D or CoV-O in Supplementary Fig. 2. n = 3 to 5. d Binding of pooled serum from bleed 2 to Wuhan and Omicron S determined by ELISA. n = 3 per group. e Serum neutralizing titers of mice one day after receiving pooled serum IP from naive mice or mice immunized twice with MVA, rMVA-W or rMVA-O. n = 3. Recovery of f SARS-CoV-2 and g sgRNAs from the lungs at 4 days after challenge of passively immunized mice. h Timeline of C57BL/6 mice were immunized twice with rMVA-W and boosted with rMVA-W, -B, -D, or -O. n = 9 for rMVA-W and n = 10 for other rMVAs. i Neutralization of pseudoviruses rVSV-W, -B, -D and -O by serum obtained after the third immunization (bleed 4). n = 9 for rMVA-W and n = 10 for other rMVAs. j…

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  Fig. 6. Protection of mice immunized IM with rMVAs to matched or mismatched CoVs. a Timeline showing IM immunizations of K18-hACE2 mice (n = 9 per group) with 2×107 PFU of MVA, rMVA-W or rMVA-O and matched and mismatched challenges with 104 TCID50 of CoV-W and 5x104 TCID50 of CoV-O. Mice were H.K on day 2 (n = 5) and day 4 (n = 4). b Neutralization of pseudoviruses rVSV-W and -O by serum obtained after one and two IM immunizations with MVA, rMVA-W or rMVA-O. n = 9 per group. Recovery of SARS-CoV-2 from (c) lungs and (d) nasal turbinates on days 2 and 4 after challenge with CoV-W or CoV-O. Copies of sgRNAs N and S relative to 18 S RNA from (e) lungs and (f) nasal turbinates on days 2 and 4 after challenge. Statistics: for panels b and c Mann-Whitney; d-f ANOVA with Dunnet post-hoc. *p ≤ 0.03; **p ≤ 0.002; ****p < 0.0001. Dotted line, limit of detection.

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  Fig. 7. Protection of CoV-W and CoV-O challenged mice vaccinated IN with rMVA-O. a Timeline of IN immunizations of K18-hACE2 mice (n = 13 per group) with 2 × 107 PFU of MVA or rMVA-O and matched and mismatched challenges with 104 TCID50 of CoV-W and 5 × 104 TCID50 of CoV-O. On days 2 and 4, 5 mice of each group were H.K. and 3 were followed for weight loss. b Neutralization of pseudoviruses rVSV-W and -O by serum obtained after one and two IN immunizations with MVA or rMVA-O. n = 13 per group. c Weights of mice following challenge with CoV-O or CoV-W. n = 3 per group. Recovery of d virus and e sgRNAs in the lungs of mice challenged with CoV-W or CoV-O. n = 5 per group. Recovery of f virus and g sgRNAs in the nasal turbinates of mice challenged with CoV-W or CoV-O. n = 5 per group. Fold-reduction of sgRNAs in lungs h and i nasal turbinates on day 2 for mice immunized IN and IM. Statistics: b, d–g Mann-Whitney. Error bars, S.D.; *p ≤ 0.03; **p ≤ 0.002; ***p ≤ 0.0002; ****p < 0.0001. Dotted line, limit of detection.