Upper respiratory tract immunization with Pam2Cys-adjuvanted spike protein vaccine achieves sterilizing protection against SARS-CoV-2

Erica L Stewart^1,2,3Skye Stockdale^2,3Matt Johansen⁴Lachlan Smith³Sibel Alca³Joshua WC Maxwell^5,6Duc H Nguyen⁴Stefan Miemczyk⁴Guanshu Zhao⁴Stuart Turville⁷Jamie A Triccas^2,3Megan Steain^2,3Scott Napier Byrne³Philip Michael Hansbro⁴Richard James Payne^5,6Warwick J Britton^1,8*Anneliese Sophie Ashhurst^2,3*

• ¹Tuberculosis Research Program, Centenary Institute, The University of Sydney, Sydney, Australia
• ²Sydney Institute for Infectious Diseases and the Charles Perkins Centre, The University of Sydney, Sydney, Australia
• ³School of Medical Sciences, Faculty of Medicine and Health, The University of Sydney, Sydney, Australia
• ⁴Centre for Inflammation, Centenary Institute and University of Technology Sydney, Faculty of Science, School of Life Sciences, Sydney, Australia
• ⁵School of Chemistry, Faculty of Science, The University of Sydney, Sydney, Australia
• ⁶Australian Research Council Centre of Excellence for Innovations in Peptide and Protein Science, The University of Sydney, Sydney, Australia
• ⁷The Kirby Institute, University of New South Wales, Sydney, Australia
• ⁸Department of Clinical Immunology, Royal Prince Alfred Hospital, Camperdown, Australia

Injected COVID-19 vaccines protect against severe disease, but do not induce robust mucosal immune responses. Nasal vaccines offer the advantage of local immunity to block viral infection and transmission. Previously we showed immunization of a Pam2Cys-adjuvanted SARS-CoV-2 vaccine to the upper and lower respiratory tracts (URT/LRT) induced protective immune responses in the lungs. However, URT/LRT immunization is not representative of nasal vaccines for clinical use that exclusively target the URT. Here, we show that delivery to only the URT with Pam2Cys and spike protein effectively induced strong SARS-CoV-2 specific immune responses in the nasal mucosa. When delivered in a low volume so that vaccine exposure was limited to the URT, Pam2Cys/spike protein induced local SARS-CoV-2-specific Th17 cells and neutralizing antibodies to a similar level to inhaled vaccination reaching both the URT and LRT. We compared URT versus URT/LRT delivery as booster vaccinations following parenteral immunization and found that URT vaccination concentrated the immune response to the URT rather than the lungs. Importantly, URT immunization or boosting induced sterilizing immunity in K18-hACE2 mice challenged with homologous SARS-CoV-2. Thus, booster vaccination to the URT alone with Pam2Cys/spike achieved robust nasal immunity against SARS-CoV-2 and is a promising strategy for clinical development.