Single Immunization with Recombinant ACAM2000 Vaccinia Viruses Expressing the Spike and the Nucleocapsid Proteins Protects Hamsters against SARS-CoV-2-Caused Clinical Disease.

Increasing cases of SARS-CoV-2 breakthrough infections from immunization with current spike protein-based COVID-19 vaccines highlight the need to develop alternative vaccines using different platforms and/or antigens. In this study, we expressed SARS-CoV-2 spike and nucleocapsid proteins based on a novel vaccinia virus (VACV) ACAM2000 platform (rACAM2000). In this platform, the vaccinia virus host range and immunoregulatory gene E3L was deleted to make the virus attenuated and to enhance innate immune responses, and another host range gene, K3L, was replaced with a poxvirus ortholog gene, taterapox virus 037 (TATV037), to make virus replication competent in both hamster and human cells. Following a single intramuscular immunization, the rACAM2000 coexpressing the spike and nucleocapsid proteins induced significantly improved protection against SARS-CoV-2 challenge in comparison to rACAM2000 expressing the individual proteins in a hamster model, as shown by reduced weight loss and shorter recovery time. The protection was associated with reduced viral loads, increased neutralizing antibody titer, and reduced neutrophil-to-lymphocyte ratio. Thus, our study demonstrates that rACAM2000 expressing a combination of the spike and nucleocapsid antigens is a promising COVID-19 vaccine candidate, and further studies will investigate if the rACAM2000 vaccine candidate can induce a long-lasting immunity against infection by SARS-CoV-2 variants of concern.