Immunogenic fusion proteins induce neutralizing SARS-CoV-2 antibodies in the serum and milk of sheep

Antigen-specific polyclonal immunoglobulins derived from the serum, colostrum, or milk of immunized ruminant animals have potential as scalable therapeutics for the control of viral diseases such as COVID-19. Enhancing the efficacy of vaccine antigens to induce robust and specific antibody responses remains central to developing highly effective formulations. The direct fusion of immunoglobulin (IgG) Fc domains or other immune-stimulating proteins to antigens has shown promise in several mammalian species but has not yet been tested and optimized in commercially-relevant ruminant species. Here we show that the immunization of sheep with fusions of the receptor binding domain (RBD) of SARS-CoV-2 to ovine IgG2a Fc domains promotes significantly higher levels of antigen-specific antibodies compared to native RBD or full-length spike antigens. This antibody population was shown to contain elevated levels of neutralizing antibodies that suppress binding between the RBD and soluble hACE2 receptors in vitro. The parallel evaluation of a second immune-stimulating fusion candidate, Granulocyte-macrophage colony-stimulating factor (GM-CSF), induced high neutralizing responses in select animals but narrowly missed achieving significance at the group level. Furthermore, we demonstrate that the antibodies induced by these fusion antigens are transferred from maternal serum into colostrum/milk. These antibodies also demonstrate cross-neutralizing activity against diverse SARS-CoV-2 variants including delta and omicron. Our findings highlight a new pathway for recombinant antigen design in ruminant animals with applications in immune milk production and animal health.