Synergistic effect of AddaS03 and poly I:C adjuvants on antibody titers and Th1-biased T cell responses following RBD vaccination in BALB/c mice.
BACKGROUND The COVID-19 pandemic caused over seven million deaths, and disease control relied largely on mass immunization campaigns. Although diverse vaccine platforms were used, most formulations targeted the Spike protein because of its central role in virus entry. Persistent viral evolution and global vaccine inequity highlight the need for alternative strategies, particularly for low- and middle-income countries. In this scenario, recombinant protein-based vaccines are attractive due to low cost, established manufacturing pipelines and well-characterized safety profiles. The receptor binding domain (RBD) of Spike is an appealing antigen due to its relatively simple structure, proven immunogenicity - especially in multimerized formats - and because it harbors most neutralizing antibody epitopes. Protein-based vaccines, however, rely on the use of adjuvants to optimize antigen-specific immunity. METHODS We used an RBD dimer derived from the ancestral Wuhan strain as a well-characterized protein to evaluate the humoral and cellular immunogenicity of combined AddaS03 and Poly I:C adjuvants in BALB/c mice. RESULTS The adjuvant combination preserved the strong humoral responses typically induced by AddaS03 while conferring the Th1-skewing properties of Poly I:C, producing a synergistic effect that exceed those of each adjuvant alone. Specifically, the combination improved antibody affinity, increased the generation of antibody- secreting cells in spleen and bone marrow, and enhanced the induction of polyfunctional, cytokine-producing CD4+ T cells. Additionally, we mapped and characterized three class II T cell epitopes within RBD: peptide 6 (351-YAWNRKRISNCVADYSV-367), peptide 19 (442-SKVGGNYNYLYRLFRK-458) and peptide 20 (449-YNYLYRLFRKSNLKPFE-465). CONCLUSIONS Combining AddaS03 and Poly I:C adjuvants is a potent strategy to modulate and enhance immune responses elicited by an RBD-based vaccine.