Boosting the immune response in COVID-19 vaccines via an Alum:CpG complex adjuvant

Selecting appropriate adjuvants is crucial for developing an effective vaccine. However, studies on the immune responses triggered by different adjuvants in COVID-19 inactivated vaccines are scarce. Herein, we evaluated the efficacy of Alum, CpG HP021, Alum combined with CpG HP021 (Alum/CpG), or MF-59 adjuvants with COVID-19 inactivated vaccines in K18-hACE2 mice, and compared the different immune responses between K18-hACE2 and BALB/c mice. In K18-hACE2 mice, the Alum/CpG group produced a 6.5-fold increase in anti-receptor-binding domain (RBD) IgG antibody titers compared to the Alum group, and generated a comparable level of antibodies even when the antigen amount was reduced by two-thirds, possibly due to the significant activation of germinal center (GC) structures in the central region of the spleen. Different adjuvants induced a variety of binding antibody isotypes. CpG HP021 and Alum/CpG were biased towards Th1/IgG2c, while Alum and MF-59 were biased toward Th2/IgG1. Cytokines IFN-γ, IL-2, and TNF-α were significantly increased in the culture supernatants of splenocytes specifically stimulated in the Alum/CpG group. The antibody responses in BALB/c mice were similar to those in K18-hACE2 mice, but with lower levels of neutralizing antibodies (NAbs). Notably, the Alum/CpG-adjuvanted inactivated vaccine induced a higher number of T cells secreting IFN-γ and IL-2, increased the percentage of effector memory T (TEM) cells among CD8⁺ T cells, and effectively protected K18-hACE2 mice from Delta variant challenge. Our results showed that Alum/CpG complex adjuvant significantly enhanced the immune response to inactivated COVID-19 antigens and could induce a long-lasting immune response.