Advancing subunit vaccine efficacy against ricin toxin with a novel DNA hydrogel adjuvant tandem assembly by CpG motifs.

The development of effective countermeasures against ricin toxin, a potent biological threat, remains a pressing public health challenge. Subunit vaccines based on the ricin toxin A subunit offer a favorable safety profile but often suffer from limited immunogenicity, necessitating advanced adjuvants to elicit protective immunity. Here, we report a novel injectable CpG-motif-based hydrogel that serves as both adjuvant and delivery vehicle for a mutated and truncated RTA antigen. This CpG-gel adjuvant, synthesized via rolling circle amplification, incorporated with guanine-rich sequences that could self-assemble into stable G-quadruplex networks to form a structurally robust scaffold with flower-like microarchitecture. Notably, the CpG-gel significantly outperformed conventional CpG 1826 by enhancing antigen persistence and promoting sustained immune activation, leading to more potent and durable humoral and cellular responses. In murine studies, the CpG-gel elicited robust high titers of antigen-specific IgG antibodies, achieved complete protection against lethal-dose RT challenge. These results demonstrated that the CpG-gel adjuvant could significantly enhances the immunogenicity of subunit vaccines, highlighting its potential for broad implications for next-generation vaccine development.