Dibenzocyclooctyne Conjugation Enhances Antigen Cross-Presentation and T-Cell Killing for Potent Cancer Vaccines

Cancer vaccines have gained considerable attention for tumor prevention and therapy; however, their clinical efficacy remains limited by insufficient activation of antigen-specific cytotoxic T lymphocytes (CTLs). This limitation is primarily due to the inability of conventional vaccine components to effectively promote dendritic-cell-mediated cross-presentation and activation. While adjuvants are often used to enhance this interaction, many conventional adjuvants suffer from poor biocompatibility and systemic toxicity, limiting their clinical utility. Here, we report a vaccine strategy that leverages the bioorthogonal click chemistry reagent dibenzocyclooctyne (DBCO) as an adjuvant-like immune enhancer to promote cross-presentation and a CTL response. DBCO modification to antigen significantly increased antigen uptake by dendritic cells <i>in vitro</i> and promoted MHC class I-mediated cross-presentation via activation of the NF-κB/iNOS signaling pathway. In murine tumor models, systemic administration of DBCO-conjugated protein vaccines elicited enhanced CD8⁺ T-cell activation, leading to improved tumor control and long-term immune memory. Furthermore, DBCO-modified protein vaccines significantly suppressed tumor metastasis when combined with anti-PD-1 treatment. Collectively, our findings extend the immunotherapeutic potential of click chemistry reagents and establish a promising platform for enhancing CTL-mediated cancer vaccine efficacy in clinical settings.