Adenovirus-Derived Virus-Like Particles Enable Rapid Encapsulation and Delivery of Functional Antigen Cargo

Abstract Adenoviral vectors have been widely validated as effective vaccine platforms for both infectious diseases and cancer due to their strong immunostimulatory properties. However, their application to personalized cancer vaccines and other time-sensitive antigens is limited by the prolonged production time required for conventional vector generation. Here, we developed an adenovirus-derived virus-like particle (Ad5VLP) platform that enables modular incorporation of antigen cargo without reconstruction of a complete antigen-encoding adenoviral genome. Ad5VLPs successfully packaged and delivered foreign mRNA, although mRNA-derived antigen expression remained insufficient to induce robust antigen-specific immunity in vivo. In contrast, direct encapsulation of protein or peptide antigens substantially improved intracellular antigen delivery and major histocompatibility complex (MHC) class I antigen presentation. Following subcutaneous prime-boost immunization in mice, antigen-loaded Ad5VLPs induced robust antigen-specific interferon-γ (IFN-γ) responses, with functional activity preferentially enriched within CD44hi CD8+ T cells and accompanied by effector-associated CD8+ T-cell differentiation. Importantly, robust cellular immune responses were also observed using an E0771 tumor neoantigen, demonstrating that the platform extends beyond the model ovalbumin (OVA) antigen. Protein-loaded Ad5VLPs also elicited antigen-specific and antivector humoral responses. Vaccination was well tolerated over the 14 day study period, with no substantial changes in body weight, clinical observations, serum markers of hepatic or renal injury and systemic inflammation, or broad splenic inflammatory transcriptional signatures. Together, these findings demonstrate that Ad5VLPs enable modular delivery of protein and peptide antigens and induce robust cellular immune responses, supporting further investigation for personalized cancer vaccination and other time-sensitive vaccine applications.