A Self-assembly Carrier-Free Nanovaccine based on Fluorinated CpG and Neoantigen Peptides for Hepatocellular Carcinoma Immunotherapy

Neoantigen peptides evade central thymic tolerance due to their absence in normal tissues, making neoantigen vaccines a leading strategy for personalized cancer immunotherapy. Although nanoparticle-based delivery systems are widely used to enhance neoantigen immunogenicity, traditional nanovaccines often incorporate multiple adjuvants and inert carriers, which could hinder efficient antigen delivery and raise biosafety concerns. Herein, we report a carrier-free neoantigen nanovaccine (CpG-2'F/Neo_WD), assembled via fluorination-induced self-assembly for hepatocellular carcinoma (HCC) immunotherapy. This nanovaccine is formed by the co-assembly of fluorinated CpG (CpG-2'F) with the HCC neoantigen peptide (Neo_WD), yielding nanoparticles of approximately 69 nm with a high Neo_WD loading efficiency of 67.18 wt%. In vitro studies demonstrate that the nanovaccine potently facilitates dendritic cell uptake and promotes endosomal/lysosomal escape, enhancing antigen cross-presentation efficiency by 2.05-fold compared to the antigen group. In vivo, by efficiently targeting lymph nodes (LNs), the nanovaccine orchestrated both innate and adaptive immune responses against cancer cells, leading to the pronounced expansion of antigen-specific CD8⁺ T cells and memory T cells. In both prophylactic and therapeutic Hepa1-6 HCC models, this nanovaccine effectively suppressed the progression of poorly immunogenic HCC. Therefore, this study presents a facile, safe and potently immunogenic carrier-free nanovaccine platform for HCC immunotherapy.