Self-adjuvanting alpha-helical polypeptide simultaneously delivers neoantigen mRNAs and activates dendritic cells to eradicate tumors

mRNA-based vaccines have demonstrated tremendous success during the era of COVID-19, but their therapeutic potential for treating cancer, especially poorly immunogenic solid tumors, remains under-achieved. Herein we report a class of self-adjuvanting alpha-helical polypeptides that can markedly improve the antitumor efficacy of tumor neoantigen-encoding mRNAs. Ideal carriers for antigen-encoding mRNAs should not only facilitate antigen expression in DCs but also activate DCs and induce effective processing and presentation of expressed antigens. Herein we report the development of cationic alpha-helical polypeptides as a self-adjuvanting delivery vehicle for antigen-encoding mRNAs. The alpha-helical polypeptides facilitate intracellular delivery of mRNAs into dendritic cells (DCs) via temporary membrane disruption, simultaneously activate DCs by regulating NF-κB and IRF pathways, and improve the ability of DCs to process and present mRNA-encoded neoantigens. The cationic polypeptide condenses and stabilizes mRNAs, promotes internalization via its cell-penetrating property, and increases the expression of activation markers (CD86, MHCII, CD40, and CCR7) on DCs by upregulating NF-κB, IRF, and STING pathways. Molecular docking and simulation confirm the stable complexation between mRNA and alpha-helical polypeptides. Upon subcutaneous administration, the polyplex migrates to draining lymph nodes, where it transfects and activates DCs, eliciting a strong neoantigen-specific cytotoxic T lymphocyte (CTL) response in vivo. Compared to mRNA-SM102 lipid nanoparticles or Lipofectamine 3000 lipoplexes, the polyplex enhances antigen expression and presentation by DCs, leading to significantly improved priming of antigen-specific CD8⁺ T cells in vitro and in vivo. The polyplex achieves 83.3% and 33.3% tumor-free survival in E.G7-OVA lymphoma and 4T1 triple-negative breast cancer (TNBC), respectively. This is far exceeding the 0% tumor-free survival of conventional formulations. Polyplexes composed of alpha-helical L-PPOB50-G and mRNAs encoding 4T1 TNBC neoantigens elicit potent neoantigen-specific CD8⁺ T cell responses while showing no detectable toxicity in major organs. The polyplex also reprograms the immunosuppressive tumor microenvironment by enriching DCs, M1-phenotype CD86⁺ macrophages, and CD8⁺ T cells in tumors. We also observed increased PD-1 expression on intratumoral CD8⁺ T cells and PD-L1 on 4T1 tumor cells after polyplex treatment and demonstrated synergistic effects between the polyplex vaccine and anti-PD-1 therapy. Our polyplex system provides a facile and generalizable approach to developing robust mRNA-based cancer vaccines.