The variety and functionality of current clinical vaccine adjuvants remain limited. Conventional aluminum-based adjuvants predominantly induce Th2-biased humoral immunity but exhibit a limited capacity to elicit Th1-mediated cellular immune responses, particularly tumor antigen-specific cytotoxic CD8 + T lymphocytes (CTLs), which are essential for effective cancer vaccine performance. Inspired by natural biomolecular condensates, we developed a versatile noncovalent protein self-assembly strategy distinct from traditional approaches requiring structural domain modifications or bifunctional crosslinkers. Our methodology employs amphiphilic molecules (sodium myristate/SMA and sodium dodecyl thiolate/SDT) as molecular bridges to mediate protein‒protein interactions through hydrophobic forces and disulfide bond formation. This process generates nanoscale protein condensate (PCD) vaccines with exceptional stability. As a novel adjuvant system, these synthetic condensates significantly enhance antigen cross-presentation by optimizing key parameters: antigen loading capacity, lymph node targeting, cytosolic delivery, and lysosomal escape. Consequently, they induce robust antigen-specific CTL responses and humoral immunity, demonstrating potent antitumor efficacy. Importantly, we found that the synthetic protein condensate (PCD) alone can act as a nanoadjuvant. By increasing mitochondrial membrane permeability, PCD induces mitochondrial DNA leakage into the cytosol, activating the cGAS‒STING pathway and promoting DC maturation. This safe and scalable platform eliminates the need for complex covalent modifications or genetic engineering, and it facilitates the design of diverse modular antigens, including neoantigens and viral antigens. Given its straightforward manufacturing process and superior immunogenicity, this synthetic PCD vaccine adjuvant has significant potential for clinical application and translation.
Mengyue Pei,Rong Xu,Chuangnian Zhang,Xiaoli Wang,Chen Li,Yunzhang Hu
Cancer immunotherapy is an important tumor therapy option for prevention and treatment of tumors and has attracted tremendous interests. However, the therapeutic outcomes are limited by insufficient antigen uptake and presentation by antigen-presenting cells such as dendritic cells (DCs). In this study, mannose-functionalized antigen nanoparticles with endosome escape activity were designed for targeted DCs, accelerated endosomal escape and enhanced MHC-I antigen presentation for cancer immunotherapy. Mannose was selected as DCs targeting ligand to enhance antigen uptake. Model antigen ovalbumin (OVA) was directly conjugated with mannose to obtain DCs targeting antigen, which was then complexed with polyethylenimine (PEI) through electrostatic interaction to form mannose-functionalized antigen nanoparticles (MAN-OVA/PEI NPs). Flow cytometry analysis revealed that the MAN-OVA/PEI NPs greatly increased antigen uptake by DCs compared with OVA/PEI NPs. Confocal laser scanning microscopy further demonstrated that MAN-OVA/PEI NPs enhanced cytosolic antigen release. Moreover, MAN-OVA/PEI NPs significantly promoted cytokine production and DCs maturation in vitro . More importantly, MAN-OVA/PEI NPs treated DCs exhibited enhanced cross-presentation to B3Z T cell hybridoma in vitro . This work suggests that mannose-functionalized antigen nanoparticles provide a versatile delivery vehicle for targeted DCs, accelerated endosomal escape and enhanced MHC-I antigen presentation for cancer immunotherapy.
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