Dendritic cell-based vaccine prepared with recombinant Lactococcus lactis eukaryotic-prokaryotic dual expressing OVA enhances antitumor efficacy by both direct and cross-presentation

Abstract

Probiotic Lactococcus lactis (L.L) can act as an antigen delivery carrier to cross-present (XPT) exogenous antigens in dendritic cells (DCs). To optimize antigen XPT in a DC-based vaccine for improved antitumor efficacy, we produced recombinant L.L strains expressing the model antigen OVA via distinct methods: prokaryotic expression alone, eukaryotic expression alone, or dual prokaryotic-eukaryotic expression (designated pOVA-L.L, eOVA-L.L, and dOVA-L.L, respectively). These strains were used to deliver antigens into DCs. Although all three recombinant OVA-L.L strains significantly enhanced DCs' ability to promote OT-I cell proliferation (with dOVA-L.L-treated DCs (dOVA-L.L-DCs) showing the strongest effect), only dOVA-L.L-DCs demonstrated significantly more potent antitumor efficacy in the B16-OVA tumor mouse model, consistent with the in vitro data. Further investigation into the mechanisms underlying the enhanced antigen XPT and anti-tumor efficacy of dOVA-L.L-DCs revealed that pOVA-L.L and dOVA-L.L resided both inside and outside lysosomes, whereas eOVA-L.L was located exclusively outside lysosomes. Consequently, dOVA-L.L-DCs could XPT higher amounts of MHC I-OVA peptide complex on their surface. Furthermore, these DCs could also directly present lysosome-derived OVA peptide-MHC II complexes to stimulate antigen-specific CD4⁺ T cell responses in vivo. Collectively, these mechanisms induced more persistent cellular immune responses than either of the other two recombinant L.L-DCs. In conclusion, we developed an enhanced antigen delivery system using probiotic food-grade bacteria. This system promotes the prolonged expression of exogenous antigens and significantly enhances the XPT capacity of DC vaccines for tumor immunotherapy.