Abstract
BACKGROUND: Cancer in the cervical region of the uterus is a major health issue for women. A live bacterial vector therapeutic vaccine is genetically engineered to stimulate a targeted immune response by integrating the target antigen gene into a live, attenuated vector. Listeria has become a focal point for bacterial vaccine vectors, owing to its distinctive intracellular parasitism and ability to process and present antigens. Antibiotic-resistant plasmids in vaccines present a dual risk, including the promotion of antibiotic resistance and in vivo instability. The implementation of a balanced lethal system can effectively mitigate both challenges. METHODS: We constructed therapeutic vaccinations for cervical cancer with dual-species Listeria vectors-attenuated Listeria monocytogenes (LM) with act A/plc B/dal/dat deletion (LM∆∆dd) and recombinant Listeria ivanovii (LI) whose hemolysin gene (ilo) is substituted with the hly gene of LM and with dal/dat deletion (LI∆ilo::hly∆∆dd). Recombinant Listeria-vectored therapeutic vaccines for cervical cancer, expressing HPV6/11/16/18 E6E7 proteins, were constructed based on the balanced lethal system. The basic biological characteristics of the vaccines were described. The vaccinations were evaluated for biosafety, stability, and immunogenicity in mice, and their efficacy was assessed in subcutaneous HPV16/18 E6/E7-expressing tumor models. RESULTS: This study demonstrates that recombinant Listeria expressing multivalent HPV E6/E7 proteins exhibit broad-spectrum anti-tumor effects against cervical cancer models. The study showed that the vaccination strains effectively expressed multivalent HPV antigen, exhibited plasmid stability in vitro, and had no significant impact on biochemical reactions and hemolytic activity. In mice, the vaccine strain was biologically safe, and the plasmid remained stable in vivo. A heterologous immunization strategy-LM prime-LI boost-LM final-was employed to immunize the mice, resulting in a robust, antigen-specific cellular immune response. In the subcutaneous HPV16/18 E6/E7-expressing tumor models, this vaccination protocol achieved complete tumor cure rates of 30% and 40%, respectively, and significantly extended the median survival time. CONCLUSIONS: Our objective was to evaluate the efficacy of the vaccine against subcutaneous HPV16/18 E6/E7-expressing tumor models, enhance its safety profile, and co-express multivalent HPV antigens to target a broader range of cervical cancer types.