Targeting Fusobacterium Nucleatum With Mrna-LNP Vaccines Restores Pd-1 Sensitivity and Reprograms the Tumor Microenvironment in Esophageal Squamous Cell Carcinoma

Background F. nucleatum (Fn) has been implicated in the poor response to immunotherapy in esophageal squamous cell carcinoma (ESCC). In a humanized Fn-colonized ESCC mouse model, Fn promoted tumor progression and induced resistance to PD-1 blockade, suggesting a potential role for Fn in modulating the immune response and contributing to immune evasion in ESCC. Methods Two mRNA-LNP vaccines targeting the Fn membrane protein FomA were developed: one encoding the full-length FomA protein (seq1) and another encoding only the extracellular repeat domains (seq2). The efficacy of these vaccines, administered alone or in combination with anti-PD-1 therapy, was evaluated in Fn-colonized ESCC mice. Metabolomic analysis and single-cell RNA sequencing were performed to assess alterations in the tumor microenvironment following treatment. Results The combination of Fn mRNA vaccines with anti-PD-1 therapy significantly inhibited tumor growth compared to the control groups, with seq1 and seq2 vaccines both effectively reversing Fn-induced PD-1 resistance. Histological examination showed increased infiltration of CD8+ T cells and reduced tumor-associated macrophages, indicating enhanced anti-tumor immunity. Metabolomic analysis revealed alterations in lipid metabolism and amino acid biosynthesis pathways, suggesting an improved metabolic profile conducive to immune activation. Single-cell transcriptomic analyses revealed that the vaccine treatment induced substantial reprogramming of immune cell activity and metabolic pathways within the tumor microenvironment, thereby augmenting the anti-tumor immune response. Conclusion Fn plays a critical role in inducing PD-1 resistance in ESCC, and targeted vaccination against Fn using mRNA-LNP technology effectively restores PD-1 sensitivity. This approach offers a novel therapeutic strategy to overcome microbial-driven immune resistance and modulate the tumor microenvironment to enhance the efficacy of immunotherapy in ESCC.