FLT3L combined with GM-CSF induced dendritic cells drive broad tumor-specific CD8(+) T cell responses and remodel the tumor microenvironment to enhance anti-tumor efficacy.

BACKGROUND: Dendritic cells (DCs) play a crucial role in anti-tumor immunity by capturing, processing, and presenting tumor antigens to T cells, making DC-based immunotherapy a promising approach for cancer treatment. However, the most commonly used clinical strategy still relies on inducing DCs in vitro using granulocyte-macrophage colony-stimulating factor (GM-CSF) and interleukin-4 (IL - 4) (GM/IL4-DCs), which often results in a heterogeneous cell population with suboptimal anti-tumor function. Here, we compared DCs generated by co-stimulating with FMS-like tyrosine kinase 3 ligand (FLT3L) and GM-CSF (FL/GM-DCs) with the conventional GM/IL4-DCs. METHOD: To compare the functional differences of DCs induced by different methods, we conducted a comprehensive study. Mouse bone marrow cells were continuously cultured for 9 days in a FLT3L/GM-CSF-containing medium. After cell collection, we analyzed the composition, subpopulations, and status of FL/GM-DCs using flow cytometry and scRNA-seq. Flow cytometry was also used to assess their antigen presentation and ability to stimulate T cells. In vivo experiments were performed to examine their distribution, anti-tumor effects, and therapeutic responses in tumor models. Finally, combining scRNA-seq and scTCR-seq, we explored the mechanisms by which FL/GM-DCs reshape the tumor microenvironment. RESULTS: The results showed that FL/GM-DCs exhibited a unique subpopulation distribution, characterized by an abundance of conventional cDC subpopulations, and demonstrated enhanced cross-antigen presentation capabilities. Notably, FL/GM-DCs were able to induce a broader and more tumor-specific CD8(+) T cell response, effectively reshaping the tumor microenvironment by promoting the infiltration of cytotoxic T lymphocytes (CTLs) and reducing immunosuppressive components. In contrast, GM/IL4-DCs contained fewer cDC subpopulations, eliciting a weaker initial CD8(+) T cell response and yielding relatively inferior anti-tumor effects. CONCLUSION: In summary, FLT3L combined with GM-CSF induced DCs, through their unique subpopulation composition and functional state, can more effectively expand tumor-specific CD8(+) T cells and reshape the tumor microenvironment, thereby achieving superior immunotherapy outcomes. This study highlights the potential of FL/GM-DCs as a next-generation DC platform, paving the way for improved clinical translation of DC-based adoptive cancer immunotherapies.