CXCL13-expressing CD4⁺ T cells coordinate the lymphocytes triad to promote the anti-tumor immunity in NSCLC

CD4⁺ T cells are indispensable for CD8⁺ T cells-mediated anti-tumor immunity, while little is known about how CD4⁺ T cells coordinate with other cells to promote CD8⁺ T cells activity. In this study, by coupling single-cell RNA sequencing (scRNA-seq) with multiplex immunohistochemistry staining (mIHC), CXCL13-expressing CD4⁺ T cells were explored to recruit CXCR5⁺ B cells to form CXCL13⁺CD4⁺ T cells:: CXCR5⁺ B cells:: CD8⁺ T cells triad, promoting the anti-tumor immunity and heralding a favorable prognosis. A CXCL13-expressing subset of CD4⁺ T cells was identified to be associated with better prognosis and tumor-reactive hallmarks. The further cell types interaction analysis revealed a specific interaction between CXCL13⁺CD4⁺ T cells and CXCR5⁺ B cells in comparison with other cell types. The mIHC verified that an elevated level of CXCR5⁺ B cells infiltration in the proximity of CXCL13⁺CD4⁺ T cells. The subsequent Entropy analysis, which characterizes the colocalization among more than two cell types, was employed to reveal the spatial distribution pattern. The results revealed that CXCL13⁺CD4⁺ T cells and CXCR5⁺ B cells co-localized with cytotoxic CD8⁺ T cells, forming a functional lymphocyte triad, whose existence heralded a better prognosis. Collectively, by integrating scRNA-seq with mIHC, we demonstrated that the recruitment of CXCR5⁺ B cells by neoantigen reacting to CXCL13⁺CD4⁺ T cells through CXCL13-CXCR5 signaling contributed to the lymphocyte triad, leading to an enhanced anti-tumor immunity and heralding a favorable prognosis in non-small-cell lung cancer (NSCLC).