Interferon Regulatory Factor 4 Recruits Immature B Cells to Signal Tertiary Lymphoid Structure Immaturity and Progression of Clear Cell Renal Cell Carcinoma

Background: Tertiary lymphoid structures (TLSs), organized clusters of immune cells within non-lymphoid tissues, significantly influence tumor progression and therapeutic response. However, their prognostic relevance and underlying regulatory mechanisms in clear cell renal cell carcinoma (ccRCC) remain insufficiently characterized. Methods: We integrated transcriptomic and clinical data from 928 ccRCC patients to construct a TLS-related prognostic RiskScore using machine learning algorithms. TLS maturation heterogeneity was characterized via immunohistochemistry and multiplex immunofluorescence analyses. The functional role of interferon regulatory factor 4 (IRF4), a key regulator within the TLS gene network, was investigated using in vitro assays. Single-cell RNA sequencing (scRNA-seq) and spatial transcriptomics were employed to dissect the involvement of IRF4 in TLS formation and maturation. Results: The derived TLS-associated signature RiskScore, comprising CCL22, LOXL1, LIPA, ADAM8, and SAA1, effectively stratified patients into distinct prognostic groups and showed robust associations with clinical parameters, tumor microenvironment (TME) features, and predicted immunotherapy responses. Functional assays demonstrated that IRF4 significantly enhanced the malignant phenotype of 786-O and 769-P ccRCC cells. Clinically, elevated IRF4 expression independently predicted worse patient outcomes, characterized by a predominance of immature TLS phenotypes, reduced TLS density, and diminished CD8⁺ T cell infiltration. Mechanistically, scRNA-seq analyses revealed that active IRF4 signaling was predominantly confined to immature B cell states and was inversely associated with TLS maturation trajectories. Spatial transcriptomics further confirmed IRF4 enrichment within TLS regions, notably spatially segregated from high endothelial venules (HEVs) and mature TLS compartments. Conclusion: In conclusion, this study establishes a robust TLS-related prognostic signature for ccRCC and elucidates the mechanistic role of IRF4 in promoting TLS immaturity and immune dysfunction. By potentially recruiting immature B cells while impairing their maturation, IRF4 contributes to an ineffective anti-tumor immune landscape, offering a promising target for therapeutic intervention.