CD4(+)CXCL13(+) exhausted T cells drive immune microenvironment divergence in synchronous double primary lung adenocarcinoma with different degrees of invasiveness.

BACKGROUND: Synchronous double primary lung adenocarcinoma (sDPLA) is a distinct subtype of lung adenocarcinoma characterized by the co-existence of two independent lesions in the same patient. We conducted the first comprehensive analysis of the immune microenvironment of sDPLA lesions with different degrees of invasiveness to examine immune evolution during early lung adenocarcinoma progression. METHODS: In total, 10 sDPLA patients undergoing synchronous surgical resection were enrolled in the study. The minimally invasive adenocarcinoma (MIA) and invasive adenocarcinoma (IAC) lesions were analyzed by next-generation sequencing (NGS) or single-cell RNA sequencing (scRNA-seq), focusing on T/natural killer (NK) cell subsets. The key cell subsets and signaling pathways were identified using Mfuzz clustering, CellChat, and Monocle, and validated by multiplex immunofluorescence and flow cytometry. RESULTS: We identified 11 T/NK cell subsets, among which the CD4(+) exhausted T (Tex) cell CXCL13 population was significantly enriched in the IAC lesions. These cells exhibited high expression of PD-1 and TIM-3, enhanced interactions with epithelial cells, and enrichment of the JAK-STAT and PI3K-AKT pathways, suggesting a central role in immune suppression. Validation confirmed the preferential accumulation of this subset of cells in the IAC tissues. CONCLUSIONS: This study found significant immune heterogeneity between sDPLA lesions with different degrees of invasiveness and identified CD4(+) Tex CXCL13 cells as key drivers of tumor immune progression. Our findings provide new insights into early immune evolution and may inform precision immunotherapy strategies.