Multimodal single-cell and spatial profiling reveals altered T cell-mediated immunity and B-cell follicular architecture in non-metastatic lymph nodes of patients with aggressive non-small cell lung cancer
Regional lymph nodes (LNs) in the thoracic cavity serve as essential immunological hubs that coordinate humoral and cell-mediated responses against the development and progression of non-small cell lung cancer (NSCLC). To investigate immune dysregulation in the non-metastatic regional LNs of patients with aggressive NSCLC, we performed multimodal profiling on 36 LNs from 11 patients undergoing curative-intent resection including CITE-seq, scRNA-seq, and Imaging Mass Cytometry (IMC). Regional N1 LNs from patients with more aggressive disease (stage IB–IIIA) exhibited a significant enrichment of dysfunctional CD8? T cells and regulatory T cells (Tregs) compared to N2 LNs and LNs from patients with less aggressive disease (stage IA). These immune subsets were spatially co-localized with mature regulatory dendritic cells (mregDCs; CD1c?, TIM3?, LAMP3?), forming an immunosuppressive niche uniquely enriched in the N1 LNs of higher-stage patients. Concurrently, higher-stage N1 LNs contained larger number of “decorticated” B-cell follicles characterized by decreased encapsulation of the mantle zone layer surrounding the germinal centers. This mantle zone disorganization was associated with increased spatial niches involving Tregs, CD68? CD163? TIM3? Macrophages, CD163? TIM3dim Monocytic-Myeloid Derived Suppressor Cells (M-MDSC), plasma B cells, and a decrease in spatial niches involving CD4? T helper cells and fibroblastic reticular cells (FRCs). Together, our findings reveal parallel alterations in humoral and cell-mediated immunity within the regional LNs of patients with aggressive NSCLC. Overall design: Fine needle aspirate samples of lymph nodes, primary tumors, and normal lung tissue were collected from surgical specimens of patients undergoing curative-intent resection at Boston Medical Center. Mediastinal LN dissection was performed as per the Commission on Cancer guideline, removing, when possible, 3 N2 station LNs and at least 1 N1 station LN. Pathologic stage was determined per 8th edition of TNM Staging.