A80-2-09 Tumor Heterogeneity and Distinct Immune Microenvironment of Multiple Primary Lung Cancer

Background Multiple primary lung cancer (MPLC) exhibits distinct clinicopathological features and poor response to EGFR-TKIs, yet its underlying biology remains unclear. Methods We performed single-cell RNA sequencing (scRNA-seq) on 29 samples (14 tumors, 9 normal tissues) from 13 individuals (7 MPLC, 6 solitary primary lung cancer [SPLC] patients). Integrated analyses included copy number variation (CNV) inference, tumor cell annotation, immune cell annotation, and cell-cell communication mapping. Results MPLC demonstrated significant CNVs in histologically normal lung tissues (P<0.001), supporting the field cancerization hypothesis. Tumor cells in MPLC exhibited lower malignancy, evidenced by enrichment of alveolar differentiation genes (*SFTPA1/SFTPA2*; favorable prognosis in TCGA-LUAD, P<0.05); preferential oxidative phosphorylation (OXPHOS) over glycolysis (P<0.01), linked to DNMT1high/NNMTlow expression; reduced immunosuppressive Tregs and expanded cytotoxic GZMB+ CD8+ T cells and CD16+ NK cells (P < 0.001); MPLC-specific TNFA-NFKB pathway activation was observed in T cells and macrophages (CCL3+/AZU1+ subsets). In contrast, SPLC showed fibroblast-dominated stroma and glycolytic metabolism. Conclusion MPLC is characterized by field cancerization-driven tumorigenesis, OXPHOS dependency, and a unique immunocompetent microenvironment. Therefore, this study reveals the difference of tumorigenesis between MPLC and SPLC and provides a potential immunotherapy strategy for MPLC.