Dynamic tumor microenvironment remodeling from laryngeal leukoplakia to carcinoma revealed by single-cell transcriptomics.

Laryngeal leukoplakia represents the most frequent precancerous lesion in laryngeal carcinogenesis, yet its transformation mechanisms remain elusive. By performing scRNA-seq on ten clinical specimens (five leukoplakia lesions across pathological stages, four early carcinomas, and one control), we established the first single-cell atlas of this malignant progression. Computational analysis revealed dynamic microenvironmental shifts dominated by epithelial cells, fibroblasts, and mononuclear phagocytes. We identified two critical epithelial subpopulations: Epi_4 (tumor-like cells), a high-grade dysplasia-specific subpopulation with high malignant potential, and Epi_5 (tumor cells) in carcinoma, which carries a favorable prognostic gene signature (Module 3). Furthermore, Epi_4 showed preferential communication with cancer-associated fibroblasts (CAFs) and tumor-associated macrophages (TAMs) via the JAG1-NOTCH4 and CXCL5-CXCR1 axes, suggesting actionable therapeutic targets. We also observed the progressive activation of genes involved in redox processes (NQO1, GSTM3, UCHL1, NTRK2) via the KEAP1-NRF2 pathway. This work systematically characterizes the cellular and molecular landscape during laryngeal leukoplakia malignant transformation, providing a framework for future mechanistic studies and early detection strategies.