Identification and functional validation of GREM2 and PIK3C2G as senescence-associated genes in esophageal squamous cell carcinoma.

Esophageal squamous cell carcinoma (ESCC) is an aggressive malignancy with dismal clinical outcomes, and how cellular senescence modulates ESCC biology remains poorly understood. Here, we integrated transcriptomic profiles from the TCGA-ESCC cohort with a curated panel of cellular senescence-related genes to identify and functionally validate senescence-associated regulators in ESCC. Univariate Cox regression and consensus clustering stratified ESCC patients into two molecular subtypes with divergent survival. Through differential expression analysis, random forest prioritization, and independent validation in the GSE20347 cohort, we identified GREM2 and PIK3C2G as lead candidate genes. Both were consistently downregulated in ESCC tumors and cell lines, and low expression predicted poorer overall survival. Gain- and loss-of-function experiments demonstrated that GREM2 and PIK3C2G drive p16/p21-dependent cellular senescence, trigger the senescence-associated secretory phenotype (SASP), and restrict proliferation in esophageal epithelial cells. Immune correlation analyses linked both genes to immune cell infiltration, chemokine signaling, and MHC molecule expression in the tumor microenvironment. These findings establish GREM2 and PIK3C2G as senescence-associated tumor suppressors in ESCC, offering candidate biomarkers and a rationale for exploring senescence-directed therapies in this disease.