Gli1, a potential regulator of esophageal cancer stem cell, is identified as an independent adverse prognostic factor in esophageal squamous cell carcinoma

Abstract

The hedgehog (Hh) pathway is involved in cancer stem cell (CSC) maintenance in various tumors. Glioma-associated oncogene homolog 1 (Gli1) is a key mediator of the Hh pathway; however, its expression and clinical significance in esophageal squamous cell carcinoma (ESCC) have not been reported. In this study, we aimed to reveal clinical significance of Gli1 expression in ESCC and further investigate the potential of Gli1 as a CSC regulator of ESCC by comparing its expression with expressions of other stemness genes in ESCC. We assessed the expressions of Gli1, Sox9, CD44, Sox2, LSD1, and Oct4 in 127 patients' tissue specimens of ESCC using immunohistochemistry and in ESCC cell lines using Western blotting. The relationship of Gli1 expression with clinic-pathologic parameters as well as cell-cycle-regulating genes was investigated. We also investigated the biological pathways that are activated in Gli1-high ESCC using The Cancer Genome Atlas (TCGA) data. Gli1 expression was observed in 28.3% of ESCC, and its expression was correlated with the expression of stemness genes, Sox9 (P=0.003) and CD44 (P=0.012). And Gli1, CD44, and Sox9 were highly expressed in more poorly differentiated ESCC cell lines such as TE8 and TE1 cells. Notably, Gli1 expression was positively associated with distant metastasis (P=0.011), increased microvessel density (MVD) (P=0.002), and expression of cell cycle regulators such as p21, cyclin D1, cyclin E1, and NF-κB (P<0.05). Sox9 and CD44 expressions in ESCC were also significantly associated with unfavorable clinic-pathologic parameters such as increased MVD, advanced tumor (pT) stage, and higher TNM stage. Moreover, all three potential CSC markers such as Gli1, Sox9, and CD44 were strongly linked to worse clinical outcome and independent poor prognostic factors in overall survival and disease-free survival in ESCC. Gene set enrichment analysis revealed that the Gli1-high-expressing ESCC patients' group was strongly enriched for gene expression signature of Hh signaling pathway, epithelial-mesenchymal transition, and cancer stem cell. Targeting Gli1, a potential diagnostic marker of ESCC stem cells, will have a profound therapeutic and prognostic value.