5-azacytidine inhibits the tumorigenesis of esophageal cancer cells through the suppression of NOTCH1 signaling

Abstract Background: Esophageal squamous carcinoma (ESCC) and esophageal adenocarcinoma (EAC) are different pathological types of esophageal cancer (EC) with low patient survival. The methyltransferase inhibitor 5-azacytidine (5-azaC) has been approved to treat hematological malignancies and malignant solid tumors for years. NOTCH1 pathway plays an important role in both hematological and esophageal cancer and previous studies demonstrated a NOTCH1/IL-7/IL-7R signal in other cancers. Methods: TE-1 and OE33 cells were employed to represent ESCC and EAC respectively. The effects of 5-azaC on cells were evaluated by CCK8, wound healing, Transwell assay, and flow cytometry. Pyrosequencing was performed to detect changes of 18 CpG units in cells after being treated with 5-azaC. Western blot and Quantitative Real-time PCR were conducted respectively to test expressions of NOTCH1/IL-7/IL-7R signal for exploring the mechanisms. siRNA transfections were performed to inhibit IL-7R. Results: 5-azaC showed anticancer effects and NOTCH1 signaling was also downregulated in both cell lines. Although there were abundant CpG islands in NOTCH1, no change was observed in its methylation level. Moreover, the combination of 5-azaC with NOTCH1 signaling inhibitor DAPT had a synergistic inhibiting effect in EAC but ESCC cells. We proved the existence of the NOTCH1/IL-7/IL-7R signal in the ESCC cell line. Additionally, the activation or inactivation of the IL-7/IL-7R pathway could mitigate or potentiate the potency of 5-azaC on ESCC cells as well. Conclusions: Our findings showed a possibility of treating esophageal cancer with 5-azaC combining inhibitors of NOTCH1/IL-7/IL-7R signal, hoping to provide novel therapeutic strategies for EC.