Effects of the combination of As(2)O(3) and AZT on proliferation inhibition and apoptosis induction of hepatoma HepG2 cells following silencing of Egr-1

CONTEXT: Previous studies have demonstrated that 3′-azido-3′-deoxythymidine (AZT) and arsenic trioxide (As(2)O(3)), traditional chemotherapy agents, can synergically inhibit the growth of hepatocellular carcinoma cells. However, the molecular mechanisms underlying As(2)O(3) and AZT anti-hepatoma activity are unknown. OBJECTIVE: This study aimed to investigate the role of early growth response protein 1 (Egr-1) in the process of As(2)O(3) combined with AZT inhibiting proliferation and inducing apoptosis of human hepatocellular carcinoma HepG2 cells, and explore the possible mechanism. MATERIALS AND METHODS: The expression of Egr-1 was silenced using siRNA, and then HepG2 cells were treated with As(2)O(3) (2 μM) and AZT (20 μM). The rates of cell inhibition and apoptosis were determined by the 3-(4,5-Dimethylthiazol-2-yl)-2,5-diphenyltetrazoliumbromide (MTT) method and flow cytometry, respectively. The mRNA and protein expression of p53, caspase-3, and Egr-1 were detected by real-time quantitative polymerase chain reaction and Western blotting, respectively. RESULTS: The inhibitory rate of As(2)O(3) (2 μM) combined with AZT (20 μM) on proliferation of HepG2 cells was significantly higher than that of As(2)O(3) alone. The combination index (CI) values were 0.2<CI<0.4, showing strong synergic effect. After silencing Egr-1, the proliferation inhibition and proapoptotic ability of As(2)O(3) combined with AZT on HepG2 cells were decreased, and the CI value was greater than 1, showing antagonistic effect. In addition, the expression of p53 and caspase-3 mRNA/protein was also significantly decreased. CONCLUSION: The present results show that AZT could increase the sensitization of As(2)O(3) for inhibiting proliferation and promoting apoptosis of HepG2 cells through regulating the expression of Egr-1, which may control the expression of p53 and caspase-3.