Experimental study on the radiosensitization effect of etanidazole and paclitaxel on hepatocellular carcinoma H22 cells in mice

Abstract

Objective To investigate the synergistic effect of etanidazole and paclitaxel on hepatocellular carcinoma H22 cells in mice. Methods To establish the murine xenografts, H22 cells was inoculated subcutaneously into the back of BALB/c mice. Among them, 40 tumor-bearing mice were divided into 4 groups by random number table method, 10 mice in each group, and each group injected with phosphate buffered saline (PBS), etanidazole (200 mg/kg), paclitaxel (1 mg/kg) and two drugs combination. Two hours after the administration, the mice were sacrificed by dislocation, and the drugs content in blood and tumor tissues of mice was measured by high-performance liquid chromatography. Sixty-five tumor-bearing mice were selected and divided into PBS non-irradiation group, PBS irradiation group, etanidazole irradiation group, paclitaxel irradiation group and two drugs combination irradiation group by random number table method, 13 mice/group. According to the grouping, the irradiated mice received 60Co source irradiation 2 hours after administration; the survival of 10 mice was observed at different time points, and the tumor volume was measured and calculated. The survival mice were killed by cervical dislocation 180 days after radiation. The other 3 mice in each group were killed after 2 days of irradiation, the tumor tissues were taken, and the expression of hypoxia inducible factor 1 (HIF-1) in these tumor tissues was detected by using immunohistochemistry. Statistical analysis of measurement data between groups was performed by using the one-way ANOVA and t test, and the survival analysis was made by Kaplan-Meier method. Results There was no significant difference of etanidazole and paclitaxel content in blood and tumor tissues between alone and combination administration groups at 2 hours after administration [etanidazole in blood: (55.5±4.7) μg/ml vs. (50.7±5.2) μg/ml; etanidazole in tumor: (31.2±3.5) μg/g vs. (33.6±5.4) μg/g; paclitaxel in blood: (316.9±9.6) μg/ml vs. (289.5±10.3) μg/ml; paclitaxel in tumor: (161.7±7.2) μg/g vs. (181.3±11.6) μg/g; all P > 0.05]. Within 40 days from the 10th day after the irradiation, the body weight of the irradiated group at each time point was significantly lower than that of the unirradiated group (all P 0.05). On the 40th day after irradiation, the tumor inhibition rates of the PBS irradiation group, etanidazole irradiation group, paclitaxel irradiation group, and two drugs combination irradiation group were 19.2%, 33.9%, 48.7%, and 61.6%, respectively. On the 180th day after irradiation, the survival rates of the PBS non-irradiation group, PBS irradiation group, etanidazole irradiation group, paclitaxel irradiation group, and two drugs combination irradiation group were 0, 0, 0, 12.5%, and 25.0%, respectively, and the median survival interval was 30.2, 54.7, 55.6, 83.4, and 102.8 d. After 2 days of irradiation, the positive expression rate of HIF-1 in the tumors tissues of PBS irradiation group, etanidazole irradiation group, paclitaxel irradiation group and two drugs combination irradiation group was lower than that in the PBS non-irradiation group, the differences were statistically significant [(45.7±4.8)%, (40.6 ±5.9)%, (24.5±5.6)%, (17.2±3.7%)% vs. (63.1±7.2)%, all P < 0.05]. The positive expression rate of HIF-1 in the paclitaxel irradiation group and two drugs combination irradiation group was lower than that in the PBS irradiation group and etanidazole irradiation group (all P < 0.05). Conclusion The combination of etanidazole and paclitaxel has synergistic radiation sensitization effect on hepatocellular carcinoma H22 cells in mice, and the expression level of HIF-1 is decreased, reflecting the decrease of hypoxic cells. Key words: Liver neoplasms; Mice; Etanidazole; Paclitaxel; Radiation tolerance