Objective To investigate the inhibition and radiosensitization effects of Topotecan (TPT) on cervical cancer cell line HeLa, and make comparison with Cisplatin (DDP) and Taxol (TAX). Methods The effects of TPT, DDP and TAX on proliferation of cervical cancer cell line HeLa were evaluated by MTT assay. The radiosensitization effects of TPT, DDP and TAX on HeLa cells were detected by clone formation assay, and the sensitization enhancement ratios of TPT, DDP and TAX were calculated. Results The IC50 of TPT at different reaction time (24 h, 48 h and 72 h) were 8.0 μg/mL, 2.6 μg/mL and 0.8 μg/mL, respectively, those of DDP were 2.4 μg/mL, 0.7 μg/mL and 0.1 μg/mL, respectively, and those of TAX were 0.3 μg/mL, 0.1 μg/mL and 0.0 μg/mL, respectively. The apoptosis rates of tumor cells treated by radiosensitization were significantly higher than those of tumor cells treated by single radiotherapy or single chemotherapy (P<0.05). The sensitization enhancement ratios of TPT at 24 h and 48 h were 1.167 and 1.344, respectively, those of DDP were 1.314 and 1.538, respectively, and those of TAX were 1.076 and 1.316, respectively. Conclusion TPT, DDP and TAX have significant inhibition effects on cervical cancer cell line HeLa, which are time-dependant and dose-dependant. Besides, TPT, DDP and TAX have radiosensitizing effects, with DDP being the strongest and TAX being the weakest.