In order to diversify the pharmacological activity of chalcones and extend the scaffold of topoisomerase and cathepsins B and L inhibitors, we have designed and synthesized total 18 chalcone compounds and tested their biological activity. In the topoisomerase inhibition test, most analogues in group III and IV except compound 11 exhibited more efficient topoisomerase I inhibitory activity than camptothecin at 20 μM. Compounds 15, 16 and 18 in group IV showed significant cathepsin B and L inhibitory activity. Among the compounds, compound 15 was most active with IC50 values of 1.81±0.05 μM on cathepsin B and 3.15±0.07 μM on cathepsin L, respectively. Compound 15 also showed most potent cytotoxic activity against T47D and SNU638 cells with IC50 values of 1.37±0.05 μM and 0.62±0.01 μM, respectively. Overall, although more compounds should be tested and analyzed for clear SAR against topoisomerase I and cathepsin B and L, compound 15 showed consistent inhibitory ability on the tested assays, which can implicate the cytotoxic activity of compound 15 on topoisomerase I and cathepsin B and L inhibitory pathways.