Cytotoxic activity-guided fractionation studies on Glycyrrhiza echinata roots led to the isolation of eight compounds (1–8). Chemical structures of the isolates were identified by NMR and MS analysis. Among the tested molecules, retrochalcones namely echinatin (3) (IC50=23.45–41.83 μM), licochalcone B (4) (IC50=36.04–39.53 μM) and tetrahydroxylmethoxychalcone (5) (IC50=7.09–80.81 μM) were the most active ones against PC3, MCF7 and HepG2 cells. Moreover, 5 exhibited selectivity on prostate cancer cells (SI: 5.19). Hoechst staining and Annexin V/PI binding assays as well as cell cycle analysis on the compounds 3 (23 μM) and 5 (5 and 7 μM) demonstrated that these retrochalcones induced apoptosis and significantly suppressed cell cycle in G1 and G2/M phases. Furthermore, 3 and 5 showed antimigratory effects on PC3 cells by wound healing assay. The results indicated that tested retrochalcones most particularly 5 could be potential anticancer drug candidates that prevent proliferation and migration of cancer cells.