Objective Tumor necrosis factor related apoptosis-inducing ligand(TRAIL)has been demonstrated that se-lectively induced cancer cells, but facilitated prosurvival response leading to resistance. It has been reported that genistein could inhibit nuclear factor-κB(NF-κB)exprotein and activaty. The purpose of this study was to examine the effects of genistein or TRAIL,or both on apoptosis of liver cancer stem-like cells(LCSLCs)derived from SMMC-7721 cell line. Methods The third passage sphere-forming cells (SFCs)derived from SMMC-7721 cell line which was also called LCSLCs were obtained though suspended culture in ultra-low adhesion plate with stem cell-conditioned medium. LCSLCs were treated with Genistein (10μM)and TRAIL (10 ng/mL)alone or in combination for 72 h. Histone/DNA fragment was tested using enzyme-linked immunosorbent assay (ELISA). And apoptosis rate was detected by flow cy-tometry (FCM) with propidium iodide(PI) staining; the protein expression of NF-κB(p65) was analyzed by western blot. Results The level of histone/DNA fragment(100%vs 152.6%)and apoptosis rate[(2.71±0.64)%vs (5.14±0.76)%] was increased moderately by genistein(10 μM) in LCSLCs(P<0.05), but TRAIL(10 ng/mL) has no effect on histone/DNA fragment level(100% vs 110%) and apoptosis rate[(2.71±0.64)% vs (3.15±0.72)%]. After preincubation with 10μM genistein for 1h, the combination with 10 ng/mL TRAIL lead to significantly increase of histone/DNA fragment level(100% vs 568%) and apoptosis rate[(2.71±0.64)% vs (17.3±1.27)%] (P<0.05). 10 ng/mL TRAIL induced NF-κB(p65)expression in LCSLCs. 10μM genistein could suppress the protein expression of NF-κB(p65). And the pro-tein expression of NF-κB (p65) was significantly attenuated by the combination of genistein and TRAIL. Conclusion The apoptosis of LCSLCs which is co-induced by genistein and TRAIL is associated with the inhibition of NF-κB.