Lupiwighteone (Lup) is a kind of natural isoflavone, but its pharmacological effect and active mechanism are rarely reported. This study aimed to investigate the anticancer and cancer preventive effects of Lup on human neuroblastoma (SH-SY5Y) cells. We found that Lup could inhibit SH-SY5Y cells growth in a concentration- and time-dependent manner. Further studies suggested that Lup could induce G2/M phase arrest associated with an evident decrease in cyclin B1/D1 and cyclin dependent kinase (CDK) 1/2/4/6 protein expressions. Moreover, Lup could regulate the changes of mitochondrial membrane potential and increase intracellular reactive oxygen species (ROS) production. After the cells were treated with Lup, topical morphological characteristics were observed; apoptosis-related protein expressions, such as Bax, cytochrome c, cleaved caspase-9, cleaved caspase-3 and cleaved PARP-1 were increased; and protein expressions, such as Bcl-2, procaspase-9, PARP-1 and P-Akt were decreased. These changes were observed simultaneously. In addition, Nrf2 transcription factor activation was detected by an ARE-GFP reporter assay. Nrf2 nuclear localization was then investigated using a fluorescence microscope. Furthermore, Nrf2 and Keap1 protein levels were determined by western blot. Our results may provide a scientific basis for the application of the anticancer and cancer preventive effects of Lup on SH-SY5Y cells.