OBJECTIVE To investigate the inhibitory effects of Bruceine A (BA) on colon cancer and its underlying mechanisms.

METHODS Human colon cancer HT-29 and HCT116 cells were treated with various concentrations of BA (0, 1, 2, 5, 10, 20, 40, 80 μmol ·L^-1). Cell viability was assessed using the Cell Counting Kit-8 (CCK-8). Flow cytometry, wound healing assays, and Transwell assays were employed to evaluate the effects of BA on cell apoptosis, cell cycle, invasion, and migration. Molecular docking simulations were used to assess the binding of BA to GSK-3β protein, and Western blot analysis was used to examine protein expression related to the cell cycle, epithelial-mesenchymal transition, and the Wnt/β-catenin signaling pathway. An HT-29 cell subcutaneous xenograft mouse model was established. After tumor formation, mice were randomly divided into three groups (six mice per group): a blank group, a low-dose BA group (0.1 mg ·kg^-1), and a high-dose BA group (0.2 mg ·kg^-1). Mice were administered the drug for 19 d, then sacrificed, and tumor tissues were collected. Tumor volume changes over time were observed; Ki67 immunohistochemistry was used to assess cell proliferation in tumor tissues; Western blot analysis of Wnt/β-catenin signaling pathway protein expression was conducted.

RESULTS Compared with the blank group, BA could significantly inhibit the proliferation of HT-29 and HCT116 cells, with IC₅₀ values of 10.80 μmol ·L^-1 and 17.96 μmol ·L^-1, respectively. Flow cytometry results showed that BA significantly induced apoptosis of HT-29 cells (P < 0.01, P < 0.001), and arrested the cell cycle at the S phase, accompanied by decreased expression of cycle-related proteins CDK2 and Cyclin A (P < 0.05, P < 0.01, P < 0.001). BA inhibited cell migration and invasion ability (P < 0.05, P < 0.01, P < 0.001), reduced the expression of EMT-related proteins Snail, Vimentin, and N-Cadherin(P < 0.01, P < 0.001), and upregulated the expression of E-Cadherin protein. In addition, BA inhibited the expression of β-catenin and p-GSK3β proteins. Wnt agonist LiCl could significantly antagonize the anti-colon cancer effect of BA; Wnt inhibitor XAV939 could enhance the anti-colon cancer effect of BA. In the in vivo experiment, compared with the blank group, the tumor volume of the low-dose and high-dose BA groups was significantly reduced (P < 0.05, P < 0.001). Immunohistochemistry results showed that compared with the blank group, the expression of Ki67 in tumor tissues of the low-dose and high-dose BA groups was significantly reduced (P < 0.001). Western blot results further proved that BA inhibited the Wnt/β-catenin signaling pathway.

CONCLUSION BA inhibits the viability, invasion, and migration of colon cancer HT-29 cells, induces apoptosis, and causes cell cycle arrest. Additionally, it significantly suppresses the growth of subcutaneous HT-29 cell xenografts in vivo, possibly related to the Wnt/β-catenin signaling pathway.