Abstract
OBJECTIVES: To synthesize new bakuchiol aminoguanidine derivatives and test their effect on viability and apoptosis of human triple-negative breast cancer (TNBC) cells. METHODS: Two bakuchiol derivatives 1 and 2 were obtained by formylation and Shiff base reaction of bakuchol. The structures of derivatives 1 and 2 were identified by (1)H-NMR, (13)C-NMR, and high-resolution electrospray ionization mass spectrometry (HR-ESI-MS) analysis. Human TNBC MDA-MB-231 cells were treated with bakuchiol and its derivatives and cell viability was measured by MTT assay. Apoptosis was detected by fluorescence microscopy and flow cytometry with Annexin V-FITC/PI staining. The expressions of apoptosis-related proteins were analyzed with Western blotting. The JC-1 and reactive oxygen species (ROS) assay kits were used to determine the effect of new bakuchiol derivatives on mitochondrial function. RESULTS: Based on spectroscopic analysis, a new bakuchiol schiff base derivative was elucidated as 2-{(E)-5-[(S, E)-3, 7-dimethyl-3-vinylocta-1, 6-dien-1-yl]-2-hydroxylbenzylidene} hydrazine-1-carboximidamide (derivative 2). Bakuchiol and its derivatives 1 and 2 all showed cytotoxic activity against the MDA-MB-231 cells. Derivative 2 exhibited the most potent cytotoxic activity to MDA-MB-231 cell with IC(50) of (13.11±1.09), (6.91±1.78), and (2.23±1.32) μmol/L after 24, 48, and 72 h. It had low toxicity to normal mouse liver (AML-12) cells with IC(50) of (31.23±1.58) μmol/L at 72 h. Fluorescence microscopy and flow cytometry demonstrated apoptosis in breast cancer cells after treating with derivative 2 in a concentration dependent manner. Western blotting showed that after derivative 2 treatment, the expression of apoptosis-related proteins cytochrome C, cleaving caspase-3 and Bax/Bcl-2 radio in MDA-MB-231 cells increased; in addition, apoptosis was associated with the decreased mitochondrial membrane potential and increased reactive oxygen species accumulation. CONCLUSIONS: The novel bakuchiol aminoguanidine derivative (derivative 2) is capable of inducing apoptosis in MDA-MB-231 cells, but has low toxicity to normal liver cells, suggesting that it may be used as a lead compound for an anti-TNBC agent.