Global Metabolomic and Lipidomic Analysis Reveal the Synergistic Effect of Bufalin in Combination with Cinobufagin against HepG2 Cells

Chansu, which is prepared from the skin secretions of toad (Bufo bufo gargarizans Cantor), is widely used in traditional Chinese medicine (TCM). Being the principal bioactive constituents of Chansu, bufalin (BFL) and cinobufagin (CBF) have been shown to possess anticancer properties. TCM confers bioactivities through the synergistic effect between potential active ingredients, so as to interfere with the development of the disease, and ultimately achieve the therapeutic effect. We found that the anticancer effect was significantly potentiated by cotreatment with BFL and CBF compared to monotreatment, suggesting their synergistic interaction. To reveal their synergistic mechanisms, metabolomic and lipidomic profiling based on liquid chromatography–mass spectrometry (LC–MS) was utilized to delineate the responses in HepG2 cells after treatment with BFL and CBF individually or in combination. Metabolic pathways, including methionine metabolism, energy metabolism, lipid metabolism, and amino acid metabolism, were modulated and subsequently led to apoptosis and cell cycle arrest of HepG2 cells. In particular, the discrepant regulation of methionine metabolism between the monotreatment and cotreatment with BFL and CBF may account for their synergistic effect. Our study provided novel insights into the mechanistic links between cellular metabolism and the synergistic effect, which may ultimately lead to better treatments for hepatoma.