Gambogic Acid Inhibits Liver Cancer Via Disrupting YAP/TEAD Protein-protein Interaction

Objective:As an anti-cancer drug,gambogic acid(GA)is known to inhibit tumor progression by inducing cell apoptosis,inhibiting migration and invasion,and suppressing angiogenesis,representing potent anti-tumor activity.The Hippo signaling pathway is composed of a series of kinase cascades,which is a highly conservative pathway that regulates the homeostasis of tissues and organs.Its function mainly depends on the formation of a transcription complex composed by hippo signlaing effector YAP and TEAD proteins.Therefore,targeted inhibition of YAP/TEAD protein-protein interaction is a strategy for cancer treatment.This project aims to clarify whether gambogic acid can exert its anti-cancer properties by affecting the Hippo signaling pathway and/or disrupting the YAP/TEAD protein-protein interaction.Methods:Following treatment of GA,the GST pull-down and immunoprecipitation assays were employed to verify the effects of gambogic acid on the interaction of YAP and TEAD at exogenous and endogenous levels in liver cancer Hep G2 and Huh7 cells.Real-time PCR and Western blot assays were used to detect the expression and regulation of gambogic acid on the Hippo signaling pathway and YAP/TEAD regulatory molecules at the gene and protein levels.In addition,the dual luciferase reporter assay was carried out to detect the effect of gambogic acid on the transcriptional activity of the YAP/TEAD complex.In terms of functional verification,a stable YAP(S94A)cell line without TEAD binding ability was constructed,and cell proliferation,migration,and invasion assays were conducted to investigate whether gambogic acid can exert its anti-tumor activity associated with disrupting YAP/TEAD protein-protein interaction.Result:1.GA does not affect the expression phosphorylation of major members in the Hippo signaling pathway.2.GA inhibits the expression of target genes of YAP/TEAD transcription complex.3.GA suppresses the transcriptional activity of YAP/TEAD transcription complex.4.GA disurpts YAP/TEAD protein-protein interaction.5.Inhibition the interaction of YAP and TEAD is associated with the suppression of proliferation,migration and invasion by GA in liver cancer cells.Conclusion:Gambogic acid inhibits cell proliferation,migration,and invasion in liver cancer cells,which is associated with disrupting YAP/TEAD protein-protein interaction,inhibiting YAP/TEAD complex transcriptional acivity,as well as the down-regulation of the target genes that promoting cancer progression.