Identification of the Constituent Brusatol as a EGFR-Tyrosine Kinase Inhibitor in a Cell-Free Assay.

Inhibitors of the tyrosine kinase (TK) activity of the epidermal growth factor receptor (EGFR) are routinely used in cancer therapy. However, there is a need to discover a new TK inhibitor. This study evaluated extracts from and its components for their potential as novel EGFR-TK inhibitors. The cytotoxic effect of a aqueous extract and its fractions was assessed by MTT assays with A549 lung cancer cells. The two fractions with the highest cytotoxicity were analyzed by LC/MS and H NMR. Brusatol was identified as the main constituent of these fractions, and its cytotoxic and pro-apoptotic activities were confirmed in A549 cells. To elucidate the inhibitory activity of brusatol against EGFR-TK, a specific ADP-GloTM kinase assay was used. In this assay, the IC value for EGFR-TK inhibition was 333.1 nM. Molecular dynamic simulations and docking experiments were performed to identify the binding pocket of brusatol to be located in the intracellular TK-domain of EGFR. This study demonstrates that brusatol inhibits EGFR-TK and therefore harbors a potential as a new therapeutic drug for the therapy of EGFR-depending cancers.