A series of triazinic inhibitors of focal adhesion kinase (FAK) have been recently shown to exert antiangiogenic activity against HUVEC cells and anticancer efficacy against several cancer cell lines. We report herein that we further explored the heterocyclic core of these inhibitors by a fused imidazole ring with the triazine to provide imidazo[1,2-a][1,3,5]triazines. Importantly, these new compounds displayed 10(-7)-10(-8) M IC50 values, and the best inhibitor showed IC50 value of 50 nM against FAK enzymatic activity. Several inhibitors potently inhibited the proliferation of a panel of cancer cell lines expressing high levels of FAK. Apoptosis analysis in U87-MG and HCT-116 cell lines suggested that these compounds delayed cell cycle progression by arresting cells in the G2/M phase of the cell cycle, retarding cell growth. Further investigation demonstrated that these compounds strongly inhibited cell-matrix adhesion, migration, and invasion of U87-MG cells.

Focal Adhesion Kinase signaling pathway and its functions have been involved in the development and aggressiveness of tumor malignancy, it then presents a promising cancer therapeutic target. Several reversible FAK inhibitors have been developed and are being conducted in clinical trials. On the other hand, irreversible covalent inhibitors would bring many desirable pharmacological features including high potency and increased duration of action. Herein we report the structure-guided development of the first highly potent and irreversible inhibitor of the FAK kinase. This inhibitor showed a very potent decrease of autophosphorylation of FAK in squamous cell carcinoma. A cocrystal structure of the FAK kinase domain in complex with this compound revealed the inhibitor binding mode within the ATP binding site and confirmed the covalent linkage between the targeted Cys427 of the protein and the inhibitor.