The TEAD family of transcription factors are the pivotal downstream effectors that regulate the final transcriptional output of the Hippo signal cascade. Although emerging clinical evidence has implicated the tumorigenic role of TEAD-YAP, drug discovery against Hippo pathway has been unsuccessful. It remains challenging to develop small molecule activators for the main kinase module and TEAD-YAP interface has been intractable to conventional drug discovery approaches due to the flat interface and high binding affinity for PPI. Recently, there is biochemical evidence demonstrating that TEAD proteins undergo auto-palmitoylation on a conserved cysteine under physiological conditions which is essential for its function and stability making it an attractive site for pharmacological intervention. Herein, through screening against rationally designed TEAD-focused covalent fragment library based on reported non-covalent lipid scaffold flufenamic acid, we identified a novel Y-shaped pocket which could be pharmacologically targeted by MYF-03-69, a potent TEAD palmitoylation inhibitor. MYF-03-69 significantly inhibited the TEAD palmitoylation activity while the non-covalent counterpart showed minimal inhibition. Further crystallographic investigation revealed the basis for molecular recognition and provided guidance for further structure-guided optimization efforts. In cells, MYF-03-69 significantly inhibited TEAD transcription activity and influenced canonical downstream gene expression including CTGF, CYR61, NRG1, KRT34, IGFBP3, NPPB and pro-apoptotic gene BMF by abolishing endogenous TEAD-YAP interaction. Besides, MYF-03-69 retarded the cell proliferation of YAP-activated NCI-H226 (NF2-deficient) and MSTO-211H (LATS1-PSEN1 fusion) mesothelioma cells by arresting cell cycle at G1 phase and exhibited excellent selectivity over normal mesothelium cells (MeT-5A) and YAP-insensitive NF2-intact mesothelioma cells (NCI-H2452). Our work highlights TEAD as an exploitable molecular vulnerability in mesothelioma and provides an efficient paradigm for combining covalent fragment screening with structure-based medicinal chemistry to develop new covalent prototype drugs.

Discovery of a covalent, cell-active pan-TEADs palmitoylation inhibitor

ACS Spring Meeting 2021

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