Discovery of 1‑Benzoyl 4‑Phenoxypiperidines as Small-Molecule Inhibitors of the β‑Catenin/B-Cell Lymphoma 9 Protein–Protein Interaction

Structure-based
design and optimization were performed to develop
small-molecule β-catenin/B-cell lymphoma 9 (BCL9) inhibitors
and improve their inhibitory activities. Compound ZL3138 with a novel 1-benzoyl 4-phenoxypiperidine scaffold was discovered
to disrupt the β-catenin/BCL9 protein–protein interaction
(PPI) with a Ki of 0.96 μM in AlphaScreen
competitive inhibition assays and displayed good selectivity for β-catenin/BCL9
over β-catenin/E-cadherin PPIs. The binding mode of new inhibitors
was characterized by structure–activity relationship and site-directed
mutagenesis studies. Protein pull-down assays indicate that this series
of compounds directly binds with β-catenin. Cellular target
engagement and co-immunoprecipitation experiments demonstrate that ZL3138 binds with β-catenin and disrupts the β-catenin/BCL9
interaction without affecting the β-catenin/E-cadherin interaction
in living cells. Further cell-based studies show that ZL3138 selectively suppresses transactivation of Wnt/β-catenin signaling,
regulates transcription and expression of Wnt target genes, and inhibits
the growth of Wnt/β-catenin-dependent cancer cells.