We describe a general method for the mimicry of one face of an alpha-helix based on a terphenyl scaffold that spatially projects functionality in a manner similar to that of two turns of an alpha-helix. The synthetic scaffold reduces the flexibility and molecular weight of the mimicked protein secondary structure. We have applied this design to the development of antagonists of the alpha-helix binding protein Bcl-x(L). Using a sequential synthetic strategy, we have prepared a library of terphenyl derivatives to mimic the helical region of the Bak BH3 domain that binds Bcl-x(L). Fluorescence polarization assays were carried out to evaluate the ability of terphenyl derivatives to displace the Bcl-x(L)-bound Bak peptide. Terphenyl 14 exhibited good in vitro affinity with a K(i) value of 0.114 muM. These terphenyl derivatives were more selective at disrupting the Bcl-x(L)/Bak over the HDM2/p53 interaction, which involves binding of the N-terminal alpha-helix of p53 to HDM2. Structural studies using NMR spectroscopy and computer-aided docking simulations suggested that the helix binding area on the surface of Bcl-x(L) is the target for the synthetic ligands. Treatment of human embryonic kidney 293 (HEK293) cells with terphenyl derivatives resulted in the disruption of the binding of Bcl-x(L) to Bax in intact cells.
A series of Bcl-x(L)/Bak antagonists, based on a terephthalamide scaffold, was designed to mimic the alpha-helical region of the Bak peptide. These molecules showed favorable in vitro activities in disrupting the Bcl-x(L)/Bak BH3 domain complex (terephthalamides 9 and 26, K(i) = 0.78 +/- 0.07 and 1.85 +/- 0.32 microM, respectively). Extensive structure-affinity studies demonstrated a correlation between the ability of terephthalamide derivatives to disrupt Bcl-x(L)/Bak complex formation and the size of variable side chains on these molecules. Treatment of human HEK293 cells with the terephthalamide derivative 26 resulted in disruption of the Bcl-x(L)/Bax interaction in whole cells with an IC(50) of 35.0 microM. Computational docking simulations and NMR experiments suggested that the binding cleft for the BH3 domain of the Bak peptide on the surface of Bcl-x(L) is the target area for these synthetic inhibitors.
The Bcl-x(L)/Bak protein-protein interaction has emerged as an important target for cancer therapy due to its role in apoptosis. Inhibition of this interaction by small-molecule antagonists induces apoptosis in unhealthy cells. Bak, a pro-apoptotic Bcl-2 protein, projects four hydrophobic side chains (V74, L78, I81, and I85), corresponding to the i, i+4, i+7, and i+11 positions of an alpha-helix, into a hydrophobic cleft on Bcl-x(L). Herein, we present a novel family of rationally designed alpha-helix mimetics with improved solubility and synthetic feasibility based on a benzoylurea scaffold. These benzoylurea derivatives favor a linear conformation stabilized by an intramolecular hydrogen bond, and are able to mimic the spatial projection of the i, i+4, and i+7 residues of an alpha-helix. The binding of the benzoylurea derivatives to Bcl-x(L) was assessed using fluorescence polarization competition assays, isothermal titration calorimetry, and (15)N-HSQC experiments. These experiments showed that these agents bind to and disrupt Bcl-x(L) with low micromolar inhibition and dissociation constants, with (15)N-HSQC experiments confirming binding to the hydrophobic pocket of Bcl-x(L) normally occupied by the Bak helix.
Turn Bak: We present rationally designed scaffolds that mimic the spatial projection of the i, i+4, i+7, and i+11 residues of an α‐helix. A library of biphenyl derivatives was shown by competition fluorescence polarization and ITC to mimic Bak and disrupt the Bak/Bcl‐xL protein–protein interaction. 15N HSQC experiments confirmed that the surface of Bcl‐xL normally occupied by Bak was the target area of our new synthetic inhibitors.magnified image
scite is a Brooklyn-based organization that helps researchers better discover and understand research articles through Smart Citations–citations that display the context of the citation and describe whether the article provides supporting or contrasting evidence. scite is used by students and researchers from around the world and is funded in part by the National Science Foundation and the National Institute on Drug Abuse of the National Institutes of Health.