Title: Small Molecule Modulators of Apoptosis
Description: Control of cell survival relies on a delicate balance between pro-apoptotic and anti-apoptotic signalling. In humans, the key regulatory proteins are those of the BCL-2 family, which include effector proteins such as BAX and BAK, anti-apoptotic proteins including BCL-2 and MCL-1, and pro-apoptotic proteins including BID and BIM. Dysregulation of apoptosis is among the Hallmarks of Cancer, and modulation of apoptosis holds promise as an effective therapeutic strategy for a range of malignancies. This thesis advances new strategies for modulating apoptosis using small molecules.

The first section explores the properties of stapled peptides. These molecules incorporate two non-natural amino acids with olefin sidechains that are then covalently linked. The resulting “staple” modifies the biophysical properties of the molecule. This chapter shows how stapling results in greater serum stability of the peptides, improves binding affinity for anti-apoptotic targets, and allows for facile transformation of a native sequence into an improved peptide, as demonstrated by stapling a SOS1 peptide to target KRAS.

The second section targets MCL-1, an antiapoptotic BCL-2 family protein that has emerged as a major pathogenic factor in human cancer. MCL-1 bears a surface groove whose function is to sequester the BH3 killer domains of proapoptotic BCL-2 family members, but successful drugging of this groove has not been achieved. This chapter develops an alternative strategy using a small molecule that covalently modifies C286 at a novel interaction site distant from the BH3-binding groove. This allosteric mechanism results in reduced BH3 binding capacity of MCL-1 and impairs the oncogenic anti-apoptotic activity of the protein.

The final chapter targets BAX, a critical executioner protein in the apoptotic pathway whose oligomerization causes permeabilization of the mitochondrial outer membrane. Using STD-NMR, a library of nearly 1,000 fragments was screened for binding to full-length BAX. This resulted in the discovery of a compound BIF-44 that sensitizes BAX by engaging a noncanonical hydrophobic pocket formed by the junction of the α3-α4 and α5-α6 hairpins. Biochemical and structural analyses indicate that the molecule sensitizes BAX by allosterically mobilizing the α1-α2 loop, a mechanism implicated in the initiation of BH3-mediated direct BAX activation. The identified compound thus informs the mechanism for initiation of BAX activation, and provides a new opportunity to reduce the apoptotic threshold for potential therapeutic benefit.