Files
Abstract
The transduction of external stimuli to cellular responses is vital for the function of living systems. Protein-protein interactions (PPIs) play a pivotal role in responding to stimuli and, via signaling cascades, achieving a cellular response. Aberrant regulation of signaling pathways in cells is responsible for numerous pathologies, and these pathways serve as viable targets for pharmaceutical development. The extensive binding interfaces of proteins present a challenge for conventional small molecule drug targeting. A strategy to target these interfaces is via the mimicry of key binding regions between proteins. α-helices comprise the largest class of secondary structure of proteins and are responsible for a major proportion of PPIs. Protein therapeutics, capable of specifically binding large protein interfaces, are amenable to targeting extracellular proteins such as receptors involved in signaling pathways, however, are limited by their permeability. Peptides, relatively short sequences of amino acids, are amenable to targeting these large interfaces. However, outside of the larger protein context, peptides are unlikely to adopt the proper α-helical conformation needed for target binding and face the risk of proteolytic degradation.Modification of the peptide via incorporation of synthetic olefin-bearing residues and ring-closing metathesis to form a macrocyclic brace has demonstrated stabilization of α-helical structures, increased proteolytic resistance, and reduced entropic penalty of target binding. These all-hydrocarbon stapled peptides are an emerging class of pharmaceuticals capable of disrupting PPIs conventionally deemed “undruggable” and in doing so provide a unique strategy to target signaling pathways in disease. The goal of this research was to develop and characterize novel all-hydrocarbon stapled peptide-based compounds to target proteins and PPIs involved in signaling pathways. These include a novel stapled-peptide PROTAC targeting Protein Kinase A for proteolytic degradation and inhibition of downstream phosphorylation, as well as BRK-derived mimics targeting the WASF Regulatory Complex that regulates actin cytoskeleton dynamics and drives metastasis in triple-negative breast cancer. Furthermore, we characterized a novel lipid encapsulation strategy in vitro to deliver proteins including trastuzumab, which disrupts HER2 signaling, across the cell membrane for access to intracellular targets.