Targeting Undruggable Oncoprotein Epitopes with Protein Catalyzed Capture Agents

Citation

Henning, Ryan Kenneth (2017) Targeting Undruggable Oncoprotein Epitopes with Protein Catalyzed Capture Agents. Dissertation (Ph.D.), California Institute of Technology. doi:10.7907/Z9CJ8BJG. https://resolver.caltech.edu/CaltechTHESIS:06022017-070813473

Abstract

The protein catalyzed capture (PCC) agent platform provides a new strategy to develop peptide-based ligands for difficult protein targets. This approach utilizes the target-guided in situ click reaction to allow the protein of interest to assemble its own binder. Developing a PCC agent begins with an epitope targeting strategy to develop anchor candidates against a specific region of interest on the target protein. This approach has been used to target diverse epitopes including unstructured hydrophobic regions, allosteric enzyme sites, and single amino acid point mutations. The process can then be iterated to expand a monoligand into a multiligand binder with affinity and selectivity that rivals monoclonal antibodies.

One disease-associated protein of particular importance is the serine/threonine kinase Akt. Akt is a key regulator of signal transduction pathways and is implicated in many disease such as cancer, diabetes, and neurodegeneration. Several ligands for Akt have been developed recently with the PCC agent screening approach. PCC agents now exist that can alter Akt enzymatic activity, detect its position in the cell, identify mutations within the protein, and even cause its destruction within the cell. The first part of this thesis summarizes the prior efforts to develop PCC agents against Akt and then describes new applications for these reagents while the latter part describes efforts to develop new PCC agents against another interesting target.

Chapter 1 provides a summary of the technology and describes how it has be utilized thus far. Chapter 2 describes how a PCC agent was used as an imaging probe capable of detecting Akt membrane localization. Chapter 3 provides several examples of the modularity of PCC agents and demonstrates how they can be used to influence a target protein in cells. A pair of allosteric Akt modulators were functionalized with a cell penetrating peptide for cellular delivery and were subsequently used to activate or inhibit Akt enzymatic activity. PCC agents can also be used as a targeting moiety to deliver a specific signal to a protein. When functionalized with a degradation tag the Akt-binding capture agents caused the protein to be degraded. This provides another demonstration of the usefulness of Proteolysis Targeting Chimeric Molecules, or PROTACs, in destroying disease-associated proteins. Finally, Chapter 4 describes the development of PCC agents against the oncoprotein K-RasG12D and how these molecules can be used to target this protein in new ways.

Thesis Files