Advancing the Protein-Catalyzed Capture Agent Technology to New Frontiers

Citation

McCarthy, Amy Michelle (2018) Advancing the Protein-Catalyzed Capture Agent Technology to New Frontiers. Dissertation (Ph.D.), California Institute of Technology. doi:10.7907/HHP5-1Z83. https://resolver.caltech.edu/CaltechTHESIS:06012018-005336099

Abstract

Protein-catalyzed capture (PCC) agents are a nascent synthetic aptamer technology that was first disclosed in 2009. In addition to reviewing the different classes of peptide-based aptamers in chapter 1, this thesis records efforts to advance the PCC technology in two ways. First, in chapter 2 the development of a barcoded-rapid assay platform (B-RAP) technology enables the parallel analysis of up to fifteen PCC agents at once as well as dramatically shortening the time required to characterize the binding affinity for a pool of ligands from weeks to a couple of days. Secondly, the capture agent technology was utilized to target difficult proteins. Kirsten rat sarcoma (KRas) protein is a GTPase that acts as a light switch for several important cellular signaling pathways. Oncogenic variants of KRas are responsible for driving roughly 20-25% of all cancers, but KRas is considered “undruggable” from a small molecule targeting point of view. We report the identification of PCC ligands that bind to conserved allosteric switches on KRas and inhibit the protein’s GTPase enzymatic activity. The biomarker Plasmodium falciparum Histidine Rich Protein II (HRP2) presents an unusual challenge as it is a highly variable, unstructured and sticky protein. In chapter 3 we report on efforts to develop low nM binding capture agents against highly prevalent epitopes of HRP2, and the use of medicinal chemistry optimization to prepare structurally related variants of the lead capture agent for probing the structure-activity relationship and how it affects binding to HRP2.

Thesis Files