Discovery of molecules that modulate protein-protein interactions in the context of human proliferating cell nuclear antigen-associated processes of DNA replication and damage repair

Matthew David Bartolowits, Purdue University

Abstract

Integral to cell viability is the homotrimeric protein complex Proliferating Cell Nuclear Antigen (PCNA) that encircles chromatin-bound DNA and functionally acts as a DNA clamp that provides topological sites for recruitment of proteins necessary for DNA replication and damage repair. PCNA has critical roles in the survival and proliferation of cells, as disease-associated dysregulation of associated functions can have dire effects on genome stability, leading to the formation of various malignancies ranging from non-Hodgkin’s lymphoma to skin, laryngeal, ocular, prostate and breast cancers. Here, a strategy was explored with PCNA as a drug target that may have wider implications for targeting protein-protein interactions (PPIs) as well as for fragment-based drug design. A design platform using peptidomimetic small molecules was developed that maps ideal surface binding interaction sites at a PPI interface before considering detailed conformations of an optimal ligand. A novel in silico multi-fragment, combinatorial screening approach was used to guide the selection and subsequent synthesis of tripeptoid ligands, which were evaluated in a PCNA-based competitive displacement assay. From the results, some of the peptoid-based compounds that were synthesized displayed the ability to disrupt the interaction between PCNA and a PIP box-containing peptide. The IC50 values of these compounds had similar or improved affinity to that of T2AA, an established inhibitor of PCNA-PIP box interactions. The information gained here could be useful for subsequent drug lead candidate identification. As a second goal of this study, the known anti-tumor agent, EGFR antagonist, gefitinib, was chemically modified and incorporated into subcellularly-targeted peptoid-peptide hybrid complexes. It is hypothesized that targeting this agent into the nucleus of tumor cells can allow for bypassing mechanisms of drug resistance observed in various tumor types, particularly triple negative breast cancer. Enhanced cellular and nuclear uptake was observed with the drug conjugates. Furthermore, some of the compounds demonstrated the ability to selectively downregulate STAT3 phosphorylation, while not affecting the phosphorylation of ERK1/2, and had enhanced antiproliferative activity in gefitinib-resistant cells. Further studies are needed to assess the mechanism of action of these molecules, but they hold promise as agents that can be used to treat drug-resistant cancers.

Degree

Ph.D.

Advisors

Davisson, Purdue University.

Subject Area

Biochemistry|Organic chemistry|Pharmacy sciences

COinS