Author ORCID Identifier

https://orcid.org/0000-0001-8572-0020

Date of Graduation

8-2020

Document Type

Dissertation (PhD)

Program Affiliation

Medical Physics

Degree Name

Doctor of Philosophy (PhD)

Advisor/Committee Chair

Steven Millward, Ph.D.

Committee Member

Seth Gammon, Ph.D.

Committee Member

Pratip Bhattacharya, Ph.D.

Committee Member

Richard Wendt, Ph.D.

Committee Member

Robert Bast Jr., M.D.

Committee Member

David Piwnica-Worms, M.D., Ph.D.

Abstract

In recent decades it has become increasingly clear that induction of autophagy plays an important role in the development of treatment resistance and dormancy in many cancer types. Chloroquine (CQ) and hydroxychloroquine (HCQ), two autophagy inhibitors in clinical trials, suffer from poor pharmacokinetics and high toxicity at therapeutic dosages. This has prompted intense interest in the development of targeted autophagy inhibitors to re-sensitize disease to treatment with minimal impact on normal tissue. We utilized Scanning Unnatural Protease Resistant (SUPR) mRNA display to develop macrocyclic peptides targeting the autophagy protein LC3. The resulting peptides bound LC3A and LC3B—two essential components of the autophagosome maturation machinery—with mid-nanomolar affinities and disrupted protein-protein interactions (PPIs) between LC3 and its binding partners in vitro. LC3-binding SUPR peptides re-sensitized platinum-resistant ovarian cancer cells to cisplatin treatment and triggered accumulation of the adapter protein p62 suggesting decreased autophagic flux through successful disruption of LC3 PPIs in cell culture. In mouse models of metastatic ovarian cancer, treatment with LC3-binding SUPR peptides and carboplatin substantially reduced tumor growth after four weeks of treatment. These results indicate that SUPR peptide mRNA display can be used to develop cell-penetrating macrocyclic peptides that target and disrupt the intracellular PPIs that govern the autophagic machinery.

Keywords

directed evolution, macrocyclic peptides, peptides, mrna display, autophagy, ovarian cancer

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