Raul Iglesias

Graduation Year

2016

Document Type

Dissertation

Degree

Ph.D.

Degree Name

Doctor of Philosophy (Ph.D.)

Degree Granting Department

Chemical Engineering

Major Professor

Piyush Koria, Ph.D.

Committee Member

Nathan Gallant, Ph.D.

Committee Member

Subhra Mohapatra, Ph.D.

Committee Member

Venkat Bhethanabotla, Ph.D.

Committee Member

Mark Jaroszeski, Ph.D.

Keywords

ELP, Lytic Peptide, Growth Factor, Receptor

Abstract

Over the past few years, researchers have focused their attention on the development of targeted cancer therapies to minimize the side effects associated with non-targeted treatments such as chemotherapy. Specifically, these approaches have focused on blocking growth factor receptors (GFR) that are overexpressed in cancer cells. In this thesis, we also focus on targeting overexpressed GFR; however, instead of blocking the GFR, our novel approach aims at using them to selectively enhance the endocytotic process of macropinocytosis to deliver peptides that either disrupts the mitochondria or inhibits glycolysis.

Herein, we show the selective enhancement of macropinocytosis by the fusion protein comprised of the keratinocyte growth factor (KGF) fused to elastin like polypeptide (ELP), KGF-ELP. Furthermore, we report the synthesis of the fusion protein consisting of mitochondriotoxic peptide (KLAKLAK)2 with ELP, (KLAKLAK)2-ELP. We show that (KLAKLAK)2-ELP forms nanoparticles (NPs) that are internalized via macropinocytosis and their internalization is facilitated by the interaction between the ELP domain and heparan sulfate proteoglycan (HSPG) on the cell surface. This internalization results in mitochondrial swelling, depolarization and subsequent cell death. Moreover, we show that heterogeneous NPs comprising of the two fusions KGF-ELP and (KLAKLAK)2-ELP selectively kill lung cancer cells expressing the keratinocyte growth factor receptor (KGFR).

We also report the synthesis of the fusion consisting of peptides derived from a phosphorylated domain of the glycolytic enzyme phophoglycerate mutase (PGM) and ELP, PGM-ELP. We demonstrate that this fusion inhibits the step in glycolysis that converts 3-phosphoglycerate (3PG) to 2-phosphoglycerate (2PG); the results show that cell death occurred preferentially in lung cancer cells compared to normal cells. Additionally, the heterogeneous NPs comprising of KGF-ELP and PGM-ELP selectively enhanced killing in lung cells with high levels KGFR. Finally, the synthesis of a fusion proteins consisting of four PGM domains fused to ELP, (PGM)4-ELP, exhibits higher cytotoxic effect and efficiency when compared to the single PGM domain fusion, PGM-ELP.

Overall, we conclude that targeting overexpressed growth factor receptors to stimulate macropinocytosis can be a tremendously selective therapy for the treatment of lung cancer. This can result in attenuating side effects and improvement of the patient’s prognosis

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