DOI

10.17077/etd.s2w0teuh

Document Type

Dissertation

Date of Degree

Spring 2018

Degree Name

PhD (Doctor of Philosophy)

Degree In

Pharmacy

First Advisor

Rice, Kevin G.

First Committee Member

Strack, Stefan

Second Committee Member

Duffel, Michael W.

Third Committee Member

Roman, David

Fourth Committee Member

Doorn, Jonathan A.

Abstract

Gene therapy can potentially treat a wide range of diseases ranging from inherited diseases to cancer. The successful use of nucleic acids to treat genetic diseases is limited by rapid capture and degradation of the nanoparticle by Kupffer cells in the liver. Scavenger receptors on the cell surface, capture both viral and non-viral nanoparticles leading to reduced efficacy. PEG-peptides were found to inhibit scavenger receptors on the surface of Kupffer cells by forming albumin nanoparticles when intravenously dosed. This work explores the development of potent, low-molecular weight PEG-peptide inhibitors. In order to study the in vivo activity of the nanoparticle, an in vivo assay was developed to directly assess the potency of inhibition. High molecular weight polylysine peptides (33.5 kDa) inhibited liver uptake with an IC50 of 18 μM. Incorporation of four leucine residues, to improve albumin binding, allowed for a decrease in PEG molecular weight and number of lysine residues, resulting in PEG5kda-Cys-Tyr-Lys-(Leu-Lys4)3-Leu-Lys (7.4 kDa) that inhibited scavenger receptors with an IC50 = 20 μM. Further decrease in the PEG molecular weight resulted in the discovery of PEG2kDa- Cys-Tyr- (Leu-Lys4)3-Leu-Lys (4.4 kDa) with potency of 3 μM. The increase in potency could be attributed to a decrease in the zeta potential of the albumin nanoparticle resulting in more efficient scavenger receptor mediated uptake. Co- administration of PEG2kDa- Cys-Tyr-(Leu-Lys4)3-Leu-Lys with a stable PEGylated polyacridine DNA polyplex resulted in inhibition of rapid polyplex uptake by the liver with an IC50 = 11 μM. Other properties including spatial distribution of leucine, hydrophobicity and peptide length were also explored to determine their effect on liver uptake. Hydrophobic peptides resulted in the formation of micelles which were inactive as scavenger receptor inhibitors and exhibited increased liver uptake upon dose escalation. Reduction in the peptide length resulted in peptides that were not captured by the liver. Inhibition scavenger receptors has the potential to improve the efficacy of viral and non-viral nanoparticles. The findings of this work provide a framework for the development of PEG-peptide inhibitors capable of blocking live uptake of viral and non-viral nanoparticles.

Keywords

Gene delivery, Kupffer Cells, Liver, PEG-peptide, Scavenger Receptor

Pages

xxi, 166 pages

Bibliography

Includes bibliographical references (pages 155-166).

Copyright

Copyright © 2018 Rondine Joni-Ann Allen

Share

COinS