Graduation Date
12-2018
Document Type
Master's Thesis
Degree Name
Master of Science
Department or Program
Biological Sciences
Department or Program Chair
Meredith Protas, PhD
First Reader
Manuel Lopez, PhD
Second Reader
Kristylea Ojeda, PhD
Abstract
Cyclodextrin (CD) is a cyclic oligosaccharide that has common use clinically as an excipient for drug stability and enhanced delivery. Currently, a highly soluble CD, hydroxypropyl-β-cyclodextrin (HP-bCD), is in phase 2b/3 clinical trials to treat patients with Niemann-Pick Type C disease (NPC). NPC is a lysosomal storage disorder where cholesterol is depleted from the cell membrane, yet it accumulates in the lysosomes/late endosomes along with other lipids and sterols. We have engineered a variety of functionalized versions of bCD to trace its function within the cell and improve upon its therapeutic performance. The modified bCDs have aided in demonstrating bCD cellular internalization and localization to the NPC1 late endosomal/lysosomal compartment. Filipin staining for free cholesterol accumulation and bCD mass spectrometry analysis suggest that the rate of intracellular uptake of bCD is limited by the concentration of bCD on the exterior of the cell. The amount of storage reduction seen in NPC cells is in turn dependent on the internal cellular concentration of bCD. Thus, bCDs designed for enhanced membrane or receptor mediated uptake were more potent at reducing cholesterol storage in a NPC1 null cell line.