Author Type

Graduate Student

Date of Award

2024

Document Type

Dissertation

Publication Status

Version of Record

Submission Date

October 2026

Thesis/Dissertation Advisor [Chair]

Stawikowski, Maciej J.

Abstract

Cholesterol is a pivotal component of mammalian cell membranes and homeostasis. Due to its high concentration and heterogenous distribution in the brain, cholesterol is tightly regulated and its dyshomeostasis is frequently implicated in several neurodegenerative diseases. This dissertation reports the design, synthesis, and application of ten cholesterol naphthalimide probes (CND) to study cholesterol trafficking in live cells. The CND series was rationally designed by incorporating several of the structural features of endogenous cholesterol onto the naphthalimide (ND) scaffold conjugated via an ester bond. The modularity of the ND scaffold enabled all analogs to have the aliphatic tail of cholesterol, which is lacking in the most ubiquitously utilized probe, BODIPY-Cholesterol, a.k.a. Top-Fluor® Cholesterol (TFC). The CNDs were demonstrated to be optimal probes for cholesterol due to the ability to fluorescence exclusively in hydrophobic/membrane environments and exhibit low fluorescence in hydrophilic/aqueous environments. By incorporating a protonatable piperazine group on the C4 position of the ND scaffold, CND2 – CND4 possessed pH sensing capabilities, which were demonstrated to monitor intracellular vesicle turnover in neurons. The potential of the CNDs to bind to the lysosomal sterol transport protein NPC2 was investigated by molecular docking and molecular dynamics (MD) simulations. The docking pose with the CND’s aliphatic tail positioned inside the hydrophobic binding pocket was essential for mimicking endogenous cholesterol’s interactions and stabilizing the NPC2-ligand complex. Fluorescence confocal microscopy demonstrated a structure-dependent and cell-dependent intracellular distribution of the CND series in live cells.

Extent

273 p.

Comments

Includes bibliography.|Dissertation (PhD)--Florida Atlantic University, 2024.|FAU Electronic Theses and Dissertations Collection|Includes bibliography.

Identifier

FA00014430

Format

pdf

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