Date of Award
Doctor of Philosophy
Michael D. Best
John E Bartmess, Brian Long, Todd Reynolds
Liposomal drug delivery shows great promise for developing smart delivery materials, especially when a cytotoxic drug needs to be administered. The liposome membrane bilayer structure reduces toxicity through encapsulation, circumventing side effects. Current work in this field is aimed at selectively targeting delivery to diseased cells and controlling the release of encapsulated drugs.
In the first project of this dissertation, we describe the synthesis of a clickable and photocleavable lipid analog for efficient drug delivery to cells and subsequent triggered release. This lipid analog contains a cyclooctyne moiety by which azide-containing drug molecules or fluorophores can be attached using copper-free click chemistry and a photocleavable group for light-driven release. We have validated conjugation of this lipid analog via click chemistry using fluorescence-based liposome derivatization studies and a liposome immobilization microplate assay. Next, fluorescence microscopy was used to show successful delivery of fluorescent dyes to cells and subsequent release. Finally, this lipid was applied for liposomal delivery of camptothecin to cancer cells, which led to a dose-dependent decrease in cell viability.
Metabolic labeling of biomolecules is another topic of interest described in this dissertation. In this project, we sought to design, synthesize and study click-tagged serine analogs that could produce labeled versions of phosphatidylserine in cells to investigate lipid production, trafficking, and to understand the role of lipids in the instigation of cancer and other diseases. In studies, either Saccharomyces cerevisiae or human cells were treated with these probes, followed by incubation with a clickable fluorescent reagent and confocal fluorescence microscopy to detect labeled products. To determine the labeled lipids, the extracted cell samples were subjected to tandem mass spectrophotometry and a number of phospholipids were detected. Lastly, we also showed that click labeling of PS, a lipid upregulated on the outer leaflet of cancer cell membranes, can be utilized as a tool for delivery the delivery of liposomes bearing partner clickable tags, which has been confirmed via a confocal microscope. Finally, we have synthesized a number of L-serine analogs as potential PS synthase inhibitors and worked to evaluate the efficacy of these compounds using a PS synthase assay.
Alam, Shahrina, "Development of New Methods for Liposomal Drug Delivery and the Labeling of Cellular Lipids. " PhD diss., University of Tennessee, 2016.