Date of Award

2015

Document Type

Dissertation

Degree Name

Doctor of Philosophy (PhD)

Department

Cell and Molecular Pharmacology and Experimental Therapeutics

College

College of Graduate Studies

First Advisor

Richard Drake

Second Advisor

Lauren Ball

Third Advisor

Sebastiano Gattoni Celli

Fourth Advisor

Steven A. Rosenzweig

Abstract

A deeper understanding of dysregulated glycosylation in pancreatic cancer can provide insights into disease mechanisms and the identification of novel disease markers. Recent improvements in mass spectrometry techniques have been instrumental in profiling biologically relevant tissue sections in order to identify disease marker candidates, but have either not yet been adopted for studying glycosylation or applied directly to pancreatic cancer. In the dissertation herein, new methods have been developed and adapted to the study of aberrant glycosylation in pancreatic cancer, with the ultimate goal of identifying novel disease marker candidates. For the first time, we describe a mass spectrometry imaging approach to study the localization of N-glycans. This technique demonstrated a histology-derived localization of N-glycans across tissue sections, with identifications displaying remarkable consistency with documented studies. Furthermore, the technique provides superior structural information compared to preexisting methodologies. In the analysis of diseased specimen, changes in glycosylation can be linked to aberrations in glycosyltransferase expression. When applied to pancreatic cancer in a high-throughput and high-dimensional analysis, panels of glycans displayed an improved ability to differentiate tumor from non-tumor tissues compared to current disease markers. Furthermore, the data suggest that glycosylation can identify premalignant lesions, as well as differentiate between malignant and benign conditions. These observations overcome significant limitations that hinder the efficacy of current disease markers. In an effort to link aberrant glycosylation to the modified protein, a subset of glycosylated proteins were enriched and analyzed by mass spectrometry to identify proteins that are integral to disease progression and can be probed for the early detection of pancreatic cancer. Known disease markers were among the glycoproteins identified, validating the utility of the enrichment and detection strategy outlined. This approach also differentiated the role of N- and O-glycosylation in antigen expression. Finally, we outline an integrated workflow that takes advantage of the unique capabilities of high resolution mass spectrometers. This workflow can capitalize on prior glycomic and proteomic experiments to provide a comprehensive analysis of dysregulated protein glycosylation in pancreatic cancer.

Rights

All rights reserved. Copyright is held by the author.

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