Theses and Dissertations at Montana State University (MSU)
Permanent URI for this collectionhttps://scholarworks.montana.edu/handle/1/733
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Item When a lectin binds a sugar, and other sweet tales(Montana State University - Bozeman, College of Letters & Science, 2019) Bernhard, Samuel Pruitt; Chairperson, Graduate Committee: Mary J. Cloninger; Mackenzie S. Fricke was an author and Katharina Achazi, Paul Hillman, Willy Totten, Rainer Haag and Mary J. Cloningerwere co-authors of the article, 'The toxicity, uptake, and impact on galectin-3 mediated apoptosis of lactose functionalized dendrimers' submitted to the journal 'Biomolecules Special Issue: Moving Forward with Dendrimers' which is contained within this dissertation.The current state of chemotherapy and cancer treatment leaves much to be desired. Treatment is generally non-specific and relies on high dosage to achieve therapeutically relevant concentrations at target sites. Glycopolymer-drug conjugates, featuring targeting molecules and therapeutic prodrug on a water-soluble polymeric scaffold, offer a solution to these contemporary problems. Here, the complexity of glycopolymer design is explored through the lens of a biologically significant carbohydrate-binding receptor. In particular, galectin-3 is a complex Beta-galactoside binding lectin that experiences altered expression in many cancer pathologies and is implicated in metastasis, angiogenesis and poor overall prognosis. Galectin-3 mediates undesired cancer promoting processes through carbohydrate binding and oligomerization. A more complete understanding of the role galectin-3 plays in cancer progression will guide development of methods in the therapeutic intervention of these processes. In the interest of understanding galectin-3 and using it as a targeted receptor, its binding characteristics have been assessed through fluorescence lifetime and dynamic light scattering measurements. Employment of carbohydrates and glycopolymers including mannose, lactose, and lactose functionalized poly(amidoamine) (PAMAM) dendrimers, dendritic polyglycerols (dPG), and linear polymers (LP) provided insight into the carbohydrate binding avidity of galectin-3 and its propensity to oligomerize or form micron scale aggregates. A relationship between scaffold size and receptor recruitment was observed, which sheds light into multivalent binding motifs initiated by these glycopolymers and establishes a threshold for minimum requisite lactose functionality on lactose functionalized dendritic polyglycerols. In vitro cell based glycopolymer studies with AlexaFluor 647 and lactose functionalized PAMAM dendrimers revealed size-dependent uptake and demonstrated that accumulation occurs within the lysosome. Cellular aggregation experiments revealed that lactose functionalized LPs and dPGs influence galectin-3 mediated homotypic cellular aggregation and, in fact, augment this aggregation through receptor recruitment and cross-linking. The results reported here have provided a more fundamental understanding of galectin-3 binding interactions and have laid the groundwork for optimized glycopolymer-drug conjugate design.Item Glycodendrimer mediation of galectin-3 cancer processes and indium(III) as a glycosylation promoter(Montana State University - Bozeman, College of Letters & Science, 2014) Michel, Anna Kaczmarek; Chairperson, Graduate Committee: Mary J. CloningerGalectin-3 is a carbohydrate-binding protein that is found inside and at the surface of most healthy cells, where it plays a role in cellular differentiation, proliferation, and death. Galectin-3 is also overexpressed in many forms of cancer, where it interacts with beta-galactosides on the surface of the transmembrane protein Mucin 1 (MUC1), which is also upregulated in many cancers. The interaction between MUC1 and galectin-3 leads to enhanced tumor formation, invasion, and metastasis. Glycodendrimers were synthesized and used as tools to probe the galectin- 3/MUC1 mediated cancer cellular aggregation. Cellular aggregation assays were designed and performed using three different cancer cell lines. Results indicate that cellular aggregation can either be inhibited or intensified depending on the size of the dendrimer and on the number of carbohydrate endgroups present. Confocal micrographs of cancer cells confirm that the pattern of galectin-3 on the outside of the cell is altered in the presence of glycodendrimers. These finding indicate that glycodendrimers mediate cellular aggregation properties, giving insight into the mechanism of action. The results also establish glycodendrimers as potential targeting agents and prodrug delivery systems when considering cancer drug design. A dendrimer bearing a matrix metalloproteinase (MMP) substrate was synthesized to explore dendritic potential as drug delivery systems. Results show that the substrate can be cleaved from the dendrimer when incubated with cancer cells known to overexpress MMP, indicating a potential for glycodendrimers as a prodrug carrier. A novel method for glycosylation was also explored. Indium(III) provides many advantages over standard glycosylation promoters (such as BF 3 x OEt 2) in that it is not air-or water-sensitive, and there is no need for additional purification of In(III) before use. Carbohydrate donors with a variety of protecting groups were tested against a range of glycosyl acceptors, including a carbohydrate. Results show that using In(III) as a glycosylation promoter gives comparable yields in less time when compared to a common glycosylation promoter.