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Changes in glycosylation have been well-documented with regard to the onset and progression of cancer. Pervasive amongst these is the overexpression of cell surface sialic acid, a shared characteristic of myriad cancer types. Hypersialylation has been noted as a defining feature of the malignant phenotype, playing numerous functional roles in promoting cancer cell survival and invasivity, and a great body of research has sought to unravel its multifaceted contributions to metastatic advancement. A complete understanding of the impact elevated sialic acid has on cancer progression has been hindered by a lack of tools for its direct study. Glycan biosynthesis is not genomically encoded, disqualifying commonplace molecular biology techniques such as mutagenesis and fluorescent protein fusions. This has motivated the development of novel chemical approaches to fill this void. In particular, bioorthogonal chemical methods have accomplished the direct tagging and imaging of sialylated glycans, enabling precise tracking within complex cellular milleu. As an additional challenge, the cancer disease state comprises intimate interactions between tumor and host cells; a comprehensive grasp of sialic acid’s involvement in cancer must be obtained through the use of animal models to recapture the physiological environment. Current in vivo models, primarily mice, are severely limited in their ability to observe cancer behavior at the cellular scale. An alternative model, the embryonic zebrafish, surmounts this barrier and permits the real-time detailed observation of cancer cell dynamics in vivo. Utilization of the zebrafish model’s imaging capabilities may deepen our mechanistic understanding of sialic acid’s role in cancer. The work presented in this dissertation catalogs efforts to study cancer sialylation in zebrafish using bioorthogonal chemistry and other approaches. The first chapter reviews advancements made in the field of in vivo bioorthogonal cancer labeling. Recent years have seen the emergence of varied approaches to target cancer in mouse models for imaging and therapy, using both metabolic incorporation and antibody-pretargeting strategies. This survey of the literature highlights the potential for continually evolving bioorthogonal technology to play a role in human health, and also illuminates the challenges associated with bioorthogonal chemistry-based approaches within mice, creating a framework for discussion of bioorthogonal labeling as applied to a zebrafish model of cancer.Chapter 2 describes the development of methods for the bioorthogonal labeling of cancer-associated sialic acid in a zebrafish xenograft model. Through the use of metabolic incorporation of modified sialic acid precursors and two widely popular bioorthogonal reactions, copper-free click chemistry and the tetrazine ligation, selective labeling of the cancer cell surface was achieved with greater sensitivity and spatial resolution than has been demonstrated in mouse tumor models. These efforts lay groundwork for the application of bioorthogonal chemistry to monitor sialic acid distribution on the subcellular scale in zebrafish throughout different stages of cancer progression.Chapter 3 investigates the interplay between cancer cells and the innate immune system in zebrafish. Sialic acid-mediated interactions between cancer and immune cells have been observed previously and their biochemical basis is beginning to be understood. How these interactions impact the behavior of cancer and immune cells in vivo is an ongoing area of research. Embryonic zebrafish are a powerful tool in this regard, as they enable the facile monitoring of cancer and immune cell dynamics through real-time imaging. Through sialidase-treatment of cancer xenografts, we examined the role sialic acid plays in cancer cell survival and recruitment of macrophages and neutrophils. Sialic acid was found to have a significant influence on these processes, the mechanisms of which are currently being deciphered.Finally, Chapter 4 represents an excursion outside the realm of zebrafish cancer biology. Genetically encodable self-labeling protein tags are an invaluable tool for protein imaging, and offer greater temporal control over traditional fluorescent protein tags. However, their large size, often in excess of 20 kDa, can interfere with native interactions. Described herein is a 10 kDa riboflavin-binding protein rationally engineered from riboflavin synthase. This protein undergoes a self-labeling reaction with novel acrylamide-modified flavin fluorophores, creating a covalently bound fluorescent protein tag. Preliminary results indicate that this system is suitable for labeling within cellular environments with a potential application as a small, genetically encodable tag for fluorescence anisotropy measurements of protein hydrodynamics.

A new class of tunable heterophosphole dimeric ligands have been designed and synthesized., A new class of tunable heterophosphole dimeric ligands have been designed and synthesized. These ligands have enabled the first examples of Cu-catalyzed hydrogenation of 2-substituted-1-tetralones and related heteroaryl ketones via dynamic kinetic resolution, simultaneously creating two contiguous stereogenic centers with up to >99 : 1 dr and 98 : 2 er. The ligand-Cu complexes were isolated and characterized by single crystal X-ray, and DFT calculations revealed a novel heteroligated dimeric copper hydride transition state.

This work presents shock tube experiments and kinetic modeling efforts on the pyrolysis and combustion of JP-10. The experiments were performed at 6–8 atm using 2000 ppm of JP-10 over a temperature range of 1000–1600 K for pyrolysis and oxidation equivalence ratios from 0.14 to 1.0. This work distinguishes itself from previous studies as GC/MS was used to identify and quantify the products within the shocked samples, enabling the tracking of product yield dependence on equivalence ratio as well as identifying several new intermediates that form during JP-10’s decomposition. A detailed, comprehensive model of JP-10’s combustion and pyrolysis kinetics was constructed with the help of RMG, an open-source reaction mechanism generation software package. The resulting model, which includes 691 species reacting in 15,518 reactions, was extensively validated against the shock tube experimental dataset as well as newly published flow tube pyrolysis data from Ghent. Most of the important rate coefficients were computed using quantum chemistry. The model succeeds in identifying all major pyrolysis and combustion products and captures key trends in the product distribution. Simulated ignition delays agree within a factor of 4 with most experimental ignition delay data gathered from literature. The presented experimental work and modeling efforts yield new insights on JP-10’s complex decomposition and oxidation chemistry and identify key pathways towards aromatics formation., Naval Air Warfare Center (U.S.) (Contract N68335-09-C-0367), Naval Air Warfare Center (U.S.) (Contract N68335-10-C-0534), United States. Department of Energy. Office of Basic Energy Sciences (Grant DE-FG02-98ER14914), National Science Foundation (U.S.) (Grant 0535604), National Science Foundation (U.S.) (Grant 0312359), United States. Department of Energy. Office of Basic Energy Sciences (DE-SC0001198), Richard Burnes