The mechanism or a thermal amino group transfer-fragmentation reaction yielding carboxyamides from mixed phosphoric-carboxylic anhydrides (RO(R¹R²N)P(O)OC(O)R³; R = R¹ = alkyl; R² = H, alkyl, aryl; R³ = alkyl, aryl) was elucidated from structure reactivity studies using a model system, R = R¹ = R² = Me, R³ = Ph. Kinetic data was obtained using ¹H nmr spectroscopy; MNDO molecular orbital and molecular mechanics calculations, and the crystal structure or N-methyl-2-benzoyloxy-2-oxo-1, 3, 2-oxazaphosphorinane (Pna2₁; a = 22.229(6)Å, b = 7.597(2)Å, c = 7.210(2)Å; v = 1217.6(6)ų. Final R = 3. 08% for 1037 reflections with I (rel )> 2αI (rel) and 15 7 parameters) were userul in providing additional in formation about the reaction mechanism .

The methods available for the calculation of ratios of partition functions for isotopic molecules were examined critically and compared. A number of calculations were made for carbon as well as for selenium systems using the exact method rather than an approximate one. Improvements were made to the sample handling system of the mass spectrometer to prevent sample cross-contamination and to reduce the time needed for sample changeover. Four sets of experiments were performed. The kinetic isotope effects in the reductions to Se° of Se^V1 and Se^1V were studied as were the isotope exchange reactions between Se° and Se^1V and between Se^1V and Se^V1. Estimates were made of the ratios of rate constants and isotope exchange equilibrium constants for 82Se and 76Se in these systems and these were compared with the theoretical values. / Thesis / Doctor of Philosophy (PhD)

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Background: Although lymphoma is one of the most common malignancies in dogs, remission rates and survival times remain stagnant. Treatment with a multi-agent chemotherapy protocol induces remission for less than one year and the majority of patients relapse. Fewer than 25% of dogs live longer than two years with the currently available treatments. Targeted immunotherapy using checkpoint molecule blockade of PD-1 and PD-L1 shows promise for various types of human cancer, including relapsed/refractory lymphoma; however, little is known regarding the role of these checkpoint molecules in canine lymphoma. Objectives: To determine the patterns of expression of mRNAs encoding PD-1 and its ligands PD-L1 and PD-L2 in lymphoma and reactive lymphoid hyperplasia controls. Methods: Retrospective: formalin-fixed paraffin-embedded (FFPE) tissue from dogs with untreated lymphoma (n=10) and reactive lymphoid hyperplasia (n=10). Prospective: fine-needle aspirates (FNAs) from dogs with untreated lymphoma (n=10) and reactive lymphoid hyperplasia (n=10). Total RNA was extracted, and expression of PD-1, PD-L1, and PD-L2 was measured using qRT-PCR analysis of random-primed cDNA. Checkpoint molecule expression levels were determined using the 2^∆∆CT method. Lymphoma immunophenotype was assessed using immunohistochemical analysis of CD3 and CD79a (FFPE) and review of patient medical records (FNA). Data analysis included Wilcoxon ranksum tests, Dunn's procedure of multiple comparisons, Kruskal-Wallis tests, and regression within an ANOVA. Significance at P < 0.05. Results: PD-1, PD-L1, and PD-L2 expression (normalized internally to 18S rRNA) was lower in lymphoma compared to reactive lymphoid hyperplasia (FFPE); the difference was significant for PD-1 and PD-L2. PD-1 and PD-L2 expression was lower in lymphoma compared to reactive lymphoid hyperplasia (FNA); the difference was significant for PD-1. PD-1, PD-L1, and PD-L2 expression was lower in B cell lymphoma compared to reactive lymphoid hyperplasia (FFPE); this difference was significant for PD-1 and PD-L2. PD-1 and PD-L2 expression was lower in B cell lymphoma compared to reactive lymphoid hyperplasia (FNA); the difference was significant for PD-1. The higher relative abundance of PD-L1 vs PD-1 and PD-L2 vs PD-1 was significantly different between lymphoma and reactive lymphoid hyperplasia (FFPE and FNA). Conclusions: In this study, checkpoint molecule expression was not upregulated in canine lymphoma relative to canine reactive lymphoid hyperplasia, suggesting a limited application of PD-1 and PD-L1 blockade in canine lymphoma. The ligand:receptor relative abundance imbalances reflect the lower PD-1 expression relative to PD-L1 and PD-L2 in lymphoma. Although these results do not suggest that checkpoint inhibitors would be useful for treatment, they give insight into the mechanisms of unchecked lymphocyte proliferation in canine lymphoma. / Master of Science / Lymphoma, a cancer of the white blood cells in the body, is one of the most common malignancies in dogs. Although treatment with a multi-agent chemotherapy protocol results in high remission rates, the remission duration is usually less than one year, with the majority of patients relapsing. In an effort to improve remission rates and survival times, scientists have been working to develop therapeutic interventions that target specific points in the development and replication cycle of a cancer cell. One such strategy, targeting checkpoint molecules programed death (PD)-1 and PD-L1, has shown promise for several different types of human cancers, including lymphoma. PD-1 is a receptor on T cells, which together with its ligands, PD-L1 and PD-L2, decreases lymphocyte function when activated. This is a protective mechanism, acting to inhibit sustained harmful inflammation in a normal healthy dog. Some cancers have taken advantage of this pathway, increasing expression of PD-L1 or L-L2 in order to evade detection by the immune system. To date, little is known regarding the role and expression of these immune checkpoint molecules in dogs with lymphoma. We sought to evaluate if PD-1, PD-L1 and PD-L2 expression is significantly increased in canine lymphoma compared to reactive lymphoid hyperplasia controls. Tissue samples were collected from two sources. Cytology samples of lymphoma and reactive lymphoid hyperplasia were collected by fine needle aspiration from clinical patients. Formalin fixed paraffin embedded tissue samples of lymphoma and reactive lymphoid hyperplasia were collected from the archived tissue bank. Using a molecular analysis technique called quantitative reverse transcription PCR (qRT-PCR) we measured the amount of messenger RNA (mRNA) encoding PD-1 and its ligands PD-L1 and PD-L2 in lymphoma and in reactive lymphoid hyperplasia controls. In our results we did not observe an upregulation in the expression of checkpoint molecules in canine lymphoma relative to canine reactive lymphoid hyperplasia. This suggests there may be a limited therapeutic application for PD-1 and PD-L1/PD-L2 blockade in canine lymphoma. Although these results do not suggest that checkpoint inhibitors would be useful for treatment, they give insight into the mechanisms of unchecked lymphocyte proliferation in canine lymphoma.

N cadherin is one member of the cadherin superfamily. It like other classes of cell adhesion molecules (CAMs and FAKs) is responsible for cell-cell adhesion ... specifically during early morphogenesis. · N cadherin is the designated cell adhesion molecule for neural tissue. It, in concert with several signal transduction pathways, allows an early neurite to follow chemical and adhesive gradients from its origin to its destination. N cadherin can only operate as an affector of neurite or growth cone guidance when in complex. As a mediator of information from the outside environment to the inside cytoskeleton, N cadherin must rely on cooperation with PTPµ ( a protein tyrosine phosphatase), ~ and a catenin to make firm connection with actin. With that connection established, an early growth cone can make decisions based on chemical and adhesive cues. Movement of the growth cone, as a function of adhesion or "homophilic binding," is termed "haptotaxis." Intracellular concentrations of catenin pools and the activity of two pathways (Src and Wnt) regulate the entire system. Based on studies and experimental data, a model, with respect to complex orientation and behavior is proposed.

Inelastic collisions between dipolar molecules, assumed to be trapped in a static electric field at cold (> 10−3K) temperatures, are investigated and compared with elastic collisions. For molecules with a Λ-doublet energy-level structure, a dipole moment arises because of the existence of two nearly degenerate states of opposite parity, and the collision of two such dipoles can be solved entirely analytically in the energy range of interest. Cross sections and rate constants are found to satisfy simple, universal formulas. In contrast, for molecules in a Σ electronic ground state, the static electric field induces a dipole moment in one of three rotational sublevels. Collisions between two rotor dipoles are calculated numerically; the results scale simply with molecule mass, rotational constant, dipole moment, and field strength. It might be expected that any particles interacting only under the influence of the dipole-dipole interaction would show similar behavior; however, the most important and general result of this research is that at cold temperatures inelastic rate constants and cross sections for dipoles depend strongly upon the internal structure of the molecules. The most prominent difference between the Λ-doublet and rotor molecules is variation of the inelastic cross section with applied field strength. For Λ-doublet dipoles, cross sections decrease with increasing field strength. For rotor dipoles, cross sections increase proportionally with the square of field strength. Furthermore, the rate constants of the two types of molecules depend very differently on the angular orientations of the dipoles in the electric field.

A 10.6 ym C02 laser operating a power range S P 200 watts was used to pump some select vibrational transitions in the NO2 molecule while monitoring the rotational transitions (91/9—'100/10), (232f 22 ~~"*242,23> ' (402,38 "393,37) in the (0, 0, 0) vibrational level and the (8q,8—*"^1,7) rotational transition in the (0, 1, 0) vibrational level. These rotational transitions were monitored by microwave probing to determine how the population of states in the rotational manifolds were being altered by the laser. Coincidences between some components of the V3-V2 band of N02 and the C02 infrared laser lines in the 10 um region appeared to be responsible for the strong interaction between the continuous laser beams and the molecular states.

Although many signalling molecules appear relevant to the production of complex behaviours, those that are important to the physiological regulation of behaviour, and so those that characterise individual styles of behaviour, are unknown. Vasopressin is the strongest candidate regulator of social behaviour. Experiments were carried out in consideration that vasopressin may directly regulate aggressive behaviour in lactating rats. Patterns of immediate early gene expression during/subsequent to aggressive behaviour suggested specific neural circuits may have significant direct regulatory influence over particular behaviours, and that activation of the V1b vasopressin receptor, in these circuits, may contribute to this putative regulatory signalling. In situ hybridisation studies indicated that patterns of vasopressin release, rather than receptor expression, might be important for any peripartum changes in behaviour driven by vasopressin. Although their relative importance is unknown, central actions of vasopressin may exert a strong regulatory influence over a range of behaviours, across a range of species.

published_or_final_version / Biochemistry / Doctoral / Doctor of Philosophy