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Theses and Dissertations

This collection contains theses and dissertations of graduate students of the University of Alberta. The collection contains a very large number of theses electronically available that were granted from 1947's to 2009, 90% of theses granted from 2009-2014, and 100% of theses granted from April 2014 to the present (as long as the theses are not under temporary embargo by agreement with the Faculty of Graduate Studies and Research). IMPORTANT NOTE: To conduct a comprehensive search of all UofA theses granted and in University of Alberta Libraries collections, search the library catalogue at - you may search by Author, Title, Keyword, or search by Department. To retrieve all theses and dissertations associated with a specific department from the library catalogue, choose 'Advanced' and keyword search "university of alberta dept of english" OR "university of alberta department of english" (for example). Past graduates who wish to have their thesis or dissertation added to this collection can contact the ERA Mediated HelpDesk at

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    Creator: Jamro, Ghulam M
    Description: Peat mineral soil mix (PMM) and LFH, identifiable litter (L), fragmented litter (F) and humus (H), mineral soil (MS) mix are organic capping materials commonly used over overburden (OB) and tailings sand (TS) substrate materials in oil sands reclamation. These organic capping materials have different biological properties and nutrient availabilities due to differences in the carbon to nitrogen (C to N) ratio, exogenous organic input and organic to MS ratio. Substrate materials inherently possess high pH, electrical conductivity (EC) and soil compaction. The main goal of this research was to evaluate biogeochemical and fine root processes affected by required reclamation practices for oil sands reclamation. Soil sampling was conducted from 0 to 10 and 10 to 20 cm soil depth from June to October in 2011 and 2012 and N availability and enzyme activities were analyzed. Organic substrate type and diversity effects on microbial processes including carbon dioxide (CO2) emission, enzyme activities, available N and community level physiological profiles (CLPPs) were evaluated in a laboratory experiment using three organic substrates (glucose, acetic acid, alanine). The organic substrates were applied singly and in a mixture of two or three in an LFH-MS and a PMM. Effects of organic to MS ratio on biogeochemical processes were evaluated in a laboratory experiment using five ratios of LFH or peat to MS at 0:100, 30:70, 50:50, 70:30 and 100:0. Fine root properties such as root length density, surface area, total root biomass and rates of root production, turnover and decomposition of lodgepole pine and white spruce planted on the PMM placed over TS and OB substrates, respectively and were assessed from May to October in 2011 and 2012. The N availability and N-acetyl glucosaminidase, arylamidase and protease activities were greater in LFH-MS than in PMM, decreased along the soil depth and were influenced by the time of sampling. These differences were attributed to the lower C to N ratio in LFH-MS than in PMM. The addition of fresh labile C through root exudates and litter fall likely induced the N availability and enzyme activities in fall rather than in summer. The addition of organic substrates (laboratory study) significantly segregated CLPPs from the control (no substrate) in LFH-MS and PMM. The significant increase in enzyme activities, available N through increasing the organic substrate diversity, was likely associated with changes in CLPPs and a reduction in C to N ratio with a substrate addition in LFH-MS and PMM. The mixing of organic single substrates enhanced the CO2 emission rate and NO3--N concentration only in LFH-MS and PMM. The laboratory study results revealed that the β-glucosidase, cellobiohydrolase, phenol oxidase and leucine aminopeptidase activities, CO2 emission rates and available N were increased along the increasing organic to MS ratio regardless of LFH and peat. The increase in soil processes was due to changes in C to N ratios and pH along with increase in organic to MS ratios. Fine root length density, fine root production and turnover rates were increased along the low < medium < high productivity level in pine stands, and were positively correlated with tree height and diameter at the breast height. Fine root surface area was the only parameter that was increased along the productivity gradient in spruce. These differences were attributed to negative relationships of EC and soil compaction with root properties in pine and spruce, respectively. The root decomposition did not change along the productivity level of both pine and spruce species but was affected by the time of incubation, due to differences in species and OB and TS properties. The LFH-MS is a better soil quality organic capping material than PMM due to N availability and enzyme activities; however, the availability of LFH-MS is limited for reclamation. Hence, the ratios of organic to MS for optimization, particularly LFH, help to take advantage of available LFH material. Alternatively, augmenting LFH-MS with PMM can overcome the limitations of both materials, as indicated from organic substrate diversity and organic to MS ratios in this research. An evaluation of the effects of substrate properties on fine root properties can help improve current reclamation practices.
    Subjects: Oil sands reclamation, Biogeochemistry, Fine roots, Soil Enzymes, Organic capping materials, LFH-mineral soil mix, Ratio of materials
  2. (RADA)4 Self-Assembling Peptide based Hydrogels: Design, Characterization and In-Vitro Biological Evaluation [Download]

    Title: (RADA)4 Self-Assembling Peptide based Hydrogels: Design, Characterization and In-Vitro Biological Evaluation
    Creator: Saini, Aditi
    Description: Ionic self-assembling peptides have emerged as promising nano-biomaterials, with direct applications in the fields of bioengineering and applicative medicine. (RADA)4 is a self-assembling 16 residue containing peptide, with alternating hydrophobic and hydrophilic residues. This characteristic amphiphilic nature of the peptide allows it to self-assemble into β-sheets, forming higher order three-dimensional (3D) structures. The resulting 3D hydrogels are non-immunogenic, highly hydrated (containing > 99% water), and can respond to physiological stimuli. The (RADA)4 hydrogels can serve as ECM (extra cellular matrix) scaffolds, assist in drug delivery, and can achieve rapid hemostasis. This dissertation consists of two major parts: 1) fundamental study of the self-assembly of the model ionic complementary peptide (RADA)4, and 2) in-vitro biocompatibility assessment of the (RADA)4 peptides. In the fundamental study, the secondary structure of (RADA)4 was examined with Circular Dichroism (CD), and compared to that of its two variants: (RADA)4K5 and (RADA)4S5. The effect of peptide concentration and temperature on the secondary structure was also studied. It was found that all but (RADA)4K5 peptides formed successful β-sheets, consequently forming nanofibers, whereas (RADA)4K5 resulted in the formation of aggregates rich of primarily random coil sequences. Further, studies were conducted to determine the critical concentration of (RADA)4K5 for successful nanofiber formation. The 3D nanostructure of the peptide was affected by the amino acid sequence as well as by the temperature induced denaturation of the peptide. Quantitative structural analysis of all the samples were carried out using an online DICHROWEB server, by comparing the secondary structure molar ellipticity of the peptides collected with that of seven reference proteins (data confirmed via X-ray crystallography). Single-molecule Florescence Correlation Spectroscopy (FCS) was used to confirm the molecular interaction of the pristine (RADA)4 nanofibers with 25% (RADA)4K5. The effect of hydration on these self-assembling peptides was investigated via Differential Scanning Calorimetry (DSC), over a range of temperatures. The Equilibrium Water Content (EWC) in (RADA)4 was comparable to (RADA)4S5 and (RADA)4K5, even at varying compositions. The content of non-frozen bound water increased upon appending either Lysine or Serine residues to the (RADA)4 peptide. Microscopy techniques such as Atomic Force Microscopy (AFM), and Transmission Electron Microscopy (TEM), were also employed to visually inspect the higher order structures formed by these peptides. The second part of this dissertation focuses on the in-vitro biocompatibility of the (RADA)4 based peptides. The PAC-1, CD62-P, and CD42 markers were used to study platelet activation (via Flow Cytometry) and a time based clotting analysis was conducted to evaluate the hemostatic ability of peptides. Complement C3a ELISA assay was conducted with (RADA)4 based peptides to gain more insight into the biocompatibility. The pristine (RADA)4 nanofibers caused a rapid clot formation, but yielded a low platelet activation and low C3a activation. Whereas, (RADA)4K5 peptide displayed a significantly higher complement activation, when compared to both (RADA)4S5, and the (RADA)4 peptide, likely due to the free NH3 groups and steric hindrance in packing. The overall trend of the platelet activation among the three variants of the peptides remained consistent: (RADA)4K5 activated the platelets to the highest, whereas (RADA)4S5, and (RADA)4 showed comparable platelet activation. However, it should be noted that both the platelet and complement activation by these peptides were lower than the already considered biocompatible biopolymers. Lastly, morphology analysis of the platelets in contact with the hydrogels was conducted by calculating the Morphology Score (MS) using a Kunicki scoring system, and the platelets were visualized using Scanning Electron Microscopy (SEM). An overall Kunicki morphology score of above 385 was achieved for all the peptides, where a score of less than 200 represent poor retention of morphological characteristics associated with platelets that are active. Overall, the (RADA)4 based peptides had a lower, or comparable platelet and complement activation, when compared with the already in-use biomaterials (such as poly(methyl methacrylate), and dextran), making them a desirable material to further investigate. The work in this dissertation, not only provides the fundamental knowledge to design novel biomaterials for direct application in medicine, but also provides the stepping stone for further in-vitro and in-vivo biocompatibility analysis, required for any further medical application.
    Subjects: (RADA)4, Peptide, Hydrogel, Nanofiber, Self-assembly, Biocompatibility, Biomaterial
  3. Conduction and Dielectric Relaxation Mechanisms in Oil Sands Influencing Electrical Heating [Download]

    Title: Conduction and Dielectric Relaxation Mechanisms in Oil Sands Influencing Electrical Heating
    Creator: Abraham, Tinu M
    Description: Electrical heating has been proposed in the past as an alternative to conventional water based thermal methods for reducing viscosity of bitumen in oil sands reservoirs. This could reduce or even eliminate water use and associated problems like inefficient heat transfer in the reservoir, poor bitumen recovery as well as produced water treatment issues in the oil sands processing plant. However, four decades since its initial ideation, electrical heating of oil sands is still not commercialized. The reasons are rooted in a lack of understanding about the dynamic electrical heat generation mechanisms in oil sands. This has led to over-dependence of electrical heating on water, resulting in non-uniform and discontinuous heating of the reservoir, as well as overheating of electrodes leading to failure during field trials. This research study therefore gave importance to understanding the dynamic electrical heat generation mechanisms in heterogeneous oil sands as a function of their composition, microstructural arrangement and heating temperatures. The first approach was to determine conduction and dielectric relaxation mechanism in oil sands using impedance spectroscopy studies conducted between 1Hz and 1 MHz and at temperatures between 20 and 200°C. These studies revealed an array of conduction and polarization mechanisms. When water content of oil sands was high (>5%) present as connected water channels, dc conduction was the dominant mechanism via which electrical energy was dissipated as heat. On the other hand when it was low (<5%), water was assumed to be present in isolation at the interface between bitumen and silica grains resulting in interfacial or Maxwell Wagner (MW) polarizations which showed dielectric relaxations between 1 kHz and 1 MHz. Oil sands with least water content (<1%) showed a dominance of conduction relaxations via charge hopping mechanisms following Jonscher’s law owing to the presence of silica grains having conduction through grain and grain boundaries. They also showed dominance of dipole relaxations in bitumen between 100 kHz and 1 MHz. These bitumen relaxations were present in all oil sands irrespective of their water content but were revealed only in cases where water was low. Temperature based studies revealed that beyond 120°C all oil sands behaved similarly, revealing a dominance of conduction relaxations due to silica and dipole relaxations due to bitumen, irrespective of the initial water and clay contents making them low loss heterogeneous dielectrics. Results from the second research approach linking heating patterns to the dynamic electrical behaviour of oil sands shed light on important operational strategies that could be implemented while carrying out electrical heating. A resonant autotransformer was used for electrically heating, probing and controlling the heating iii process via capacitive heating configuration. The studies revealed that joule or ohmic heating could be most suitable for high water content (>5%) oil sands having dominance of dc conduction. Frequency tuned capacitive heating would be useful for oil sands showing a dominant loss peak due to MW polarizations (1 to 5% water). Whereas capacitive or dielectric heating set at the relaxation frequency of bitumen molecules would be most suitable for oil sands with least water content (<1%). Pure capacitive heating could also be most suitable beyond 120°C for all oil sands as they showed similar electrical behavior therefore suggesting that as temperature changes, operational strategies should be varied to catch up with changing electrical behaviour of oil sands. This research study therefore sheds new light on the electrical heat generation mechanisms which could influence efficient electrical heating of oil sands. These findings are expected to improve oil sands extraction process, resulting in cost reduction coupled with reduced impact on environment due to reduction in water usage, and carbon emissions.
    Subjects: oil sands, electrical heating, electromagnetic heating, radio frequency heating, dielectric heating, conduction, polarization, dipole relaxation
  4. Transitioning to University: The Effect of Changes in Academic Engagement and Pressure and Support from Friends on Grades [Download]

    Title: Transitioning to University: The Effect of Changes in Academic Engagement and Pressure and Support from Friends on Grades
    Creator: Vargas Lascano, Dayuma Ixchel
    Description: In Canada, over half of high school graduates attend post-secondary education (PSE); however, up to 32% of these students fail to graduate. Given that PSE completion requires meeting academic performance standards and that persistence in PSE is strongly associated with academic performance, a better understanding of factors that influence PSE students’ academic performance can inform educators’, students’, and other stakeholders’ efforts to improve persistence in PSE. This dissertation longitudinally examines two aspects of students’ motivational systems proposed to impact academic performance following the Self-Systems Motivational Model: academic engagement and friends’ influences. Whereas extensive research exists on the role of academic engagement and friends on students’ academic performance during elementary and junior high school, research on these issues during high school is limited and during PSE is even scarcer. To address this gap, this dissertation examines university-level students’ academic engagement and their perceived academic pressure and support from friends across their first semester of post-secondary education, the relation of friendship and engagement to one another, and their roles in students’ academic performance (GPA). First-year full-time university students (N = 544) were tracked four times across their first semester. Latent growth curve models showed that all aspects of academic engagement changed across the semester, with some differences in patterns of change over time across different aspects of engagement. Students experienced steady declines in in-class behavioral engagement across the semester while experiencing declines in the first half of the semester followed by slower loss and a slight uptake by the end in all other aspects of engagement investigated, with slight differences in rates of change among them (out-of-class behavioral engagement, cognitive engagement, and social behavioral engagement). Different aspects of friends’ influences also showed different patterns of change across the semester, with students experiencing steady increases in school-supportive pressure throughout, decreasing school-obstructive pressure in the first half of the semester followed by slight increases, and increasing academic instrumental support in the first half of the semester followed by slight decreases. Parallel process models showed significant associations between aspects of academic engagement and aspects of friends’ influences at baseline in seven out twelve assessed models. In general, at the beginning of the semester, more academically engaged students experienced higher school-supportive pressure and academic instrumental support from friends and lower school-obstructive pressure from friends compared to less academically engaged students. Students’ experiences of friends’ influences at the beginning of the semester did not predict change over time in students’ engagement nor did students’ engagement at the beginning of the semester predict change over time in students’ experiences of friends’ influences. There were no consistent associations between change in aspects of academic engagement across the semester and changes in aspects of friends’ influences across the semester (significant in two out of twelve models). In terms of academic performance, students who were more academically engaged (all aspects except cognitive engagement) at the beginning of the semester received higher GPAs at the end of the semester. Conversely, students experiencing more school-supportive pressure from friends at the beginning of the semester received lower GPAs. Overall, first year students experienced changes in academic engagement and perceived academic pressure and support from friends across their first semester at university. How much students engage with their academics was linked to their experiences of perceived academic pressure and support from friends at the beginning of the semester and both of these predicted their academic performance outcome at the end of the semester. Students’ academic engagement and friends’ influences experiences across their first university semester were not linked to one another and did not matter for their academic performance once the impact of initial levels of engagement and friends’ influences were taken into accounted. What students bring with them to university, then, in terms of engagement and friendship support seems to impact their academic performance at the end of the semester more than changes in these two areas across the semester. Working with high school teachers and students to help students develop habits linked to academic success in PSE may help PSE institutions improve the academic performance of their students as much as providing academic support services to them once they start their PSE studies.
    Subjects: academic engagement, friend influence, academic performance, longitudinal
  5. A Proposed Experimental Methodology for Assessing the Effects of Biophysical Properties and Energy Content on Live Fuel Flammability [Download]

    Title: A Proposed Experimental Methodology for Assessing the Effects of Biophysical Properties and Energy Content on Live Fuel Flammability
    Creator: Melnik, Oleg
    Description: The effectiveness of fire management tactics and safety of firefighters strongly depend on the reliability of fire behaviour predictions that is currently limited by a lack of understanding of the flammability of live fuel. Until now fire modeling has been primarily based on the flammability of dead fuel using the assumption that combustion of the live fuel has a very limited effect on fire behaviour. However, the analysis of the existing data revealed that live fuel constituted from 48% to 60% of fuel consumed during the passage of a flame-front, meaning that live fuel plays a significant role in determining frontal fire intensity and fire behaviour. Introducing a new definition of flammability and a test method for flammability assessment, this study identifies how and to what extent live fuel and its properties may affect frontal flame intensity. By evaluating flammability directly in a flame, the proposed oxygen consumption calorimetry method better represents the high-intensity combined radiative and convective heat transfer prior to ignition as well as the conditions of oxygen deficiency and high concentrations of water vapor within the flame-front. The flammability of live fuel consumed within the flame-front was defined as energy release contribution to the frontal flame and represented the energy-generation component of the energy balance of the flame-front rather than just an energy content of separate fuel elements. The flammability was measured as the change in energy release from the flame resulting from interaction with live fuel during average flame-front residence time. Assessing the flammability of fresh shoots rather than just foliage allowed for better representation of the live plant material consumed in the flame-front. Live fuel flammability range, factors, and seasonal trend were investigated on a tree branch scale and a new high-resolution volume measuring technique was also introduced. The variation in live fuel flammability for white spruce was more than twice that measured using existing techniques suggesting that the actual changes in live fuel flammability have been underestimated by current fire modelling systems. Measured negative values of flammability for new shoots in the beginning of the season indicated a reduction in the energy release of the combined system of live fuel and frontal flame assumed to result from the high water content of live fuel and oxygen deficiency. Dry matter content and variables characterizing chemical composition of the fuel were replaced by a newly-introduced variable – energy content per unit of fresh mass or volume. Using the gravimetric approach, the energy content did not improve the prediction of flammability; however, when using the volumetric approach, variation in flammability was better explained by energy content than by water content. The proposed volumetric multivariable flammability model (adjusted R^2 = 0.87) was able to better predict flammability compared with the volumetric single-variable models (adjusted R^2 = 0.79). The Canadian Fire Behaviour Prediction System assumes only one seasonal maximum in flammability occurring in early-mid June, but it was instead observed one month earlier. Two additional spikes in flammability occurred in early July and mid-August, when the lowest seasonal values were expected. The mid-August spike in the flammability was caused by a second seasonal minimum in water content induced by drought. If applied to a known amount of live and dead fuel in a vegetative canopy, the proposed approach allows for evaluation of the combined energy release contribution to the flame-front by live and dead fuel on a vegetative canopy scale without modeling fuel consumption. This measure of the forest stand energy release response to fire conditions can further be used in the development of a numerical stand characteristics-based fuel classification and as a forest stand flammability input to fire behaviour modelling systems.
    Subjects: Live fuel flammability, Oxygen consumption calorimtry, Energy content
  6. Condition Based Maintenance Decision Making with Delay Time Modeling [Download]

    Title: Condition Based Maintenance Decision Making with Delay Time Modeling
    Creator: Lipi, Tahmina Ferdousi
    Description: This research targets to develop condition based maintenance models for better maintenance decisions. We address a condition and age based replacement decision problem using the complete history of measured condition observations to minimize long-run average cost or maximize long-run average availability, or both. To estimate the residual lifetime distribution conditional on the history of observed condition information and current age, a delay time model (DTM) based stochastic filtering process (SFP) is used. A long-run average cost model and a long-run average availability model are analyzed in order to develop the theorems necessary for determining the optimum replacement time. A multi-objective decision frontier is proposed that will help maintenance managers deal with trade-offs between the two objectives to minimize the cost and to maximize availability simultaneously. We also proposed models to integrate imperfect maintenance while making replacement decisions. Finally, in order to show the effectiveness of our proposed models, numerical examples are presented.
    Subjects: delay time, condition based maintenance, cost and availability models, net benefit model, decision frontier
  7. Model Based Control of Combustion Timing and Load in HCCI Engines [Download]

    Title: Model Based Control of Combustion Timing and Load in HCCI Engines
    Creator: Ebrahimi, Khashayar
    Description: Different model based control strategies are developed for combustion timing and load control in a single cylinder Homogeneous Charge Compression Ignition (HCCI) engine. In HCCI engines, a lean homogeneous air-fuel mixture auto-ignites due to compression and the resulting combustion occurs at lower temperatures compared to spark ignition or diesel engines. The low HCCI combustion temperatures result in low Nitrogen Oxides (NOX) levels but high unburnt Hydrocarbons (HC) and Carbon Monoxide (CO) levels. High HCCI thermal efficiency occurs when the combustion efficiency is high and the combustion timing is appropriate. In this thesis, the effects of fueling rate and valve timing on HCCI engine performance and energy distribution are described. This analysis indicates that Variable Valve Timing (VVT) with Symmetric Negative Valve Overlap (SNVO) is an effective actuator for combustion timing control. In addition, combustion timing affects combustion efficiency which has an important role in engine energy distribution. Next, a detailed multi-zone model with fuel specific kinetics is developed for HCCI engine performance analysis that captures valve timing and fueling rate dynamics. The multi-zone physics based model has 483 states, 5 inputs and 4 outputs. PI controller gains are first tuned using the detailed multi-zone model in simulation and then the controller is implemented on a single cylinder engine. Combustion timing is used as feedback to the controller and valve timing is the main actuator. Then a Feedforward/Feedback (Fdfwd/Fdbk) strategy is developed for HCCI combustion timing control. The Fdfwd/Fdbk controller is based on a model that relates combustion timing to valve timing and it is combined with an integrator feedback to zero the steady state error. A Model Predictive Control (MPC) strategy is then developed for HCCI combustion timing and load control that takes into account actuator and output constraints. A physics based approach is used for model order reduction of the detailed multi-zone model and a discrete nonlinear control oriented model is obtained with 4 states, 2 inputs and 2 outputs. This model is linearized around one operating point and the MPC is designed based on the linearized version of the 4-state control oriented model. The MPC is then implemented on the single cylinder engine and the results are compared to the PI and Fdfwd/Fdbk controller. The MPC exhibits good tracking performance for combustion timing and load. Finally, a new control oriented model is developed for combustion timing and load control considering combustion efficiency. This model can be used for future MPC design which consider combustion efficiency constraints.
    Subjects: Modeling, Control, HCCI, MPC
  8. Effects of pre-weaning plane of milk replacer and feeding frequency on glucose metabolism in dairy calves [Download]

    Title: Effects of pre-weaning plane of milk replacer and feeding frequency on glucose metabolism in dairy calves
    Creator: MacPherson, Jayden A R
    Description: Feeding dairy calves large milk meal sizes at a low feeding frequency has been associated with reduced insulin sensitivity in previous literature. Therefore, the aim of this thesis was to investigate the influence of feeding an elevated plane of milk on glucose metabolism in calves pre- and post-weaning. To assess insulin sensitivity in calves in chapter 2 and 3, postprandial plasma glucose and insulin concentrations (pre-weaning), as well as insulin response to a glucose tolerance test (GTT; pre and post-weaning) were evaluated. In addition, in chapter 2 the rate of abomasal emptying was characterized pre-weaning to determine the extent of its control over glucose appearance in the blood. Results from chapter 2 where calves were fed a low (4 L/day) or a high (8 L/day) plane of milk twice daily indicated that calves fed either treatment did not experience hyperglycemia or hyperinsulinemia. Additionally, responses to a GTT were similar pre- or post-weaning suggesting both treatments had similar glucose tolerance at all ages. Abomasal emptying was reduced in calves fed a larger meal size (4 L) which indicates it can be used to modulate the appearance of glucose in the blood to prevent hyperglycemia. In chapter 3, feeding an elevated plane of milk (8 L/day) fed over four (4x; meal size 2 L) or two meals per day (2x; meal size 4 L) was compared. Neither treatment resulted in a state of hyperglycemia or hyperinsulinemia, and responses to the GTT were similar indicating similar glucose tolerance. Overall findings from this thesis suggest that calves fed an elevated plane of milk do not experience reduced insulin sensitivity when fed at differing frequencies of 2 or 4 times a day or when compared to calves fed a low plane of milk (4 L/day). These results have significant implications for the dairy industry as this means dairy operations can feed calves more milk, up to 8 L fed over two meals a day, allowing for greater pre-weaning growth without compromising glucose metabolism pre- or post-weaning.
    Subjects: calf nutrition, glucose and insulin, abomasal emptying, plane of nutriton, feeding frequency
  9. From G551D to F508del: An Inquiry Into The Development Of Targeted Therapeutics For The Treatment Of Cystic Fibrosis [Download]

    Title: From G551D to F508del: An Inquiry Into The Development Of Targeted Therapeutics For The Treatment Of Cystic Fibrosis
    Creator: Mohammad, Sohaib
    Description: Cystic Fibrosis (CF) is a fatal inherited disease caused by mutations in the gene encoding the Cystic Fibrosis Transmembrane Regulator (CFTR) protein. CFTR plays an integral role in salt and water transport across the epithelial membrane of major organs, such as the lungs. CFTR-targeted therapeutic strategies can theoretically reduce the effects of CFTR ion dysfunction through potentiation, correction, or both. Potentiators work by increasing the length of time CFTR channels remain open following activation while correctors work by increasing the cell surface density of CFTR. In 2012, regulatory approval by the United Stated Food and Drug Administration (FDA) and European Medicines Agency (EMA) was granted to a potentiator compound, Ivacaftor (trade named Kalydeco). In 2015, the FDA and EMA granted regulatory approval to a corrector-potentiator combination, Lumacaftor-Ivacaftor (trade named Orkambi). The regulatory approval of these compounds has been met with both excitement and concern. For the first time since the discovery of CFTR gene in 1989, an agent which works by directly targeting the CFTR channel has been developed, and this in turn has paved the way towards a potential cure. On the other hand, Kalydeco and Orkambi are not curative, and the amount of clinical benefit seen in clinical trials ranges from, at best, an absolute improvement of 12.5% (Kalydeco) and 3.6% (Orkambi) from baseline for one measure of lung function, Percent Predicted Forced Expiratory Volume in 1 Second (FEV1% predicted). Given that they need to be taken throughout a patient’s lifetime, there has been some concern regarding the cost-effectiveness of these treatments, which range from $259,000 USD (Orkambi) to over $300,000 USD (Kalydeco) per patient per year This thesis research involves three investigations pertaining to the development of Kalydeco and Orkambi for the treatment of CF: 1) A review of Phase II/III clinical trials which have lead to the regulatory approval of Kalydeco and Orkambi; 2) An evaluation of clinical trials that have studied Ivacaftor, Lumacaftor, or their combination in patients homozygous for the F508del mutation, which affects nearly half of all CF patients; and 3) A study of gaps that existed in CF treatment when Kalydeco first received regulatory approval and which gaps remain since Orkambi’s regulatory approval. Our first study is important to understand the historical development of Kalydeco and Orkambi, and in particular, to gain a better understanding of the underlying biology of CFTR as well as the clinical endpoints used in clinical trials. The results of our second investigation suggest that, although Lumacaftor-Ivacaftor combination therapy appears to be superior to Lumacaftor monotherapy, studies of longer duration that are adequately powered towards key clinical endpoints, like FEV1% predicted, are needed to distinguish Lumacaftor-Ivacaftor as being superior to Ivacaftor monotherapy. The results of our third study indicate that several gaps in CF treatment through the use of these targeted agents have been fully or partially filled, but there are certain key gaps which remain. In particular, there is still uncertainty pertaining to the clinical benefit of Ivacaftor in certain sub-populations eligible to receive the treatment, the cost effectiveness of Ivacaftor, as well as the usefulness of sweat chloride concentration as a clinical endpoint in clinical trials.
    Subjects: Cystic Fibrosis, Kalydeco, Orkambi
  10. The Discovery and Application of Bacteriophage Receptor Binding Proteins [Download]

    Title: The Discovery and Application of Bacteriophage Receptor Binding Proteins
    Creator: Simpson, David James
    Description: Bacteriophages are considered to be the most abundant and potentially the most diverse form of life on earth. Phage receptor binding proteins (RBPs), which allow phages to specifically target their host bacteria, consequently represent a massive diversity of bacterial targeting proteins. These RBPs can bind to bacteria with strong affinity and show considerable resistance to proteases and detergents. In recent years, a number of new technologies have been developed for pathogen detection through the attachment of RBPs to surfaces or beads. This thesis describes new techniques that take advantage of these proteins that act as surrogate antibodies. Recent years have seen a resurgence in bacteriophage research, often in the form of phage therapy, due to the recent rise in antibiotic resistance. However, bacteriophage RBPs remain difficult to identify based on homology alone due to their considerable diversity. While these proteins share a trimeric structure, they can be very dissimilar on the sequence level. In order to further exploit the use of RBPs, I have developed an assay for discovering RBPs using phage genome expression libraries and protein screens to identify binding partners that recognize the host bacterium. Briefly, the phage DNA is sheared and ligated into an expression library, this library is transferred to a nitrocellulose membrane where the colonies are induced to express the inserts, the cells are then lysed and the membrane is probed with the host bacteria. When the Salmonella enterica serovar Typhimurium phage P22 was screened using this assay, Gp9 was the only RBP discovered, confirming previous predictions that this is the sole RBP encoded by this phage. I then examined the Escherichia coli O157:H7 typing phage 1 using this assay and identified a previously undescribed RBP, Gp145. This general approach has the potential to assist in the identification of RBPs from other relevant bacteriophages. In previous studies the bacteriophage P22 RBP has also been shown to be able to reduce colonization of S. Typhimurium in chickens. In order to exploit this finding further, the protein was expressed in plants with the aim of creating an inexpensively produced selective antimicrobial feed. An elastin like polypeptide (ELP) tag was added to the P22 RBP to increase expression of the protein in Nicotiana benthamiana. This thesis demonstrates that the RBP containing plant extract was capable of capturing S. Typhimurium on a nitrocellulose membrane, and moderately reducing the ability of S. Typhimurium to colonize chickens. RBPs bind bacteria with high affinity, to make use of this trait in a diagnostics-based platform, the cellulose binding module CBM9, that enables proteins to bind to paper, was added to Gp9 and Gp145. The N-terminally tagged Gp9 and Gp145 constructs were spotted on paper and are able to capture S. Typhimurium and E. coli O157:H7, respectively on paper. Gp145 was further characterized and shown to bind to the lipopolysaccharide of E. coli O157:H7 and interestingly also binds to S. Typhimurium, likely through a protein receptor. Taken together, these results demonstrate that RBPs represent an exciting new technology for microbial detection and treatment which are inexpensive, easy to use and readily scalable.
    Subjects: Bacteriophage, Receptor Binding Protein, Salmonella