The tumour suppressor p53-binding protein 1 (53BP1) is phosphorylated following DNA double strand breaks (DSBs); however, little is understood about the upstream signaling pathways that control this phosphorylation. Additionally, it is not known how these processes combine with 53BP1 to control the survival of cells following DNA damage such as that imparted by ionizing radiation (IR), which is the basis of radiotherapy. In this thesis, I have shown that 53BP1 is phosphorylated specifically in S-phase cells, but not relocalized to intranuclear foci, in response to severe oxygen stress. This occurs with only partial dependence on the ATM kinase (Chapter 2). Following IR, I find that both ATM and DNA-PKcs contribute to intranuclear phosphorylated 53BP1 foci, but that this phosphorylation is independent of proximal signaling molecules that control the localization of 53BP1 to initial DSBs (Chapter 3). Furthermore, I show that 53BP1 loss confers sensitivity to IR and this can be further augmented by inhibition of ATM and DNA-PKcs kinases suggesting that there are both 53BP1-dependent and -independent pathways of survival from IR (Chapter 4). These findings may have important implications for molecular pathology and personalized medicine as 53BP1 has recently been found to be activated or lost in subsets of human tumours. I have collaborated to initiate the development of a novel system to interrogate the implications of 53BP1 loss as traditional siRNA approaches in human cancer cells were not feasible (Chapter 5 and Appendix 2). This system can be used in vivo as tumour xenografts to further understand how 53BP1 and the tumour microenvironment interact endogenously and in response to IR. I also present the possibility and proof of concept for the use of 53BP1 as a biomarker in primary human prostate cancer tissue where little is known about 53BP1 biology (Chapter 5).

Bilateral implantation of a cochlear implant (CI) after a >2 year period of unilateral hearing with a second implant has been shown to result in altered latencies in brainstem responses in children with congenital deafness. In this thesis, a neural source localization method was developed to investigate the effects of unilateral CI use on cortical development after the implantation of a 2nd CI. The electroencephalography (EEG) source localization method is based on the linearly constrained minimum variance (LCMV) vector beamformer and utilizes null constraints to minimize the electrical artifact produced by the CI. The accuracy of the method was assessed and optimized through simulations and comparisons to beamforming with magnetoencephalography (MEG) data. After using cluster analyses to ensure that sources compared across subjects originate from the same neural generators, a study was done to examine the effects of unilateral CI hearing on hemispheric lateralization to monaural responses. It was found that a >2 year period of unilateral hearing results in expanded projections from the 1st implanted ear to the contralateral auditory area that is not reversed by implantation of a 2nd CI. A subsequent study was performed to examine the effects of unilateral CI hearing on the contributions of the 1st and 2nd implanted ears to the binaural response. It was found that in children with > 2 years of unilateral hearing, the binaural response is dominated by the 1st implanted ear. Together, these results suggest that the delay between the 1st and 2nd CI should be minimized in bilateral implantation to avoid dominance of auditory pathways from the 1st implanted ear. This dominance limits developmental competition from the 2nd CI and potentially contributes to poorer performance in speech detection in noise tasks.

Microbubble contrast agents for ultrasound are widely used in numerous medical applications, both diagnostic and therapeutic. Due to their size, similar to that of red blood cells, microbubbles are able to traverse the entire vascular bed, enabling their utilization for applications such as tumour diagnosis. Vaporizable submicron droplets of liquid perfluoro- carbon potentially represent a new generation of extravascular contrast agents for ultrasound. Droplets of a few hundred nanometers in diameter have the ability to extravasate selectively in regions of tumour growth while staying intravascular in healthy tissues. Upon extravasation, these droplets may be vaporized with ultrasound and converted into gas bubbles. In this thesis we argue that vaporized submicron perfluorocarbon droplets possess the necessary stability and acoustic characteristics to be potentially applicable as a new gener- ation of extravascular ultrasound contrast agents. We examine, separately, the ultrasound conditions necessary for vaporization of the droplets into microbubbles, the size and stability of these bubbles following vaporization, on timescales ranging from nanoseconds to minutes, and the bubbles’ acoustic response to incident diagnostic ultrasound. We show that submicron droplets may be vaporized into bubbles of a few microns in diameter using single ultrasound pulse within the diagnostic range. The efficiency of conversion is shown to be on the order of at least 10% of the exposed droplets converting into stable microbubbles. The bubbles are shown to be stabilized by the original coating material encapsulating the droplet precursors, and be stable for at least minutes following vaporization. Finally, vaporized droplets are shown to be echogenic, with acoustic characteristics comparable to these of the commercially available ultrasound contrast agents. The results presented here show that vaporized droplets possess the necessary stability properties and echogenicity required for successful application as contrast agents, suggesting potential for their future translation into clinical practice.

Microbubble ultrasound (US) contrast agents have been used to monitor the progression of anti-angiogenic chemotherapies. However, US backscatter measurements used in contrast imaging are inherently variable, given the presence of many microbubbles of random position and size. A model was developed to investigate the influence of US scanner and microbubble characteristics on these variable measurements. The Coefficient of Variation was used to measure variability. It was found that an optimum excitation frequency exists that minimizes this variability. In the case of DefinityTM, a 2.25 MHz centre-frequency pulse yielded a less variable measurement than at 5 MHz. Conversely, decreasing microbubbble concentration was found to significantly increase variability. Evidence suggests that microbubbles are no longer Rayleigh scatterers at sufficient low concentrations. Post-processing was found to aid in reducing measurement variability by averaging samples where microbubble positions are uncorrelated. As well, reduction can be achieved by averaging about a region-of-interest of uniform perfusion.

The use of complementary non-invasive imaging modalities has been proposed to track disease progression, particularly cancer, while simultaneously evaluating therapeutic efficacy. A major obstacle is a limited ability to compare parameters obtained from different modalities, especially those from exogenous contrast agents or tracers. We hypothesize that combining Magnetic Resonance Imaging (MRI) and Ultrasound (US) will improve characterization of the tumour microenvironment. In this study, we describe a co-registration apparatus that facilitates the acquisition of a priori co-registered MR and US images in vivo. This apparatus was validated using phantom data and it was found that the US slices can be selected to an accuracy of +/- 100µm translationally and +/- 2 degrees rotationally. Additionally, it was shown that MRI and US may provide complimentary information about the tumour microenvironment, but more work needs to be done to assess repeatability of dynamic contrast enhanced MRI and US.

Obesity is an adverse factor in the development and severity of breast cancer. Obesity is often accompanied by an increase in circulating insulin, which is also associated with poor BC prognosis. Although not expressed in normal breast tissue, the insulin receptor (IR) is highly expressed in BC, therefore insulin signaling in BC cells may be responsible for the negative prognostic effects associated with hyperinsulinemia. This thesis describes the development of a cell-based system to study how insulin affects BC. My work shows that MCF10A, untransformed human breast epithelial cells that express the IR, require insulin for normal proliferation and morphology. Interestingly, I discovered hyperactivation of ERK1/2 in MCF10A cells in response to insulin withdrawal, resulting in a loss of epithelial phenotype. Unexpectedly, while losing epithelial phenotype, MCF10A cells depleted of insulin failed to migrate. In conclusion, breast cells that express IR require insulin for migration and maintenance of epithelial characteristics.

Monitoring treatment efficacy is a large area of cancer research as it can increase the effectiveness of therapy regimens. Diffusion weighted Magnetic Resonance imaging (DWI), allows assessment of tissue microstructure without exogenous contrast agents. In this thesis, two different DWI techniques were used to acquire data from acute myeloid leukemia cells undergoing apoptosis, and data was fitted to an analytical model of re- stricted diffusion. Results indicated a decrease in average restriction size from 6.4 to 2.7μm, and an increase in the restricted diffusion coefficient from 0.17 to 0.82μm^2/ms in untreated versus treated cells. The free diffusion coefficient was constant indicating changes in restrictions, rather than any intrinsic changes in the intra-cellular or extra- cellular fluid. This combination of techniques has the potential for use in preclinical and clinical settings as it demonstrates that apoptotic changes may be measured consistently.

A number of interventional procedures such as cardiac catheterization, angiography and the deployment of endovascular devices are routinely performed using x-ray fluoroscopy. To minimize the patient’s exposure to ionizing radiation, each fluoroscopic image is acquired using a very low x-ray exposure (~ 1 uR at the detector). At such an exposure, most semiconductor-based digital flat panel detectors (FPD) are not x-ray quantum noise limited (QNL) due to the presence of electronic noise which substantially degrades their imaging performance. The goal of this thesis was to investigate how a FPD based on amorphous selenium (a-Se) with internal avalanche multiplication gain could be used for QNL fluoroscopic imaging at the lowest clinical exposures while satisfying all of the requirements of a FPD for interventional radiology. Towards this end, it was first determined whether a-Se can reliably provide avalanche multiplication gain in the solid-state. An experimental method was developed which enabled the application of sufficiently large electric field strengths across the a-Se. This method resulted in avalanche gains as high as 10000 at an applied field of 105 V/um using optical excitation. This was the first time such high avalanche gains have been reported in a solid-state detector based on an amorphous material. Secondly, it was investigated how the solid-state a-Se avalanche detector could be used to image X-rays at diagnostic radiographic energies (~ 75 kVp). A dual-layered direct-conversion FPD architecture was proposed. It consisted of an x-ray drift region and a charge avalanche multiplication region and was found to eliminate depth-dependent gain fluctuation noise. It was shown that electric field strength non-uniformities in the a-Se do not degrade the detective quantum efficiency (DQE). Lastly, it was determined whether the solid-state a-Se avalanche detector satisfies all of the requirements of interventional radiology. Experimental results have shown that the total noise produced by the detector is negligible and that QNL operation at the lowest fluoroscopic exposures is indeed possible without any adverse effects occurring at much larger radiographic exposures. In conclusion, no fundamental obstacles were found preventing the use of avalanche a-Se in next-generation solid-state QNL FPDs for use in interventional radiology.

Dose management in CT is an increasingly important issue as the number of CT scans per capita continues to rise. One proposed approach for enhanced dose management is to allow the spatial pattern of x-ray fluence delivered to the patient to change dynamically as the x-ray tube rotates about the patient. The changes in incident fluence could be guided using a patient model and optimization method in order to deliver user-defined image quality criteria while minimizing dose. This approach is referred to as fluence field modulated CT (FFMCT). In this work, a framework and optimization method was developed for evaluating the dose and image quality benefits of FFMCT, both in simulated and experimental data. Modulated fluence profiles were optimized for different objects and image quality criteria using a simulated annealing algorithm. Analysis involved comparing predicted image quality maps and dose outcomes to those using conventional methods. Results indicated that image quality distributions using FFMCT agreed better with prescribed image qualities than conventional techniques allow. Dose reductions ranged depending on the task and object of interest. Simulation studies using a simulated anthropomorphic phantom of the chest suggest an average dose reduction of at least 20% compared to conventional techniques is possible, where local dose reductions may be greater than 60%. Across different imaging tasks and objects, integral dose reductions ranged from 20-50% when compared to a conventional bowtie filter. The results of this study suggest that given a suitable collimator approach, FFMCT could reap significant benefits in terms of reducing dose and optimizing image quality. Though the tradeoff between image quality and imaging dose may not be eliminated, it may be better managed using an FFMCT approach.

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