Gene therapy is advancing rapidly, with Recombinant Adeno-associated virus (rAAV) being investigated for potential use in treating cancer and neurological disorders. Plasmid DNA transfection and viral infection are standard methods for producing large-scale rAAV vectors. However, improving yield production requires careful monitoring and control of process state variables, which can be expensive and time-consuming. This thesis proposes a model predictive control (MPC) model that can efficiently monitor, predict, and optimize the final product by controlling state variables like DOT and temperature. The model relies on an unstructured mechanistic kinetic model designed explicitly based on rAAV upstream production. Monitoring viral vector production based on substrate or biomass concentration enhances bioprocess production efficiency. However, other state variables like dissolved oxygen (DO), pH, and temperature should also be considered. The objective of this thesis is to enhance cell growth in bioreactors by regulating dissolved oxygen and temperature levels using a Model Predictive Control (MPC) system. This model can be employed in different processes to enhance cell growth and examine the impact of control measures. The goal is to achieve a high cell density, increase productivity, and lower costs in a shorter duration. Simulink, a software tool developed by MATLAB, seamlessly integrates Ordinary Differential Equations (ODEs) to optimize bioprocesses in bioreactors. The Model Predictive Control (MPC) controller expertly regulates Dissolved Oxygen Tension (DOT) and temperature, thereby increasing cell growth concentrations. This sophisticated controller efficiently manages multiple variables simultaneously and exceeds the Proportional Integral Derivative (PID) controller. The model is straightforward to comprehend and promptly responds to anomaly data. To evaluate the suggested resolution, we conducted tests on both PID and MPC controllers by introducing measurement noise to the DOT. Our analysis indicated that MPC demonstrated superior performance based on the ISE (Integral of Squared Error), IAE (Integral of Absolute Error), and ITAE (Integral of Time-weighted Absolute Error), all of which were substantially higher for the PID controller. Regardless of changing conditions, MPC adeptly tracks the setpoint and optimizes the variable to enhance production efficiency.

In Canada alone, there were an estimated 800,000 visually impaired people in 2007, costing the federal government an annual amount of $15.8 billion in services, treatments and lost revenue. These costs are estimated to double by the year 2032, as the population ages. The leading causes of visual impairment and blindness is retinal degeneration, characterized by the progressive death of retinal cells. The research presented in this PhD thesis aimed to prevent retinal degeneration by over-expressing the X-linked Inhibitor of Apoptosis (XIAP) in retinal cells using plasmid and adeno-associated viral vectors. The work is divided into four sequential chapters targeted at developing an anti-apoptotic gene therapy strategy to prevent retinal cell death. The first chapter examines XIAP gene therapy in the treatment of Leber’s Hereditary Optic Neuropathy (LHON). In vitro studies using the 661W cone-photoreceptor cell line showed that XIAP over-expression significantly lowers cell death when 661W cells are exposed to a number of apoptotic stimuli. In a mouse model of Leber’s Hereditary Optic Neuropathy (LHON), XIAP expression in retinal ganglion cells (RGCs) protected the ultrastructure of the RGC axons within the optic nerve, in addition to providing evidence of functional protection. The second and third chapters further examine the potential for XIAP gene therapy in the treatment of retinal disease by developing an in vivo model of retinal detachment in cats, followed by evaluating the efficacy of XIAP gene therapy intervention. When XIAP was over-expressed in the photoreceptor cells, there was significant structural protection and trends in preservation of function in this model of degeneration. Finally, the fourth chapter explores an alternate method to viral gene therapy by evaluating the efficacy and toxicity of chitosan microparticles as a protein delivery system to the retina. Results show that chitosan microparticles are mucosal-adhesive and are non-toxic at low concentrations in vitro in 661W cells and in vivo in rats. This thesis work provides strong evidence that XIAP gene therapy is an effective method for preventing retinal degeneration, and works as a broad spectrum gene therapy strategy that can be applied to different forms of retinal degeneration.

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