Irreversible electroporation is a minimally invasive technique for the non-thermal destruction of cells in a targeted volume of tissue, using brief electric pulses, (~100 µs long) delivered through electrodes placed into or around the targeted region. These electric pulses destabilize the integrity of the cell membrane, resulting in the creation of nanoscale defects that increase a cell’s permeability to exchange with its environment. When the energy of the pulses is high enough, the cell cannot recover from these effects and dies in a non-thermal manner that does not damage neighboring structures, including the extracellular matrix. IRE has been shown to spare the major vasculature, myelin sheaths, and other supporting tissues, permitting its use in proximity to these vital structures. This technique has been proposed to be harnessed as an advantageous non-thermal focal ablation technique for diseased tissues, including tumors. IRE electric pulses may be delivered through small (ø ≈ 1 mm) needle electrodes, making treatments minimally invasive and easy to apply. There is sub-millimeter demarcation between treated and unaffected cells, which may be correlated with the electric field to which the tissue is exposed, enabling numerical predictions to facilitate treatment planning. Immediate changes in the cellular and tissue structure allow real-time monitoring of affected volumes with imaging techniques such as computed tomography, magnetic resonance imaging, electrical impedance tomography, or ultrasound. The ability to kill tumor cells has been shown to be independent of a functioning immune system, though an immune response seems to be promoted by the ablation. Treatments are unaltered by blood flow and the electric pulses may be administered quickly (~ 5 min). Recently, safety and case studies using IRE for tumor therapy in animal and human patients have shown promising results. Apart from these new studies, previous work with IRE has involved studies in healthy tissues and small cutaneous experimental tumors. As a result, there remain significant differences that must be considered when translating this ablation technique towards a successful and reliable therapeutic option for patients. The dissertation work presented here is designed to develop irreversible electroporation into a robust, clinically viable treatment modality for targeted regions of diseased tissue, with an emphasis on tumors. This includes examining and creating proving the efficacy for IRE therapy when presented with the many complexities that present themselves in real-world clinical patient therapies, including heterogeneous environments, large and irregular tumor geometries, and dynamic tissue properties resulting from treatment. The impact of these factors were theoretically tested using preliminary in vitro work and numerical modeling to determine the feasibility of IRE therapy in heterogeneous systems. The feasibility of use was validated in vivo with the successful treatment of human mammary carcinomas orthotopically implanted in the mammary fat pad of mice using a simple, single needle electrode design easily translatable to clinical environments. Following preliminary theoretical and experimental work, this dissertation considers the most effective and accurate treatment planning strategies for developing optimal therapeutic outcomes. It also experimentally characterizes the dynamic changes in tissue properties that result from the effects of IRE therapy using ex vivo porcine renal cortical tissue and incorporates these into a revised treatment planning model. The ability to use the developments from this earlier work is empirically tested in the treatment of a large sarcoma in a canine patient that was surgically unresectable due to its proximity to critical arteries and the sciatic nerve. The tumor was a large and irregular shape, located in a heterogeneous environment. Treatment planning was performed and the therapy carried out, ultimately resulting in the patient being in complete remission for 14 months at the time of composing this work. The work presented in this dissertation finishes by examining potential supplements to enhance IRE therapy, including the presence of an inherent tumor-specific patient immune response and the addition of adjuvant therapeutic modalities. / Ph. D.