Since its conception in 2005, irreversible electroporation (IRE), a non-thermal tumor ablation modality, was investigated for safety and efficacy in clinical applications concerning different organs. IRE utilizes high voltage (~3kV), short duration (~100us) pulses to create transient nanoscale defects in the plasma membrane to cause cell death due to irreversible defects, osmotic imbalances and ATP loss. More recently, high-frequency irreversible electroporation (H-FIRE), which employs narrow bipolar pulses (~0.5-10us) delivered in bursts (on time ~100us), was invented to provide benefits such as the mitigation of intense muscle contractions associated with IRE-based treatments. Furthermore, H-FIRE exhibits the potential to improve lesion predictability in homogeneous and heterogeneous tissue masses. Therapeutic IRE and H-FIRE utilize source and sink electrodes inserted into or around the tumor to deliver the treatment. Prediction of the ablation size, for a set of parameters, can be achieved by the use of pre-treatment planning algorithms that calculate the induced electric field distribution in the target tissue. An electric field above a certain threshold induces cell death and parameters are tuned to ensure complete tumor coverage while sparing the nearby healthy tissue. IRE studies have shown that the underlying field is influenced by the increase in tissue conductivity due to enhanced membrane permeability, and treatment outcome can be improved when this nonlinearity is accounted for in numerical models. Since IRE pulses far exceed the time constant of the cell (~1us), the tissue response can be treated as essentially DC a static approximation can be used to predict the field distribution. Alternately, as H-FIRE pulses are on the order of the time constant of the membrane, the tissue response can no longer be treated as DC. The complexity of the H-FIRE-induced field distribution is further enhanced due to the dispersion and non-linearity in biological tissue impedance during treatment. In this dissertation, we have studied the electromagnetic fields induced in tissue during H-FIRE using several experimental and modeling techniques. In addition, we have characterized the nonlinearity and dispersion in tissue impedance during H-FIRE treatments and proposed simpler methods to predict the field distribution to enable easier translation to the clinic. / Ph. D.

Remote measurement of electric field generated by lightning has played a major role in understanding the lightning phenomenon. Even though other measurements such as photographic and channel base current have contributed to this research field, due to practical reasons remote measurements of electric field is considered as the most useful tool in lightning research. This thesis discusses the remotely measured radiation field component of electric field generated by cloud flashes (ICs) and narrow bipolar pulses (NBPs). The associated HF radiation of these events at 3 MHz and 30 MHz are also discussed. To understand the initiation process of these discharges, a comparative study of the initial pulse of cloud flashes against the initial pulse of cloud to ground flashes was conducted. The result suggests that both discharges might have been initiated by similar physical processes inside the thunderclouds. Comparing the features of initial pulse of cloud and ground flashes with that of pulses that appeared in the later stages of cloud flashes suggests that the initiation process involved in both flashes are not very much different from the initiation of cloud flashes at the later stage. The average spectral amplitudes of electric field of full duration cloud flashes (180 ms) showed f  -1 frequency dependence within the interval of 10 kHz to approximately 10 MHz. This is in contrast to the standard f  -2 decrement (or even steeper ) at high frequency region for other lightning processes such as return strokes. It was suggested that small pulses which repeatedly appeared at the later stage of cloud flashes might have contributed to enhance the spectral amplitude at higher frequencies. Electric fields generated by Narrow Bipolar Pulses (NBPs), which are considered as one of the strongest sources of HF radiation, were measured in the tropics of Malaysia and Sri Lanka.  Their features were also studied and show a good agreement with previously published observations of NBPs from other geographical regions. Thorough analyses and observations of these pulses found previously unreported sharp, fine peaks embedded in the rising and decaying edge of the electric field change of NBPs. Therefore it was suggested that these fine peaks are mostly responsible for the intense HF radiation at 30 MHz.

Glioblastoma multiforme (GBM) is the most common and lethal primary brain cancer with an average survival time of 15 months. GBM is considered incurable with even the most aggressive multimodal therapies and is characterized by near universal recurrence. Irreversible electroporation (IRE) is a cellular ablation method currently being investigated as a therapy for a variety of cancers. Application of IRE involves insertion of electrodes into tissue to deliver pulsed electric fields (PEFs), which destabilize the cell membrane past the point of recovery, thereby inducing cell death. While this treatment modality has numerous advantages, the lack of selectivity for malignant cells limits its application in the brain where damage to healthy tissue is especially deleterious. In this dissertation we hypothesize that a form of IRE therapy, high-frequency IRE (H-FIRE), may be able to act as a selective targeted therapy for GBM due to its ability to create an electric field inside a cell to interact with altered inner organelles. Through a comprehensive investigation involving experimental testing combined with numerical modeling, we have attained results in strong support of this hypothesis. Using tissue engineered hydrogels as our platform for therapy testing, we demonstrate selective ablation of GBM cells. We develop mathematical models that predict the majority of the electric field produced by H-FIRE pulses reach the inside of the cell. We demonstrate that the increased nuclear to cytoplasm ratio (NCR) of malignant GBM cells compared to healthy brain—evidenced in vivo and in in vitro tissue mimics—is correlated with greater ablation volumes and thus lower electric field thresholds for cell death when treated with H-FIRE. We enhance the selectivity achieved with H-FIRE using a molecularly targeted drug that induces an increase in NCR. We tune the treatment pulse parameters to increase selective malignant cell killing. Finally, we demonstrate the ability of H-FIRE to ablate therapy-resistant GBM cells which are a focus of many next-generation GBM therapies. We believe the evidence presented in this dissertation represents the beginning stages in the development of H-FIRE as a selective therapy to be used for treatment of human brain cancer. / Ph. D.

Irreversible electroporation has recently emerged as an effective focal ablation technique. When performed clinically, the procedure involves placing electrodes into, or around, a target tissue and applying a series of short, but intense, pulsed electric fields. Oftentimes, patient specific treatment plans are employed to guide procedures by merging medical imaging with algorithms for determining the electric field distribution in the tissue. The electric field dictates treatment outcomes by increasing a cell's transmembrane potential to levels where it becomes energetically favorable for the membrane to shift to a state of enhanced permeability. If the membrane remains permeabilized long enough to disrupt homeostasis, cells eventually die. By utilizing this phenomenon, irreversible electroporation has had success in killing cancer cells and treating localized tumors. Additionally, if the pulse parameters are chosen to limit Joule heating, irreversible electroporation can be performed safely on surgically inoperable tumors located next to major blood vessels and nerves. As with all technologies, there is room for improvement. One drawback associated with therapeutic irreversible electroporation is that patients must be temporarily paralyzed and maintained under general anesthesia to prevent intense muscle contractions occurring in response to pulsing. The muscle contractions may be painful and can dislodge the electrodes. To overcome this limitation, we have developed a system capable of achieving non-thermal irreversible electroporation without causing muscle contractions. This progress is the main focus of this dissertation. We describe the theoretical basis for how this new system utilizes alterations in pulse polarity and duration to induce electroporation with little associated excitation of muscle and nerves. Additionally, the system is shown to have the theoretical potential to improve lesion predictability, especially in regions containing multiple tissue types. We perform experiments on three-dimensional in vitro tumor constructs and in vivo on healthy rat brain tissue and implanted tumors in mice. The tumor constructs offer a new way to rapidly characterize the cellular response and optimize pulse parameters, and the tests conducted on live tissue confirm the ability of this new ablation system to be used without general anesthesia and a neuromuscular blockade. Situations can arise in which it is challenging to design an electroporation protocol that simultaneously covers the targeted tissue with a sufficient electric field and avoids unwanted thermal effects. For instance, thermal damage can occur unintentionally if the applied voltage or number of pulses are raised to ablate a large volume in a single treatment. Additionally, the new system for inducing ablation without muscle contractions actually requires an elevated electric field. To ensure that these procedures can continue to be performed safely next to major blood vessels and nerves, we have developed new electrode devices that absorb heat out of the tissue during treatment. These devices incorporate phase change materials that, in the past, have been reserved for industrial applications. We describe an experimentally validated numerical model of tissue electroporation with phase change electrodes that illustrates their ability to reduce the probability for thermal damage. Additionally, a parametric study is conducted on various electrode properties to narrow in on the ideal design. / Ph. D.

This work investigates the use of irreversible electroporation (IRE) for tissue engineering applications and as a cancer ablation therapy. IRE uses short, high-intensity electric pulses to create pores in a cell's membrane and disrupt its stability. At a certain energy level, damage to the cell becomes too great and it leads to cell death. The particular mechanisms that drive this response are still not completely understood. Thus, further characterization of this behavior for cell death induced by pulsed electric fields (PEFs) will advance the understanding of these types of therapies and encourage their use to treat unresectable tumors that can benefit from the non-thermal mechanism of action which spares critical blood vessels and nerves in the surrounding area. We evaluate the response to PEFs by different cell types through experimental testing combined with computer simulations of these treatments. We show that IRE can be used to kill a specific type of bacteria that produce cellulose which can be used as an implantable material to repair damaged tissues. By killing these bacteria at particular times and locations during their cellulose production, we can create conduits in the overall structure of this material for the transport of oxygen and nutrients to the cells within the area after implantation. The use of tissue models also plays a key role in the investigation of various cancer treatments by providing a controlled environment which can mimic the state of cells within a tumor. We use tumor models comprised of a mix of collagen and cancer cells to evaluate their response to IRE based on the parameters that induce cell death and the time it takes for this process to occur. The treatment of prostate and pancreatic cancer cells with standard monopolar (only positive polarity) IRE pulses resulted in different time points for a full lesion (area of cell death) to develop for each cell type. These results indicate the presence of secondary processes within a cell that induce further cell death in the border of the lesion and cause the lesion to increase in size several hours after treatment. The use of high-frequency irreversible electroporation (H-FIRE)--comprised of short bursts of high-intensity, bipolar (both positive and negative polarity) pulses--can selectively treat cancer cells while keeping healthy cells in the neighboring areas alive. We show that H-FIRE pulses can target tumor-initiating cells (TICs) and late-stage, malignant cancer cells over non-malignant cells using a mouse ovarian cancer model representative of different stages of disease progression. To further explore the mechanisms that drive this difference in response to IRE and H-FIRE, we used more complex tumor models. Spheroids are a type of 3D cell culture model characterized by the aggregation of one or more types of cells within a single compact structure; when embedded in collagen gels, these provide cell-to-cell contact and cell-to-matrix adhesion by interactions of cells with the collagen fibers (closely mimicking the tumor microenvironment). The parameters for successful ablation with IRE and H-FIRE can be further optimized with the use of these models and the underlying mechanisms driving the response to PEFs at the cellular level can be revealed. / Ph. D. / This work investigates the use of irreversible electroporation (IRE) for tissue engineering applications and as a cancer ablation therapy. IRE uses short, high-intensity electric pulses to create pores in a cell’s membrane and disrupt its stability. At a certain energy level, damage to the cell becomes too great and it leads to cell death. The particular mechanisms that drive this response are still not completely understood. Thus, further characterization of this behavior for cell death induced by pulsed electric fields (PEFs) will advance the understanding of these types of therapies and encourage their use to treat unresectable tumors that can benefit from the non-thermal mechanism of action which spares critical blood vessels and nerves in the surrounding area. We evaluate the response to PEFs by different cell types through experimental testing combined with computer simulations of these treatments. We show that IRE can be used to kill a specific type of bacteria that produce cellulose which can be used as an implantable material to repair damaged tissues. By killing these bacteria at particular times and locations during their cellulose production, we can create conduits in the overall structure of this material for the transport of oxygen and nutrients to the cells within the area after implantation. The use of tissue models also plays a key role in the investigation of various cancer treatments by providing a controlled environment which can mimic the state of cells within a tumor. We use tumor models comprised of a mix of collagen and cancer cells to evaluate their response to IRE based on the parameters that induce cell death and the time it takes for this process to occur. The treatment of prostate and pancreatic cancer cells with standard monopolar (only positive polarity) IRE pulses resulted in different time points for a full lesion (area of cell death) to develop for each cell type. These results indicate the presence of secondary processes within a cell that induce further cell death in the border of the lesion and cause the lesion to increase in size several hours after treatment. The use of high-frequency irreversible electroporation (H-FIRE)—comprised of short bursts of high-intensity, bipolar (both positive and negative polarity) pulses—can selectively treat cancer cells while keeping healthy cells in the neighboring areas alive. We show that H-FIRE pulses can target tumor-initiating cells (TICs) and late-stage, malignant cancer cells over non-malignant cells using a mouse ovarian cancer model representative of different stages of disease progression. To further explore the mechanisms that drive this difference in response to IRE and H-FIRE, we used more complex tumor models. Spheroids are a type of 3D cell culture model characterized by the aggregation of one or more types of cells within a single compact structure; when embedded in collagen gels, these provide cell-to-cell contact and cell-to-matrix adhesion by interactions of cells with the collagen fibers (closely mimicking the tumor microenvironment). The parameters for successful ablation with IRE and H-FIRE can be further optimized with the use of these models and the underlying mechanisms driving the response to PEFs at the cellular level can be revealed.