Background: Despite promising treatments for breast cancer, mortality rates remain high and treatments for metastatic disease are limited. High-frequency irreversible electroporation (H-FIRE) is a novel tumor ablation technique that utilizes high-frequency bipolar electric pulses to destabilize cancer cell membranes and induce cell death. However, there is currently a paucity of data pertaining to immune system activation following H-FIRE and other electroporation based tumor ablation techniques. Methods: Here, we utilized the mouse 4T1 mammary tumor model to evaluate H-FIRE treatment parameters on cancer progression and immune system activation in vitro and in vivo. Findings: H-FIRE effectively ablates the primary tumor and induces a pro-inflammatory shift in the tumor microenvironment. We further show that local treatment with H-FIRE significantly reduces 4T1 metastases. H-FIRE kills 4T1 cells through non-thermal mechanisms associated with necrosis and pyroptosis resulting in damage associated molecular pattern signaling in vitro and in vivo. Our data indicate that the level of tumor ablation correlates with increased activation of cellular immunity. Likewise, we show that the decrease in metastatic lesions is dependent on the intact immune system and H-FIRE generates 4T1 neoantigens that engage the adaptive immune system to significantly attenuate tumor progression. Interpretation: Cell death and tumor ablation following H-FIRE treatment activates the local innate immune system, which shifts the tumor microenvironment from an anti-inflammatory state to a pro-inflammatory state. The non-thermal damage to the cancer cells and increased innate immune system stimulation improves antigen presentation, resulting in the engagement of the adaptive immune system and improved systemic anti-tumor immunity.

Treatment of intracranial disorders suffers from the inability to accumulate therapeutic drug concentrations due to protection from the blood–brain barrier (BBB). Electroporation-based therapies have demonstrated the capability of permeating the BBB, but knowledge of the longevity of BBB disruption (BBBD) is limited. In this study, we quantify the temporal, high-frequency electroporation (HFE)-mediated BBBD in an in vivo healthy rat brain model. 40 male Fisher rats underwent HFE treatment; two blunt tipped monopolar electrodes were advanced into the brain and 200 bursts of HFE were delivered at a voltage-to-distance ratio of 600 V/cm. BBBD was verified with contrast enhanced T1W MRI (gadopentetate dimeglumine) and pathologically (Evans blue dye) at time points of 1, 24, 48, 72, and 96 h after HFE. Contrast enhanced T1W scans demonstrated BBBD for 1 to 72 h after HFE but intact BBB at 96 h. Histologically, tissue damage was restricted to electrode insertion tracks. BBBD was induced with minimal muscle contractions and minimal cell death attributed to HFE. Numerical modeling indicated that brief BBBD was induced with low magnitude electric fields, and BBBD duration increased with field strength. These data suggest the spatiotemporal characteristics of HFE-mediated BBBD may be modulated with the locally applied electric field.

The proposed physics-based pre-treatment plan through finite element analysis and system for actively monitoring resistance changes can be paired to significantly reduce ablation times and risk of thermal effects during IRE procedures for LAPC.

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Purpose: To determine the safety and feasibility of percutaneous high-frequency irreversible electroporation (HFIRE) for primary liver cancer and evaluate the HFIRE-induced local immune response. Materials and Methods: HFIRE therapy was delivered percutaneously in 3 canine patients with resectable hepatocellular carcinoma (HCC) in the absence of intraoperative paralytic agents or cardiac synchronization. Pre-and post-HFIRE biopsy samples were processed with histopathology and immunohistochemistry for CD3, CD4, CD8, and CD79a. Blood was collected on days 0, 2, and 4 for complete blood count and chemistry. Numeric models were developed to determine the treatment-specific lethal thresholds for malignant canine liver tissue and healthy porcine liver tissue. Results: HFIRE resulted in predictable ablation volumes as assessed by posttreatment CT. No detectable cardiac interference and minimal muscle contraction occurred during HFIRE. No clinically significant adverse events occurred secondary to HFIRE. Microscopically, a well-defined ablation zone surrounded by a reactive zone was evident in the majority of samples. This zone was composed primarily of maturing collagen interspersed with CD3 þ /CD4 À /CD8 À lymphocytes in a proinflammatory microenvironment. The average ablation volumes for the canine HCC patients and the healthy porcine tissue were 3.89 cm 3 ± 0.74 and 1.56 cm 3 ± 0.16, respectively (P ¼ .03), and the respective average lethal thresholds were 710 V/cm ± 28.2 and 957 V/cm ± 24.4 V/cm (P ¼ .0004). Conclusions: HFIRE can safely and effectively be delivered percutaneously, results in a predictable ablation volume, and is associated with lymphocytic tumor infiltration. This is the first step toward the use of HFIRE for treatment of unresectable liver tumors.

Purpose: This study evaluates the effects of various pulsing paradigms, on the irreversible electroporation (IRE) lesion, induced electric current, and temperature changes using a perfused porcine liver model. Materials and methods: A 4-monopolar electrode array delivered IRE therapy varying the pulse length and inter-pulse delay to six porcine mechanically perfused livers. Pulse paradigms included six forms of cycled pulsing schemes and the conventional pulsing scheme. Finite element models provided further insight into the effects of cycled pulsing on the temperature and thermal injury distribution. Results: 'Single pulse cycle with no interpulse delay' deposited maximum average energy (2.34 ± 0.35 kJ) and produced the largest ratio of thermally damaged tissue area and IRE ablation area from all other pulse schemes (18.22% 6 8.11, p < .0001 all pairwise comparisons). These compared favorably to the conventional algorithm (2.09 ± 0.37 kJ, 3.49% 6 2.20, p < .0001, all comparisons). Though no statistical significance was found between groups, the '5 pulse cycle, 0 s delay' pulse paradigm produced the largest average IRE ablation cross sectional area (11.81 ± 1.97 cm 2), while conventional paradigm yielded an average of 8.90 ± 0.91 cm 2. Finite element modeling indicated a '10 pulse cycle, 10 s delay' generated the least thermal tissue damage and '1 pulse cycle, 0 s delay' pulse cycle sequence the most (0.47 vs. 3.76 cm 2), over a lengthier treatment time (16.5 vs. 6.67 minutes). Conclusions: Subdividing IRE pulses and adding delays throughout the treatment can reduce white tissue coagulation and electric current, while maintaining IRE treatment sizes.