Poster S9 - Analysis of Tissue Impedance Changes following Electroporation

Saturday, October 26, 2024

10:00 AM - 11:00 AM EST

Location: Exhibit Hall E, F & G

Presenting Author(s)

Varad Gurude

Graduate Student
University of North Carolina at Chapel Hill
Milton, Georgia, United States

Co-Author(s)

Robert H. Williamson, BS (he/him/his)

Graduate Research Assistant
North Carolina State University, United States
JD

Jewels Darrow

Research Specialist
North Carolina State University, United States

Primary Investigator(s)

MS

Michael Sano, PhD

Associate Professor
North Carolina State University, United States

Introduction: Cancer continues to stand as a leading cause of death worldwide, presenting formidable challenges in managing inoperable solid tumors due to limited treatment efficacy and severe side effects. Among the most difficult types to treat are liver and pancreatic cancers, with low 5-year survival rates of 18% and 10%, respectively. Integrated Time Nanosecond Pulse Irreversible Electroporation (INSPIRE) is a promising emerging solution, utilizing ultrashort electrical pulses to selectively target malignant cells while sparing healthy tissues. This approach addresses several challenges associated with traditional electroporation treatments including the induction of intense muscle stimulation, the need for cardiac synchronization, and accidental induction of thermal damage to tissues adjacent to the applicators.

This study using the INSPIRE system aims to investigate the correlation between tissue impedance fluctuations during and post-electroporation treatments and the overall viability of tissues. By exploring these dynamics, we aim to establish informed treatment endpoints. The investigation will employ fixed doses that are currently standard in clinical practice to assess the efficacy of various methodologies. Our approach will focus on enhancing the monitoring of electroporation treatments through the analysis of impedance spectra and the establishment of pertinent treatment endpoints. Understanding the interplay between tissue impedance changes and cellular reactions is crucial for refining the treatment procedure and maximizing its effectiveness.

Materials and

Methods: Fresh liver and tumor tissues were collected from veterinary patients and clinical research animals, placed in cold phosphate-buffered saline (PBS), and stored on ice until experimentation. Using a punch biopsy, cylindrical tissue samples approximately 10mm in length were prepared. Each sample's dimensions were measured and recorded, then stored in cold PBS in 24-well plates on ice until the experiment. For both experimental and control groups, 20 samples (N=20) were prepared. To assess impedance changes following INSPIRE treatments, the impedance analyzer was calibrated using a 50-ohm precision resistor. Baseline impedance testing involved placing samples in an electrode holder and measuring impedance from 10 kHz to 10 MHz, with 10 measurements averaged by software. Values for 10 kHz, 100 kHz, and 1 MHz were recorded, and impedance plots were saved.

INSPIRE treatments were administered using the electroporation system with specific waveform, voltage, dose, and temperature settings. Pre-treatment temperatures were noted, and treatments were delivered. Post-treatment, samples were allowed to return to pretreatment temperatures before impedance measurements were repeated. Two energy delivery methods were evaluated: fixed and temperature controlled. Fixed energy treatments tested parameters including waveforms (1-1-1, 2-1-2, or 100-10-0), rates (500 µs/s, 750 µs/s, 1000 µs/s), a 0.02s dose, and 1000V. Each of the 4 parameters and controls had 4 replicates, totaling 20 samples. Temperature-controlled treatments used the same waveforms, voltage, and dose but were delivered at an initial rate of 300 µs/s which was algorithmically controlled to ensure target temperatures did not exceed set points of either 35℃, 45℃, and 55℃.

Results, Conclusions, and Discussions: The comparison of conductivity values before and after electroporation treatments in liver tissue biopsies demonstrated varying levels of statistical significance across different protocols. The control group, with p-values of 0.0559, 0.1441, and 0.6457 at frequency level of 10 kHz, 100 kHz, and 1 MHz respectively, showed no significant changes in conductivity, indicating that the electroporation treatments were responsible for the observed differences in the other protocols. In contrast, the treatment protocol with a waveform of 1-1-1, rate of 500 µs/s, dose of 0.02 s, and voltage of 1000 V the first two p-values were below the significance threshold of 0.05, indicating significant changes in conductivity. The third p-value, however, did not reach significance, suggesting variability in response. Similarly, the two remaining protocols with waveform 1-1-1, rate of 750 µs/s and 1000 µs/s, dose of 0.02 s, and voltage of 1000 V showed similar statistical results. In addition, as shown in Figure 1 for the protocol with a rate of 1000 µs/s, the graph comparing impedance revealed that pre-treatment values had higher magnitudes and greater standard deviation compared to post-treatment values. A similar pattern was observed for the additional protocols, indicating that electroporation consistently reduced impedance magnitude across different treatment settings.

The significant changes observed in several protocols underscore the effectiveness of electroporation in altering tissue properties. The non-significant p-values observed for the 1 MHz rate protocol could be attributed to several factors. Higher frequencies might affect tissue conductivity differently compared to lower frequencies, possibly due to less effective electroporation or variations in how the frequency interacts with tissue properties. The parameters used (waveform 1-1-1, dose 0.02 s, and voltage 1000 V) might not have been optimal for the 1 MHz frequency, leading to suboptimal treatment effects.

Future studies should re-examine these protocols across an increased sample size to enhance the reliability of these findings. Additional future studies will expand upon these findings to investigate INSPIRE therapy across various organs such as kidney, pancreas, and lungs. This broader exploration aims to clear how tissue-specific responses to electroporation can inform treatment strategies tailored to different anatomical structures.

Acknowledgements (Optional): Research reported in this publication was supported by the National Cancer Institute of the National Institutes of Health under award numbers 1R01CA276232 and R01CA272550, by the North Carolina Biotechnology Center under award number 2021-TRG-6708, and by the Joint Biomedical Engineering Department of the University of North Carolina Chapel Hill and North Carolina State University