Abstract 819: Investigating the impacts of membrane potential on nano-electroporation efficacy
Authors: Shotbolt Max , Andre Victoria , Conlan Skye , Liang Ping , Khizroev Sakhrat 
Published: April 2023

Abstract
Electroporation has been shown to be an effective method of intracellular drug delivery and tissue ablation, however it faces significant challenges not only in clinical application due to the risks associated with the necessity of high voltage electrodes, but also due to a lack of fundamental research into the electrical characteristics of cancer cells. In order to improve outcomes in the clinic when using irreversible electroporation(IRE), it will be crucial to create a deeper understanding of the mechanisms underlying electroporation, such as the impacts of resting membrane potential on IRE outcomes. Magnetoelectric-nanoparticles (MENPs)have proven their ability to generate electric fields when stimulated by an external magnetic field and thus may be an effective alternative to electrodes for in-vivo electroporation and electro-chemotherapy. The ability of MENPs to electroporate cells is limited by the strength of their magnetoelectric (ME)effect, as such it is important to optimize MENPs fabrication for future applications. Improving this technology will require a deeper understanding of the membrane potential and other electrical properties of cancer cells.We fabricated MENPs consisting of a magnetostrictive CoFe2O4 core coupled with a piezoelectric BaTiO3shell. SKOV-3 and HOMEC cells were seeded in culture plates and given a 50ul dose of particle suspension, and 10ul of propidium iodide(PI). The cell samples were then stimulated by a 10 minute exposure to an alternating magnetic field of 400Oe at 100Hz. Fluorescence-microscope images were taken at 5 minute intervals during, and for 10 minutes following, the magnetic stimulation. These images were processed in MATLAB to quantify total fluorescence, and thus PI uptake, over time for each cell culture-particle combination.MENPs coupled with magnetic field stimulation were able to cause PI uptake in both HOMEC and SKOV-3 cells, indicating electroporation had occurred. By adding PI at time intervals after stimulation, we were able to determine whether electroporation had been reversible or irreversible by showing dye uptake would stop soon after stimulation had ended in the case of reversible EP but not IRE. We found SKOV-3 to have been reversibly electroporated, and HOMEC to have undergone IRE. To understand these results we used Oxonol uptake in a flow cytometry measurement to estimate resting membrane potential (RMP). HOMEC cells were shown to have a RMP approximately 20mV smaller than SKOV-3, which corresponded to the occurrence of IRE in HOMEC compared to reversible electroporation in SKOV-3.These results demonstrate the efficacy of MENPs to electroporate mammalian cells in the presence of a magnetic field. They also indicate a link between RMP and reversibility of electroporation. Further research into the electrical properties of cancer cells will be needed to formulate a complete model of the interactions between the nanoparticles and mammalian cells.
Citation Format: Max Shotbolt, Victoria Andre, Skye Conlan, Ping Liang, Sakhrat Khizroev. Investigating the impacts of membrane potential on nano-electroporation efficacy [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 819.