Abstract 1053: Combination therapy with nonthermal plasma and tirapazamine selectively targets glioblastoma cells and regulates the gap junction phenotype
Authors: Yehl Matthew , Nicolais Andrew , Gerardi Samuel , Dianat Ahmad , Kucharski Olivia , Hutcherson Timothy C. , Zucker Shoshanna N. 
Published: June 2022

Abstract
Considering the side effects common to most chemotherapeutic agents, a novel approach was developed to selectively treat cancer by targeting the oxidative stress pathway which is induced in many tumor types. Our previous studies demonstrated that nonthermal plasma (NTP) selectively causes apoptotic cell death in melanoma cells while preserving the normal surrounding keratinocytes. The prodrug, tirapazamine, was shown to potentiate the effects of NTP while maintaining selectivity because tirapazamine only activates in hypoxic conditions, an environment common to many tumors. This patented combination therapy has an additive-to-synergistic effect on inducing apoptosis in melanoma. Previous studies have also demonstrated efficacy in a broad range of tumor cells including cervical carcinoma, colon cancer, and pancreatic adenocarcinoma. This report demonstrates for the first time that while NTP alone kills a targeted area of glioblastoma cells and tirapazamine alone kills most of the susceptible cells, the combination of NTP and tirapazamine kills all the cells throughout the tissue culture plate. The mechanism for the increase in the surface area of cell death was identified as the passage of reactive oxygen species (ROS) by intercellular communication through gap junctions. The IC50 dose of tirapazamine required to target several glioblastoma cell lines was less than that for melanoma cells. Through parameter optimization with varied plasma conditions and altered drug dosing, the selective killing of glioblastoma cells was achieved without affecting normal primary astrocytes. Primary astrocytes and glioblastoma cells were co-cultured to observe the effect on the optimal targeting dose of tirapazamine. Glioblastoma cells had an IC50 at hypoxia which was reduced as compared to astrocytes in their normoxia state. The Glioblastoma IC50 values were further decreased when co-cultured with astrocytes. This study documents an increase in the Connexin43 (Cx43) gap junction protein, upon treatment with NTP for both astrocytes and glioblastoma cells. However, treatment with tirapazamine alone ameliorates this effect in glioblastoma cells only and not in astrocytes. The combination therapy of NTP and tirapazamine increased this effect to levels comparable to the original expression levels. The regulation of Cx43 may reduce the metastatic potential of the glioblastoma cells while enabling the combination therapy to show efficacy both between glioblastoma cells and in the context of communication with astrocytes. By this mechanism, the astrocytes may act as conduits to transport ROS between pockets of glioblastoma cells. In summary, the results demonstrate that the novel combination therapy of NTP and tirapazamine is an effective method to selectively target glioblastoma cells with a potential for clinical applications with limited adverse reactions.
Citation Format: Matthew Yehl, Andrew Nicolais, Samuel Gerardi, Ahmad Dianat, Olivia Kucharski, Timothy C. Hutcherson, Shoshanna N. Zucker. Combination therapy with nonthermal plasma and tirapazamine selectively targets glioblastoma cells and regulates the gap junction phenotype [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 1053.