Abstract 1382: Targeting Akt signaling pathway potentiates the antitumor effect of Tumor Treating Fields (TTFields) in vitro
Authors: Klein-Goldberg Anat , Voloshin Tali , Zemer-Tov Efrat , Paz Rom , Koren Lilach , Volodin Alexandra , Brant Boris , Giladi Moshe , Weinberg Uri , Palti Yoram 
Published: July 2021

Abstract
Introduction Tumor Treating Fields (TTFields) therapy is a clinically applied anti-neoplastic treatment modality, approved for the treatment of glioblastoma and malignant pleural mesothelioma. TTFields are low intensity (1-3 V/cm) alternating electric fields within the intermediate frequency range (100-300 kHz), that are applied loco-regionally and non-invasively to the tumor site. Although TTFields therapy has demonstrated improved progression free survival and overall survival, tumor recurrence still occurs in some patients. To evaluate the cellular mechanisms of treatment resistance to TTFields, we analyzed changes in signaling pathway mediators in treated human glioma (U-87 MG) and ovarian (A2780) cancer cell lines. Also, we examined the in vitro efficacy of TTFields in combination with therapies that target molecular candidates that may confer resistance to TTFields.
Methods To establish TTFields resistant glioma and ovarian cancer cell lines in vitro, 2 approaches were tested: 1) cyclic repetitions, where TTFields treatment was repeatedly applied for 72 hours with 24 hours breaks between treatment cycles; and 2) continuous, long-duration application of TTFields (1 or 2 weeks, depending on the cancer cell line type) without treatment breaks. Luminex assay followed by Western blot analysis were used to study changes in signaling pathways following TTFields treatment. Finally, we examined the in vitro efficacy of continuous, long-duration application of TTFields in combination with BGT226 (a phosphatidylinositol-3-kinase (PI3K)/mammalian target of rapamycin (mTOR) dual inhibitor).
Results Application of TTFields using cyclic repetition significantly decreased proliferation of human glioma and ovarian cancer cells, whereas cells treated using continuous, long-duration application of TTFields showed reduced sensitivity to TTFields. Luminex analysis clearly showed activation of the PI3K/mTOR/AKT signaling pathway in response to TTFields treatment. To clarify the importance of this signaling pathway on regulating treatment resistance to TTFields, we examined the effect of TTFields, alone and in combination, with a PI3K/mTOR inhibitor (BGT226) on cancer cell proliferation, apoptosis, and activation of AKT. While long-duration application of TTFields led to Akt activation, TTFields in combination with BGT226 resulted in reduction of p-AKT S473, with a corresponding blockade of proliferation, induction of apoptosis, and decrease in the clonogenic potential.
Conclusions We propose here a new mechanism of resistance to prolonged TTFields treatment mediated by the PI3K/mTOR/AKT signaling pathway in glioma and ovarian cancer cells. We demonstrate that combination therapy of TTFields and a targeted PI3K/mTOR dual inhibitor (BGT226) inhibited AKT S473 phosphorylation and sensitized cancer cells to “long-duration” TTFields application.
Citation Format: Anat Klein-Goldberg, Tali Voloshin, Efrat Zemer-Tov, Rom Paz, Lilach Koren, Alexandra Volodin, Boris Brant, Moshe Giladi, Uri Weinberg, Yoram Palti. Targeting Akt signaling pathway potentiates the antitumor effect of Tumor Treating Fields (TTFields) in vitro [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2021; 2021 Apr 10-15 and May 17-21. Philadelphia (PA): AACR; Cancer Res 2021;81(13_Suppl):Abstract nr 1382.