Abstract 2030: N-cadherin-mediated activation of PI3K/Akt pathway following application of tumor treating fields (TTFields)
Authors: Klein-Goldberg Anat , Voloshin Tali , Zemer-Tov Efrat , Paz Rom , Koren Lilach , Wainer-Katsir Kerem , Volodin Alexandra , Koltun Bella , Brant Boris , Barsheshet Yiftah , Kan Tal , David Cfir , Khalil Tharwat Haj , Haber Adi , Giladi Moshe , Weinberg Uri , Palti Yoram 
Published: March 2024

Abstract
Introduction: Tumor Treating Fields (TTFields) therapy is FDA-approved for the treatment of glioblastoma and unresectable pleural mesothelioma, and has recently demonstrated benefit (together with an immune checkpoint inhibitor) for the treatment of metastatic non-small cell lung carcinoma (NSCLC) progressing on or after platinum-based therapy. The current study aimed to uncover cellular response mechanisms to TTFields.
Methods: A2780 ovarian cancer cells and H1299 NSCLC cells were treated with TTFields for 72 h (200 and 150 kHz, respectively; 1.7 V/cm RMS). Changes in signaling pathways were analyzed using the Luminex multiplex assay and validated by protein expression. Fluorescence microscopy, calcium switch assay, antibody neutralization assay, and immunoprecipitation were employed to investigate the mechanism of action. The efficacy of concomitant treatment with TTFields and a potential inhibitor was tested in cell lines (cytotoxic and clonogenic effects) and two orthotopic mouse models: MOSE-L-FFL ovarian cancer and LL/2 lung cancer. Immunohistochemistry was performed on tumor sections from both models.
Results: Increased AKT phosphorylation was observed in the cancer cells treated with TTFields. N-cadherin, known to be involved in the activation of the PI3K/AKT pathway, was elevated in the membranes of cells treated with TTFields. Elimination of calcium ions required for N-cadherin homophilic ligation abrogated TTFields-induced AKT phosphorylation, whereas calcium supplementation restored AKT phosphorylation. Inhibition of N-cadherin-mediated cell-cell contacts by neutralizing antibody resulted in a significant reduction in TTFields-induced AKT phosphorylation. Pull-down assays with an anti-N-cadherin antibody demonstrated increased recruitment of the p85 regulatory subunit of PI3K to N-cadherin complexes following TTFields application. TTFields-induced activation of AKT could be mitigated in cell cultures as well as in animal models by application of the PI3K inhibitor alpelisib. Co-treatment with alpelisib also enhanced the effectiveness of TTFields in vitro and in vivo.
Conclusions: The PI3K/AKT signaling pathway, activated via cell-cell N-cadherin interactions, plays a role in the cancer cell response to TTFields treatment. Inhibiting the PI3K/AKT pathway may potentially sensitize tumors to TTFields.
Citation Format: Anat Klein-Goldberg, Tali Voloshin, Efrat Zemer-Tov, Rom Paz, Lilach Koren, Kerem Wainer-Katsir, Alexandra Volodin, Bella Koltun, Boris Brant, Yiftah Barsheshet, Tal Kan, Cfir David, Tharwat Haj Khalil, Adi Haber, Moshe Giladi, Uri Weinberg, Yoram Palti. N-cadherin-mediated activation of PI3K/Akt pathway following application of tumor treating fields (TTFields) [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 2030.