Tumor Treating Fields (TTFields) Hinder Cancer Cell Motility through Regulation of Microtubule and Actin Dynamics

SIMPLE SUMMARY: Tumor Treating Fields (TTFields), encompassing alternating electric fields within the intermediate frequency range, is an anticancer treatment delivered to the tumor region through transducer arrays placed non-invasively on the skin. Although established as an anti-mitotic treatment modality, the anti-metastatic potential of TTFields and their effect on rapid cytoskeletal dynamics during cellular motility warrant further investigation. In this study, we report that TTFields application induces changes in microtubule organization leading to interference with the directionality and robustness of cancer cell migration. We show that these changes in microtubule organization result in activation of GEF-H1/RhoA/ROCK signaling pathway, and the consequent formation of focal adhesions and changes in actin cytoskeleton architecture. Together, these results propose a novel mechanism by which TTFields induce changes in microtubule and actin organization and dynamics, thereby disrupting processes important for polarity generation and motility in cancer cells. ABSTRACT: Tumor Treating Fields (TTFields) are noninvasive, alternating electric fields within the intermediate frequency range (100–300 kHz) that are utilized as an antimitotic cancer treatment. TTFields are loco-regionally delivered to the tumor region through 2 pairs of transducer arrays placed on the skin. This novel treatment modality has been FDA-approved for use in patients with glioblastoma and malignant pleural mesothelioma based on clinical trial data demonstrating efficacy and safety; and is currently under investigation in other types of solid tumors. TTFields were shown to induce an anti-mitotic effect by exerting bi-directional forces on highly polar intracellular elements, such as tubulin and septin molecules, eliciting abnormal microtubule polymerization during spindle formation as well as aberrant cleavage furrow formation. Previous studies have demonstrated that TTFields inhibit metastatic properties in cancer cells. However, the consequences of TTFields application on cytoskeleton dynamics remain undetermined. In this study, methods utilized in combination to study the effects of TTFields on cancer cell motility through regulation of microtubule and actin dynamics included confocal microscopy, computational tools, and biochemical analyses. Mechanisms by which TTFields treatment disrupted cellular polarity were (1) interference with microtubule assembly and directionality; (2) altered regulation of Guanine nucleotide exchange factor-H1 (GEF-H1), Ras homolog family member A (RhoA), and Rho-associated coiled-coil kinase (ROCK) activity; and (3) induced formation of radial protrusions of peripheral actin filaments and focal adhesions. Overall, these data identified discrete effects of TTFields that disrupt processes crucial for cancer cell motility.