LAUSR

Changes in the mechanical microenvironment and mechanical signals are observed during tumor progression, malignant transformation, and metastasis. In this context, understanding the molecular details of mechanotransduction signaling may provide unique therapeutic targets. Here we report that normal breast epithelial cells are mechanically sensitive, responding to mechanical stimuli through a two-part calcium signaling mechanism. We observed an immediate, robust rise in intracellular calcium (within seconds) followed by a persistent extracellular calcium influx (up to 30 minutes). This persistent calcium was sustained via microtubule-dependent mechano-activation of NADPH oxidase 2 (NOX2)-generated reactive oxygen species (ROS), which acted on TRPM8 channels to prolong calcium signaling. In contrast, the introduction of a constitutively-active oncogenic KRas mutation inhibited the magnitude of initial calcium signaling and severely blunted persistent calcium influx. The identification that oncogenic KRas suppresses mechanically-induced calcium at the level of ROS provides a novel mechanism for how KRas could alter cell responses to tumor microenvironment mechanics and may reveal new chemotherapeutic targets for cancer. Moreover, we find that expression changes in both NOX2 and TRPM8 mRNA predicts poor clinical outcome in estrogen receptor (ER) negative breast cancer patients, a population with limited available treatment options. The clinical and mechanistic data demonstrating disruption of this mechanically-activated calcium pathway in breast cancer patients and by KRas activation reveal novel signaling alterations that could influence cancer cell responses to the tumor mechanical microenvironment and impact patient survival. Citation Format: Stephen JP Pratt, Rachel M. Lee, Erick O. Hernandez-Ochoa, Eleanor C. Ory, Keyata N. Thompson, Patrick C. Bailey, Trevor J. Mathias, Julia A. Ju, Michele I. Vitolo, Martin F. Schneider, Joseph P. Stains, Christopher W. Ward, Stuart S. Martin. Mechanoactivation of NOX2-generated ROS elicits persistent TRPM8 Ca2+ signals that are inhibited by oncogenic KRas [abstract]. In: Proceedings of the Annual Meeting of the American Association for Cancer Research 2020; 2020 Apr 27-28 and Jun 22-24. Philadelphia (PA): AACR; Cancer Res 2020;80(16 Suppl):Abstract nr 2575.