Abstract 1212: Disruption of Gαs/cAMP-mediated acid sensing by breast tumor cells blunts tumor-induced immune suppression and decreases bone metastases by enhancing cytotoxic T cell function
Authors: Kaylee M. O'Donnell, Xinming Su, Netra Navadkar, Yalin Xu, Kristin A. Kwakwa, Mahta Mardani, Will A. Mattock, Nicole M. White, Christopher A. Maher, Katherine N. Weilbaecher
Journal: Cancer Research (2025)

Abstract Patients with bone metastases treated with immune checkpoint blockade (ICB) often have decreased response rates compared to patients with visceral metastases only. Increased proton extrusion by tumor cells due to the Warburg effect is associated with tumor progression, metastasis, reduced efficacy of standard-of-care treatments, and impaired immune cell function. Attempts to decrease tumor and systemic pH have not been successful in part due to homeostatic mechanisms and poor tolerability. Since the discovery of proton sensing GPCRs, we and others have studied the implications of acidosis in the tumor microenvironment on signaling and transcriptional changes in different cell types. We previously demonstrated that cyclic AMP (cAMP) signaling in response to acidic extracellular pH (pHe) synergizes with granulocyte-macrophage colony stimulating factor (GM-CSF), a myeloid cell supportive cytokine, signaling to increase Arginase 1 expression in tumor infiltrating myeloid cells (TIMs). We demonstrated that GM-CSF and low pHe are both required for immune suppression and resistance to ICB in breast cancer bone tumor models. Further, we found that TNF and IL-1β are expressed by immune suppressive TIMs and these cytokines significantly augment GM-CSF production in breast tumor cell lines, but not lung or melanoma. Here we examined the role of acid-induced signaling through Gαs/cAMP in breast cancer-mediated immunosuppression. We found that GM-CSF production increases synergistically with TNF or IL-1β treatment and low pH culture media in syngeneic murine PyMT-derived luminal B ER+ BO1 and TNBC 4T1 breast cancer cell lines in vitro. Treatment with inhibitors of adenylyl cyclase (MDL-12, 330A) and downstream Epac1/2 (ESI-09) reduced GM-CSF production. We then knocked out G-protein Gαs (Gnas) in these cells and found that while the growth kinetics and acid production were the same as wildtype (WT) controls, GnasKO cells had significantly decreased GM-CSF expression compared to GnasWT cells at all pH treatments. When intracardially injected into immune competent mice, GnasKO tumors had a 23-fold lower burden in hindlimbs than GnasWT cells and caused less osteolysis at terminal endpoint. GnasWT and GnasKO tumor burden was equivalent in CD8-depleted mice, implicating tumor proton sensing with cytotoxic T cell function. We propose that breast tumor cell pHe sensing through cAMP signaling synergizes with inflammatory cytokine signaling to produce GM-CSF, a necessary cytokine for myeloid immune modulation. Further, CD8+ T cell function was dependent upon tumor cell cAMP signaling, whether directly or indirectly through other immune or stromal cell communication. Limiting acid-induced signaling by tumor and myeloid cells may bolster an antitumor effect and increase the efficacy of ICB in breast cancer bone metastases. Citation Format: Kaylee M. O'Donnell, Xinming Su, Netra Navadkar, Yalin Xu, Kristin A. Kwakwa, Mahta Mardani, Will A. Mattock, Nicole M. White, Christopher A. Maher, Katherine N. Weilbaecher. Disruption of Gαs/cAMP-mediated acid sensing by breast tumor cells blunts tumor-induced immune suppression and decreases bone metastases by enhancing cytotoxic T cell function [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2025; Part 1 (Regular Abstracts); 2025 Apr 25-30; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2025;85(8_Suppl_1):Abstract nr 1212.