Abstract 4236: The metabotropic Glutamate receptor-1 is a glutamatergic checkpoint that integrates with TCR to augment anti-tumor CD8+ T cell activation, metabolism and effector function

Abstract Background: Glutamate receptors (GluRs), classically linked to neuronal signaling, are also recognized as key immunomodulators. T cell activation requires precise coordination of TCR-driven signaling microclusters and calcium-dependent pathways that shape T cell effector function. Emerging evidence shows that glutamatergic cues influence T cell metabolism, activation, and fate, yet their role within TCR signaling remains poorly defined. Here, we show how GluR-TCR interactions govern CD8+ T cell activation and function, particularly within the metabolic and functional constraints of the tumor microenvironment. Methods: Tumors and lymphoid tissue single-cell suspensions were analyzed for the expression of ionotropic (GluA3, NR1, NR2B) and metabotropic (mGluR1, mGluR5) receptors on CD4+ and CD8+ TILs by flow cytometry. CD8+ T cells were activated with anti-CD3/CD28 or PMA/ionomycin ± IL-2 in the presence or absence of NBQX (GluA3) and CPCCOEt (mGluR1) inhibitors. Expression of glutamate transporters, viability, and extracellular glutamate were assessed by fluorometric assays. CPCCOEt-treated cells were analyzed for mGluR1-TCR colocalization and phosphorylation of TCR-signaling proteins by confocal microscopy and flow cytometry. Metabolic activity was measured by Seahorse XF96 (OCR/ECAR). Intracellular Ca2+ dynamics were evaluated using Fura-2 AM ratiometric imaging. CD8+ T cell proliferation was assessed by CFSE, and cytotoxicity was tested in vitro and in vivo using a pulmonary metastasis model; activated CD8+ T cells ± antagonists were transferred retro-orbitally. Results: We found that mGluR1 and GluA3 are expressed at higher levels in T cells compared with other immune cell lineages, and their expression increases during CD8+ T cell activation. Conversely, pharmacologic inhibition with CPCCOEt significantly impaired CD8+ T cell activation, disrupting mGluR1 colocalization with the TCR-Vβ8.1 in the cell membrane and reducing phosphorylation of key proteins involved in TCR-driven signaling, metabolic pathways, inducing a pronounced dysregulation of Ca2+ flux. Functionally, CPCCOEt-treated CD8+ T cells showed a reduction in proliferation to levels resembling naïve T cells and a marked decrease in cytotoxic activity. In vivo, these impairments translated into an increased burden of secondary metastatic pulmonary nodules, highlighting the essential role of mGluR1 signaling in sustaining effector CD8+ T cell function and antitumor immunity. Conclusion: These findings identify mGluR1 as a critical glutamatergic checkpoint that integrates with the TCR to augment CD8+ T cell activation, metabolism, and effector function. Its disruption collapses signaling strength, Ca2+ responses, and cytotoxic competence, revealing a potential target to boost T cell potency in solid tumor microenvironments. Citation Format: Salvador Gonzalez Ochoa, Maria Teresa P. de Aquino, Thomas W. Hodo, Thanigaivelan Kanagasabai, Muna A. Mohammed, Jane Tonello, Alla Ivanova, Anil Shanker, . The metabotropic Glutamate receptor-1 is a glutamatergic checkpoint that integrates with TCR to augment anti-tumor CD8+ T cell activation, metabolism and effector function [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2026; Part 1 (Regular Abstracts); 2026 Apr 17-22; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2026;86(7 Suppl):Abstract nr 4236.