Metabolic inhibition of glutamate-cysteine ligase increases dendritic cell-mediated antitumor immunity in melanoma

BACKGROUND: ) in melanocytes to investigate if aberrant glutamate metabolism drives tumor formation and affects immune cell function. METHODS: We performed liquid chromatography-tandem mass spectrometry (LC-MS/MS)-based metabolomic analyses and RNA sequencing on tumor-free and tumor-bearing tg(Grm1)EPv tissues to characterize metabolic alterations associated with tumor progression. Flow cytometry was used to examine changes in immune cell subsets within the TME. To assess the functional relevance of glutamate metabolism, we inhibited glutathione metabolism using L-buthionine-(S,R)-sulfoximine (BSO), an inhibitor of glutamate-cysteine ligase that depletes cellular glutathione levels. RESULTS: LC-MS/MS-based metabolomic analyses and RNA sequencing revealed changes in glutamate and glutamine metabolism, a glycolytic shift (Warburg effect), and reduced ATP levels in advanced tumors compared with tumor-free tissue, suggesting respiratory chain dysfunction. These metabolic changes in the TME are advantageous for the tumor cells and unfavorable for immune cells, such as dendritic cells (DC). Indeed, flow cytometry analysis of myeloid subsets during tumor progression showed a decline in tumor-infiltrating conventional type 2 DC and macrophages, alongside an increase in neutrophil and monocyte populations in advanced lesions. Interference with glutamate metabolism using BSO induced immunogenic cell death, namely ferroptosis, in an tg(Grm1)EPv-derived cell line in vitro. Therefore, we evaluated the combination of this inhibitor with immunotherapy as a promising new approach for the treatment of tumors in the tg(Grm1)EPv mouse model. We observed that tumor growth could be delayed in vivo when BSO was combined with a therapy regimen boosting DC numbers and activation. This inhibition of tumor growth was supported by the infiltration of activated T cells. CONCLUSION: Overall, our findings provide novel insights into the importance of combining metabolic intervention with immunotherapy for the treatment of patients with melanoma, particularly those bearing glutamate pathway-active or immunologically cold tumors. This knowledge can drive the design of novel therapeutic strategies for patients with cancer.