2787 - MTA3 and Radiosensitivity of Non-Small Cell Lung Cancer through Enhancing Glutamine Synthetase-Mediated Cancer Stemness

L. Du¹, S. Zeng², M. Wang², Y. Jiang², D. Tao¹, D. Yang², and W. Zhou¹; ¹Radiation Oncology Center, Chongqing University Cancer Hospital, Chongqing, China, ²Department of Radiation Oncology, Chongqing University Cancer Hospital, Chongqing, China

Purpose/Objective(s): Metastasis-associated protein 3 (MTA3) exhibits dual roles as an oncogene or tumor suppressor, varying across cancer types. However, The impact of MTA3 on the radiosensitivity of non-small cell lung cancer (NSCLC) patients remains unclear. We thus investigated the role of MTA3 in modulating radiosensitivity in NSCLC, the predominant pathological variant of lung cancer, constituting around 85% of total cases. Materials/

Methods: To assess the influence of MTA3 on NSCLC radiosensitivity, colony formation assays and xenograft tumor assays in nude mice were employed. The role of glutaminolysis was examined using glutamine consumption and glutamate production assay kits, alongside Glutaminase (GLS) and Glutamine Synthetase (GS) Activity Assay Kits for metabolic enzyme activity analysis. The regulatory mechanism of glutaminolysis by MTA3 was confirmed using Chromatin immunoprecipitation assay and Gaussia luciferase assay. The expression levels of MTA3 and GS in NSCLC primary tissues were evaluated using immunohistochemistry. Survival outcomes were analyzed using Kaplan-Meier curves and log-rank tests.
Results: The colony formation assay showed that MTA3 depletion and overexpression caused significantly lower and higher clonogenic survival following different doses of irradiation (IR), respectively. Xenograft tumors in nude mice from MTA3 knockdown and overexpressed cells were notably smaller and larger after IR, respectively. These findings indicate MTA3s potential to reduce radiosensitivity in vitro and in vivo. Meanwhile, knockdown and overexpressed MTA3 can repress and expedite glutamine consumption and glutamate production uniformly, respectively. To determine how MTA3 acts on glutaminolysis, the activity of two specific metabolic enzymes dominate this metabolism, GS and GLS, were evaluated. It found that MTA3s knockdown and overexpression predominantly inhibited and enhanced GS activity, respectively, with minimal impact on GLS. Notably, the radiosensitivity reduction mediated by MTA3 overexpression was significantly mitigated by GS inhibition, highlighting GSs critical role in MTA3-regulated radiosensitivity reduction. Mechanistically, Chromatin immunoprecipitation assay and Gaussia luciferase assay showed that MTA3 was recruited to the promoter of GS and promoted GS transcription. However, knockdown of EBF1 abolished MTA3s promoting effect on GS and inhibited the MTA3s occupation on the promoter region of GS. These results collectively demonstrated that, in NSCLC cells, MTA3 is recruited by EBF1 to augment GS expression, thereby enhancing glutaminolysis and diminishing radiosensitivity. Finally, we showed that the NSCLC patients in the MTA3^high/GS^high group is signi?cantly associated with shorter overall survival.
Conclusion: MTA3 is capable of suppressing radiosensitivity through upregulating GS and MTA3^high/GS^high might be a potential prognostic factor for NSCLC patients.