2667 - Targeting the GOT2-TOMM22 Achilles' Heel: High-Throughput Screening and the Antidepressant Vortioxetine as a Radiosensitizer in Esophageal Cancer

Purpose/Objective(s): Radiotherapy (RT) resistance remains a major clinical challenge in esophageal squamous cell carcinoma (ESCC). Vortioxetine is a novel antidepressant with a favorable safety profile. Through high-throughput screening of an FDA-approved drug library combined with functional validation, we identified vortioxetine as a potent radiosensitizer in ESCC; however, the underlying mechanism remains unclear. This study aims to elucidate how vortioxetine targets the GOT2-TOMM22 axis to suppress mitochondrial metabolism, thereby limiting DNA damage repair and enhancing radiosensitivity.

Materials/Methods: To identify the direct target of vortioxetine, probe modification and pull-down assays coupled with mass spectrometry were performed. The interaction was validated by drug affinity responsive target stability (DARTS) and surface plasmon resonance (SPR). GST pull-down combined with mass spectrometry and co-immunoprecipitation (Co-IP) assays identified TOMM22 as a key GOT2-interacting protein. GOT2 knockdown was performed to assess its role in glutamine metabolism and TCA cycle activity via [¹³C5]-glutamine metabolic flux analysis. To mechanistically validate the hypothesis that vortioxetine disrupts the GOT2-TOMM22 interaction, we employed SPR with purified proteins, site-directed mutagenesis to map the binding interface. Radiosensitizing effects were assessed by clonogenic survival, ?H2AX foci, and reactive oxygen species (ROS) detection. In vivo efficacy was evaluated in radioresistant-cell derived xenograft models treated with vortioxetine combined with fractionated RT. Clinical relevance was examined by correlating GOT2/TOMM22 expression with RT response in ESCC patient specimens.

Results: Preliminary data demonstrate that vortioxetine directly binds to Leu97 ? Asn318 in NMT1. GOT2 physically interacts with TOMM22, a mitochondrial outer membrane translocase. Functional studies show that GOT2 knockdown significantly suppresses glutamine metabolism and TCA cycle activity, indicating that the GOT2-TOMM22 axis is critical for maintaining mitochondrial metabolic function.

Conclusion: This study will elucidate a novel mechanism by which vortioxetine targets the GOT2-TOMM22 axis to overcome RT resistance in ESCC. By repurposing a safe antidepressant, this research provides a readily translatable strategy for radiosensitization and establishes the GOT2-TOMM22 interaction as a potential therapeutic target in ESCC.