AFF3 regulates ferroptosis and malignant progression in bladder cancer cells via the NRF2/GPX4 signaling axis
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Bladder cancer (BCa) is a common malignancy of the urinary system, and its malignant progression is closely associated with ferroptosis. As a transcriptional regulator in the AFF family, AFF3 participates in the regulation of gene expression in multiple diseases; however, its role in BCa and its relationship with ferroptosis remain unclear. This study aimed to investigate the role of AFF3 in BCa progression and the molecular mechanism through which it regulates ferroptosis. AFF3 expression was examined through bioinformatics analysis and experiments in clinical tissues and cells. AFF3 was overexpressed in UM-UC-3 cells, and cell proliferation and migration were assessed using CCK-8, colony formation, and Transwell assays. Ferroptosis was evaluated using BODIPY 581/591 C11, MDA, and ferroptosis-related protein assays, and rescue experiments were performed with the ferroptosis inhibitor Fer-1. A subcutaneous xenograft model was established in nude mice, and the NRF2 activator TBHQ was used to verify the in vivo role and mechanism of AFF3 in regulating ferroptosis. Our results revealed that AFF3 was expressed at low levels in BCa. Functionally, AFF3 overexpression inhibited BCa cell proliferation, colony formation, and migration; promoted the accumulation of lipid ROS and MDA; downregulated GPX4 and SLC7A11 expression; and increased sensitivity to ferroptosis. Notably, Fer-1 partially reversed these effects. Mechanistically, AFF3 upregulated KEAP1 and inhibited NRF2 nuclear translocation, thereby reducing GPX4 expression; conversely, TBHQ partially reversed these molecular and functional changes. In vivo animal experiments further confirmed that AFF3 overexpression inhibited xenograft growth and promoted ferroptosis-related changes, whereas TBHQ attenuated the regulatory effects of AFF3. Overall, these findings demonstrate that AFF3 exerts tumor-suppressive effects in BCa by promoting KEAP1 expression and inhibiting NRF2 nuclear translocation and GPX4-mediated antioxidant defense, thereby enhancing the sensitivity of tumor cells to ferroptosis and suppressing the malignant progression of BCa.
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Ethics Approval
The study involving human bladder cancer tissues was reviewed and approved by the Ethics Committee of the Second Affiliated Hospital of Kunming Medical University, All animal experiments were reviewed and approved by the Institutional Animal Care and Use Committee of the Second Affiliated Hospital of Kunming Medical University Animal CenterCRediT authorship contribution
Song Chen, conceptualization, methodology, investigation, writing – original draft, writing – review & editing. Jiansong Wang, conceptualization, methodology, investigation, writing – original draft, writing – review & editing. Xiangyun Li, data curation, formal analysis, writing – review & editing. Peiqin Zhan, data curation, formal analysis, writing – review & editing. Tianjiao Luo, formal analysis, writing – review & editing. All authors read and approved the final manuscript.
Data Availability Statement
The datasets used and analyzed during the current study are available from the corresponding author on reasonable request.
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