Abstract Background: Radiotherapy is is a pivotal treatment for locally advanced or inoperable non-small cell lung cancer (NSCLC), however its efficacy is limited by radioresistance. Approximately 44%-46% of patients develop recurrence or metastasis after chemoradiotherapy, with stage III patients showing a 30%-40% local recurrence rate within five years. Identification of effective therapeutic targets to enhance radiosensitivity remains a critical challenge in NSCLC management. This study aims to elucidate the molecular basis of radioresistance and develop novel approaches to enhance tumor radiosensitivity. Methods: Through RNA transcriptome sequencing of treatment-naive NSCLC patients (n=87) stratified by RECIST into radiotherapy-sensitive (n=55) and -resistant (n=32) cohorts, and based on histopathological and clinical diagnostic data from Tianjin Medical University Cancer Hospital (2015-2020), we analyzed the association between CYLD expression and radiotherapy outcomes through RNA sequencing and tissue microarrays. The impact of CYLD on radiosensitivity was assessed through flow cytometry and colony formation assays, and further verified in nude mouse xenograft models. To investigate CYLD-related pathways, we performed next-generation sequencing and examined its role in ferroptosis through Western blotting, qPCR, glutathione assays, malondialdehyde measurements, lipid peroxidation assessment, and transmission electron microscopy. Potential CYLD-interacting proteins were identified via mass spectrometry, immunofluorescence, and co-immunoprecipitation assays, while CYLD-mediated deubiquitination of ALOX12B was characterized using ubiquitination immunoprecipitation.Furthermore, we combined database predictions with Western blot, qPCR, chromatin immunoprecipitation, and luciferase reporter assays to establish EGR1 as a transcriptional regulator CYLD expression. Result: Our findings identify CYLD as a positive regulator of radiosensitivity in NSCLC, We demonstrate that CYLD enhances radiation sensitivity in NSCLC by stabilizing ALOX12B. Mechanistically, CYLD enhances ALOX12B protein stability by deubiquitinating K63- and K48-linked ubiquitin chains, thereby suppressing its proteasomal degradation. This stabilization potentiates cellular ferroptosis and consequently increases radiosensitivity. Furthermore, we identified EGR1 as an upstream transcriptional activator of CYLD that promotes ferroptosis in a CYLD-dependent manner. Conclusion: Our study demonstrates that CYLD, driven by its transcriptional activator EGR1, promotes ferroptosis and enhances radiosensitivity in NSCLC by mediating the deubiquitination and stabilization of ALOX12B. This study identifies CYLD as both a valuable prognostic biomarker and a potential molecular target for radiosensitization in NSCLC. Citation Format: Yihan Xu, Jiazhuo Yan, Huiwen Yu, Lu Zhang, Jinpu Yu, Zhiyong Yuan, . CYLD induces ferroptosis through ALOX12B to enhance radiosensitivity in non-small cell lung cancer 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 6620.
Xu et al. (Fri,) studied this question.