Abstract Osteosarcoma is the leading primary malignant bone tumor in children and young adults. Despite multimodal therapy involving surgery and multi-agent chemotherapy, patient survival rates have remained stagnant over the past four decades, necessitating the development of novel, effective therapeutic strategies. Ferroptosis, an iron-dependent programmed cell death, is a promising therapeutic target in osteosarcoma. Our recent findings indicate that osteosarcoma cells may circumvent GPX4-mediated ferroptosis by promoting iron detoxification through the ferroportin (FPN) exporter. This study was designed to define the molecular mechanisms by which osteosarcoma cells withstand redox stress associated with excess intracellular iron and to assess the potential impact of this regulation on the tumor immune microenvironment. First, we found that the TFRC gene, encoding the transferrin receptor responsible for cellular iron uptake, was associated with poor survival time in cervical cancer, glioblastoma, and liver cancer, and renal cancer in TCGA datasets. In contrast, SLC40A1 encoding FPN showed favorable clinical associations in liver cancer, pancreatic adenocarcinoma, and renal cancer. In human osteosarcomas (n = 88) from the Target Osteosarcoma database, SLC40A1 expression was positively correlated with overall immune gene scores (r = 0.241, p = 0.024) and M1 macrophage transcripts (r = 0.340, p = 0.001). Comparative transcriptomic analysis in canine osteosarcomas (n = 43) revealed that iron-related genes, including FTH1 (r = 0.659, p 0.0001), SLC39A8 (r = 0.665, p 0.0001), and ALAS1 (r = 0.573, p 0.0001), were associated with immune scores. In DOUG canine osteosarcoma cells, treatment with the GPX4 inhibitor RSL3 significantly altered the expression signature of genes involved in tissue damage and remodeling, including those in in the ECM-receptor interaction, PI3K-AKT, and AGE-RAGE signaling pathway. Specifically, RSL3 treatment resulted in the marked upregulation of ARNT (encoding HIF-1β) and ALDH3A1, both critical components of detoxification and oxidative stress response pathways. Furthermore, Erastin, another ferroptosis inducer, substantially increased HMOX1, a key antioxidant defense gene, in both canine DOUG and COS31 and DOUG canine osteosarcoma cells. These data collectively suggest that osteosarcoma cells employ robust regulatory mechanisms governing iron detoxification and antioxidant defense, potentially influencing tumor immunity. Furthermore, our ongoing studies are investigating the mechanistic role of the GPX4-ARNT-FTH1 axis in immunomodulatory processes in osteosarcomas. This work will also seek to identify a conserved cross-species mechanism that regulates the interplay between iron metabolism and tumor immunity, enabling translational models using naturally occurring canine osteosarcoma. Citation Format: Md Abdullah, Donghee Lee, Jong Hyuk Kim. GPX4-FTH1-ARNT signaling regulates iron efflux to suppress ferroptotic cell death and promote immune remodeling in osteosarcoma 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 5677.
Abdullah et al. (Fri,) studied this question.