Impact of reoxygenation on Prolyl hydroxylase domain 3 expression and cisplatin sensitivity in breast cancer stem cells after prolonged hypoxia

Background: Targeting cancer stem cells (CSCs) within the tumor microenvironment has emerged as a critical focus in the development of novel oncological therapies. CSCs are frequently subjected to prolonged hypoxic conditions; while subsequent reoxygenation has been shown to significantly enhance the efficacy of radiotherapy across various cancer cell lines, its impact on chemotherapeutic resistance remains poorly understood. Methods: In this study, an in vitro model was utilized where breast cancer stem cells were treated with cobalt chloride (CoCl₂) for 72 hours to induce hypoxia, followed by a 24-hour incubation in CoCl₂-free medium to simulate rapid reoxygenation. Alterations in hypoxia-related gene expression were quantified using RT-qPCR, and changes in BCSC chemoresistance were evaluated via alamarBlue-based IC50 assays. Results: Both prolonged hypoxia and rapid reoxygenation significantly modulated the expression of the hypoxia-responsive gene HIF-1α and its regulatory target, PHD3. Notably, the expression of the drug efflux transporter ABCG2 was markedly downregulated upon reoxygenation following hypoxic preconditioning. Despite this molecular alteration, the associated reduction in cisplatin resistance did not reach statistical significance. Conclusion: Collectively, these findings provide preliminary evidence that integrating oxygen-dependent therapeutics or enhancing oxygen delivery to hypoxic tumor cells influences resistance-associated gene expression, though further optimization is needed to significantly improve chemotherapy efficacy.