HILPDA, Regulated by WTAP in an m6A-Dependent Manner, Enhances Osteosarcoma Development by Inhibiting Ferroptosis and Promoting Glycolysis.

Osteosarcoma (OS) is the most common primary malignant bone cancer. Hypoxia-inducible lipid droplet-associated protein (HILPDA) promotes tumor growth, migration, and ferroptosis resistance in OS. However, the oncogenic mechanisms of HILPDA in OS are largely unexplored. Glycolytic activity was assessed by measuring glucose uptake and lactate release. Ferroptosis was evaluated by measuring ROS, MDA, Fe2+, and total iron contents. Cell proliferation was assessed via EdU incorporation assay, and tube formation ability was analyzed. Animal experiments were used to test the function in vivo. Expression analysis was performed using quantitative PCR, immunoblotting, and immunohistochemistry. The regulation of Wilms' tumor 1-associating protein (WTAP) in HILPDA was detected by methylated RNA immunoprecipitation (MeRIP) assay. HILPDA and WTAP were upregulated in OS tissues. Overexpression of HILPDA or WTAP was associated with poor prognosis in OS patients. HILPDA deficiency impaired OS cell proliferation and glycolysis while promoting ferroptosis, concurrently attenuating HUVEC tube formation in vitro. Moreover, HILPDA depletion constrained the in vivo growth of HOS OS xenografts. Mechanistically, WTAP regulated HILPDA expression through an m6A-dependent manner. Additionally, HILPDA re-expression reversed sh-WTAP-mediated alterations in OS cell glycolysis and ferroptosis and HUVEC tube formation. Our findings identify the WTAP/HILPDA axis as a key regulator of OS development through coordinated control of glycolysis, ferroptosis, and angiogenesis, offering new avenues for multi-targeted therapeutic intervention in OS.