Background <p>Although 15-hydroxyprostaglandin dehydrogenase (HPGD) is known to regulate the metabolism of prostaglandins and lipoxin A4, and its dysregulation has been implicated in various cancers, its role in esophageal squamous cell carcinoma (ESCC) has not been determined. This study is the first to comprehensively characterize HPGD expression in ESCC and establish its clinical relevance in predicting patient outcomes. Furthermore, we elucidated the previously unrecognized molecular mechanisms through which HPGD suppresses ESCC progression and its potential as a novel therapeutic target.</p> Methods <p>Transcriptome sequencing was performed on paired tumor and adjacent normal tissues from deceased patients with ESCC to identify differentially expressed genes. The differential expression of the HPGD gene was subsequently validated in two independent, large-scale ESCC patient cohorts, and its prognostic significance was evaluated. To evaluate the functional role of HPGD in ESCC, the enzyme was overexpressed in ESCC cell lines, and a series of in vitro assays were conducted to assess its effects on proliferation, apoptosis, invasion, and migration. To elucidate the molecular mechanisms underlying the effects of HPGD, we performed transcriptomic sequencing to profile gene expression changes in ESCC cells. Through multiple analyses, including measurements of lipid peroxidation, intracellular ferrous ion and reactive oxygen species (ROS) levels, dual-fluorescence flow cytometry for autophagy, phosphoprotein microarrays, biotin pull-down assays, and chromatin immunoprecipitation (ChIP), we demonstrated that HPGD regulates the malignant phenotype of ESCC cells primarily by inducing ferroptosis and autophagy. Finally, the impact of HPGD on ESCC tumor growth was validated in vivo using a subcutaneous xenograft model in nude mice.</p> Results <p>HPGD expression was significantly lower in ESCC tissues than in normal tissues and was negatively correlated with tumor cell differentiation and patient outcomes. HPGD overexpression inhibited ESCC cell proliferation, invasion, and migration in vitro and in xenograft tumor growth in vivo. In vitro experiments demonstrated that HPGD suppresses ERK1/2 activation by facilitating lipoxin A4 (LXA4) degradation. This inhibition facilitates binding of the RNA-binding protein U2AF2 to the promoter region of the transferrin receptor (TFRC), thereby increasing TFRC expression. Consequently, these alterations lead to intracellular iron accumulation and initiate ferroptosis. Excessive generation of ROS during ferroptosis results in hyperactivation of autophagy via the AMPK/mTOR signaling pathway. Mitigating the HPGD-induced upregulation of TFRC or reducing ROS production effectively reverses ferroptosis, prevents excessive autophagy, and ameliorates malignant cell phenotypes.</p> Conclusions <p>HPGD exerts its antitumor effects by promoting ferroptosis through the LXA4-ERK1/2-U2AF2 signaling axis, which in turn induces autophagy hyperactivation via the AMPK-mTOR pathway. These findings suggest that HPGD is a promising therapeutic target for esophageal squamous cell carcinoma (ESCC) and reveal the nonclassic role of the RNA-binding protein U2AF2 in regulating the expression of TFRC by acting like a transcription factor.</p>