Abstract
<title>Abstract</title> <p> Hepatocellular carcinoma (HCC) has a high recurrence rate and a poor prognosis. Although the transcription factor thymocyte selection-associated high mobility group box factor 3 (TOX3) has diverse roles in cancer, its function in HCC remains unclear. The present study defined TOX3’s mechanistic role and therapeutic potential in HCC progression. TOX3 expression and its correlation with prognosis were analyzed in clinical HCC cohorts. TOX3 expression was modulated using small interfering RNA-mediated knockdown and plasmid-mediated overexpression (OE) in HCC cell lines. Comprehensive <italic>in vitro</italic> functional studies (including proliferation, migration, invasion, apoptosis and cell cycle assays) were conducted. In vivo validation was performed through mouse xenograft models, and transcriptome sequencing was used to explore the molecular mechanism of TOX3 in HCC progression. Clinical analysis revealed that TOX3 was upregulated in human HCC tissues, and its elevated expression correlated negatively with patient prognosis. <italic>In vitro</italic> , TOX3 OE promoted proliferation, migration and invasion, and inhibited apoptosis in HCC cells; its knockdown induced G <sub>2</sub> /M arrest and reversed these effects. In vivo, TOX3 OE enhanced tumor growth in xenografts, while knockdown of TOX3 suppressed it. Transcriptome sequencing revealed that TOX3 OE downregulated the p53-p21-Rb pathway. In conclusion, the present study revealed that TOX3 functions as an oncogene in HCC, promoting tumor progression by inhibiting the p53-p21-Rb pathway, thereby disrupting cell cycle progression, and enhancing cell proliferation, migration and invasion. High TOX3 expression was correlated with poor prognosis. The present findings establish the role and mechanism of TOX3 in HCC progression, highlighting its potential as a novel prognostic biomarker and therapeutic target. </p>