Targeting the E2F6-TOP2A-DKK1 axis: a novel therapeutic strategy for EMT-driven hepatocellular carcinoma progression.

BACKGROUND: Hepatocellular carcinoma (HCC) has a poor prognosis, and identifying key driver genes and their molecular mechanisms is crucial for improving patient outcomes. While TOP2A is dysregulated in multiple cancers, its role and regulatory network in HCC remain incompletely understood.

METHODS: Integrated bioinformatics analyses were performed using TCGA, ICGC, and GEO datasets. TOP2A expression and prognostic value were validated in 120 clinical samples via immunohistochemistry. Functional assays (including CCK-8, Transwell, and wound healing in vitro, as well as xenograft tumor models in vivo) were conducted to assess phenotypic effects. Mechanisms were explored using ChIP-PCR, WB, and RT-qPCR. Prognostic models were developed using multivariate regression and evaluated using calibration curves, ROC, and DCA.

RESULTS: TOP2A was significantly upregulated in HCC tissues, with an AUC of 0.935 for diagnosis, and high expression correlated with advanced stage and poor survival (p<0.05). A prognostic model incorporating TOP2A expression, TNM stage, and tumor grade showed robust predictive accuracy for 1-, 3-, and 5-year survival (AUCs: 0.77, 0.85, 0.75). Knockdown of TOP2A suppressed proliferation, migration, and invasion. Mechanistically, E2F6 transcriptionally activated TOP2A by binding to its promoter, while TOP2A promoted EMT by upregulating DKK1 and activating β-catenin signaling. High TOP2A expression predicted poor response to TACE, sorafenib, and immunotherapy. The candidate targeted agent, A-443654, alone or combined with anti-PD-L1, potently suppressed tumor growth in vivo.

CONCLUSION: This study delineates the E2F6-TOP2A-DKK1 axis as a key mechanism in HCC progression. TOP2A serves as a diagnostic and prognostic biomarker, and targeting this pathway offers a promising therapeutic strategy for HCC.