Liver hepatocellular carcinoma (LIHC) is one of the most common and aggressive malignancies worldwide, with high mortality rates. PLOD family genes (PLOD1, PLOD2, and PLOD3) play a pivotal role in extracellular matrix (ECM) remodeling, which contributes to cancer progression and metastasis. This study explores the diagnostic, prognostic, and therapeutic potential of PLOD1, PLOD2, and PLOD3 in LIHC.

Seven LIHC cell lines and five normal liver tissue cell lines were cultured. RT-qPCR was used to assess the expression of PLOD genes. The GSCA and TCGA databases were used for gene expression, pathway analysis, survival, and enrichment analysis. Promoter methylation was analyzed via GSCA and OncoDB, and mutational and CNV analyses were conducted using the cBioPortal database. The KM plotter tool assessed the prognostic significance of PLOD genes, while miRNA-mRNA network analysis was performed using the mirNET and UALCAN databases. Functional assays, including siRNA-mediated knockdown, Western blotting, cell proliferation, colony formation, and wound healing assays, were used to assess the biological roles of PLOD2 and PLOD3 in LIHC.

The expression of PLOD1, PLOD2, and PLOD3 was significantly (p-value < 0.05) higher in LIHC cell lines compared to normal controls. Validation in the TCGA cohort confirmed upregulation of PLOD genes in LIHC tumors, with PLOD2 and PLOD3 showing progressively higher expression across pathological stages. PLOD genes were associated with the activation of oncogenic pathways such as epithelial-to-mesenchymal transition (EMT) and poor survival outcomes. Promoter methylation analysis revealed lower methylation in tumor samples. Kaplan-Meier survival analysis indicated that higher PLOD expression correlated with poorer overall survival (OS) and relapse-free survival (RFS) in LIHC patients. The miRNA-mRNA network identified three miRNAs (hsa-miR-503-5p, hsa-miR-195-5p, and hsa-miR-193a-3p) regulating PLOD genes, with aberrant miRNA expression linked to poor prognosis. PLOD2/3 knockdown in HepG2 and Hep3B cells significantly reduced cell proliferation, colony formation, and migration, highlighting their critical roles in tumor progression.

Our study demonstrates that PLOD1, PLOD2, and PLOD3 are significantly upregulated in LIHC and correlate with poor prognosis. The PLOD genes may serve as valuable biomarkers for diagnosis and prognosis in LIHC. Moreover, targeting PLOD genes could provide new therapeutic strategies to hinder tumor progression and metastasis in liver cancer.