Identification of EFNA family as new potential prognostic biomarkers correlated with immune cell infiltration in hepatocellular carcinoma.

BACKGROUND

Accumulating evidence has revealed that the EFNA family contributes to the initiation and development of multiple malignancies. However, the roles of EFNAs in gene expression, function, clinical relevance, and prognosis in hepatocellular carcinoma (HCC) are still not well understood. The aim of this study is to investigate the relationship between EFNAs expression and clinical outcomes and immune infiltration in HCC patients.

METHODS

We utilized several public databases, including GEPIA, UALCAN, The Cancer Genome Atlas (TCGA), Gene Expression Omnibus (GEO), International Cancer Genome Consortium (ICGC), Human Protein Atlas (HPA), Kaplan-Meier plotter, cBioPortal, LinkedOmics, Metascape, and Tumor Immune Estimation Resource (TIMER), alongside the Cell database, to conduct bioinformatics analysis and explore the relationships between each EFNA family member and HCC. Immunofluorescence was utilized to validate the expression level of EFNA family members in HCC clinical tissues.

RESULTS

EFNA1/3/4 messenger RNAs (mRNAs) and protein expression levels were considerably upregulated in HCC, while the expression levels of EFNA5 mRNA and protein were significantly reduced in HCC compared to normal liver tissue. Overexpression of EFNA1/3/4/5 was closely related to higher tumor stages and poorer tumor grades. Receiver operating characteristic (ROC) analysis showed strong diagnostic performance for EFNA1 [area under the curve (AUC) =0.885], EFNA3 (AUC =0.928), and EFNA4 (AUC =0.963), outperforming alpha-fetoprotein (AFP) in several tumor (T) stages. Higher EFNA1/3/4/5 expression correlated with worse overall survival [GEPIA hazard ratio (HR) range 1.7-2.6; Kaplan-Meier Plotter HR range 1.57-2.31], and EFNA1/4/5 remained independent prognostic factors in multivariable Cox models (P<0.05). Additionally, a genetically altered rate of up to 50% of EFNA genes has been found in HCC patients. We analyzed EFNA and its 50 frequently altered adjacent genes in HCC for functional prediction and pathway enrichment. The results indicated that these genes were primarily involved in DNA-binding transcription factors, the PI3K-Akt signaling pathway, and pathways related to HCC. We revealed that the expressions of EFNA family members were remarkably linked with immune cell infiltration, suggesting that EFNA family members may reflect immune status. Finally, we further verified the expression of EFNA family members in HCC samples and found the results to be consistent with the databases.

CONCLUSIONS

Our findings suggest that EFNA1, EFNA3, EFNA4, and EFNA5 could serve as potential biomarkers for prognosis and promising targets for precision therapy in HCC patients.