Identification of EFNA3 as candidate prognosis marker and potential therapeutic target for adrenocortical carcinoma

Adrenocortical carcinoma (ACC) is a rare, but highly aggressive endocrine malignancy with poor prognosis and limited treatment options. Identifying novel biomarkers and therapeutic targets is essential for improving patient outcomes. The present study aimed to systematically characterize ephrin-A3 (EFNA3) expression patterns, its prognostic and diagnostic value, and its functional role in ACC progression through multi-omics bioinformatics and <i>in vitro</i> validation. Transcriptomic, epigenetic and pharmacogenomic data were obtained from The Cancer Genome Atlas, Genotype-Tissue Expression, Genomics of Drug Sensitivity in Cancer, Cancer Therapeutics Response Portal and MethSurv databases. Expression, survival, immune infiltration, methylation and drug sensitivity analyses were conducted using the R software and online tools (GEPIA2, CIBERSORT and cBioPortal). competitive endogenous RNA (ceRNA) networks were constructed based on microRNA (miRNA)/long non-coding RNA (lncRNA) predictions. Functional assays, including CCK-8, flow cytometry, Transwell assays were performed on the ACC cell lines, SW-13 and NCI-H295R, to validate EFNA3 function. EFNA3 was significantly upregulated in numerous types of cancer and associated with poor prognosis. In ACC, upregulated EFNA3 was associated with a poor prognosis [Overall survival (OS), hazard ratio (HR)=3.14, 95% CI, 1.49-7.81; disease-specific survival, HR=4.27, 95% CI, 1.70-10.72; progression-free interval, HR=6.24, 95% CI, 2.94-13.23; P<0.05] and diagnostic efficiency (area under the curve=0.829, 95% CI, 0.760-0.897). EFNA3-mutated cases had significantly worse OS in ACC specifically (OS, HR=2.97, 95% CI, 1.12-7.90, P=0.029; disease-free survival, HR=8.65, 95% CI, 2.14-34.93, P=0.002). β-catenin (CTNNB1) was among most frequently co-mutated genes ACC with EFNA3 (P=4.6×10^-4). Genetic amplification and DNA methylation alterations were observed in the ACC cohort. EFNA3 expression negatively correlated with immune infiltration and positively correlated with several m6A/m5C regulators. ceRNA network analysis demonstrated key lncRNA-miRNA-EFNA3 axes. Drug sensitivity profiling indicated that EFNA3 expression was associated with statin and proteasome inhibitor responses. The co-expression of positively correlated gene enrichment results suggested that Wnt signaling pathway and β-catenin/T-cell factor complex may be involved in the progression of ACC mediated by EFNA3. Functionally, EFNA3 promoted ACC cell proliferation and migration <i>in vitro</i>. The present study demonstrated that EFNA3 acts as an oncogene in ACC and may contribute to tumor aggressiveness via β-catenin activation and glycolytic reprogramming, and thus may serve as a potential biomarker for prognosis, immunotherapy sensitivity and drug repurposing, particularly involving statins.