Adrenocortical carcinoma (ACC) is a rare disease associated with a poor prognosis. Furthermore, the underlying molecular mechanism of carcinogenesis is poorly understood, and prognostic prediction of ACC has low accuracy. In the present study, a bioinformatics approach was used to investigate the molecular mechanisms and prognosis of ACC. Samples of adrenal tumors were collected from patients undergoing adrenalectomy at the Department of Urology, the First Hospital of China Medical University. The analyzed gene datasets were downloaded from the Gene Expression Omnibus and The Cancer Genome Atlas (TCGA) database. Following this, the differentially expressed genes (DEGs) were included in Gene Ontology enrichment, and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway, protein‑protein interaction network and survival analyses. MTT colorimetric assays, colony formation assays and 5‑ethynyl‑20‑deoxyuridine incorporation assays were also conducted to evaluate ACC cell proliferation. The identified DEGs included 20 downregulated genes and 51 upregulated genes, which were highly associated with the cell cycle, organelle fission, chromosome segregation, cell division and spindle stability. The top 14 hub genes were subsequently confirmed by reverse transcription‑quantitative polymerase chain reaction in ACC and adrenocortical adenoma samples. It was identified that the nuclear division cycle 80, cyclin B2 and topoisomerase 2‑α may serve important roles in adrenocortical tumor development. Furthermore, these three genes predicted overall survival and recurrence‑free survival in patients with ACC from the TCGA cohort. The findings identified three novel genes that have important roles in carcinogenesis and in the prognostic prediction of ACC.