Gastric cancer (GC) is one of the most common malignancies worldwide; however, understanding of its development and carcinogenesis is currently limited. Centromere protein O (CENPO), is a newly discovered constitutive centromeric protein, associated with cell death. The expression of CENPO in human cancers, including GC, is currently unknown. The aim of the present study was to investigate the clinical association between CENPO and GC, and to elucidate the potential mechanisms of CENPO in the process of GC progression. The results demonstrated that CENPO was expressed at high levels in GC and was correlated with p‑TNM stage. In addition, CENPO was observed to be a marker of poor prognosis in patients with GC. Knockdown of CENPO contributed to GC cell growth inhibition and apoptosis induction. In addition, downregulation of CENPO expression suppressed GC cell growth in vivo. Furthermore, CENPO knockdown decreased ataxia telangiectasia mutated (ATM), cyclin D1 and c‑Jun expression, indicating that the ATM signaling pathway may be involved in CENPO‑mediated regulation of GC cell growth. In conclusion, increased CENPO expression may be associated with the aggressive tumor biology of GC and CENPO may present a novel therapeutic target and prognostic biomarker for patients with GC.