〔Abstract〕 Objective To investigate the role of Disco Interacting Protein 2 Homolog B (DIP2B) in the proliferation, migration, and invasion processes of gastric cancer cells. Methods Bioinformatic analysis was performed using public datasets, including The Cancer Genome Atlas (TCGA), GEPIA and TIMER2.0, to assess DIP2B expression and its clinical prognostic value in gastric cancer. DIP2B expression levels in gastric cancer cell lines were validated by Western blot and RT-qPCR. ADIP2B knockdown cell model was established via siRNA mediated gene silencing, and the knockdown efficiency was confirmed. Cell proliferation was assessed using real-time cellular analysis (RTCA) and colony formation assays. Cell migration and invasion were evaluated by Transwell assays, while flow cytometry was employed to detect changes in apoptosis and cell cycle distribution. Results DIP2B was significantly upregulated in gastric cancer cells and correlated with poor prognosis in patients (P<0.05). The experimental results revealed that DIP2B knockdown markedly suppressed the proliferation, migration, and invasion of gastric cancer cells, while promoting apoptosis and inducing cell cycle arrest (P<0.05). Conclusion DIP2B is highly expressed in gastric cancer and associated with unfavorable prognosis. Knockdown of DIP2B inhibits the proliferation, migration, and invasion of gastric cancer cells and promotes apoptosis, suggesting that DIP2B may serve as a potential prognostic biomarker and therapeutic target for gastric cancer.