Nek2 (NIMA-related kinase 2) is involved in cell division and mitotic regulation by centrosome splitting. We previously reported that Nek2 depletion causes growth suppression and cell death in cholangiocarcinoma and breast cancer cells. In this report, we examine the effect of a combination treatment using Nek2 siRNA with the cytotoxic chemotherapeutic agent cisplatin (CDDP) on colorectal cancer. Nek2 was overexpressed in all colorectal cancer cell lines examined (HCT-15, DLD-1, Colo205, and Colo320). Nek2 short-interfering RNA (siRNA) resulted in the inhibition of cell proliferation and the induction of apoptosis in vitro. Nek2 siRNA suppressed tumor growth compared to control siRNA in a xenograft mouse model. To investigate the potential utility of Nek2 siRNA for clinical cancer therapy, we examine the effect of a combination treatment using Nek2 siRNA with CDDP on colorectal cancer. The combined administration of both Nek2 siRNA and CDDP inhibited cell proliferation and induced apoptotic cell death in vitro. Furthermore, the combined administration of both Nek2 siRNA and CDDP suppressed tumor growth compared to either the single administration of Nek2 siRNA or the combined administration of control siRNA and CDDP. Our results suggest that combination treatment using Nek2 siRNA and chemotherapeutic agents may be an effective therapeutic option for colorectal cancer.