TOPK (T–lymphokine-activated killer cell–originated protein kinase) is a protein found in a wide range of human cancers and plays a vital role in the mitosis of cancer cells. OTS964 inhibited TOPK kinase activity with an IC50 value of 28 nM and maintained its specificity against TOPK. OTS964 exhibited excellent in vivo efficacy with a liposomal formulation and caused complete regression of tumors without any detectable toxicity in LU-99 lung cancer xenograft models. Tumor analysis revealed that treatment with the liposomal formulation of OTS964 effectively induced apoptosis in cancer cells. These results suggest that the liposomal OTS964 may be a promising molecular-targeted therapy, which could be applied to treat cancer patients whose tumors express high amounts of TOPK.