The Forkhead Box M1 (FOXM1) transcription factor has been considered as a potential target for the prevention and/or therapeutic intervention in human carcinomas because of its roles in tumorigenesis and tumor progression through regulating the expression of genes relevant to cell proliferation and transformation. In this study, FOXM1 was found to express strongly in both clinical tissue specimens and human hepatocellular carcinoma (HCC) cell lines such as Huh-6, Huh-7 and HepG2. The knockdown of FOXM1 expression through an adenovirus vector (named AdFOXM1shRNA), which expresses a short hairpin RNA to downregulate FOXM1 expression specifically, diminished the proliferation of Huh-7 and HepG2 cells and anchorage-independent growth of Huh-7 cells. Furthermore, we assessed the efficacy of AdFOXM1shRNA for tumor gene therapy with the Huh-7 cell xenograft mouse model and found that the tumor growth was significantly suppressed when inoculated mice were injected with AdFOXM1shRNA in the tumors. Together, our results suggest that FOXM1 is a potential therapeutic target for HCC and AdFOXM1shRNA may be an additional gene therapeutic intervention for HCC treatment.