Hyperplasia suppressor gene (HSG) is a novel gene that markedly suppresses the mitogenetic stimuli or injury mediated by vascular smooth muscle cell proliferation. Herein we provide experimental evidence to show that HSG can also play a key role in tumor proliferation. Down-regulation of HSG protein in hepatocellular carcinoma tissues compared to adjacent tissues. Overexpression of HSG suppressed the growth of liver cancer cell lines, resulted in cell cycle arrest in the G0/G1 phase, increased expression of the cyclin dependent kinase inhibitors (CKIs), and reduced expression of proliferating cell nuclear antigen (PCNA). It also showed that adenovirus-mediated HSG overexpression induced apoptosis. Up-regulation of HSG by adenovirus also significantly suppressed the growth of subcutaneous tumors in nude mice both ex vivo and in vivo. Collectively, our data suggest that HSG is a potential therapy for tumors and possibly other proliferative diseases as well and it has antitumor efficacy on hepatocellular carcinoma by using adenovirus vectors, which may be a new therapeutic target for liver cancer prevention.