Hepatocellular carcinoma (HCC) is a significant contributor to cancer-related mortality in the United States and worldwide. Current treatments are largely ineffective: single agent and combination chemotherapy regimens have low success rates in HCC patients. Therefore, surgery and liver transplantation remain the only potential curative options for HCC. However, surgery and transplantation are not viable options for most patients because of significant liver cirrhosis, invasion into extra-hepatic structures or dissemination to distant tissues. Therefore, it is imperative to understand the mechanisms that regulate HCC cell migration, invasion and metastasis. By gene expression profiling, we identified kruppel-like transcription factor 6 (KLF6) as a potential regulator of HCC cell migration. KLF6 is commonly inactivated, through a variety of mechanisms, in human cancers including HCC. Interestingly, decreased KLF6 mRNA levels are associated with vascular invasion in human HCC. Moreover, we found that KLF6 knockdown increases HCC cell migration in vitro, and deletion of a single Klf6 allele enhances tumor formation, increased metastasis to the lungs, and decreased survival in a HCC mouse model. Together, these data indicate that KLF6 suppresses HCC development and metastasis. To identify the potential mechanisms through which KLF6 regulates HCC cell migration, we combined gene expression profiling and chromatin immunoprecipitation coupled to deep sequencing (ChIP-Seq) to identify novel KLF6 transcriptional targets. Among the genes identified were VAV3 and CDC42EP3, both of which regulate the activity of RHO family GTPases. Indeed, RAC1 and CDC42 activity is increased in HCC cells following KLF6 knockdown, and RAC1 inhibition reduces HCC cell migration. Moreover, VAV3 knockdown reduces RAC1 activity and HCC cell migration. Together, our data identify a novel KLF6-VAV3-Rac1 axis that regulates HCC cell migration and dissemination.