Changes in oligosaccharide structures alter the biological functions of cancer cells. α1-6 fucosyltransferase (α1-6FucT) catalyzes the transfer of fucose to the innermost GlcNAc in N-glycans. Although α1-6FucT is barely detected in normal liver, it is enhanced during rat hepatocarcinogenesis and in human hepatoma. To understand the biological meaning of the α1-6FucT in hepatoma, especially in terms of metastasis, we established human hepatoma cell lines, which express high levels of α1-6FucT by transfection of the α1-6FucT gene and investigated intrahepatic metastasis after splenic injection to athymic mice. Tumor formation in the liver was dramatically suppressed in the α1-6FucT transfectants (1 of 9 and 1 of 10 in α1-6FucT transfectants versus 6 of 9 and 6 of 9 in controls). Although there were no differences in terms of cell invasiveness to a Matrigel or in terms of cytotoxicity to interleukin 2-treated lymphocytes between α1-6FucT transfectants and control cells, cell adhesion to mice hepatocytes and nonparenchymal liver cells in culture was significantly inhibited in α1-6FucT transfectants, compared to the controls. Attachment of α1-6FucT transfectants to a fibronectin-coated dish was decreased compared to controls because α5β1 integrin was more strongly α1-6 fucosylated in the α1-6FucT transfectants. Two-dimensional electrophoresis followed by lectin blot showed that certain glycoproteins (M_r 50,000–150,000, pI 4.8–5.5) were α1-6 fucosylated and might be linked to suppression of intrahepatic metastasis. This is the first demonstration of the biological significance of α1-6 fucosylation on N-glycans in hepatoma cells under in vivo conditions.