Two beta 4-N-acetylgalactosaminyltransferases (beta 4GalNAcTs), beta 4GalNAcT3 and beta 4GalNAcT4, have been shown to be involved in the synthesis of the GalNAc beta 1 -> 4GlcNAc (LacdiNAc) group expressed on the outer branches of N- and/or O-glycans, and only beta 4GalNAcT4 is expressed in human mammary gland. We found that the expression level of the LacdiNAc group decreases as human breast cancers progress. To investigate biological significances of this disaccharide in human breast cancers, we transfected the FLAG-tagged beta 4GalNAcT4 cDNA into MDA-MB-231 cells, and obtained several clones showing enhanced expression of the gene. Clones 1 and 2 showed 15 and 9 times more transcript than mock-transfected cells. The FLAG-beta 4GalNAcT4 protein and its product, the LacdiNAc group, were detected in clone 1 and 2 cells. No change was observed in their growth rates while significant decreases in colony forming and invasive abilities were observed for clone I and 2 cells. When clone 1 cells were transplanted subcutaneously into nude mice, no tumors were formed while tumors were formed with mock-transfected cells. These results indicate that the expression of the LacdiNAc group is quite important for the suppression of malignancies of the MDA-MB-231 cells. (C) 2015 Elsevier Inc. All rights reserved.