Abstract
BackgroundLewis y antigen is difucosylated oligosaccharide and is carried by glycoconjugates at cell surface. Elevated expression of Lewis y has been found in 75% of ovarian tumor, and the high expression level is correlated to the tumor's pathological staging and prognosis. This study was to investigate the effect and the possible mechanism of Lewis y on the proliferation of human ovarian cancer cells.MethodsWe constructed a plasmid encoding α1,2-fucosyltransferase (α1,2-FT) gene and then transfected it into ovarian carcinoma-derived RMG-I cells with lowest Lewis y antigen expression level. Effect of Lewis y on cell proliferation was assessed after transfection. Changes in cell survival and signal transduction were evaluated after α-L-fucosidase, anti-Lewis y antibody and phosphatidylinositol 3-kinase (PI3K) inhibitor treatment.ResultsOur results showed that the levels of α1,2-FT gene and Lewis y increased significantly after transfection. The cell proliferation of ovarian carcinoma-derived RMG-I cells sped up as the Lewis y antigen was increased. Both of α-L-fucosidase and anti-Lewis y antibody inhibited the cell proliferation. The phosphorylation level of Akt was apparently elevated in Lewis y-overexpressing cells and the inhibitor of PI3K, LY294002, dramatically inhibited the growth of Lewis y-overexpressing cells. In addition, the phosphorylation intensity and difference in phosphorylation intensity between cells with different expression of α1,2-FT were attenuated significantly by the monoantibody to Lewis y and by the PI3K inhibitor LY294002.ConclusionsIncreased expression of Lewis y antigen plays an important role in promoting cell proliferation through activating PI3K/Akt signaling pathway in ovarian carcinoma-derived RMG-I cells. Inhibition of Lewis y expression may provide a new therapeutic approach for Lewis y positive ovarian cancer.
Highlights
Lewis y antigen is difucosylated oligosaccharide and is carried by glycoconjugates at cell surface
Elevated expression of Lewis y has been found in 70-90% of the human carcinomas of epithelial cell origin, including breast, ovary, prostate, colon cancers, and the high expression level is correlated to the tumor's pathological staging and prognosis [4,5,6]
The following reagents were purchased from commercial sources: expression vector pcDNA3.1(-) and a TA cloning kit from Invitrogen (San Diego, CA, USA); E. coli JM109 from Toyobo (Tokyo, Japan); restriction endonucleases, BamHI, EcoRI, and G418 from Gibco; cell transfection and NucleoBond plasmid kits from GE Healthcare (Piscataway, NJ, USA); AmpliTaq GoldTM and a BigdyeTM terminator cycle sequencing ready reaction kit from Perkin-Elmer/Applied Biosystems (Foster City, CA, USA); DMEM and fetal bovine serum (FBS) from Hyclone (Logan, UT, USA); trypsin, ethylenediamine tetraacetic acid (EDTA), dimethyl sulfoxide (DMSO) and 3-(4,5dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) from Amresco (Solon, OH, USA); SABC test kit from Boshide Biotech Co (Wuhan, China); α-L-fucosidase and methylene blue from Sigma
Summary
Lewis y antigen is difucosylated oligosaccharide and is carried by glycoconjugates at cell surface. Elevated expression of Lewis y has been found in 75% of ovarian tumor, and the high expression level is correlated to the tumor's pathological staging and prognosis. Lewis y antigen is carried by glycoconjugates (glycoproteins and glycolipids) at cell surface. It is an oligosaccharide with two fucoses, and its chemical structure is Fucα1 → 2Galβ1 → 4 [Fucα1 → 3]GlcNAcβ1 → R, belonging to the A, B, H, Lewis blood group antigens family with specific fucosylation of the terminal end of carbohydrate structure catalyzed by the α1,2-fucosyltransferase [1,2]. Elevated expression of Lewis y has been found in 70-90% of the human carcinomas of epithelial cell origin, including breast, ovary, prostate, colon cancers, and the high expression level is correlated to the tumor's pathological staging and prognosis [4,5,6]. It has been reported that the Lewis y antigen was expressed on a number of different molecular carriers, including 2 major ovarian cancer antigens (CA125 and MUC-1), suggesting the high incidence of Lewis y in ovarian cancer [7]
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