MONOCLONAL-ANTIBODY LU-BCRU-G7 AGAINST A BREAST TUMOR-ASSOCIATED GLYCOPROTEIN RECOGNIZES THE DISACCHARIDE GAL-BETA-1-3GLCNAC

The monoclonal antibody LU-BCRU-G7, that was generated by in vitro imm unization, shows clinical value as a prognostic marker in early stage breast carcinoma, It has now been characterized with regard to its bin ding epitope, Using a recently described method based on the construct ion of N-substituted polyacrylamide (PAA) derivatives of carbohydrates (pseudopolysaccharides), the structure of the epitope for the monoclo nal antibody LU-BCRU-G7 has been determined, Competitive binding assay s and inhibitory enzyme-linked inmunosorbent assays (ELISAs) using the se pseudopolysaccharides have shown the LU-BCRU-G7 epitope to be a dis accharide Gal beta 1-3GlcNAc (Le(c); where Gal is D-galactose, Glc is D-glucose and GlcNAc is N-acetyl-D-glucosamine). Both galactose and N- acetyl glucosamine moieties are essential for binding, Substitution on C-2 or C-3 of the terminal galactose abolished binding, as did galact ose-alpha terminated oligosaccharides, The galactose moiety alone, as expressed by the Gal beta-PAA conjugate, appeared to be a more importa nt feature of the epitope than the GlcNAc-PAA conjugate, which failed to bind or inhibit the LU-BCRU-G7 antibody, In the N-acetyl glucosamin e moiety, binding was decreased but not eliminated by fucose substitut ion, as in Le(a), or change in configuration of C-4, as in Gal beta 1- 3GlcNAc. Omission of the NAc group resulted in complete loss of activi ty, The tetrasaccharide lacto-N-tetraose, although containing the term inal Le(c) disaccharide, does not react with the antibody, suggesting conformational interference of the binding site, These findings show t hat the monoclonal antibody LU-BCRU-G7 recognizes a terminal isolactos amine fragment on a tumour-associated glycoprotein, which we have prev iously shown to be inversely related to survival in breast cancer.