Novel O-glycan arrays to characterize human cancer-associated epithelial antigens

Abstract

Aberrantly expressed carbohydrates occur on cancer cells and are antigenic. This thesis is focused on studies to elucidate two elusive hybridoma-defined, cancer-associated carbohydrate antigens: a prostate cancer-associated antigen F77 and a broadly distributed epithelial cancer-associated antigen AE3. The key experiments performed are microarray analyses with mucin-type glycoproteins and generation of designer arrays, multidimensional chromatographies and mass spectrometry of O-glycomes. Antigen-positive sequences assigned are corroborated with focused arrays of natural and chemically synthetized oligosaccharides or glycolipids and products of glycosidase treatments. The F77 antigen is assigned as blood group H type 2 on a 6-linked branch of a poly-N-acetyllactosamine backbone (structure a). This sequence is shown to occur on O-glycans and on glycolipids. The F77 antibody can bind, with lower intensities, to the blood group A and B analogues of structure a. The minimum F77 antigenic sequence is shown to be a pentasaccharide (underlined in structure a). The close association of F77 antigen with prostate cancer is proposed to be a consequence of up-regulation of branching enzymes together with persistent expression of H antigen. This may account for the prevalence of F77 antigen in prostate cancers irrespective of the patients’ ABO blood group status. [Scheme appears here. To view, please open pdf attachment]. There appear to be two distinct forms of AE3 antigen as evidenced by the ability of AE3 antibody to bind: (i) the O-glycan structure b, known as T antigen, which is joined by α-linkage to serine on mucin-type glycoproteins, and (ii) a sulphated glycolipid known as SM1a (structure c) in which “T” sequence is joined by β-linkage to galactose. An O-glycan analogue of structure c (structure d), has not been reported so far. Chemo-enzymatic synthesis of structure d will shortly be attempted for antigenic analysis. [Scheme appears here. To view, please open pdf attachment]. With knowledge of details of these two antigens, the biosynthetic pathways, the biological functions and clinical applications can now be rationally pursued.