Poster Presentation HUPO 2019 - 18th Human Proteome Organization World Congress

MUC1 glycopeptides by chemoenzymatic synthesis revealed distinct specificities of anti-MUC1 antibodies (#472)

Yasunori Chiba 1 , Yayoi Yoshimura 1 2 , Kaori Denda-Nagai 3 , Yoshie Takahashi 1 , Izuru Nagashima 1 , Hiroki Shimizu 1 , Toshimitsu Kishimoto 2 , Miki Noji 3 , Tatsuro Irimura 3
  1. National Institute of Advanced Industrial Science and Technology, Tsukuba, IBARAKI, Japan
  2. Japan Bioindustry Association, Tokyo, Japan
  3. Graduate School of Medicine, Juntendo University, Tokyo, Japan

Mucin 1 (MUC1) has been known as a carcinoma-associated mucin-like glycoprotein and anti-MUC1 antibodies have been used as serum markers for tumor burden to monitor therapeutic outcomes of breast cancer and as a serum marker for lung fibrosis. Though some of the anti-MUC1 antibodies were already investigated, the specificities of many anti-MUC1 monoclonal antibodies, particularly their specificities toward glycoforms at different positions remain unclear. In this study, we produced a total of 20 glycopeptides representing a unit of the tandem repeat domain of MUC1 with GalNAc (Tn-antigen), Galβ1-3GalNAc (T-antigen), NeuAcα2-3Galβ1-3GalNAc (sialyl T-antigen), or NeuAcα2-6GalNAc (sialyl Tn-antigen) at each threonine or serine residue by chemoenzymatic synthesis to analyze their capacity to bind 13 monoclonal antibodies specific for MUC1. First, glycopeptides with a GalNAc residue at each one of the five possible O-glycosylation sites were chemically synthesized and then extended with Gal and/or NeuAc residues using glycosyltransferases with donor sugar nucleotides. In the case of synthesis of NeuAcα2-6GalNAc attached to the serine at position 9 or 19 of the peptide, Galβ1-3(NeuAcα2-6)GalNAc was prepared and then β1-3,4 galactosidase was used to remove galactose. Next, the binding capacity of 13 antibodies for the glycopeptides was analyzed by Enzyme-Linked ImmunoSorbent Assay. The results indicated that anti-MUC1 monoclonal antibodies can be classified into several subgroups based on their specificity toward different glycopeptides having specific glycan structures and glycosylation sites.  Considering that some of these anti-MUC1 antibodies were developed to aid cancer diagnosis and therapy, the present results are potentially useful as the basis for further clinical use of various anti-MUC1 antibodies.