title: Fast epitope mapping for the anti-MUC1 monoclonal antibody by combining a one-bead-one-glycopeptide library and a microarray platform.
abstract: Anti-MUC1 monoclonal antibodies (mAbs) are powerful tools that can be used to recognize cancer-related MUC1 molecules, the O-glycosylation status of which is believed to affect binding affinity. We demonstrate the feasibility of using a rapid screening methodology to elucidate those effects. The approach involves i) "one-bead-one-compound"-based preparation of bilayer resins carrying glycopeptides on the shell and mass-tag tripeptides coding O-glycan patterns in the core, ii) on-resin screening with an anti-MUC1 mAb, iii) separating positive resins by utilizing secondary antibody conjugation with magnetic beads, and (iv) decoding the mass-tag that is detached from the positive resins pool by using mass spectrometric analysis. We tested a small library consisting of 27 MUC1 glycopeptides with different O-glycosylations against anti-MUC1 mAb clone VU-3C6. Qualitative mass-tag analysis showed that increasing the number of glycans leads to an increase in the binding affinity. Six glycopeptides selected from the library were validated by using a microarray-based assay. Our screening provides valuable information on O-glycosylations of epitopes leading to high affinity with mAb.
authors.affiliation: Faculty of Advanced Life Science and Graduate School of Life Science, Hokkaido University, N21, W11, Kita-ku, Sapporo 001-0021 (Japan), Fax: (+81) 11-706-9042.
authors.forename: Fayna
authors.initials: F
authors.lastname: Garcia-Martin
authors.forename: Takahiko
authors.initials: T
authors.lastname: Matsushita
authors.forename: Hiroshi
authors.initials: H
authors.lastname: Hinou
authors.forename: Shin-Ichiro
authors.initials: S
authors.lastname: Nishimura
mesh_terms: D000595:Amino Acid Sequence; D000911:Antibodies, Monoclonal / Q000737:chemistry / Q000276:immunology; D020650:Combinatorial Chemistry Techniques; D018604:Epitope Mapping; D000939:Epitopes / Q000276:immunology*; D006020:Glycopeptides / Q000737:chemistry*; D006031:Glycosylation; D006801:Humans; D018396:Mucin-1 / Q000737:chemistry* / Q000276:immunology; D010446:Peptide Fragments / Q000737:chemistry* / Q000276:immunology; D019151:Peptide Library; D060327:Solid-Phase Synthesis Techniques
chemical_list: D000911:Antibodies, Monoclonal; D000939:Epitopes; D006020:Glycopeptides; D018396:Mucin-1; D010446:Peptide Fragments; D019151:Peptide Library
keywords: antibodies; combinatorial chemistry; glycopeptides; microarrays; solid-phase synthesis