A combined computational-experimental approach to define the structural origin of antibody recognition of sialyl-Tn, a tumor-associated carbohydrate antigen

Scientific Reports
Ron AmonVered Padler-Karavani

Abstract

Anti-carbohydrate monoclonal antibodies (mAbs) hold great promise as cancer therapeutics and diagnostics. However, their specificity can be mixed, and detailed characterization is problematic, because antibody-glycan complexes are challenging to crystallize. Here, we developed a generalizable approach employing high-throughput techniques for characterizing the structure and specificity of such mAbs, and applied it to the mAb TKH2 developed against the tumor-associated carbohydrate antigen sialyl-Tn (STn). The mAb specificity was defined by apparent KD values determined by quantitative glycan microarray screening. Key residues in the antibody combining site were identified by site-directed mutagenesis, and the glycan-antigen contact surface was defined using saturation transfer difference NMR (STD-NMR). These features were then employed as metrics for selecting the optimal 3D-model of the antibody-glycan complex, out of thousands plausible options generated by automated docking and molecular dynamics simulation. STn-specificity was further validated by computationally screening of the selected antibody 3D-model against the human sialyl-Tn-glycome. This computational-experimental approach would allow rational design of potent ant...Continue Reading

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Citations

Sep 27, 2019·Frontiers in Immunology·Anoopjit Singh KoonerXi Chen
Dec 6, 2020·Annales d'endocrinologie·Nathalie GilliéronUrs E Nydegger
Jun 14, 2021·Journal of Molecular Biology·Aliza Borenstein-KatzRon Diskin

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Datasets Mentioned

BETA
M35669.1

Methods Mentioned

BETA
glycosylation
PCR
PCRs

Software Mentioned

IgBLAST
Mestrenova
NEBaseChanger
Carb
Web
ClustalW2
PIGS
Genepix
Autodock Vina
GLYCAM

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