Glycosylations versus conformational preferences of cancer associated mucin core.
Synthetic oligosaccharide vaccines based on core STn (sialyl alpha2-6 GalNAc) carbohydrate epitopes are being evaluated by a number of biopharmaceutical firms as potential immunotherapeutics in the treatment of mucin-expressing adenocarcinomas. The STn carbohydrate epitopes exist as discontinuous clusters, O-linked to proximal serine and threonine residues within the mucin sequence. In an effort to probe the structure and dynamics of STn carbohydrate clusters as they may exist on the cancer-associated mucin, we have used NMR spectroscopy and MD simulations to study the effect of O-glycosylation of adjacent serine residues in a repeating (Ser)n sequence. Three model peptides/glyco-peptides were studied: a serine trimer containing no carbohydrate groups ((Ser)3 trimer); a serine trimer containing three Tn (GalNAc) carbohydrates alpha-linked to the hydroxyls of adjacent serine sidechains ((Ser.Tn)3 trimer); and a serine trimer containing three STn carbohydrates alpha-linked to the hydroxyls of adjacent serine sidechains ((Ser.STn)3 trimer). Our results demonstrate that clustering of carbohydrates shifts the conformational equilibrium of the underlying peptide backbone into a more extended and rigid state, an arrangement that could function to optimally present the clustered carbohydrate antigen to the immune system. Steric effects appear to drive these changes since an increase in the size of the attached carbohydrate (STn versus Tn) is accompanied by a stronger shift in the equilibrium toward the extended state. In addition, NMR evidence points to the formation of hydrogen bonds between the peptide backbone NH protons and the proximal GalNAc groups in the (Ser.Tn)3 and (Ser.STn)3 trimers. The putative peptide-sugar hydrogen bonds may also play a role in influencing the conformation of the underlying peptide backbone, as well as the orientation of the O-linked carbohydrate. The significance of these results will be discussed within the framework of developing clustered STn-based vaccines, capable of targeting the clustered STn epitopes on the cancer-associated mucin.
Schuman
J
J
Department of Medicinal Chemistry, School of Pharmacy, University of Washington, Seattle, WA 98195, USA.
Qiu
D
D
Koganty
R R
RR
Longenecker
B M
BM
Campbell
A P
AP
D000230:Adenocarcinoma / Q000737:chemistry* / Q000276:immunology / Q000628:therapy; D019496:Cancer Vaccines / Q000737:chemistry / Q000276:immunology; D000939:Epitopes / Q000737:chemistry / Q000276:immunology / Q000378:metabolism; D006031:Glycosylation; D006860:Hydrogen Bonding; D009682:Magnetic Resonance Spectroscopy; D008958:Models, Molecular; D008968:Molecular Conformation; D009077:Mucins / Q000737:chemistry* / Q000276:immunology / Q000378:metabolism*; D012694:Serine / Q000737:chemistry / Q000378:metabolism; D012997:Solvents; D013696:Temperature; D013816:Thermodynamics; D014867:Water / Q000737:chemistry / Q000378:metabolism
D019496:Cancer Vaccines; D000939:Epitopes; D009077:Mucins; D012997:Solvents; D014867:Water; D012694:Serine