Synthesis and antibody recognition of synthetic antigens from MUC1

Research output: Contribution to journalArticle

3 Citations (Scopus)

Abstract

In the altered form of MUC1 mucin associated with breast cancer, the highly immunogenic sequence PDTRPAP is exposed, and may be an immunologically relevant target for the development of diagnostics or cancer immunotherapy. In this study, we report the preparation and antibody binding properties of monomeric and dimeric MUC1 peptides containing the epitope region recognized by monoclonal antibody (mAb) C595. Peptides contained a single or two copies of the whole 20-mer repeat unit (VTSAPDTRPAPGSTAPPAHG) of MUC1 protein. MUC1 40-mer peptides were prepared by the condensation of semi-protected fragments of the repeat unit, in solution or by chemical ligation. In the first case, cyclohexyl-type protecting groups were used for the synthesis of semi-protected fragments by the Boc/Bzl strategy. Unprotected fragments were used in the chemical ligation to produce thioether linkages. In one of the fragments, a Gly residue was replaced by Cys at the C-terminus and the other fragment was chloroacetylated at the N-terminus. In addition, the short peptide APDTRPAPG, and its disulfide dimer, (APDTRPAPGC)2 were produced. The antibody binding properties of these MUC1 peptide constructs were tested by competition enzyme-linked immunosorbent assay (ELISA). The short epitope region peptide, APDTRPAPG and its dimer (APDTRPAPGC)2 showed higher IC50 values (IC50 = 56.3 and 53.2 μmol/l, respectively). While the 20-mer peptide (IC50 = 25.9 μmol/l) and more markedly its 40-mer dimers (IC50 = 0.62 and 0.78 μmol/l) were recognized better. CD data obtained in water or in TFE indicated no significant conformational differences between the 20-mer and 40-mer peptides. We found a high level of similarity between the binding properties of the 40-mer peptides with amide or thioether links, providing a new possibility to build up oligomeric MUC1 peptides by thioether bond formation.

Original languageEnglish
Pages (from-to)610-616
Number of pages7
JournalJournal of Peptide Science
Volume14
Issue number5
DOIs
Publication statusPublished - May 2008

Fingerprint

Synthetic Vaccines
Peptides
Antibodies
Inhibitory Concentration 50
Sulfides
Dimers
Ligation
Epitopes
Immunosorbents
Polytetrafluoroethylene
Mucins
Amides
Disulfides
Immunotherapy
Condensation
Assays
Enzyme-Linked Immunosorbent Assay
Monoclonal Antibodies
Breast Neoplasms

Keywords

  • Antibody binding
  • Antibody recognition of mucin 1 tandem repeat
  • Chemical ligation
  • Mucin 1 epitope region
  • Solution conformation
  • Synthetic antigen

ASJC Scopus subject areas

  • Analytical Chemistry
  • Biochemistry, Genetics and Molecular Biology(all)
  • Biochemistry

Cite this

Synthesis and antibody recognition of synthetic antigens from MUC1. / Bánóczi, Z.; Mező, G.; Windberg, Emoke; Uray, K.; Majer, Zs.; Hudecz, F.

In: Journal of Peptide Science, Vol. 14, No. 5, 05.2008, p. 610-616.

Research output: Contribution to journalArticle

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AU - Hudecz, F.

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AB - In the altered form of MUC1 mucin associated with breast cancer, the highly immunogenic sequence PDTRPAP is exposed, and may be an immunologically relevant target for the development of diagnostics or cancer immunotherapy. In this study, we report the preparation and antibody binding properties of monomeric and dimeric MUC1 peptides containing the epitope region recognized by monoclonal antibody (mAb) C595. Peptides contained a single or two copies of the whole 20-mer repeat unit (VTSAPDTRPAPGSTAPPAHG) of MUC1 protein. MUC1 40-mer peptides were prepared by the condensation of semi-protected fragments of the repeat unit, in solution or by chemical ligation. In the first case, cyclohexyl-type protecting groups were used for the synthesis of semi-protected fragments by the Boc/Bzl strategy. Unprotected fragments were used in the chemical ligation to produce thioether linkages. In one of the fragments, a Gly residue was replaced by Cys at the C-terminus and the other fragment was chloroacetylated at the N-terminus. In addition, the short peptide APDTRPAPG, and its disulfide dimer, (APDTRPAPGC)2 were produced. The antibody binding properties of these MUC1 peptide constructs were tested by competition enzyme-linked immunosorbent assay (ELISA). The short epitope region peptide, APDTRPAPG and its dimer (APDTRPAPGC)2 showed higher IC50 values (IC50 = 56.3 and 53.2 μmol/l, respectively). While the 20-mer peptide (IC50 = 25.9 μmol/l) and more markedly its 40-mer dimers (IC50 = 0.62 and 0.78 μmol/l) were recognized better. CD data obtained in water or in TFE indicated no significant conformational differences between the 20-mer and 40-mer peptides. We found a high level of similarity between the binding properties of the 40-mer peptides with amide or thioether links, providing a new possibility to build up oligomeric MUC1 peptides by thioether bond formation.

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KW - Chemical ligation

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KW - Solution conformation

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