Journal article
Characterization of Binding Epitopes of CA125 Monoclonal Antibodies
The most used cancer serum biomarker is the CA125 immunoassay for ovarian cancer that detects the mucin glycoprotein MUC16. Several monoclonal antibodies (mAbs) including OC125 and M11 are used in CA125 assays. However, despite considerable efforts, our knowledge of the molecular characteristics of the recognized epitopes and the role played by glycosylation has remained elusive.
Here a comprehensive set of recombinant MUG 16 tandem repeats (TRs) expressed in glycoengineered mammalian cells and E. coil, together with overlapping peptides, was used to probe antigen-binding epitopes. We present a complete analysis of N- and O-glycosylation sites of a MUC16 TR expressed in CHO cells and demonstrate that neither N- nor O-glycosylation appear to substantially influence binding of OC125 and M11 mAbs.
A series of successive N- and C-terminal truncations of a MUC16 TR construct expressed in E. coli narrowed down the epitopes for OC125 and M11 to a segment containing parts of two consecutive SEA domains with a linker. Thus, a complete SEA domain is not required. These findings suggest that binding epitopes of mAbs OC125 and M11 are dependent on conformation but not on glycosylation.
The availability of recombinant TR constructs with and without aberrant glycosylation now opens the way for vaccine studies.
Language: | English |
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Year: | 2014 |
Pages: | 3349-3359 |
ISSN: | 15353907 and 15353893 |
Types: | Journal article |
DOI: | 10.1021/pr500215g |
ORCIDs: | 0000-0003-1428-5695 , 0000-0003-1629-187X , 0000-0001-7012-9836 , 0000-0002-4976-0647 and 0000-0002-0915-5055 |
02506, Cytology - Animal 03506, Genetics - Animal 10060, Biochemistry studies - General 10062, Biochemistry studies - Nucleic acids, purines and pyrimidines 12504, Pathology - Diagnostic 16506, Reproductive system - Pathology 24001, Neoplasms - Diagnostic methods 24004, Neoplasms - Pathology, clinical aspects and systemic effects 31000, Physiology and biochemistry of bacteria 31500, Genetics of bacteria and viruses BIOCHEMICAL Biochemistry and Molecular Biophysics CA-125 EPITHELIAL OVARIAN ETD Facultatively Anaerobic Gram-Negative Rods Eubacteria Bacteria Microorganisms (Bacteria, Eubacteria, Microorganisms) - Enterobacteriaceae [06702] Escherichia coli species expression system GALNT GLYCOPROTEOME ISOBM TD-1 WORKSHOP MUC16 NM024690.2 GenBank, EMBL, DDJB nucleotide sequence NP078966.2 GenBank, EMBL, DDJB nucleotide sequence O-GLYCOSYLATION OC125 monoclonal antibody OVARIAN-CANCER ANTIGEN Rodentia Mammalia Vertebrata Chordata Animalia (Animals, Chordates, Mammals, Nonhuman Vertebrates, Nonhuman Mammals, Rodents, Vertebrates) - Cricetidae [86310] CHO cell line cell_line Chinese hamster ovary cells SDG 3 - Good Health and Well-being SEA DOMAIN TANDEM-REPEAT TETHERED MUCINS Tumor Biology antigen-binding epitope cancer vaccine mass spectrometry ovarian cancer Ovarian Neoplasms (MeSH) neoplastic disease, reproductive system disease/female diagnosis recombinant MUG16 tandem repeat expression N-glycosylation site, O-glycosylation site vaccine study