Isolation and characterization of ovarian cancer antigen CA 125 using a new monoclonal antibody (VK-8): identification as a mucin-type molecule.
A new murine monoclonal antibody (MAb VK-8), detecting the CA 125 ovarian cancer antigen, was used to purify this antigen from OVCAR-3 ovarian cancer cells by affinity chromatography. The biochemical properties of the purified antigen are characteristic of a mucin-type glycoprotein: (1) the molecule is highly glycosylated (77% w/w), mainly with galactose, N-acetylglucosamine, and N-acetylgalactosamine, (2) the protein moiety is rich in serine, threonine and proline, (3) many of the serine and threonine residues are glycosylated, (4) the glycan chains are almost entirely O-linked, with core 2 [Galbeta1 --> 3(GlcNAcbeta1 --> 6)GalNAc] structures predominating and (5) these chains carry fucosylated Type 2 (Le(y) and Le(x) and H type 2) blood group structures. The antigen exhibited a very high m.w. (> 10(3) kDa) in aqueous buffer as well as in urea, but was degraded by proteolytic enzymes to smaller fragments that no longer reacted with the antibody. Although this result, and other immunochemical data, indicate that OC125, the original MAb to CA125, and VK-8 antibodies detect epitopes on the protein portion of the molecule, the involvement of carbohydrate cannot be ruled out. Further insight into the structure and function of the CA125 antigen will come from cloning the gene coding for the peptide backbone, and from more detailed carbohydrate structural analysis.
Lloyd
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KO
Immunology Program, Memorial Sloan-Kettering Cancer Center, New York, NY 10021, USA. klloyd@ski.mskcc.org
Yin
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BW
Kudryashov
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V
D000116:Acetylgalactosamine / Q000032:analysis; D000117:Acetylglucosamine / Q000032:analysis; D000818:Animals; D000911:Antibodies, Monoclonal / Q000276:immunology; D000918:Antibody Specificity; D018394:CA-125 Antigen / Q000737:chemistry* / Q000276:immunology / Q000302:isolation & purification*; D002236:Carbohydrate Conformation; D004591:Electrophoresis, Polyacrylamide Gel; D004797:Enzyme-Linked Immunosorbent Assay; D005260:Female; D005643:Fucose / Q000378:metabolism; D005690:Galactose / Q000032:analysis; D006031:Glycosylation; D006801:Humans; D007163:Immunosorbent Techniques; D007457:Iodine Radioisotopes; D007983:Lewis Blood Group Antigens; D051379:Mice; D009077:Mucins / Q000737:chemistry* / Q000276:immunology; D010051:Ovarian Neoplasms / Q000276:immunology*
D000911:Antibodies, Monoclonal; D018394:CA-125 Antigen; D007457:Iodine Radioisotopes; D007983:Lewis Blood Group Antigens; D009077:Mucins; D005643:Fucose; D000116:Acetylgalactosamine; D000117:Acetylglucosamine; D005690:Galactose