Preparation and Characterization of Monoclonal Antibodies Directed to the Tumor-associated O-linked Sialosyl-2→6 α-N-Acetylgalactosaminyl (Sialosyl-Tn) Epitope
T Kjeldsen, H Clausen, S Hirohashi, T Ogawa… - Cancer …, 1988 - aacrjournals.org
T Kjeldsen, H Clausen, S Hirohashi, T Ogawa, H Iijima, S Hakomori
Cancer research, 1988•aacrjournals.orgTwo monoclonal antibodies, TKH1 and TKH2, directed toward the sialosyl-Tn structure
(NeuAcα2→ 6GalNAcα1→ O-Ser or Thr), which display a remarkable immunohistological
tumor specificity, were generated by immunization with ovine submaxillary mucin. The
reactivity of these antibodies was monitored by solid phase enzyme-linked immunosorbent
assay with different native and glycosidase-treated mucins and glycoproteins. Binding of the
antibody to ovine submaxillary mucin glycoprotein was strongly inhibited by the O-linked …
(NeuAcα2→ 6GalNAcα1→ O-Ser or Thr), which display a remarkable immunohistological
tumor specificity, were generated by immunization with ovine submaxillary mucin. The
reactivity of these antibodies was monitored by solid phase enzyme-linked immunosorbent
assay with different native and glycosidase-treated mucins and glycoproteins. Binding of the
antibody to ovine submaxillary mucin glycoprotein was strongly inhibited by the O-linked …
Abstract
Two monoclonal antibodies, TKH1 and TKH2, directed toward the sialosyl-Tn structure (NeuAcα2→6GalNAcα1→O-Ser or Thr), which display a remarkable immunohistological tumor specificity, were generated by immunization with ovine submaxillary mucin. The reactivity of these antibodies was monitored by solid phase enzyme-linked immunosorbent assay with different native and glycosidase-treated mucins and glycoproteins. Binding of the antibody to ovine submaxillary mucin glycoprotein was strongly inhibited by the O-linked disaccharide NeuAcα2→6GalNAcα1→O-serine, less strongly by NeuAcα2→6GalNAcβ1→O-propyl, and weakly by the monosaccharide GalNac. The reactivity was compared with previously established anti-Tn antibodies B72.3, NCC-Lu-35, and NCC-Lu-81. The antibody B72.3 was prepared previously after immunization with metastatic breast adenocarcinoma and its epitope was claimed to be GalNAcα1→O-Ser (or - Thr) by Springer and associates [Springer, G. F., et al. In: T. Dao, et al. (eds.), Tumor Markers and Their Significance in the Management of Breast Cancer, pp. 47–70. New York: A. R. Liss, 1986]. The antibody was found to show very similar reactivity as that of TKH1/TKH2, and its reactivity to ovine submaxillary mucin was inhibited specifically by NeuAcα2→6GalNAcα1→O-serine, indicating that the antibody is clearly directed to sialosyl-Tn antigen. Immunohistological study of the distribution of this antigen in various normal human tissues and carcinomas by TKH1/TKH2 antibodies, as well as B72.3 and monoclonal antibodies NCC-Lu-35/81, which are directed to GalNAcα1→O-Ser or Thr (Tn), was performed. The sialosyl-Tn antigen was not found in normal tissue except for a weak expression in Leydig cells of the testis, goblet cells of the colon, and parietal cells of the stomach. In contrast, the sialosyl-Tn antigen was strongly expressed in a large number of adenocarcinomas. As expected from the specificity studies, B72.3 shows the same reactivity as TKH1 and TKH2. Thus, both sialosyl-Tn (NeuAcα2→6GalNAcα1→O-Ser/Thr) and Tn (GalNAcα1→O-Ser/Thr) are good tumor markers, and combined use of antibodies directed to these structures might be useful in the screening and classification of cancer.
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