Identification of the gastrointestinal and pancreatic cancer-associated antigen detected by monoclonal antibody 19-9 in the sera of patients as a mucin

Abstract

Monoclonal antibody 19-9, produced by a hybridoma prepared from spleen cells of a mouse immunized with a human colon carcinoma cell line, detects an antigen in the serum from most patients with gastrointestinal and pancreatic cancer (M. Herlyn, H.F. Sears, Z. Steplewski, and H. Koprowski, J. Clin. Immunol., 2: 135-140, 1982). The epitope of this antibody is a carbohydrate with the sugar sequence (formula; see text) in which NeuNAc is N-acetylneuraminic acid, Gal is galactose, GlcNAc is N-acetylglucosamine, and Fuc is fucose. In the colon carcinoma cell line and many gastrointestinal and pancreatic cancers, this sequence occurs in a monosialoganglioside containing a sialylated Lea-active pentasaccharide (sialylated lacto-N-fucopentaose II, IV3-alpha-NeuNAc-III4-alpha-Fuc-LcOse4, in which LcOse4 is Gal beta 1-3GlcNAc beta 1-3Gal beta 1-4Glc) (J. L. Magnani et al. J. Biol. Chem., 257: 14365-14369, 1982). However, the antigen in the sera of patients occurs mainly as a mucin, not a ganglioside, based on the following evidence. Little antigen is extracted by organic solvents from sera, and that which is extracted remains at the origin under conditions of thin-layer chromatography where the ganglioside antigen migrates up the plate. Upon gel filtration of serum on Sephacryl S-400, the antigen is eluted in the void volume, indicating a molecular weight of greater than or equal to 5 X 10(6). Incubation for 5 hr at 37 degrees in 0.1 N NaOH destroys the serum antigen but does not affect the ganglioside antigen. The density of the serum antigen as determined in a CsCl gradient is 1.50 g/ml, while in 4 M guanidine. HCl its density is 1.43 g/ml. Finally, antigen affinity purified by antibody 19-9 from the serum of a cancer patient belonging to the Le(a-b+) blood group contains Leb antigen, consistent with the multiple antigenic specificities exhibited by mucins.