Inhibition of MT-450 rat mammary tumour growth by antibodies recognising subtypes of blood group antigen B.

Using subtractive immunization to identify cell surface epitopes expressed in a metastasis-specific fashion on cells of the rat MT-W9 mammary carcinoma model, we generated a monoclonal antibody called M-N#1. This antibody binds specifically to metastasizing cells of the MT-W9 series and also to certain other metastasizing rat mammary carcinoma cell lines. We demonstrate that the M-N#1 antibody recognizes a fucosylated N-glycosyl sugar modification, and furthermore show that the epitope specificity of the M-N#1 antibody is for blood group antigen B subtypes 2, 3 and 4 with slight cross-reactivity with blood group antigen A subtype 2. The expression of these carbohydrate epitopes on MT-450 cells is functionally important, because the M-N#1 antibody efficiently inhibits MT-450 tumour growth in spontaneous metastasis assays. These results suggest that expression of the subtypes of blood group antigen B recognized by the M-N#1 antibody does not directly participate in the metastatic cascade but rather confers a growth or survival advantage on the tumour cells.

Characterization of a monoclonal antibody against a mucin-type glycoprotein in human sweat.

We report the preparation and characterization of an IgG2 monoclonal antibody (MAb), HSMA, prepared against a human pooled sweat extract (HPSE). The major component of HPSE was a mucin-type molecule, as revealed by sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) stained with periodic acid-Schiff (PAS) reagent. By immunoblotting, HSMA revealed a smear in the high molecular weight range, typical of mucins. In enzyme-linked immunosorbent assay (ELISA), HSMA failed to react with HPSE fractions isolated after anionic exchange gel chromatography. Similarly, radio-immunobinding assays demonstrated no reactivity between HSMA and A, B, H, and Lewis blood group-related structures. The immunohistological labeling on normal skin showed that HSMA reacted with the cells of eccrine sweat glands, and to a lesser extent, with sebaceous glands and epidermal cells. Periodate treatment in situ abolished these reactions, thus suggesting the carbohydrate structure of the HSMA-epitope. In indirect immunofluorescence (IF) studies, HSMA also reacted with other exocrine glands, e.g. mammary glands, sublingual glands, mixed sero-mucous glands of the trachea, and in the pancreas. Sparse positive cells were also observed in the testis, kidney, thyroid and digestive tract.

The Lewis antigens and secretor status.

The coincidence of the presence of ABH active substances in the saliva of persons with those individuals who possess either LEWIS-d or LEWIS-b antigens on epithelial cells of their stomach tissues is examined in terms of the three-dimensional conformations of the oligosaccharide structures which form the ABH and LEWIS antigenic determinants. The H activity in saliva is normally established by examining the inhibition of the agglutination of human H red cells, which possess H (Type 2) determints at their surface, by the lectin Ulex europaeus. As was to be expected, the LEWIS-d (H-Type 1) (alpha LFuc (1 leads to 2) beta DGal (1 leads to 3) beta DGlcNAcOR, R = (CH2)8COOCH3) and LEWIS-b (alpha LFuc (1 leads to 2) beta DGal (1 leads to 3) [alpha LFuc (1 leads to 4)] beta DGlcNAcOR) determinants, obtained by chemical synthesis, do not bind Ulex. However, these structures in the de-N-acetylated form proved to do so as evidenced by their ability to inhibit the agglutination. These results together with results obtained in inhibition studies involving chemically modified H (Type 1) and H (Type 2) structures are rationalized in terms of conformational analysis and raise the possibility that the LEWIS-b and d antigens in saliva are present with the determinants in the amine form. The circumstantial evidence obtained in support of this possibility is, as yet, inconclusive. The main thrust of the paper is to indicate how modern conformational analysis may play an important role in the improvement of knowledge in carbohydrate-receptor site interaction.