Harada Y, Ohuchi N, Masuko T, Funaki Y, Mori S, Satomi S, Hashimoto Y
Second Department of Surgery, Tohoku University School of Medicine, Sendai, Japan.
Tohoku J Exp Med. 1996 Nov;180(3):273-88. doi: 10.1620/tjem.180.273.
We have characterized a new tumor-associated antigen defined by monoclonal antibody (MAb) generated against HMA-1 breast cancer cell line. MAb AM-1 was selected based on its preferential reactivity to breast cancer cells versus to normal or benign epithelial cells by immunofluorescence and immunohistochemical assays of cultured, or fresh specimens. AM-1 demonstrated strong reactivity to breast cancer cell lines including HMA-1, YMB-1-E, YMB-1 and MDA-MB-231 in flow cytometry. In immunoprecipitation, AM-1 recognized high molecular weight components of 160-210 kDa and > 370 kDa. Reactivity with HMA-1 cells was diminished markedly when treated by heat, protease or periodate, suggesting that the antigenic epitope is composed with carbohydrates and peptides. Enzyme digestion of precipitated antigens demonstrated that the antigen contains O-linked and N-linked carbohydrates with neuraminic acid structures. Furthermore, binding inhibition and sandwich ELISA assays using MAbs reactive with known breast cancer-associated antigens and synthetic MUC1 core peptide (PDTRPAPGSTAPPAHGVTSAPDTR) demonstrated that the antigen is distinct from CEA, TAG-72 or MUC1, while the antigen conjoins with MUC1 and TAG-72 as a trimmer form in HMA-1 cells. These results suggest that AM-1 recognizes a novel glycoprotein which is abundant in breast cancer, and may be utilized in the management of breast cancer patients.
我们已经鉴定出一种新的肿瘤相关抗原，该抗原由针对HMA-1乳腺癌细胞系产生的单克隆抗体（MAb）所定义。通过对培养的或新鲜标本进行免疫荧光和免疫组织化学分析，基于MAb AM-1对乳腺癌细胞相对于正常或良性上皮细胞的优先反应性，筛选出了该抗体。在流式细胞术中，AM-1对包括HMA-1、YMB-1-E、YMB-1和MDA-MB-231在内的乳腺癌细胞系表现出强烈反应。在免疫沉淀中，AM-1识别出分子量为160 - 210 kDa和> 370 kDa的高分子量成分。用热、蛋白酶或高碘酸盐处理后，与HMA-1细胞的反应性显著降低，这表明抗原表位由碳水化合物和肽组成。对沉淀抗原进行酶消化表明，该抗原含有具有神经氨酸结构的O-连接和N-连接碳水化合物。此外，使用与已知乳腺癌相关抗原反应的单克隆抗体和合成MUC1核心肽（PDTRPAPGSTAPPAHGVTSAPDTR）进行结合抑制和夹心ELISA分析表明，该抗原与癌胚抗原（CEA）、TAG-72或MUC1不同，而在HMA-1细胞中，该抗原以三聚体形式与MUC1和TAG-72结合。这些结果表明，AM-1识别一种在乳腺癌中丰富的新型糖蛋白，并且可能用于乳腺癌患者的管理。
