Characterization of the ETSA-21 Antigen,a Glycosylphosphatidyl-Inositol Anchor Glycoprotein Identified in Breast Cancer Cells Using Monoclonal Antibody B21

Mab B21 is a monoclonal antibody (Mab) that recognizes an epithelial tumor surface antigen (ETSA–B21) from diverse human tumor cell lines including breast, ovary, uterus, and their cognate carcinoma tissues. A lower reactivity has been observed in normal breast tissue and benign hyperplesia. In this study, the characteristics of the ETSA–B21 antigen have been examined in greater detail in the MCF–7, SK-BR–3, and MDA–MB–453 breast cancer cell lines. Treatment with phosphatidylinositol-phospholipase C, but no neuraminidase were found to partially remove the ETSA–B21 signal from the cell surface as revealed by immunofluorescence microscopy. Inhibition of the N-glycosylation pathway by tunicamycin resulted in a decreased ETSA–B21 signal on the cell membrane. In addition, the antigen–antibody complex was internalized in breast cancer cells as demonstrated by an acidic wash internalization assay evaluated using immunofluorescence. In conclusion, this study suggests that ETSA–B21 is a GPI anchor N-glycosylated protein promoting specific antibody internalization in breast cancer cells.