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We describe the generation and characterization of a monoclonal antibody (MAb), designated D612, with selective reactivity for malignant and normal gastro-intestinal epithelium. MAb D612, a murine IgG2a, was generated using a membrane-enriched fraction of a human colon carcinoma biopsy as immunogen. Employing radioimmunoassays (RIAs) of biopsy extracts to a range of normal and neoplastic tissues, and both immunofluorescence and immunoperoxidase assays on frozen sections of a range of normal and neoplastic tissues, we have shown that MAb D612 binds to 82% of colorectal carcinomas tested (n = 67) and to normal gastro-intestinal epithelium, but does not bind similarly to either neoplastic or normal tissues from a wide range of other sites. Western blotting has shown MAb D612 to react with a high-molecular-weight antigen. Live cell RIAs and FACS analyses demonstrate the reactive epitope to be present on the surface of colon carcinoma cells. Immunohistochemical studies have shown intense membrane staining of colon adenocarcinomas with MAb D612; the vast majority of both primary and metastatic colon adenocarcinomas from a variety of sites were positive with many lesions showing homogeneous staining of virtually all cells present. Using human effector cells, we also showed that MAb D612 mediates antibody-dependent cell-mediated cytotoxicity (ADCC) of human colon carcinoma cells; this activity was enhanced in the presence of interleukin (IL-2). Radiolabelled D612-purified IgG selectively binds a human colon carcinoma xenograft in situ. The pattern of membrane-associated staining, the molecular weight of the reactive antigen, the IgG2a isotype, the ability to mediate ADCC in the presence of IL-2, and the immunohistochemical and RIA studies demonstrating highly restricted reactivity to malignant and normal gastro-intestinal tissue, all distinguish MAb D612 from other MAbs thus far described.
Muraro et al. (Sat,) studied this question.