Generation and characterization of B72.3 second generation monoclonal antibodies reactive with the tumor-associated glycoprotein 72 antigen

Monoclonal antibody (MAb) B72.3 was generated using a membrane-enriched fraction of a human mammary carcinoma biopsy. It has demonstrated reactivity to the majority of human adenocarcinomas including colorectal, gastric, pancreatic, ovarian, endometrial, mammary, and non-small cell lung cancer and no or weak reactivity to normal adult tissues, with the exception of secretory endometrium. The B72.3-reactive antigen, termed tumor-associated glycoprotein (TAG)-72, has been purified and used as an immunogen to generate B72.3 second generation MAbs. Since the source of purified TAG-72 was a human colon cancer (CC) xenograft, these MAbs have been given a CC designation. Twenty-eight CC MAbs, all immunoglobulin Gs, have been generated and shown to be reactive with TAG-72 and via both radioimmunoassay and immunohistochemical analyses show differential reactivity to carcinoma versus normal adult tissue biopsies. Nine CC MAbs (CC11, 15, 29, 30, 40, 46, 49, 83, and 92) were selected for further characterization. As a result of analyses using direct-binding radioimmunoassay to a range of human carcinomas, Western blotting, live cell surface binding assays, five liquid competition radioimmunoassays, and Ka measurements, all nine CC MAbs could be distinguished from each other and from B72.3. The Ka of B72.3 was determined to be 2.54 X 10(9) M-1; all the CC MAbs demonstrated higher KaS with MAbs CC92, 49, and 83 having KaS of 14.26, 16.18, and 27.72 X 10(9) M-1, respectively. These studies thus demonstrate that one or more of the anti-TAG-72 CC MAbs may be more efficient than B72.3, or useful in combination with B72.3, toward the further study of human carcinoma cell population and the diagnostic and therapeutic procedures presently utilizing MAb B72.3.