Distribution and physical properties of BCA200, a Mr 200,000 glycoprotein selectively associated with human breast cancer.

Of 122 mouse monoclonal antibodies selective for human breast cancer, 13 immunoprecipitated an acidic glycoprotein from SK-Br-3 and ZR-75-30 human breast cancer cells. The antigen (BCA200) migrates with an apparent molecular weight of 200,000 on reducing and 180,000 on nonreducing sodium dodecyl sulfate-polyacrylamide gel electrophoresis, suggesting a single polypeptide chain with a folded domain stabilized by a disulfide bond. Cross-blocking and sandwich immunoassays detected at least three distinct antigenic determinants on BCA200. Scatchard experiments measured 1,000,000 to 5,000,000 antigen copies per SK-Br-3 cell. The tissue distribution of BCA200 was studied using two monoclonals to different epitopes. Neither antibody stained any cells in human blood. When frozen sections of 20 normal human tissues were immunoperoxidase stained, the only positive structures were mucinous glands of colon, transitional epithelium of bladder, sweat glands of skin, and acinar epithelium of breast. Antibody 454C11 stained 16 of 21 breast tumor frozen sections and 9 of 12 breast cancer cell lines, while antibody 520C9 stained 5 of 20 breast tumors and 4 of 10 breast cancer lines. Cross-reaction was observed with lung, prostatic, pancreatic, endometrial, and ovarian cancer, but not with lymphoma, melanoma, colon, stomach, bladder, or esophageal cancer. When conjugated to ricin A chain, 10 of 13 antibodies produced immunotoxins selectively cytotoxic to SK-Br-3 breast cancer cells.

Tissue distribution of breast cancer-associated antigens defined by monoclonal antibodies.

Balb/c mice were immunized with membrane preparations of primary and metastatic breast carcinomas or with live breast cancer cell lines. Sixty-two fusions were performed between immune splenocytes and SP2/0 mouse myeloma cells, and 107 hybridomas were cloned that produced antibody reactive with breast cancer membrane extract, cell lines, and frozen sections, but not with normal tissue membrane extracts or a human fibroblast line. Ninety-four monoclonal antibodies were purified and tested for binding to 16 normal tissue frozen sections and five blood cell types. Thirty-five of the 94 antibodies were also tested on breast cancer sections from 21 patients, 14 breast cancer cell lines, and 11 nonbreast tumor sections. Two of 94 antibodies showed no reactivity to any of the 21 normal tissues or 11 nonbreast neoplasms studied. These two antibodies, 451B7 and 452F2, bound 60% of the breast cancer cell lines and 25% of the breast cancer tissue sections. Eight additional antibodies bound three or fewer of 21 normal tissues. These antibodies bound 25-85% of breast cancer sections, 0-75% of breast cancer cell lines, and many of the nonbreast cancers. A comparison of normal tissues and nonbreast tumors bound by the breast cancer-reactive antibodies indicated that most of the cross-reacting normal tissue structures were epithelial in origin, and that the most cross-reactive nonbreast cancers were those of secondary sex organs (uterus, prostate, ovary).