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Characterization of the Shed Form of the Human Tumor-associated Glycoprotein (TAG-72) from Serous Effusions of Patients with Different Types of Carcinomas

Laboratory of Tumor Immunology and Biology, National Cancer Institute, National Institutes of Health, Bethesda, Maryland 20892

Monoclonal antibody (MAb) B72.3 binds a high molecular weight tumor-associated glycoprotein designated TAG-72. This study reports the isolation and characterization of secreted TAG-72 directly from effusions of ovarian, colorectal, pancreatic, and endometrial carcinoma patients and compares them to TAG-72 derived from the LS-174T colon carcinoma xenograft. The B72.3-reactive antigen, TAG-72, was used as immunogen to produce second generation anti-TAG-72 MAbs. One of these second generation MAbs, CC49, had a higher affinity than that of B72.3 and was utilized as an affinity reagent in a procedure to purify the TAG-72 present in the serous effusions of carcinoma patients. A three-step purification procedure, utilizing heat extraction, CC49 antibody affinity chromatography, and gel filtration chromatography, resulted in 1000- to 4400-fold purifications of the TAG-72 derived from effusions, as analyzed using a double-determinant radioimmunoassay. Radiolabeled TAG-72 from each of the effusions demonstrated similar high molecular weight bands on sodium dodecyl sulfate-polyacrylamide gel electrophoresis. Similar results from the various effusions were also obtained in Western blotting analyses. Analyses of TAG-72 from the different effusions in radioimmunoassay using five different anti-TAG-72 MAbs revealed similar binding patterns. The results of these studies thus indicate that TAG-72 obtained directly from patients with ovarian, colorectal, endometrial, and pancreatic carcinomas and the LS-174T xenograft are highly similar in terms of immunochemical properties and antigenic profile.

¹ To whom requests for reprints should be addressed, at the Laboratory of Tumor Immunology and Biology, National Institutes of Health, Building 10, Room 8B07, Bethesda, MD 20892.

Received 2/ 1/90. Revised 5/15/90.

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