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MUC1/X Protein Immunization Enhances cDNA Immunization in Generating Anti-MUC1 /ß Junction Antibodies that Target Malignant Cells

¹ Food and Drug Administration, Silver Spring, Maryland; Departments of ² Cell Research and Immunology and ³ Molecular Microbiology and Biotechnology, Tel Aviv University, Ramat Aviv, Israel; ⁴ Biomodifying, LLC, San Diego, California; ⁵ Antibody Unit, Weizmann Institute of Science, Rechovot, Israel; and ⁶ Department of Hematology, Sourasky Medical Center, Tel Aviv, Israel

Requests for reprints: Daniel H. Wreschner, Department of Cell Research and Immunology, Tel Aviv University, Tel Aviv 69978, Israel. Phone: 972-3-6407425; Fax: 972-3-6422046; E-mail: danielhw{at}post.tau.ac.il or c/o A. Greenboim, Biomodifying LLC, San Diego, CA 92122. Phone: 858-678-0731; Fax: 858-678-0791.

MUC1 has generated considerable interest as a tumor marker and potential target for tumor killing. To date, most antibodies against MUC1 recognize epitopes within the highly immunogenic chain tandem repeat array. A major shortcoming of such antibodies is that the MUC1 chain is shed into the peripheral circulation, sequesters circulating antitandem repeat array antibodies, and limits their ability to even reach targeted MUC1-expressing cells. Antibodies recognizing MUC1 epitopes tethered to the cell surface would likely be more effective. MUC1 subunit binding the membrane-tethered ß subunit provides such an epitope. By use of a novel protocol entailing immunization with cDNA encoding full-length MUC1 (MUC1/TM) followed by boosting with the alternatively spliced MUC1/X isoform from which the tandem repeat array has been deleted, we generated monoclonal antibodies, designated DMC209, which specifically bind the MUC1 /ß junction. DMC209 is exquisitely unique for this site; amino acid mutations, which abrogate MUC1 cleavage, also abrogate DMC209 binding. Additionally, DMC209 specifically binds the MUC1 /ß junction on full-length MUC1/TM expressed by breast and ovarian cancer cell lines and on freshly obtained, unmanipulated MUC1-positive malignant plasma cells of multiple myeloma. DMC209 is likely to have clinical application by targeting MUC1-expressing cells directly and as an immunotoxin conjugate. Moreover, the novel immunization procedure used in generating DMC209 can be used to generate additional anti-MUC1 /ß junction antibodies, which may, analogously to Herceptin, have cytotoxic activity. Lastly, sequential immunization with MUC1/TM cDNA acting as a nonspecific adjuvant followed by protein of interest may prove to be a generalizable method to yield high-titer specific antibodies. (Cancer Res 2006; 66(23): 11247-53)