Abstract #1579: Characterization of monoclonal antibodies to glycodelin showing clinical utility in the identification of ovarian cancer
Authors: Jeffrey Baker, Robert Cheek, John Groelke, Qin He, George Brough, Lorraine King, Steven Knapp, Karen Lenz, Stephen Simkins, Ingrid Felton, Patricia Gallagher, Eric Dixon, Clark Whitehead
Journal: Cancer Research (2009)

Ovarian cancer, the fourth leading cause of cancer death, is diagnosed in more than 22,000 women in the U.S each year. The only established biomarker for ovarian cancer is CA125, which shows low sensitivity for detecting early stage ovarian cancer. The prognosis for late stage diagnosis is poor due to the high rate of metastasis of ovarian cancer. In order to improve the clinical management of ovarian cancer, new biomarkers must be identified and used to create novel  in vitro  diagnostic tools. Glycodelin or PAEP (progestagen-associated endometrial protein) is a member of the kernel lipocalin family of proteins. Glycodelin is normally expressed in women during ovulation and the first trimester of pregnancy. After menopause, no significant level of glycodelin has been reported in normal serum. Our group has previously reported the use of a research use only glycodelin kit to predict the presence of ovarian cancer. The utility of the kit led us to develop an in-house assay for  in vitro  quantitation of glycodelin in human serum. In order to produce a recombinant protein, the coding sequence for glycodelin was fused in frame to a C-terminal Histidine tag in a mammalian expression vector. Recombinant glycodelin was secreted from transiently transfected HEK cells and purified by nickel chromatography directly from conditioned media. Mice were then immunized with recombinant glycodelin to produce monoclonal antibodies. In order to increase the epitopic diversity of glycodelin presented to the immune system, mice were immunized with two injection protocols. Following immunization, B cells were harvested and fused with the myeloma cell line P3X63-Ag-653/Bcl-2. Initial screening of hybridoma supernatants against the recombinant protein resulted in a library of polydoma antibodies specific to glycodelin. Analysis of the supernatents by ELISA reduced the pool of highly specific polydomas to eight, which were subsequently cloned. Two of these clones, one from each immunization were chosen as capture and detector antibodies for further analysis. The binding site for one of the antibodies mapped to the C-terminal region of glycodelin, while the second binds natively folded full-length glycodelin. The two clones also showed differences in binding affinity determined by calorimetry. A serum cohort consisting of 150 benign, 100 normal, 78 cancer, 15 borderline, and 12 interfering pathology samples has been analyzed with the in house glycodelin assay in combination with CA125. The data demonstrate a significant predictive value in addition to CA125 in serum from women older than 55 (p=0.0057). The increased predictive value corresponds to 9 cancer samples identified by glycodelin out of the 30 patients that were determined normal by CA125 alone. This analysis indicates that glycodelin may significantly improve the clinical management of ovarian cancer.   Citation Information:  In: Proc Am Assoc Cancer Res; 2009 Apr 18-22; Denver, CO. Philadelphia (PA): AACR; 2009. Abstract nr 1579.