To Discover New Tumor Biomarkers By Antibodies

Scientists at the Harvard Medical School discovered the mouse counterpart ofHuman Carcinoma Antigen (HCA) and named it epiglycanin. Epiglycanin hasseveral remarkable attributes including expression only on cancer cells,unusual size and complexity, and a potentially cancer-promoting alteration ofthe immune response. It was discovered that monoclonal antibodies specificfor mouse epiglycanin displayed the striking ability to react strongly withhuman cancer tissues, but only weakly, if at all, with normal tissues. Finallyinvestigators observed that elevated levels of the molecule they later calledHCA could be measured in tissues and body fluids from patients withcarcinomas, but not in fluids from normal patients. The observation that HCA iselevated in the sera and body fluids of cancer patients has led to significantdiagnostic product concepts for confirmation and monitoring of a variety ofcancers. The structure and the function of HCA are not well recognized. Andthere is only one IgM antibody can react with HCA.In the present study, we described our strategies to generate monoclonalantibodies for HCA using HCA isolated from total carcinoma tissue proteinsand total cancer cell line proteins as the immunogens. The antibodies againstHCA were selected by Western blot, ELISA and immunohistochemistry. 90monoclonal antibodies against HCA were generated. 20 monoclonalantibodies recognized proteins of human liver cancer tissue and onemonoclonal antibody could specifically react with human prostate cancer tissue but not normal tissue by Western blot. Two monoclonal antibodiesrecognized proteins of liver cancer tissue but not normal tissue while onemonoclonal antibody reacted with proteins in prostate cancer tissue but notproteins in benign prostatic hyperplasia tissue by immunohistochemistry. Wefound that both the molecular weight of antibody-recognized proteins and thesubcellular localization of the antigens were different by Western blot andimmnohistochemistry. A panel of antibodies was also tested for their ability tocapture the targeting proteins and complexes from the total cancer tissueproteins. The antibody-recognized proteins were identified by massspectrometry following immunoprecipitation (IP). Finally, we found that both90K recognized by monoclonal antibody BAF112 and Annexinâ…¡recognizedby monoclonal antibody BCE075 elevated in cancer tissue. We believe thesewell-characterized antibodies would be useful in finding cancer biomarkers andmore proteins specific to cancer will be found in the near future.