Ectopic Beta-Human Chorionic-Gonadotropin Production by A Human Hepatoma-Cell Line (focus) - Isolation and Immunochemical Characterization

We have established a human hepatocellular carcinoma cell line designed FOCUS that produces and secretes the beta-subunit of hCG. In the study of beta hCG production by FOCUS cells, we have developed and employed a series of monoclonal immunoradiometric assays (IRMAs) that detect epitopes unique to beta hCG, alpha hCG, hCG, and sequence-specific regions of the carboxyl-terminal peptide of beta hCG. The cells secrete approximately 15 ng beta hCG/10(6) cells and per 24 h; however, we were unable to detect either hCG or the alpha-subunit. The ectopic beta hCG was subsequently affinity purified from the culture medium and partially characterized by sodium dodecyl sulfate-polyacrylamide gel electrophoresis and autoradiography. The mol wt of ectopic beta hCG was approximately 35,000 daltons. Immunochemical analysis of ectopic beta hCG by sequence-specific monoclonal antibodies revealed that epitopes corresponding to amino acid sequences 109-115, 121-145, 134-140, and 139-145 of the carboxyl-terminal peptide were present. FOCUS cells also demonstrate an intracytoplasmic localization of beta hCG by immunoperoxidase-staining techniques. Taken together, these findings suggest that FOCUS cells produce and secrete only beta hCG and that thus far, its physical properties appear indistinguishable from those of the native subunit. This unique cell line will be useful to study beta hCG gene(s) regulation as well as the mechanisms of ectopic beta hCG production and secretion.