[entrez_report] --------------------------------------------------------------------------- Links: [124 medline neighbors] Cancer 78: 2388-2399 (1996) Metastatic phenotype correlates with high expression of membrane-associated complete beta-human chorionic gonadotropin in vivo. Acevedo HF, Hartsock RJ Department of Pathology and Laboratory Medicine, Allegheny-Singer Research Institute, Allegheny General Hospital, MCP-Hahnemann Medical School, Allegheny University of the Health Sciences, Pittsburg, Pennsylvania 15212-9986, USA. BACKGROUND: Investigations using living human cancer cells and the nude mouse model were conducted to evaluate the expression of human chorionic gonadotropin (hCG) in various cancers grown in vitro and in vivo. The aim was to determine whether membrane-associated hCG in any of its forms is a characteristic metastatic marker, and at what levels or ratios. METHODS: Human cancer cell lines known to produce tumors that metastasize spontaneously when grown in nude mice (n = 4) were compared with those that do not produce such tumors (n = 4) using analytical (quantitative) flow cytometry. Monoclonal antibodies directed to epitopes of intact hCG (hCG-holo) and its subunits, including beta-human chorionic gonadotropin with its carboxy-terminal peptide (hCG beta-CTP), allowed for the determination of hCG beta-CTP/hCG-holo ratios. RESULTS: No significant difference in hCG beta-CTP/hCG-holo ratios was found between the cultured human cancer cells that do not metastasize spontaneously (ratio = 2.39) and those that do (ratio = 2.13), and no difference was seen in their growth rate in nude mice. However, the cells isolated from tumors that do not metastasize spontaneously showed a decrease in their ratios to values less than 1. They reverted to their original values after reestablishment in culture and subsequent passages. In contrast, the ratios shown by cells isolated from tumors that metastasize spontaneously increased to 3 to 6 times their original values in culture, then reverted to their original values after reestablishment in culture and subsequent passages. CONCLUSIONS: To our knowledge, these data demonstrate the following for the first time: 1) There is a direct in vivo correlation between human cancer cells that metastasize spontaneously in nude mice and the expression of membrane-associated complete hCG beta (hCG beta-CTP); and the correlation identifies this molecule as a characteristic metastatic phenotype marker. 2) The marked ratio variations under different conditions indicate that the metastatic phenotype is an unstable event. 3) Growth and local invasion in vivo correlates with the expression of hCG-holo. PMID: 8941011, MUID: 97095987 --------------------------------------------------------------------------- Links: [155 medline neighbors] Cancer 76: 1467-1475 (1995) Human chorionic gonadotropin-beta subunit gene expression in cultured human fetal and cancer cells of different types and origins Acevedo HF, Tong JY, Hartsock RJ Department of Pathology and Laboratory Medicine, Allegheny-Singer Research Institute, Allegheny General Hospital, Medical College of Pennsylvania, Pittsburgh 15212-9986, USA. BACKGROUND. The authors' previous investigations using living cultured human cancer cells and cells isolated from cancer tissues, analytical flow cytometry, and monoclonal antibodies directed to epitopes located in five different sites of the human chorionic gonadotropin (hCG) molecule, identified the presence of membrane-associated hCG, its subunits and fragments, by cells from all cancers, irrespective of type and origin, indicating that the expression of these sialoglycoproteins is a common phenotypic characteristic of cancer. Although benign neoplasms do not express these compounds, cultured human embryonic and fetal cells also express the same materials. To corroborate these findings, five fetal cell lines and 28 cancer cell lines were randomly selected from those previously studied, to determine the presence of translatable levels of hCG-beta (hCG beta) mRNA. METHODS. All cell lines were grown under identical conditions. Determination of hCG beta mRNA was made by extracting the total RNA from the cells, followed by synthesis of cDNA with RNase H- reverse transcriptase and polymerase chain reaction amplification using specific hCG beta-luteinizing hormone-beta (hLH beta) primers. The presence of amplified hCG beta cDNA was corroborated by hybridization of the product with an hCG beta-specific oligonucleotide and Southern blot analyses of the hybridization products. Gestational choriocarcinoma cells and HeLa adenocarcinoma of cervical cells, known producers of biologically active hCG, were positive control subjects, and human pituitary cells were used as negative control subjects. RESULTS. The results showed single and multiple hCG beta gene activation by the fetal cells and the different types of cancer, indicating that at any given time, there is the possibility of activation of as many as four genes of the six genes of the hCG beta-hLH beta gene cluster, even though alternative gene splicing cannot be ruled out. CONCLUSIONS. In addition to the authors' previous findings, the results of these studies support the concept that cancer is a problem of development and differentiation, and, to the authors' knowledge, prove definitively for the first time that synthesis and expression of hCG, its subunits, and its fragments, is a common biochemical denominator of cancer, providing the scientific basis for studies of its prevention and/or control by active and/or passive immunization against these sialoglycoproteins. PMID: 8620425, MUID: 96223203 --------------------------------------------------------------------------- Links: [200 medline neighbors] Cancer 69: 1829-1842 (1992) Expression of membrane-associated human chorionic gonadotropin, its subunits, and fragments by cultured human cancer cells. Acevedo HF, Krichevsky A, Campbell-Acevedo EA, Galyon JC, Buffo MJ, Hartsock RJ Department of Pathology and Laboratory Medicine, Allegheny General Hospital, Pittsburgh, PA 15212. The expression of human chorionic gonadotropin (hCG), its subunits, and fragments on the cell membrane of cultured human cancer cells was investigated using a flow cytometric method. This method uses living cells; a double-antibody reaction; a flow cytometer with an argon laser, standard settings, and filters for fluorescein isothiocyanate; commercially available software; the American Type Culture Collection (ATCC) CCL 2 HeLa cell line as cell control and overall quality control; polyclonal rabbit antisera raised against the hCG dimer, its alpha subunit (hCG alpha), and its beta subunit (hCG beta); and a panel of monoclonal antibodies (MoAb) recognizing different epitopes on the intact hCG molecule, its subunits, and fragments. The purified immunoglobulin G fractions from the polyclonal antisera were used to estimate the total expression of the membrane-associated glycoproteins; the MoAb were used to detect the expression of epitopes of the hCG dimer, its subunits, and fragments. The results of the analyses done on cells from 74 established cancer cell lines of different types and origins (including 52 carcinomas, 10 sarcomas, 4 leukemias, 6 lymphomas, and 2 retinoblastomas) showed variable degrees of reactivity in a great percentage of cells in all cell lines studied with MoAb directed against different conformational epitopes of intact hCG (hCG-holo), hCG beta, hCG beta-free, the carboxy terminal peptide (CTP) of hCG beta, and an epitope of hCG alpha. The expression of the membrane-associated epitopes of hCG and its subunits was found to be a phenotypic marker characteristic of all evaluated cultured human cancer cell lines, irrespective of their type or origin. There were, however, quantitative and qualitative differences in the expression of the different epitopes. Thus, hCG beta, free and as part of hCG-holo, recognized by the MoAb against hCG beta-CTP, was expressed by a high percentage of cells of most cell lines. There was great variability in the expression of hCG-holo, recognized by MoAb B109. For this reason some groups of cancers expressed larger amounts of incompetent hCG alpha and/or hCG beta than others. Cell lines derived from adenocarcinomas of the lung were the only exception to this general finding; the expression of small amounts of hCG-holo was caused by a low degree of hCG alpha synthesis. PMID: 1372528, MUID: 92200333 --------------------------------------------------------------------------- Cancer 69: 1818-1828 (1992) Flow cytometry method for the analysis of membrane-associated human chorionic gonadotropin, its subunits, and fragments on human cancer cells. Acevedo HF, Krichevsky A, Campbell-Acevedo EA, Galyon JC, Buffo MJ, Hartsock RJ Department of Pathology and Laboratory Medicine, Allegheny General Hospital, Pittsburgh, PA 15212. A quantitative flow cytometry method for the analysis of membrane-associated human chorionic gonadotropin (hCG), its subunits, and fragments on human cancer cells was developed using a double-antibody reaction; a flow cytometry with a 2-W argon laser, standard settings, and filters for fluorescein isothiocyanate use; commercially available software; and the ectopic hCG producer CCL 2 HeLa cells from the American Type Culture Collection (ATCC) as a cell control to standardize the reagents and for overall quality control. Twenty-two monoclonal antibodies (MoAb) and immunoglobulin G fractions from three rabbit polyclonal antisera were tested for effects of antibody concentration (titration), reproducibility at different levels of epitope expression, and variability of epitope expression to select appropriate primary antibodies. Based on the results of the various tests, three polyclonal immunoglobulin G antibodies and a panel of nine MoAb directed to epitopes located in five different regions on the hCG molecule were selected as first antibodies. Their specificity was determined by using two unrelated MoAb of the same isotype at the same concentration to replace the primary MoAb and by a competition experiment. The unrelated MoAb also were used for the selection of the appropriate control fluorescence profile needed for the software. The unique characteristics of this method were: the use of living cells, standardized reagents, internal and external quality control, and the highest sensitivity, which could detect as few as 10(3) molecules of fluorochrome per cell. Serial analyses of the ATCC CCL 2 HeLa cells and two of its variants and of the eutopic hCG producer JEG-3 choriocarcinoma cells revealed the expression of membrane-associated epitopes of intact hCG, its subunits, and fragments by a high percentage of the cells, indicating that the expression of these sialoglycoproteins by these two different types of cancer cells is a common phenotypic characteristic. MeSH Terms: * Adenocarcinoma/pathology * Adenocarcinoma/chemistry* * Antibodies, Monoclonal * Binding, Competitive * Cell Membrane/physiology * Cell Survival/physiology * Epitopes/immunology * Epitopes/analysis * Female * Flow Cytometry/methods* * Gonadotropins, Chorionic/physiology * Gonadotropins, Chorionic/immunology * Gonadotropins, Chorionic/analysis* * Hela Cells * Human * Neoplasms/pathology * Neoplasms/chemistry* * Peptide Fragments/analysis* * Reproducibility of Results * Sensitivity and Specificity * Support, Non-U.S. Gov't * Support, U.S. Gov't, P.H.S. * Time Factors Substances: * Peptide Fragments * Gonadotropins, Chorionic * Epitopes * Antibodies, Monoclonal PMID: 1372527, MUID: 92200332