Cell surface coatings and membrane potentials of malignant and nonmalignant cells.

A positive surface potential indicating a cell coating is common to malignant cells, lymphocytes, and normal and malignant trophoblastic cells. This characteristic was not generally found for other normal cell types tested by microelectrode penetration.

Human hormone production in vitro.

A new in vitro hormone-synthesizing cell system, line BeWO, derived from human gestational choriocarcinoma, has been permanently established; a high degree of synthesis of functional hormone has been maintained for 18 months in tissue culture. Predominantly glycolytic metabolism, dependent on the level of glucose in the medium, has been observed. Many facets of placental and tumor-cell metabolism, growth, and differentiation are found in this unique multipotential cell.

Tumor antigen and human chorionic gonadotropin in CaSki cells: a new epidermoid cervical cancer cell line.

Epidermoid cervical carcinoma cells (CaSki line) have been established in continuous culture. When leukocytes from cervical cancer patients were incubated with CaSki culture fluid concentrates, inhibition of leukocyte migration was observed in more than 70 percent of the patients tested. By contrast, significantly less inhibition was observed with normal donor leukocytes or leukocytes from patients with other types of cancer. These results were consistent with the expression of tumor-associated antigen by CaSki cells. Analysis of the serum from the donor of the cell line at the time of tumor biopsy, and of CaSki culture fluids, demonstrated the presence of the beta subunit of human chorionic gonadotropin.

Expression of cell-mediated immunity and blocking factor using a new line of ovarian cancer cells in vitro.

A cell-mediated cytotoxicity test, quantitated by postlabeling with tritiated thymidine, was used to asses immune reactivity of cancer patients to the HeW cell line derived from serous cystadenocarcinoma of the ovary. Lymphocytes from 71.4% of serous and mucinous cystadenocarcinoma patients demonstrated a cytotoxic response towards the HeW cells, whereas no reactivity was observed towards target cells derived from nonovarian cancer. These observations indicated that the HeW cells express tumor-associated antigen. In some patients bearing similar tumors, cytotoxicity was blocked by ascitic fluid from other patients with cystadenocarcinoma. In addition, antigen obtained from the spent culture fluid of HeW cells exhibited blocking activity in a typical dose-response fashion, suggesting that blocking factor may be free tumor-associated antigen or an antigen-specific suppressor molecule. Thus, blocking of the lymphocytotoxic response of cystadenocarcinoma patients towards HeW cells may be utilized to monitor the isolation of ovarian carcinoma-associated antigen.

A distinctive form of human chorionic gonadotropin beta-subunit-like material produced by cervical carcinoma cells.

The DoT and CaSki human cervical carcinoma cell lines ectopically produce material immunologically similar to the beta-subunit of human chorionic gonadotropin (hCG beta). Culture fluids were analyzed by gel filtration chromatography and radioimmunoassay (RIA) using (a) antiserum directed to conformation-specific (core-directed) determinants not involving the carboxyl-terminal peptide (CTP) in hCG beta purified from urinary hCG (i.e., standard hCG beta) or (b) antiserum directed to the CTP in standard hCG beta. CTP-directed RIA recognized a peak of hCG beta-like immunoreactive material that eluted in the same position as standard hCG beta. However, core-directed RIA recognized additional hCG beta-like material (i.e., ectopic beta-II), most of which eluted before standard hCG beta. CaSki cells were incubated with [3H]mannose, [3H]proline, and [3H] leucine, and the spent medium was immunoprecipitated and analyzed by gel electrophoresis. Several labeled peaks were detected in the lane from the anti-hCG beta X Sepharose immunoprecipitate, one of which corresponded in mobility to standard hCG beta, with two more intense components migrating at higher apparent molecular weights. Carboxypeptidase Y digestion released only 0.2 mol equivalents each of [3H]proline and [3H]leucine from the labeled CaSki material immunoprecipitated with anti-hCG beta X Sepharose, compared to 1 mol equivalent each in similar analysis of standard hCG beta. These findings were consistent with the absence of the 4-carboxy-terminal amino acids from 80% of the hCG beta-like immunoreactive material secreted by CaSki cells. The affinity purified ectopic beta-II failed to combine with standard hCG alpha under conditions in which combination of standard hCG beta with standard hCG alpha was essentially complete. Neither aggregation nor proteolytic degradation was the cause of failure of ectopic beta-II to combine with hCG alpha. We conclude that both the DoT and CaSki cervical carcinoma cell lines secrete a distinctive form of hCG beta-like material, ectopic beta-II. Lack of recognition by CTP-directed antisera and amino acid analysis suggest that ectopic beta-II may lack the CTP, despite its apparent larger size relative to standard hCG beta.

Absence of the COOH-terminal peptide on ectopic human chorionic gonadotropin beta-subunit (hCG beta).

A comparison of the physical properties and immunoreactivity of ectopic hCG beta from cultured cervical cancer cells and standard hCG beta was performed. The heterogeneity of the preparations was examined by ion-exchange chromatography, and the molecular size by analytical gel filtration. Two distinct forms of ectopic hCG beta were identified. One form was indistinguishable from standard hCG beta, while the other, although larger, lacked the characteristic COOH-terminal peptide (CTP). This was shown by the failure of antisera specific for determinants on the cTP to recognize this molecule, and by the apparent absence of the 0-linked oligosaccharides and thermolysin cleavage site normally found in this region.

Human chorionic gonadotropin in BeWo trophoblastic cells after 12 years in continuous culture: retention of intact human chorionic gonadotropin secretion in mechanically versus enzyme-dispersed cells.

Gel filtration and radioimmunological studies were used to demonstrate that BeWo malignant trophoblastic cells, subcultured for more than 12 yr as roller tube colonies using dissection by scalpel, retained the ability to secrete large quantities of intact hCG. By contrast, BeWo cells that had been subcultured weekly for 12 hr using proteolytic enzyme dispersion lost their ability to secrete intact hCG. Rather, a large form of hCG-alpha was the major secretory product, along with lesser amounts of heterogeneous low molecular weight forms of hCG-beta. The gradual loss of hormone secretory ability and fidelity of hormone product by cells in continuous culture is a well known phenomenon. Our results suggest that the habitual use of trypsin-EDTA to subculture cells may accelerate this process.

Regulation of human chorionic gonadotropin (hCG) secretion by serum and dibutyryl cyclic AMP in malignant trophoblast cells in vitro.

Human chorionic gonadotropin (hCG) secretion was studied in human malignant trophoblast cells (Jar line) in continuous culture. Radioimmunoassayable hCG in the cells and in the daily culture fluid was increased for at least 3 days by 10% newborn calf serum and by 1 mN N-6, 0-2'-dibutyryl cyclic adenosine 3', 5'-monophosphoric acid (dbcAMP) plus 1 mM theorphylline. Incubation of the cells in the presence of serum, dbcAMP and theorphylline resulted in additive stimulation of hCG secretion after 1 day. After 2 or 3 days, a supra-additive stimulation was observed, which was prevented by Actinomycin D. When cells were preincubated for 2 days in the absence of serum but with dbcAMP plus theorphylline, subsequent addition of serum resulted in supra-additive stimulation of hCG secretion within a single day; the continued presence of dbcAMP plus theophylline was not necessary for this stimulation by serum. These findings suggested that dbcAMP activates a slow process requiring RNA synthesis, resulting in increased hCG secretion. Serum appears to protect the hCG synthesizing system from degradation.

A human parathyroid carcinoma that produces parathyroid hormone: long term maintenance in tissue culture.

Parathyroid carcinoma cells from a pulmonary metastasis of a patient with a serum Ca of 17 mg/dl and an immunoreactive parathyroid hormone (PTH) of 6.4 ng eq bovine (b) PTH/ml (normal 40--400 pg/ml) have been maintained in tissue culture for more than 2 1/2 years. The cells secrete PTH into the culture media that 1) during immunoassay dilutes in parallel to human hyperparathyroid serum, 2) has a molecular weight similar to intact highly purified bPTH, and 3) stimulates bone resorption in a manner that is equivalent and additive to synthetic bPTH-(1--34).

Phosphorus-32-chromic phosphate for ovarian cancer: I. Fractionated low-dose intraperitoneal treatments in conjunction with platinum analog chemotherapy.

UNLABELLED: For many years, 32P-chromic phosphate (32P-CP) intraperitoneal instillations and platinum analogue chemotherapy have been used to treat disseminated ovarian cancer. To investigate possible enhancement of 32P-CP irradiation due to the concomitant administration of chemotherapy, in vitro studies were undertaken. Based on those laboratory investigations, a clinical regimen of combined 32P-CP and platinum analogue chemotherapy was developed. METHODS: In vitro enhancement of 32P-CP cytotoxicity by cisplatin was studied in cultured human ovarian adenocarcinoma (CHOA) cell lines and in a fibroblast cell strain. In addition, ovarian cancer cells obtained from the malignant abdominal ascites and pleural effusions of 10 individual patients were also studied ex vivo. As part of routine clinical care, 30 patients with disseminated ovarian adenocarcinoma underwent up to eight monthly cycles of platinum analogue chemotherapy with concomitant intraperitoneal instillation of 5 mCi of 32P-CP at each monthly chemotherapy cycle. RESULTS: There was an enhanced and possibly supra-additive effect of cisplatin on the cytotoxicity from 32P-CP irradiation. For the 30 patients, the survival rate at 3 yr was 63%. CONCLUSION: Phosphorus-32 CP low-dose intraperitoneal treatments in conjunction with platinum analogue chemotherapy is a promising approach for the treatment of disseminated intraperitoneal ovarian cancer.

Carcinoembryonic antigen and L-fucose in malignant and benign mammary disease.

A prospective study of 123 patients was undertaken to determine whether carcinoembryonic antigen (CEA) or L-fucose could be used as "tumor markers" to distinguish patients who had mammary cancer (41) from patients who had benign mammary disease (41) and patients who did not have mammary cancer (41). The predictive value of positive test results for both L-fucose and CEA was lower for patients who smoked than for those who did not. The sensitivities of CEA and L-fucose indicators are both below 55%. There were frequent false-positive and false-negative results. A substantial overlap in the levels of CEA and L-fucose was found for the patients who had mammary cancer and those who had benign mammary disease. CEA and L-fucose performed similarly in distinguishing cancer from benign disease. For patients who smoked, however, the predictive value of L-fucose was only 44%. The effect of smoking on L-fucose levels may account for discrepancies in L-fucose data reported in previous studies of mammary cancer. Neither CEA nor L-fucose were found to be of value in screening for mammary cancer.

Effect of Crude and Purified hCG on Lymphocyte Blastogenesis.

Crude APL hCG completely inhibited blastogenesis induced by phytohemagglutinin, pokeweed mitogen, or concanavalin A in normal lymphocytes and lymphocytes obtained from cancer patients. Phenol in quantities comparable to its concentration in commercial lyophilized APL hCG also completely inhibited all three mitogenic responses. By contrast, similar doses of highly purified hCG had no inhibitory effect; higher concentrations produced only slight inhibition. Beta-hCG had a partial (less than 50%) inhibitory effect that was dose-independent and nonreproducible.

Production of tumor-associated antigen, TA-4, by the CaSki cervical carcinoma cell line.

Previous Japanese studies described the purification of TA-4 from homogenates of tumor tissues excised from squamous cell carcinomas of the uterine cervix, and the development of a radioimmunoassay to detect TA-4 in sera of patients with this disease. The aim of the present investigation was to determine if TA-4 was produced by the CaSki cell line, established in culture ten years ago from epidermoid carcinoma of the uterine cervix. The radioimmunoassay detected the TA-4 antigen in the CaSki cells, but not in cell lines derived from either choriocarcinoma or breast carcinoma. The TA-4 concentration in the CaSki cell lysate exceeded that in the CaSki culture fluid by more than twentyfold, and exceeded the concentration of human chorionic gonadotropin beta-like immunoreactive material by nearly two orders of magnitude. Antiserum to TA-4 was used to immunoprecipitate biosynthetically labeled TA-4 from CaSki cultures that had been incubated with [3H]leucine. After electrophoresis and autoradiography, the immunoprecipitated material showed a major band corresponding in apparent molecular weight (48,000 daltons) to TA-4 originally isolated from squamous cell carcinoma tumors. It is concluded that the CaSki cell line constitutes an ideal model with which to investigate the biosynthesis and regulation of TA-4, and a source for large-scale production of TA-4 for characterization studies as well as development of clinical diagnostic reagents.

Alpha subunit and human chorionic gonadotropin in normal pregnancy and gestational trophoblastic disease.

The reported incidence of gestational trophoblastic disease is an order of magnitude higher in Nigeria than in the United States. Sera from a total of 283 pregnant black patients, 138 United States and 148 Nigerian pregnant patients, were analyzed for their serum levels of alpha subunit and human chorionic gonadotropin (hCG). The patterns of hCG secretion were similar in the two populations during normal pregnancy. However, the level of alpha subunit was persistently higher in Nigerian women than in comparable pregnant United States patients. A statistically significantly higher alpha subunit level in the Nigerian patients was found only in the ten- to 13-week gestational period (P less than .005). The higher level of alpha subunit in pregnancy in Nigerian women may signal a population of trophoblastic cells which may be at higher risk for malignancy development in the Nigerian woman.

Trophoblastic neoplasms: occult disease.

Occult choriocarcinoma may be associated with low or undetectable levels of human chorionic gonadotropin. This article reviews the various methods of identifying the foci of choriocarcinoma that are currently available.

Gestational trophoblastic disease and human chorionic gonadotropin measurement.

Improved radioimmunometric assays for the glycoprotein human chorionic gonadotropin, its whole molecule and free beta and alpha subunits have improved the capability for trophoblast tumor detection and monitoring. New heights in survival rates have been reached with these improvements, particularly in high-risk disease.

The hCG assay in the treatment of trophoblastic disease.

The most important criterion in monitoring the treatment of patients with trophoblastic disease is the quantitative measurement of hCG levels. It is particularly important to have a sensitive hCG-detection procedure that reliably distinguishes low positive hCG levels from negative ones. Reliable monitoring of serum hCG levels minimizes unnecessary chemotherapy in patients entering remission and provides an early indication for additional chemotherapy if and when the hCG levels again become detectable. We evaluated a two-site monoclonal-antibody immunoradiometric assay (IRMA) for its reliability in the management of trophoblastic disease. The assay utilizes a solid-phase, monoclonal antibody that recognizes the alpha subunit of complete hCG and a 125I monoclonal antibody that recognizes the beta subunit. The minimal effective concentration of hCG detectable with the IRMA is 3-4 mIU/ml. The study population consisted of 6 choriocarcinoma patients, 49 hydatidiform mole patients, 37 patients being monitored after spontaneous abortion or blighted ovum and 80 patients in whom the possibility of trophoblastic disease was being ruled out. Each serum specimen was analyzed with the IRMA and in one or more of a number of different radioimmunoassays. The results were correlated with the patient's clinical course. Of the procedures evaluated, the IRMA was the most reliable in identifying trophoblastic-disease patients who required additional chemotherapy. Furthermore, the IRMA yielded no persistent low positive hCG values in patients without clinical evidence of trophoblastic disease. Therefore, the two-site IRMA is recommended for accurately distinguishing a clinically relevant low positive hCG level from an undetectable one.

Bacillus Calmette-Guérin immunotherapy in ovarian cancer.

Forty-six advanced ovarian cancer patients treated with conventional modalities with the addition of bacillus Calmette-Guérin (BCG) immunotherapy showed prolonged survival when compared to controls not given BCG. Although the data suggest enhancement of survival with the addition of BCG to conventional treatment, the fact remains that disease recurrence ultimately claims the lives of most of these patients. Nonetheless, patients are surviving longer in the face of advanced disease.

Endometriosis with a pleural effusion and ascites. Report of a case treated with nafarelin acetate.

BACKGROUND: Although endometriosis is generally confined to the pelvis, it may occur at remote sites with unusual manifestations. Rare examples include pulmonary endometriosis and endometriosis associated with ascites. These complications represent diagnostic and therapeutic dilemmas, especially when both occur concurrently in the same patient. CASE: A 21-year-old nulligravida had a history of endometriosis and complained of severe dyspnea and weight loss. Pleural effusion and pelvic ascites associated with recurrent endometriosis were found. The patient was treated with thoracentesis and nafarelin acetate nasal spray. A remarkable improvement in her condition occurred, and the side effects of the spray were minimal during treatment. CONCLUSION: Gonadotropin-releasing hormone agonist therapy is recommended as the initial treatment for endometriotic ascites and/or pleural effusion.