Expression of human placental-type alkaline phosphatase in primary breast cancer

A monoclonal antibody (H317) with specificity for the heat-stable placental-type alkaline phosphatase (PI-ALP) isoenzyme has been used to investigate the occurrence of PI-ALP in patients with primary breast carcinoma. All pre-operative plasma samples were negative for PI-ALP in a sensitive solid-phase enzyme immunoassay with a lower limit of detection of 0.1 U/l. In contrast, using a peroxidase-anti-peroxidase staining technique on fixed tissue sections, as well as enzyme immunoassay on fresh tissue extracts, PI-ALP could be demonstrated in the carcinomatous tissue of all seven patients investigated.     

Detection of placental-type alkaline phosphatase in ovarian cancer

A monoclonal antibody, H317, has been used for the sensitive and specific detection of placental-type alkaline phosphatase (PLAP) in sera, solubilized tissue extracts and fixed tumour tissue sections from patients representing a variety of ovarian tumours. PLAP was detected in over 30% of these sera and in most solubilized tumour tissue extracts. There was no association between circulating PLAP levels and either tissue extract levels or immunohistological staining of ovarian tumour tissue sections with H317. Nevertheless, immunohistology demonstrated the heterogeneity of cellular localization of PLAP within different tumours, and can often be of value in localizing tumour tissue.     

Electrophoretic heterogeneity of alkaline phosphatase isozymes in seminoma and normal testis

Electrophoretic patterns of seminoma- and normal-testis-derived alkaline phosphatase isozymes, the placental alkaline phosphatase (PLAP)-like enzyme and the tissue-nonspecific (liver) alkaline phosphatase (LAP), were studied on starch gel and isoelectric focusing (IEF). Different migration patterns of the PLAP-like enzyme were observed with respect to both seminomas and normal testes on starch gel electrophoresis. On IEF, seminomas showed different staining patterns among different tumors; however, a common main activity was focused at pIs of 4.3-4.6, corresponding to pIs of PLAP. Normal testes showed two enzyme-staining regions, at pIs of 4.1 and 5.0-5.2, which were discriminated from pIs of PLAP and the PLAP-like enzyme in seminoma. The PLAP-like enzyme in seminoma was differentiated from PLAP by digestion with neuraminidase. Neuraminidase treatment simplified the distribution patterns of the PLAP-like enzyme in normal testis, but did not alter the pattern of microheterogeneity in seminoma. Two factors other than sialylation, namely structural modification of the carbohydrate moiety and variation of hydrophobicity, were shown to contribute to the microheterogeneity of the PLAP-like enzyme in seminoma. LAP in seminoma and in normal testis also showed marked electrophoretic heterogeneity and differences in pI distributions from LAP of liver. However, the migration patterns after desialylation were very similar to each other. The findings imply that electrophoretic heterogeneity demonstrated in LAP in seminoma and in normal testis is caused by a difference in sialic acid content in the molecule, and the heterogeneity of the PLAP-like enzyme in seminoma is considerable.     

Phenotypes of placental-type alkaline phosphatase in seminoma sera as defined by monoclonal antibodies

PLAP-like enzymes could be detected in serum of patients with primary testicular tumors, in particular seminomas. The use of a panel of monoclonal antibodies (MAbs) permitted typing into 6 different testicular serum phenotypes, of which one appeared similar to a placental type (II) and 2 have not been previously described. Most tumor sera belonged to type I, as described for seminoma tissues. With a more advanced tumor the mean serum PLAP-like levels increased. After operation, after radiotherapy or with no evidence of disease lower or non-detectable enzyme levels were found. In typing the tissue PLAP-like antigen in serum of the same patient or sera from recurrences of a seminoma in the same patient, the same phenotypes of PLAP-like antigen were usually but not always found. None of the 6 phenotypes appeared to confer a poorer prognosis. We conclude that the expression of PLAP-like antigen is eutopic and is enhanced by testicular malignancy, especially in seminoma cells.     

Hydrophobicity and lectin affinity of alkaline phosphatase isozymes in seminoma and normal testis

Isozymes of alkaline phosphatases (ALP) in seminoma and normal testis were separated by use of high-performance liquid chromatography and a TSK-gel phenyl-5PW column. The tissue-nonspecific (liver) ALP (LAP) was the dominating isozyme, consisting of more than 90% ALP activity. The placental ALP (PLAP)-like enzyme contributed to 4-8% of the total ALP activity. The intestinal isozyme (IAP) could not be identified. The glycosylation patterns of the isozymes were studied using concanavalin A (Con A) affinity chromatography and batch elution with competing sugar. All PLAP activity in placental extracts and LAP activity in liver extracts was bound to Con A-Sepharose. In the tumor extracts, only 50-70% of the PLAP-like enzyme and 20-50% of the LAP activity from seminomas were bound to Con A-Sepharose. A similar binding pattern of the PLAP-like enzyme and LAP was also seen in the normal testes. This variability in Con A reactivity with PLAP or the PLAP-like enzyme was also reflected in serum of seminoma patients and of pregnant women. Thus, ALP expressed in seminoma has different lectin affinity characteristics compared with the same isozyme from placenta and liver, but almost identical to ALP in the normal testes. These findings imply that the PLAP-like enzyme and LAP in the testis can be discriminated from PLAP of placenta and LAP of liver by carbohydrate lectin affinity. It also supports the concept that the increased amounts of ALP in seminomas result from the enhanced eutopic expression of enzymes normally expressed in the testis.     

Immunohistochemical localization of placental alkaline phosphatase, carcinoembryonic antigen, and cancer antigen 125 in normal and neoplastic human lung

Human placental alkaline phosphatase (HPLAP), carcinoembryonic antigen (CEA), and cancer antigen 125 (CA 125) were localized immunohistochemically in paraffin sections of normal lung tissue from 16 patients, using monoclonal antibodies and an indirect avidin-biotin-peroxidase staining procedure. HPLAP and CEA were present in epithelial cells of respiratory bronchioli and alveolar type I pneumocytes. CEA was also observed in the tracheal, bronchial, and bronchiolar epithelium. CA 125 was present in the tracheal, bronchial, bronchiolar, and terminal bronchiolar epithelium; in the tracheal and bronchial glands; and in the pleural mesothelium. Normal and hyperplastic type II pneumocytes were negative for HPLAP, CEA, and CA 125 but were histochemically positive for nonspecific alkaline phosphatase. Fetal lung tissue between 11 and 15 weeks of gestation was negative for HPLAP, CEA, and CA 125. The fetal tracheal and bronchial epithelium, tracheal glands, and pleural mesothelium were positive for CA 125. For ten malignant pulmonary tumors investigated, HPLAP staining was observed in five, CEA in nine, and CA 125 in seven. The localization of HPLAP, CEA, and CA 125 in apparently normal constituents of all pulmonary specimens is in disagreement with the concept that the expression of these substances in the lung is indicative of abnormal cellular activity.     

Expression of a Nagao-type, phosphatidylinositol-glycan anchored alkaline phosphatase in human choriocarcinomas

The alkaline phosphatase (AP) synthesized by human tumor cells closely resembles human placental AP (PLAP). Little is known about the molecular events that lead to the expression of a placenta-like AP in tumor cells. The complementary DNA encoding the AP expressed by a choriocarcinoma cell line, BeWo, was isolated and characterized. The complementary DNA is the product of the germ cell AP (Nagao isozyme) gene and not of the term PLAP gene. Like placental AP, the tumor AP can be released from the cell membrane by a phosphaditylinositol-specific phospholipase C and has a phosphaditylinositol-glycan (PI-glycan) moiety at the COOH terminus. Immunoprecipitation of phosphaditylinositol-specific phospholipase C-treated AP and analysis by polyacrylamide gel electrophoresis or isoelectric focusing demonstrates that at least 95% of the AP contains PI-glycan. Two-dimensional gel electrophoresis reveals two precursors of the mature AP. One of these does not bind an antibody against the Trypanosoma variable surface glycoprotein cross-reacting determinant and probably does not contain PI-glycan. This precursor had a shorter half-life than the more prominent PI-glycan-containing precursor in pulse-chase experiments, suggesting a precursor-product relationship between the two proteins. These data demonstrate that BeWo AP is the product of a gene normally expressed in testis, thymus, and germ cells, but not in placenta. Thus, the expression of BeWo AP results from the repression of the PLAP gene and derepression of the germ cell AP gene and, as such, the expression is ectopic. The BeWo AP (Nagao isozyme) is modified with PI-glycan that is added soon after translation, not cotranslationally.     

Levels of alkaline phosphatase isozymes in human seminoma tissue

The three human isozymes of alkaline phosphatases were quantitatively determined in normal testis and seminoma tissues. The highly selective assays were based on isozyme specific monoclonal antibodies. In the normal testis approximately 90% of the catalytic activity originates from the tissue unspecific alkaline phosphatase, and the remaining activity was due to trace expression of both intestinal (approximately 5%) and placental alkaline phosphatase (PLAP) or PLAP-like isozyme (approximately 5%). In homogenates of seminoma tissues, highly increased levels of all three isozymes were identified. Both the tissue unspecific alkaline phosphatase and PLAP-like enzymes displayed relative increases of 10- to 100-fold and intestinal alkaline phosphatase 2- to 10-fold compared with normal testis. This finding indicates that the entire genome coding for alkaline phosphatases may be activated in seminomas. The PLAP-like enzyme from seminoma cells comprises a heterogenous population of molecules demonstrating partial heat sensitivity and microheterogeneity upon starch gel electrophoresis in contrast to the pregnancy related PLAP. These findings have implications for the different PLAP assays used in the clinical monitoring of seminoma patients.     

Demonstration of placental alkaline phosphatase in human breast cancer.

The screening of a series of 11 metastatic breast tumors for the presence of the placental isoenzyme of alkaline phosphatase (EC3.1.3.1) by RIA revealed one strong producer. The alkaline phosphatase of this tumor was characterized with respect to its immunochemical cross-reactivity, inhibition by L-phenylalanine and levamisole, subunit molecular weight (Mr) and isoelectric point (pl) in two-dimensional electrophoresis, and one-dimensional peptide map. In all parameters of the characterization, the tumor alkaline phosphatase, except for subunit molecular weight which was slightly lower (60,000 versus 64,000 for the placental isoenzyme). No strong placental alkaline phosphatase producers were found among 16 primary tumors examined by RIA. The screening of patients' sera for the placental alkaline phosphatase using RIA indicated elevated levels over post-menopausal controls in 20% of the metastatic patients. Only 3% of the primary patients had elevated serum levels. These results suggest that the placental isoenzyme of alkaline phosphatase may be a useful tumor marker for recurrent breast cancer.     

The occurrence of human placental alkaline phosphatase (PLAP) in extracts of normal, benign and malignant tissues of the female genital tract

In addition to its presence in extracts of carcinomatous tissues from the vulva, endometrium and ovary, PLAP can also be found in tissue extracts of benign conditions of the endometrium and myometrium. We detected slightly elevated levels of PLAP in patients with myoma, raised levels in 2/2 patients with endometrial polyps and high values in 2/2 patients with glandulocystic hyperplasia. Moreover, normal endometrium and fragments of normal Fallopian tube also contained fairly high amounts of endogenous PLAP. These results have important implications with regard to the possible use of PLAP as a tumour marker or of anti-PLAP monoclonal antibodies in radioimmunolocalization or radioimmunotherapy.     

Serum alpha-fetoprotein and variant alkaline phosphatase in human hepatocellular carcinoma

Alpha-fetoprotein and variant alkaline phosphatase were studied in a group of patients with hepatoma. The incidence of positive alpha-fetoprotein was 67.6% in confirmed cases. Variant alkaline phosphatase was present only in patients with positive fetoprotein test, and it appeared in only 28.6% of the cases.     

Identification and characterization of alkaline phosphatase isozymes in human colorectal adenocarcinomas

Using alkaline phosphatase isozyme-specific immunocatalytical assays, the content of isozymes was determined in normal mucosas and adenocarcinomas from human colon or rectum. Tumor levels of both the tissue (liver)-unspecific and the placental-like alkaline phosphatase (PLAP-like) were elevated compared to normal mucosas of the same patients. Such elevations have been reported previously, particularly in seminomas and ovarian tumors. In several tumors, moreover, the intestinal isozyme was expressed in lesser amounts than in the adjacent mucosa. The present results indicate that the activation of two of the phosphatase isozymes, including expression of the typical germ cell line phosphatase (the PLAP-like isozyme), may occur even in nongonadal tumors. This may reflect an induction pattern of phosphatase isozymes, with implications for malignant transformation also in other tumors.     

Carcinoma of the kidney, testis, and rare urologic malignancies

The purpose of this symposium was to provide a forum for discussion on current information on the etiology and diagnosis of, and therapy for, tumors of the kidney, testis, and several uncommon malignancies of the genitourinary tract. The most recent contributions in epidemiology and molecular genetics were discussed with specific reference to their importance for clinical practice. Contemporary treatment approaches with the emphasis on multidisciplinary patient management of tumors commonly seen in the clinic as well as those that are only rarely diagnosed by urooncologists were presented. Major stress was given to the management optimization as it pertains to short- and long-term quality of life issues of patients treated for these tumors. Methods to reduce treatment toxicity including carcinogenic potential of chemotherapy, radiotherapy, or their combination were found to be of nearly equal importance to patient survival. Symposium participants reached consensus on a number of important points: 1) The management of patients with several malignancies discussed requires the presence of a multidisciplinary team of specialist who are interested in diagnosis and treatment of genitourinary tumors; 2) Patients managed in such an environment are expected to have optimal survival and the best possible quality of life; 3) Real advances in the management of patients can be best obtained through well-designed prospective clinical trials; and 4) There is a need for timely introduction of relevant advances in epidemiology and molecular genetics to clinics.