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A procedure for the estimation of nuclear androgen receptors in benign prostatic hyperplastic tissue is described, which employs extraction of receptors from nuclei with buffers containing heparin. Extraction of a nuclear pellet with a heparin-containing (1 g/l) buffer appeared to have definite advantages over 0.4 mol/l KCl extraction. Heparin appeared to be twice as efficient in extracting androgen receptors. In addition aggregated receptor proteins, formed after storage at -80 degrees C, were partly deaggregated by heparin. Specific isolation of the androgen receptor was performed using either agar gel electrophoresis, protamine sulphate precipitation or LH-20 gel filtration. A comparison was made between the amounts of estimated receptors with these different techniques. Protamine sulphate precipitation resulted in the highest estimates of receptor-bound 5 alpha-[3H]dihydrotestosterone (3H-DHT). Treatment of the labelled nuclear extracts with a charcoal suspension prior to the receptor assay resulted in lower amounts of estimated androgen receptors. A method for routine evaluation of nuclear androgen receptors in prostatic tissue has been evaluated, which involves extraction of nuclear pellets with a heparin-containing (1 g/l) buffer, exchange labelling of the nuclear extracts for 20 h at 10 degrees C and quantification of the receptors with protamine sulphate precipitation.

Oestrogen (ER) and progestin receptors (PR) were assayed in human meningioma cytosol by radioligand binding assay with Scatchard plot analysis and by monoclonal antibody based enzyme immunoassays. For comparison, human breast cancer tissues were used. Results of both assays agreed very well. For human breast cancer, receptor levels assayed with both methods showed a highly significant (ER: r = 0.96; n = 74 and PR: r = 0.95; n = 19). Also for meningioma cytosols a good agreement was observed between the result of both assays. Thus, most meningiomas were devoid of ER but contained significant concentrations of PR. PR levels in meningioma determined with the enzyme immunoassay correlated well with those found by Scatchard plot analysis. After logarithmic transformation of the data the regression line was; PR(EIA) = 0.83 X PR(Scatchard) + 0.36 (r = 0.917; n = 24). The recognition of the progestin binder in meningioma by a monoclonal antibody against the progestin receptor is a further indication that progestin binding in meningioma occurs to a true receptor.

Prolactin is associated with the development of mammary tumors in rats. The aim of the present study was to evaluate whether strain differences in susceptibility to the development of mammary tumors could be explained by genetic differences in the response of the pituitary to chronic stimulation by estrogens. Prolactin levels were measured in plasma from rats of the Sprague-Dawley, Wistar WAG/Rij and Brown Norway BN/BiRij strains before and at different times after subcutaneous implantation of estradiol-17 beta in cholesterol/paraffin pellets. In all strains plasma prolactin was elevated from the second week after implantation of the pellet, although there were quantitative differences between the responses. At 32 weeks after implantation of the pellets the plasma level of prolactin in Sprague-Dawley rats was 1247 +/- 367 ng NIAMDD prolactin RP-1/ml (mean +/- S.E.M), whereas Wistar WAG/Rij and Brown Norway BN/BiRij had plasma prolactin levels of 679 +/- 211 and 182 +/- 19 ng/ml respectively. Between 52 and 104 weeks after implantation these values rose to 4016 +/- 1116, 5004 +/- 1053 and 808 +/- 129 ng/ml respectively. The plasma concentration of prolactin of rats in this age group was strongly associated with the occurrence of pituitary adenomas in all three strains. In untreated rats, the concentration of prolactin in the plasma increased with age to only 200-400 ng/ml at 12-24 months of age but no significant differences were observed between the three rat strains. It is concluded that observed differences in spontaneous and estrogen-mediated mammary tumor development in these rat strains cannot be explained by genetic differences in the plasma concentration of prolactin.

Three patients with metastatic prostatic cancer were treated for 10, 6 and 2 months with the potent LHRH-agonist Buserelin (Hoe 766) as a first-line agent. All showed a fall of elevated prostatic acid phosphatase levels (nearly undetectable after treatment in 2 patients) parallel to plasma testosterone with a relief of complaints after 3-4 weeks of treatment. Two patients had an increment of appetite and body weight. In one patient radiological evidence for objective tumour regression was found by CT scan of the prostate (decrease of 41% in prostate volume), skeletal X-rays and bone scan. In this patient plasma alkaline phosphatase showed a transient increase parallel to disappearance of osteolytic bone lesions (indicating new bone formation) followed by a normalization. It is concluded that LHRH-agonist treatment is effective in patients with metastatic prostatic carcinoma in the absence of serious side-effects.

Because of a well-established mechanism of action, tissue concentrations of steroid hormones are thought to be more closely related than blood levels to the biological effects exerted by these hormones. The results of studies on oestrogen and androgen concentrations in malignant and normal breast tissues are presented. Normal fatty and epithelial breast tissues and malignant tumour samples which had been obtained from pre- and postmenopausal women of two countries (Poland and The Netherlands) differing in the incidence of this malignancy were studied. In both countries highly comparable oestradiol concentrations in the breast were found. The median hormone levels in tumour tissue of 0.65 pmol/g tissue did not change with age. They were significantly higher than in normal epithelial (0.48 and 0.25 pmol/g in pre- and postmenopausal women) and fatty tissues (0.54 and 0.19 pmol/g respectively). Particularly in postmenopausal women, hormone levels in tumour tissue were much higher than plasma concentrations, which are comparable in both populations. Oestrone levels decreased with age in normal and malignant breast tissues. In both countries median levels in normal and fatty tissues of premenopausal women were similar (1.10 pmol/g tissue) but higher than those in postmenopausal patients (0.45 pmol/g tissue. Significantly lower levels were found in the malignant tissue samples of Polish premenopausal women (0.70 pmol/g) than in Dutch women (1.05 pmol/g); similarly, after menopause the tissue concentrations were higher in Dutch (0.55 pmol/g) than in Polish (0.31 pmol/g) patients. Thus lower oestrone tissue levels were observed in tumours from the country with the lower incidence for breast cancer. In a comparable study of uterine tissues, obtained from pre- and postmenopausal women, higher oestradiol concentrations than in the breast were found, whereas estrone levels were very similar. The levels in the uterus did not correlate with those in the plasma; no relation with histology was observed. The results of androgen measurements in breast tissues were in agreement with the concept that, particularly, androstenedione and testosterone could play a role as substrates for local aromatization. Lower concentrations were observed in the tumours than in the normal and fatty tissues. More extensive investigations will be needed to clarify the role of local formation (aromatization, hydrolysis by sulphatase) of oestrogens in tissues and of the interconversion of less active (oestrone) to more active (oestradiol) oestrogens.

In the present study the occurrence of estrogen and progestin receptors in the lymphoid tissue of 20 canine non-Hodgkin's Lymphoma patients was investigated. Lesions in all dogs were histologically classified according to criteria used in human patients. No progestin receptors could be demonstrated. Estrogen receptors were found only in the cytosol of two canine lymphomas. Antiestrogen binding sites were present in all 18 cases examined (range 37-356, median 110 fmol/mg protein). No correlation with age, sex or histological classification was found. A pilot study was carried out in five of these dogs that were negative for estrogen receptors and positive for antiestrogen binding sites. These dogs were treated with the antiestrogen tamoxifen (20-30 mg/day) for 2-3 weeks, but the treatment had no apparent effect. It was concluded that if antiestrogens do play a role in the treatment of non-Hodgkin's lymphoma in the dog, it is most likely not by means of antiestrogen binding sites.

We studied oestrogen binding sites in blood mononuclear cells from healthy blood donors, patients with leukaemia or systemic lupus erythematosus, and in thymocytes, using the dextran-coated charcoal assay and Scatchard analysis of binding data. Using 3H-labelled oestradiol as ligand, inaccurate results were obtained which could be related to the low amounts of binding sites. Using 125I-labelled ligand, saturable oestradiol binding sites could be demonstrated in low amount (mean value 2.1 fmol/mg of cytosolic protein) and high affinity (mean Kd value 0.26 nM; mean Ka value 3.85 X 10(9) M-1). The binding could be inhibited by unlabelled oestradiol but not with oestrone, dihydrotestosterone, cortisol and the progestin-receptor ligand Org 2058. We conclude that blood mononuclear cells and thymocytes contain true oestrogen receptors. This conclusion supports current hypotheses on the involvement of such receptors in oestrogen-mediated modulation of the immune system.

A case-control study incorporated within a cohort study was undertaken to test the hypothesis that anovulation or decreased luteal function is associated with increased risk of breast cancer. A large population (n = 12,000) of apparently healthy women aged 40-49 collected a specimen of urine on day 22 of 3 consecutive menstrual cycles, or, in cases of amenorrhea, on 3 arbitrarily chosen days. These samples were stored at -20 degrees C. Subsequent screening of the women by mammography revealed 34 cases of breast cancer, and over the next 4 years a further 34 cases were reported to the cancer registry. Urine samples from 53 of these cases were assayed for pregnanediol and creatinine, and the results were compared with those for an equal number of matched controls. No differences between these groups were found in the pregnanediol/creatinine ratio. We conclude that our results do not support the hypothesis that women with luteal insufficiency are at increased risk for breast cancer. However, since breast cancer patients tend to have a later menopause than controls, the possibility cannot be excluded that breast cancer patients continue to have ovulatory cycles for a longer period during reproductive life. Such a finding--opposite to the working hypothesis--would complicate the interpretation of the results.

Estrogen and progestin receptor levels (ER and PgR) in tumors from 506 patients with primary breast cancer diagnosed in 1979, 1980, and 1981 were measured by a Scatchard plot analysis. At a median follow-up time of 3.5 years the prognostic value of the receptor levels was evaluated and compared with other tumor and patient characteristics. No relation was found between receptor levels and tumor, lymph node, metastasis (TNM) classification or location of the primary tumor. A significant positive rank correlation was observed between ER and PgR levels (rs = 0.57) and between ER level and age of the patients (rs = 0.39, P less than 0.001). The observed association between ER level and menopause status could not be maintained after correction for age. Independent prognostic factors for overall survival were tumor size (P = 0.002), the number of positive lymph nodes (P less than 0.001), age at primary surgery (P less than 0.001), the PgR level of the tumor (P less than 0.001), but not ER level. Independent prognostic factors for relapse were tumor size (P = 0.003), number of positive lymph nodes (P less than 0.001), age (P = 0.006), menopause status (P = 0.02), PgR level (P = 0.007), but not ER level. Finally, for death rate after relapse the following prognosticators were identified: size of the primary tumor (P = 0.03), number of positive lymph nodes (P = 0.03), age (P = 0.003), site of relapse (P less than 0.001), ER level (P = 0.02), and PgR level (P = 0.04). Patients with tumors containing low positive PgR levels (10 to 20 fmol/mg protein) had a slightly better prognosis than patients with PgR-negative tumors. It is concluded that the PgR level of the primary tumor is a better prognosticator than the ER level. The ER offered no additional ability for discriminating between low- and high-risk patients once PgR was included in the model. In contrast, PgR was capable of improving on the discriminating ability of ER. In addition, patients with tumors containing both PgR and ER showed the best prognosis. Therefore, it is recommended that ER and PgR should be assayed in all breast cancer biopsies.

The occurrence of oestrogen and progestin receptors in cytosols from human intracranial meningiomas was studied with a dextran-coated charcoal assay and Scatchard plot analysis. [3H]-Oestradiol and [3H]-promegestone (17 alpha, 21-dimethyl-19-norpregna-4,9-diene-3,20-dione, R-5020) were used as tracers. Using this method, no high-affinity binding sites for oestradiol were observed, whereas progestin binding was identified in 18 out of 20 meningioma cytosols. The number of progestin binding sites was identical in meningioma cytosols obtained from female patients (192 +/- 57 fmol/mg protein, mean +/- S.E.M., n = 12) and those obtained from male patients (230 +/- 57 fmol/mg protein, n = 6), as was the dissociation constant of the complex (1.5 +/- 0.3 vs 1.4 +/- 0.3 nmol/l respectively). Only progestins (progesterone, R-5020 and megestrol acetate) competed successfully with tritiated R-5020. Oestrogens, androgens and cortisol showed no appreciable cross-reaction. It was concluded that the cytosols from human intracranial meningiomas contain progesterone receptors in the absence of oestrogen receptors. The presence of these progesterone receptors may indicate that (anti)-progestational treatment could be of potential value in cases which cannot be treated by surgery alone.