Estrone sulfate and sulfatase activity in human breast cancer and endometrial cancer

Estrone sulfate (E1-S) in the serum and tissues of patients with breast cancer or endometrial cancer was measured by a direct radioimmunoassay without hydrolysis. The concentration of E1-S in breast cancer tissue was 1.64 +/- 0.28 ng/g wet wt (+/- SE), lower than in surrounding normal breast tissue (4.46 +/- 1.23). Estradiol-17 beta(E2)/E1-S was higher in endometrial cancer tissue than normal endometrial tissue. Estrone sulfatase activity in breast cancer tissue was 0.81 +/- 0.23 nmol/h/mg protein, higher than in surrounding normal breast tissue (0.35 +/- 0.11). These results suggest that E1-S, which is abundant in the peripheral circulation, is hydrolyzed by sulfatase in breast cancer tissue or endometrial cancer tissue and liberates free estrogens, which may stimulate the growth of these malignant tumors.     

Steroid sulfate sulfatase in human benign prostatic hyperplasia: characterization and quantification of the enzyme in epithelium and stroma

Characteristics and activities of estrone sulfate (E1S) and dehydroepiandrosterone sulfate (DHAS) sulfatases were studied in epithelium and stroma of benign hyperplastic tissues from human prostates. Tissues were obtained by suprapubic prostatectomy, and epithelium and stroma were separated mechanically by standard techniques. The assay procedure comprised homogenization in Tris-buffer, incubation of the homogenate with [3H]E1S or [3H]DHAS, separation of free steroids from nonhydrolyzed steroid sulfates by extraction with ether, and their final quantification by LSC. The main results were: (1) The pH-optimum of the sulfatase was found at pH 7.0. (2) The highest specific sulfatase activity was found in the epithelium and was associated with its nuclear fraction. (3) Michaelis-Menten constants Km (microM) were 8.7 +/- 1.4 (7) and 4.3 +/- 0.8 (5), maximum velocity rates Vmax (nmol/h x mgDNA) were 47.4 +/- 8.8 (7) and 8.4 +/- 1.5 (5) for E1S and DHAS, respectively (means +/- SEM (n]. (4) The enzymatic cleavage of E1-sulfate was competitively inhibited by DHA-sulfate and vice versa with inhibition constants Ki (microM) of 4.0 +/- 0.5 (2) for E1S and 2.7 +/- 0.4 (2) for DHAS. On the basis of these findings, possible roles of steroid sulfate-sulfatases in forming precursors of active androgens and estrogens from the high amounts of E1S and DHAS in blood are discussed.     

Uterine estrogen sulfatase activity at the time of blastocyst implantation in the rat

The conversion of estrone sulfate (E1S) to estrone (E1) was measured during the in vitro incubation of the labeled sulfoconjugate with implantation sites (IS) and nonimplanted regions (NIS) of uterine horns from 6-day pregnant rats. Extensive metabolism of E1S occurred in both tissues, being noticeably less (29.31%) in IS than in NIS. Estrogen sulfatase activity present in the uterus of ovariectomized virgin rats was found to be higher than in both uterine regions of the pregnant rats. We suggest that E1S present in uterine fluids may be accessible to be metabolized into unconjugated estrogens by both intrauterine tissues of 6-day pregnant rats. This metabolism could be locally modulated in IS through the participation of the estrogen sulfatase, the activity of which is in turn controlled by the presence of free estrogens, possibly synthesized and/or secreted by the embryo, which has been shown to inhibit the sulfohydrolase activity.     

Estrogen metabolism in neural tissues of rabbits: 17 beta - hydroxysteroid oxidoreductase activity.

Incubation of adult rabbit neural tissue homogenates with (3H)-estrone and (3H)-estradiol revealed that the conversion of estrone to estradiol is higher in both male and female animals than estradiol to estrone. Both 17 oxidation and reduction are higher in male animals than in females. However, it is observed that the quotient of estrone leads to estradiol/estradiol leads to estrone for pituitary tissue and hypothalamus are higher in females than in males. There is no such dimorphism in cerebral cortex. The overall metabolism in pituitary is higher than in hypothalamus and cortex in both sexes. These results suggest that 17 beta hydroxysteroid oxidoreductase activity may play an important role in the regulation of estrogen function in neuroendocrine tissues.     

Norelgestromin as selective estrogen enzyme modulator in human breast cancer cell lines. Effect on sulfatase activity in comparison to medroxyprogesterone acetate

Human breast cancer tissue contains enzymes (estrone sulfatase, 17beta-hydroxysteroid dehydrogenase, aromatase) involved in the last steps of estradiol (E(2)) formation. In this tissue, E(2) can be synthesized by two main pathways: (1) sulfatase-transforms estrogen sulfates into bioactive E(2), and the (2) aromatase-converts androgens into estrogens. Quantitative assessment of E(2) formation in human breast tumors indicates that metabolism of estrone sulfate (E(1)S) via the sulfatase pathway produces 100-500 times more E(2) than androgen aromatization.In the present study, we demonstrated in T-47D and MCF-7 human breast cancer cells that norelgestromin (NGMN) (a metabolite of norgestimate) is a potent inhibitory agent of the estrone sulfatase activity. After 24h incubation of physiological concentrations of E(1)S (5 x 10(-9)mol/l) the inhibitory effect of NGMN at concentrations of 5 x 10(-9), 5 x 10(-7) and 5 x 10(-5)mol/l was 43+/-7, 74+/-4 and 97+/-2%, respectively, in T-47D cells; 25+/-4, 57+/-5 and 96+/-2% respectively, in MCF-7 cells. Comparative studies using medroxyprogesterone acetate (MPA) showed that this progestin also has an inhibitory effect on sulfatase activity, but significantly less intense than that of NGMN. The inhibition for MPA at concentrations of 5 x 10(-9), 5 x 10(-7) and 5 x 10(-5)mol/l was 31+/-5, 47+/-3 and 61+/-3%, respectively, for T-47D cells; 6+/-3, 20+/-3 and 63+/-4%, respectively, for MCF-7 cells.In conclusion, the present data show that NGMN is a very potent inhibitory agent for sulfatase activity in the hormone-dependent breast cancer cells, resulting in decreased tissue concentration of E(2). The clinical significance of this finding remains to be elucidated.     

Comparison of estrogen concentrations, estrone sulfatase and aromatase activities in normal, and in cancerous, human breast tissues

In the present study, the concentrations of estrone (E(1)), estradiol (E(2)) and their sulfates (E(1)S and E(2)S), as well as the sulfatase and aromatase activities, were evaluated in post-menopausal patients with breast cancer. Comparative studies of the evaluation of these parameters were carried out in (a) tumor tissue, (b) areas surrounding the tumor, and (c) areas distant from the tumor (glandular tissue) which were considered as normal tissue. The levels (in pm/g; mean +/- SEM) were: for E(1) in the (a) area: 320+/-95; in (b): 232+/-86; and in (c): 203+/-71; for E(2) in the (a) area: 388+/-106; in (b): 224+/-48; and in (c): 172+/-80; for E(1)S in the (a) area: 454+/-110; in (b): 259+/-90; and in (c): 237+/-65; for E(2)S in the (a) area:318+/-67; in (b): 261+/-72; and in (c): 232+/-75, respectively. The values of E(1)S and E(2) were significantly higher in the tumor tissue than in the area considered as normal. In all the tissues studied, the sulfatase activity was much higher than aromatase (130-200). In addition, the sulfatase levels were significantly higher in the peripheral and in the tumor tissue than in the area considered as normal. The levels of aromatase were significantly higher in tumoral than in normal tissue. The present data extend the "intracrine concept" for breast cancer tumors. The physiopathology and clinical significance as promoter parameters in breast cancer is to be explored.     

Paradoxical effect of estradiol: it can block its own bioformation in human breast cancer cells

The great majority of breast cancers are in their early stage hormone-dependent and it is well accepted that estradiol (E(2)) plays an important role in the genesis and evolution of this tumor. Human breast cancer tissues contain all the enzymes: estrone sulfatase, 17 beta-hydroxysteroid dehydrogenase (17 beta-HSD), aromatase, involved in the last steps of E(2) bioformation in this tissue. Quantitative data show that the 'sulfatase pathway', which transforms estrogen sulfates into the bioactive unconjugated E(2), is 100-500 times higher than the 'aromatase pathway' which converts androgens into estrogens. In this paper we explore the effect of E(2) on the sulfatase activity using two hormone-dependent human breast cancer cells: MCF-7 and T-47D. The action of E(2) on the sulfatase activity was evaluated by the conversion of estrone sulfate (E(1)S) into E(2). The cells were incubated in Minimal Essential Medium (MEM) containing 5% steroid-depleted fetal calf serum and incubated with physiological concentrations of [(3)H]E(1)S (5 x 10(-9) M) alone (control) or in the presence of E(2) (5 x 10(-10) to 5 x 10(-5) M) for 24 h at 37 degrees C. It was found that E(2) is a potent inhibitory agent of the estrone sulfatase activity in both cell lines. A low concentration of E(2): 5 x 10(-9) M decreases the sulfatase activity by 67% in MCF-7 cells and 57% in T-47D cells. More than 80% of the decrease in the formation of E(2) was obtained with the dose of 5 x 10(-7) M in both cell lines. It is concluded that this paradoxical effect of E(2) adds a new biological response of this hormone and could be related to estrogen replacement therapy in which it was observed to have either no effect or to decrease breast cancer mortality in postmenopausal women. Preliminary results are indicated in the Proceedings of the 14th International Symposium of the Journal of Steroid Biochemistry & Molecular Biology (Quebec, Canada, 24-27 June 2000) [J. Steroid Biochem. Molec. Biol. 76 (2001) 95-104](1) and presented at the 83rd Annual Meeting of the Endocrine Society (Denver, USA, 20-23 June 2001 (abstract no. P2-615).     

The Distribution of Estrone Sulphatase, Dehydroepiandrosterone Sulphatase, and Arylsulphatase C in the Primate (Macaca radiata) Brain and Pituitary

Abstract: Estrone sulphatase, arylsulphatase C and dehydroepiandrosterone sulphatase were measured in the pituitary, hypothalamus-preoptic area, amygdala, hippocampus, midbrain, septum, frontal cortex and occipital cortex of monkey ( Macaca radiata ) brain. All the regions showed measurable activities of all three enzymes. In all the animals tested, either the midbrain or hypothalamus-preoptic area showed the greatest activity of all three enzymes. In particular, estrone sulphatase showed the highest specific activity in either of the above two regions in all animals. Subcellular distribution studies in the hypothalamus preoptic area showed a similarity in the distribution profile between arylsulphatase C and estrone sulphatase and a significant difference of dehydroepiandrosterone sulphatase from the others.     

Inhibition of steryl sulfatase activity in LNCaP human prostate cancer cells

The enzyme steryl sulfatase may help support the growth of hormone-dependent tumors, including prostate cancers, by facilitating the conversion of circulating precursor steroids to active hormones. We sought to determine the presence of steryl sulfatase activity in the androgen-dependent human prostate cancer cell line LNCaP, and to determine if this activity was inhibited by known steryl sulfatase inhibitors. Intact LNCaP cultures had steryl sulfatase activity, as determined by conversion of [3H]estrone sulfate (E(1)S) to unconjugated steroids. The level of steryl sulfatase activity was relatively low (4.6 pmol/18 h/million cells) compared to MDA-MB-231 breast cancer cells (284.0 pmol/18 h/million cells). The observed activity in both cell lines was blocked by addition of 1 microM estrone sulfamate (EMATE), an active-site-directed, steroidal inhibitor of steryl sulfatase. Steryl sulfatase activity was also inhibited by Danazol, and by (p-O-sulfamoyl)-tetradecanoyl tyramine (C2-14), a non-steroidal inhibitor. Microsomes prepared from LNCaP cultures also showed steryl sulfatase activity, as determined by hydrolysis of [3H]E(1)S and [3H]dehydroepiandrosterone sulfate (DHEAS) to unconjugated forms. LNCaP and MDA-MB-231 microsomes both hydrolyzed E(1)S about two times faster than DHEAS. Hydrolysis of E(1)S in LNCaP and MDA-MB-231 microsomes was blocked by steryl sulfatase inhibitors with the following relative potencies: EMATE>C2-14>Danazol. These data demonstrate that LNCaP prostate cancer cells contain a steryl sulfatase with properties similar to that found in human breast cancer cells, and that the activity of this enzyme can be blocked by known steryl sulfatase inhibitors. Steryl sulfatase inhibitors may be useful as an adjuvant to androgen deprivation therapy for prostate cancer.     

Estrone sulfatase activity in rat brain and pituitary: effects of gonadectomy and the estrous cycle

Estrone sulfatase activity was characterized in microsomal preparations from rat brain and anterior pituitary. No differences in apparent Km were found in hypothalamic-preoptic area between male (7.5 microM) and female (7.4 microM) rats. Apparent Km's of anterior pituitaries from males (14.5 microM) and females (22.5 microM) were higher than those found in brain. Estrone sulfatase activity was equally inhibited by estradiol-17 beta-3-sulfate, dehydroepiandrosterone-3-sulfate and estrone-3-sulfate indicating a broad range of substrate specificity for this enzyme. Sulfatase activity in female anterior pituitary was found to be twice that of male. Sulfatase activity was distributed similarly in brain tissues between sexes with cerebellum greater than or equal to medial basal hypothalamus greater than preoptic area = cortex. Following gonadectomy, sulfatase activity in anterior pituitary of males was significantly greater than activity found in intact animals (P less than 0.05). This increase in activity, however, was unaffected by treatment with testosterone, dihydrotestosterone or estradiol-17 beta. Gonadectomy did not change sulfatase activity in brains of males or females or in pituitaries of females. However, sulfatase activity in pituitary glands of females changed significantly (P less than 0.05) with stages of the estrous cycle (metestrus less than diestrus less than proestrus less than estrus). These data indicate sulfatase activity in rat anterior pituitary gland may be controlled by gonadal factors while sulfatase activity in brain is regulated differently.     

Effect of nomegestrol acetate on estrogen biosynthesis and transformation in MCF-7 and T47-D breast cancer cells

Although ovaries serve as the primary source of estrogen for pre-menopausal women, after menopause estrogen biosynthesis from circulating precursors occurs in peripheral tissues by the action of several enzymes, 17beta-hydroxysteroid dehydrogenase 1 (17beta-HSD1), aromatase and estrogen sulfatase. In the breast, both normal and tumoral tissues have been shown to be capable of synthesizing estrogens, and this local estrogen production can be implicated in the development of breast tumors. In these tissues, estradiol (E(2)) can be synthesized by three pathways: (1) estrone sulfatase transforms estrogen sulfates into bioactive estrogens, (2) 17beta-HSD1 converts estrone (E(1)) into E(2), (3) aromatase which converts androgens into estrogens is also present and contributes to the in situ synthesis of active estrogens but to a far lesser extent than estrone sulfatase. Quantitative assessment of E(2) formation in human breast tumors indicates that metabolism of estrone sulfate (E(1)S) via the sulfatase pathway produces 100-500 times more E(2) than androgen aromatization. Breast tissue also possesses the estrogen sulfotransferase involved in the conversion of estrogens into their sulfates that are biologically inactive. In the present review, we summarized the action of the 19-nor-progestin nomegestrol acetate (NOMAC) on the sulfatase, 17beta-HSD1 and sulfotransferase activities in the hormone-dependent MCF-7 and T47-D human breast cancer cell lines. Using physiological doses of substrates NOMAC blocks very significantly the conversion of E(1)S to E(2). It inhibits the transformation of E(1) to E(2). NOMAC has a stimulatory effect on sulfotransferase activity in both cell lines, with a strong stimulating effect at low doses but only a weak effect at high concentrations. The effects on the three enzymes are always stronger in the progesterone-receptor rich T47-D cell line as compared with the MCF-7 cell line. Besides, no effect is found for NOMAC on the transformation of androstenedione to E(1) in the aromatase-rich choriocarcinoma cell line JEG-3. In conclusion, the inhibitory effect provoked by NOMAC on the enzymes involved in the biosynthesis of E(2) (sulfatase and 17HSD pathways) in estrogen-dependent breast cancer, as well as the stimulatory effect on the formation of the inactive E(1)S, can open attractive perspectives for future clinical trials.     

Evidence for sulfatase and 17beta-hydroxysteroid dehydrogenase type 1 activities in equine epididymis and uterus

Our previous work showed that stallion testis produces high amounts of estrogens which are subsequently found in the ejaculate. These estrogens are mainly synthesized by testicular aromatase, and the major estrogen produced is estrone sulfate (E1S). The objective of this study was to investigate the potential role of E1S as a source of estrogens in the male and female horse reproductive tracts by determining whether both estrone sulfatase (Sulf) and 17beta-hydroxysteroid dehydrogenase type I (17beta-HSD1) activities were present in equine testes, epididymis and uterus. We assessed E1S bioconversion into estrone (E1) and estradiol (E2) in these tissues. Both Sulf and 17beta-HSD1 activities were well detected in the cauda epididymis and uterus. Additionally, Sulf activity was present in the distal corpus of the epididymis, and 17beta-HSDI in the proximal corpus. In contrast, aromatase gene expression, measured as an internal control of endogenous estrogen production, had high activity only in the testis. We found that seminal E1S of testicular origin can be metabolized to E2, especially in the cauda epididymis and uterus. Because E2 appears to play a major role in male and female reproduction, we propose that the bioconversion of seminal E1S could affect male and female fertility.     

17-oxidoreduction of 17beta estradiol, estrone and their 3-sulfates by kidney slices from guinea pig and human.

Slices of whole kidney and kidney cortex from the female guinea pig catalyzed a marked reduction of estrone 3-sulfate (E13S) and estrone (E1) to 17beta-estradiol 3-sulfate (E23S) and 17beta-estradiol (E2), respectively, as well as the reverse (dehydrogenation) reactions. Slices of medulla did not appear active in E23S-E13S interconversion but did possess the ability to interconvert E2 and E1, besides possessing considerable sulfatase activity. The use of [3H-55S]E13S and [3H-55S]E23S as substrates, together with a demonstrated lack of estrogen sulfate synthesis by the tissue slices, provided ample evidence that the intact sulfates were involved in direct oxidoreduction. Slices of human kidney cortex catalyzed the reduction of E13S to a very limited extent. Slices of whole kidney and of cortex from guinea pig formed small amounts of estrogen glucuronide(s).     

In vitro and in vivo models for the evaluation of new inhibitors of human steroid sulfatase,devoid of residual estrogenic activity

The goal of our research project is to develop a new class of orally active drugs, estrone sulfatase inhibitors, for the treatment of estrogen-dependent (receptor positive) breast cancer. Several compounds were synthesized and their pharmacological potencies explored. Based on encouraging preliminary results, three of them, TX 1299, TX 1492 and TX 1506 were further studied in vitro as well as in vivo. They proved to be strong inhibitors of estrone sulfatase when measured on the whole human JEG-3 choriocarcinoma and MCF-7 breast cancer cells and their IC(50)s found to be in the range of known standard inhibitors. Their residual estrogenic activity was checked as negative in the test of induction of alkaline phosphatase (APase) activity in whole human endometrial adenocarcinoma Ishikawa cells. In addition, their effect on aromatase activity in JEG-3 cells was also examined, since the goal of inhibiting both sulfatase and aromatase activities appears very attractive. However, it has been unsuccessful so far. Then, in vivo potencies of TX 1299, the lead compound in our chemical series, were evaluated in comparison with 6,6,7-COUMATE, a non-steroidal standard, in two different rat models and by oral route. First, the absence of any residual estrogenic activity for these compounds was checked in the uterotrophic model in prepubescent female rats. Second, antiuterotrophic activity in adult ovariectomized rat supplemented with estrone sulfate (E(1)S), showed that both compounds were potent inhibitors, the power of TX 1299 relative to 6,6,7-COUMATE being around 80%. This assay was combined with uterine sulfatase level determination and confirmed the complete inhibition of this enzyme within the target organ.Preliminary studies indicated that other non-steroid compounds in the Théramex series were potent in vitro and in vivo inhibitors of estrone sulfatase in rats and further studies are in progress.     

Steroid sulfatase and 17 beta-hydroxysteroid oxidoreductase activities in mouse tissues

The metabolism of estrone sulfate and dehydroisoandrosterone sulfate to the free, unconjugated steroids, estrone and dehydroisoandrosterone, was demonstrated in more than thirty different tissues from male and female BALB/c mice. The activity of steroid sulfatase, when expressed per mg tissue, was greatest in both the pituitary gland and the adrenal glands. The pituitary gland, however, had the lowest capacity for hydrolysis of steroid sulfates while the liver had the greatest capacity. 17 beta-Hydroxysteroid oxidoreductase activity also was demonstrated in all mouse tissues by the formation of estradiol-17 beta when using estrone sulfate as the substrate. The highest apparent activity for 17 beta-hydroxysteroid oxidoreductase was found in lung tissue, and the greatest capacity to form estradiol-17 beta from estrone sulfate was found in liver, lungs, kidneys and testes. This study demonstrates that the majority of mouse tissues have steroid sulfatase and 17 beta-hydroxysteroid oxidoreductase activities.     

Steroid sulfatase activity and expression in mammary myoepithelial cells

PURPOSE: This investigation examined mRNA expression and enzymatic activity of steroid sulfatase (STS) in human mammary myoepithelial cells (MMECs) and MCF-7 cells and assessed the effects of 17-beta estradiol on the activity of STS. METHODS: The mRNA level of STS in MMECs was determined by RT-PCR analysis using specific primers for STS. STS enzymatic activity prior to and after treatment with 17-beta estradiol was determined by measuring 3H-metabolites formed after exposure to [3H]estrone 3-sulfate (E1S) and [3H]dehydroepiandrosterone-sulfate (DHEA-S). RESULTS: Our data demonstrate the presence of STS in the MMECs. Based on RT-PCR analysis, MMECs had slightly lower levels of STS compared to MCF-7 cells. However, sulfatase activity was about 120 times greater in the MMECs than the MCF-7 cells (E1S V(max)=2640nmol/(mg DNAh) compared to 20.9nmol/(mg DNAh)). Exposure to 17-beta estradiol was associated with 70% reduction in E1S sulfatase activity in the MCF-7 cells and 9% increase in the MMECs after 6 days. DISCUSSION: Our studies indicate for the first time the presence of STS in MMECs. This is suggestive of a previously undetermined role for MMECs in converting precursor hormones into active steroid hormones within mammary tissue. In addition, differential response of the MMECs and the MCF-7 cells to estrogen demonstrates differences in hormone metabolism between these two cell types, perhaps related to the absence of estrogen receptors in the MMECs and their presence in the MCF-7 cells. The MMECs may have an important role in hormonal regulation within mammary tissue.     

Estrone sulfate sulfohydrolase in the developing brain and pituitary of rat, mouse and guinea pig

The pattern of estrone sulfate sulfohydrolase (estrogen sulfatase) development in the brain of rat, mouse and guinea pig has been established by assaying whole homogenates. Activity was measurable in each species from the fetal state to adulthood. Maximum brain content was reached at about 20 days of age in rat, 14 days in mouse and 15 days in guinea pig. A considerable decrease occurred between 14 days and adulthood in mouse and lesser decreases were seen in rat and guinea pig. The subcellular distribution of enzyme in rat and mouse brain appeared to change from the immature to the adult state. No major differences in enzyme activity occurred between the sexes at any age. Tissue concentration of enzyme in the hypothalamic-preoptic area of rat and mouse was similar to that in the remainder of the brain. In guinea pig the brain concentration was slightly lower than that of the hypothalamic-preoptic region. Sulfatase content of the pituitary was low in all 3 species but the tissue concentration was considerably higher than that of brain, particularly in rat and mouse. Apparent Km values for brain sulfatase were in the range 6-17 microM, with no striking sex difference. Apparent Km's for pituitary sulfatase of immature rat and guinea pig were similar to those for brain in the same animals but that for mouse pituitary (0.9 microM) was much lower. It is unlikely that brain or pituitary sulfatase is by itself, a major factor in making available potentially active estrogen for use during differential sex development in these species.     

Monoamine oxidase activity in the hypothalamus and various other brain areas and in some endocrine glands of the rat during the estrus cycle

—The activity of monoamine oxidase (MAO, EC 1.4.3.4) was measured in the entire hypothalamus and different hypothalamic regions, in the amygdala, frontal and lateral cerebral cortex, in the pituitary, adrenals and genital organs of male rats and of female rats during the estrus cycle. Activity of MAO changed cyclically in the hypothalamus, amygdala, adrenals and ovaries. The highest levels in the hypothalamus occurred at 10 a.m. on the day of proestrus and during estrus. The lowest levels occurred at 6 p.m. on the day of proestrus, of metestrus and during diestrus. Cyclical variations similar to those found in the whole hypothalamus were also observed in anterior, posterior and lateral portions and the median eminence of the hypothalamus. Activity in the median eminence was greater than that of the whole hypothalamus or its various other portions. The amygdala exhibited less marked cyclical activity which followed the pattern of the hypothalamus by increasing at 10 a.m. and peaking at 3 p.m. on the day of proestrus. At the‘post-critical’period of proestrus, when the activity of MAO in the hypothalamus and amygdala decreased, the activity of MAO in the ovaries and adrenals rose. During the estrus cycle much lower levels of activity of MAO were demonstrated in other regions of the brain (frontal and lateral cerebral cortex), in the pituitary and in the uterus, none of which showed cyclical changes. The changes in activity of MAO in cerebral tissues, endocrine glands and genital organs have been discussed in relation to the probable participation of monoamines in the mechanism(s) of secretion of gonadotrophins by the hypothalamus.     

Sulfamoyloxy-substituted 2-phenylindoles: antiestrogen-based inhibitors of the steroid sulfatase in human breast cancer cells

Estrone sulfate (E1S) is an endogenous prodrug that delivers estrone and, subsequently, estradiol to the target cells following the hydrolysis by the enzyme estrone sulfatase which is active in various tissues including hormone dependent breast cancer cells. Blockade of this enzyme should reduce the estrogen level in breast cancer cells and prevent hormonal growth stimulation. Sulfamates of a variety of phenolic compounds have been shown to be inhibitors of estrone sulfatase. Our rational is based on findings that these inhibitors can undergo hydrolysis and the pharmacological effects of the free hydroxy compounds contribute to the bioactivity of the sulfamates. A desirable action of the metabolites would be an estrogen antagonism to block stimulatory effects of residual amounts of estrogens. Thus, we synthesized a number of sulfamoyloxy-substituted 2-phenylindoles with side chains at the indole nitrogen that guarantee antiestrogenic activity. All of the new sulfamates were studied for their inhibitory effects on the enzyme estrone sulfatase from human breast cancer cells and their (anti)hormonal activities in stably transfected human MCF-7/2a mammary carcinoma cells. The hormonal profile of the sulfamates was partly reflected by the properties of the corresponding hydroxy precursors. Some of the sulfamoylated antiestrogens strongly inhibited estrone sulfatase activity with IC₅₀ values in the submicromolar range. They were devoid of agonist activity and suppressed estrone sulfate-stimulated gene expression mainly by blocking the enzyme. Examples are the disulfamates of the indoles ZK 119, 010 and ZK 164, 015. Their IC₅₀s for sulfatase inhibition were 0.3 and 0.2 μM, respectively, and 50 and 80 nM, respectively, for the inhibition of E1S-stimulated luciferase expression in transfected MCF-7 cells. With some of the new sulfamates an additional direct antiestrogenic effect was noticed which might be due to a partial hydrolysis during incubation and would improve the growth inhibitory effect on estrogen-sensitive breast cancer cells.     

Estrone sulfate and dehydroepiandrosterone sulfate concentrations in normal subjects and men with cirrhosis.

Circulation levels of estrone sulfate (E1S) and dehydroepiandrosterone sulfate (DHAS) have been measured in plasma using a radioimmunoassay for estrone and dehydroepiandrosterone following extraction and hydrolysis of the sulfate. The mean +/- SE concentrations of E1S and DHAS in normal men were 458 +/- 25 pg/ml and 1.45 +/- 0.19 micrograms/ml, respectively. In normal women the values for days 5-7 of the cycle were 880 +/- 117 pg/ml and 1.25 +/- 0.12 micrograms/ml which were not different than the values for days 20-22 of 1195 +/- 176 pg/ml and 1.58 +/- 0.29 micrograms/ml. The mean values in post-menopausal women were 250 +/- 33 pg/ml and 0.47 +/- 0.07 micrograms/ml, both lower than the values in young women. In a group of cirrhotic men the mean values were 325 +/- 55 pg/ml and 0.38 +/- 0.12 micrograms/ml, both significantly lower than the normal values. This suggests a defect in sulfurylation in men with hepatic cirrhosis.