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Per Eystein Lønning - One of the best experts on this subject based on the ideXlab platform.

  • an optimised highly sensitive radioimmunoassay for the simultaneous measurement of Estrone estradiol and Estrone Sulfate in the ultra low range in human plasma samples
    The Journal of Steroid Biochemistry and Molecular Biology, 2008
    Co-Authors: Jurgen Geisler, Per Eystein Lønning, Dagfinn Ekse, Hildegunn Helle, Nhat K Duong
    Abstract:

    Abstract Following the introduction of potent aromatase inhibitors for the treatment of breast cancer patients, highly sensitive methods have become mandatory to evaluate the influence of these drugs on plasma estrogen levels. Commercially available kits for estrogen measurements are not suitable for these kinds of evaluations due to their detection limits that are close to baseline estrogen levels in postmenopausal women. We describe here an optimised radioimmunoassay suitable for the simultaneous measurement of plasma Estrone (E 1 ), estradiol (E 2 ) and Estrone Sulfate (E 1 S) levels in the ultra-low range. Following incubation with [ 3 H]-labelled estrogens as internal standards, crude estrogen fractions were separated by ether extraction. The E 1 S fraction was hydrolysed with sulfatase followed by eluation on a Sephadex column. Free estrogens (E 1 , E 2 ) were separated by chromatography (LH-20). Estrone and E 1 S (following hydrolysis) were converted into E 2 , and each estrogen fraction was measured by the same highly sensitive and specific radioimmunoassay using estradiol-6-(O-carboxymethyl)-oximino-2-(2-[ 125 I]-iodo-histamine) as ligand. Although several purification steps were involved, the internal recovery values for tritiated estrogens were found to be 88%, 90%, and 49% for E 1 , E 2 and E 1 S, respectively. The intra-assay coefficient of variation was 1 , 0.67 pmol/L for E 2 , and 0.55 pmol/L for E 1 S, respectively. The intra-assay coefficient of variation (CV) was found to be 3.4% for E 1 , 5.1% for E 2 and 6.1% for E 1 S, while the inter-assay CV was 13.6%, 7.6% and 7.5% for E 1 , E 2 , and E 1 S, respectively. Considering normal plasma levels for E 2 (15 pmol/L), E 1 (80 pmol/L) and E 1 S (400 pmol/L) in postmenopausal women, the method allows theoretically to detect suppression of plasma E 2 , E 1 and E 1 S levels by 95.5%, 98.6% and 99.9% when starting from average, normal postmenopausal levels. Thus, the method presented here is to our knowledge the currently most sensitive assay available for plasma estrogen measurements in the ultra-low range and, as such, a reliable tool for a proper evaluation of potent aromatase inhibitors and other potential drugs influencing on plasma estrogen levels.

  • an optimised highly sensitive radioimmunoassay for the simultaneous measurement of Estrone estradiol and Estrone Sulfate in the ultra low range in human plasma samples
    The Journal of Steroid Biochemistry and Molecular Biology, 2008
    Co-Authors: Jurgen Geisler, Per Eystein Lønning, Dagfinn Ekse, Hildegunn Helle, Nhat K Duong
    Abstract:

    Following the introduction of potent aromatase inhibitors for the treatment of breast cancer patients, highly sensitive methods have become mandatory to evaluate the influence of these drugs on plasma estrogen levels. Commercially available kits for estrogen measurements are not suitable for these kinds of evaluations due to their detection limits that are close to baseline estrogen levels in postmenopausal women. We describe here an optimised radioimmunoassay suitable for the simultaneous measurement of plasma Estrone (E1), estradiol (E2) and Estrone Sulfate (E1S) levels in the ultra-low range. Following incubation with [3H]-labelled estrogens as internal standards, crude estrogen fractions were separated by ether extraction. The E1S fraction was hydrolysed with sulfatase followed by eluation on a Sephadex column. Free estrogens (E1, E2) were separated by chromatography (LH-20). Estrone and E1S (following hydrolysis) were converted into E2, and each estrogen fraction was measured by the same highly sensitive and specific radioimmunoassay using estradiol-6-(O-carboxymethyl)-oximino-2-(2-[125 I]-iodo-histamine) as ligand. Although several purification steps were involved, the internal recovery values for tritiated estrogens were found to be 88%, 90%, and 49% for E1, E2 and E1S, respectively. The intra-assay coefficient of variation was <5% for all recovery measurements. The detection limits were calculated following repeated blank measurements and found to be 1.14 pmol/L for E1, 0.67 pmol/L for E2, and 0.55 pmol/L for E1S, respectively. The intra-assay coefficient of variation (CV) was found to be 3.4% for E1, 5.1% for E2 and 6.1% for E1S, while the inter-assay CV was 13.6%, 7.6% and 7.5% for E1, E2, and E1S, respectively. Considering normal plasma levels for E2 (15 pmol/L), E1 (80 pmol/L) and E1S (400 pmol/L) in postmenopausal women, the method allows theoretically to detect suppression of plasma E2, E1 and E1S levels by 95.5%, 98.6% and 99.9% when starting from average, normal postmenopausal levels. Thus, the method presented here is to our knowledge the currently most sensitive assay available for plasma estrogen measurements in the ultra-low range and, as such, a reliable tool for a proper evaluation of potent aromatase inhibitors and other potential drugs influencing on plasma estrogen levels.

  • influence of dexaminoglutethimide an optical isomer of aminoglutethimide on the disposition of Estrone Sulfate in postmenopausal breast cancer patients
    The Journal of Clinical Endocrinology and Metabolism, 1998
    Co-Authors: Jurgen Geisler, S Lundgren, Hildegunn Berntsen, J L Greaves, Per Eystein Lønning
    Abstract:

    Aminoglutethimide (AG) has been the most widely used aromatase inhibitor in breast cancer patients to date. Commercially, AG (Orimeten) is available as a racemate (dl-AG). Previous studies suggested the stereoisomers of AG (d-AG and l-AG) to differ considerably in their affinities and potencies to inhibit different cytochrome P-450-dependent enzymes, with d-AG being the potent aromatase inhibitor. dl-AG, apart from being an aromatase inhibitor, is known to enhance the metabolism of plasma Estrone Sulfate (E1S). In the present study we compared the effects of d-AG (500 mg daily) and dl-AG (1000 mg daily) on plasma estrogen levels and Estrone (E1) and E1S clearance rates, determined after the injection of [14C]E1 and[ 3H]E1S, in a cross-over study involving 12 postmenopausal breast cancer patients. Treatment with dl-AG and d-AG suppressed plasma E1S to 18.6% and 15.0% of pretreatment levels, whereas E1 and estradiol E2 levels fell to 18.6% and 23.4% of their pretreatment levels during treatment with dl-AG a...

  • profound suppression of plasma estrogens by megestrol acetate in postmenopausal breast cancer patients
    Clinical Cancer Research, 1996
    Co-Authors: S Lundgren, S I Helle, Per Eystein Lønning
    Abstract:

    Twelve postmenopausal women suffering from advanced breast cancer had plasma estrogens, androgens, cortisol, and gonadotropins determined before therapy and during treatment with megestrol acetate (MA) in oral doses escalated from 40 to 160 mg. The plasma clearance and production rate of Estrone and Estrone Sulfate were determined before treatment and after 4 weeks of therapy with 160 mg MA. Treatment with MA suppressed plasma levels of dehydroepiandrosterone Sulfate, androstenedione, and cortisol in a dose-dependent manner to <10% of pretreatment values. Plasma testosterone, estradiol, Estrone, and Estrone Sulfate were suppressed to 18-29% of pretreatment values, whereas the gonadotropins were suppressed to 35-52%. The plasma clearance rates of Estrone and Estrone Sulfate were increased by a mean value of 23.7% (P < 0.01) and 23.5% (P < 0.025), whereas the production rates were reduced by 76.7% (P < 0.0005) and 76.1% (P < 0.0005), respectively. Our findings indicate that MA causes profound suppression of adrenal steroid production but in addition suppresses ovarian secretion of androgens in postmenopausal breast cancer patients. The reduction in plasma estrogens is comparable to values obtained with commonly used aromatase inhibitors and may be responsible for its antitumor effects in breast cancer.

Giuseppe Corona - One of the best experts on this subject based on the ideXlab platform.

  • Effect of one-month treatment with vaginal promestriene on serum Estrone Sulfate levels in cancer patients: A pilot study
    Maturitas, 2012
    Co-Authors: L. Del Pup, D. Postruznik, Giuseppe Corona
    Abstract:

    Abstract Vaginal promestriene was tested in gynecological cancer patients who suffered from severe vaginal dryness and dyspareunia. This form of estrogen has a low level of vaginal absorption and proved to be effective for vaginal atrophy. Method 17 patients were treated with a 10 mg soft vaginal suppository daily for one month. Plasma levels of Estrone Sulfate (E1S), used as the marker of overall estrogenicity, were measured by liquid chromatography in combination with mass spectrometry. Results Mean E1S levels changed from 533 (22–2920) to 374 (81–856) pg/ml ( p  = 0.39). Conclusion In highly symptomatic gynecological cancer patients the level of circulating Estrone Sulfate was not significantly affected by vaginal promestriene treatment overall, but a wide range of levels was noted pre and post treatment in individual patients.

  • Liquid chromatography tandem mass spectrometry assay for fast and sensitive quantification of Estrone-Sulfate.
    Clinica Chimica Acta, 2010
    Co-Authors: Giuseppe Corona, Caterina Elia, Bruno Casetta, Alessandro Da Ponte, Lino Del Pup, Enzo Ottavian, Giuseppe Toffoli
    Abstract:

    Abstract Background The circulating pool of Estrone-Sulfate is considered as a “reservoir” of slowly-metabolized estrogen that can be exploited for assessing overall individual estrogenicity. The aim of this study was to develop a rapid and sensitive liquid chromatography–tandem mass spectrometry assay for the determination of Estrone-Sulfate, suitable for routine clinical investigations. Methods The proposed assay is based on a simple protein precipitation procedure and on a fast measurement with a triple–quadrupole mass spectrometer operating in negative ion mode and in multiple reaction monitoring. The method was assessed for intra- and inter-day precision, accuracy, recovery, and clinical suitability. A comparison with available radioimmunoassay was also performed. Results The LC–MS/MS method is able to detect Estrone-Sulfate concentrations ≤1 pg/mL and has a low limit of quantification of 7.8 pg/mL. Intra- and inter-day precision and accuracy were less than 10.5% and 5.0% respectively. The recovery was in the range of 93%–110%. When compared with radioimmunoassay the method resulted more accurate and therefore more suitable for quantifying the Estrone-Sulfate in different clinical settings, including patients treated with aromatase inhibitors. Conclusions The proposed LC–MS/MS method represents a convincing alternative to the immunoassay for a fast, cost-effective and reliable measurement of Estrone-Sulfate in routine clinical investigations and in large epidemiological studies. It may contribute in shedding a new light on the diagnostic value of Estrone-Sulfate in normal and pathological conditions.

J R Pasqualini - One of the best experts on this subject based on the ideXlab platform.

  • concentrations of Estrone estradiol and Estrone Sulfate and evaluation of sulfatase and aromatase activities in pre and postmenopausal breast cancer patients
    The Journal of Clinical Endocrinology and Metabolism, 1996
    Co-Authors: J R Pasqualini, L Delalonde, M C Feinstein, Gérard Chetrite, C Blacker, M Talbi, C Maloche
    Abstract:

    This report concerns the evaluation of various estrogens, Estrone (El), estradiol (E2), and Estrone Sulfate (E1S), as well as E1S-sulfatase and aromatase activities in pre- and postmenopausal women with breast cancer. The levels (in picomoles per g; mean +/- SEM) of the various estrogens in the breast tissue from premenopausal patients (n = 11) are: El, 1.4 +/- 0.5; E2, 1.2 +/- 0.6; and E1S, 1.2 +/- 0.3. In postmenopausal patients (n = 23), the values are, respectively, 1.0 +/- 0.4, 1.4 +/- 0.7, and 3.3 +/- 1.9. These concentrations of estrogens in the tumors of postmenopausal patients are significantly higher than those found in plasma. The activity of E1S-sulfatase in both pre- and postmenopausal patients was 50-200 times higher than that of aromatase. E1S-sulfatase and aromatase activities are significantly higher in post-menopausal than in cycling patients. It is concluded that despite the low levels of circulating estrogens in postmenopausal patients, the tissue concentrations of these steroids are s...

  • Estrone Sulfate sulfatase and 17β hydroxysteroid dehydrogenase activities a hypothesis for their role in the evolution of human breast cancer from hormone dependence to hormone independence
    The Journal of Steroid Biochemistry and Molecular Biology, 1995
    Co-Authors: J R Pasqualini, Bachlien Nguyen, C Maloche, L Delalonde, Marwa Talbi, M C Feinstein, Joaquin Botella, Catherine Blacker, Gérard Chetrite, J. Paris
    Abstract:

    Abstract The evaluation of estrogens (Estrone, estradiol, and their Sulfates) in the breast tissue of post-menopausal patients with breast cancer indicates high levels, particularly of Estrone Sulfate (E 1 S) which is 15–25 times higher than in the plasma. Breast cancer tissue contains the enzymes necessary for local synthesis of estradiol and it was demonstrated that, despite the presence of the sulfatase and its messenger in hormone-dependent and hormone-independent breast cancer cells, this enzyme operates particularly in hormone-dependent cells. Different progestins: Nomegestrol acetate, Promegestone, progesterone, as well as Danazol, can block the conversion of E 1 S to E 2 very strongly in hormone-dependent breast cancer cells. The last step in the formation of estradiol is the conversion of E 1 to this estrogen by the action of 17β-hydroxysteroid dehydrogenase. This activity is preferentially in the reductive direction (formation of E 2 ) in hormone-dependent cells, but oxidative (E 2 → E 1 ) in hormone-independent cells. Using intact hormone-dependent cells it was observed that Nomegestrol acetate can block the conversion of E 1 to E 2 . It is concluded, firstly, that in addition to ER mutants other factors are involved in the transformation of hormone-dependent breast cancer to hormone-independent, this concerns the enzymatic activity in the formation of E 2 ; it is suggested that stimulatory or repressive factor(s) involved in the enzyme activity are implicated as the cancer evolves to hormone-independence; secondly, different drugs can block the conversion of E 1 S to E 2 . Clinical trials of these “anti-enzyme” substances in breast cancer patients could be the next step to investigate new therapeutic possibilities for this disease.

  • action of danazol on the conversion of Estrone Sulfate to estradiol and on the sulfatase activity in the mcf 7 t 47d and mda mb 231 human mammary cancer cells
    The Journal of Steroid Biochemistry and Molecular Biology, 1993
    Co-Authors: B L Nguyen, I Ferme, G Chetrite, J R Pasqualini
    Abstract:

    Abstract In the present studies the action of Danazol on the conversion of Estrone Sulfate (E 1 S) to estradiol (E 2 ) as well as on the sulfatase activity in the MCF-7 and T-47D, hormone-dependent, and MDA-MB-231, hormone-independent, mammary cancer cell lines was explored. Using intact cells we observed that Danazol blocks very significantly the radioactivity uptake and the conversion of [ 3 H]E 1 S to E 2 in all the cells studied. In particular, a very strong effect (85% decrease of these parameters versus the control values) is observed in the T-47D cells. In another series of studies using cell homogenates it is observed that Danazol inhibits the sulfatase activity in all these cell lines. The effect of Danazol is dose-dependent and significant from a concentration of 1 μM. At concentrations of 8 μM E 1 S, 10 −5 M Danazol, the inhibition of sulfatase activity is 38% in MCF-7, 36% in MDA-MB-231, and 27% in T-47D cells. Analysis by Lineweaver-Burk plot shows that the inhibitory effect is competitive. As E 1 S is one of the main sources of E 2 in human mammary tumors, the present data could open new possibilities for therapeutic applications in hormone-dependent breast cancer.

Dirk Grundemann - One of the best experts on this subject based on the ideXlab platform.

  • hyperosmolarity stimulates transporter mediated insertion of Estrone Sulfate into the plasma membrane but inhibits the uptake by slc10a1 ntcp
    Biochemical Pharmacology, 2021
    Co-Authors: Julian Peter Muller, Lena Keufgens, Dirk Grundemann
    Abstract:

    Abstract Many drugs are largely hydrophobic molecules; a transporter might conceivably insert these into the plasma membrane. At least 18 transporters from diverse families have been reported to transport the model compound Estrone Sulfate alias Estrone-3-Sulfate (E3S). Out of these, we recently examined SLC22A11 (OAT4). We concluded from a comparison of E3S and uric acid transport that SLC22A11 does not translocate E3S into the cytosol, but into the plasma membrane. Here we present a hyperosmolarity alias hypertonicity assay to differentiate transport mechanisms. Human transporters were expressed heterologously in 293 cells. Solute uptake into intact cells was measured by LC-MS. Addition of mannitol or sucrose led to rapid cell shrinkage, but cell viability after 60 min in hyperosmolar buffer was not impaired. A decrease in substrate accumulation with increasing osmolarity as observed here for several substrates and the transporters SLC22A11, ETT (SLC22A4), OCT2 (SLC22A2), OAT3 (SLC22A8), and MATE1 (SLC47A1) suggests regular substrate translocation into the cytosol. An increase as observed for E3S transport by SLC22A11, OAT3, MATE1, SLC22A9, and SLC10A6 implies insertion into the membrane. In marked contrast to the other E3S transporters, the bile acid transporter SLC10A1 (NTCP, Na+ taurocholate co-transporting polypeptide) showed a decrease in the accumulation of E3S in hyperosmolar buffer; the same was observed with taurocholic acid. Indeed, our data from several functional assays strongly suggest that the transport mechanism is identical for both substrates. Apparently, a unique transport mechanism has been established for SLC10A1 by evolution that ensures the transport of amphipathic, detergent-like molecules into the cytosol.

  • a novel mode of operation of slc22a11 membrane insertion of Estrone Sulfate versus translocation of uric acid and glutamate
    Biochemical Pharmacology, 2017
    Co-Authors: Peter Skwara, Edgar Schomig, Dirk Grundemann
    Abstract:

    Abstract Estrone Sulfate alias Estrone-3-Sulfate (E3S) is considerably larger and much more hydrophobic than typical substrates of SLC22 transporters. It is puzzling that many otherwise unrelated transporters have been reported to transport E3S. Here we scrutinized the mechanism of transport of E3S by SLC22A11 (alias OAT4), by direct comparison with uric acid (UA), an important physiological substrate. Heterologous expression of SLC22A11 in human 293 cells gave rise to a huge unidirectional efflux of glutamate (Glu) and aspartate, as determined by LC–MS/MS. The uptake of E3S was 20-fold faster than the uptake of UA. Yet, the outward transport of Glu was inhibited by extracellular E3S, but not by UA. The release of E3S after preloading was trans -stimulated by extracellular dehydroepiandrosterone Sulfate (DHEAS), but neither by UA nor 6-carboxyfluorescein (6CF). The equilibrium accumulation of E3S was enhanced 3-fold by replacement of chloride with gluconate, but the opposite effect was observed for UA. These results establish that SLC22A11 provides entirely different transport mechanisms for E3S and UA. Therefore, E3S must not be used as a substitute for UA to assay the function of SLC22A11. In equilibrium accumulation experiments, the transporter-mediated uptake was a linear function of the concentration of UA and 6CF. By contrast, in the same concentration range the graph for E3S was hyperbolic. This suggests that SLC22A11 inserts E3S into a small volume with limited capacity, the plasma membrane. Our data support the notion that the reverse process, extraction from the membrane, is also catalyzed by the carrier.

Bachlien Nguyen - One of the best experts on this subject based on the ideXlab platform.

  • Estrone Sulfate sulfatase and 17β hydroxysteroid dehydrogenase activities a hypothesis for their role in the evolution of human breast cancer from hormone dependence to hormone independence
    The Journal of Steroid Biochemistry and Molecular Biology, 1995
    Co-Authors: J R Pasqualini, Bachlien Nguyen, C Maloche, L Delalonde, Marwa Talbi, M C Feinstein, Joaquin Botella, Catherine Blacker, Gérard Chetrite, J. Paris
    Abstract:

    Abstract The evaluation of estrogens (Estrone, estradiol, and their Sulfates) in the breast tissue of post-menopausal patients with breast cancer indicates high levels, particularly of Estrone Sulfate (E 1 S) which is 15–25 times higher than in the plasma. Breast cancer tissue contains the enzymes necessary for local synthesis of estradiol and it was demonstrated that, despite the presence of the sulfatase and its messenger in hormone-dependent and hormone-independent breast cancer cells, this enzyme operates particularly in hormone-dependent cells. Different progestins: Nomegestrol acetate, Promegestone, progesterone, as well as Danazol, can block the conversion of E 1 S to E 2 very strongly in hormone-dependent breast cancer cells. The last step in the formation of estradiol is the conversion of E 1 to this estrogen by the action of 17β-hydroxysteroid dehydrogenase. This activity is preferentially in the reductive direction (formation of E 2 ) in hormone-dependent cells, but oxidative (E 2 → E 1 ) in hormone-independent cells. Using intact hormone-dependent cells it was observed that Nomegestrol acetate can block the conversion of E 1 to E 2 . It is concluded, firstly, that in addition to ER mutants other factors are involved in the transformation of hormone-dependent breast cancer to hormone-independent, this concerns the enzymatic activity in the formation of E 2 ; it is suggested that stimulatory or repressive factor(s) involved in the enzyme activity are implicated as the cancer evolves to hormone-independence; secondly, different drugs can block the conversion of E 1 S to E 2 . Clinical trials of these “anti-enzyme” substances in breast cancer patients could be the next step to investigate new therapeutic possibilities for this disease.

  • effect of the progestagen r5020 promegestone and of progesterone on the uptake and on the transformation of Estrone Sulfate in the mcf 7 and t 47d human mammary cancer cells correlation with progesterone receptor levels
    Cancer Letters, 1992
    Co-Authors: Jorge R. Pasqualini, Claire Varin, Bachlien Nguyen
    Abstract:

    In the present study we have explored the actions of the progestagen R5020 (Promegestone: 17 alpha, 21-dimethyl-19-nor-pregna-4, 9-diene-3,20-dione) and progesterone on the uptake of [3H]Estrone Sulfate ([3H]E1S) and its conversion to estradiol (E2) by two hormone-dependent mammary cancer cell lines: MCF-7 and T-47D. R5020 or progesterone significantly decreased the uptake of [3H]E1 and its conversion to (E2). In the cells of the two lines, R5020 or progesterone (5 x 10(-6) M) decreased the E2 concentrations by 2-3 times in relation to the levels in untreated cells. E1S (1 x 10(-7) M) also increased expression of the progesterone receptor (PR) and both R5020 (5 x 10(-6) M) and progesterone (5 x 10(-6) M) blocked this stimulatory action of E1S in cells of both cell lines. As E2 is one of the main factors of cancerization in the breast and Estrone Sulfate is quantitatively the most important precursor of E2 in this tissue, the decrease of E2 by these progestagens could open new possibilities for the control of E2 in the breast cancer tissue.

  • Estrone sulfatase activity and effect of antiestrogens on transformation of Estrone Sulfate in hormone-dependent vs. independent human breast cancer cell lines
    Breast Cancer Research and Treatment, 1991
    Co-Authors: Jorge R. Pasqualini, Bachlien Nguyen
    Abstract:

    The effect of the anti-estrogens ICI 164,384 and tamoxifen on the estradiol (E_2) concentration after incubation of Estrone Sulfate (E_1-S) with different hormone-dependent (MCF-7 and T-47D) and hormone-independent (MDA-MD-231 and MDA-MB-436) mammary cancer cells, as well as the Estrone sulfatase activity in these various cell lines, are presented. The anti-estrogen ICI 164,384 decreased very significantly the concentration of E_2 after incubation of E_1-S with MCF-7 (control, mean ±SE: 100±24 pg/mg DNA; + ICI 164,384 [10^−6M]: 7±2 pg/mg DNA). This effect was much more intense than with tamoxifen. A similar effect was observed with T-47D cells. However, no significant effect was observed in the hormone-independent cells. In the intact cell, Estrone sulfatase activity was very intense in the hormone-dependent cells, but very small in the hormone-independent cells. However, this activity became very strong after homogenization in the hormone-independent cells. The data suggest that Estrone Sulfate can play an important role on the bioavailability of E_2 in hormone-dependent breast cancer, and that understanding the control of Estrone sulfatase activity can open new knowledge of the estrogen responses and new possibilities of therapeutic application in breast cancer.