Design and synthesis of 3-benzylaminocoumarin-7-O-sulfamate derivatives as steroid sulfatase inhibitors.

Steroid sulfatase (STS) is a sulfatase enzyme that catalyzes the conversion of sulfated steroid precursors to free steroid. The inhibition of STS could abate estrogenic steroids that stimulate the proliferation and development of breast cancer, and therefore STS is a potential target for adjuvant endocrine therapy. In this study, a series of 3-benzylaminocoumarin-7-O-sulfamate derivatives targeting STS were designed and synthesized. Structure-relationship activities (SAR) analysis revealed that attachment of a benzylamino group at the 3-position of coumarin improved inhibitory activity. Compound 3j was found to have the highest inhibition activity against human placenta isolated STS (IC50  0.13 µM) and MCF-7 cell lines (IC50 1.35 µM). Kinetic studies found compound 3j to be an irreversible inhibitor of STS, with KI and kinact value of 86.9 nM and 158.7 min-1, respectively.

The Role of Steroid Sulfatase as a Prognostic Factor in Patients with Endometrial Cancer.

PURPOSE: The aim of the study was to determine steroid sulfatase (STS) expression in endometrial cancer patients and its correlation with disease prognosis. MATERIALS AND METHODS: We conducted a retrospective study in 59 patients who underwent surgery with histologically confirmed endometrial cancer from January 2000 to December 2011 at Hanyang University Hospital. Immuno-histochemical staining of STS was performed using rabbit polyclonal anti-STS antibody. RESULTS: Sixteen of the 59 patients (27.1%) were positive for STS expression. Disease free survival (DFS) was 129.83±8.67 [95% confidence interval (CI): 112.84-146.82] months in the STS positive group (group A) and 111.06±7.17 (95% CI: 97.01-125.10) months in the STS negative group (group B) (p=0.92). Overall survival (OS) was 129.01±9.38 (95% CI: 110.63-147.38) months and 111.16±7.10 (95% CI: 97.24-125.07) months for the groups A and B, respectively (p=0.45). Univariate analysis revealed that FIGO stage and adjuvant therapy are significantly associated with DFS and OS. However, in multivariate analysis, FIGO stage and adjuvant therapy did not show any statistical significance with DFS and OS. STS was also not significantly associated with DFS and OS in univariate and multivariate analysis. CONCLUSION: STS expression was not significantly associated with DFS and OS, despite positive STS expression in 27% of endometrial cancer patients. Therefore, the role of STS as a prognostic factor in patients with endometrial cancer remains unclear and requires further research.

The Role of Steroid Sulfatase as a Prognostic Factor in Patients with Endometrial Cancer

PURPOSE: The aim of the study was to determine steroid sulfatase (STS) expression in endometrial cancer patients and its correlation with disease prognosis. MATERIALS AND METHODS: We conducted a retrospective study in 59 patients who underwent surgery with histologically confirmed endometrial cancer from January 2000 to December 2011 at Hanyang University Hospital. Immuno-histochemical staining of STS was performed using rabbit polyclonal anti-STS antibody. RESULTS: Sixteen of the 59 patients (27.1%) were positive for STS expression. Disease free survival (DFS) was 129.83±8.67 [95% confidence interval (CI): 112.84-146.82] months in the STS positive group (group A) and 111.06±7.17 (95% CI: 97.01-125.10) months in the STS negative group (group B) (p=0.92). Overall survival (OS) was 129.01±9.38 (95% CI: 110.63-147.38) months and 111.16±7.10 (95% CI: 97.24-125.07) months for the groups A and B, respectively (p=0.45). Univariate analysis revealed that FIGO stage and adjuvant therapy are significantly associated with DFS and OS. However, in multivariate analysis, FIGO stage and adjuvant therapy did not show any statistical significance with DFS and OS. STS was also not significantly associated with DFS and OS in univariate and multivariate analysis. CONCLUSION: STS expression was not significantly associated with DFS and OS, despite positive STS expression in 27% of endometrial cancer patients. Therefore, the role of STS as a prognostic factor in patients with endometrial cancer remains unclear and requires further research.

Steroid sulfatase in the human MG-63 preosteoblastic cell line: Antagonistic regulation by glucocorticoids and NFκB.

Steroid sulfatase (STS) converts sulfated steroids into active forms in cells. Preosteoblastic cells possess STS, but its role and regulation in bone are unclear. We examined STS activity and gene expression during differentiation of human MG-63 preosteoblasts. STS activity and gene expression were decreased during differentiation in cells treated with osteogenic supplement containing dexamethasone (DEX). DEX also inhibited STS activity and expression in undifferentiated cells, and the glucocorticoid antagonist RU486 reversed DEX inhibition of STS. These data may have implications for glucocorticoid-induced osteoporosis. The NFκB activators lipopolysaccharide and phorbol myristate acetate increased STS expression in undifferentiated and differentiated MG-63 cells, while the NFκB inhibitor BAY-11-7082 partially blocked these responses. The antagonistic actions of glucocorticoids and NFkB on STS expression are similar to the regulation of inflammatory response proteins. We propose a model of STS regulation whereby inflammation leads to increased STS, resulting in increased estrogen, which modulates the inflammatory response.

Anti-tumor effect of Shu-Gan-Liang-Xue decoction in breast cancer is related to the inhibition of aromatase and steroid sulfatase expression.

ETHNOPHARMACOLOGICAL RELEVANCE: Shu-Gan-Liang-Xue Decoction (SGLXD), a traditional Chinese herbal formula used to ameliorate the hot flushes in breast cancer patients, was reported to have anti-tumor effect on breast cancer. Estrogen plays a critical role in the genesis and evolution of breast cancer. Aromatase and steroid sulfatase (STS) are key estrogen synthesis enzymes that predominantly contribute to the high local hormone concentrations. The present study was to evaluate the anti-tumor effect of SGLXD on estrogen receptor (ER) positive breast cancer cell line ZR-75-1, and to investigate its underlying mechanisms both in vitro and in vivo. MATERIALS AND METHODS: The anti-tumor activity of SGLXD in vitro was investigated using the MTT assay. The in vivo anti-tumor effect of SGLXD was evaluated in non-ovariectomized and ovariectomized athymic nude mice. The effect of SGLXD on enzymatic activity of aromatase and STS was examined using the dual-luciferase reporter (DLR) based on bioluminescent measurements. Aromatase and STS protein level were assessed using Western blot assay. RESULTS: SGLXD showed dose-dependent inhibitory effect on the proliferation of ZR-75-1 cells with IC50 value of 3.40 mg/mL. It also suppressed the stimulating effect on cell proliferation of testosterone and estrogen sulfates (E1S). Oral administration of 6 g/kg of SGLXD for 25 days resulted in a reduction in tumor volume in non-ovariectomized and ovariectomized nude mice. The bioluminescent measurements confirmed that SGLXD has a dual-inhibitory effect on the activity of aromatase and STS. Western blot assay demonstrated that the treatment of SGLXD resulted in a decrease in aromatase and STS protein levels both in vitro and in vivo. CONCLUSION: Our results suggested that SGLXD showed anti-tumor effect on breast cancer cells both in vitro and in vivo. The anti-tumor activity of SGLXD is related to inhibition of aromatase and STS via decreasing their expression. SGLXD may be considered as a novel treatment for ER positive breast cancer.

Shu-Gan-Liang-Xue Decoction, a Chinese herbal formula, down-regulates the expression of steroid sulfatase genes in human breast carcinoma MCF-7 cells.

ETHNOPHARMACOLOGICAL RELEVANCE: Shu-Gan-Liang-Xue Decoction (SGLXD), a traditional Chinese herbal formula, has been used to ameliorate hot flushes symptom in breast cancer patients for decades. AIM OF THE STUDY: Steroid sulfatase (STS) has a crucial role in regulating the estrogen biosynthesis within breast tumors. We aimed to investigate whether SGLXD could modulate STS expression in human breast carcinoma MCF-7 cells. MATERIALS AND METHODS: By semi-quantitative/quantitative reverse transcription-PCR, we investigated the transcript level of STS in MCF-7 cells treated with various concentrations of SGLXD. By using transient cotransfection with estrogen dependent plasmid pERE-TK-Luc and internal control plasmid pRL-TK in MCF-7 cells and dual luciferase reporter (DLR) based bioluminescent measurements, we evaluated the enzymatic activity of STS after SGLXD treatment. RESULTS: By RT-PCR and real time PCR, the mRNA level of STS was decreased by SGLXD treatment, in the dose-dependent manner, compared to negative control (p<0.01). By DLR assay, different concentrations of SGLXD significantly inhibited the enzymatic activity of STS in MCF-7 cells dose-dependently (p<0.05). CONCLUSIONS: SGLXD could act as a selective estrogen enzyme modulator by down-regulating the STS expression in MCF-7 cells.

Inactivation of recombinant human steroid sulfatase by KW-2581.

Steroid sulfatase (STS) catalyses the hydrolysis of the sulfate esters of 3-hydroxy steroids, which are inactive transport or precursor forms of the active 3-hydroxy steroids. STS inhibitors are expected to block the local production and, consequently to reduce the active steroid levels; therefore, they are considered as potential new therapeutic agents for the treatment of estrogen- and androgen-dependent disorders such as breast and prostate cancers. KW-2581 is a novel steroidal STS inhibitor. In the present study, we found KW-2581 inhibited recombinant human STS (rhSTS) activity with an IC(50) of 2.9 nM when estrone sulfate was used as a substrate. The potency of KW-2581 was approximately 5-fold higher than that of a non-steroidal STS inhibitor, 667 COUMATE. KW-2581 was able to equally inhibit rhSTS activity when dehydroepiandrosterone sulfate was used as another substrate. KW-2581 inhibited rhSTS activity in a time- and concentration-dependent manner (k(inact), 0.439 min(-1); K(i, app), 15 nM), suggesting that it is an active site-directed irreversible inhibitor. Both decrease of KW-2581 concentration and increase of the des-sulfamoylated form's concentration were simultaneously observed during the reaction in a time-dependent manner with corresponding to the decrease of STS activity. Our findings for the first time demonstrated the production of des-sulfamoylated form of the compound as a consequence of STS inactivation.

Effects of black cohosh on estrogen biosynthesis in normal breast tissue in vitro.

OBJECTIVES: To investigate the effect of black cohosh on the estrogen biosynthesis in the breast in vitro. METHODS: Steroid sulfatase (STS) activity was studied in normal breast tissue obtained from pre- and postmenopausal women undergoing reduction mammoplasty. STS protein expression was studied by immunohistochemistry and western blotting. Breast tissue was incubated in vitro without or with black cohosh (iCR) at concentrations ranging from 0.1mg/ml to 1 ng/ml. STS activity was evaluated by incubating homogenized breast tissue with [3H]-estrone sulfate, separating the formed products, estrone (E1) and estradiol (E2), by thin layer chromatography and measuring the amounts of E1 and E2 by scintillation counting. RESULTS: STS protein expression and enzymatic activity were detected in all specimens investigated. In all groups, significantly more E1 than E2 was produced. Local estrogen formation was decreased in premenopausal breast tissue by treatment with iCR at 0.1mg/ml (p

Phytoestrogens are potent inhibitors of estrogen sulfation: implications for breast cancer risk and treatment.

We investigated the ability of 37 flavonoids and flavonoid sulfoconjugates, including some abundant dietary constituents, to act as substrates and/or inhibitors of the sulfotransferase and sulfatase enzymes that interconvert active estrogens and inactive estrogen sulfates in human tissues. The enzymes studied include estrogen sulfotransferase, the thermostable phenolsulfotransferase that acts on a range of substrates including estrogens; steroid sulfatase; and two related enzymes, monoamine phenolsulfotransferase and arylsulfatase A. Several dietary flavonoids, including the soy isoflavones genistein and daidzein, were sulfated by these human sulfotransferases. Many flavonoids were potent inhibitors of thermostable phenolsulfotransferase. Genistein and equol were potent mixed inhibitors of hepatic estrogen sulfotransferase, with inhibitory constant values of 500 nM and 400 nM, respectively. Monoamine phenolsulfotransferase activity was relatively unaffected by flavonoids, but this enzyme was mainly responsible for the sulfation of flavonoids at concentrations greater than 1 micro M. Of the compounds tested, only daidzein 4,7-bisulfate, a trace metabolite in humans, significantly inhibited steroid sulfatase in the micromolar concentration range. Hence, dietary flavonoids may be able to influence the bioavailability of endogenous estrogens, and disrupt endocrine balance, by increasing the ratio of active estrogens to inactive estrogen sulfates in human tissues.