Investigation of 1,3-dipolar cycloaddition reactions of unsaturated nitrones in ionic liquids.

The effect of the media (achiral and chiral ionic liquids) on the stereochemistry of intramolecular 1,3-dipolar cycloaddition reactions of D-galactose-derived omega-unsaturated nitrones, leading to bicyclic isoxazolidines, has been investigated.

An intramolecular 1,3-dipolar cycloaddition reaction towards the synthesis of chiral azetidine nucleoside analogues: the D-gluco case.

The diastereoselective intramolecular 1,3-dipolar cycloaddition reaction of unsaturated nitrones, derived from methyl alpha-D-glucopyranoside with 2-furaldehyde and 2-(benzyloxy)acetaldehyde has been studied In our pevious studies with 2-furaldehyde, the cycloaddition resulted 3 diastereoisomers in a 3:1:1 ratio. In this article, how the number of the possible isomers generated by 1,3-cycloaddition could be reduced from 4 to 1 when 2-(benzyloxy)acetaldehyde was employed as an aldehyde is shown.

Steroidal and nonsteroidal sulfamates as potent inhibitors of steroid sulfatase.

Synthetic routes to potent steroidal and nonsteroidal sulfamate-based active site-directed inhibitors of the enzyme steroid sulfatase, a topical target in the treatment of postmenopausal women with hormone-dependent breast cancer, are described. Novel compounds were examined for estrone sulfatase (E1-STS) inhibition in intact MCF-7 breast cancer cells and placental microsomes. Reaction of the sodium salt of estrone with sulfamoyl chloride gave estrone 3-O-sulfamate (EMATE, 2) which inhibits E1-STS activity potently (> 99% at 0.1 microM in intact MCF-7 cells, IC50 = 65 pM) in a time- and concentration-dependent manner, suggesting that EMATE is an active site-directed inhibitor. EMATE is also active in vivo orally. 5,6,7,8-Tetrahydronaphthalene 2-O-sulfamate (7) and its N-methylated derivatives (8 and 9) were synthesized, and 7 inhibits the E1-STS activity in intact MCF-7 cells by 79% at 10 microM. 4-Methylcoumarin 7-O-sulfamate (COUMATE) and its derivatives (14, 16, and 18) were prepared to extend this series of nonsteroidal inhibitors, and COUMATE reduces the E1-STS activity in placental microsomes by > 90% at 10 microM. Although the orally active COUMATE is less potent than EMATE as an active site-directed inhibitor, it has the important advantage of being nonestrogenic. Analogues (20, 22, 24, 26, 27, 31, 33, 39, and 44) of COUMATE were synthesized to study its structure-activity relationships, and sulfamates of tetralones (46 and 48) and indanones (49, 51, and 53) were also prepared. While most of these compounds were found to inhibit E1-STS activity less effectively than COUMATE, one analogue, 3,4-dimethylcoumarin 3-O-sulfamate (24), was found to be some 12-fold more potent than COUMATE as an E1-STS inhibitor in intact MCF-7 cells (IC50 = 30 nM for 24, cf. 380 nM for COUMATE). Hence, highly potent sulfamate-based inhibitors of steroid sulfatase, such as EMATE, COUMATE, and 24, possess therapeutic potential and will allow the importance of estrogen formation in breast tumors via the E1-STS pathway to be assessed. A pharmacophore for active site-directed sulfatase inhibition is proposed.

Estrone sulfonates as inhibitors of estrone sulfatase.

In our continuing quest to design efficient inhibitors of estrone sulfatase activity and to assess the recognition of estrone sulfate surrogates by estrone sulfatase, we synthesized and evaluated several sulfonate derivatives of 5,6,7,8-tetrahydronaphth-2-ol and estrone. 5,6,7,8-Tetrahydronaphth-2-methanesulfonate (11), and 5,6,7,8-tetrahydronaphth-2-(p-toluene)sulfonate (12) were found not to inhibit estrone sulfatase activity; estrone-3-methane-sulfonate (5), estrone-3-ethanesulfonate (6), estrone-3-butanesulfonate (7), and estrone-3-[(+)10-camphor]sulfonate (8) all weakly inhibited estrone sulfatase, and the best inhibitor, from this class of compounds, was estrone-3-(p-toluene)sulfonate (9). At 10 microM, it inhibited estrone sulfatase activity by 91%. These results emphasize some of the requirements needed for high-affinity binding to the enzyme.

Inactivation of steroid sulfatase by an active site-directed inhibitor, estrone-3-O-sulfamate.

Steroid sulfatases are responsible for the hydrolysis of 3beta-hydroxy steroid sulfates, such as cholesterol and pregnenolone sulfate, and have an important role in regulating the synthesis of estrogenic steroids, from estrone sulfate and dehydroepiandrosterone sulfate, in endocrine-dependent tumors. Although little is known about the mechanism by which the sulfate group is removed from a steroid nucleus, an active site-directed sulfatase inhibitor has been developed. This inhibitor, estrone-3-O-sulfamate (EMATE), was synthesized by treating the sodium salt of estrone with sulfamoyl chloride. This compound inhibited not only estrone sulfatase but also dehydroepiandrosterone sulfatase activity in placental microsomes and in intact MCF-7 breast cancer cells. Pretreatment of MCF-7 cells or placental microsomes with EMATE, followed by extensive washing or dialysis indicated irreversible inhibition. This was confirmed by showing that EMATE inhibited estrone sulfatase activity in placental microsomes in a time-, concentration-, and pH-dependent manner. The enzyme is protected from inactivation by estrone sulfate, which is also consistent with active site-directed inhibition. EMATE is proposed to inactivate estrone sulfatase by irreversible sulfamoylation of the enzyme. Maximum enzyme activity was detected at pH 8.6, and the maximum rate of enzyme inactivation by EMATE also occurred at this pH. The pKa values of the enzymatic reaction and pKa of inactivation were 7.2 and 9.8, providing evidence that two active site residues are being modified by EMATE. As the phenolic pKa of tyrosine (9.7) and the pKa of histidine will allow the roles that (6.8) are similar to the pKa values of inactivation, these amino acid residues may play a role in the catalytic mechanism.(ABSTRACT TRUNCATED AT 250 WORDS)

Inhibition of steroid sulphatase activity by steroidal methylthiophosphonates: potential therapeutic agents in breast cancer.

The hydrolysis of steroid sulphates, by steroid sulphatase, is an important source of oestrogenic steroids (oestrone, oestradiol and 5-androstene-3 beta,17 beta-diol) which are found in tumours. In the present study, we have examined the effect of dehydroepiandrosterone-3-O-methylthiophosphonate (DHA-3-MTP), pregnenolone-3-O-methylthiophosphonate (pregnenolone-3-MTP) and cholesterol-3-O-methylthiophosphonate (cholesterol-3-MTP) on the inhibition of oestrone sulphatase as well as DHA sulphatase activities in intact MCF-7 breast cancer cells and in placental microsomes. All three methylthiophosphonates significantly (P < 0.01) inhibited the hydrolysis of oestrone sulphate (E1 S) in intact MCF-7 cells (31-85% inhibition at 1 microM and 53-97% inhibition at 10 microM). Significant inhibition of DHA sulphatase was also achieved. At a concentration of 50 microM, all three compounds inhibited the hydrolysis of dehydroepiandrosterone sulphate (DHAS) by > 95%. Using human placental microsomes, the Km and Vmax of E1S were determined to be 8.1 microM and 43 nmol/h/mg protein. The corresponding Ki values for DHA-3-MTP, pregnenolone-3-MTP and cholesterol-3-MTP were found to be 4.5, 1.4 and 6.2 microM, respectively. Such inhibitors which are resistant to metabolism may have considerable potential as therapeutic agents and may have additional advantage over aromatase inhibitors in also reducing tumour concentrations of the oestrogenic steroid, 5-androstene-3 beta,17 beta-diol, by inhibiting the hydrolysis of DHAS.

Inhibition of estrone sulfatase activity by estrone-3-methylthiophosphonate: a potential therapeutic agent in breast cancer.

Many breast tumors are hormone dependent, and there is evidence that hydrolysis of estrone sulfate (E1S) to estrone, by estrone sulfatase, is an important source of the estrogen which is found in tumors. In this study, we have developed a novel pathway for the synthesis of estrone-3-methylthiophosphonate (E1-3-MTP) and examined its ability to inhibit estrone sulfatase activity in MCF-7 breast cancer cells and human placental and breast tumor preparations. In MCF-7 breast cancer cells, E1-3-MTP, 100 nM and 10 microM, inhibited estrone sulfatase activity by 52 and > 98%, respectively. The apparent Km and Vmax for E1S were 4.8 microM and 148 pmol/min/mg for placental and 16.9 microM and 38 pmol/min/mg for breast tumor preparations. Kinetic studies revealed that E1-3-MTP inhibited estrone sulfatase in a competitive manner with the Ki values for placental and tumor preparations being 14.6 and 32.8 microM, respectively. A comparison of the metabolism of [3H]E1S and [3H]E1-3-MTP by human placenta or rat liver revealed that, whereas 50-60% of [3H]E1S was converted to [3H]estrone, < 3% of [3H]E1-3-MTP was hydrolyzed. The development of an efficient inhibitor of estrone sulfatase, which is resistant to metabolism, will allow the importance of the estrone sulfatase pathway of estrogen formation in breast tumors to be assessed and such an inhibitor may have considerable potential as a therapeutic agent.