3β-Sulfamate Derivatives of C19 and C21 Steroids Bearing a t -Butylbenzyl or a Benzyl Group: Synthesis and Evaluation as Non-estrogenic and Non-androgenic Steroid Sulfatase Inhibitors

Abstract

A series of C19 and C21 steroids bearing one or two inhibiting groups (3b-sulfamate and 17a-or 20(S)-tbutylbenzyl or benzyl) were synthesized and tested for inhibition of steroid sulfatase activity. When only a sulfamate group was added to dehydroepiandrosterone, androst-5-ene-3b,17b-diol, pregnenolone and 20-hydroxy-pregnenolone, no significant inhibition of steroid sulfatase occurred at concentrations of 0.3 and 3 mM. With only a t-butylbenzyl or a benzyl group, a stronger steroid sulfatase inhibition was obtained in the androst-5-ene than in the pregn-5-ene series. Comparative results from the screening tests and the IC 50 values have shown that the effect of a sulfamate moiety as a second inhibiting group can be combined to the t-butylbenzyl or benzyl effect in the C19 and C21 steroid series. The 3b-sulfamoyloxy-17a-t-butylbenzyl-5-androsten-17b-ol (10) was thus found to be the most active compound with IC 50 values of 46 6 8 and 14 6 1 nM; respectively for the transformations of E 1 S to E 1 and DHEAS to DHEA. The IC 50 values of compound 10 are similar to that of 17a-t-butylbenzyl-estradiol, which was previously reported by our group as a good steroid sulfatase reversible inhibitor, but remains higher than that of the potent inactivators estrone-3-O-sulfamate (EMATE) and 17a-t-butylbenzyl-EMATE. However, contrary to these two latter inhibitors, compound 10 did not induce any proliferative effect on estrogensensitive ZR-75-1 cells nor on androgen-sensitive Shionogi cells at concentrations tested, suggesting that this steroid sulfatase inhibitor is non estrogenic and non androgenic.

Steroid sulfatase inhibitors

“…7). 94,101 The same negative result we obtained with the sulfamates of pregnenolone, androsterone, and epiandrosterone. In contrast, steroidal sulfamates with an aromatic A-ring are potent inhibitors of STS, as already known for estrone sulfamate (EMATE), but they act by an irreversible mechanism (see below).…”

“…67,68,105,106 Furthermore, androgenicity of inhibitors was tested using an androgen-sensitive mammary carcinoma cell line. 101 …”

“…97,101 The most active compound out of this series was the androstenediol analog 5 ( Fig. 6) with 79% inhibition of STS activity in HEK-293 cell homogenates at a concentration of 3 mM.…”

“…69 Finally, inhibition of E1S-stimulated uterine growth in ovariectomized mice was used as a model system. 100,101,103 However, such model systems are quite artificial, since they rely on the addition/ administration of sulfated precursors to cell cultures/animals as estrogenic stimulus. Growth of breast tumor cells in human patients will be stimulated both by active estrogens from the circulation and by estrogen precursors (E1S, DHEAS) converted to active estrogens via the sulfatase pathway in the tumor; thus, experiments with STS inhibitors in ovariectomized mice receiving E1S as the only source of estrogens cannot predict anything about efficacy in human disease.…”

“…5 for structure), a prototype STS inhibitor; 99 this stimulated an intense search for non-estrogenic STS inhibitors. Various test systems have been used to test for estrogenicity of STS inhibitors: (i) binding to the isolated ER-a and -b; 100 (ii) stimulation of estrogen-dependent breast cancer cell growth 64,[101][102][103] ; (iii) induction of alkaline phosphatase in endometrial carcinoma cells; 69,104 (iv) stimulation of uterine weight in ovariectomized rats. 67,68,105,106 Furthermore, androgenicity of inhibitors was tested using an androgen-sensitive mammary carcinoma cell line.…”