Estrogens play an important role in the development of breast cancer. Inhibiting 17β-hydroxysteroid dehydrogenase type 1 (17β-HSD1)—the enzyme responsible for the last step in the biosynthesis of the ...most potent estrogen, estradiol (E
2)—would thus allow hindering the growth of estrogen-sensitive tumors. Based on a previous study identifying 16β-benzyl-E
2 (
1) as a lead compound for developing inhibitors of the transformation of estrone (E
1) into E
2, we modified the benzyl group of
1 to improve its inhibitory activity. Three strategies were also devised to produce compounds with less residual estrogenic activity: (1) replacing the hydroxy group by a hydrogen at position 3 (C3); (2) adding a methoxy at C2; and (3) adding an alkylamide chain known to be antiestrogenic at C7. In order to test the inhibitory potency of the new compounds, we used the human breast cancer cell line T-47D, which exerts a strong endogenous 17β-HSD1 activity. In this intact cell model, 16β-
m-carbamoylbenzyl-E
2 (
4m) emerged as a potent inhibitor of 17β-HSD1 with an IC
50 value of 44
nM for the transformation of
14C-E
1 (60
nM) into
14C-E
2 (24-h incubation). In another assay aimed at assessing the unwanted estrogenic activity, a 10-day treatment with
4m at a concentration of 0.5
μM induced some proliferation (38%) of T-47D estrogen-sensitive (ER
+) breast cancer cells. Interestingly, when
4m (0.5
μM) was given with E
1 (0.1
nM) in a 10-day treatment, it blocked 62% of the T-47D cell proliferation induced by E
1 after its reduction to E
2 by 17β-HSD1. Thus, in addition to generating useful structure–activity relationships for the development of 17β-HSD1 inhibitors, our study demonstrates that using such inhibitors is a valuable strategy for reducing the level of E
2 and consequently its proliferative effect in T-47D ER
+ breast cancer cells.
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17Beta-hydroxysteroid dehydrogenase type 3 (17β-HSD3) is a steroidogenic enzyme that catalyzes the transformation of 4-androstene-3,17-dione (Δ
4-dione) into androgen testosterone ...(T). To provide effective inhibitors of androgen biosynthesis, we synthesized two different series (amines and carbamates) of 3β-substituted-androsterone derivatives and we tested their inhibitory activity on 17β-HSD3. From the results of our structure–activity relationship study, we identified a series of compounds producing a strong inhibition of 17β-HSD3 overexpressed in HEK-293 cells (homogenized cells). The most active compound when tested in intact HEK-293 transfected cells, namely (3α,5α)-3-{
trans-2,5-dimethyl-4-{2-(trifluoromethyl)phenyl sulfonyl}piperazin-1-ylmethyl}-3-hydroxyandrostan-17-one (
15b), shows an IC
50 value of 6
nM, this compound is thus eight times more active than our reference compound D-5-2 (IC
50
=
51
nM). This new improved inhibitor did not stimulate the proliferation of androgen-sensitive Shionogi cells, suggesting a non-androgenic profile. Compound
15b is thus a good candidate for further in vivo studies on rodents.
•Lactone- and lactol-estradiol derivatives were synthesized and characterized.•Lactone E-ring was diversified by adding a hydroxymethyl, a methylcarboxylate, a carboxy or an allyl group.•A chemical ...approach was developed to introduce a chemical group on hindered beta-steroid face.•Lactone and lactol derivatives inhibited 17β-HSD1 (34–60%) similarly as the natural substrate estrone (53%).
To control estradiol (E2) formation, we are interested in synthesizing inhibitors of 17β-hydroxyteroid dehydrogenase type 1 (17β-HSD1). Since the results of docking experiments have shown that E2-lactone derivatives substituted in position 19 or 20 (E-ring) could generate interactions with the active site of the enzyme, we carried out their chemical synthesis. After having prepared the 16β,17β-γ-lactone-E2 in four steps starting from estrone (E1), we introduced the molecular diversity by adding a hydroxymethyl, a methylcarboxylate, a carboxy or an allyl group. The allyl derivative was used as a key intermediate to generate a hydroxyethyl side chain in α or β position. Two lactols were also obtained from two hydroxyalkyl lactones. Enzymatic assays revealed that lactone and lactol derivatives weakly inhibited 17β-HSD1 in homogenized HEK-293 cells overexpressing 17β-HSD1 (34–60% at 1μM) and in intact T-47D cells expressing 17β-HSD1 (10–40% at 10μM).
This article is part of a Special Issue entitled “Synthesis and biological testing of steroid derivatives as inhibitors”.
The involvement of aromatase, steroid sulfatase (STS) and reductive 17β-hydroxysteroid dehydrogenases (17β-HSDs) in the production of estrogens was determined in four cell lines of endometrial cancer ...(Ishikawa, HEC-1A, HEC-1B and RL-95) and one cell line of cervix cancer (Hela) in culture. After incubation with 4-androstene-3,17-dione (4-dione), there are no estrogens, estrone (E1) and estradiol (E2), detected suggesting that the pathway of aromatase is not important in these cell lines. In whole cells, the results show low percentages of transformation of estrone sulfate (E1S) into E1 suggesting that the entrance of E1S is difficult. However, in homogenized cells the STS activity was much higher and fully blocked by an inhibitor. Using selective inhibitors for each reductive 17β-HSD (types 1, 5, 7 and 12), alone or in combination, we did not succeed in completely blocking the conversion of E1 into E2, suggesting that another 17β-HSD (known or unknown) is involved in the formation of E2 from E1.
•17β-HSD3 is involved in the formation of androgenic hormone testosterone (T).•RM-532-105 is an androsterone derivative inhibiting 17β-HSD3.•RM-532-105 inhibits the formation of T in homogenized and ...whole HEK-293 cells overexpressing 17β-HSD3.•RM-532-105 (10mg/kg, s.c.) reached a plasma concentration of 250ng/mL at 7h in rat.
17β-Hydroxysteroid dehydrogenase type 3 (17β-HSD3 or HSD17B3) catalyzes the last step in the biosynthesis of the potent androgen testosterone (T), by stereoselectively reducing the C17 ketone of 4-androstene-3,17-dione (4-dione), with NADPH as cofactor. Since T plays an important role in androgen-sensitive diseases, this enzyme is thus an interesting therapeutic target. In an attempt to design compounds to lower the level of T, we synthesized androsterone derivatives substituted at position 3 as inhibitors of 17β-HSD3, and selected one of the most potent compounds for additional studies. In an enzymatic assay in homogenized and whole HEK-293 cells overexpressing 17β-HSD3, the inhibitor RM-532-105 efficiently inhibited the conversion of natural substrate 4-dione (50nM) into T with an IC50 of 26nM and 5nM, respectively. Moreover, the inhibitor RM-532-105 (10mg/kg) reached a plasma concentration of 250ng/mL at 7h (AUC 24h: 3485ngh/mL) after subcutaneous (s.c.) injection in the rat. In order to mimic the human situation in which 4-dione is converted to T in the testis, we used intact rats. Treatment for 7 days with 17β-HSD3 inhibitor RM-532-105 by s.c. injection or oral gavage exerted no effect on the testis, prostate and seminal vesicle weight and no modification in the levels of plasma steroids. However, after this treatment, the concentration of inhibitor in plasma increased depending on the dose. We thereafter determined the concentration of inhibitor in the testis and we discovered that the compound was slightly present. In fact, at 10mg/kg, the inhibitor RM-532-105 seems to have difficulty penetrating inside the testis and was found to be concentrated in the testicular capsule, and therefore unable to inhibit the 17β-HSD3 located inside the testis. However, with a higher dose of 50mg/kg injected s.c. in rats, RM-532-105 significantly decreased the level of T and dihydrotestosterone measured in plasma at 2h.
•17β-HSD3 is involved in the formation of androgenic hormone testosterone (T).•RM-532-105 is an androsterone derivative inhibiting 17β-HSD3.•RM-532-105 inhibits the formation of T in homogenized and ...whole HEK-293 cells overexpressing 17β-HSD3.•RM-532-105 (10mg/kg, s.c.) reached a plasma concentration of 250ng/mL at 7h in rat.
17β-Hydroxysteroid dehydrogenase type 3 (17β-HSD3 or HSD17B3) catalyzes the last step in the biosynthesis of the potent androgen testosterone (T), by stereoselectively reducing the C17 ketone of 4-androstene-3,17-dione (4-dione), with NADPH as cofactor. Since T plays an important role in androgen-sensitive diseases, this enzyme is thus an interesting therapeutic target. In an attempt to design compounds to lower the level of T, we synthesized androsterone derivatives substituted at position 3 as inhibitors of 17β-HSD3, and selected one of the most potent compounds for additional studies. In an enzymatic assay in homogenized and whole HEK-293 cells overexpressing 17β-HSD3, the inhibitor RM-532-105 efficiently inhibited the conversion of natural substrate 4-dione (50nM) into T with an IC50 of 26nM and 5nM, respectively. Moreover, the inhibitor RM-532-105 (10mg/kg) reached a plasma concentration of 250ng/mL at 7h (AUC 24h: 3485ngh/mL) after subcutaneous (s.c.) injection in the rat. In order to mimic the human situation in which 4-dione is converted to T in the testis, we used intact rats. Treatment for 7 days with 17β-HSD3 inhibitor RM-532-105 by s.c. injection or oral gavage exerted no effect on the testis, prostate and seminal vesicle weight and no modification in the levels of plasma steroids. However, after this treatment, the concentration of inhibitor in plasma increased depending on the dose. We thereafter determined the concentration of inhibitor in the testis and we discovered that the compound was slightly present. In fact, at 10mg/kg, the inhibitor RM-532-105 seems to have difficulty penetrating inside the testis and was found to be concentrated in the testicular capsule, and therefore unable to inhibit the 17β-HSD3 located inside the testis. However, with a higher dose of 50mg/kg injected s.c. in rats, RM-532-105 significantly decreased the level of T and dihydrotestosterone measured in plasma at 2h.
The stereoselective synthesis of furanic-steroid derivatives involving ring-closing metathesis and catalytic hydrogenation as key steps is described. The synthetic strategy was first illustrated by ...the synthesis of the furanic-estrane derivative
1 in seven steps starting from estrone and 2-methylene-propane-1,3-diol. This compound initially targeted as a potential inhibitor of 17β-hydroxysteroid dehydrogenase type 1 by a docking experiment was found to inhibit the enzyme. The scope of this new strategy was also extended to furanic-androstane derivatives by synthesizing compound
20.
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The involvement of aromatase, steroid sulfatase (STS) and reductive 17beta-hydroxysteroid dehydrogenases (17beta-HSDs) in the production of estrogens was determined in four cell lines of endometrial ...cancer (Ishikawa, HEC-1A, HEC-1B and RL-95) and one cell line of cervix cancer (Hela) in culture. After incubation with 4-androstene-3,17-dione (4-dione), there are no estrogens, estrone (E1) and estradiol (E2), detected suggesting that the pathway of aromatase is not important in these cell lines. In whole cells, the results show low percentages of transformation of estrone sulfate (E1S) into E1 suggesting that the entrance of E1S is difficult. However, in homogenized cells the STS activity was much higher and fully blocked by an inhibitor. Using selective inhibitors for each reductive 17beta-HSD (types 1, 5, 7 and 12), alone or in combination, we did not succeed in completely blocking the conversion of E1 into E2, suggesting that another 17beta-HSD (known or unknown) is involved in the formation of E2 from E1.
Estrogens play an important role in the development of breast cancer. Inhibiting 17beta-hydroxysteroid dehydrogenase type 1 (17beta-HSD1)--the enzyme responsible for the last step in the biosynthesis ...of the most potent estrogen, estradiol (E2)--would thus allow hindering the growth of estrogen-sensitive tumors. Based on a previous study identifying 16beta-benzyl-E2 (1) as a lead compound for developing inhibitors of the transformation of estrone (E1) into E2, we modified the benzyl group of 1 to improve its inhibitory activity. Three strategies were also devised to produce compounds with less residual estrogenic activity: (1) replacing the hydroxy group by a hydrogen at position 3 (C3); (2) adding a methoxy at C2; and (3) adding an alkylamide chain known to be antiestrogenic at C7. In order to test the inhibitory potency of the new compounds, we used the human breast cancer cell line T-47D, which exerts a strong endogenous 17beta-HSD1 activity. In this intact cell model, 16beta-m-carbamoylbenzyl-E2 (4m) emerged as a potent inhibitor of 17beta-HSD1 with an IC50 value of 44 nM for the transformation of 14C-E1 (60 nM) into 14C-E2 (24-h incubation). In another assay aimed at assessing the unwanted estrogenic activity, a 10-day treatment with 4m at a concentration of 0.5 microM induced some proliferation (38%) of T-47D estrogen-sensitive (ER+) breast cancer cells. Interestingly, when 4m (0.5 microM) was given with E1 (0.1 nM) in a 10-day treatment, it blocked 62% of the T-47D cell proliferation induced by E1 after its reduction to E2 by 17beta-HSD1. Thus, in addition to generating useful structure-activity relationships for the development of 17beta-HSD1 inhibitors, our study demonstrates that using such inhibitors is a valuable strategy for reducing the level of E2 and consequently its proliferative effect in T-47D ER+ breast cancer cells.
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