Characteristics of human types 1, 2 and 3 17β-hydroxysteroid dehydrogenase activities: Oxidation/reduction and inhibition

doi:10.1016/0960-0760(95)00209-X

The Journal of Steroid Biochemistry and Molecular Biology

Volume 55, Issues 5–6, December 1995, Pages 581-587

https://doi.org/10.1016/0960-0760(95)00209-X Get rights and content

Abstract

Following transfection of types 1, 2 and 3 17β-hydroxysteroid dehydrogenase (17β-HSD) cDNAs into transformed embryonal kidney (293) cells, we have characterized the selective directional and inhibitory characteristics of these activities. While homogenates of transfected cells could catalyze interconversion of the substrate and product, in agreement with the general belief on the activity of these enzymes, the same activities measured in intact cells, in order to better reflect the physiological conditions, showed an unidirectional reaction. Types 1 and 3 17β-HSD catalyzed the reduction of estrone to estradiol and 4-androstenedione to testosterone, respectively, while type 2 17β-HSD catalyzed the oxidative transformation of both testosterone and 17β-estradiol to 4-androstenedione and estrone, respectively. In addition, types 1, 2 and 3 17β-HSD activities showed different pH optima. While types 1 and 3 showed pH optimum values centered at around 5 and 6, respectively, type 2 17β-HSD activity, which preferentially catalyzes the oxidation reaction, has higher activity at an alkaline pH (8–10). Differences in the optimum incubation temperatures were also observed: type 1 17β-HSD shows a relatively high temperature tolerance (55°C). In contrast, type 2 and 3 functioned best at 37°C. Types 1, 2 and 3 17β-HSD activities could be also differentiated by their sensitivity toward various specific inhibitors: type 1 was potently inhibited by an estradiol derivative containing a bromo/or iodopropyl group at position 16α. On the other hand a derivative of estrone containing a spiro-γ-lactone at position 17 showed a potent inhibitory effect on type 2 17β-HSD, whereas type 3 was strongly inhibited by 1,4-androstadiene-1,6,17-trione.

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Citation Excerpt :
Physiologically, testosterone is predominantly formed in the testicular Leydig cells by carbonyl reduction of its precursor androstenedione at position C17 (Fig. 1). This final step of the biosynthesis is catalyzed by the microsomal enzyme 17β-hydroxysteroid dehydrogenase type 3 (17βHSD3), which in turn needs the cofactor NADPH as a hydride provider [5–7]. Under pathological conditions, prostate cancer cells are able to express a noteworthy amount of 17βHSD3 which leads to higher testosterone concentrations in the malignant tissue.
Testosterone depletion is a common aim in the treatment of hormone-dependent prostate cancer, since the steroid boosts the tumor's proliferation. Therefore, inhibition of 17β-hydroxysteroid dehydrogenase type 3 (17βHSD3), which catalyzes the carbonyl reduction of androstenedione to testosterone, represents an expedient therapeutic drug target. Among the compounds targeting 17βHSD3, tetrahydrodibenzazocines have been reported to be highly potent inhibitors. Thus, we hypothesized that structural analogs to the tetrahydrodibenzazocine scaffold, namely diazocines, which contain an azo group instead of the ethylene moiety, are also able to inhibit 17βHSD3. Diazocines consist of a photoresponsive core and can be isomerized from Z into E configuration by irradiation with a specific wavelength. In the present study, 17βHSD3 inhibition by diazocine photoisomers was examined in transfected human embryonic kidney 293 cells (HEK-293) and isolated microsomes. For this purpose, cells or microsomes were treated with androstenedione and incubated for 2 or 24 h in the presence or absence of irradiated and non-irradiated diazocines. Testosterone formation was determined by uHPLC. We report a weak inhibition of 17βHSD3 activity by diazocines in HEK-293 cells and microsomes. Furthermore, we found no significant difference between samples treated with irradiated and non-irradiated diazocines in terms of inhibition. However, we detected a new compound by HPLC analysis, which only appeared in light-treated samples, indicating a chemical modification of the photoswitched diazocines, presumably rendering them ineffective. Further investigations revealed that this modification occurs in the presence of reducing agents like dithiothreitol and glutathione. A preliminary mass-spectrometric analysis suggests that the N–N double bond is reduced, resulting in a dianiline derivative. Nevertheless, optimized photoswitchable diazocine derivatives, which are stable in a cellular environment, might serve as potent 17βHSD3 inhibitors, effective only in irradiated tissue.
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Functional aspects of 17β-estradiol dehydrogenaseCharacteristics of human types 1, 2 and 3 17β-hydroxysteroid dehydrogenase activities: Oxidation/reduction and inhibition

Abstract

J. Biol. Chem.

J. Steroid Biochem. Molec. Biol.

J. Biol. Chem.

J. Steroid Biochem.

J. Biol. Chem.

J. Steroid Biochem.

FEBS Lett.

Molec. Cell. Endocr.

Bioorg. Med. Chem. Lett.

Bioorg. Med. Chem. Lett.

Structure of two in tandem human 17β-hydroxysteroid dehydrogenase genes

Molec. Endocr.

Steroid modulation of 17β-hydroxysteroid oxidoreductase activities in human placental villi in vitro

J. Clin. Endocr. Metab.

Human ovarian 17β-hydroxysteroid oxidoreductase activity

A comparison of normal and polycystic ovarian tissues

J. Clin. Endocr. Metab.

Testicular 17β-hydroxysteroid dehydrogenase: molecular properties and reaction mechanism

Steroids

Male pseudohermaphroditism caused by mutations of testicular 17β-hydroxysteroid dehydrogenase 3

Nature Genet.

The in vitro and in vivo uptake and metabolism of steroid in human adipose tissue

J. Clin. Endocr. Metab.

Quantitative studies on the uptake and metabolism of estrogens and progesterone by human endometrium

Endocrinology

Metabolism of oestrone and oestradiol-17β in human liver in vitro

Nature

Functional aspects of 17β-estradiol dehydrogenase
Characteristics of human types 1, 2 and 3 17β-hydroxysteroid dehydrogenase activities: Oxidation/reduction and inhibition