Abstract

Tibolone is used to treat climacteric complaints and prevent osteoporosis. These beneficial effects are exerted via its 3α-and 3β-hydroxymetabolites. Undesirable stimulation of the breast and endometrium is not apparent. Endometrial stimulation is prevented by the progestogenic activity of its Δ⁴-ene metabolite. The enzymes responsible for the formation of these active metabolites are unknown. Human aldo-keto reductase (AKR)1C isoforms have been shown to act as 3α/3β-hydroxysteroid dehydrogenases (HSDs) on 5α-dihydrotestosterone (5α-DHT). We show that AKR1Cs also efficiently catalyze the reduction of the Δ⁵⁽¹⁰⁾-3-ketosteroid tibolone to yield 3α- and 3β-hydroxytibolone. Homogeneous recombinant AKR1C1, AKR1C3, and AKR1C4 gave similar catalytic profiles to those observed with 5α-DHT. AKR1C1 catalyzed exclusively the formation of 3β-hydroxytibolone, AKR1C3 showed weak 3β/3α-HSD activity, and AKR1C4 acted predominantly as a 3α-HSD. Whereas AKR1C2 acted as a 3α-HSD toward 5α-DHT, it functioned exclusively as a 3β-HSD on tibolone. Furthermore, strong substrate inhibition was observed for the AKR1C2 catalyzed reduction of tibolone. Using NAD⁺, the 3-hydroxymetabolites were efficiently oxidized by homogeneous recombinant AKR1C2 and AKR1C4. However, because of potent inhibition of this activity by NADPH, AKR1Cs will probably act only as 3-ketosteroid reductases in vivo. Molecular docking simulations using crystal structures of AKR1C1 and AKR1C2 explained why AKR1C2 inverted its stereospecificity from a 3α-HSD with 5α-DHT to a 3β-HSD with tibolone. The preference for AKR1C1 and AKR1C2 to form 3β-hydroxytibolone, and the preference of the liver-specific AKR1C4 to form 3α-hydroxytibolone, may explain why 3β-hydroxytibolone is the major metabolite in human target tissues and why 3α-hydroxytibolone is the major circulating metabolite.