Author/Editor     Starčević, Š; Kocbek, P; Hribar, G; Lanišnik-Rižner, L; Gobec, S
Title     Biochemical and biological evaluation of novel potent coumarin inhibitor of 17beta-HSD type 1
Type     članek
Source     Chem Biol Interact
Vol. and No.     Letnik 191, št. 1-3
Publication year     2011
Volume     str. 60-5
Language     eng
Abstract     Human 17beta-hydroxysteroid dehydrogenase (17beta-HSD) type 1 is an enzyme that acts at the pre-receptor level. It catalyzes the NADPH-dependent reduction of the weak estrogen estrone into the most potent estrogen 17beta-estradiol, which exerts proliferative effects via estrogen receptors. Overexpression of 17beta-HSD type 1 in estrogen-responsive tissues is related to the development of hormone-dependent diseases, such as breast cancer and endometriosis. 17beta-HSD type 1 thus represents an attractive target for development of new drugs. Recently, we discovered that substituted coumarin derivatives potently and selectively inhibit 17beta-HSD type 1. In the present study, we have performed additional biochemical and biological evaluation of the most promising coumarin derivative. First, we used an efficient method for isolation and purification of the active, soluble recombinant human 17beta-HSD type 1 from Escherichia coli. This 17beta-HSD type 1 showed a specific activity of 0.64+/-0.08mumolmin(-1)mg(-1) for estrone reduction in the presence of NADPH at pH 6.5, and a K(m) of 62nM for estrone. Next, we evaluated the best of the coumarin-derivative inhibitors, showing its reversible and competitive inhibition of 17beta-HSD type 1 reductive activity with a K(i) of 53nM. We confirmed that this coumarin inhibitor acts not only in a cell-free assay, but also decreases endogenous 17beta-HSD type 1 activity in human T-47D breast cancer cells. This inhibitor also reduced estrone dependent growth of T-47D cells after 48h of incubation.
Descriptors     COUMARINS
17-HYDROXYSTEROID DEHYDROGENASES
BREAST NEOPLASMS
TUMOR CELLS, CULTURED
CELL DIVISION
KINETICS
ESTROGENS
BIOLOGICAL ASSAY