Oestradiol (E2) stimulates the growth of hormone-dependent breast cancer. 17b-hydroxysteroid dehydrogenases (17b-HSDs) catalyse the pre-receptor activation/inactivation of hormones and other substrates. 17b-HSD1 converts oestrone (E1) to active E2, but it has recently been suggested that another 17b-HSD, 17b-HSD12, may be the major enzyme that catalyses this reaction in women. Here we demonstrate that it is 17b-HSD1 which is important for E2 production and report the inhibition of E1-stimulated breast tumor growth by STX1040, a non-oestrogenic selective inhibitor of 17b-HSD1, using a novel murine model. 17b-HSD1 and 17b-HSD12 mRNA and protein expression, and E2 production, were assayed in wild type breast cancer cell lines and in cells after siRNA and cDNA transfection. Although 17b-HSD12 was highly expressed in breast cancer cell lines, only 17b-HSD1 efficiently catalysed E2 formation. The effect of STX1040 on the proliferation of E1-stimulated T47D breast cancer cells was determined in vitro and in vivo. Cells inoculated into ovariectomised nude mice were stimulated using 0.05 or 0.1 lg E1 (s.c.) daily, and on day 35 the mice were dosed additionally with 20 mg/kg STX1040 s.c. daily for 28 days. STX1040 inhibited E1-stimulated proliferation of T47D cells in vitro and significantly decreased tumor volumes and plasma E2 levels in vivo. In conclusion, a model was developed to study the inhibition of the major oestrogenic 17b-HSD, 17b-HSD1, in breast cancer. Both E2 production and tumor growth were inhibited by STX1040, suggesting that 17b-HSD1 inhibitors such as STX1040 may provide a novel treatment for hormone-dependent breast cancer.

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Multidrug resistance mediated by P-glycoprotein (Pgp) or multidrug-resistance-associated protein (MRP) remains a major obstacle for successful treatment of cancer. Inhibition of Pgp and MRP transport is important for high efficacy of anticancer drugs. While several Pgp inhibitors have entered clinical trials, the development of specific MRP1 inhibitors is still in its infancy. In our screening program, we have identified a pyrrolopyrimidine (4) as a novel and selective MRP1 inhibitor. Subsequent SAR work on the 4-position of the template revealed the phenethylpiperazine side chain as a potent replacement of the benzylthio group of the lead molecule. Introduction of groups at the 2-position seems to have no detrimental effect on activity. Modifications to the nitrile group at the 7-position resulted in the identification of analogues with groups, such as amides, with superior pharmacokinetic profiles. In vivo efficacy has been demonstrated by xenograft studies on selected compounds.

The 17beta-hydroxysteroid dehydrogenases (17beta-HSDs) catalyze the interconversion between the oxidized and reduced forms of androgens and estrogens at the 17 position. The 17beta-HSD type 1 enzyme (17beta-HSD1) catalyzes the reduction of estrone to estradiol and is expressed in malignant breast cells. Inhibitors of this enzyme thus have potential as treatments for hormone dependent breast cancer. Here we report the syntheses and biological evaluation of novel inhibitors based on the estrone or estradiol template. These have been investigated by modification at the 6, 16 or 17 positions or combinations of these in order to gain information about structure-activity relationships by probing different areas in the enzyme active site. Activity data have been incorporated into a QSAR with predictive power, and the X-ray crystal structures of compounds 15 and 16c have been determined. Compound 15 has an IC50 of 320 nM for 17beta-HSD1 and is selective for 17beta-HSD1 over 17beta-HSD2. Three libraries of amides are also reported that led to the identification of inhibitors 19e and 20a, which have IC50 values of 510 and 380 nM respectively, and 20 h which, having an IC50 value of 37 nM, is the most potent inhibitor of 17beta-HSD1 reported to date. These amides are also selective for 17beta-HSD1 over 17beta-HSD2.