High affinity 17alpha-substituted estradiol derivatives: synthesis and evaluation of estrogen receptor agonist activity

We synthesized four derivatives of 17beta-estradiol (E2) with an azide substitution on a 17alpha-side chain of varying length, namely 17alpha-(azidopropargyl)-3,17beta-estradiol (5), its 17beta-azido derivative (diazide 7), 17alpha-(5-azido-pent-1-ynyl)-3,17beta-estradiol (6) and 17alpha-(azidopentyn-2-yl)-3,17beta-estradiol (10). While most of the derivatives had low (7) or marginal (6 and 10) relative binding affinity (RBA) for both types of estrogen receptor (ERalpha and ERbeta), the RBAalpha and RBAbeta of 5 were practically identical to those of E2. The estrogenic activity of the derivatives was assessed using estrogen-responsive breast (MCF-7) and endometrial cancer (Ishikawa) cells. While 5 was a potent and effective inducer of alkaline phosphatase in Ishikawa cells and 7 was less potent but as effective as 5, 6 was marginally active and 10 was totally inactive in this respect. In the presence of 0.1 nM E2, however, 6 exhibited some ER antagonist activity at the highest concentration tested (1 microM). Similar results were obtained as regards the potency and efficacy of stimulation of MCF-7 cell proliferation and induction of luciferase gene expression in MCF-7:D5L cells, a clone stably transfected with an estrogen-responsive form of the gene. These data suggest that, while 5, 6, 7 and 10 interact with either type of ER in isolation, only 5 and 7 exhibit substantial ER agonist activity in the different estrogen-target cells examined, which could provide for photoaffinity labelling of the receptor in the cell as well as in isolation.