Estrogenic properties of naturally occurring prenylated isoflavones in U2OS human osteosarcoma cells: Structure-activity relationships.

Eight isoflavones derivatives, with isoprenyl and/or 7-methoxy substitution, isolated from Erythrina poeppigiana (Fabaceae) have been investigated for their estrogenic properties in receptor subtype-specific reporter gene assays. First we focused on their estrogen receptor alpha and beta (ERalpha and ERbeta) selectivity, second we addressed structure-activity relationships, using bone-derived human osteosarcoma cell line (U2OS cells) stably expressing ERalpha or transiently expressing ERbeta. Our results show that a substitution at position 3' together with a 7-methoxy substitution on the genistein skeleton is associated with a statistically significant activation of the ERalpha- and ERbeta-dependent reporter gene expression in U2OS cells starting from 0.1nM. Particularly, the 7-methoxy-3'-isoprenyl (1) and the 7-methoxy-3'-(3-methyl-2-hydroxybuten-3-yl) (3) derivatives of genistein induces an ERalpha- and ERbeta-coupled luciferase activity at a concentration ten times lower than that of genistein, for which a statistically significant effect was observable at 1nM. On the other hand, isoprenyl substitution at position 6 of the A ring, compound 5, seems to have very little impact on the genistein ability to induce ER-coupled luciferase activity in U2OS cells, while a double prenylation at positions 8 and 3', compound 7, is associated with an almost complete loss of function on the reporter gene activation in U2OS-ERalpha, but in ERbeta expressing system the effectiveness remains on a statistically significant level, demonstrating an "exclusive ERbeta-selectivity" in U2OS human osteosarcoma cells, and therefore 7 can be considered as an isotype-selective ER ligand. Finally all the tested isoflavones derivatives appear to exhibit a slightly pronounced ERbeta preference, depending upon the position and the nature of the substituent moiety on the isoflavone skeleton. The estrogen-like effect of these prenylated isoflavone derivatives could be inhibited by the pure ER antagonist ICI 182 780, indicating that these effects were primarily mediated through ERs.