Substituted C19 steroid analogs as inhibitors of aromatase

Various C19-steroidal derivatives have been synthesized and evaluated in biochemical assays for their ability to inhibit the biosynthesis of estrogens. Steroids with substitutions on the A or B ring were prepared by Michael addition of various thiol reagents to appropriate dienone intermediates. An in vitro assay using human placental microsomes was used to evaluate aromatase-inhibitory properties. Synthesized compounds that exhibited high inhibitory activity were further evaluated under initial velocity conditions to determine apparent Ki values. Several 7 alpha-substituted androstenediones were effective competitive inhibitors and have apparent Ki values ranging from 18 to 69 nM, with the apparent Km for androstenedione being 63 nM. The most effective competitive inhibitor tested is 7 alpha-(4'-amino)phenylthioandrost-4-ene-3,17-dione with an apparent Ki of 18 nM. Derivatives of this 7 alpha-thioether compound that contain alkylating moieties have been prepared as potential irreversible enzyme inhibitors and demonstrate varying abilities to inactivate the aromatase enzyme. The results of these studies demonstrate that large chemical functionalities such as an aromatic ring with polar substituents can be accommodated in or near the active site of aromatase and, in some cases, can enhance the affinity of the enzyme for the inhibitors.