Selective estrogen receptor modulator BC-1 activates antioxidant signaling pathway in vitro via formation of reactive metabolites

Benzothiophene compounds are selective estrogen receptor modulators (SERMs), which are recently found to activate antioxidant signaling. In this study the molecular mechanisms of antioxidant signaling activation by benzothiophene compound BC-1 were investigated.

Benzothiophene compound BC-1 activates ARE signaling via reactive metabolite formation that is independent of estrogen receptors.

HepG2 cells were stably transfected with antioxidant response element (ARE)-luciferase reporter (HepG2-ARE cells). The expression of nuclear factor erythroid 2-related factor 2 (Nrf2) in HepG2-ARE cells was suppressed using siRNA. The metabolites of BC-1 in rat liver microsome incubation were analyzed using LC-UV and LC-MS.

Addition of BC-1 (5 μmol/L) in HepG2-ARE cells resulted in a 17-fold increase of ARE-luciferase activity. Pretreatment with the estrogen receptor agonist E2 (5 μmol/L) or antagonist ICI 182,780 (5 μmol/L) did not affect BC-1-induced ARE-luciferase activity. However, transfection of the cells with anti-Nrf2 siRNA suppressed this effect by 79%. Addition of BC-1 in rat microsome incubation resulted in formation of di-quinone methides and o-quinones, followed by formation of GSH conjugates. BC-1 analogues with hydrogen (BC-2) or fluorine (BC-3) at the 4' position did not form the di-quinone methides. Both BC-2 and BC-3 showed comparable estrogenic activity with BC-1, but did not induce ARE-luciferase activity in HepG2-ARE cells.