A bis-sulphamoylated estradiol derivative induces ROS-dependent cell cycle abnormalities and subsequent apoptosis

作者：Visagie Michelle Helen; van den Bout Iman; Joubert Anna Margaretha

来源：PLos One, 2017, 12(4): e0176006.

DOI：10.1371/journal.pone.0176006

Clinical trials have revealed that the potential anticancer agent, 2-methoxyestradiol (2ME2) has limitations due to its low bioavailability. Subsequently, 2ME2 derivatives including (8R, 13S, 14S, 17S)-2-ethyl-13-methyl-7,8,9,11,12,13,14,15,16,17-decahydro-6H-cyclopenta[ a] phenanthrane-3,17-diyl bis(sulphamate) (EMBS) have shown improved efficacies in inducing apoptosis. However, no conclusive data exist to explain the mode of action exerted by these drugs. This study investigated the mode of action used by EMBS as a representative of the sulphamoylated 2ME2 derivatives. Hydrogen peroxide and superoxide production was quantified using dichlorofluorescein diacetate and hydroethidine. Cell proliferation and mitochondrial metabolism were investigated using crystal violet and Alamar Blue. Apoptosis was assessed using Annexin V-FITC while mitochondrial integrity was assessed using Mitocapture. Autophagy was visualised using LC3B II antibodies. The effects of EMBS on H2A phosphorylation and nuclei were visualised using phospho H2A antibody and 4', 6-diamidino-2-phenylindole, dihydrochloride. Data showed that EMBS exposure leads to increased reactive oxygen species (ROS) production which is correlated with loss of cell proliferation, mitochondrial membrane damage, decreased metabolic activity, G(2)/M arrest, endoreduplication, DNA double stranded breaks, micronuclei and apoptosis induction. Treatment of EMBS-exposed cells with the ROS scavenger, N-acetyl cysteine, abrogated ROS production, cell cycle arrest and apoptosis implying an essential role for ROS production in EMBS signaling. The inhibition of c-Jun N-terminal kinase (JNK) activity also inhibited EMBS-induced apoptosis suggesting that EMBS triggers apoptosis via the JNK pathway. Lastly, evaluation of LC3IIB protein levels indicated that autophagy is not activated in EMBS-exposed cells. Our data shows that EMBS targets a pathway that leads to increased ROS production as an early event that culminates in G(2)/M arrest and apoptosis by means of JNK-signaling in cancer cells. This study suggests a novel oxidative stress-dependent mode of action for sulphamoylated derivatives.

• 出版日期2017-4-14