NF-E2-related factor-2 (Nrf2) is a transcription factor regulating major cellular defense mechanisms ubiquitously expressed at low levels in all human tissues and maintained in cytoplasm under normal conditions. Its rapid activation in inducing phase II enzymes expression is important for preventing cancer and other cellular diseases. Under redox stress, Nrf2 is phosphorylated and translocated into the nucleus, activating genes through AREs. High constitutive activation of Nrf2 and its downstream gene expression is reported in many tumors and cancer cell lines; it protects them from cytotoxic effects of anticancer therapies. Dual role of Nrf2 (redox signaling and apoptotic regulation) discovery in cells brought safety concerns with respect to Nrf2 activators use for disease prevention; however, Nrf2 is induced transiently in normal cells, and functional Keap1 reduces Nrf2 to basal levels upon redox balance restoration.

Conversion of Nrf2 mediated redox signaling to apoptosis induction during cancer therapy is a novel strategy to improve therapeutic response in cancer without developing drug resistance. This requires understanding of Nrf2 regulation and identification of its activators that may play a role in cancer cell sensitization to chemotherapy. We developed Nrf2-luciferase fusion protein to measure nuclear Nrf2 translocation in response to its activators to examine the efficiency of redox signaling during chemotherapy in cancer, constructed luciferase fusion with N-terminal Nrf2 fragments , and measured luciferase activity in transfected cells in response to Nrf2 activators, and in the presence of anticancer drugs, evaluated redox signaling and apoptosis. We paid attention to exposure times and activators' concentrations, when cancer cells response switches from protective to apoptotic as a result of Nrf2 activation, by simultaneously measuring redox and apoptosis effect of antioxidant/drug combination.

Redox signaling in cells treated with EGCG, PTS, Rx001 or cisplatin remained unchanged when compared to baseline, while it at least doubled or tripled when treated with Rx001/EGCG or cisplatin/EGCG. When treated with cisplatin and Rx001 in combination with increasing concentrations of Nrf2 activators (EGCG and PTS) apoptosis significantly increased in as quickly as 24h when co-treated with EGCG, while PTS protected cells. Expression of SOD, the downstream target of Nrf2, rose upon exposure to anticancer drugs/activators applied in combination, as compared to anticancer drug treatment alone. Nrf2-Fluc2 fusion construct can be a valuable tool for monitoring anti-cancer therapy by using tumor cell xenografts expressingit to study Nrf2-ARE signaling role in cancer therapy. Antioxidant mediated chemosensitization requires separate evaluation for every type of cancer and chemotherapeutic drug/antioxidant combination, but is a promising approach for a personalized therapy.

Citation Format: Kira Foygel, Thillai V. Sekar, Ramasamy Paulmurugan. Monitoring antioxidant induced chemosensitization in triple negative breast cancer cells by Nrf2-luciferase fusion protein. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 5120. doi:10.1158/1538-7445.AM2014-5120