Abstract P6-07-05: Estrogen-induced carcinogenesis of normal breast cells via regulating NRF2-mediated pathway

Abstract

Abstract Background: Breast cancer is the most common malignancy in women and is known to be associated with steroid estrogen (E2). The importance of E2 in the etiology of breast cancer has been widely recognized. Most postmenopausal breast cancer is E2-dependent, mainly due to the continuous growth and proliferation of potent biological E2. E2-induced oxidative stress is involved in this carcinogenic process. The objective of the present study was to characterize the mechanism of E2-induced carcinogenesis of normal breast cells.Methods: MCF-10A and MCF-12A cells were treated with different doses of E2 for 48h, the cells were harvested and extracted for MTT assay to detect cell viability. MCF-10A and MCF-12A cells were treated with E2 for up to 48h. The 8-Hydroxydeoxyguanosine (8-OhdG) and lactate dehydrogenase (LDH) leakage were measured using the Oxiselect Oxidative DNA damage ELISA kit and LDH assay kits, respectively. The Annexin V Apoptosis Detection Kit FITC and terminal deoxynucleotidyl transferase dUTP nick end labeling (TUNEL) assay were used to detect cell apoptosis. Colony formation and migration assays were performed to examine the effect on colony formation and migration of cells by E2. Intracellular reactive oxygen species (ROS) status was measured using the 2',7'-dichlorodihydrofluorescein diacetate (DCFH-DA) method. Levels of oxidative stress markers were determined by measuring total superoxide dismutase (SOD), glutathione peroxidase (GSH-Px), catalase (CAT) and total antioxidant capacity (TAC) using commercial kits. Western blot analysis and RT-qPCR were conducted to respectively evaluate protein and mRNA expressions of the nuclear factor erythroid 2 like 2 (NRF2), HO-1 and NQO1.Results: In this study, we explored the E2-induced carcinogenesis of normal breast cells and uncovered the signaling pathways involved in these processes. Our results demonstrate that 5nM E2 treatment induces cell viability, colony formation, and migration, reduces cell apoptosis. The results of LDH and 8- OhdG tests reveal that E2 could reduce LDH and 8-ohdg levels. The results of ROS level and related oxidative stress indicators identity that E2 could reduce the expression of ROS, SOD, GPX, CAT, and TAC. Furthermore, the Western blot analysis and RT-qPCR results show that E2 could induce the expression of NRF2, HO-1, and NQO1.Conclusions: Taken together, these results suggest that E2-induced breast carcinogenesis via induction of NRF2-mediated pathways.Keywords: Estrogen, MCF-10A, MCF-12A, ROS, NRF2. Funding: Guangdong Natural Science Foundation No. 2017A030313719;Foundation Project of Guangzhou University of Traditional Chinese Medicine, No. XKP2019002. Citation Format: Dandan Chen, Yuzhu Zhang, Ling Zhu, Renxiong Wei, Qianjun Chen. Estrogen-induced carcinogenesis of normal breast cells via regulating NRF2-mediated pathway [abstract]. In: Proceedings of the 2019 San Antonio Breast Cancer Symposium; 2019 Dec 10-14; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2020;80(4 Suppl):Abstract nr P6-07-05.