Abstract 1119: Role of BTG2 in the antioxidant response in breast cancer cells

Proceedings: AACR 101st Annual Meeting 2010‐‐ Apr 17‐21, 2010; Washington, DC This study shows a cytoprotective role of B-cell translocation gene 2, BTG2, in breast carcinoma cell (MCF7 and T47D) in response to oxidative stress, partly by upregulation of antioxidant response. We specifically propose that BTG2 may enhance the repair of oxidative DNA damage that in effect would be predicted to decrease the gene mutations. Our studies exploring the cytoprotective mechanism of BTG2 against oxidative stress include: the effects of BTG2 on the cellular redox state under basal conditions and in response to an oxidant; H2O2, as well as its effects on the antioxidant enzyme activities [superoxide dismutase 1 & 2, catalase, and glutathione peroxidases]; if BTG2 can stimulate the repair of oxidative DNA lesions (8-oxoguanine) and, if so, can BTG2 enhance the activity of the base excision repair (BER) pathway, the major pathway for the processing oxidative DNA damage. Briefly, the cellular levels of BTG2 are manipulated via transient transfection of wtBTG2 or BTG2-specific siRNAs. The cells are assayed for antioxidant enzyme activities using commercially available kits. Additionally, a set of transfected cells is exposed to a range of H2O2 doses then assayed for the cellular redox state using a commercially available kit. The measurement of the OGG1 enzyme activity, a commonly used assay to determine the efficiency of the BER pathway, is performed. RT-PCR and Western blotting confirmed the mRNA and protein levels of antioxidant/DNA repair enzymes. Our previous work indicates BRCA1 protects cells against oxidative stress partly by stimulating the expression of antioxidant genes (eg., GSTs) and by shifting the cellular redox balance. A new study suggests BRCA1 can stimulate the repair of 8-oxoguanine DNA lesions. Here, we evaluate the role of BTG2 as an intermediary for cytoprotective role of BRCA1. Specifically, if BRCA1 effects on antioxidant gene expression, cellular redox state, and repair of oxidized DNA are dependent upon the endogenous BTG2 and alternatively if wtBTG2 can restore these effects in the BRCA1-deficient mammary epithelial cells; investigating the structural basis of BRCA1/BTG2. We manipulated the levels of BRCA1 and BTG2 as noted above. The antioxidant response is measured as noted above. Additionally, we used a set of GST-BRCA1 and GST-BTG2 fusion proteins to establish the physical interaction between BRCA1 and BTG2. We found that wtBTG2 protects against oxidative stress in breast tumor cells. Additionally, antioxidant enzymes as well as their enzyme activity, i.e. catalase, superoxide dismutase 1 & 2 and glutathione peroxidases were upregulated by wtBTG2. wtBTG2 can protect BRCA1-deficient mammary epithelial cells from oxidative stress. Our studies will establish a role of BTG2 as a cytoprotective protein against oxidative DNA damage. It will also identify BTG2 as a potential molecular target for breast cancer prevention. Ultimately, BTG2 gene may also serve as a marker for the efficacy of tumor preventive agents. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 1119.