Background: Breast cancer is a leading cause of cancer-related death for women in the United States. Thus, there is a need to investigate novel prognostic markers and therapeutic strategies. Inflammation raises. Challenges to both treating and preventing the spread of breast cancer. Specifically, the nuclear factor kappa b (NFkB) pathway contributes to cancer progression by stimulating proliferation and preventing apoptosis. One target gene of this pathway is PTGS2, the gene that encodes for cyclooxygenase 2 (COX-2), which is upregulated in 40% of human breast carcinomas. COX-2 is an enzyme involved in inflammation. Here we investigate the effect of Singleminded 2s, a transcriptional tumor suppressor that is implicated in inhibition of tumor growth and metastasis, in regulating NFkB and COX-2. Methods: We utilized in vitro reporter assays, immunoblot analyses, qPCR and Immunohistochemical analysis to dissect the relationship between NFk, SIM2s, and COX-2. Furthermore, we utilized COX-2 targeting strategies to determine tumor suppressive activities. Results: Our results reveal that SIM2s attenuates the activation of a NFkB via luciferase reporter assays. Furthermore, immunostaining of lysates from breast cancer cells over expressing SIM2s showed reduction in various NFkB signaling proteins, whereas knockdown of SIM2 revealed increases in the same NFkB signaling proteins. Additionaly, by increasing NFkB translocation to the nucleus in DCIS.com cells, we show that NFkB signaling can act in a reciprocal manner to decrease expression of SIM2s. Likewise, suppressing NFkB translocation in DCIS.com cells increases SIM2s expression. We also found that NFkB/p65 represses SIM2 in via dose-dependent manner and when NFkB is suppressed the effect on SIM2s is negated. Additionally, our CHIP analysis confirms that NFkB/p65 binds directly to the SIM2 promoter site and that the NFkB sites in the SIM2 promoter are required for NFkB-mediated suppression of SIM2s. Finally over expression of SIM2s decreases PTGS2 in vitro and COX-2 staining in vivo while decreasing PTGS2 and/or Cox-2 activity results in re-expression of SIM2. Our findings identify a novel role for SIM2s in NFkB signaling and COX-2 expression. Conclusions: These findings provide evidence for a mechanism where SIM2s may represses COX-2 expression to provide an overall better prognosis for breast cancer patients.

Cross-talk between SIM2s and NFkB regulates cyclooxygenase 2 expression in breast cancer