Abstract 1411: Regulation of osteoprotegerin expression in breast cancer cells by nuclear factor-kappaB

Abstract

Abstract Acquisition of osteoprotegerin (OPG) expression by breast epithelial cells appears to be linked to the formation of malignant breast neoplasia. Previous in vitro studies demonstrate that OPG expression by breast cancer cells increases resistance to tumor necrosis factor related apoptosis inducing ligand (TRAIL) mediated apoptosis, suggesting a tumor promoting role for OPG. Recently we have observed reduced ability to metastasize in the chick embryo model by MDA-MB-231 breast cancer cells after reducing OPG expression using RNAi methods, further substantiating a tumor promoting role of OPG. In order to direct inhibitory strategies, we investigated the regulation of OPG expression in human breast cancer cell lines. Based on regulation of OPG in other cell types, we hypothesized that OPG expression is regulated by the transcription factor Nuclear Factor-kappaB (NF-kappaB). OPG expression was examined in MDA-MB-231 (basal, triple negative), MCF-7 (luminal, ER-positive) and SK-BR-3 (HER-2 positive) breast cancer cells. We used real time RT-PCR and ELISA to confirm that all cell lines expressed OPG mRNA and protein, respectively. NF-kappaB activity was modulated using thymoquinone (TQ; 10 µM) and BMS345541 (1 µM; both inhibitors) and Tumor Necrosis Factor-alpha (TNF-alpha) and Interleukin-1-beta (IL-1-beta; 10 ng/ml, both activators). Specificity for these modulators was determined by NF-kappaB promoter driven luciferase reporter assay. In MDA-MB-231 cells, inhibition of NF-kappaB lowered basal levels of OPG mRNA and protein expression, suggesting constitutive NF-kappaB activity plays a role in OPG expression. TNF-alpha and IL-1-beta treatment increased OPG mRNA and protein expression significantly above basal levels. Inhibition of NF-kappaB ameliorated TNF-alpha and IL-1-beta induced OPG expression to baseline. In MCF-7 and SK-BR-3 cells, basal levels of OPG mRNA and protein expression were not reduced by the NF-kappaB inhibitors. TNF-alpha and IL-1-beta treatment increased OPG mRNA and protein expression significantly above baseline, however inhibition of NF-kappaB did not ameliorate the induction of OPG mRNA and protein expression. Our data suggests that the mechanisms regulating OPG expression in breast cancer cells may depend on breast cancer subtype. It appears in the basal cell line, MDA-MB-231, NF-kappaB's tumor promoting effects may be modulated in part by increasing OPG expression. The regulation of OPG expression in cells from other breast cancer subtypes requires further investigation. Citation Format: Michael Weichhaus, Linda Connelly. Regulation of osteoprotegerin expression in breast cancer cells by nuclear factor-kappaB. [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 1411. doi:10.1158/1538-7445.AM2014-1411