Previous studies have found that AhR is overexpressed and constitutively activated in several breast carcinoma cell lines and the expression levels showed strong correlation with the degree of the tumor malignancy. The objective of this study is to examine the effect of reducing the expression levels of AhR in metastatic human breast carcinoma (HBC) cell lines on their tumorigenic properties; specifically cellular growth, invasion into matrigel, and migration. We designed a series of short hairpin RNA (shRNA) targeting AhR and cloned them in pSuper retroviral vectors, and viruses expressing shRNA were used to infect a battery of HBC cell lines including MDA-MB231, MDA-MB468, BT549 and MT2 to generate cell lines that permanently express siRNA targeting AhR. Clonal cell lines with 50%-90% knockdown of AhR were isolated and characterized in vitro for their tumorigenic properties in comparison to the scramble RNA-expressing control cell lines. Cell cycle analysis by fluorescence-activated cell sorting showed that knock down of AhR expression decreases transition of cells from G1 to S and their progression to G2/M phases of cell cycle. This was seen as an increase of cells in G0/G1 (49% to 61%) and a decreased number of cells in S (26% to 21%) and G2/M phases (25% to 17%). This effect was coupled with a prolonged doubling time and a decreased cellular proliferation. Cells with AhR knock-down exhibited a decreased anchorage independent growth; forming less numbers of colonies in soft agar than control cells. Modified Boyden Chamber invasion assay coupled with fluorescence detection demonstrated that clones of HBC with AhR knock down levels exhibited substantially reduced invasiveness and diminished potential to invade matrigel. These clonal cell lines further exhibited reduced ability to migrate, as measured by scratch wound test. These observations collectively demonstrate a major role for the AhR in regulating tumorigenic phenotypes of metastatic breast cancer. Currently breast cancer prevention research focuses on the identification of novel approaches and/or novel targets. Our findings may establish the potential of targeting AhR as one of the prevention strategies for breast cancer.

Supported by RR03032 15, CA91408-01 and DAMD17-02-01-0483 grants.

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 4077.

©2010 American Association for Cancer Research
2010