Combination Treatment with Luteolin and Quercetin Enhances Antiproliferative Effects in Nicotine-Treated MDA-MB-231 Cells by Down-regulating Nicotinic Acetylcholine Receptors

Large-scale epidemiological cohort studies performed in the United States indicate that breast cancer risk is associated with active and passive smoking. As of yet, however, there is no direct evidence of antitumor effects by agents that block the effect of tobacco compound nicotine (Nic) on relevant nicotinic receptors (nAChR) involved in breast tumorigenesis. In the present study, the expression profiles of different nAChR subunits in the human breast cancer cell line (MDA-MB-231) were characterized by RT-PCR. Nic (>0.1 mu M, 6 h) significantly increased alpha 9-nAChR mRNA and protein expression levels in human breast cancer Cells (MDA-MB-231 cells). On the other hand, combined treatment with luteolin (Lut, 0.5 mu M),and quercetin (Que, 0.5 mu M) profoundly decreased MDA-MB-231 proliferation by down-regulating alpha 9-nAChR expression. MDA-MB-231 cells were cultured in soft agar to evaluate anchorage-independent colony formation; combined treatment of Lut + Que inhibited Nic-induced MDA-MB-231 colony formation. Interestingly, the number of colonies formed was profoundly reduced in alpha 9-nAChR knockdown (Si alpha 9) cells in the combined (Lut + Que)-treated group as compared to the relevant control groups. Such results show that Lut- or Que-induced antitransforming activities were not limited to specific inhibition of the alpha 9-nAChR receptor. Both alpha 5- and alpha 9-nAChR appear to be important molecular targets for Lut- and Que-induced antitumor effects in human breast cancer cells.