Paclitaxel induces the apoptosis in MCF10AT series of human breast epithelial cells: A model for proliferative breast disease and carcinoma

3217 Human proliferative breast disease (PBD) refers to a sequence of progressive morphologic changes, including hyperplasia that occurs in the breast tissue prior to breast cancer development. MCF10AT cell series of human breast epithelial cell includes normal MCF10A (10A), premalignant MCF10AT (10AT) and MCF10ATG3B (10ATG3B), and fully malignant MCF10CA1a (10CA1a) cells and serves as a unique model system for development of PBD and carcinoma. The effects of paclitaxel (PTX), anti-microtubule agent, in MCF10AT cell lineage were examined. PTX inhibited the proliferation of MCF10AT cell lineage in a time-dependent (24, 48 and 72 h) and a concentration-dependent (0~10 nM) manners. PTX (10 nM) inhibited the proliferation of 10A, 10AT, 10ATG3B and 10CA1a cells by 65, 56, 43 and 52%, respectively, at 48 h. Treatment of cells with PTX (10 nM) for 24 h induced the apoptosis with the percentage of cells in sub-G 1 phase being 23.6, 26.1, 25.2 and 10.2% in 10A, 10AT, 10ATG3B, and 10CA1a cells, respectively, showing less sensitiveness to 10CA1a tumor cells. However, further treatment of cells with PTX for 48 h resulted in G 1 -phase arrest. Treatment of cells with PTX (0~10 nM) for 24 h showed the appearance of DNA fragmentation (a hallmark of apoptosis) with less sensitiveness to 10CA1a tumor cells. PTX increased the protein expression of p53 by 87, 102, 812 and 84% in 10A, 10AT, 10ATG3B and 10CA1a cells, respectively, and induced cleaved poly(ADP-ribose) polymerase (PARP) fragmentation. These results correlated well to PTX-mediated induction of apoptosis with the decreased sensitivity of 10CA1a cells. In accordance with the G 1 -phase arrest, retinoblastoma (Rb), phospho-Rb and cyclin-dependent kinase (cdk)-2 proteins significantly decreased after PTX treatment. PTX increased the protein expression of cdk inhibitor, p21 Waf1 , by 2.57, 1.53 and 2.48 fold in 10A, 10AT and 10ATG3B cells, respectively, with the negligible detection in 10CA1a cells. The protein expression of p27 Kip1 , another cdk inhibitor, also significantly increased by 2.73, 2.22, 2.40 and 1.71 fold for 10A, 10AT, 10ATG3B and 10CA1a cells, respectively, after the treatment of PTX. Collectively, these data show that PTX inhibited cell proliferation by G 1 cell cycle arrest in MCF10AT cell lineage. PTX also induced the apoptosis of 10A, 10AT and 10ATG3B cells, whereas 10CA1a tumor cells were less inducible to PTX effects. The effects of PTX on DNA fragmentation, p53 protein expression and PARP cleavage contributed to the induction of apoptosis in 10A, 10AT and 10AT3B cells, with a lesser effect noted for 10CA1a tumor cells.
 Supported by Research Fund by Kangnung National University and Regional Technology Innovation (RTI 05-01-02) Program of the Ministry of Commerce, Industry and Energy (MOCIE)
 Keywords: MCF 10AT cells; paclitaxel; apoptosis; DNA fragmentation; p53; PARP