A115

The present study investigated the synergistic inhibitory effects of polyphenon E (PPE, a standardized green tea polyphenol preparations containing appromately 65% EGCG), and atorvastatin (ATST) in a 4-(methylnitrosaminao)-1-(3-pyridyl)-1-butanone (NNK)-induced lung tumorigenesis model and H1299 human lung cancer cell line. Female A/J mice were given two weekly i.p. injections of NNK (150 mg/kg total dose), and one week later, they were treated with PPE (0.25% or 0.5% in drinking fluid), ATST (200 ppm or 400 ppm in diet), or PPE (0.25%) plus ATST (200ppm) for 16 weeks. With the single agents, only the high doses were effective in inhibiting lung tumorigenesis. However, the combination of PPE/ATST significantly reduced both the tumor multiplicity and tumor burden (by 56% and 55%, respectively). Synergistic reduction of tumor multiplicity (p = 0.0006) and tumor burden (p = 0.0009) by this combination was indicated by the isobologram analysis, which is used for studying the interaction of two agents. This conclusion was further supported by an isobologram analysis of the inhibitory effect of this combination in H1299 human lung cancer cells, where the combination of these two agents synergistically decreased the number of viable cells.
 >Immunohistochemistry analyses on the lung tumor tissues demonstrated that the combination of low doses PPE and ATST significantly induced apoptosis in tumors (3.2 fold of the control); whereas 0.5% PPE or 400ppm ATST alone increased the apoptotic indexes by 1.8 fold and 1.4 fold respectively. Induction of apoptosis was also found in H1299 cells by the TUNEL assay, and the combination of PPE/ATST was more efficient in enhancing the cleaved-caspase 3, 9, and PRPP levels than the single agent treatment. The induced apoptosis by the combination of PPE/ATST was possibly due to the reduced levels of anti-apoptotic proteins Mcl-1 and Bax. In both the NNK-induced lung tumors and H1299 cells, the Mcl-1 and Bax levels were reduced by PPE/ATST combination by Western blot and immunohistochemistry. The reductions of Mcl-1 and Bax levels by the combination were more significant than PPE or ATST alone. Cyclin D1 and hyper-phosphorylated Rb protein levels were also significantly reduced by this combination in H1299 cells, suggesting the inhibition of cell proliferation. The present work demonstrated that PPE and ATST synergistically inhibited NNK-induced lung tumorigenesis in mice and the growth of lung cancer H1299 cells. This effect is most likely due to the induction of cell apoptosis through inhibition of Mcl-1 (supported by NIH grant CA88961).

Sixth AACR International Conference on Frontiers in Cancer Prevention Research-- Dec 5-8, 2007; Philadelphia, PA