Inhibition of cyclin-dependent kinases 2 and 4 activities as well as induction of cdk inhibitors p21 and p27 during growth arrest of human breast carcinoma cells by (?)-epigallocatechin-3-gallate

(-)-Epigallocatechin-3-gallate (EGCG) potently inhibits cell proliferation and suppresses tumor growth both in vitro and vivo, but little is known regarding the cell cycle regulatory proteins mediating these effects. This study investigated the effects of EGCG and other catechins on the cell cycle progression. DNA flow cytometric analysis indicated that 30 microM of EGCG blocked cell cycle progression at G1 phase in asynchronous MCF-7 cells. In addition, cells exposed to 30 microM of EGCG remained in the G1 phase after release from aphidicolin block. Over a 24-h exposure to EGCG, the Rb protein changed from hyper- to hypophosphorylated form and G1 arrest developed. The protein expression of cyclin D1, and E reduced slightly under the same conditions. Immunocomplex kinase experiments showed that EGCG inhibited the activities of cyclin-dependent kinase 2 (Cdk2) and 4 (Cdk4) in a dose-dependent manner in the cell-free system. As the cells were exposed to EGCG (30 microM) over 24 h a gradual loss of both Cdk2 and Cdk4 kinase activities occurred. EGCG also induced the expression of the Cdk inhibitor p21 protein and this effect correlated with the increase in p53 levels. The level of p21 mRNA also increased under the same conditions. In addition, EGCG also increased the expression of the Cdk inhibitor p27 protein within 6 h after EGCG treatment. These results suggest that EGCG either exerts its growth-inhibitory effects through modulation of the activities of several key G1 regulatory proteins such as Cdk2 and Cdk4 or mediates the induction of Cdk inhibitor p21 and p27.