Caffeine Overcomes Genistein-Induced G2/M Cell Cycle Arrest in Breast Cancer Cells

Abstract

Although inhibition of tumor cell growth by genistein is mediated by different types of cell cycle arrest, its regulation of genes related to the cell cycle is not clear. In this study, genistein caused a concentration-dependent growth inhibition in the hormone-independent cell line MDA-MB-435S. Flow cytometric analysis showed that genistein induced a concentration-dependent accumulation of cells in the G2/M phase of the cell cycle. Caffeine enhanced the inhibition of cell proliferation induced by genistein. Caffeine alone did not have an appreciable effect on the phases of the cell cycle, but caffeine at 3 mM completely eliminated genistein-induced G2/M cell cycle arrest. cDNA microarrays were used to investigate the mechanism of genistein-induced, caffeine-negated G2/M arrest. We identified 12 genes, which had opposite responses to genistein and caffeine treatments. Among these, 5 genes were upregulated by genistein and downregulated by caffeine; 7 genes were downregulated by genistein and upregulated by caffeine. Reversal by caffeine of genistein-induced G2/M phase arrest in breast cancer cell lines could increase their sensitivity to genistein and enhance genistein-induced inhibition of cell growth. Genes that have opposite responses to genistein and caffeine may be involved in regulation of the G2/M phase of the cell cycle.

ACKNOWLEDGMENTS

We thank Ruiwen Zhang, Donald L. Hill, and Hui Wang for help in preparation of the manuscript. This work was supported by Grant 30271122 and 30671769 from National Natural Science Foundation of China and Grant BK2002027 from Natural Science Foundation of Jiangsu Province (to Z. Li). All authors have no personal or financial conflict of interest and have not entered into any agreement that could interfere with our access to the data on the research or on our ability to analyze the data independently, to prepare manuscripts, and to publish them.

Notes

^a Values are fold change to control. Fold change values represent the average of 2 independent microarray experiments. All genes listed are upregulated at least twofold by genistein. All genes listed are downregulated at least 2.5-fold (i.e., < 0.5 of control level) by caffeine.

^a :Abbreviations are as follows: CON, control; GE, genistein (20 μ M); CAFF, caffeine. GE + CAFF is 20 μ M GE plus 3 mM CAFF.

^b :Fold overexpression for microarray data based on ratio of fluorescence for treated group probe compared to control probe.

^c :Fold overexpression for northern blots based on densitometry from treated messenger RNA (mRNA) compared to control mRNA using β -actin levels for normalization.