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Research Articles: Therapeutics, Targets, and Development

Induction of growth arrest and apoptosis in human breast cancer cells by 3,3-diindolylmethane is associated with induction and nuclear localization of p27^kip

¹ Department of Pathology, Karmanos Cancer Institute, Wayne State University School of Medicine, Detroit, Michigan and ² Medical Oncology, Peninsula Regional Medical Center, Salisbury, Maryland

Requests for reprints: Fazlul H. Sarkar, Department of Pathology, Karmanos Cancer Institute, Wayne State University School of Medicine, 9374 Scott Hall, 540 East Canfield, Detroit, MI 48201. Phone: 313-576-8327; Fax: 313-576-8389; E-mail: sarkarf{at}karmanos.org

Abstract

3,3'-Diindolylmethane (DIM) is a stable condensation product of indole-3-carbanol, a potential breast cancer chemoprevention agent. Human breast cancer cell lines were studied to better understand its mechanisms. In vitro experiments were done in MCF-7, T47D, BT-20 and BT-474 cells using MTT, ELISA, immunoblotting assays, reverse transcription-PCR, protein half-life, confocal microscopy, cell fractionation, and immunoprecipitation assays. We found that DIM inhibited the growth of all four breast cancer cell lines (IC₅₀s, 25-56 µmol/L). Because BT-20 and BT-474 overexpressed Her-2 and activated Akt, and BT-20 lacks estrogen receptor, these were studied further. In both cell lines, DIM appeared to induce expression of p27^kip protein before the loss of cell viability and apoptosis. In BT-20 cells, DIM also inhibited expression of activated Akt, but this appeared after p27^kip induction. In both cell lines, DIM induced p27^kip transcript expression within 6 h. DIM prolonged the p27^kip protein half-life in BT-20 but not BT-474 cells. We also showed, for the first time, that DIM induced nuclear localization of p27^kip in both cell lines. Moreover, in BT-20 cells, DIM induced a decrease in p27^kip phosphorylation at Thr¹⁸⁷, and its association with the 14-3-3 protein, which helped to explain the protein half-life increase and nuclear localization, respectively. DIM modulates p27^kip through transcription, prolongation of protein half-life, and nuclear localization. These effects appear to be independent of Her-2, Akt, or estrogen receptor status and should support further study for its chemoprevention potential in breast cancer. [Mol Cancer Ther 2008;7(2):341–9]

The costs of publication of this article were defrayed in part by the payment of page charges. This article must therefore be hereby marked advertisement in accordance with 18 U.S.C. Section 1734 solely to indicate this fact.

Note: Z. Wang and B.W. Yu equally contributed to this work.

³ Unpublished data.

Received 7/16/07; revised 11/16/07; accepted 12/28/07.