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Research Articles: Therapeutics, Targets, and Development

Reprogramming epigenetic silencing: artificial transcription factors synergize with chromatin remodeling drugs to reactivate the tumor suppressor mammary serine protease inhibitor

¹ Department of Pharmacology and Lineberger Comprehensive Cancer Center, University of North Carolina at Chapel Hill, Chapel Hill, North Carolina; and ² Department of Biochemistry and Biophysics, Yale University Medical Center, New Haven, Connecticut

Requests for reprints: Pilar Blancafort, Department of Pharmacology and Lineberger Comprehensive Cancer Center, University of North Carolina at Chapel Hill, Chapel Hill, NC 27599-7365. Phone: 919-966-1614. E-mail: pilar_blancafort{at}med.unc.edu

Abstract

Mammary serine protease inhibitor (maspin) is an important tumor suppressor gene whose expression is associated not only with tumor growth inhibition but also with decreased angiogenesis and metastasis. Maspin expression is down-regulated in metastatic tumors by epigenetic mechanisms, including aberrant promoter hypermethylation. We have constructed artificial transcription factors (ATFs) as novel therapeutic effectors able to bind 18-bp sites in the maspin promoter and reactivate maspin expression in cell lines that harbor an epigenetically silenced promoter. In this article, we have investigated the influence of epigenetic modifications on ATF-mediated regulation of maspin by challenging MDA-MB-231 breast cancer cells, comprising a methylated maspin promoter, with different doses of ATFs and chromatin remodeling drugs: the methyltransferase inhibitor 5-aza-2'-deoxycytidine and the histone deacetylase inhibitor suberoylanilide hydroxamic acid. We found that the ATFs synergized with both inhibitors in reactivating endogenous maspin expression. The strongest synergy was observed with the triple treatment ATF-126 + 5-aza-2'-deoxycytidine + suberoylanilide hydroxamic acid, in which the tumor suppressor was reactivated by 600-fold. Furthermore, this combination inhibited tumor cell proliferation by 95%. Our data suggest that ATFs enhance the efficiency of chromatin remodeling drugs in reactivating silenced tumor suppressors. Our results document the power of a novel therapeutic approach that combines both epigenetic and genetic (sequence-specific ATFs) strategies to reactivate specifically silenced regions of the genome and reprogram cellular phenotypes. [Mol Cancer Ther 2008;7(5):1080–90]

Grant support: NIH/National Cancer Institute grant 1R01CA125273-01, American Lung Association and LUNGevity Foundation grant LD-17098-N, V-Foundation Award, and Department of Defense Idea Award BC051475 (P. Blancafort) and National Cancer Institute grant 1 R21 CA116079-01 (P.M. Lizardi).

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.

³ Supplementary material for this article is available at Molecular Cancer Therapeutics (http://mct.aacrjournals.org/).

Received 8/ 2/07; revised 1/18/08; accepted 3/24/08.