Histone Deacetylase Inhibitor Induction of P-Glycoprotein Transcription Requires Both Histone Deacetylase 1 Dissociation and Recruitment of CAAT/Enhancer Binding Protein β and pCAF to the Promoter Region

Although histone deacetylase (HDAC) inhibitors are appreciated as a promising class of anticancer drugs, recent reports show that P-glycoprotein (P-gp) is induced by HDAC inhibitor treatment in cancer cells, resulting in multidrug resistance of cancer cells to other chemotherapeutic agents. In this study, we investigated the molecular mechanism of HDAC inhibitor induction of P-gp expression. HDAC inhibitor treatment causes cell type–specific induction of P-gp expression without changes in the CpG methylation status of the promoter region. In addition, our data show that HDAC inhibitor does not alter the DNA binding activity of Sp1 but facilitates both the recruitment of a coactivator complex that includes CAAT/enhancer binding protein β and pCAF and the dissociation of the repressive complex, HDAC1, to the Sp1 binding region. Subsequently, the hyperacetylated histone H3 becomes enriched in the promoter region, leading to RNA polymerase II recruitment to activate P-gp gene transcription. Furthermore, specific down-regulation of HDAC1, but not HDAC2, by RNA silencing was enough to induce P-gp expression in HeLa cells, strongly supporting the essential role of HDAC1 in HDAC inhibitor induction of P-gp. Concomitantly, cell type–specific induction of P-gp expression seems to be dependent on phosphatidylinositol 3-kinase activity. Taken together, our findings show that HDAC inhibitor treatment leads to an increase in P-gp expression through dynamic changes in chromatin structure and transcription factor association within the promoter region. (Mol Cancer Res 2009;7(5):735–44) Introduction It has been well shown that histone deacetylase (HDAC) inhibitors alter the transcription of many tumor-associated genes and exhibit therapeutic activity against a variety of human malignancies, leading to phenotypic changes in the tumor, including growth arrest, morphologic reversion, differentiation, and apoptosis (1-6). It is also generally accepted that more active chromatin regions are associated with histone hyperacetylation and recruitment of histone acetyltransferases; histone deacetylation, which is associated with recruitment of HDACs, often restores these genomically active regions to a more repressed and condensed chromatin state. Thus, HDAC inhibitors are emerging as a promising class of chemotherapeutic agents against cancer. Several HDAC inhibitors, such as vorinostat [suberoylanilide hydroxamic acid (SAHA); refs. 7, 8], MS275 (9, 10), phenylbutyrate (11), and depsipeptide FK228 (12-14), have shown potent antitumor activities and are currently in phase I and II clinical trials. Recently, vorinostat (Zolinza, Merck & Co., Inc.) was approved for the treatment of cutaneous T-cell lymphoma. Multidrug resistance (MDR), in which cells are resistant to many structurally and functionally unrelated drugs, is a serious obstacle to cancer chemotherapy (15). The MDR phenotype is often due to overexpression of drug efflux pumps in the plasma membranes of cancer cells. Overexpression of P-glycoprotein (P-gp), an integral membrane protein encoded by the ABCB1 (MDR1) gene, is likely to be both a predominant contributor to this MDR phenotype and a marker for poor prognosis of several human cancers (16, 17). P-gp functions as a drug efflux pump that actively transports drugs from the inside to the outside of cells and causes a defect in the intracellular accumulation of drugs necessary for killing of cancer cells (16, 18, 19). Several reports show that P-gp expression can be silenced by epigenetic modifications involving methylation of CpG islands in the promoter region (20-24). Hypermethylations of CpG dinucleotides in the P-gp gene promoter region are associated with low expression of P-gp in various cell lines and tissues from patients (21, 23-25). This silencing can be reversed by treatment with a DNA methylation inhibitor (20, 21). In addition, accumulating evidence shows that HDAC inhibitors increase the expression of P-gp in cancer cells (26, 27), as well as peripheral blood mononuclear cells from patients (28), leading to chemoresistance in clinical applications and possibly limiting the clinical use of HDAC inhibitors. Recently, our group reported that apicidin, a HDAC inhibitor with broadspectrum antiproliferative activity against various cancer cell Received 6/24/08; revised 1/2/09; accepted 1/29/09; published OnlineFirst 5/12/09. Grant support: Korea Research Foundation Grant funded by the Korean Government (MOEHRD, Basic Research Promotion Fund; KRF-2007-314E00054 and KRF-2007-355-E00006). 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: Supplementary data for this article are available at Molecular Cancer Research Online (http://mcr.aacrjournals.org/). S-N. Kim and N.H. Kim contributed equally to this work. Requests for reprints: Yong Kee Kim, Department of Pharmacology, Kwandong University College of Medicine, 522 Naegok-dong, Gangneung, Gangwon-do 210-701, Korea. Phone: 82-33-649-7479; Fax: 82-33-641-1074. E-mail: [email protected] Copyright © 2009 American Association for Cancer Research. doi:10.1158/1541-7786.MCR-08-0296 Mol Cancer Res 2009;7(5). May 2009 735 on April 13, 2017. © 2009 American Association for Cancer Research. mcr.aacrjournals.org Downloaded from Published OnlineFirst May 12, 2009; DOI: 10.1158/1541-7786.MCR-08-0296