help button home button The Oncologist http://theoncologist.alphamedpress.org/misc/eLetters.shtml
HOME HELP CONTACT US SUBSCRIPTIONS ARCHIVE SEARCH TABLE OF CONTENTS
The Oncologist, Vol. 11, No. 1, 1-8, January 2006; doi:10.1634/theoncologist.11-1-1
© 2006 AlphaMed Press

This Article
Right arrow Abstract Freely available
Right arrow Full Text (PDF)
Right arrow eLetters: Submit a response to this article
Right arrow Alert me when this article is cited
Right arrow Alert me when eLetters are posted
Right arrow Alert me if a correction is posted
Right arrow Citation Map
Services
Right arrow E-mail this article link to a friend
Right arrow Similar articles in this journal
Right arrow Similar articles in PubMed
Right arrow Alert me to new issues of the journal
Right arrow Download to citation manager
Right arrow Reprints/Permissions
Citing Articles
Right arrow Citing Articles via HighWire
Right arrow Citing Articles via Google Scholar
Google Scholar
Right arrow Articles by Giacinti, L.
Right arrow Articles by Giordano, A.
Right arrow Search for Related Content
PubMed
Right arrow PubMed Citation
Right arrow Articles by Giacinti, L.
Right arrow Articles by Giordano, A.

Epigenetic Information and Estrogen Receptor Alpha Expression in Breast Cancer

Laura Giacintia,b, Pier Paolo Claudioa, Massimo Lopezb, Antonio Giordanoa,c

a Sbarro Institute for Cancer Research and Molecular Medicine, College of Science and Technology, Temple University, Philadelphia, Pennsylvania, USA; b Division of Medical Oncology B, Regina Elena Cancer Institute, Rome, Italy; c Department of Human Pathology and Oncology, Laboratory of Biomedical Technology and Oncology, Siena, Italy

Key Words. Breast cancer • Gene expression regulation • Estrogen receptors • Anticancer drug combinations

Correspondence: Antonio Giordano, M.D., Ph.D., Sbarro Institute for Cancer Research and Molecular Medicine, Center for Biotechnology, BioLife Sciences Bldg. 333, Temple University, 1900 North 12th Street, Philadelphia, Pennsylvania 19122-6009, USA. Telephone: 215-204-9520; Fax: 215-204-9522; e-mail: giordano{at}temple.edu

Received December 29, 2004; accepted for publication October 11, 2005.


   LEARNING OBJECTIVES
Top
 Learning Objectives
Abstract
 Introduction
 Epigenetic Information and...
 Regulation of ER-Promoter...
 Conclusion
 Disclosure of Potential...
 References
 
After completing this course, the reader will be able to:

  1. Describe the role of epigenetic information in the regulation of gene expression and in the development of cancer.
  2. Explain the important role of estrogen receptor expression in breast cancer as a prognostic marker and predictive factor of response to endocrine therapy.
  3. Evaluate the current and potential role that the comprehension of the molecular biology of a tumor may have in finding a new therapeutic approach in cancer treatment.

Access and take the CME test online and receive 1 AMA PRA category 1 credit at CME.TheOncologist.com

The CME activity for this article is forthcoming.


   ABSTRACT
Top
 Learning Objectives
 Abstract
Introduction
 Epigenetic Information and...
 Regulation of ER-Promoter...
 Conclusion
 Disclosure of Potential...
 References
 
In industrialized countries, breast cancer is the most common tumor in women. The tumor expression of estrogen receptors (ERs) is a very important marker for prognosis and a marker that is predictive of response to endocrine therapy. The loss of ER expression portends a poor prognosis and, in a significant fraction of breast cancers, this repression is a result of the hypermethylation of CpG islands within the ER-{alpha} promoter. Hypermethylation is one of the best known epigenetic events in mammalian cells. Over the last few years, many studies have found that other epigenetic events, such as deacetylation and methylation of histones, are involved in the complex mechanism that regulates promoter transcription. The exact interplay of these factors in transcriptional repression activity is not yet well understood. Inhibitors of some of these are currently being studied as new drugs able to restore ER-{alpha} protein expression in ER-{alpha}–negative breast cancer cells and to promote apoptosis and differentiation. Demethylating agents and histone deacetylase (HDAC) inhibitors are candidates for becoming potent new drugs in cancer therapy. This paper reviews the current understanding of the role of epigenetic information in the development of cancer and its significance in breast cancer as predictive markers of ER status and as new targets of anticancer therapy.


   INTRODUCTION
Top
 Learning Objectives
 Abstract
 Introduction
Epigenetic Information and...
 Regulation of ER-Promoter...
 Conclusion
 Disclosure of Potential...
 References
 
In industrialized countries, breast cancer is the most common tumor in women, and it is the most important cause of cancer-associated morbidity and mortality. Thanks to early diagnosis and the discovery of new drugs, cancer-associated mortality has decreased. Over the last few years, new molecular markers have been discovered as important targets for diagnosis and therapy. One goal is to identify a specific molecular assay for each breast tumor and then to obtain an individualized therapeutic approach.

Today, a variety of molecular factors is being used in clinical practice to predict the prognosis and response to therapy of breast cancer patients, and the tumor expression of estrogen receptors (ERs) is one of the most important. ERs are members of the nuclear receptor (NR) superfamily that mediates the effects of the steroid hormone estrogen in a diverse range of physiological processes. Estrogens regulate growth, differentiation, and homeostasis in eukaryotic cells but have been associated pathologically with a higher risk for breast and endometrial cancer [1]. In fact, breast cancer hormone dependence is correlated with tumor progression and patient prognosis [2]. As such, tumor ER expression plays an important role in the clinical care of breast cancer patients as a prognostic factor and therapeutic target. ER absence is correlated with a more malignant disease and poor prognosis.

Binding of 17-ß-estradiol (E2) to the active form of ERs (Fig. 1Go) induces tumor growth in ER-positive breast cancers. Therefore, reducing estrogen levels or altering the activity of the receptor can induce these cancers to regress. E2 action involves ligand-mediated activation of nuclear ER-{alpha} and ER-ß, which interact directly with estrogen response elements (EREs) in the promoter and/or enhancer regions of target genes and recruit various coactivators to mediate transcriptional regulation [3]. ER regulation is not limited to direct ligand binding. It can also be modulated by several other pathways that include the epidermal growth factor (EGF) and insulin-like growth factor 1 (IGF-1) mitogenic pathways and the second messengers cyclic AMP and dopamine [1, 4, 5].



View larger version (25K):
[in this window]
[in a new window]
  		Figure 1. Estrogen receptors trigger gene activation. Estrogen receptors normally reside in the cell’s nucleus. When an estrogen molecule enters a cell and binds to its receptor, this estrogen-receptor complex then binds to specific DNA sites, called estrogen response elements, near genes that are controlled by estrogens. This mechanism activates gene transcription and then synthesis of specific proteins that can influence cell behavior in different ways, depending on the cell type involved.

 
To date, two isoforms of ER (ER-{alpha} and ER-ß) have been identified, encoded by independent genes. Both ER isoforms show modular structure characteristics of the NR superfamily and, although both ER-{alpha} and ER-ß display similar binding affinities for E2, they have different roles in the regulation of gene expression [1]. ER-{alpha} was believed for many years to be the only ER. ER-ß was identified more recently, and it is still subject to ongoing research as to whether mutations and splice variance of this receptor play any role in breast cancer development and/or prognosis and in response to endocrine therapy [6–8]. The role of ER-ß in breast cancer remains elusive, but the presence of ER-{alpha} at the time of diagnosis is an indication for endocrine therapy.

Sixty percent of primary breast cancers are ER-positive and two thirds of advanced ER-positive breast cancers respond to therapy with antiestrogens (Fig. 2Go), such as tamoxifen (Nolvadex®; AstraZeneca Pharmaceuticals, Wilmington, DE, http://www.astrazenecaus.com) [6]. However, up to one third of breast cancers lack ER-{alpha} at the time of diagnosis, and a fraction of breast cancers that are initially ER-{alpha}–positive lose ER expression during tumor progression [9]. This deprives us of an important possibility of tumor patient care by endocrine therapy and increases poor clinical outcome. In a significant fraction of breast cancers, ER absence is a result of aberrant methylation of CpG islands, cytosine–guanine-rich areas that are located in the 5' regulatory regions of the ER-{alpha} gene [9–11].



View larger version (25K):
[in this window]
[in a new window]
  		Figure 2. Antiestrogen. Antiestrogens work by binding to estrogen receptors and blocking estrogen from binding to these receptors. This also blocks estrogen from activating specific genes and blocks the expression of the relative specific proteins.

 
Methylation of a specific site on a gene is an epigenetic event that causes transcriptional repression of that site. The genome contains two types of information: genetic and epigenetic; the epigenetic information provides instructions on how, where, and when the genetic information should be used. The most important known form of epigenetic information in mammalian cells is DNA methylation, the covalent addition of a methyl group to the 5' position of cytosine, predominantly within the CpG dinucleotide [9, 12, 13]. CpG islands are GpC- and CpG-rich regions of 1kilobase(kb) that are usually associated with the promoter or 5' end of genes. If a promoter of a tumor suppressor gene is hypermethylated, the expression of the associated gene is silenced and this event provides the cell with a growth advantage, as well as deletions or mutations [13].

Hypermethylation events can occur early in tumorigenesis, involving the disruption of pathways that may predispose cells to malignant transformation. Preneoplastic lesions often show aberrant methylation, and the frequency of aberrations increases with the progression of disease [13–15]. CpG island methylation is not random. The genes that are more susceptible to hypermethylation within their CpG islands are likely to be those that are involved in the regulation of cell growth, and so the cells that lack them could have a growth advantage [16]. ER-{alpha} promoter methylation is an example of cell growth advantage.

Breast cancer is usually a hormone-dependent tumor. Estrogens can regulate the growth of breast cells by binding with ERs. This means that exposure to estrogens could increase breast cancer incidence and proliferation, but also that tumor progression could be controlled by antiestrogenic drugs. Loss of ER expression means that breast cancer cells are no longer being regulated by estrogen in their growth and that they cannot be stopped by endocrine therapy. This results in higher tumor aggressiveness and poor prognosis. Therefore, ER-{alpha} is a critical growth regulatory gene in breast cancer, and its expression in breast cancer cells is critical for tumor progression.

ER transcriptional regulation is a result of a very complex mechanism, which is still poorly defined. Our comprehension of the mechanism that regulates transcriptional repression could allow us to interfere with it and to restore gene expression. This would permit us to re-establish cancer cell growth control by endocrine therapy. It could also be interesting to find new markers that may be predictive of lacking ER expression in primary breast cancers with an ER-positive phenotype. This finding could guide physicians in their choice between chemotherapy and endocrine therapy as a better treatment for patients with ER-positive breast cancer, in addition to the other prognostic factors and factors predicting therapy response used in clinical practice.


   EPIGENETIC INFORMATION AND REPRESSION OF TRANSCRIPTION
Top
 Learning Objectives
 Abstract
 Introduction
 Epigenetic Information and...
Regulation of ER-Promoter...
 Conclusion
 Disclosure of Potential...
 References
 
DNA methylation in mammalian cells is established and maintained by a complex interplay among three DNA methyltransferases, DNMT1, DNMT3a, and DNMT3b. DNMT1 is the most abundant and catalytically active DNA methyltransferase; it is thought to function as a maintenance methyltransferase by copying DNA methylation patterns from the parental to the daughter strand following DNA replication. DNMT3a and DNMT3b are referred to as de novo DNA methyltransferases. The function of a fourth DNMT family member, DNMT2, remains unknown [17].

Histone deacetylase (HDAC) family members remove acetyl groups from lysine residues of core histones, particularly H3 and H4, increasing ionic interactions between positively charged lysines of histones and negatively charged DNA, which results in a more compact nucleosome structure that limits transcription [9]. DNMT1 has been found to interact physically with either HDAC1 or HDAC2 through its N-terminus, thereby forming a transcriptionally inactive chromatin structure that represses transcription. Thus, DNA methylation and histone deacetylation function through a common mechanistic pathway to repress transcription [9, 18–21].

A direct interaction between the plant homeodomain (PHD) domain of DNMT3a and HDAC1 has been demonstrated, which is consistent with the finding that the majority of the repressive activity mediated by DNMT3a can be relieved by treatment with the HDAC inhibitor trichostatin A (TSA). The role of the interaction of DNMT3a and HDAC1 remains unclear, but it may involve targeting DNMT3a to transcriptionally silenced regions to carry out de novo methylation [17, 22]. Thus, the association of these two activities is likely to be critical for the establishment and maintenance of heterochromatic regions.

DNA methylation represses genes partly by recruitment of methyl-CpG-binding domain (MBD) proteins, which selectively recognize methylated CpG dinucleotides and tend to cluster at certain chromosomal loci [15, 23, 24]. An abundant chromosomal MBD protein, MeCP2, was the first protein identified as linking methylated DNA with HDAC-containing transcriptionally repressive complexes for gene silencing [25–27]. Subsequently, several MBD proteins have been identified that, similar to MeCP2, couple methylated DNA to HDAC [28, 29]. The recruitment of MeCP2 to methylated CpG dinucleotides represents a major mechanism by which DNA methylation can repress transcription [30, 31]. However, the inhibition of histone deacetylase activity using drugs such as TSA only partially relieves MeCP2-mediated transcriptional repression. This partial relief indicates that additional mechanisms of repression by MeCP2 likely exist in addition to the recruitment of histone deacetylase [25, 27, 32].

Recent findings show that MeCP2 acts as a bridge between DNA methylation and histone methylation, reinforcing the repressive chromatin state [32]. Histone methylation is an important epigenetic mechanism for the organization of chromatin structure and the regulation of gene expression. In particular, methylation at lysine 9 of histone H3 resulting from the activity of SUV39H1, a human homologue of the Drasophila position effect variegation modifier Su(rar)3–9, is associated with gene silencing [33]. MeCP2-mediated repression might also include a second stage involving histone methylation, in addition to histone deacetylation [32, 34, 35]. MeCP2 could be considered as a protein that can connect a repressive modification on DNA to a repressive modification on histones. Recent evidence from Neurospora crassa and Arabidopsis thaliana indicates that the reverse is also possible: methylation of histone H3 at lysine 9 leads to methylation of DNA [36, 37]. MeCP2 may set up a self-reinforcing cycle of repression by promoting further rounds of DNA methylation following histone methylation, and thus, these two global epigenetic events act together to perpetuate and maintain a repressed chromatin state [32]. In DNMT1 knockout human cancer cells (KO1), the loss of DNMT1 results in an overall change in the histone H3 modification pattern. The effect of the loss of DNMT1 is an increase in the acetylation of histone H3 and a decrease in the dimethylation and trimethylation of lysine 9 (TriMetK9H3). TriMetK9H3 is a marker for constitutive heterochromatin. This means that the loss of DNMT1 in human cells may be associated with the disorganization of heterochromatin. In addition, in these cells, we can observe that HDACs and HP1 (methyl-lysine binding proteins) no longer interact with histone H3 and pericentrometric repetitive sequences (satellite 2). Interaction of HP1 and MetK9H3 is involved in the silencing of specific promoters and in the formation of heterochromatin. In KO1 cells, this complex is totally abrogated. The hyperacetylation of H3 in KO1 is not the result of a loss of HDAC activity, but it may likely be the result of a loss of targeted HDAC activity at specific DNA sequences. It is possible that a change in DNA methylation also causes histone modification changes and that DNMT1 activity is important to preserve the correct organization of chromatin domains inside the cell nucleus [38].

Recent data also show that DNMT3a associates primarily with histone H3 Lys9 methyltransferase but also, to a lesser extent, with histone H3 Lys4 activity [39]. The mechanism by which histone H3 Lys4 methylation is linked to transcriptional silencing is currently unknown. The effect of histone H3 Lys4 methylation on transcription may well depend on its combination with other histone modifications, whereby methylation at Lys4 of histone H3 might function in concert with DNA methylation to repress transcription [32].

SUV39H1 is also involved in transcriptional repression by the retinoblastoma protein Rb [34, 40, 41]. SUV39H1 creates a high-affinity binding site for proteins of the HP1 family, and its activity is required for the proper localization of HP1 at heterochromatic sites [34, 42, 43]. Both HP1 localization and repression by Rb also require, at least in part, histone deacetylases. SUV39H1 can physically interact with HDAC1, HDAC2, and HDAC3, suggesting that transcriptional repression by SUV39H1 could be the consequence of histone deacetylase recruitment [34]. The N-terminal transcriptional repression domain of SUV39H1 binds the so-called "core histone deacetylase complex," composed of HDAC1, HDAC2, and the Rb-associated proteins [34]. SUV39H1 interacts physically with Rb and functions as a corepressor of activity. E2F is a family of transcription factors indicated in cell cycle control. E2F-containing promoters are repressed by the members of the Rb family, which are recruited by a physical interaction with the E2F protein. It is possible that transcriptional repression by Rb and its cousins involves the recruitment of HDACs to E2F-containing promoters [20, 44, 45]. Inhibition of histone deacetylase activity by TSA inhibits Rb-mediated repression, and this finding involves only a determined subset of genes, demonstrating that the requirement of HDACs for Rb-mediated transcriptional repression is promoter specific [45, 46].

The order and the timing of these modifications and their impact on transcription inactivation are less well understood. In a transgenic model, it was recently shown that histone H3 and histone H4 hypoacetylation and loss of methylation at histone H3 lysine 4 all occurred during the same window of time and that these histone modifications were the primary events in gene silencing [47]. New investigations are required to better understand the correct action of each regulating factor, its interplay, and the temporal sequence of its binding to DNA.

Epigenetic lesions may affect all the molecular pathways involved in malignant transformation, and they are often an early event in tumorigenesis. Aberrant methylation of certain genes reflects their very selected involvement in specific tumor types; therefore, it is possible to propose a specific "methylotype" for each kind of tumor. In breast cancer, we can observe promoter hypermethylation and silencing in different kinds of genes. Some of the most important are cell cycle inhibitor genes (p16ink4a); DNA repair genes (BRCA1); genes that encode BRCA1-binding protein (SRBC), tyrosine kinase (SYK), metabolic enzymes (GSTP1), and estrogen and progesterone receptors (ER, PR); genes involved in cell adherence and the metastatic process (CDH1-E cadherin, CDH13-H cadherin) and in histone/protein methyltransferase (RIZ1); and proapoptotic genes (TMS1). The specific "methylotype" of a tumor could be a marker for prognosis and response to therapy in clinical practice. The profile of genes that have been hypermethylated and then inactivated may provide information about the biological behavior of a particular disease and may enable clinicians to predict the response to chemotherapy or hormonal therapy in a patient with that specific malignancy [48, 49].


   REGULATION OF ER-PROMOTER TRANSCRIPTION AND BREAST CANCER THERAPY
Top
 Learning Objectives
 Abstract
 Introduction
 Epigenetic Information and...
 Regulation of ER-Promoter...
Conclusion
 Disclosure of Potential...
 References
 
ER-{alpha} promoter transcription is also involved in the complex interplay of transcriptional regulating factors. Up to one third of breast cancers lack ER-{alpha} at the time of diagnosis, and a fraction of breast cancers that are initially ER-{alpha} positive lose ER during tumor progression [50]. In a significant fraction of breast cancers, the loss of ER protein expression is a result of the hypermethylation of the CpG islands within the ER-{alpha} promoter [50, 51]. The loss of ER expression causes tumor growth that is no longer under estrogen control, which leads to greater cancer aggressiveness and the failure of endocrine therapy.

The role of DNMT and HDAC activity in the repression of ER-{alpha} transcription has been investigated in a variety of studies in which the re-expression of ER was a result of the use of DNMT and HDAC inhibitors. Treatment of ER-negative human breast cancer cells with the methyltransferase inhibitor 5-aza-2'-deoxycytidine (5-aza-dC) led to partial demethylation of the ER CpG island, re-expression of ER mRNA, and synthesis of functional ER protein [52, 53]. Inhibition of DNMT1 by antisense oligonucleotides also caused ER gene re-expression and the restoration of estrogen responsiveness in ER-negative breast cancer cells [54]. These results have been confirmed by recent findings showing that selective depletion of DNMT1 by either antisense oligonucleotides or siRNA induces global and gene-specific demethylation and re-expression of tumor suppressor genes in human cancer cells and also enhances 5-aza-dC activity in the re-expression of silenced genes [55]. However, some other evidence shows that inhibiting the activity of DNMT1 alone is not sufficient to obtain demethylation and re-expression of a hypermethylated gene. Recent findings in colorectal cancer cell lines demonstrate that DNMT1 cooperates with DNMT3b to silence genes and maintain DNA methylation. The lack of single DNMT1 and DNMT3b genes (single DNMT knockout) in colorectal cancer cell lines has only a small effect in reducing global DNA methylation, while the disruption of both DNMT1 and DNMT3b induces a decrease in DNA methylation by greater than 95% and the reactivation of the genes that were transcriptionally silenced by CpG island hypermethylation [56, 57].

HDAC1 interacts with ER-{alpha} in vitro and in vivo and suppresses ER-{alpha} transcriptional activity. The interaction of HDAC1 with ER-{alpha} is mediated by the activation function-2 (AF-2) domain and DNA-binding domain of ER-{alpha}. An endogenous interaction of HDAC1 with ER-{alpha} in breast cancer cells has been observed, and this interaction is decreased in the presence of estrogen. Furthermore, treatment of ER-negative breast cancer cells with the HDAC-specific inhibitor TSA induced re-expression of ER-{alpha} mRNA and protein. These findings strongly suggest that HDAC1 affects breast cancer progression by promoting cellular proliferation in association with a reduction in both ER-{alpha} protein expression and transcriptional activity. Thus, HDAC1 may be a potential target for therapeutic intervention in the treatment of a subset of ER-negative breast cancers [53, 58].

Cotreatment with DNMT and HDAC inhibitors, such as TSA, can synergistically induce ER gene expression in ER-negative breast cancer cells. Induced expression of ER mRNA and protein is associated with expected ER function on estrogen-responsive targets. Synergistic induction of ER occurs in conjunction with reduced soluble DNMT1 expression and DNMT activity, partial demethylation of the ER CpG island, and increased acetylation of histones H3 and H4. These data suggested that pharmacological intervention against both DNMT and HDAC can synergistically reactivate the methylated ER gene expression and restore ER function [9].

A novel HDAC inhibitor, scriptaid, has been shown to inhibit tumor growth in vivo and to cause re-expression of functional ER in vivo in conjunction with 5-aza-dC. Scriptaid treatment of three ER-negative cell lines (MDA-MB-231, MDA-MB 435, Hs578t) resulted in significant growth inhibition and increased acetylation of H3 and H4 histone tails. As seen with TSA and 5-aza-dC, scriptaid and 5-aza-dC cotreatment was more effective in inducing ER expression than either scriptaid or 5-aza-dC alone [59].

Several studies in the last year have shown that ER transcription is regulated by a complex mechanism of interplay among a variety of factors that are not yet well understood. Recent results show that p53 is involved in this regulation pathway. p53 upregulates ER-{alpha} gene expression by increasing transcription of the gene through elements located upstream of ER-{alpha} promoter. The p53-induced increase in ER-{alpha} gene transcription is not dependent on the ability of p53 to bind to DNA but on its ability to interact with other proteins.

The ability of p53 to control ER-{alpha} expression suggests that specific p53 mutations in breast tumors may contribute not only to oncogenesis and drug resistance but also to the more aggressive phenotype associated with the loss of ER expression. Interestingly, a high percentage of breast tumors with p53 mutations are ER-negative. These studies suggest that p53 not only regulates the ER-{alpha} promoter but may play a role in determining ER status. It may be postulated that p53 plays a role in the progression of breast cancer from a hormone-dependent phenotype to a more aggressive phenotype that is unresponsive to hormones [60]. Furthermore, it has been found that the treatment of MCF-7 cells with paclitaxel (Taxol®; Bristol-Myers Squibb, Princeton, NJ, http://www.bms.com) results in the induction of ER{alpha} gene transcription, which may be mediated through the induction of p53 [61].

pRb2/p130 has been shown to have an important role in the transcriptional regulation of the ER-{alpha} promoter as well. We recently showed a new molecular mechanism of ER-{alpha} gene inactivation mediated by pRb/p130 in ER-negative breast cancer cells. We investigated, in vivo, the occupancy of the ER-{alpha} promoter by pRb2/p130-E2F4/5-HDAC1-SUV39H1-p300 and pRb2/p130-E2F4/5-HDAC1-SUV39H1-DNMT1 complexes and provided a link between pRb2/p130 and chromatin-modifying enzymes in the regulation of ER-{alpha} transcription in a physiological setting. These findings suggest that pRb2/p130 can be key elements in the regulation of ER-{alpha} gene expression and may be viewed as promising targets for the development of novel therapeutic strategies in the treatment of breast cancer, especially for those tumors that are ER-negative [62]. The possibility of re-establishing ER expression in ER-negative breast cancers opens new therapeutic perspectives.

In a recent study, it was found that TSA sensitizes ER-{alpha}–negative, antihormone-unresponsive breast cancer cells to tamoxifen treatment, possibly by upregulating ER-ß activity. We can, therefore, explore a new strategy for inducing apoptosis, differentiation, and/or cell growth arrest in human breast and other cancer cell lines [19].

Inhibition of HDAC activities in cancer cells leads to cell cycle arrest and the induction of apoptosis. To date, only a few inhibitors of HDAC are known. TSA, a natural product, and other synthetic compounds, such as suberoyl-anilide hydroxamic acid (SAHA) and analogues, have been reported as antitumor agents that inhibit proliferation of tumor cells by inducing terminal differentiation of tumor cells [63–66]. TSA and SAHA also cause cytoplasmic {alpha}-tubulin acetylation, and this finding could indicate new roles for HDAC inhibitors in other cellular processes, in addition to transcriptional repression [67].

SAHA induces differentiation in the ER-negative cell line SKBr-3 and in the Rb-negative cell line MDA-468. SAHA exhibits a profound antiproliferative activity by causing these cells to undergo cell cycle arrest and differentiation that is dependent on the presence of SAHA [68]. The mechanism of SAHA also leads to induction of pRb-2/p130 interaction and nuclear translocation with E2F-4, causing a significant repression of E2F-1 and PCNA (Proliferating cell nuclear anfigem) nuclear levels, which leads to inhibition of DNA synthesis in mammary epithelial cell lines [69]. HDAC inhibitors such as SAHA may provide an alternative therapeutic approach for the treatment of breast cancer.

TSA and SAHA also promote apoptosis in different drug-resistant cells. Both HDAC inhibitors promote endogenous downregulation of P-glycoprotein, which is overexpressed in drug-resistant cells. This result suggests a therapeutic potential for these drugs in the treatment of cancer with acquired resistance [70].

SAHA also significantly enhances the DNA damage induced by topoisomerase-II inhibitors; however, synergy is dependent on the sequence of drug administration and the expression of the target. This finding may have an impact on the clinical development of combining HDAC inhibitors with DNA-damaging agents. [71].


   CONCLUSION
Top
 Learning Objectives
 Abstract
 Introduction
 Epigenetic Information and...
 Regulation of ER-Promoter...
 Conclusion
Disclosure of Potential...
 References
 
A better understanding of the epigenetic mechanisms that cause transcriptional repression has allowed researchers to find new agents that are very effective in inducing apoptosis, differentiation, and/or cell growth arrest in human lung cancer, breast cancer, thoracic cancer, leukemia, and colon cancer cell lines [19]. The combination of demethylating agents and HDAC inhibitors has been demonstrated to be synergic in the re-expression of ER-{alpha} in ER-{alpha}–negative breast cancer cells, and their combination with commonly used hormonal drugs or other chemotherapeutic agents opens new possibilities for a molecularly targeted approach to treatment [19].


   DISCLOSURE OF POTENTIAL CONFLICTS OF INTEREST
Top
 Learning Objectives
 Abstract
 Introduction
 Epigenetic Information and...
 Regulation of ER-Promoter...
 Conclusion
 Disclosure of Potential...
References
 
The authors indicate no potential conflicts of interest.


   ACKNOWLEDGMENT
Top
 Learning Objectives
 Abstract
 Introduction
 Epigenetic Information and...
 Regulation of ER-Promoter...
 Conclusion
 Disclosure of Potential...
 References
 
We thank Ms. M. Basso for her assistance in editing the manuscript. This work was supported by Sbarro Health Research Organization, NIH, and AIRC grants (AG) and by the WW Smith Charitable Trust (PPC). LG is supported by a fellowship from the University of Rome "La Sapienza," Italy.


   REFERENCES
Top
 Learning Objectives
 Abstract
 Introduction
 Epigenetic Information and...
 Regulation of ER-Promoter...
 Conclusion
 Disclosure of Potential...
 References
 

  1. Shao W, Brown M. Advances in estrogen receptor biology: prospects for improvements in targeted breast cancer therapy. Breast Cancer Res 2004;6:39–52.[CrossRef][Medline]
  2. Lapidus RG, Nass SJ, Davidson NE. The loss of estrogen and progesterone receptor gene expression in human breast cancer. J Mammary Gland Biol Neoplasia 1998;3:85–94.[CrossRef][Medline]
  3. McDonnell DP, Norris JD. Connections and regulation of the human estrogen receptor. Science 2002;296:1642–1644.[Abstract/Free Full Text]
  4. Smith CL. Cross-talk between peptide growth factor and estrogen receptor signaling pathways. Biol Reprod 1998;58:627–632.[Abstract/Free Full Text]
  5. Driggers PH, Segars JH. Estrogen action and cytoplasmic signaling pathways. Part II: the role of growth factors and phosphorylation in estrogen signaling. Trends Endocrinol Metab 2002;13:422–427.[CrossRef][Medline]
  6. Hanstein B, Djahansouzi S, Dall P et al. Insights into the molecular biology of the estrogen receptor define novel therapeutic targets for breast cancer. Eur J Endocrinol 2004;150:243–255.[Abstract]
  7. Shaaban AM, O’Neill PA, Davies MP et al. Declining estrogen receptor-beta expression defines malignant progression of human breast neoplasia. Am J Surg Pathol 2003;27:1502–1512.[Medline]
  8. Iwase H. Molecular action of the estrogen receptor and hormone dependency in breast cancer. Breast Cancer 2003;10:89–96.[Medline]
  9. Yang X, Phillips DL, Ferguson AT et al. Synergistic activation of functional estrogen receptor (ER)-alpha by DNA methyltransferase and histone deacetylase inhibition in human ER-alpha-negative breast cancer cells. Cancer Res 2001;61:7025–7029.[Abstract/Free Full Text]
  10. Lapidus RG, Nass SJ, Butash KA et al. Mapping of ER gene CpG island methylation-specific polymerase chain reaction. Cancer Res 1998;58:2515–2519.[Abstract/Free Full Text]
  11. Bird AP. CpG-rich islands and the function of DNA methylation. Nature 1986;321:209–213.[CrossRef][Medline]
  12. Ferguson AT, Vertino PM, Spitzner JR et al. Role of estrogen receptor gene demethylation and DNA methyltransferase. DNA adduct formation in 5-aza-2'deoxycytidine-induced cytotoxicity in human breast cancer cells. J Biol Chem 1997;272:32260–32266.[Abstract/Free Full Text]
  13. Robertson KD. DNA methylation, methyltransferases, and cancer. Oncogene 2001;20:3139–3155.[CrossRef][Medline]
  14. Belinsky SA, Nikula KJ, Palmisano WA et al. Aberrant methylation of p16(INK4a) is an early event in lung cancer and a potential biomarker for early diagnosis. Proc Natl Acad Sci U S A 1998;95:11891–11896.[Abstract/Free Full Text]
  15. Nuovo GJ, Plaia TW, Belinsky SA et al. In situ detection of the hypermethylation-induced inactivation of the p16 gene as an early event in oncogenesis. Proc Natl Acad Sci U S A 1999;96:12754–12759.[Abstract/Free Full Text]
  16. Costello JF, Fruhwald MC, Smiraglia DJ et al. Aberrant CpG-island methylation has non-random and tumour-type-specific patterns. Nat Genet 2000;24:132–138.[CrossRef][Medline]
  17. Ling Y, Sankpal UT, Robertson AK et al. Modification of de novo DNA methyltransferase 3a (Dnmt3a) by SUMO-1 modulates its interaction with histone deacetylases (HDACs) and its capacity to repress transcription. Nucleic Acids Res 2004;32:598–610.[Abstract/Free Full Text]
  18. Rountree MR, Bachman KE, Baylin SB. DNMT1 binds HDAC2 and a new co-repressor, DMAP1, to form a complex at replication foci. Nat Genet 2000;25:269–277.[CrossRef][Medline]
  19. Zhu WG, Otterson GA. The interaction of histone deacetylase inhibitors and DNA methyltransferase inhibitors in the treatment of human cancer cells. Curr Med Chem Anti-Canc Agents 2003;3:187–199.[CrossRef]
  20. Robertson KD, Ait-Si-Ali S, Yokochi T et al. DNMT1 forms a complex with Rb, E2F1 and HDAC1 and represses transcription from E2F-responsive promoters. Nat Genet 2000;25:338–342.[CrossRef][Medline]
  21. Dobosy JR, Selker EU. Emerging connections between DNA methylation and histone acetylation. Cell Mol Life Sci 2001;58:721–727.[CrossRef][Medline]
  22. Fuks F, Burgers WA, Godin N et al. Dnmt3a binds deacetylases and is recruited by a sequence-specific repressor to silence transcription. EMBO J 2001;20:2536–2544.[CrossRef][Medline]
  23. Ballestar E, Paz MF, Valle L et al. Methyl-CpG binding proteins identify novel sites of epigenetic inactivation in human cancer. EMBO J 2003;22:6335–6345.[CrossRef][Medline]
  24. Bird A. DNA methylation patterns and epigenetic memory. Genes Dev 2002;16:6–21.[Free Full Text]
  25. Nan X, Ng HH, Johnson CA et al. Transcriptional repression by the methyl-CpG-binding protein MeCP2 involves a histone deacetylase complex. Nature 1998;393:386–389.[CrossRef][Medline]
  26. Meehan RR, Lewis JD, McKay S et al. Identification of a mammalian protein that binds specifically to DNA containing methylated CpGs. Cell 1989;58:499–507.[CrossRef][Medline]
  27. Jones PL, Veenstra GJ, Wade PA etal. Methylated DNA and Me CP2 recruit histone deacetylase to repress transcription. Nat Genet 1998;19:187–191.[CrossRef][Medline]
  28. Fujita N, Watanabe S, Ichimura T et al. Methyl-CpG binding domain 1 (MBD1) interacts with the Suv39h1-HP1 heterochromatic complex for DNA methylation-based transcriptional repression. J Biol Chem 2003;278:24132–24138.[Abstract/Free Full Text]
  29. El-Osta A, Wolffe AP. DNA methylation and histone deacetylation in the control of gene expression: basic biochemistry to human development and disease. Gene Expr 2000;9:63–75.[Medline]
  30. Nan X, Campoy FJ, Bird A. MeCP2 is a transcriptional repressor with abundant binding sites in genomic chromatin. Cell 1997;88:471–481.[CrossRef][Medline]
  31. Lewis JD, Meehan RR, Henzel WJ et al. Purification, sequence, and cellular localization of a novel chromosomal protein that binds to methylated DNA. Cell 1992;69:905–914.[CrossRef][Medline]
  32. Fuks F, Hurd PJ, Wolf D et al. The methyl-CpG-binding protein MeCP2 links DNA methylation to histone methylation. J Biol Chem 2003;278:4035–4040.[Abstract/Free Full Text]
  33. Kouzarides T. Histone methylation in transcriptional control. Curr Opin Genet Dev 2002;12:198–209.[CrossRef][Medline]
  34. Vaute O, Nicolas E, Vandel L et al. Functional and physical interaction between the histone methyl transferase Suv39H1 and histone deacetylases. Nucleic Acids Res 2002;30:475–481.[Abstract/Free Full Text]
  35. Rea S, Eisenhaber F, O’Carroll D et al. Regulation of chromatin structure by site-specific histone H3 methyltransferases. Nature 2000;406: 593–599.[CrossRef][Medline]
  36. Tamaru H, Selker EU. A histone H3 methyltransferase controls DNA methylation in Neurospora crassa. Nature 2001;414:277–283.[CrossRef][Medline]
  37. Jackson JP, Lindroth AM, Cao X et al. Control of CpNpG DNA methylation by the KRYPTONITE histone H3 methyltransferase. Nature 2002;416:556–560.[CrossRef][Medline]
  38. Espada J, Ballestar E, Fraga MF et al. Human DNA methyltransferase 1 is required for maintenance of the histone H3 modification pattern. J Biol Chem 2004;279:37175–37184.[Abstract/Free Full Text]
  39. Fuks F, Hurd PJ, Deplus R et al. The DNA methyltransferases associate with HP1 and the SUV39H1 histone methyltransferase. Nucleic Acids Res 2003;31:2305–2312.[Abstract/Free Full Text]
  40. Nielsen SJ, Schneider R, Bauer UM et al. Rb targets histone H3 methylation and HP1 to promoters. Nature 2001;412:561–565.[CrossRef][Medline]
  41. Vandel L, Nicolas E, Vaute O et al. Transcriptional repression by the retinoblastoma protein through the recruitment of a histone methyltransferase. Mol Cell Biol 2001;21:6484–6494.[Abstract/Free Full Text]
  42. Lachner M, O’Carroll D, Rea S et al. Methylation of histone H3 lysine 9 creates a binding site for HP1 proteins. Nature 2001;410:116–120.[CrossRef][Medline]
  43. Bannister AJ, Zegerman P, Partridge JF et al. Selective recognition of methylated lysine 9 on histone H3 by the HP1 chromo domain. Nature 2001;410:120–124.[CrossRef][Medline]
  44. Ferreira R, Magnaghi-Jaulin L, Robin P et al. The three members of the pocket proteins family share the ability to repress E2F activity through recruitment of a histone deacetylase. Proc Natl Acad Sci U S A 1998;95:10493–10498.[Abstract/Free Full Text]
  45. Brehm A, Miska EA, McCance DJ et al. Retinoblastoma protein recruits histone deacetylase to repress transcription. Nature 1998;391:597–601.[CrossRef][Medline]
  46. Siddiqui H, Solomon DA, Gunawardena RW et al. Histone deacetylation of RB-responsive promoters: requisite for specific gene repression but dispensable for cell cycle inhibition. Mol Cell Biol 2003;23:7719–7731.[Abstract/Free Full Text]
  47. Mutskov V, Felsenfeld G. Silencing of transgene transcription precedes methylation of promoter DNA and histone H3 lysine 9. EMBO J 2004;23:138–149.[CrossRef][Medline]
  48. Esteller M, Corn PG, Baylin SB et al. A gene hypermethylation profile of human cancer. Cancer Res 2001;61:3225–3229.[Abstract/Free Full Text]
  49. Esteller M. CpG island hypermethylation and tumor suppressor genes: a booming present, a brighter future. Oncogene 2002;21:5427–5440.[CrossRef][Medline]
  50. Hortobagyi GN. Treatment of breast cancer. N Engl J Med 1998;339: 974–984.[Free Full Text]
  51. Weigel RJ, deConinck EC. Transcriptional control of estrogen receptor in estrogen receptor-negative breast carcinoma. Cancer Res 1993;53:3472–3474.[Abstract/Free Full Text]
  52. Ferguson AT, Lapidus RG, Davidson NE. Demethylation of the progesterone receptor CpG island is not required for progesterone receptor gene expression. Oncogene 1998;17:577–583.[CrossRef][Medline]
  53. Yang X, Ferguson AT, Nass SJ et al. Transcriptional activation of estrogen receptor alpha in human breast cancer cells by histone deacetylase inhibition. Cancer Res 2000;60:6890–6894.[Abstract/Free Full Text]
  54. Yan L, Nass SJ, Smith D et al. Specific inhibition of DNMT1 by anti-sense oligonucleotides induces re-expression of estrogen receptor-alpha (ER) in ER-negative human breast cancer cell lines. Cancer Biol Ther 2003;2:552–556.[Medline]
  55. Robert MF, Morin S, Beaulieu N et al. DNMT1 is required to maintain CpG methylation and aberrant gene silencing in human cancer cells. Nat Genet 2003;33:61–65.[CrossRef][Medline]
  56. Paz MF, Wei S, Cigudosa JC et al. Genetic unmasking of epigenetically silenced tumor suppressor genes in colon cancer cells deficient in DNA methyltransferases. Hum Mol Genet 2003;12:2209–2219.[Abstract/Free Full Text]
  57. Rhee I, Bachman KE, Park BH et al. DNMT1 and DNMT3b cooperate to silence genes in human cancer cells. Nature 2002;416:552–556.[CrossRef][Medline]
  58. Kawai H, Li H, Avraham S et al. Overexpression of histone deacetylase HDAC1 modulates breast cancer progression by negative regulation of estrogen receptor alpha. Int J Cancer 2003;107:353–358.[CrossRef][Medline]
  59. Keen JC, Yan L, Mack KM et al. A novel histone deacetylase inhibitor, scriptaid, enhances expression of functional estrogen receptor alpha (ER) in ER negative human breast cancer cells in combination with 5-aza 2'-deoxycytidine. Breast Cancer Res Treat 2003;81:177–186.[CrossRef][Medline]
  60. Angeloni SV, Martin MB, Garcia-Morales P et al. Regulation of estrogen receptor-alpha expression by the tumor suppressor gene p53 in MCF-7 cells. J Endocrinol 2004;180:497–504.[Abstract]
  61. Martin MB, Angeloni SV, Garcia-Morales P et al. Regulation of estrogen receptor-alpha expression in MCF-7 cells by taxol. J Endocrinol 2004;180:487–496.[Abstract]
  62. Macaluso M, Cinti C, Russo G et al. pRb2/p130-E2F4/5-HDAC1-SUV39H1-p300 and pRb2/p130-E2F4/5-HDAC1-SUV39H1-DNMT1 multimolecular complexes mediate the transcription of estrogen receptor-alpha in breast cancer. Oncogene 2003;22:3511–3517.[CrossRef][Medline]
  63. Marks PA. The mechanism of the anti-tumor activity of the histone deacetylase inhibitor, suberoylanilide hydroxamic acid (SAHA). Cell Cycle 2004;3:534–535.[Medline]
  64. Bouchain G, Leit S, Frechette S et al. Development of potential antitumor agents. Synthesis and biological evaluation of a new set of sulfonamide derivatives as histone deacetylase inhibitors. J Med Chem 2003;46: 820–830.[Medline]
  65. Bouchain G, Delorme D. Novel hydroxamate and anilide derivatives as potent histone deacetylase inhibitors: synthesis and antiproliferative evaluation. Curr Med Chem 2003;10:2359–2372.[Medline]
  66. Marks PA, Richon VM, Kelly WK et al. Histone deacetylase inhibitors: development as cancer therapy. Novartis Found Symp 2004;259:269–281, discussion 281–288.[Medline]
  67. Koeller KM, Haggarty SJ, Perkins BD et al. Chemical genetic modifier screens: small molecule trichostatin suppressors as probes of intracellular histone and tubulin acetylation. Chem Biol 2003;10:397–410.[CrossRef][Medline]
  68. Munster PN, Troso-Sandoval T, Rosen N et al. The histone deacetylase inhibitor suberoylanilide hydroxamic acid induces differentiation of human breast cancer cells. Cancer Res 2001;61:8492–8497.[Abstract/Free Full Text]
  69. Said TK, Moraes RC, Sinha R et al. Mechanisms of suberoylanilide hydroxamic acid inhibition of mammary cell growth. Breast Cancer Res 2001;3:122–133.[Medline]
  70. Castro-Galache MD, Ferragut JA, Barbera VM et al. Susceptibility of multidrug resistance tumor cells to apoptosis induction by histone deacetylase inhibitors. Int J Cancer 2003;104:579–586.[CrossRef][Medline]
  71. Marchion DC, Bicaku E, Daud AI et al. Sequence-specific potentiation of topoisomerase II inhibitors by the histone deacetylase inhibitor suberoylanilide hydroxamic acid. J Cell Biochem 2004;92:223–237.[CrossRef][Medline]



This article has been cited by other articles:


Home page
 J. Biol. Chem.Home page
J.-J. Zhao, J. Lin, H. Yang, W. Kong, L. He, X. Ma, D. Coppola, and J. Q. Cheng
MicroRNA-221/222 Negatively Regulates Estrogen Receptor{alpha} and Is Associated with Tamoxifen Resistance in Breast Cancer
J. Biol. Chem., November 7, 2008; 283(45): 31079 - 31086.
[Abstract] [Full Text] [PDF]

Home page
 Mol Hum ReprodHome page
H. Asada, Y. Yamagata, T. Taketani, A. Matsuoka, H. Tamura, N. Hattori, J. Ohgane, N. Hattori, K. Shiota, and N. Sugino
Potential link between estrogen receptor-{alpha} gene hypomethylation and uterine fibroid formation
Mol. Hum. Reprod., September 1, 2008; 14(9): 539 - 545.
[Abstract] [Full Text] [PDF]

Home page
 Mol Cancer ResHome page
Y. Han, L. Yang, F. Suarez-Saiz, S. San-Marina, J. Cui, and M. D. Minden
Wilms' Tumor 1 Suppressor Gene Mediates Antiestrogen Resistance via Down-Regulation of Estrogen Receptor-{alpha} Expression in Breast Cancer Cells
Mol. Cancer Res., August 1, 2008; 6(8): 1347 - 1355.
[Abstract] [Full Text] [PDF]

Home page
 Cancer Res.Home page
N. Kondo, T. Toyama, H. Sugiura, Y. Fujii, and H. Yamashita
miR-206 Expression Is Down-regulated in Estrogen Receptor {alpha}-Positive Human Breast Cancer
Cancer Res., July 1, 2008; 68(13): 5004 - 5008.
[Abstract] [Full Text] [PDF]

Home page
 Clin. Cancer Res.Home page
S. Lopez-Tarruella and R. Schiff
The Dynamics of Estrogen Receptor Status in Breast Cancer: Re-shaping the Paradigm
Clin. Cancer Res., December 1, 2007; 13(23): 6921 - 6925.
[Full Text] [PDF]

Home page
 Mol. Endocrinol.Home page
B. D. Adams, H. Furneaux, and B. A. White
The Micro-Ribonucleic Acid (miRNA) miR-206 Targets the Human Estrogen Receptor-{alpha} (ER{alpha}) and Represses ER{alpha} Messenger RNA and Protein Expression in Breast Cancer Cell Lines
Mol. Endocrinol., May 1, 2007; 21(5): 1132 - 1147.
[Abstract] [Full Text] [PDF]

Home page
 Molecular Cancer TherapeuticsHome page
V. F. Chekhun, N. Y. Lukyanova, O. Kovalchuk, V. P. Tryndyak, and I. P. Pogribny
Epigenetic profiling of multidrug-resistant human MCF-7 breast adenocarcinoma cells reveals novel hyper- and hypomethylated targets
Mol. Cancer Ther., March 1, 2007; 6(3): 1089 - 1098.
[Abstract] [Full Text] [PDF]

This Article
Right arrow Abstract Freely available
Right arrow Full Text (PDF)
Right arrow eLetters: Submit a response to this article
Right arrow Alert me when this article is cited
Right arrow Alert me when eLetters are posted
Right arrow Alert me if a correction is posted
Right arrow Citation Map
Services
Right arrow E-mail this article link to a friend
Right arrow Similar articles in this journal
Right arrow Similar articles in PubMed
Right arrow Alert me to new issues of the journal
Right arrow Download to citation manager
Right arrow Reprints/Permissions
Citing Articles
Right arrow Citing Articles via HighWire
Right arrow Citing Articles via Google Scholar
Google Scholar
Right arrow Articles by Giacinti, L.
Right arrow Articles by Giordano, A.
Right arrow Search for Related Content
PubMed
Right arrow PubMed Citation
Right arrow Articles by Giacinti, L.
Right arrow Articles by Giordano, A.

HOME HELP CONTACT US SUBSCRIPTIONS ARCHIVE SEARCH TABLE OF CONTENTS
THE ONCOLOGIST STEM CELLS CME ALPHAMED PRESS JOURNALS