Introduction Epigenetic alterations play an important
Epigenetic alterations play an important role in carcinogenesis. Aberrant promoter CpG island hypermethylation causes transcriptional gene silencing , and has been documented for a number of genes in prostate cancer (PCA) . Histone modifications are also important regulators of transcriptional activity; so far, they have been less comprehensively studied in PCA despite the fact that changes in DNA methylation are closely related to histone alterations and both mechanisms interact in the regulation of gene expression. Posttranslational modifications (e.g. acetylation and methylation) occur on the N-terminal tail of the histone, and function by disrupting AG-120 contacts and affecting the recruitment of various proteins to the chromatin, thereby modifying the transcriptional activity. For example, histone H3 lysine 9 methylation (H3K9me) and histone H3 lysine 27 methylation (H3K27me) are linked with transcriptional repression, whereas histone H3 lysine 4 methylation (H3K4me) is connected with transcriptional activation. In addition, histone lysine methylation exists in the form of mono (me1), di (me2) and tri (me3) methylation. Histone lysine acetylation in general leads to gene activation .
Epigenetic alterations are reversible. Numerous studies demonstrated re-expression of epigenetically silenced genes after treatment with DNA-methyltransferase (DNMT) inhibitors (e.g. decitabine) and histone deacetylase (HDAC) inhibitors (e.g. vorinostat) in cancer cell lines , , . However, side effects limit the use of these classical inhibitors.
A novel HDAC inhibitor, depsipeptide (romidepsin, FK228), was recently approved for the treatment of cutaneous T-cell-lymphoma and is relatively well tolerated . Depsipeptide induces histone acetylation by inhibiting HDAC class I . The acetylation of the histones alters the chromatin structure and the recruitment of enzymes. Depsipeptide reduces the binding of DNMT1 to the promoter region of tumor suppressor genes and the expression of histone methyltransferases . Thus, depsipeptide may also be useful for epigenetic therapy of PCA.
Epigenetic alterations in PCA are best characterized for the GSTP1 gene: The CpG island at the promoter of GSTP1 is unmethylated in the normal prostate epithelium, but methylated in PCA cells . DNA hypermethylation is associated with a repressive histone modification pattern (i.e. H3K9me2) at the GSTP1 promoter in LNCaP cells .
In this study we explore the effect of depsipeptide on the cell cycle in different prostate cell lines. We chose GSTP1 as a model gene to analyzed how depsipeptide influences DNA methylation, histone modifications and GSTP1 gene expression.
Materials and methods
Discussion HDAC inhibitors promote histone acetylation, chromatin uncoiling and transcription of variety of genes and demonstrate significant anticancer activity, including growth arrest, differentiation and apoptosis . The HDAC inhibitor depsipeptide was already approved for the treatment of cutaneous T-cell lymphoma and peripheral T-cell lymphoma, but it is poorly studied in non-haematological diseases. A small phase-II study demonstrated minimal antitumor activity in patients with metastatic PCA , although PCA cell lines are sensitive to depsipeptide . We assume that a better understanding of depsipeptide effects could help to optimize its application in the treatment of PCA. We therefore used a cell culture model to analyze the effect of depsipeptide on cell cycle, DNA methylation and histone modification patterns at the GSTP1 promoter, and the resulting effect on GSTP1 mRNA expression. The cell cycle analysis demonstrated an increase of the sub G1 fraction, whereas the G1-fraction decreased in all cell lines. Thus, depsipeptide treatment leads to apoptosis. Similarly, Piekarz et al. showed that depsipeptide induces apoptosis in T-cell lymphoma cell lines . Different modes of action were also reported: in A459 lung cancer cells, depsipeptide caused G2 and mitotic arrest . It is of interest that depsipeptide influences the expression of at least 27 proteins with multiple biological functions (e.g. signal transduction, transcriptional regulation or protein folding) . Thus, diverse mechanism may be involved in depsipeptide induced apoptosis.