A Small-Molecule Probe of the Histone Methyltransferase G9a Induces Cellular Senescence in Pancreatic Adenocarcinoma

Chemical Biology Program, Broad Institute, Cambridge, Massachusetts 02142, United States.
ACS Chemical Biology (Impact Factor: 5.33). 04/2012; 7(7):1152-7. DOI: 10.1021/cb300139y
Source: PubMed


Post-translational modifications of histones alter chromatin structure and play key roles in gene expression and specification of cell states. Small molecules that target chromatin-modifying enzymes selectively are useful as probes and have promise as therapeutics, although very few are currently available. G9a (also named euchromatin histone methyltransferase 2 (EHMT2)) catalyzes methylation of lysine 9 on histone H3 (H3K9), a modification linked to aberrant silencing of tumor-suppressor genes, among others. Here, we report the discovery of a novel histone methyltransferase inhibitor, BRD4770. This compound reduced cellular levels of di- and trimethylated H3K9 without inducing apoptosis, induced senescence, and inhibited both anchorage-dependent and -independent proliferation in the pancreatic cancer cell line PANC-1. ATM-pathway activation, caused by either genetic or small-molecule inhibition of G9a, may mediate BRD4770-induced cell senescence. BRD4770 may be a useful tool to study G9a and its role in senescence and cancer cell biology.

Download full-text


Available from: Bridget K Wagner,
  • Source
    • "Knockdown of G9a by RNAi mediated gene silencing inhibits growth and further promotes apoptosis of breast cancer cells [8,12]. Pharmacological inhibition of G9a disturbs the cell cycle and induces senescent phenotypes, further inhibiting growth in prostate and pancreatic cancer [13,14]. Furthermore, inhibition of G9a may lead to autophagy in breast and colorectal cancers [15], suggesting that G9a may control cell growth in a variety of cancer types via multiple routes. "
    [Show abstract] [Hide abstract]
    ABSTRACT: Head and neck squamous cell carcinoma (HNSCC) is a common cancer worldwide. Emerging evidence indicates that alteration of epigenetics might be a key event in HNSCC progression. Abnormal expression of histone methyltransferase G9a, which contributes to transcriptional repression of tumor suppressors, has been implicated in promoting cancerous malignancies. However, its role in HNSCC has not been previously characterized. In this study, we elucidate the function of G9a and its downstream mechanism in HNSCC. We investigated the clinical relevance of G9a in HNSCC using immunohistochemistry (IHC) staining. In vitro cell proliferation and tumorigenesis ability of G9a-manipulated HNSCC cells were examined with MTT assays, clonogenic assays, and soft agar assays. We examined different routes of cell death in HNSCC cells induced by G9a-depletion or enzymatic inhibition by immunoblot, flow cytometry, fluorescent and transmission electron microscopy analysis. Specific targets of G9a were identified by affymetrix microarray and quantitative reverse transcription-polymerase chain reaction (qRT-PCR). Lastly, functions of G9a in vivo were confirmed with a xenograft tumor model. G9a expression is positively correlated to proliferation marker Ki-67 and to poor prognosis in HNSCC patients. Genetic or pharmacological inhibition of G9a reduced cell proliferation without inducing necrosis or apoptosis. Instead, autophagic cell death was the major consequence, and our investigation of mechanisms suggested it is mediated via the dual specificity phosphatase-4 (DUSP4) dependent ERK inactivation pathway. An orthotopic tumor model further confirmed the growth inhibiting effect and induction of autophagy that followed suppression of G9a. In this study, we provide evidence that G9a confers the survival advantage of HNSCC. Genetic or pharmacological inhibition of G9a induces autophagic cell death; this finding provides a basis for new therapeutic targets for treating HNSCC.
    Molecular Cancer 07/2014; 13(1):172. DOI:10.1186/1476-4598-13-172 · 4.26 Impact Factor
  • Source
    • "Small molecule inhibitors targeting these modifiers and readers might effectively block EMT and cancer metastasis. Such inhibitors include: BRD4770 (Yuan et al. 2012), Entinostat (Kummar et al. 2007), Romidepsin (Saijo et al. 2012), TCP derivatives (Harris et al. 2012; Schenk et al. 2012), GSK- J1 (Kruidenier et al. 2012), GSK- 126 (McCabe et al. 2012), MM- 102 (Senisterra et al. 2013) and EPZ004777 (Daigle et al. 2011) Cell Tissue Res (2014) 356:617–630 625 effects. For example, one recent study demonstrated that the treatment of prostate cancer cells with the HDACi suberoylanilide hydroxamic acid (SAHA) and TSA results in the induction of EMT and increased expression of the EMT-TFs ZEB1, ZEB2 and SLUG and of the mesenchymal makers vimentin, N-cadherin and fibronectin; this was accompanied by tumorigenicity and invasiveness (Kong et al. 2012). "
    [Show abstract] [Hide abstract]
    ABSTRACT: The epithelial-to-mesenchymal transition (EMT) is a reversible change in cell phenotype that plays a crucial role during normal development and cancer metastasis. EMT imparts embryonic epithelial cells with the ability to migrate and to give rise to organs or tissues at distant sites. During cancer progression, the same developmental process is utilized in an analogous manner to enable cancer cells to move to distant organs and form metastases. The reversion of EMT via the mesenchymal-to-epithelial transition (MET) appears to be required for the formation of secondary tumors at distal sites. The plasticity of epigenomic modifications that control the transcriptional program of cells enables cells to switch back and forth from epithelial and mesenchymal phenotypes during these transitions. Here, we review the interplay between complex epigenomic regulatory mechanisms and various transcription factors involved in EMT leading to changes in gene expression and cell phenotype. We also discuss the way that a deeper understanding of the epigenomic regulation of EMT might shed light onto the process of cancer progression and reveal new targets for novel and more specific anticancer epigenomic therapies.
    Cell and Tissue Research 05/2014; 356(3). DOI:10.1007/s00441-014-1912-y · 3.57 Impact Factor
  • Source
    • "Furthermore, LSD1/KDM1A inhibition suppresses acute myeloid leukemia (AML) stem cell activity (Harris et al. 2012; Schenk et al. 2012), and disruption of the chromatin binding of Brd4 by a BET bromodomain inhibitor blocks c-Myc expression and the proliferation of leukemic cells (Delmore et al. 2011; Zuber et al. 2011). To date, several inhibitors of histone methyltransferases and demethylases have been reported, including those that target G9a and GLP with high specificity (Kubicek et al. 2007; Vedadi et al. 2011; Yuan et al. 2012). G9a/GLP uniquely catalyze mono-and dimethylation of histone 3 on Lys9 (H3K9me1/2) (Tachibana et al. 2002, 2005), a highly abundant chromatin mark in mammalian cells. "
    [Show abstract] [Hide abstract]
    ABSTRACT: Chromatin modulators are emerging as attractive drug targets, given their widespread implication in human cancers and susceptibility to pharmacological inhibition. Here we establish the histone methyltransferase G9a/EHMT2 as a selective regulator of fast proliferating myeloid progenitors with no discernible function in hematopoietic stem cells (HSCs). In mouse models of acute myeloid leukemia (AML), loss of G9a significantly delays disease progression and reduces leukemia stem cell (LSC) frequency. We connect this function of G9a to its methyltransferase activity and its interaction with the leukemogenic transcription factor HoxA9 and provide evidence that primary human AML cells are sensitive to G9A inhibition. Our results highlight a clinical potential of G9A inhibition as a means to counteract the proliferation and self-renewal of AML cells by attenuating HoxA9-dependent transcription.
    Genes & development 02/2014; 28(4):317-27. DOI:10.1101/gad.236794.113 · 10.80 Impact Factor
Show more