Anti-AML Activity of Combined Epigenetic Therapy with Novel DNMT1 Inhibitors SGI-110 or SGI-1036 and Histone Deacetylase Inhibitor Panobinostat.

Blood ◽  
2008 ◽  
Vol 112 (11) ◽  
pp. 3355-3355
Author(s):  
Warren Fiskus ◽  
Pace Johnston ◽  
Rajeshree Joshi ◽  
Rekha Rao ◽  
Celalettin Ustun ◽  
...  
Keyword(s):  
Dna Methyltransferase ◽  
Cell Cycle Progression ◽  
Dna Methyltransferases ◽  
Parp Cleavage ◽  
Epigenetic Therapy ◽  
Gene Promoters ◽  
Dna Hypermethylation ◽  
Dnmt Inhibitors ◽  
Protein Levels ◽  
Core Proteins

Abstract Lysine specific histone methylation and deacetylation and DNA hypermethylation are involved in the epigenetic silencing of tumor suppressor genes (TSG), e.g., p15 and p16. DNA methyltransferase (DNMT) inhibitors 5-azacytidine and 5-aza-2’-deoxycytidine demethylate the CpG dinucleotide islands in or near gene promoters, leading to derepression of TSGs in AML. SGI-110 (S110) (Cancer Res.2007; 67:6400) and SGI-1036 (SuperGen, Inc.) are novel, DNMT inhibitors, which also deplete DNMT1 levels. SGI-110 is a dinucleotide containing 5-aza-2’-deoxycytidine and SGI-1036 is a non-nucleoside heterocycle. The multi-protein complex PRC (polycomb repressive complex) 2 that contains the three core proteins EZH2, SUZ12 and EED, has intrinsic histone methyltransferase (HMTase) activity. This is mediated by the SET domain of EZH2, which induces trimethylation of histone H3 on lysine (K)-27. We recently reported that treatment with the pan-HDAC inhibitor panobinostat (LBH589, Novartis Pharmaceutical Corp) acetylates and inhibits the ATP binding and chaperone function of hsp90, as well as depletes the levels of EZH2, Suz12 and EED in cultured and primary AML cells (Mol Cancer Ther.2006; 5:3096). Within the PRC2 complex, EZH2 was shown to interact with and modulate the DNA methyltransferases DNMT1, DNMT3a and DNMT3b, which affects their binding to the EZH2-targeted gene promoters. In the present studies we determined the effects of SGI-110 or SGI-1036 and LBH589 on the PRC2 proteins EZH2 and SUZ12, and DNMT1, in the cultured (HL-60, OCI-AML3 and K562) and primary AML cells. Treatment with SGI-110 (0.5 to 2.0 μM) or SGI-1036 (0.5 and 1.0 μM) for 24 hours depleted protein levels of DNMT1 and EZH2 in the cultured and primary AML cells. SGI-110 and SGI-1036 promoted proteasomal degradation of DNMT1 and EZH2 since co-treatment with bortezomib significantly restored DNMT1 and EZH2 levels in the AML cells. Following treatment with SGI-110 or SGI-1036, bisulfite modification and methylation specific PCR demonstrated increase in unmethylated promoter DNA of p15 and JunB. This was associated with induction of the mRNA and protein levels of p15 and JunB, as well as caused inhibition of cell cycle progression (% of cells increased in G1 and increased in S phase) and colony growth in the soft agar. Treatment with 1.0 μM of SGI-110 or SGI-1036 also induced PARP cleavage activity of caspases and induced morphologic evidence of apoptosis in the AML cells. Co-treatment with 10 to 50 nM panobinostat enhanced SGI-110 or SGI-1036 mediated depletion of DNMT1 and EZH2, with more de-repression of the p15 and JunB and significant increase in apoptosis of AML cells. Collectively, these findings indicate that, SGI-110 and SGI-1036 deplete DNMT1 and EZH2 levels, as well as exert potent anti-AML activity. Additionally, combined epigenetic therapy consisting of SGI-110 or SGI-1036 in combination with panobinostat may represent a promising novel treatment of AML.

Synergistic Pre-Clinical Activity of Combined Epigenetic Therapy with the Novel Histone Methyltransferase EZH2 Inhibitor 3-Deazaneplanocin and Histone Deacetylase Inhibitor Panobinostat against Human AML Cells.

Blood ◽  
2008 ◽  
Vol 112 (11) ◽  
pp. 3356-3356
Author(s):  
Warren Fiskus ◽  
Yongchao Wang ◽  
Anand Jillella ◽  
Celalettin Ustun ◽  
Pace Johnston ◽  
...  
Keyword(s):  
Histone Deacetylase ◽  
Hox Genes ◽  
Histone Methyltransferase ◽  
Dna Methyltransferases ◽  
Parp Cleavage ◽  
Epigenetic Therapy ◽  
Clinical Activity ◽  
Gene Promoters ◽  
Core Proteins ◽  
Induced Apoptosis

Abstract Lysine specific histone methylation and deacetylation and DNA hypermethylation are involved in the epigenetic silencing of tumor suppressor genes (TSG), e.g., p15 and p16. The multi-protein complex PRC (polycomb repressive complex) 2 that contains the three core proteins EZH2, SUZ12 and EED, has intrinsic histone methyltransferase (HMTase) activity. This is mediated by the SET domain of EZH2, which induces tri-methylation (3Me) of lysine (K)-27 on histone H3, regulates the expression of HOX genes and promotes cell proliferation and aggressiveness of neoplastic cells. In the present studies we demonstrate that treatment with the S-adenosylhomocysteine hydrolase inhibitor 3-Deazaneplanocin A (DZNep) dose-dependently (200 nM to 2.0 uM) depletes EZH2, SUZ12 and EED levels, inhibits 3Me K27 on H3 while inducing K27 H3 acetylation in the cultured human AML HL60 and OCI-AML3 cells and in primary, patient-derived AML blasts. DZNep treatment also induced the levels of p16, p21, p27 and FBXO32 while depleting cyclin E and HOXA9 levels. Treatment with DZNep induced PARP cleavage activity of the caspases and apoptosis in the cultured and primary AML cells. DZNep promoted proteasomal degradation of EZH2 and SUZ12, since co-treatment with bortezomib significantly restored EZH2 and SUZ12 levels in the AML cells. We had previously reported that treatment with the pan-histone deacetylase (HDAC) inhibitor panobinostat (PS) (LBH589, Novartis Pharmaceutical Corp) also depletes the levels of EZH2, SUZ12 and EED in cultured and primary AML cells (Mol Cancer Ther.2006; 5:3096). Within the PRC2 complex, EZH2 bound and recruited the DNA methyltransferases DNMT1, and treatment with PS also disrupted the interaction of EZH2 with DNMT1, attenuated DNMT1 levels and its binding to the EZH2-targeted gene promoters, e.g., p16 and JunB. Here, we also demonstrate that, as compared to treatment with either agent alone, co-treatment with DZNep and PS caused more depletion of EZH2, SUZ12 and EED, more induction of p16, p21 and p27, as well as synergistically induced apoptosis of AML cells (combination indices < 1.0). Additionally, DZNep induced apoptosis of HL-60/LR cells that are resistant to HDACs including PS, as well as sensitized HL-60/LR cells to PS. Taken together, these findings indicate that targeting EZH2 and the PRC2 complex is an effective epigenetic therapy of AML that also overcomes resistance to HDAC inhibitors. Additionally, combined epigenetic therapy with DZNep and PS exerts synergistic in vitro activity against human AML cells, suggesting that this combination may be a promising novel treatment for AML.

Combined Epigenetic Therapy with the Novel Histone Methyl Transferase EZH2 Inhibitor 3-Deazaneplanocin and Histone Deacetylase Inhibitor Panobinostat Exerts Synergistic Activity against Human Mantle Cell Lymphoma Cells

Blood ◽  
2008 ◽  
Vol 112 (11) ◽  
pp. 3622-3622
Author(s):  
Warren Fiskus ◽  
Yongchao Wang ◽  
Anand Jillella ◽  
Pace Johnston ◽  
Rajeshree Joshi ◽  
...  
Keyword(s):  
Cell Proliferation ◽  
Mantle Cell Lymphoma ◽  
Histone Deacetylase ◽  
Cell Lymphoma ◽  
Parp Cleavage ◽  
Epigenetic Therapy ◽  
Synergistic Activity ◽  
Gene Promoters ◽  
Core Proteins ◽  
Mantle Cell

Abstract Lysine specific histone methylation and deacetylation and DNA hypermethylation are involved in the epigenetic silencing of tumor suppressor genes (TSG), e.g., p16 and JunB. The multi-protein complex PRC (polycomb repressive complex) 2 that contains the three core proteins EZH2, SUZ12 and EED, has intrinsic histone methyltransferase (HMTase) activity. This is mediated by the SET domain of EZH2, which induces tri-methylation (3Me) of lysine (K)-27 on histone H3, as well as promotes cell proliferation and aggressiveness of neoplastic cells. EZH2 is preferentially overexpressed in proliferating but not resting Mantle Cell Lymphoma (MCL) cells. In the present studies we demonstrate that treatment with the S-adenosylhomocysteine hydrolase inhibitor 3-Deazaneplanocin A (DZNep) dose-dependently (500 nM to 2.0 uM) depletes EZH2, SUZ12 and EED levels, as well as inhibits 3Me K27 on H3 while inducing K27 H3 acetylation. DZNep treatment also induces the levels of p21, p27, JunB and FBXO32, while depleting cyclin D1 and cyclin E levels in the cultured human MCL Jeko-1, MO2058 and Z138 cells and in primary patient-derived MCL cells. Treatment with DZNep induces PARP cleavage activity of the caspases and apoptosis in the cultured and primary MCL cells. DZNep promoted proteasomal degradation of EZH2 and SUZ12, since co-treatment with bortezpmib significantly restored EZH2 and SUZ12 levels in the MCL cells. We had previously reported that treatment with the pan-histone deacetylase (HDAC) inhibitor panobinostat (PS) (LBH589, Novartis Pharmaceutical Corp) depletes the levels of EZH2, SUZ12 and EED in cultured and primary AML cells (Mol Cancer Ther.2006; 5:3096). Within the PRC2 complex, EZH2 bound and recruited the DNA methyltransferases DNMT1, and treatment with PS also disrupted the interaction of EZH2 with DNMT1, attenuated DNMT1 levels and its binding to the EZH2-targeted gene promoters, e,g, JunB. Here, we also demonstrate that, PS treatment depletes DNMT1 levels and induces JunB levels in cultured MCL cells. As compared to treatment with either agent alone, co-treatment with DZNep and PS caused more depletion of EZH2 and SUZ12, but not of DNMT1, more induction of JunB, p21 and p27, as well as synergistically induced apoptosis of cultured MCL cells (combination indices < 1.0). Taken together, these findings indicate that DZNep and PS mediated targeting of EZH2 and the PRC2 complex is an effective epigenetic therapy of MCL, which also results in undermining several molecular determinants of MCL cell proliferation and survival. Additionally, combined epigenetic therapy with DZNep and PS exerts synergistic in vitro activity against human MCL cells, suggesting that this combination may be a promising novel treatment for MCL.

scholarly journals DNA Methyltransferase 3A Is Involved in the Sustained Effects of Chronic Stress on Synaptic Functions and Behaviors

2020 ◽  
Author(s):  
Jing Wei ◽  
Jia Cheng ◽  
Nicholas J Waddell ◽  
Zi-Jun Wang ◽  
Xiaodong Pang ◽  
...  
Keyword(s):  
Dna Methylation ◽  
Dna Methyltransferase ◽  
Epigenetic Modification ◽  
Substantial Reduction ◽  
Dna Methyltransferases ◽  
Male Rats ◽  
Neural Pathways ◽  
Dnmt Inhibitors ◽  
Synaptic Functions

Abstract Emerging evidence suggests that epigenetic mechanisms regulate aberrant gene transcription in stress-associated mental disorders. However, it remains to be elucidated about the role of DNA methylation and its catalyzing enzymes, DNA methyltransferases (DNMTs), in this process. Here, we found that male rats exposed to chronic (2-week) unpredictable stress exhibited a substantial reduction of Dnmt3a after stress cessation in the prefrontal cortex (PFC), a key target region of stress. Treatment of unstressed control rats with DNMT inhibitors recapitulated the effect of chronic unpredictable stress on decreased AMPAR expression and function in PFC. In contrast, overexpression of Dnmt3a in PFC of stressed animals prevented the loss of glutamatergic responses. Moreover, the stress-induced behavioral abnormalities, including the impaired recognition memory, heightened aggression, and hyperlocomotion, were partially attenuated by Dnmt3a expression in PFC of stressed animals. Finally, we found that there were genome-wide DNA methylation changes and transcriptome alterations in PFC of stressed rats, both of which were enriched at several neural pathways, including glutamatergic synapse and microtubule-associated protein kinase signaling. These results have therefore recognized the potential role of DNA epigenetic modification in stress-induced disturbance of synaptic functions and cognitive and emotional processes.

scholarly journals Novel Quinoline Compounds Active in Cancer Cells through Coupled DNA Methyltransferase Inhibition and Degradation

Cancers ◽  
2020 ◽  
Vol 12 (2) ◽  
pp. 447
Author(s):  
Clemens Zwergel ◽  
Rossella Fioravanti ◽  
Giulia Stazi ◽  
Federica Sarno ◽  
Cecilia Battistelli ◽  
...  
Keyword(s):  
Protein Degradation ◽  
Cancer Cell ◽  
Dna Methyltransferase ◽  
Dna Methyltransferases ◽  
Biological Evaluation ◽  
Solid Cancer ◽  
Egfp Gene ◽  
Design Synthesis ◽  
Dnmt Inhibitors ◽  
Cancer Cell Survival

DNA methyltransferases (DNMTs) play a relevant role in epigenetic control of cancer cell survival and proliferation. Since only two DNMT inhibitors (azacitidine and decitabine) have been approved to date for the treatment of hematological malignancies, the development of novel potent and specific inhibitors is urgent. Here we describe the design, synthesis, and biological evaluation of a new series of compounds acting at the same time as DNMTs (mainly DNMT3A) inhibitors and degraders. Tested against leukemic and solid cancer cell lines, 2a–c and 4a–c (the last only for leukemias) displayed up to submicromolar antiproliferative activities. In HCT116 cells, such compounds induced EGFP gene expression in a promoter demethylation assay, confirming their demethylating activity in cells. In the same cell line, 2b and 4c chosen as representative samples induced DNMT1 and -3A protein degradation, suggesting for these compounds a double mechanism of DNMT3A inhibition and DNMT protein degradation.

scholarly journals Early epigenetic changes and DNA damage do not predict clinical response in an overlapping schedule of 5-azacytidine and entinostat in patients with myeloid malignancies

Blood ◽  
2009 ◽  
Vol 114 (13) ◽  
pp. 2764-2773 ◽  
Author(s):  
Tamer E. Fandy ◽  
James G. Herman ◽  
Patrick Kerns ◽  
Anchalee Jiemjit ◽  
Elizabeth A. Sugar ◽  
...  
Keyword(s):  
Bone Marrow ◽  
Dna Damage ◽  
Clinical Response ◽  
Clinical Efficacy ◽  
Dna Methyltransferase ◽  
Hdac Inhibitors ◽  
Hematologic Malignancies ◽  
Epigenetic Therapy ◽  
Dnmt Inhibitors ◽  
Sequential Administration

Abstract Sequential administration of DNA methyltransferase (DNMT) inhibitors and histone deacetylase (HDAC) inhibitors has demonstrated clinical efficacy in patients with hematologic malignancies. However, the mechanism behind their clinical efficacy remains controversial. In this study, the methylation dynamics of 4 TSGs (p15INK4B, CDH-1, DAPK-1, and SOCS-1) were studied in sequential bone marrow samples from 30 patients with myelodysplastic syndrome (MDS) or acute myeloid leukemia (AML) who completed a minimum of 4 cycles of therapy with 5-azacytidine and entinostat. Reversal of promoter methylation after therapy was observed in both clinical responders and nonresponders across all genes. There was no association between clinical response and either baseline methylation or methylation reversal in the bone marrow or purified CD34+ population, nor was there an association with change in gene expression. Transient global hypomethylation was observed in samples after treatment but was not associated with clinical response. Induction of histone H3/H4 acetylation and the DNA damage–associated variant histone γ-H2AX was observed in peripheral blood samples across all dose cohorts. In conclusion, methylation reversal of candidate TSGs during cycle 1 of therapy was not predictive of clinical response to combination “epigenetic” therapy. This trial is registered with http://www.clinicaltrials.gov under NCT00101179.

scholarly journals Site-Specific DNA Demethylation as a Potential Target for Cancer Epigenetic Therapy

2020 ◽  
Vol 13 ◽  
pp. 251686572096480
Author(s):  
Sultan Abda Neja
Keyword(s):  
Cpg Island ◽  
Causal Effect ◽  
Dna Methyltransferases ◽  
Dna Demethylation ◽  
Epigenetic Therapy ◽  
In Vivo Model ◽  
Transcription Activator ◽  
Dna Hypermethylation ◽  
Site Specific

Aberrant promoter DNA hypermethylation is a typical characteristic of cancer and it is often seen in malignancies. Recent studies showed that regulatory cis-elements found up-stream of many tumor suppressor gene promoter CpG island (CGI) attract DNA methyltransferases (DNMT) that hypermethylates and silence the genes. As epigenetic alterations are potentially reversible, they make attractive targets for therapeutic intervention. The currently used decitabine (DAC) and azacitidine (AZA) are DNMT inhibitors that follow the passive demethylation pathway. However, they lead to genome-wide demethylation of CpGs in cells, which makes difficult to use it for causal effect analysis and treatment of specific epimutations. Demethylation through specific demethylase enzymes is thus critical for epigenetic resetting of silenced genes and modified chromatins. Yet DNA-binding factors likely play a major role to guide the candidate demethylase enzymes upon its fusion. Before the advent of clustered regulatory interspaced short palindromic repeats (CRISPR), both zinc finger proteins (ZNFs) and transcription activator-like effector protein (TALEs) were used as binding platforms for ten-eleven translocation (TET) enzymes and both systems were able to induce transcription at targeted loci in an in vitro as well as in vivo model. Consequently, the development of site-specific and active demethylation molecular trackers becomes more than hypothetical to makes a big difference in the treatment of cancer in the future. This review is thus to recap the novel albeit distinct studies on the potential use of site-specific demethylation for the development of epigenetic based cancer therapy.

scholarly journals STAT3 activation in large granular lymphocyte leukemia is associated with cytokine signaling and DNA hypermethylation

Leukemia ◽  
2021 ◽  
Author(s):  
Daehong Kim ◽  
Giljun Park ◽  
Jani Huuhtanen ◽  
Bishwa Ghimire ◽  
Hanna Rajala ◽  
...  
Keyword(s):  
T Cells ◽  
Nk Cells ◽  
Cd8 T Cells ◽  
Dna Methyltransferase ◽  
Large Granular Lymphocyte ◽  
Cytokine Signaling ◽  
Dna Hypermethylation ◽  
Gain Of Function ◽  
Large Granular Lymphocyte Leukemia ◽  
Dnmt Inhibitors

AbstractLarge granular lymphocyte leukemia (LGLL) is characterized by somatic gain-of-function STAT3 mutations. However, the functional effects of STAT3 mutations on primary LGLL cells have not been studied in detail. In this study, we show that CD8+ T cells isolated from STAT3 mutated LGLL patients have high protein levels of epigenetic regulators, such as DNMT1, and are characterized by global hypermethylation. Correspondingly, treatment of healthy CD8+ T cells with IL-6, IL-15, and/or MCP-1 cytokines resulted in STAT3 activation, increased DNMT1, EZH2, c-MYC, l-MYC, MAX, and NFκB levels, increased DNA methylation, and increased oxidative stress. Similar results were discovered in KAI3 NK cells overexpressing gain-of-function STAT3Y640F and STAT3G618R mutants compared to KAI3 NK cells overexpressing STAT3WT. Our results also confirm that STAT3 forms a direct complex with DNMT1, EZH2, and HDAC1. In STAT3 mutated LGLL cells, DNA methyltransferase (DNMT) inhibitor azacitidine abrogated the activation of STAT3 via restored SHP1 expression. In conclusion, STAT3 mutations cause DNA hypermethylation resulting in sensitivity to DNMT inhibitors, which could be considered as a novel treatment option for LGLL patients with resistance to standard treatments.

scholarly journals Identification of Novel Bacterial M.SssI DNA Methyltransferase Inhibitors

2012 ◽  
Vol 18 (3) ◽  
pp. 348-355 ◽  
Author(s):  
Marlinda Hupkes ◽  
Rita Azevedo ◽  
Hans Jansen ◽  
Everardus J. van Zoelen ◽  
Koen J. Dechering
Keyword(s):  
Dna Methylation ◽  
Dna Methyltransferase ◽  
Homology Model ◽  
Binding Mode ◽  
Dna Methyltransferases ◽  
Docking Studies ◽  
Dna Methyltransferase Inhibitors ◽  
Dnmt Inhibitors ◽  
Starting Point

DNA methylation is an important epigenetic regulator of gene expression. Abnormalities in DNA methylation patterns have been associated with various developmental and proliferative diseases, particularly cancer. Targeting DNA methyltransferases (DNMTs) represents a promising strategy for the treatment of such diseases. Current DNMT inhibitors suffer important drawbacks with respect to their efficacy, specificity, and toxicity. In this study, we have set up a robust in vitro bacterial M.SssI DNMT activity assay to systematically screen a collection of 26 240 compounds that were predicted to compete with the S-adenosyl-L-methionine (SAM) substrate of DNMT. This resulted in the identification of a novel set of structurally distinct inhibitors of M.SssI DNMT activity. Although molecular docking studies using an M.SssI homology model suggest that these compounds might compete with SAM binding, mode of activity (MoA) assays are still needed to confirm this hypothesis. Our set of novel M.SssI DNMT inhibitors, once confirmed in an orthogonal DNMT assay, may thus serve as a starting point to identify and characterize suitable lead candidates for further drug optimization.

scholarly journals Kaiso regulates DNA methylation homeostasis

2020 ◽  
Author(s):  
D Kaplun ◽  
G Filonova ◽  
Y. Lobanova ◽  
A Mazur ◽  
S Zhenilo
Keyword(s):  
Dna Methylation ◽  
Complex Formation ◽  
Dna Methyltransferase ◽  
De Novo ◽  
Main Idea ◽  
Dna Methyltransferases ◽  
The Other ◽  
Dna Hypermethylation ◽  
Mouse Embryonic Fibroblasts ◽  
Embryonic Fibroblasts

ABSTRACTGain and loss of DNA methylation in cells is a dynamic process that tends to achieve an equilibrium. Many factors are involved in maintaining the balance between DNA methylation and demethylation. Previously, it was shown that methyl-DNA protein Kaiso may attract NcoR, SMRT repressive complexes affecting histone modifications. On the other hand, the deficiency of Kaiso resulted in slightly reduced methylation of ICR in H19/Igf2 locus and Oct4 promoter in mouse embryonic fibroblasts. However, nothing is known whether Kaiso may attract DNA methyltransferase to influence DNA methylation level. The main idea of this work is that Kaiso may lead to DNA hypermethylation attracting de novo DNA methyltransferases. We demonstrated that Kaiso regulates TRIM25 promoter methylation. It can form a complex with DNMT3b. BTB/POZ domain of Kaiso and ADD domain of DNA methyltransferase are essential for complex formation. Thus, Kaiso can affect DNA methylation.

DNA Methyltransferase Inhibitors and their Therapeutic Potential

2019 ◽  
Vol 18 (28) ◽  
pp. 2448-2457 ◽  
Author(s):  
Zehao Zhou ◽  
Huan-Qiu Li ◽  
Feng Liu
Keyword(s):  
Dna Methylation ◽  
Dna Methyltransferase ◽  
Therapeutic Potential ◽  
Biological Activities ◽  
Dna Methyltransferases ◽  
Dna Methyltransferase Inhibitors ◽  
Dnmt Inhibitors ◽  
Aberrant Dna Methylation ◽  
Cancer Tissues ◽  
Nucleoside Inhibitors

Aberrant DNA methylation at the 5-position of cytosine, catalyzed by DNA methyltransferases (DNMTs), is associated with not only various cancers by silencing of tumor suppressor genes but also other diseases. The DNMTs, especially the DNMT1, DNMT3A and DNMT3B, are often overexpressed in various cancer tissues and cell lines. DNMTs are important epigenetic targets for drug development since the DNA methylation is reversible. This review summarizes an array of nucleoside and non-nucleoside inhibitors of DNMTs, as well as their biological activities. Among these inhibitors, the nucleoside analogue azacytidine and its deoxy derivative decitabine are both irreversible DNMT inhibitors and approved for the treatment of myelodysplastic syndrome.

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