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.

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.

scholarly journals Azacitidine Blocks GATA-1-Mediated Repression of the PU.1 Gene in Human Leukemic Cells

Blood ◽  
2015 ◽  
Vol 126 (23) ◽  
pp. 5220-5220
Author(s):  
Pavel Burda ◽  
Jarmila Vargova ◽  
Nikola Curik ◽  
John Strouboulis ◽  
Giorgio Lucio Papadopoulos ◽  
...  
Keyword(s):  
Dna Methylation ◽  
Transcription Factors ◽  
Regulatory Element ◽  
Methylation Status ◽  
Erythroid Differentiation ◽  
Dna Demethylation ◽  
Leukemic Cells ◽  
Epigenetic Therapy ◽  
Myeloid Differentiation

Abstract Introduction: GATA-1 and PU.1 are two important hematopoietic transcription factors that mutually inhibit each other in progenitor cells to guide entrance into the erythroid or myeloid lineage, respectively. Expression of PU.1 is controlled by several transcription factors including PU.1 itself by binding to the distal URE enhancer (upstream regulatory element) whose deletion leads to acute myeloid leukemia (AML) (Rosenbauer F et al. 2004). Co-expression of PU.1 and GATA-1 in AML-erythroleukemia (EL) blasts prevents efficient differentiation regulated by these transcription factors. Inhibition of transcriptional activity of PU.1 protein by GATA-1 has been reported (Nerlov C et al. 2000), however it is not known whether GATA-1 can inhibit PU.1 gene in human early erythroblasts directly. We have recently found that MDS/AML erythroblasts display repressive histone modifications and DNA methylation status of PU.1 gene that respond to 5-azacitidine (AZA) leading to inhibited blast cell proliferation and stimulated myeloid differentiation (Curik N et al. 2012). We hypothesize that l eukemia blockade during early erythroid differentiation includes direct GATA-1-mediated inhibition of the PU.1 gene. Results: We herein document the GATA-1 mediated repression of the PU.1 gene in human EL cell lines (OCI-M2 and K562) together with the recruitment of DNA methyl transferase I (DNMT1) to the URE known to guide most of the PU.1 gene transcription. Repression of the PU.1 gene involves both DNA methylation at the URE and methylation/deacetylation of the histone H3 lysine-K9 residue and methylation of H3K27 at additional DNA elements and the PU.1 promoter. Inhibition of GATA-1 by siRNA as well as the AZA treatment in AML-EL led to the significant DNA-demethylation of the URE thorough the mechanism of DNMT1 depletion leading to upregulation of the PU.1 expression. Conclusions: Our data indicate that GATA-1 binds to the PU.1 gene at the URE and initiate events leading to the PU.1 gene repression in human ELs. The mechanism includes repressive epigenetic remodeling of the URE that is important for the PU.1 downregulation and leukemogenesis and that is also simultaneously sensitive to the DNA demethylation treatment with AZA. The GATA-1-mediated inhibition likely contributes to the PU.1 downregulation during progenitor cell differentiation that could be employed during leukemogenesis. Importantly, we also observed important differences between murine and human ELs and found that repression of the PU.1 gene in human ELs can become reverted by the epigenetic therapy with AZA. Our work also suggests that hypomethylating therapy using DNA methylation inhibitors in MDS/AML may become potentially effective in MDS/EL patients. We think that during early erythroid differentiation the GATA-1 binds and represses the PU.1 gene, however this is not fully completed in EL and therefore the erythroid as well as myeloid differentiation are blocked. Grants: GACR P305/12/1033, UNCE 204021, PRVOUK-P24/LF1/1. Disclosures Off Label Use: Azacitidine, DNA demethylation agens tested in vitro in AML/MDS treatment. Stopka:Celgene: Research Funding.

DNA Methylation Analysis of 807 Genes in Chronic Myeloid Leukemia and Acute Promyelocytic Leukemia.

Blood ◽  
2007 ◽  
Vol 110 (11) ◽  
pp. 2122-2122 ◽  
Author(s):  
Hyang-Min Byun ◽  
Shahrooz Eshaghian ◽  
Jia Yi Jiang ◽  
Si Ho Choi ◽  
John Soussa ◽  
...  
Keyword(s):  
Dna Methylation ◽  
Cpg Island ◽  
Blast Crisis ◽  
Bimodal Distribution ◽  
Epigenetic Therapy ◽  
Dna Hypermethylation ◽  
Cpg Sites ◽  
Genetic Translocation ◽  
Methylator Phenotype ◽  
Hypermethylated Genes

Abstract DNA methylation changes are a common finding in leukemia, and hypermethylation of CpG island promoters is associated with aberrant gene silencing. Some abnormal cancer related methylation changes have been associated with clinical phenotype including pathologic features, prognosis, and treatment response. However, other DNA methylation changes do not appear to have phenotypic consequences and may reflect a stochastic event or a downstream event of tumorogenesis, such as the CpG island methylator phenotype (CIMP). In order to obtain a better understanding of the DNA methylation changes found in leukemia we analyzed 18 acute promyelocytic leukemia (APL) and 36 chronic myeloid leukemia (CML) patients. We specifically chose to study APL and CML as these leukemia are initiated by specific genetic translocation events, t(15:17) and t(9:22) respectively. To measure the DNA methylation status, we used the GoldenGate Assay for Methylation and BeadArray technology from Illumina, Inc. The Standard Methylation Cancer Panel I from Illumina interrogates 1505 CpG sites, selected from 807 genes (231 genes contain one CpG site per gene, 463 genes contain two CpG sites and 114 genes have three or more CpG sites). In our study we found 142 and 269 genes that were hypermethylated in CML and APL. 31 genes were uniquely hypermethylated in CML, 158 genes were hypermethylated only in APL, and 111 genes were hypermethylated in both leukemias. There was a unique pattern of hypermethylated genes in each cancer; such there was a high concordance of hypermethylated genes within each leukemia type. These data suggest that the epigenetic events were a result of the genetic translocation BCR/ABL or PML/RARα (associated with chromosomal aberrations t(9:22) or t(15:17)) that initiates these leukemias. Analysis of the number of hypermethylated genes in these two leukemias showed a bimodal distribution suggestive of CIMP, however, closer examination showed that this bimodal distribution could be attributed to the two different types of leukemia. APL patients had mean of 280 genes hypermethylated while CML patients only had a mean of 193 genes hypermethylated. APL had a stronger methylator phenotype than CML for the loci studied, which underscores the possible relationship of CIMP to a genetic phenotype. Subset analysis of our CML samples by chronic phase (23 patients), accelerated phase (5 patients), and blast crisis (8 patients) revealed 42 genes that became hypermethylated with progression of CML. It is possible that hypermethylation of these genes are clinically important in the leukemia phenotype, and maybe targets for epigenetic therapy. We examined the DNA methylation changes induced by the DNA methylation inhibitor, azacitidine, in a patient with blast crisis CML and refractory to imatinib mesylate therapy. Azacitidine could reverse the aberrant hypermethylation associated with progression of CML to blast crisis and supports the use of this drug as an epigenetic therapy. Our data show that the majority of DNA hypermethylation events in leukemia are dependent on genetic events, but there is a subset of DNA hypermethylation events that are involved in the progression of leukemia and may be therapeutically reversed by DNA methylation inhibitors.

202 Combination of transcription activator-like effector nucleases and homology-independent target integration strategy gene editing technologies for knock-in of recombinant human factor IX Under the β-casein native promoter in bovine IVF embryos

2019 ◽  
Vol 31 (1) ◽  
pp. 226
Author(s):  
V. Savy ◽  
R. J. Bevacqua ◽  
N. G. Canel ◽  
V. Alberio ◽  
L. D. Ratner ◽  
...  
Keyword(s):  
Human Factor ◽  
Factor Ix ◽  
Regulatory Sequences ◽  
Sequencing Analysis ◽  
Transcription Activator ◽  
Site Specific ◽  
Human Factor Ix ◽  
Recombinant Human Factor Ix ◽  
Cytoplasmic Injection

Precise DNA modification is a crucial approach for gene function elucidation, biomedical model development, and transgenic bioreactor generation. In livestock, its application was extremely challenging until the development of engineered nucleases such as zinc-finger nucleases, transcription activator-like effector nucleases (TALEN), and CRISPR/Cas9. Still, precise knock-in (KI) techniques remain inefficient. Recently, the homology-independent target integration (HITI) strategy was developed, allowing precise insertion of transgenes in mammalian cells in an easier fashion. The HITI technique allows site-specific gene insertion by means of cleavage of both the target sequence in the genome and the donor plasmid, followed by DNA repair by nonhomologous end joining. Here, we evaluated the use of TALENs to generate precise knockout (KO) alleles of the β-casein gene (CSN2) by creating small insertions or deletions, and precise insertion of recombinant human factor IX (rhFIX) under bovine CSN2 regulatory sequences, using HITI via cytoplasmic injection of bovine IVF zygotes. First, 2 TALEN pairs (Tn1 and Tn2) targeting exon 2 of bovine CSN2 were designed and their activity was confirmed by primary fibroblasts transfection followed by Surveyor assay at Day 3. Then, both TALEN pairs were evaluated for KO embryo generation by zygote cytoplasmic injection of in vitro-transcribed mRNA encoding for Tn1, Tn2, or a mix containing Tn1+Tn2, at 100ng μL−1. A non-injected control (NIC) was also included. Embryos were in vitro cultured until Day 7 and independently analysed by whole-genome amplification followed by PCR and sequencing. Neither the blastocyst rate [28.8% (n=73), 33.8% (n=71), 32.4% (n=74), and 54.3% (n=127) for Tn1, Tn2, Tn1+Tn2, and NIC, respectively] nor the proportion of edited embryos [44% (n=9), 20% (n=10), and 33% (n=9) for Tn1, Tn2, and Tn1+Tn2, respectively] differed between injected groups (Fisher test, P<0.05), demonstrating efficient editing in bovine embryos by TALENs. Finally, to achieve precise CSN2 KI embryos, the rhFIX open reading frame was PCR amplified with a forward primer containing the Tn1 recognition sequence to obtain the HITI donor and bovine IVF zygotes were co-injected with the Tn1 mRNA and the HITI donor. Embryos were in vitro cultured until Day 7 and individually analysed by nested PCR at both the 5′ and 3′ ends of HITI donor. The PCR-based results indicate HITI donor integration in 7% of embryos analysed (n=14). Sanger sequencing analysis is currently in progress to confirm site-specific integration of HITI and possible rearranged DNA integration in other embryos. To our knowledge, this is the first report on the use of TALEN and HITI for gene modification. Our results indicate that TALEN combined with HITI may constitute an easy strategy for precise production of pharmaceuticals in the milk of livestock.

scholarly journals Vitamin C increases viral mimicry induced by 5-aza-2′-deoxycytidine

2016 ◽  
Vol 113 (37) ◽  
pp. 10238-10244 ◽  
Author(s):  
Minmin Liu ◽  
Hitoshi Ohtani ◽  
Wanding Zhou ◽  
Andreas Due Ørskov ◽  
Jessica Charlet ◽  
...  
Keyword(s):  
Vitamin C ◽  
Cellular Response ◽  
Dna Methyltransferase ◽  
Endogenous Retrovirus ◽  
Dna Demethylation ◽  
Epigenetic Therapy ◽  
Vitamin C Deficiency ◽  
Dna Methyltransferase Inhibitors ◽  
Tet Enzymes

Vitamin C deficiency is found in patients with cancer and might complicate various therapy paradigms. Here we show how this deficiency may influence the use of DNA methyltransferase inhibitors (DNMTis) for treatment of hematological neoplasias. In vitro, when vitamin C is added at physiological levels to low doses of the DNMTi 5-aza-2′-deoxycytidine (5-aza-CdR), there is a synergistic inhibition of cancer-cell proliferation and increased apoptosis. These effects are associated with enhanced immune signals including increased expression of bidirectionally transcribed endogenous retrovirus (ERV) transcripts, increased cytosolic dsRNA, and activation of an IFN-inducing cellular response. This synergistic effect is likely the result of both passive DNA demethylation by DNMTi and active conversion of 5-methylcytosine (5mC) to 5-hydroxymethylcytosine (5hmC) by ten–eleven translocation (TET) enzymes at LTR regions of ERVs, because vitamin C acts as a cofactor for TET proteins. In addition, TET2 knockout reduces the synergy between the two compounds. Furthermore, we show that many patients with hematological neoplasia are markedly vitamin C deficient. Thus, our data suggest that correction of vitamin C deficiency in patients with hematological and other cancers may improve responses to epigenetic therapy with DNMTis.

scholarly journals Epigenetic therapy of novel tumour suppressor ZAR1 and its cancer biomarker function

2019 ◽  
Vol 11 (1) ◽  
Author(s):  
Verena Deutschmeyer ◽  
Janina Breuer ◽  
Sara K. Walesch ◽  
Anna M. Sokol ◽  
Johannes Graumann ◽  
...  
Keyword(s):  
Lung Cancer ◽  
Zinc Finger ◽  
Cpg Island ◽  
Clinical Importance ◽  
Global Gene Expression ◽  
Cancer Biomarker ◽  
Tumour Suppressor ◽  
Epigenetic Therapy ◽  
Lung Carcinogenesis ◽  
Dna Hypermethylation

Abstract Background Cancer still is one of the leading causes of death and its death toll is predicted to rise further. We identified earlier the potential tumour suppressor zygote arrest 1 (ZAR1) to play a role in lung carcinogenesis through its epigenetic inactivation. Results We are the first to report that ZAR1 is epigenetically inactivated not only in lung cancer but also across cancer types, and ZAR1 methylation occurs across its complete CpG island. ZAR1 hypermethylation significantly correlates with its expression reduction in cancers. We are also the first to report that ZAR1 methylation and expression reduction are of clinical importance as a prognostic marker for lung cancer and kidney cancer. We further established that the carboxy (C)-terminally present zinc-finger of ZAR1 is relevant for its tumour suppression function and its protein partner binding associated with the mRNA/ribosomal network. Global gene expression profiling supported ZAR1's role in cell cycle arrest and p53 signalling pathway, and we could show that ZAR1 growth suppression was in part p53 dependent. Using the CRISPR-dCas9 tools, we were able to prove that epigenetic editing and reactivation of ZAR1 is possible in cancer cell lines. Conclusion ZAR1 is a novel cancer biomarker for lung and kidney, which is epigenetically silenced in various cancers by DNA hypermethylation. ZAR1 exerts its tumour suppressive function in part through p53 and through its zinc-finger domain. Epigenetic therapy can reactivate the ZAR1 tumour suppressor in cancer.

scholarly journals Genome-wide analysis of DNA methylation identifies the apoptosis-related gene UQCRH in renal cancer

2021 ◽  
Author(s):  
Kosuke Miyakuni ◽  
Jun Nishida ◽  
Daizo Koinuma ◽  
Genta Nagae ◽  
Hiroyuki Aburatani ◽  
...  
Keyword(s):  
Renal Cancer ◽  
Cancer Progression ◽  
Dna Methyltransferases ◽  
Tumor Formation ◽  
Dna Demethylation ◽  
Sequencing Data ◽  
Dna Hypermethylation ◽  
Cell Renal Cell Carcinoma ◽  
Methylation Array

Abstract Background DNA hypermethylation is frequently observed in clear cell renal cell carcinoma (ccRCC) and correlates with poor clinical outcomes. However, the detailed function is not fully uncovered. Methods A target for DNA methyltransferases (DNMTs) was explored in ccRCC cells. Highly malignant derivatives of human ccRCC cells were established via serial orthotopic inoculation in mice. Then, DNA methylated sites were genome-widely analyzed using methylation array in reference to RNA-sequencing data. Results We found that DNMT3B upregulation contributes to renal cancer progression and identified the ubiquinol cytochrome c reductase hinge protein (UQCRH) as a methylation target in advanced ccRCC. The expression of UQCRH in human ccRCC tissues was lower than that in normal adjacent tissues. Furthermore, its silencing attenuated cytochrome c release in response to apoptotic stimuli, resulting in enhanced primary tumor formation in vivo. Moreover, DNA demethylation enhanced the therapeutic efficiency of the mammalian target of rapamycin (mTOR) inhibitor everolimus in vivo. Conclusions These findings suggest that the DNMT3B-induced UQCRH methylation may contribute to renal cancer progression and that DNMT inhibitors exhibit potential clinical significance for ccRCC treatment.

Tamarix articulata (T. articulata) - An Important Halophytic Medicinal Plant with Potential Pharmacological Properties

2019 ◽  
Vol 20 (4) ◽  
pp. 285-292 ◽  
Author(s):  
Abdullah M. Alnuqaydan ◽  
Bilal Rah
Keyword(s):  
Medicinal Plant ◽  
Biological Activities ◽  
Wide Spectrum ◽  
Biological Properties ◽  
In Vivo Model ◽  
Drug Candidate ◽  
Phytochemical Analysis ◽  
Pharmacological Properties

Background:Tamarix Articulata (T. articulata), commonly known as Tamarisk or Athal in Arabic region, belongs to the Tamaricaece species. It is an important halophytic medicinal plant and a good source of polyphenolic phytochemical(s). In traditional medicines, T. articulata extract is commonly used, either singly or in combination with other plant extracts against different ailments since ancient times.Methods:Electronic database survey via Pubmed, Google Scholar, Researchgate, Scopus and Science Direct were used to review the scientific inputs until October 2018, by searching appropriate keywords. Literature related to pharmacological activities of T. articulata, Tamarix species, phytochemical analysis of T. articulata, biological activities of T. articulata extracts. All of these terms were used to search the scientific literature associated with T. articulata; the dosage of extract, route of administration, extract type, and in-vitro and in-vivo model.Results:Numerous reports revealed that T. articulata contains a wide spectrum of phytochemical(s), which enables it to have a wide window of biological properties. Owing to the presence of high content of phytochemical compounds like polyphenolics and flavonoids, T. articulata is a potential source of antioxidant, anti-inflammatory and antiproliferative properties. In view of these pharmacological properties, T. articulata could be a potential drug candidate to treat various clinical conditions including cancer in the near future.Conclusion:In this review, the spectrum of phytochemical(s) has been summarized for their pharmacological properties and the mechanisms of action, and the possible potential therapeutic applications of this plant against various diseases discussed.

Sign in / Sign up

Export Citation Format

Share Document