scholarly journals Metallothioneins Gene Regulation Through Promoter DNA Methylation According to Zn and Cu Trace Elements Status in Human Hepatocellular Carcinoma (P05-002-19)

2019 ◽  
Vol 3 (Supplement_1) ◽  
Author(s):  
Domenica De Santis ◽  
Silvia Udali ◽  
Filippo Mazzi ◽  
Andrea Ruzzenente ◽  
Greta Beschin ◽  
...  
Keyword(s):  
Hepatocellular Carcinoma ◽  
Dna Methylation ◽  
Trace Elements ◽  
Tumor Suppressor ◽  
Tumor Suppressor Genes ◽  
Funding Sources ◽  
Promoter Dna Methylation ◽  
Serum Trace Elements ◽  
Promoter Dna

Abstract Objectives Hepatocellular carcinoma (HCC) is the most frequent primary liver cancer, yet mechanisms of hepatocarcinogenesis are largely unknown. A particular interest was recently dedicated to the role of trace elements and metallothioneins (MTs), a group of proteins involved in metal ions homeostasis and detoxification, have been suggested as possible tumor suppressor genes. The study of MTs transcriptional regulation by promoter DNA methylation is the object of study as a possible mechanism responsible for gene silencing through epigenetics. Methods Twenty-seven HCC patients undergoing surgery intervention were enrolled and clinically characterized. MT1G and MT1H gene expression was performed by Real Time qPCR. DNA methylation analysis in 23 HCC and homologous non-neoplastic liver tissue (N) was performed by Bisulfite-Amplicon Sequencing (BSAS) in an overlapping region (∼400 bp) of the promoters of the two genes. Cu and Zn concentrations were measured in serum and liver tissues (HCC and N) by Inductively Coupled Plasma Mass Spectrometry (ICP-MS). Kaplan-Meier analysis of survival was performed according to serum trace elements. Results MT1G and MT1H were transcriptionally repressed in HCC tissue as compared to N. A correlation was observed between the mRNA levels of the two MTs, in particular MT1G was repressed in 23 out of 27 HCC tissue (P = 0.0366) and MT1H was repressed in 24 out of 27 HCC tissue (P = 0.0077). The promoter region resulted hypermethylated in 9 out of 19 HCC that showed MT1G and MT1H down-regulation. Serum Zn and Cu levels were within the normal range while HCC tissue exhibited significantly reduced Zn levels as compared to N (P < 0.0001). Tissue Cu levels did not show significant differences. Serum trace elements levels were also analyzed according to patients clinical features and those with Cu levels higher than the 75th percentile had a significantly poorer prognosis than those within the lowest Cu levels quartile (P < 0.05). Conclusions MT1G and MT1H are repressed in HCC tissue. In a subset of patients the downregulation was associated to promoter hypermethylation, supporting the hypothesis of MT1G and MT1H as possible tumor suppressor genes in HCC. Evidence of a correlation between serum Cu levels and survival rate provide new insights for the role of this microelement in liver carcinogenesis. Funding Sources No funding sources.

scholarly journals Hypermethylation of MT1G and MT1H CpGs Promoter Sites and High Serum Levels of Cu Affect Survival Rate of Patients Affected by Hepatocellular Carcinoma

2020 ◽  
Vol 4 (Supplement_2) ◽  
pp. 1791-1791
Author(s):  
Domenica De Santis ◽  
Silvia Udali ◽  
Andrea Ruzzenente ◽  
Greta Beschin ◽  
Patrizia Pattini ◽  
...  
Keyword(s):  
Hepatocellular Carcinoma ◽  
Dna Methylation ◽  
Trace Elements ◽  
Survival Rate ◽  
Mortality Rate ◽  
Liver Tissue ◽  
Kaplan Meier ◽  
Promoter Dna Methylation ◽  
Promoter Dna

Abstract Objectives Recent evidences suggest a principal role of trace elements and metallothioneins (MTs), proteins involved in metal ions homeostasis and detoxification, in hepatocellular carcinogenesis. The study was designed to evaluate whether serum and liver tissue concentrations of the trace elements Cu, Zn and Se are implicated in survival rate of hepatocellular carcinoma (HCC) patients and if promoter DNA methylation is involved in trace elements-related proteins regulation. Methods Cu, Zn and Se levels were determined in serum and liver tissue samples, both HCC and homologous non neoplastic tissue (N) of 27 HCC patients by Inductively Coupled Plasma Mass Spectrometry (ICP-MS). Gene expression analysis of MT1G and MT1H, was performed by Real-time qPCR in HCC and N tissue. Promoter DNA methylation of a region overlapping MT1G and MT1H promoters was assessed by bisulfite amplicon sequencing (BSAS) in HCC and N tissues of 23 patients. Kaplan-Meier survival curves were drawn using the log-rank test (Mantel-Cox test) to examine the differences in survival according to serum trace elements and to gene-specific methylation levels. Results Kaplan-Meier analysis according to serum Cu levels showed that subjects within the highest quintile had an increased mortality rate (88.9%) compared with the other four quintiles (P = 0.025). Considering the 80th percentile of Cu levels (1118 μg/L), subjects with Cu concentrations above this value had a significantly decreased survival rate (P &lt; 0.001). Se and Zn content were depleted in HCC tissues as compared to N tissues (P &lt; 0.0001). MT1G and MT1H were strongly repressed in HCC tissues and precisely, MT1H in 24 out of 27 HCC tissues (P = 0.008) and MT1G in 23 out of 27 HCC tissues (P = 0.037). Nine out of 19 HCC tissues showing a down-regulation of MTs with three CpG sites, significantly hypermethylated in HCC tissue as compared to N tissue (P &lt; 0.05). Considering the median methylation level, patients with higher methylation values showed increased mortality rate (P = 0.015). Conclusions The significant repression of MT1G and MT1H in HCC tissue is related to promoter hypermethylation and support the hypothesis of MT1G and MT1H as possible tumor suppressor genes in HCC. The evidence of promoter methylation levels and survival rate association provide new insights for the role of DNA methylation in liver carcinogenesis. Funding Sources N/A.

scholarly journals Trace Elements Status and Metallothioneins DNA Methylation Influence Human Hepatocellular Carcinoma Survival Rate

2021 ◽  
Vol 10 ◽  
Author(s):  
Silvia Udali ◽  
Domenica De Santis ◽  
Filippo Mazzi ◽  
Sara Moruzzi ◽  
Andrea Ruzzenente ◽  
...  
Keyword(s):  
Gene Expression ◽  
Hepatocellular Carcinoma ◽  
Dna Methylation ◽  
Trace Elements ◽  
Survival Rate ◽  
Inductively Coupled Plasma ◽  
Prognostic Significance ◽  
Promoter Dna Methylation ◽  
Promoter Dna

BackgroundMechanisms underlying hepatocellular carcinoma (HCC) development are largely unknown. The role of trace elements and proteins regulating metal ions homeostasis, i.e. metallothioneins (MTs), recently gained an increased interest. Object of the study was to investigate the role of promoter DNA methylation in MTs transcriptional regulation and the possible prognostic significance of serum trace elements in HCC.MethodsForty-nine HCC patients were enrolled and clinically characterized. Cu, Se, and Zn contents were measured by Inductively Coupled Plasma Mass Spectrometry in the serum and, for a subset of 27 patients, in HCC and homologous non-neoplastic liver (N) tissues. MT1G and MT1H gene expression in hepatic tissues was assessed by Real-Time RT-PCR and the specific promoter DNA methylation by Bisulfite-Amplicon Sequencing.ResultsPatients with Cu serum concentration above the 80th percentile had a significantly decreased survival rate (P &lt; 0.001) with a marked increased hazard ratio for mortality (HR 6.88 with 95% CI 2.60–18.23, P &lt; 0.001). Se and Zn levels were significantly lower in HCC as compared to N tissues (P &lt; 0.0001). MT1G and MT1H gene expression was significantly down-regulated in HCC as compared to N tissues (P &lt; 0.05). MTs promoter was hypermethylated in 9 out of the 19 HCC tissues showing MTs down-regulation and methylation levels of three specific CpGs paralleled to an increased mortality rate among the 23 patients analyzed (P = 0.015).ConclusionsMT1G and MT1H act as potential tumor suppressor genes regulated through promoter DNA methylation and, together with serum Cu concentrations, be related to survival rate in HCC.

Epigenetic Silencing of Tumor Suppressor lncRNA NKILA: Implication on NF-κB Signaling in Non-Hodgkin’s Lymphoma

2020 ◽  
Author(s):  
Min Yue Zhang ◽  
Ming Dan Deng ◽  
Lu Qian Wang ◽  
Rex K.H. Au-Yeung ◽  
Chor Sang Chim
Keyword(s):  
Dna Methylation ◽  
Cell Death ◽  
Tumor Suppressor ◽  
Hodgkin’S Lymphoma ◽  
Cellular Proliferation ◽  
Cell Lymphoma ◽  
Hodgkin's Lymphoma ◽  
Non Hodgkin's Lymphoma ◽  
Promoter Dna Methylation ◽  
Promoter Dna

Abstract Background: NKILA, localized to 20q13.31, is a negative regulator of NF-κB signaling implicated in carcinogenesis. As a CpG island is embedded in the promoter region of NKILA, we hypothesized that NKILA is a tumor suppressor lncRNA reversibly silenced by promoter DNA methylation in non-Hodgkin’s lymphoma (NHL). Results: By pyrosequencing-verified methylation-specific PCR (MSP), NKILA was unmethylated in normal healthy controls, including 10 peripheral blood buffy coats and 11 normal tonsils tissue, but completely methylated in one (10%) NHL cell line SU-DHL-6. Among the lymphoma cell lines, by semi-quantitative RT-PCR, methylation of NKILA was inversely correlated with its expression. In the completely methylated SU-DHL-6 cells, hypomethylation treatment with 5-Aza-2'-deoxycytidine resulted in promoter demethylation and re-expression of NKILA transcript. In NHL primary samples (n=102), NKILA methylation was observed none of mantle cell lymphoma (MCL) cases, but in 29 (51.79%) diffuse large-B cell lymphoma (DLBCL) and 4 (20%) peripheral T-cell lymphoma (PTCL) cases, hence preferentially methylated in DLBCL than MCL (P < 0.0001) and PTCL (P = 0.007). Mechanistically, knockdown of NKILA resulted in promoting IkBα phosphorylation, which was associated with nucleus translocation of total p65 and phosphorylated p65 in SU-DHL-1 cells, hence constitutive NF-κB activation. Functionally, knock-down of NKILA in SU-DHL-1 cells led to decreased cell death and increased cellular proliferation, indicating a tumor suppressor role of NKILA in NHL cells. Conclusions: NKILA was a tumour suppressor lncRNA frequently hypermethylated in DLBCL. Promoter DNA methylation-mediated NKILA silencing led to increase of cellular proliferation and decrease of cell death via repression of NF-κB signaling in NHL cells.

Strategies to Re-Express Epigenetically-Silenced Tumor Suppressor Genes Converge on the Requirement for Inhibition of the Histone Methyltransferase SUV39H1.

Blood ◽  
2008 ◽  
Vol 112 (11) ◽  
pp. 3357-3357
Author(s):  
Asha Lakshmikuttyamma ◽  
Stuart Scott ◽  
David P. Sheridan ◽  
John DeCoteau ◽  
Ron Geyer
Keyword(s):  
Tumor Suppressor ◽  
Cell Lines ◽  
Tumor Suppressor Genes ◽  
Fold Increase ◽  
Dna Demethylation ◽  
Dna Hypermethylation ◽  
Suppressor Genes ◽  
Promoter Dna Methylation ◽  
Promoter Dna ◽  
E Cadherin

Abstract Gene silencing mediated by aberrant promoter DNA hypermethylation represents a key mechanism by which tumor suppressor gene expression is silenced in cancer and it is associated with multiple repressive histone modifications. Histone H3 lysine 9 (H3K9) methylation is a key repressive chromatin modification with important implications for regulating cell proliferation, differentiation, and gene expression. SUV39H1 is a methyltransferase that catalyzes the addition of trimethyl groups to H3K9. SUV39H1 is associated with regions of hypermethylated CpG islands, with repressive complexes, such as RB/E2F, and with DNA-binding proteins involved in leukemogenesis, such as AML1 and PML-RAR, where its H3K9 trimethylation activity promotes heterochromatin formation and gene silencing. We studied the requirement of SUV39H1 in the epigenetic silencing of heavily methylated tumor suppressor genes p15INK4B and E-cadherin in acute myeloid leukemia (AML). Treatment of AML cell lines AML193, KG1a, and Kasumi with the DNA methyltransferase (DNMT) inhibitor 5-Aza-2’-deoxycytidine (5-Aza-dC) induces p15INK4B and E-cadeherin re-expression in association with dramatic decreases in p15INK4B and E-cadherin promoter DNA methylation and marked reductions in the levels of SUV39H1 and H3K9 trimethylation at these promoters. Interestingly, treatment of these cell lines with SUV39H1 shRNA, or the SUV39H1 inhibitor chaetocin, also induces p15INK4B and E-cadherin re-expression and H3K9 demethylation, without affecting promoter DNA methylation. Thus, re-expression of hypermethylated tumor suppressors requires histone H3K9 demethylation, which can be achieved indirectly by decreasing the amount of SUV39H1 associated with the promoter using 5-Aza-dC, or directly by inhibiting SUV39H1 expression or activity without requiring promoter DNA demethylation. Furthermore, we found that SUV39H1 shRNA or chaetocin in combination with 5-Aza-dC acts synergistically to re-express epigenetically silenced p15INK4B and E-cadherin in AML cell lines. Treatment of primary human AML blasts obtained from two patients with combinations of 5-Aza-C and chaetocin also results in synergistic re-expression of p15INK4B and E-cadherin (2–6 fold increase with 5-Aza-C or chaetocin treatment vs. 11–14 fold increase with co-treatment). Our study has important implications for developing novel epigenetic therapies of relevance to AML as it suggests that the re-expression of tumor suppressor genes silenced by aberrant promoter DNA hypermethylation converges on the requirement for SUV39H1 and H3K9 methylation inhibition but not promoter DNA demethylation. Our finding that SUV39H1 inhibition may function synergistically with DNMT inhibitors to enhance gene reactivation and chromatin changes also highlights the needs for developing more inhibitors of histone methyltransferases and for performing detailed drug interaction studies to identify the best drug combinations for optimal epigenetic therapies.

scholarly journals Epigenetic Silencing of Tumor Suppressor lncRNA NKILA: Implication on NF-κB Signaling in Non-Hodgkin’s Lymphoma

2021 ◽  
Author(s):  
Min Yue Zhang ◽  
George Calin ◽  
Ming Dan Deng ◽  
Rex K.H. Au-Yeung ◽  
Lu Qian Wang ◽  
...  
Keyword(s):  
Dna Methylation ◽  
Cell Death ◽  
Tumor Suppressor ◽  
Cell Lines ◽  
Peripheral Blood ◽  
Cellular Proliferation ◽  
Cell Lymphoma ◽  
Hodgkin's Lymphoma ◽  
Promoter Dna Methylation ◽  
Promoter Dna

Abstract Background The long non-coding RNA (lncRNA) NKILA, localized to 20q13.31, is a negative regulator of NF-κB signaling. As a CpG island is embedded in the promoter region of NKILA, NKILA is hypothesized as a tumor suppressor lncRNA reversibly silenced by promoter DNA methylation in non-Hodgkin’s lymphoma (NHL). Methods Methylation-specific PCR (MSP) and quantitative bisulfite pyrosequencing were performed to detect the methylation of NKILA in normal peripheral blood buffy coats, normal tonsils tissue, NHL cell lines and NHL primary samples. SU-DHL-6 cells were treated with 5-Aza-2'-deoxycytidine for reversal of methylation-associated NKILA silencing. Tumor suppressor properties and biological function of NKILA were demonstrated by knockdown of NKILA in SU-DHL-1 cells. Results By pyrosequencing-verified MSP, NKILA was unmethylated in normal healthy controls, including 10 peripheral blood buffy coats and 11 tonsils tissue, but completely methylated in one (10%) NHL cell line SU-DHL-6. Among the lymphoma cell lines, by semi-quantitative RT-PCR, methylation of NKILA was inversely correlated with its expression. In the completely methylated SU-DHL-6 cells, hypomethylation treatment with 5-Aza-2'-deoxycytidine resulted in promoter demethylation and re-expression of NKILA transcript. In 102 NHL primary samples, NKILA methylation was observed in none of mantle cell lymphoma (MCL) cases, but in 29 (51.79%) diffuse large-B cell lymphoma (DLBCL) and 4 (20%) peripheral T-cell lymphoma (PTCL) cases, hence preferentially methylated in DLBCL than MCL (P < 0.0001) and PTCL (P = 0.007). Mechanistically, knockdown of NKILA resulted in promoting IkBα phosphorylation associated with nucleus translocation of total p65 and phosphorylated p65 in SU-DHL-1 cells, hence constitutive NF-κB activation. Functionally, knock-down of NKILA in SU-DHL-1 cells led to decreased cell death and increased cellular proliferation, indicating a tumor suppressor role of NKILA in NHL cells. Conclusion NKILA is a tumour suppressor lncRNA frequently hypermethylated in DLBCL. Promoter DNA methylation-mediated NKILA silencing led to increase of cellular proliferation and decrease of cell death via repression of NF-κB signaling in NHL cells.

No Reversal of Demethylation after Azacitidine Treatment in Concordance with Poor Clinical Response.

Blood ◽  
2007 ◽  
Vol 110 (11) ◽  
pp. 4629-4629
Author(s):  
Huong Thi Thanh Tran ◽  
Hee Nam Kim ◽  
JaeSook Ahn ◽  
Il-Kwon Lee ◽  
Deok-Hwan Yang ◽  
...  
Keyword(s):  
Tumor Suppressor ◽  
Clinical Response ◽  
Methylation Level ◽  
Tumor Suppressor Genes ◽  
Gene Mutations ◽  
Molecular Monitoring ◽  
Suppressor Genes ◽  
Promoter Dna Methylation ◽  
History Of ◽  
Promoter Dna

Abstract The epigenetic gene silencing associated with promoter DNA methylation is as powerful as gene mutations in functionally inactivating tumor suppressor genes. Thus, a non-intensive treatment may be changed the natural history of MDS for the first time by the demethylating agent, 5-aza-deoxycytidine (Decitabine) with silenced gene expression by reversal of p15 hypermethylation and protein expression in the bone marrow in MDS. The MS-MLPA (methylation-specific multiplex ligation-dependent probe amplification) ME-001B probemix (MRC-Holland) containing 25 tumor suppressor genes has been used to detect the methylation level in the peripheral blood samples of 29 MDS before azacitidine (Vidaza) and only 6 MDS after 3–5 courses of therapy. Patients that hypermethylated at least 1 gene were 7 of 29, either the common hypermethylating genes as p15, ESR1 or the previously known FHIT in MDS also have occurred. Only two patients except one patient related to either methylation level-reducing gene or removal methylated gene (putative demethylation reversal) have in concordance with clinical response in hematological evidence. Interestingly, three other patients were high methylation level persistently or additional methylated gene after treatment (putative demethylation no reversal or more severe), two patients of these are correspond with no clinical response and one is propensity to progressing leukemia. With IGSF4 gene hypermethylation, to the best of our knowledge, there was no report in MDS. Our results suggest that methylation level possibly contributes to the dignosistic, prognosistic and a molecular monitoring marker after treatment of Azacitidine.

scholarly journals Promoter DNA Methylation Frequency and Clinicopathological Role of miR-129-2 Gene in Patients with Chronic Lymphocytic Leukemia

2020 ◽  
Vol 35 (4) ◽  
pp. e151-e151
Author(s):  
Morteza Hashemi ◽  
Mahshid Mohammadipour ◽  
Shahrbano Rostami ◽  
Mohammad Soleiman Soltanpour
Keyword(s):  
Dna Methylation ◽  
Chronic Lymphocytic Leukemia ◽  
Lymphocytic Leukemia ◽  
Methylation Frequency ◽  
Promoter Dna Methylation ◽  
Promoter Dna

Role of tissue-specific promoter DNA methylation in regulating the human EKLF gene

2018 ◽  
Vol 71 ◽  
pp. 16-22 ◽  
Author(s):  
Yihong Li ◽  
Dun Liu ◽  
Zhiming Li ◽  
Xinhua Zhang ◽  
Yuhua Ye ◽  
...  
Keyword(s):  
Dna Methylation ◽  
Tissue Specific ◽  
Promoter Dna Methylation ◽  
Tissue Specific Promoter ◽  
Promoter Dna

scholarly journals Epigenetic Silencing of Tumor Suppressor lncRNA NKILA: Implication on NF-κB Signaling in Non-Hodgkin’s Lymphoma

Genes ◽  
2022 ◽  
Vol 13 (1) ◽  
pp. 128
Author(s):  
Min-Yue Zhang ◽  
George Calin ◽  
Ming-Dan Deng ◽  
Rex K. H. Au-Yeung ◽  
Lu-Qian Wang ◽  
...  
Keyword(s):  
Dna Methylation ◽  
Cell Death ◽  
Tumor Suppressor ◽  
Cell Lines ◽  
Cellular Proliferation ◽  
Cell Lymphoma ◽  
Hodgkin's Lymphoma ◽  
Non Hodgkin's Lymphoma ◽  
Promoter Dna Methylation ◽  
Promoter Dna

The long non-coding RNA (lncRNA) NKILA, localized to 20q13.31, is a negative regulator of NF-κB signaling implicated in carcinogenesis. As a CpG island is embedded in the promoter region of NKILA, it is hypothesized as a tumor suppressor lncRNA silenced by promoter DNA methylation in non-Hodgkin’s lymphoma (NHL). By pyrosequencing-verified methylation-specific PCR, NKILA methylation was detected in 1/10 (10%) NHL cell lines, but not in normal peripheral blood buffy coats or tonsils. NKILA methylation correlated with the repression of NKILA in cell lines. Hypomethylation treatment with 5-Aza-2′-deoxycytidine resulted in promoter demethylation and the re-expression of NKILA. In 102 NHL primary samples, NKILA was methylated in 29 (51.79%) diffuse large B-cell lymphoma (DLBCL) and 4 (20%) peripheral T-cell lymphoma cases, but unmethylated in all 26 mantle cell lymphoma cases. Mechanistically, the knockdown of NKILA resulted in promoting IkBα phosphorylation, associated with nucleus translocation of total p65 and phosphorylated p65 in SU-DHL-1 cells, hence constitutive NF-κB activation. Functionally, the knockdown of NKILA in SU-DHL-1 cells led to decreased cell death and increased cellular proliferation. Collectively, NKILA was a tumor suppressor lncRNA frequently hypermethylated in DLBCL. Promoter DNA methylation-mediated NKILA silencing resulted in increased cellular proliferation and decreased cell death via the repression of NF-κB signaling in NHL.

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