Frequent epigenetic inactivation of the RASSF1A gene in hepatocellular carcinoma

Schagdarsurengin, Undraga; Wilkens, Ludwig; Steinemann, Doris; Flemming, Peer; Kreipe, Hans; Pfeifer, Gerd P.; Schlegelberger, Brigitte; Dammann, Reinhard

doi:10.1038/sj.onc.1206338

Cited by 163 publications

(150 citation statements)

References 36 publications

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“…Detection of methylated DNA has then been suggested as a potential biomarker for early detection of cancer. In this study, HCC tumor tissues demonstrated hypermethylation in RASSF1A gene (88.6%, 31/35) and sera (40%, 14/35), which is in concordance with the findings from other investigators (tissue 60-92.5%; serum 42.5-70%) [18][19][20][21][22][23]. Although the exact mechanism of how the tumor DNA enters systemic circulation is unclear, the present result can still suggest that RASSF1A methylation should be a potential marker of incipient malignancy in the human hepatocarcinogenesis since, like any ideal biomarker, it appears early in the course of disease and is detectable in biological samples that can be obtained noninvasively.…”

Section: Discussionsupporting

confidence: 92%

“…The hypermethylation of RASSF1A suggests new perspectives for the diagnosis of malignant tumor. The methylation of RASSF1A was detected in the serum DNA of HCC patients by Yeo et al [18] and Zhang et al [19], as well as in the tissue DNA of HCC by Zhang et al [20], Di Gioia et al [21], Tischoff et al [22], and Schagdarsurengin et al [23]. But different groups obtained different data for the detection of hypermethylation of RASSF1A in HCC.…”

Section: Introductionmentioning

confidence: 99%

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Clinicopathological significance of RASSF1A reduced expression and hypermethylation in hepatocellular carcinoma

Chen

et al. 2010

Hepatol Int

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Purpose Protein downregulation and hypermethylation of Ras association domain family 1A (RASSF1A) has been recognized as an important early event in different classes of carcinogenesis, but clinicopathological significance of RASSF1A protein expression and methylation in hepatocellular carcinoma (HCC) remains largely unknown. The aim of the study was to investigate the expression of RASSF1A protein and methylation in HCC and their clinical significance. Methods Immunohistochemistry was employed to detect the expression of RASSF1A proteins in liver tissue microarrays. Aberrant promoter hypermethylation of RASSF1A was investigated in DNA from HCC, matching noncancerous tissues and serum of 35 HCC patients by methylation-specific PCR. Results RASSF1A protein expression in HCC was significantly lower than that in noncancerous (p = 0.015) and paracancerous tissues (p = 0.017). In addition, reduced RASSF1A protein expression is related to TNM stage, metastasis, a-fetoprotein, portal vein embolus, capsular infiltration, and multiple tumor nodes. Furthermore, RASSF1A promoter methylation in HCC was significantly higher than that in noncancerous liver tissues (p \ 0.05). Meanwhile, RASSF1A promoter hypermethylation was detected in 14 in the serum DNA from HCC patients, whereas no hypermethylation was detected in the normal controls. Hypermethylation of RASSF1A in HCC serum and tissues was negatively correlated with the expression of RASSF1A protein expression (p \ 0.05). Conclusions The loss or abnormal protein downregulation and the promoter hypermethylation of RASSF1A could play important roles in the tumorigenesis development and metastases of HCC. The detection of the promoter hypermethylation of RASSF1A in serum DNA could be a valuable biomarker for early-stage diagnosis in populations at high risk of HCC.

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Section: Discussionsupporting

confidence: 92%

Section: Introductionmentioning

confidence: 99%

Clinicopathological significance of RASSF1A reduced expression and hypermethylation in hepatocellular carcinoma

Chen

et al. 2010

Hepatol Int

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“…17,18 In human HCC, a number of methylation-induced inactivation of genes, such as E-cadherin, p16INK4a, 14-3-3 sigma, SOCS-1 and DLC-1, have already been documented. [19][20][21][22][23][24][25][26][27] However, there is no information on the methylation-mediated silencing of Apo D in HCC. Therefore, we first analyzed the pharmacological induction of Apo D by 5-Azacitidine in human liver cancer cell lines.…”

Section: Discussionmentioning

confidence: 99%

Clinicopathologic and prognostic values of apolipoprotein D alterations in hepatocellular carcinoma

Utsunomiya

Ogawa

Yoshinaga

et al. 2005

Intl Journal of Cancer

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We previously identified apolipoprotein D (Apo D) as a novel tumor suppressor gene based on the pharmacological unmasking of epigenetic silencing. We analyzed Apo D expression using realtime reverse transcription-PCR and evaluated its expression status based on the clinicopathological parameters of 70 patients with hepatocellular carcinoma (HCC). Immunohistochemical staining was also performed. We determined the methylation status of Apo D gene promoter by methylation-specific PCR (MSP). The Apo D gene-expression in tumor tissue was significantly lower than that in nontumor tissue (p = 0.011). A low Apo D expression significantly correlated with less-differentiated HCC ( p = 0.019). Immunohistochemical studies confirmed a decreased Apo D expression in poorly differentiated tumors. The prognosis of patients with a lower Apo D gene-expression was significantly worse than that in those with a higher expression (p = 0.028). The Apo D gene-expression was an independent prognostic factor (relative risk: 2.36, p = 0.018). An MSP assay showed a low-level of methylation in well differentiated HCC and a high-level of methylation in less differentiated tumors. Apo D may be a novel tumor suppressor gene of HCC, and its expression status may be a useful biomarker for predicting the patient outcome. ' 2005 Wiley-Liss, Inc.Key words: apolipoprotein D; hepatocellular carcinoma; histological grade; prognostic factor; methylationWe recently performed a comprehensive survey of commonly inactivated tumor suppressor genes in esophageal cancer cell lines based on functional reactivation by a demethylating agent. 1 A total of 58 silenced genes were thus identified using this approach, and one of those genes was Apolipoprotein D (Apo D). In our report, we detected the promoter hypermethylation of Apo D gene using bisulfite DNA sequencing. The methylation status of Apo D correlated closely with the Apo D expression status. Moreover, the growth suppressive activity of Apo D was also demonstrated by a colony focus assay. 1 These findings thus prompted us to analyze the clinical significance of the Apo D expression status in human cancers.Apo D, belonging to the lipocalin superfamily, is a glycoprotein of about 30 kDa found in the high-density lipoprotein fraction of human plasma. 2 The functional role of Apo D remains unclear, but the observation that this protein forms complexes with lecithin-cholesterol acyltransferase suggests that Apo D may be involved in cholesterol esterification. 3 More recently, the Apo D gene-expression was reported to be induced by retinoic acid in breast cancer cell lines. The induction of the Apo D expression was accompanied by an inhibition of cell proliferation and a progression through a more differentiated phenotype. These finding suggest that the mechanisms controlling retinoic acid-induced growth arrest, cell differentiation and Apo D synthesis may be directly coordinated in human breast cancer cells. 4 Furthermore, a low expression of Apo D determined by an immunohistochemical study was significantly as...

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“…MSP was performed using specific primers and conditions already described. 2,3,8,15,19,24 Detailed primer sequences are available on request. Briefly, a 20 ml reaction volume contained 150 ng bisulfite-modified DNA, 1 Â PCR buffer, 1.5 mM MgCl 2 , 0.16 mM dNTPs, 0.25 mM specific primer mix (forward and reverse primers) and 1 unit Taq enzyme (Roche, Mannheim, Germany).…”

Section: Methylation Analysismentioning

confidence: 99%

“…5,6 Several genes located on chromosome 3p have been studied in HCC as well as CC and include RASSF1A on 3p21.31, FHIT at 3p14.2, RIZ1, VHL at 3p25. [7][8][9][10] These results directed an intensive search for possible tumor suppressor genes located in the 3p21 region for one or more genes that could function as gatekeepers in molecular pathogenesis of human cancers. A group of candidate tumor suppressor genes (designated BLU, SEMAPHORIN 3B, or RASSF1A) has recently been mapped to this critical gene-rich region.…”

mentioning

confidence: 99%

Allele loss and epigenetic inactivation of 3p21.3 in malignant liver tumors

Tischoff

Markwarth

Witzigmann

et al. 2005

Intl Journal of Cancer

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Previously, the RASSF1A, BLU and SEMAPHORIN 3B (SEMA3B) candidate tumor suppressor genes on chromosome 3p21.3 were found to be inactivated and downregulated by genetic and epigenetic changes in lung cancer. We analyzed the methylation status of RASSF1A, BLU and SEMA3B in 35 hepatocellular carcinomas (HCCs) and 15 cholangiocarcinomas (CCs) by methylationspecific PCR and loss of heterozygosity (LOH) at 3p21.3 after microdissection. The presence of mRNA transcripts was confirmed by semiquantitative PCR. SEMA3B hypermethylation was found in 29/35 HCCs (83%) and in all (15/15) patients with CC. BLU promoter hypermethylation was detected in 7/35 (20%) HCCs and 3/15 (20%) CCs. In 2 corresponding specimens of hepatitis B virus-related liver cirrhosis, BLU methylation was also observed, but not in uninvolved normal liver tissue. RASSF1A was methylated in 21/35 HCCs (60%) and in 10/15 CCs (67%). LOH at 3p21.3 occurred in 8/35 (23%) HCCs and 3/15 (20%) CCs. The presence of hypermethylation was statistically associated with LOH of SEMA3B and correlated with downregulation of mRNA transcripts. SEMA3B transcripts increased upon treatment of HCC cell lines with the demethylation compound 5-aza-2-deoxycytidine. In conclusion, our data indicate that 2-hit gene silencing of SEMA3B through epigenetic changes and allele loss is a common and important event in the carcinogenesis of malignant liver tumors. ' 2005 Wiley-Liss, Inc.Key words: hepatocellular carcinoma; cholangiocarcinoma; methylation; loss of heterozygosity; BLU; SEMA3B; RASSF1A Primary liver cancer is one of the most frequent carcinomas worldwide. Besides hepatocellular carcinoma (HCC), accounting for 80-90% of all primary liver cancers, cholangiocarcinoma (CC) is the second most common primary hepatic malignancy. The short arm of chromosome 3 has been shown to exhibit loss of heterozygosity (LOH) in several types of cancer, including ovarian, kidney, lung, nasopharyngeal and also liver cancer. 1,2 In particular, overlapping homozygous deletions in lung cancers have been identified in region 3p21.3. 3,4 In HCC, LOH of chromosome 3p occurred in about 30% of the patients. 5,6 Several genes located on chromosome 3p have been studied in HCC as well as CC and include RASSF1A on 3p21.31, FHIT at 3p14.2, RIZ1, VHL at 3p25. 7-10 These results directed an intensive search for possible tumor suppressor genes located in the 3p21 region for one or more genes that could function as gatekeepers in molecular pathogenesis of human cancers. A group of candidate tumor suppressor genes (designated BLU, SEMAPHORIN 3B, or RASSF1A) has recently been mapped to this critical gene-rich region. 2,[11][12][13] SEMAPHORIN 3B (SEMA3B) is a member of the semaphorin family, which is also located on 3p21.3. Semaphorins are a family of signaling molecules initially identified to play a role in axonal guidance and can be classified as either membrane-bound (classes 1, 4, 5 and 6) or secreted (classes 2 and 3). 14 The receptors for the class 3 semaphorins are also known as neuropilin receptors. The intr...

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Frequent epigenetic inactivation of the RASSF1A gene in hepatocellular carcinoma

Cited by 163 publications

References 36 publications

Clinicopathological significance of RASSF1A reduced expression and hypermethylation in hepatocellular carcinoma

Clinicopathological significance of RASSF1A reduced expression and hypermethylation in hepatocellular carcinoma

Clinicopathologic and prognostic values of apolipoprotein D alterations in hepatocellular carcinoma

Allele loss and epigenetic inactivation of 3p21.3 in malignant liver tumors

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