Ras Association Domain Family 1A Promoter Research Articles

Ras Association Domain Family 1A Gene Promoter Methylation as a Biomarker for Chronic Viral Hepatitis C-related Hepatocellular Carcinoma.

Background One of the most prevalent aberrant epigenetic modifications found in hepatocellular carcinoma (HCC) is abnormal DNA methylation. Our study aimed to evaluate serum Ras association domain family 1A (RASSF1A) gene promoter methylation in patients with chronic viral hepatitis C (HCV)-associated liver cirrhosis with and without HCC as a potential new marker for the early detection of HCC. Methodology The 60 participants who participated in the trial were divided into the following three groups: 20 patients with newly diagnosed primary HCC on top of HCV-related liver cirrhosis, 20 patients with HCV-related liver cirrhosis, and 20 age- and sex-matched healthy individuals as a control group. All participants underwent methylation-specific polymerase chain reaction testing to detect the blood level of the RASSF1A gene's methylated promoter. Results Methylated RASSF1A was found in 30% of patients with liver cirrhosis caused by HCV and in 65% of patients with HCC, but not in any of the controls. It was discovered that the serum methylation RASSF1A had an accuracy of 82.50% and an area under the curve (AUC) of 0.825 for separating HCC patients from healthy controls. With an AUC of 0.675 and an accuracy of 67.50%, it was able to differentiate patients with HCC from those with HCV-related liver cirrhosis. Additionally, there was no statistically significant association between RASSF1A methylation status and HCC mass size (p = 0.449). Conclusions Serum RASSF1A promoter methylation status detection could be useful for detecting HCC early, especially in high-risk individuals such as those with HCV.

REX-1 Represses RASSF1a and Activates the MEK/ERK Pathway to Promote Tumorigenesis in Prostate Cancer.

Epigenetics play an important role in the pathogenesis of prostate cancer; it is urgent to investigate vital transcription factors in methylation regulation with the aim to develop novel treatment strategies targeting prostate cancer. As a member of the zinc finger protein family, REX-1 (reduced expression-1) is a transcription factor that has been reported to be closely linked to the development of several cancers. So far, the expression level and precise function of REX-1 in prostate cancer remain largely unknown. Here, we show that REX-1 was overexpressed in prostate cancer clinical tissues, and its expression level was closely correlated with patient prognosis. REX-1 affected prostate tumor growth in vivo by MEK/ERK phosphorylation. Mechanistic studies indicated that REX-1 recruited DNMT3b (DNA methyltransferase 3b), inhibited the transcription of RASSF1a (RAS association domain family 1a), and further modulated methylation of RASSF1a promoter. Intervention of the REX-1/DNMT3b/RASSF1a axis may shed light on the development of novel therapeutic approaches for prostate cancer treatment. IMPLICATIONS: REX1 overexpression recruits DNMT3b and downregulates RASSF1a by promoter methylation, suggesting that epigenetic intervention may contribute to prostate cancer treatment.

A RASSF1A-HIF1\u03b1 loop drives Warburg effect in cancer and pulmonary hypertension

Hypoxia signaling plays a major role in non-malignant and malignant hyperproliferative diseases. Pulmonary hypertension (PH), a hypoxia-driven vascular disease, is characterized by a glycolytic switch similar to the Warburg effect in cancer. Ras association domain family 1A (RASSF1A) is a scaffold protein that acts as a tumour suppressor. Here we show that hypoxia promotes stabilization of RASSF1A through NOX-1- and protein kinase C- dependent phosphorylation. In parallel, hypoxia inducible factor-1 α (HIF-1α) activates RASSF1A transcription via HIF-binding sites in the RASSF1A promoter region. Vice versa, RASSF1A binds to HIF-1α, blocks its prolyl-hydroxylation and proteasomal degradation, and thus enhances the activation of the glycolytic switch. We find that this mechanism operates in experimental hypoxia-induced PH, which is blocked in RASSF1A knockout mice, in human primary PH vascular cells, and in a subset of human lung cancer cells. We conclude that RASSF1A-HIF-1α forms a feedforward loop driving hypoxia signaling in PH and cancer.

The relationship between RASSF1A promoter methylation and thyroid carcinoma: A meta-analysis of 14 articles and a bioinformatics of 2 databases (PRISMA).

DNA promoter methylation can suppresses gene expression and shows an important role in the biological functions of Ras association domain family 1A (RASSF1A). Many studies have performed to elucidate the role of RASSF1A promoter methylation in thyroid carcinoma, while the results were conflicting and heterogeneous. Here, we analyzed the data of databases to determine the relationship between RASSF1A promoter methylation and thyroid carcinoma. We used the data from 14 cancer-normal studies and Gene Expression Omnibus (GEO) database to analyze RASSF1A promoter methylation in thyroid carcinoma susceptibility. The data from the Cancer Genome Atlas project (TCGA) database was used to analyze the relationship between RASSF1A promoter methylation and thyroid carcinoma susceptibility, clinical characteristics, prognosis. Odds ratios were estimated for thyroid carcinoma susceptibility and hazard ratios were estimated for thyroid carcinoma prognosis. The heterogeneity between studies of meta-analysis was explored using H, I values, and meta-regression. We adopted quality criteria to classify the studies of meta-analysis. Subgroup analyses were done for thyroid carcinoma susceptibility according to ethnicity, methods, and primers. Result of meta-analysis indicated that RASSF1A promoter methylation is associated with higher susceptibility to thyroid carcinoma with small heterogeneity. Similarly, the result from GEO database also showed that a significant association between RASSF1A gene promoter methylation and thyroid carcinoma susceptibility. For the results of TCGA database, we found that RASSF1A promoter methylation is associated with susceptibility and poor disease-free survival (DFS) of thyroid carcinoma. In addition, we also found a close association between RASSF1A promoter methylation and patient tumor stage and age, but not in patients of different genders. The methylation status of RASSF1A promoter is strongly associated with thyroid carcinoma susceptibility and DFS. The RASSF1A promoter methylation test can be applied in the clinical diagnosis of thyroid carcinoma.

The loss-of-function of DNA methyltransferase 1 by siRNA impairs the growth of non-small cell lung cancer with alleviated side effects via reactivation of RASSF1A and APC in vitro and vivo.

Hypermethylation of tumor suppressor genes (TSGs) promoters by DNA methyltransferase (DNMT) can be observed in almost all cancers which represent a hallmark of carcinogenesis, including lung cancer. DNMT inhibitors (e.g.5-Aza-CR/CdR) reactivate TSGs to exert anti-cancer activity and have been applied into the clinical. However, it is cytotoxic even at low concentrations, which might be not directly related to DNA methylation. We here investigated an alternative strategy in the lung cancer therapy and aimed to estimate and compare its efficiency and side effects of knockdown of DNMT1 in vitro and in vivo. Lung cancer tissues (n=20) showed enhanced expression of DNMT1 than corresponding non-neoplastic tissues. Similar results were found in lung cancer cell lines A549 and H538. The treatment of 5-Aza-CR or knockdown of DNMT1 in vitro could inhibit the expressions of DNMT1 but restore the TSGs expressions including the Ras association domain family 1A (RASSF1A) and the adenomatous polyposis coli (APC) via the demethylation of its promoter region, which results in the decreased proliferation, increased apoptosis and impaired ability of migration. Importantly, knockdown of DNMT1 by siRNA in vivo also effectively demethylated the RASSF1A and APC promoter, elevated their expressions and limited tumor growth, which functioned like 5-Aza-CR but with alleviated side effects, suggesting that knockdown of DNMT1 might be potential strategy for the treatment of lung cancer with better tolerability.

Methylation status and protein expression of RASSF1A in endometriosis.

Ras association domain family 1A (RASSF1A) gene inactivation by promoter hypermethylation is a common event in the development of a variety of types of human cancer. Accumulated evidence has demonstrated that DNA methylation serve a critical role in the pathogenesis of endometriosis. The aim of the present study was to analyze the methylation status and protein expression of RASSF1A in endometriosis (EMS). The ectopic and corresponding eutopic endometrium tissues were collected from 45 women with EMS (EMS group) and normal endometrium tissues from 20 women without EMS (control group). The methylation status of RASSF1A was examined by methylation specific polymerase chain reaction (MSP). Immunohistochemistry was performed to measure RASSF1A protein level in endometrium tissues. In normal endometrium samples, RASSF1A protein expression was significantly higher at the secretory phase than the proliferative phase. RASSF1A protein expression in the ectopic endometrium tissues and eutopic endometrium tissues were significantly reduced than in normal endometrium (P<0.05). The frequency of aberrant methylation of RASSF1A was 55.56% in ectopic endometrium and 33.33% in paired eutopic endometrium, whereas such methylation was not detected in normal endometrium. Moreover, RASSF1A promoter hypermethylation was frequently associated with reduced expression of RASSF1A, and was common in advanced stage in ectopic endometrium of EMS. Epigenetic inactivation of RASSF1A through aberrant promoter methylation may be important in the formation and progression of EMS, and assessment of RASSF1A methylation status in eutopic endometrium may be a potentially useful biomarker to enhance the early detection of EMS.

Meta-analysis of the Association between RASSF1A Gene Promoter Methylation and Non-small Cell Lung Cancer

背景与目的肿瘤发生、发展过程中，抑癌基因启动子区域CpG岛异常甲基化起着重要的作用。已有研究显示RAS相关区域家族1A(Ras association domain family 1A, RASSF1A)基因，作为一个抑癌基因其启动子区域甲基化与非小细胞肺癌(non-small cell lung cancer, NSCLC)的发生发展密切相关。在NSCLC患者癌组织中RASSF1A基因启动子往往表现为异常高甲基化。本研究采用meta分析的方法探讨RASSF1A基因启动子甲基化与NSCLC发生之间的关系。方法通过检索Medline、EMBASE、CNKI和万方数据库，按照已拟定的纳入与剔除标准筛选收集公开发表的关于RASSF1A基因启动子甲基化与NSCLC相关性的研究。以比值比(odds ratio, OR)和95%置信区间(confidence interval, CI)为效应指标，分析RASSF1A基因启动子甲基化与NSCLC的关系。结果共有23篇文献纳入本研究，RASSF1A基因启动子甲基化率在NSCLC患者肺部组织和对照组中分别为41.50%(95%CI: 34%-49%)和5.58%(95%CI: 2%-9%)，meta分析显示肿瘤组织中的甲基化率高于对照组(OR=8.72, 95%CI:4.88-15.58, P < 0.05)；亚组分析显示:肿瘤组织中的甲基化频率高于血浆(OR=10.99, 95%CI:2.48-48.68)和正常对照组织(OR=8.74, 95%CI: 4.39-17.41)。结论NSCLC患者肺癌组织中RASSF1A基因启动子甲基化率高于对照组，组织中RASSF1A基因启动子甲基化率对肺癌的发生更具影响，RASSSF1A甲基化可能与肺癌的发生存在相关并可作为肺癌诊断的潜在标志物。

Ras association domain family 1A gene promoter methylation status and clinical significance in breast cancer

Objective To investigate the role of ras association domain family 1A(RASSF1A)promoter hypermethylation in the occurrence and development of breast cancer. Methods We analyzed the methylation status of RASSF1A promoter by methylation specific polymerase chain reaction(MS-PCR)of 44 formalin-fixed paraffin-embedded slides of sporadic primary breast cancer, collected corresponding clinicopathological data,and analyzed the relationship between methylation status and clinicopathological data. Results We found 25 out of 44 cases of breast cancer had hypermethylation with the occurrence rate being 56.8%.In this study, we found that methylation status of RASSF1A promoter was significantly associated with age of patients with breast cancer(P<0.05),but not with menarche age and number of births. There was also no significant correlation between RASSF1A promoter methylation and tumor size, lymph node metastasis, TNM stage and estrogen receptor(ER), progesterone receptor(PR)manifestation status. Conclusion Our findings demonstrated that RASSF1A promoter methylation was a frequent event in breast cancer and plays an important role in the cancerogenesis. Key words: Breast cancer; Ras association domain family 1A gene; Promoter; Hypermethylation

Association of RASSF1A promoter methylation with gastric cancer risk: a meta-analysis

Ras-association domain family 1A (RASSF1A), a candidate tumor suppressor gene, is frequently silenced and inactivated by hypermethylation of its promoter region in several human tumors. However, the association between RASSF1A promoter methylation and gastric cancer risk remains conflicting. The aim of this study was to assess the association of RASSF1A promoter methylation with gastric cancer risk by a comprehensive meta-analysis. Relevant studies were identified by searches of PubMed and Web of Science databases with no restrictions. Combined odds ratio (OR) and 95 % confidence interval (CI) were used to assess the strength of the association between RASSF1A promoter methylation and gastric cancer risk. A chi-square-based Q test and sensitivity analyses were performed to test the between-study heterogeneity and the contributions of single studies to the final results, respectively. Funnel plots were carried out to evaluate publication bias. Overall, a significant association was observed between RASSF1A promoter methylation and gastric cancer risk (OR, 12.67; 95 % CI, 8.12-19.78; p < 0.001) with no between-study heterogeneity. Subgroup analyses further revealed that gastric cancer risk was increased for individuals carrying the methylated RASSF1A compared with those with unmethylated RASSF1A. In addition, no publication bias was detected in the overall and subgroup analyses. This study identified a strong association between RASSF1A promoter methylation and risk of gastric cancer and highlighted a promising potential for RASSF1A promoter methylation in gastric cancer risk prediction.

Quantitative promoter hypermethylation analysis of RASSF1A in lung cancer: Comparison with methylation-specific PCR technique and clinical significance

Lung cancer is the major health problem and leading cause of cancer-related deaths worldwide owing to late diagnosis and poor prognosis. Aberrant promoter methylation is an important mechanism for silencing tumor-suppressor genes in cancer and a promising tool for the development of molecular biomarkers. Ras association domain family 1A (RASSF1A), a pivotal gatekeeper of cell cycle progression, is highly methylated in a wide range of human sporadic tumors, including lung cancer. However, no significant prognostic implications have been observed in most studies qualitatively analyzed by methylation-specific PCR (MSP). We found that the RASSF1A promoter was aberrantly methylated in 44.7 and 37.4% of the tumors by pyrosequencing (PS) and MSP methods, respectively. RASSF1A methylation evaluated by the two methods was more frequent in ever-smokers and tumors with TP53 mutation than in never-smokers and tumors without TP53 mutation, respectively. Univariate and multivariate analyses revealed that strong methylation was an unfavorable prognostic factor with stage I (adjusted HR, 2.25; 95% CI 1.03-4.90; P=0.003) and squamous cell carcinoma patients (adjusted HR=2.25, 95% CI 1.03-4.90, P=0.042). Taken together, these results suggested that quantitative PS could gain wider applications in clinical samples as a promising method for early detection screening and prognosis compared with MSP.

Clinicopathological significance of RASSF1A reduced expression and hypermethylation in hepatocellular carcinoma

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. 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. 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, alpha-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). 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.

Prognostic significance of DAPK and RASSF1A promoter hypermethylation in non-small cell lung cancer (NSCLC).

The epigenetic inactivation of tumor suppressor genes may play an important role in the development and progression of many cancer types, including lung cancer. Therefore, we investigated the association between the aberrant promoter methylation of 2 genes: the Death-Associated Protein Kinase (DAPK) and the Ras Association Domain Family 1A (RASSF1A) by using methylation-specific PCR, and the clinicopathological features and prognosis in 70 radically resected non-small cell lung cancers (NSCLCs). Hypermethylation of the DAPK and RASSF1A promoters was found in 24 (34%), and in 18 (26%) tumor DNA samples, respectively. Regarding different clinicopathological features of NSCLCs, the DAPK promoter methylation was more frequently observed in squamous cell carcinoma (46%) than in adenocarcinoma (25%) and large cell carcinoma (22%), but there were no significant statistical differences (p=0.3). On the other hand, a statistically significant trend was observed between the RASSF1A methylation and a histological type of tumor (p=0.06). 45% of adenocarcinoma tumors showed RASSF1A promoter methylation in comparison to 17% of squamous cell carcinomas and 22% of large cell carcinomas. When both markers were analyzed according to the tumor-node-metastasis (TNM) staging system, no statistically significant differences were observed between stage I, II and IIIa, and the DAPK (p=0.2) and RASSF1A methylation (p=0.1). In comparison, when stage I and II were grouped together and considered vs. stage IIIa, a significant association between RASSF1A methylation and the TNM was found (p=0.03). The group of patients with tumors showing DAPK promoter methylation had significantly poorer overall survival rates (p=0.02) than the patients with tumors that did not show DAPK promoter methylation. However, the association between the RASSF1A promoter methylation status and the overall survival rates was not statistically significant (p=0.48). In conclusion, this paper supports the importance of epigenetic gene regulation in lung cancer progression and prognosis.

Frequent hypermethylation of RASSF1A tumour suppressor gene promoter and presence of Merkel cell polyomavirus in small cell lung cancer

In small cell lung cancer (SCLC), hypermethylation of the tumour suppressor Ras association domain family 1A ( RASSF1A) is frequent. It is associated with SV40 polyomaviral infection in other tumours. Merkel cell polyomavirus (MCPyV) infection has been reported in Merkel cell carcinoma (MCC), a neuroendocrine carcinoma with biological similarity to SCLC. In our study, we investigated polyomavirus infection (SV40 and MCPyV) and promoter hypermethylation of the tumour suppressors RASSF1A and p16 in 18 SCLCs (14 primaries and 4 regional lymph node metastases) and 18 blood control samples. MCPyV was found in 39% (7 of 18) of the tumour tissues but not observed in controls. SV40 was not observed in the tumour tissue. RASSF1A promoter hypermethylation (94%; 17 of 18) was more frequent compared to p16 methylation (56%, 10 of 18). We found no significant correlation between RASSF1A or p16 promoter hypermethylation and infection with the investigated polyoma viruses. Our results show a high frequency of hypermethylation of the RASSF1A promoter and occurrence of MCPyV infection in the tumour tissue of SCLC. These events may contribute to the pathogenesis of SCLC.

RASSF1A promoter methylation and expression analysis in normal and neoplastic kidney indicates a role in early tumorigenesis

BackgroundEpigenetic silencing of the RAS association domain family 1A (RASSF1A) tumor suppressor gene promoter has been demonstrated in renal cell carcinoma (RCC) as a result of promoter hypermethylation. Contradictory results have been reported for RASSF1A methylation in normal kidney, thus it is not clear whether a significant difference between RASSF1A methylation in normal and tumor cells of the kidney exists. Moreover, RASSF1A expression has not been characterized in tumors or normal tissue as yet.ResultsUsing combined bisulfite restriction analysis (COBRA) we compared RASSF1A methylation in 90 paired tissue samples obtained from primary kidney tumors and corresponding normal tissue. Bisulfite sequence analysis was carried out using both pooled amplicons from the tumor and normal tissue groups and subclones obtained from a single tissue pair. Expression of RASSF1A was analyzed by the use of tissue arrays and immunohistochemistry. We found significantly increased methylation in tumor samples (mean methylation, 20%) compared to corresponding normal tissues (mean methylation, 11%; P < 0.001). Densely methylated sequences were found both in pooled and individual sequences of normal tissue. Immunohistochemical analysis revealed a significant reduced expression of RASSF1A in most of the tumor samples. Heterogeneous expression patterns of RASSF1A were detected in a subgroup of histologically normal tubular epithelia.ConclusionOur methylation and expression data support the hypothesis that RASSF1A is involved in early tumorigenesis of renal cell carcinoma.

Hypermethylation of RASSF1A in Human and Rhesus Placentas

The pseudomalignant nature of the placenta prompted us to search for tumor suppressor gene hypermethylation, a phenomenon widely reported in cancer, in the human placenta. Nine tumor suppressor genes were studied. Hypermethylation of the Ras association domain family 1 A (RASSF1A) gene was found in human placentas from all three trimesters of pregnancy but was absent in other fetal tissues. Hypermethylation of Rassf1 was similarly observed in placentas from the rhesus monkey but not the mouse. An inverse relationship between RASSF1A promoter methylation and gene expression was demonstrated by bisulfite sequencing of microdissected placental cells and immunohistochemical staining of placental tissue sections using an anti-RASSF1A antibody. Treatment of choriocarcinoma cell lines, JAR and JEG3, by 5-aza-2'-deoxycytidine and trichostatin A led to reduction in RASSF1A methylation but increased expression. These observations extend the analogy between the primate placenta and malignant tumors to the epigenetic level.

The relationship between RASSF1A aberrant methylation and survival in small sized lung adenocarcinoma

7199 Background: Aberrant methylation of CpG islands in promoter regions of tumor cells is one of the major mechanisms for silencing of tumor suppressor genes. The RAS association domain family 1A (RASSF1A) gene was isolated from the 3p21.3 region homozygously deleted in lung cancer cell lines, and it was shown to be inactivated by hypermethylation of the promoter region in lung cancers. In this study, we investigated the clinicopathological significances of RASSF1A methylation in the development and/or progression of small-sized (less than 2.0cm) lung adenocarcinoma. It is important to identify a marker for high-risk early stage patients who should benefit from new investigational adjuvant therapies. Methods: Surgically resected specimens from 77 cases of small-sized primary lung adenocarcinoma. We determined the frequency of aberrant promoter methylation of the RASSF1A genes in small-sized adenocarcinoma. Aberrant promoter methylation was examined using methylation-specific PCR (MSP). Results: Twenty-five of 77 (32.5%) tumors showed RASSF1A methylation. RASSF1A methylation was dominantly detected in smoker (P &lt; 0.03). There was no significant correlation of RASSF1A methylation with gender, age, T stage, N stage and pathological stage. RASSF1A methylation correlated with adverse survival by univariate analysis (P &lt; 0.005) as well as multivariate analysis (P = 0.0062; RR 4.251; 95% C.I., 1.507–11.993). Furthermore, RASSF1A promoter hypermethylation in resected stage I small-sized lung adenocarcinoma was associated with impaired patient survival (P &lt; 0.01). Conclusions: Aberrant promoter methylation of the RASSF1A was present in 25 of 77 (32.5%) of small-sized lung adenocarcinoma by MSP assay. These results indicated that epigenetic inactivation of RASSF1A plays an important role in the progression of small-sized lung adenocarcinoma, and that RASSF1A hypermethylation appears to be a useful molecular marker for the prognosis of patients with small-sized and stage I lung adenocarcinoma. RASSF1A is a potential tumor suppressor gene that undergoes epigenetic inactivation in lung adenocarcinoma through hypermethylation of its promoter region. RASSF1A methylation was significantly related to unfavorable prognosis in small-sized lung adenocarcinoma. No significant financial relationships to disclose.

High frequency of promoter hypermethylation of RASSF1A in tumor and plasma of patients with hepatocellular carcinoma

To determine the presence of ras association domain family 1A (RASSF1A) promoter methylation in the tumor tissues and plasma of patients with hepatocellular carcinoma (HCC). Methylation-specific polymerase chain reaction was used to detect RASSF1A methylation in DNA extracted from HCC tumors and paired plasma samples of 40 patients. The association of RASSF1A hypermethylation in tumor and plasma DNA of HCC patients with clinicopathological characteristics was also analyzed. RASSF1A promoter hypermethylation was detected in 37 of the 40 HCC tissues (92.5%). Of the paired plasma from the 40 HCC patients, aberrant methylation was detected in 17 (42.5%). No RASSF1A methylation was detected in the plasma in the absence of methylation in the corresponding tumor. The presence of RASSF1A promoter hypermethylation in plasma DNA was found to associate with HCC size of > or =4 cm (P = 0.035). RASSF1A promoter hypermethylation occurred at a high frequency in HCC. The aberrant methylation was also detectable in over 40% of matched plasma. The latter should be evaluated as a screening tool and/or prognosticator of HCC patients.

RASSF1A promoter methylation and 3p21.3 loss of heterozygosity are features of foregut, but not midgut and hindgut, malignant endocrine tumours

The Ras-association domain family 1A (RASSF1A) tumour suppressor gene is inactivated in a variety of solid tumours, usually by epigenetic silencing of the promoter and/or allelic loss of its locus at 3p21.3. RASSF1A induces cell cycle arrest through inhibition of cyclin D1 accumulation. In this work, 62 endocrine tumours from different sites in the gut were investigated for methylation of the RASSF1A promoter using the polymerase chain reaction, the presence of 3p21.3 deletions by loss of heterozygosity analysis, and cyclin D1 expression by immunohistochemistry. Methylation was found in 20/62 (32%) cases and was restricted to foregut tumours; deletion at 3p21.3 was found in 15/58 (26%) informative cases and restricted to malignant foregut tumours; cyclin D1 hyper-expression was found in 31/58 (53%) cases and correlated with RASSF1A methylation. Our data suggest that RASSF1A is involved in the development of endocrine tumours derived from the foregut only, and that the presence of both RASSF1A methylation and 3p21.3 deletion is associated with malignancy. These results may provide a rationale for foregut-targeted therapy for aggressive endocrine carcinomas entailing the use of demethylating agents.

Epigenetic inactivation of RASSF1A candidate tumor suppressor gene at 3p21.3 in brain tumors

The human Ras association domain family 1A (RASSF1A) gene, recently isolated from the lung and breast tumor suppressor locus 3p21.3, is highly methylated in primary lung, breast, nasopharyngeal and other tumors, and re-expression of RASSF1A suppresses the growth of several types of cancer cells. Epigenetic inactivation of RASSF1A by promoter hypermethylation is also important in the development of several human cancers. The methylation status of the promoter region of RASSF1A was analysed in primary brain tumors and glioma cell lines by methylation-specific polymerase chain reaction. In primary brain tumors, 25 of 46 (54.3%) gliomas and five of five (100%) medulloblastomas showed RASSF1A methylation. In benign tumors, only one of 10 (10%) schwannomas and two of 12 (16.7%) meningiomas showed RASSF1A methylation. The RASSF1A promoter region was methylated in all four glioma cell lines. RASSF1A was re-expressed in all methylated cell lines after treatment with the demethylating agent 5-aza-2'-deoxycytidine. Methylation of the promoter CpG islands of the RASSF1A may play an important role in the pathogenesis of glioma and medulloblastoma.