Research IndicatorsGraph generated 01 September 2019 using data from PubMed using criteria.
Mouse over the terms for more detail; many indicate links which you can click for dedicated pages about the topic. Tag cloud generated 01 September, 2019 using data from PubMed, MeSH and CancerIndex
Specific Cancers (5)
Data table showing topics related to specific cancers and associated disorders. Scope includes mutations and abnormal protein expression.
Note: list is not exhaustive. Number of papers are based on searches of PubMed (click on topic title for arbitrary criteria used).
OMIM, Johns Hopkin University
Referenced article focusing on the relationship between phenotype and genotype.
International Cancer Genome Consortium.
Summary of gene and mutations by cancer type from ICGC
Cancer Genome Anatomy Project, NCI
COSMIC, Sanger Institute
Somatic mutation information and related details
GEO Profiles, NCBI
Search the gene expression profiles from curated DataSets in the Gene Expression Omnibus (GEO) repository.
Latest Publications: RASSF1 (cancer-related)
Bouras E, Karakioulaki M, Bougioukas KI, et al.Gene promoter methylation and cancer: An umbrella review.
Gene. 2019; 710:333-340 [PubMed
] Related Publications
Gene promoter methylation is a common epigenetic event, taking place in the early phase of tumorigenesis, which has a great potential as a diagnostic and prognostic cancer biomarker. In this umbrella review, we provide an overview on the association between gene-promoter methylation of protein-coding genes and cancer risk based on currently available meta-analyses data on gene promoter methylation. We searched MEDLINE via PubMed and the Cochrane Database of Systematic Reviews for meta-analyses that examine the association between gene-promoter methylation and cancer, published until January 2019 in English. We used AMSTAR to assess the quality of the included studies and applied a set of pre-specified criteria to evaluate the magnitude of each association. We provide a comprehensive overview of 80 unique combinations between 22 different genes and 18 cancer outcomes, all of which indicated a positive association between promoter hypermethylation and cancer. In total, the 70 meta-analyses produced significant results under a random-effects model with odds ratios that ranged from 1.94 to 26.60, with the summary effect being in favor of the unmethylated group in all cases. Three of the strong evidence associations involve RASSF1 methylation on bladder cancer risk (OR = 18.46; 95% CI: 12.69-26.85; I
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.
Keller M, Dubois F, Teulier S, et al.NDR2 kinase contributes to cell invasion and cytokinesis defects induced by the inactivation of RASSF1A tumor-suppressor gene in lung cancer cells.
J Exp Clin Cancer Res. 2019; 38(1):158 [PubMed
] Free Access to Full Article Related Publications
BACKGROUND: RASSF1A, a tumor suppressor gene, is frequently inactivated in lung cancer leading to a YAP-dependent epithelial-mesenchymal transition (EMT). Such effects are partly due to the inactivation of the anti-migratory RhoB GTPase via the inhibitory phosphorylation of GEF-H1, the GDP/GTP exchange factor for RhoB. However, the kinase responsible for RhoB/GEF-H1 inactivation in RASSF1A-depleted cells remained unknown.
METHODS: NDR1/2 inactivation by siRNA or shRNA effects on epithelial-mesenchymal transition, invasion, xenograft formation and growth in SCID-/- Beige mice, apoptosis, proliferation, cytokinesis, YAP/TAZ activation were investigated upon RASSF1A loss in human bronchial epithelial cells (HBEC).
RESULTS: We demonstrate here that depletion of the YAP-kinases NDR1/2 reverts migration and metastatic properties upon RASSF1A loss in HBEC. We show that NDR2 interacts directly with GEF-H1 (which contains the NDR phosphorylation consensus motif HXRXXS/T), leading to GEF-H1 phosphorylation. We further report that the RASSF1A/NDR2/GEF-H1/RhoB/YAP axis is involved in proper cytokinesis in human bronchial cells, since chromosome proper segregation are NDR-dependent upon RASSF1A or GEF-H1 loss in HBEC.
CONCLUSION: To summarize, our data support a model in which, upon RASSF1A silencing, NDR2 gets activated, phosphorylates and inactivates GEF-H1, leading to RhoB inactivation. This cascade induced by RASSF1A loss in bronchial cells is responsible for metastasis properties, YAP activation and cytokinesis defects.
Naturally-occurring mixtures of phytochemicals present in plant foods are proposed to possess tumor-suppressive activities. In this work, we aimed to evaluate the antitumor effects of
Nguyen QN, Vuong LD, Truong VL, et al.Genetic and epigenetic alterations of the EGFR and mutually independent association with BRCA1, MGMT, and RASSF1A methylations in Vietnamese lung adenocarcinomas.
Pathol Res Pract. 2019; 215(5):885-892 [PubMed
] Related Publications
Genetic and epigenetic alterations importantly contribute to the pathogenesis of lung cancer. In the study, we measured the frequency and distribution of molecular abnormalities of EGFR as well as the aberrant promoter methylations of BRCA1, MGMT, MLH1, and RASSF1A in Vietnamese lung adenocarcinomas. We investigated the association between genetic and epigenetic alteration, and between each abnormality with clinicopathologic parameters. Somatic EGFR mutation that was found in 49/139 (35.3%) lung adenocarcinomas showed a significant association with young age, female gender, and non-smokers. EGFR overexpression was identified in 82 tumors (59.0%) and statistical relationships with EGFR or BRCA1 methylation but not EGFR mutation. In addition, EGFR, BRCA1, MGMT, MLH1, and RASSF1A methylations were found in 33 (23.7%), 41 (29.5%), 46 (33.1%), 28 (20.1%), and 41 (29.5%) cases of a total of 139 lung adenocarcinomas, respectively. The RASSF1A methylation was found to be linked to the smoking habit. Methylations in MGMT and RASSF1A were also found to correlate with metastasis status. Furthermore, the distribution of EGFR mutation and that of BRCA1, MGMT or RASSF1A methylation were significantly exclusive in lung adenocarcinomas. The main finding of our study demonstrate that epigenetic abnormalities might play a critical role for the lung tumorigenesis in patients with smoking history and metastasis, and partly affect the predictive value of EGFR mutations through blocking expression due to promoter EGFR hypermethylation. Mutually exclusive distribution of genetic and epigenetic alterations reflects differently biological characteristics in the etiology of lung adenocarcinomas.
Pasha HF, Mohamed RH, Radwan MIRASSF1A and SOCS1 genes methylation status as a noninvasive marker for hepatocellular carcinoma.
Cancer Biomark. 2019; 24(2):241-247 [PubMed
] Related Publications
BACKGROUND: DNA methylation status is one of the most prevalent molecular alterations in human cancers. Identification of powerful diagnostic and prognostic biomarkers for hepatocellular carcinoma (HCC) without a biopsy is urgently required.
OBJECTIVE: The purpose of this study was to determine the methylation status of RASSF1A and SOCS-1genes as a non-invasive biomarker for HCC identification and prognosis.
METHODS: Methylation specific-PCR technique was performed to recognize the methylation status of RASSF1A and SOCS-1 genes in 100 patients with HCC, 100 patients with liver cirrhosis (LC) but without HCC were considered as cirrhotic liver control group and 100 healthy control.
RESULTS: Methylation of RASSF1A and SOCS-1 genes were detected in 40% and 38% of HCC patients respectively, 14% and 20% of LC patients respectively. Methylation of SOCS-1 gene in peripheral blood of healthy control was 23%. Methylation of RASSF1A gene was associated with age, tumor size, vascular invasion and α fetoprotein (AFP), while SOCS-1 gene methylation was significantly associated with tumor size and AFP. Furthermore, using RASSF1A/ SOCS-1/ AFP panel improve diagnostic sensitivity for HCC 86% and specificity of 75%.
CONCLUSION: RASSF1A and SOCS1 genes methylation status may play an important role in the process of hepatocarcinogenesis and may be used as diagnostic and prognostic noninvasive biomarkers for HCC when combined with serum AFP.
EBV latent antigen 3C (EBNA3C) is essential for EBV-induced primary B-cell transformation. Infection by EBV induces hypermethylation of a number of tumor suppressor genes, which contributes to the development of human cancers. The Ras association domain family isoform 1A (RASSF1A) is a cellular tumor suppressor, which regulates a broad range of cellular functions, including apoptosis, cell-cycle arrest, mitotic arrest, and migration. However, the expression of RASSF1A is lost in many human cancers by epigenetic silencing. In the present study, we showed that EBNA3C promoted B-cell transformation by specifically suppressing the expression of RASSF1A. EBNA3C directly interacted with RASSF1A and induced RASSF1A degradation via the ubiquitin-proteasome-dependent pathway. SCFSkp2, an E3-ubiquitin ligase, was recruited by EBNA3C to enhance RASSF1A degradation. Moreover, EBNA3C decreased the transcriptional activity of RASSF1A promoter by enhancing its methylation through EBNA3C-mediated modulation of DNMTs expression. EBNA3C also inhibited RASSF1A-mediated cell apoptosis, disrupted RASSF1A-mediated microtubule and chromosomal stability, and promoted cell proliferation by upregulating Cyclin D1 and Cyclin E expression. Our data provides new details, which sheds light on additional mechanisms by which EBNA3C can induce B-cell transformation. This will also facilitate the development of novel therapeutic approaches through targeting of the RASSF1A pathway.
Strzelczyk JK, Krakowczyk Ł, Owczarek AJMethylation status of SFRP1, SFRP2, RASSF1A, RARβ and DAPK1 genes in patients with oral squamous cell carcinoma.
Arch Oral Biol. 2019; 98:265-272 [PubMed
] Related Publications
Our study assessed the methylation status of the SFRP1, SFRP2, RASSF1A, RARβ and DAPK1 genes, which are associated with epigenetic silencing in cancers. In a group of 75 patients with oral squamous cell carcinoma, aberrant methylation was detected using methylation-specific PCR in tumours and matched margins. Our results showed significantly higher methylation frequency in tumours than in surgical margin of SFRP2 (26.6% vs 11.9%, p < 0.05) and DAPK1 (65.3% vs 41.3%, p < 0.01) genes. Moreover, methylation of the SFRP1 and DAPK1 genes was associated with older age. Advanced tumour stages were associated with lower rates of SFRP1 gene methylation. Decreased methylation levels of the SFRP2 and RASSF1A genes were associated with positive N stage. On the contrary, lymph node metastasis were associated with higher methylation rates of RARβ and DAPK1 genes. Patients with a familial history of cancer were associated with more frequently methylated SFRP1, SFRP2 and DAPK1 genes. Hypermethylation of DAPK1 was associated with decreased risk of death in patients. Our results are suggestive, although not conclusive, that some epigenetic changes, especially frequent hypermethylation of SFRP2 and DAPK1 genes, can be useful as potential diagnostic biomarkers of oral cavity cancer. Moreover, estimating the methylation status in surgical margins could become an additional strategy for more accurate treatment methods. Further efforts are needed to identify and validate this finding on a larger patient group and using new advanced methylation testing methods.
Bashanfer SAA, Saleem M, Heidenreich O, et al.Disruption of MAPK1 expression in the ERK signalling pathway and the RUNX1‑RUNX1T1 fusion gene attenuate the differentiation and proliferation and induces the growth arrest in t(8;21) leukaemia cells.
Oncol Rep. 2019; 41(3):2027-2040 [PubMed
] Related Publications
The t(8;21) translocation is one of the most frequent chromosome abnormalities associated with acute myeloid leukaemia (AML). This abberation deregulates numerous molecular pathways including the ERK signalling pathway among others. Therefore, the aim of the present study was to investigate the gene expression patterns following siRNA‑mediated suppression of RUNX1‑RUNX1T1 and MAPK1 in Kasumi‑1 and SKNO‑1 cells and to determine the differentially expressed genes in enriched biological pathways. BeadChip microarray and gene ontology analysis revealed that RUNX1‑RUNX1T1 and MAPK1 suppression reduced the proliferation rate of the t(8;21) cells with deregulated expression of several classical positive regulator genes that are otherwise known to enhance cell proliferation. RUNX1‑RUNX1T1 suppression exerted an anti‑apoptotic effect through the overexpression of BCL2, BIRC3 and CFLAR genes, while MAPK1 suppression induced apopotosis in t(8;21) cells by the apoptotic mitochondrial changes stimulated by the activity of upregulated TP53 and TNFSF10, and downregulated JUN gene. RUNX1‑RUNX1T1 suppression supported myeloid differentiation by the differential expression of CEBPA, CEBPE, ID2, JMJD6, IKZF1, CBFB, KIT and CDK6, while MAPK1 depletion inhibited the differentiation of t(8;21) cells by elevated expression of ADA and downregulation of JUN. RUNX1‑RUNX1T1 and MAPK1 depletion induced cell cycle arrest at the G0/G1 phase. Accumulation of cells in the G1 phase was largely the result of downregulated expression of TBRG4, CCNE2, FOXO4, CDK6, ING4, IL8, MAD2L1 and CCNG2 in the case of RUNX1‑RUNX1T1 depletion and increased expression of RASSF1, FBXO6, DADD45A and P53 in the case of MAPK1 depletion. Taken together, the current results demonstrate that MAPK1 promotes myeloid cell proliferation and differentiation simultaneously by cell cycle progression while suppresing apoptosis.
Tumor suppressor genes (TSGs), including Ten-eleven translocation 1 (TET1), are hypermethylated in hepatocellular carcinoma (HCC). TET1 catalytic domain (TET1-CD) induces genome-wide DNA demethylation to activate TSGs, but so far, anticancer effects of TET1-CD are unclear. Here we showed that after HCC cells were transiently transfected with TET1-CD, the methylation levels of TSGs, namely APC, p16, RASSF1A, SOCS1 and TET1, were distinctly reduced, and their mRNA levels were significantly increased and HCC cells proliferation, migration and invasion were suppressed, but the methylation and mRNA levels of oncogenes, namely C-myc, Bmi1, EMS1, Kpna2 and c-fos, were not significantly change. Strikingly, HCC subcutaneous xenografts in nude mice remained to be significantly repressed even 54 days after transient transfection of TET1-CD. So, transient transfection of TET1-CD may be a great advance in HCC treatment due to its activation of multiple TSGs and persistent anticancer effects.
Chen T, Yang Z, Liu C, et al.Circ_0078767 suppresses non-small-cell lung cancer by protecting RASSF1A expression via sponging miR-330-3p.
Cell Prolif. 2019; 52(2):e12548 [PubMed
] Related Publications
OBJECTIVES: This study was designed to investigate the role of circ_0078767/miR-330-3p/RASSF1A in non-small-cell lung cancer (NSCLC). Bioinformatic analysis was performed to screen for the differentially expressed genes in NSCLC tissues from adjacent lung tissues.
MATERIALS AND METHODS: qRT-PCR was used to detect the RNA expression of genes in cells and tissues, and Western blot was conducted to determine the protein levels of RASSF1A in tissues and cells. A miRanda algorithm was used to predict the targeted relationship among RNAs. A dual-luciferase reporter gene assay was conducted to verify the targeted relationship. Flow cytometry was performed to investigate the effects of circ_0078767/miR-330-3p/RASSF1A on cell cycle progression and apoptosis. A CCK-8 assay was conducted to explore the effects of circ_0078767/miR-330-3p/RASSF1A on cell proliferation. A transwell invasion assay was completed to study the effects of circ_0078767/miR-330-3p/RASSF1A on cell invasion. Lastly, an in vivo assay was conducted to investigate the effects of circ_0078767/miR-330-3p/RASSF1A on tumour development.
RESULTS: Circ_0078767 and RASSF1A were downregulated, while miR-330-3p was upregulated in NSCLC tissues than that in adjacent tissues. miR-330-3p had a binding relationship with circ_0078767 and RASSF1A. The overexpression of circ_0078767 and RASSF1A or the underexpression of miR-330-3p significantly suppressed NSCLC cell viability, cell cycle progression and invasion while also significantly promoting cell apoptosis. Additionally, these modulations significantly suppressed in vivo tumour growth.
CONCLUSIONS: Circ_0078767 could suppress NSCLC progression by inhibiting miR-330-3p, which thereby increased RASSF1 levels.
Yang Z, Qi W, Sun L, et al.DNA methylation analysis of selected genes for the detection of early-stage lung cancer using circulating cell-free DNA.
Adv Clin Exp Med. 2019; 28(3):361-366 [PubMed
] Related Publications
BACKGROUND: Lung cancer is still the deadliest cancer in the world, but early diagnosis cannot be achieved because of the limitations of diagnostic methods. DNA methylation has been proven to be a potentially powerful tool for cancer detection and diagnosis over the past decade.
OBJECTIVES: We explored whether free DNA methylation in plasma can be a reliable biomarker for noninvasive lung cancer detection.
MATERIAL AND METHODS: We detected the methylation of 8 genes in plasma-free DNA of patients with pulmonary space-occupying lesions using real-time quantitative methylation-specific polymerase chain reaction (QMSP). Among the 50 selected patients, 39 were confirmed using pathological analysis as having early lung cancer and 11 had an inflammatory pseudotumor.
RESULTS: The QMSP detection showed that the methylation levels of 8 genes in the patients were significantly higher than in the non-lung cancer group. The methylation level of CALCA was the highest and the methylation level of HOXA9 was the lowest. Methylation of RASSF1A, CDKN2A and DLEC1 occured only in lung cancer patients, while methylation of CALCA, CDH13, PITX2, HOXA9, and WT1 occured not only in lung cancer patients, but also in non-lung cancers. The specificity reached 95~100%, whether for a single gene or overall, but the sensitivity was relatively low for each gene. The sensitivity can reach 72% if the methylation of any of the 8 genes is positive and the overall specificity was 91%. The positive and negative predictive values were 96% and 60%, respectively.
CONCLUSIONS: Quantitative detection of DNA methylation in plasma is a potential method for early diagnosis of lung cancer.
Up to 60% of untreated atypical hyperplastic endometrium will develop into endometrial carcinoma (EC), and for those who underwent a hysterectomy a coexisting EC is found in up to 50%. Gene promoter methylation might be related to the EC development. The aim of this study is to determine changes in gene promoter profiles in normal endometrium, atypical hyperplasia (AH) and EC in relation to K-Ras mutations. A retrospective study was conducted in patients diagnosed with endometrial hyperplasia with and without subsequent EC. Promoter methylation of APC, hMLh1, O6-MGMT, P14, P16, RASSF1, RUNX3 was analysed on pre-operative biopsies, and correlated to the final histological diagnosis, and related to the presence of K-Ras mutations. In the study cohort (n=98), differences in promoter methylation were observed for hMLH1, O6-MGMT, and P16. Promoter methylation of hMLH1 and O6-MGMT gradually increased from histologically normal endometrium to AH to EC; 27.3, 36.4% and 38.0% for hMLH1 and 8.3%, 18.2% and 31.4% for O6-MGMT, respectively. P16 promoter methylation was significantly different in AH (7.7%) compared to EC (38%). K-Ras mutations were observed in 12.1% of AH, and in 19.6% of EC cases. No association of K-Ras mutation with promoter methylation of any of the tested genes was found. In conclusion, hMLH1 and O6-MGMT promoter methylation are frequently present in AH, and thus considered to be early events in the carcinogenesis of EC, whereas P16 promoter methylation was mainly present in EC, and not in precursor lesions supporting a late event in the carcinogenesis.
Costa AL, Moreira-Barbosa C, Lobo J, et al.DNA methylation profiling as a tool for testicular germ cell tumors subtyping.
Epigenomics. 2018; 10(12):1511-1523 [PubMed
] Related Publications
AIM: Assess differential patterns of selected five genes' promoter methylation among testicular germ cell tumors (TGCT) subtypes.
MATERIALS & METHODS: CRIPTO, HOXA9, MGMT, RASSF1A and SCGB3A1 promoter methylation levels were evaluated by quantitative methylation-specific PCR in 161 TGCT and 16 controls. Associations between clinicopathological parameters and promoter methylation levels were assessed, and receiver operating characteristics curve analysis was performed.
RESULTS: Promoter methylation of CRIPTO/HOXA9/SCGB3A1 panel and RASSF1A best discriminated between controls and nonseminomatous tumors or seminomas, respectively, whereas HOXA9/RASSF1A panel displayed the best discriminative performance between nonseminomatous tumor and seminomas. Significant differences in CRIPTO, MGMT and RASSF1A methylation levels were depicted between pure forms and matched mixed components of seminomas and embryonal carcinoma. HOXA9, RASSF1A and SCGB3A1 promoter methylation significantly associated with tumor stage.
CONCLUSION: Different combinations of five genes' promoter methylation levels discriminate among TGCT subtypes. Methylation patterns may also assist in identification of more clinically aggressive tumors.
With no sharp cure, breast cancer still be the major and the most serious life-threatening disease worldwide. Colorectal
is the third most commonly occurring cancer in men and the second most commonly occurring cancer in women. In the
present investigation, colon cancer cells (CaCo-2) and breast cancer cells (MCF-7) were treated with elevated doses
of metformin (MET) for 48h. Cell count was assessed using trypan blue test, and the cytotoxicity was evaluated using
MTT assay. Methylation-specific PCR was performed on the bisulfite-treated DNA against two tumor suppressor genes;
RASSF1A and RB. Results indicated that: in breast cancer, the cell count was decreased significantly (P>0.005) after
being treated with 5, 10, 20, 50, and 100 mM of MET. The elevated concentration had increased reduction percentages
on the MCF-7 cells, as 5 mM and 100 mM have yielded 35% and 93.3% reduction in cell viability, respectively. Colon
cancer cells have responded to the doses of MET differently, as for the 5 mM and the 100 mM, it gave 88% and 60%
reduction in cells viability, respectively. Cytotoxicity assay revealed that 5 mM and 100 mM of MET caused breast
cancer cells to loss 61.53% and 85.16% of its viability, respectively, whereas colon cancer cells have responded to the
5 mM and 100 mM of MET by reducing the cells viability with 96.91% and 96.24%, respectively. No RB promoter
methylation was detected in colon cells, while RASSF1A was partially methylated. In the MCF-7 breast cancer cells,
both RASSF1A and RB were partially methylated.
Promoter DNA methylation, which occurs on cytosine nucleotides across CpG islands, results in gene silencing and represents a major epigenetic alteration in human cancer. Methylation-specific PCR can amplify these modifications as markers in cancer cells. In the present work, we rigorously review the published literatures describing DNA methylation in the promoters of critical tumor suppressor genes; detection of promoter DNA methylation in various body fluids permits early detection of cancer cells during perioperative courses of clinical treatment. The latest whole-genome comprehensive explorations identified excellent epigenetic biomarkers that could be detected at high frequency with high specificity; these biomarkers, which are designated highly relevant methylation genes (HRMG), permit the discrimination of tumor tissues from the corresponding normal tissues; these markers are also associated with unique cancer phenotypes, including dismal prognosis. In humans, HRMG include the CDO1, GSHR, RASSF1 and SFRP1 genes, with these markers permitting discrimination depending on the organs tested. The combination of several HRMG increased the early detection of cancer and exhibited reliable surveillance potential in human body fluids. Cancer clinics using such epigenetic biomarkers are entering a new era of enhanced decision-making with the potential for improved cancer prognosis.
Cell stiffness is a potential biomarker for monitoring cellular transformation, metastasis, and drug resistance development. Environmental factors relayed into the cell may result in formation of inheritable markers (e.g., DNA methylation), which provide selectable advantages (e.g., tumor development-favoring changes in cell stiffness). We previously demonstrated that targeted methylation of two tumor suppressor genes, hypermethylated in cancer 1 (
Bure I, Geer S, Knopf J, et al.Long noncoding RNA HOTAIR is upregulated in an aggressive subgroup of gastrointestinal stromal tumors (GIST) and mediates the establishment of gene-specific DNA methylation patterns.
Genes Chromosomes Cancer. 2018; 57(11):584-597 [PubMed
] Related Publications
Aberrant alterations of DNA methylation are common events in oncogenesis. The origin of cancer-associated epigenetic defects is of interest for mechanistic understanding of malignant transformation and-in the long run-therapeutic modulation of DNA methylation in a locus-specific manner. Given the ability of certain long noncoding RNAs to operate as an interface between DNA and the epigenetic modification machinery which can interact with DNA methyltransferases, we hypothesized-considering HOTAIR as an example-that this transcript may contribute to gene specificity of DNA methylation. Using gastrointestinal stromal tumors (GISTs, n = 67) as a model, we confirmed upregulation of HOTAIR in tumors with high risk of recurrence and showed high abundance of the transcript in GIST cell lines. HOTAIR knockdown in GIST-T1 cells triggered transcriptional response of genes involved in the organization and disassembly of the extracellular matrix and, notably, induced global locus-specific alterations of DNA methylation patterns. Hypomethylation was induced at a total of 507 CpG sites, whereas 382 CpG dinucleotides underwent gain of methylation upon HOTAIR depletion. Importantly, orchestrated gain or loss of methylation at multiple individual CpG sites was shown for cancer-related DPP4, RASSF1, ALDH1A3, and other targets. Collectively, our data indicate that HOTAIR enables target specificity of DNA methylation in GIST and is capable of dual (hypo- and hypermethylation) regulation by a yet to be defined mechanism. The results further suggest the feasibility of manipulating DNA methylation in a targeted manner and are of interest in the context of epigenetic cancer therapy.
Le AV, Szaumkessel M, Tan TZ, et al.DNA Methylation Profiling of Breast Cancer Cell Lines along the Epithelial Mesenchymal Spectrum-Implications for the Choice of Circulating Tumour DNA Methylation Markers.
Int J Mol Sci. 2018; 19(9) [PubMed
] Free Access to Full Article Related Publications
(1) Background: Epithelial⁻mesenchymal plasticity (EMP) is a dynamic process whereby epithelial carcinoma cells reversibly acquire morphological and invasive characteristics typical of mesenchymal cells. Identifying the methylation differences between epithelial and mesenchymal states may assist in the identification of optimal DNA methylation biomarkers for the blood-based monitoring of cancer. (2) Methods: Methylation-sensitive high-resolution melting (MS-HRM) was used to examine the promoter methylation status of a panel of established and novel markers in a range of breast cancer cell lines spanning the epithelial⁻mesenchymal spectrum. Pyrosequencing was used to validate the MS-HRM results. (3) Results:
BACKGROUND: Urine poses an attractive non-invasive means for obtaining liquid biopsies for oncological diagnostics. Especially molecular analysis on urinary DNA is a rapid growing field. However, optimal and practical storage conditions that result in preservation of urinary DNA, and in particular hypermethylated DNA (hmDNA), are yet to be determined.
AIM: To determine the most optimal and practical conditions for urine storage that result in adequate preservation of DNA for hmDNA analysis.
METHODS: DNA yield for use in methylation analysis was determined by quantitative methylation specific PCR (qMSP) targeting the ACTB and RASSF1A genes on bisulfite modified DNA. First, DNA yield (ACTB qMSP) was determined in a pilot study on urine samples of healthy volunteers using two preservatives (Ethylenediaminetetraacetic acid (EDTA) and Urine Conditioning Buffer, Zymo Research) at four different temperatures (room temperature (RT), 4°C, -20°C, -80°C) for four time periods (1, 2, 7, 28 days). Next, hmDNA levels (RASSF1A qMSP) in stored urine samples of patients suffering from bladder cancer (n = 10) or non-small cell lung cancer (NSCLC; n = 10) were measured at day 0 and 7 upon storage with and without the addition of 40mM EDTA and/or 20 μl/ml Penicillin Streptomycin (PenStrep) at RT and 4°C.
RESULTS: In the pilot study, DNA for methylation analysis was only maintained at RT upon addition of preserving agents. In urine stored at 4°C for a period of 7 days or more, the addition of either preserving agent yielded a slightly better preservation of DNA. When urine was stored at -20 °C or -80 °C for up to 28 days, DNA was retained irrespective of the addition of preserving agents. In bladder cancer and NSCLC samples stored at RT loss of DNA was significantly less if EDTA was added compared to no preserving agents (p<0.001). Addition of PenStrep did not affect DNA preservation (p>0.99). Upon storage at 4°C, no difference in DNA preservation was found after the addition of preserving agents (p = 0.18). The preservation of methylated DNA (RASSF1A) was strongly correlated to that of unmethylated DNA (ACTB) in most cases, except when PCR values became inaccurate.
CONCLUSIONS: Addition of EDTA offers an inexpensive preserving agent for urine storage at RT up to seven days allowing for reliable hmDNA analysis. To avoid bacterial overgrowth PenStrep can be added without negatively affecting DNA preservation.
MicroRNA (miR)‑181a is a member of the miR‑181 family that serves a key role in the pathogenesis of various cancer types. The present study aimed to investigate the interaction between miR‑181a and Ras association domain family protein1 isoform A (RASSF1A), and their roles in gastric carcinogenesis. The interaction between miR‑181a and RASSF1A was assessed in cell lines and cancer tissues. The direct binding of miR‑181a and RASSF1A was identified using a luciferase reporting gene system. The effects of miR‑181a and RASSF1A on gastric cancer cell growth, cell cycle and apoptosis were assessed with a Cell Counting Kit‑8 assay and flow cytometry. The effects of miR‑181a on cell division cycle 25A (CDC25A), cyclin A2, cyclin D1, p21, Bcl‑2‑associated X protein (Bax) and B‑cell lymphoma‑2 (Bcl‑2) protein levels were assessed in gastric cancer cell lines. miR‑181a directly interacted with the 3'‑untranslated region of RASSF1A and downregulated RASSF1A protein expression. In tissues from patients with gastric cancer, the miR‑181a level was significantly higher in the tumor tissues and was negatively correlated with the RASSF1A protein level. RASSF1A suppressed gastric cancer cell proliferation and G1/S transition, and promoted apoptosis; whereas miR‑181a promoted cancer cell proliferation and G1/S transition, and suppressed apoptosis. RASSF1A knockdown attenuated the effects of miR‑181a downregulation on cell proliferation and apoptosis. Furthermore, miR‑181a upregulated CDC25A, cyclin A2 and Bcl‑2, and downregulated Bax protein expression in gastric cancer cell lines. These data indicate that miR‑181a promotes gastric carcinogenesis, possibly through a direct interaction with RASSF1A.
Yi M, Wang W, Chen S, et al.Dual-functionality of RASSF1A overexpression in A375 cells is mediated by activation of IL-6/STAT3 regulatory loop.
Mol Biol Rep. 2018; 45(5):1277-1287 [PubMed
] Related Publications
The RASSF1A, a microtubule associated protein, is a well-known tumor suppressor silenced in various cancer via promoter hypermethylation. RASSF1A is implicated in the regulation of cellular proliferation and apoptosis. However, its role in melanoma A375 cells invasion and metastasis remain unclear. Here, we report an unusual dual function role of ectopic RASSF1A in A375 cells. RASSF1A suppressed A375 cells proliferation but enhanced cells migration, invasiveness and metastatic potential in vivo. We demonstrated RASSF1A simultaneously up-regulated p21 and vimentin expression in A375 cells. Increase of vimentin expression contributes to RASSF1A mediated enhancement of cells mobility and invasion. Transcriptome assay unclosed that RASSF1A promoted IL-6 expression in A375 cells, which in turn activate JAK2/STAT3 signaling. Treatment with recombinant IL-6 enhanced both p21 and vimentin protein level in the empty vector transfected A375 cells to similar level as RASSF1A expressing cells. In contrast, knockdown IL-6 expression by siRNAs decreased p21 and vimentin level in RASSF1A expressing cells. Blockade of JAK2/STAT3 signaling by use of JAK2 inhibitor WP1066 led to decrease of IL-6, p21 and vimentin protein in RASSF1A expressing cells. Our findings unclosed a unusual dual functionality of ectopic RASSF1A overexpression in A375 cells by regulating IL-6/STAT3 regulatory loop, suggesting it should be cautious about the safety of RASSF1A-based gene therapy.
Objective: This study investigated the DNA promoter methylation profiles of BRCA1, RASSF1A and GSTP1 genes,
both individually and in an integrative manner in order to clarify their correlation with clinicopathological parameters of
breast cancer from Vietnamese patients, and establish new potential integrative methylation biomarkers for breast cancer
detection. Material and methods: The methylation frequencies of BRCA1, RASSF1A and GSTP1 were analyzed by
methylation specific polymerase chain reaction (MSP) in 70 specimens of breast carcinomas and 79 pairs of tumor and
matched adjacent normal tissues from breast cancer patients. Results: All the three analyzed genes showed a concordance
concerning their promoter methylation in tumor and adjacent normal tissue. The methylation of BRCA1, RASSF1A
and GSTP1 was found in 58.23 %, 74.68 % and 59.49 % of tumor tissues and 51.90 %, 63.29 % and 35.44 % of
corresponding adjacent tissues, respectively. When each gene was assessed individually, only the methylation of
GSTP1 was significantly associated with tumor tissues (p=0.003). However, the methylation frequency of at least one of
the three genes and the methylation frequency of all the three genes both showed significant association with tumor
(p=0.008 and p=0.04, respectively). The methylation of BRCA1 was found to be significantly associated with tumor
grade (p=0.01). Conclusion: This study emphasized that the panel of the three genes BRCA1, RASSF1A and GSTP1
can be further developed as potential biomarkers in diagnosis and classification of breast cancer in Vietnamese women.
Blanchard TG, Lapidus R, Banerjee V, et al.Upregulation of RASSF1A in Colon Cancer by Suppression of Angiogenesis Signaling and Akt Activation.
Cell Physiol Biochem. 2018; 48(3):1259-1273 [PubMed
] Related Publications
BACKGROUND/AIMS: Silencing of tumor suppressor genes (TSGs) and promotion of angiogenesis are associated with tumor development and metastasis. However, little is known if angiogenic molecules directly control TSGs and vice versa.
METHODS: A regulatory link between angiogenesis and down regulation of TSGs was evaluated using an anti-cancer agent, andrographolide (AGP) in cancer cells, mouse xenograft tissues and patient derived organoids through gene/protein expression, gene silencing, and immunohistochemical analyses.
RESULTS: AGP treatment demonstrated significant expression of RASSF1A and PTEN TSGs in colon cancer and other cancer cells, mouse tissues and organoids. Depletion of RASSF1A with siRNA limited cyclin D1 and BAX expression. SiRNA depletion of PTEN, upstream regulator of RASSF1A resulted in a 50% reduction in RASSF1A expression. Histopathological analysis of the AGP treated tumor sections showed significant reduction in vessel size, microvascular density and tumor mitotic index suggesting suppression of angiogenesis. This was corroborated by protein analysis demonstrating significant reductions in angiogenesis signaling pathway molecules VEGF165, FOXM1, and pAkt, but significant elevation of the endogenous angiogenesis inhibitor Tsp-2. Treatment of cells with exogenous VEGF prevented the suppression of angiogenesis signaling by AGP, resulting in sustained expression of pAkt, an upstream down-regulator of RASSF1A. RASSF1A expression remained low in VEGF treated cells despite the addition of AGP.
CONCLUSION: Our results demonstrate for the first time that AGP induces RASSF1A expression in colon cancer cells and is dependent on angiogenesis signaling events. Therefore, our research may facilitate novel therapeutic options for advanced colon cancer therapy.
Kozomara Z, Supic G, Krivokuca A, et al.Promoter hypermethylation of p16, BRCA1 and RASSF1A genes in triple-negative breast cancer patients from Serbia.
J BUON. 2018 May-Jun; 23(3):684-691 [PubMed
] Related Publications
PURPOSE: In order to investigate if aberrant promoter methylation of p16, BRCA1 and RASSF1A genes contributes to biological behavior of triple-negative breast cancer (TNBC), marked as the most aggressive phenotype of breast cancer, we compared the hypermethylation pattern between TNBC and ER+PR+Her2- breast cancer.
METHODS: 131 patients with histologically confirmed breast cancers were included - 61 TNBC and 70 ER+PR+Her2- cases. The patients were followed up for 1-87 months (median 78). DNA from tumor tissues was isolated by the salting out procedure. The methylation status was assessed by nested methylation-specific PCR after bisulfite modification of DNA.
RESULTS: The frequency of p16 hypermethylated breast cancer cases was significantly higher in TNBC than in ER+PR+Her2- group (33; 54.1% vs. 20; 28.6%, p=0.00298). Co-methylated p16 and RASSF1A genes were more frequent in the TNBC than in ER+PR+Her2- group (20; 32.8% vs. 10; 14.3%, p=0.0225). The same result was observed when hypermethylated BRCA1 gene was added in the analysis: 12; 19.7% vs. 3; 4.3%, p=0.00791. Although there was significant difference in disease-free survival (DFS) and overall survival (OS) between TNBC and ER+PR+Her2- group, further analysis of co-methylation of p16 and RASSF1A (p16+RASSF1A+) showed that DFS was significantly shorter in the patients with both genes co-methylated in TNBC than in ER+PR+Her-2- group (8/20; 40% vs. 2/10; 20%, p=0.03272).
CONCLUSIONS: The obtained data indicate that hypermethylated p16 and RASSF1A cell-cycle inhibitor genes might be considered as biomarkers for bad prognosis in breast cancer. Hypermethylation of these genes may influence the clinical disease course, distinguishing a particular group of TNBC patients with even more aggressive phenotype.
Ma HS, Wang EL, Xu WF, et al.Overexpression of DNA (Cytosine-5)-Methyltransferase 1 (DNMT1) And DNA (Cytosine-5)-Methyltransferase 3A (DNMT3A) Is Associated with Aggressive Behavior and Hypermethylation of Tumor Suppressor Genes in Human Pituitary Adenomas.
Med Sci Monit. 2018; 24:4841-4850 [PubMed
] Free Access to Full Article Related Publications
BACKGROUND Alteration of DNA methylation of tumor suppressor genes (TSGs) is one of the most consistent epigenetic changes in human cancers. DNMTs play several important roles in DNA methylation and development of cancers. Regarding DNMTs protein expressions, little is known about the clinical significance and correlation with promoter methylation status of TSGs in human pituitary adenomas. MATERIAL AND METHODS We analyzed the protein expression of 3 DNMTs using immunohistochemistry and assessed DNA hypermethylation of RASSF1A, CDH13, CDH1, and CDKN2A (p16) in 63 pituitary adenomas. We examined associations between DNMTs expression and clinicopathological features or promoter methylation status of TSGs. RESULTS Overexpression of DNMTs was detected in pituitary adenomas. Frequencies of DNMT1 overexpression were significantly higher in macroadenomas, invasive tumors, and grade III and IV tumors. DNMT3A was frequently detected in invasive tumors and grade IV tumors. In addition, DNMT1 and DNMT3A were frequently detected in high-methylation tumors. Furthermore, in multivariate logistic regression, the significant association between DNMT1 or DNMT3A and high-methylation status persisted after adjusting for clinicopathological features. CONCLUSIONS Our findings suggested that tumor overexpression of DNMT1 and DNMT3A is associated with tumor aggressive behavior and high-methylation status in pituitary adenomas. Our data support a possible role of DNMT1 and DNMT3A in TSG promoter methylation leading to pituitary adenoma invasion and suggest that inhibition of DNMTs has the potential to become a new therapeutic approach for invasive pituitary adenoma.
Boyne DJ, King WD, Brenner DR, et al.Aerobic exercise and DNA methylation in postmenopausal women: An ancillary analysis of the Alberta Physical Activity and Breast Cancer Prevention (ALPHA) Trial.
PLoS One. 2018; 13(6):e0198641 [PubMed
] Free Access to Full Article Related Publications
Physical activity is associated with a lower risk of breast, colon, and endometrial cancer. Epigenetic mechanisms such as changes in DNA methylation may help to explain these protective effects. We assessed the impact of a one year aerobic exercise intervention on DNA methylation biomarkers believed to play a role in carcinogenesis. The Alberta Physical Activity and Breast Cancer Prevention (ALPHA) Trial was a two-armed randomized controlled trial in 320 healthy, inactive, postmenopausal women with no history of cancer. In an ancillary analysis, frozen blood samples (n = 256) were reassessed for levels of DNA methylation within LINE-1 and Alu repeats as well as within the promoter regions of APC, BRCA1, RASSF1, and hTERT genes. Differences between the exercise and control arm at 12-months, after adjusting for baseline values, were estimated within an intent-to-treat and per-protocol analysis using linear regression. No significant differences in DNA methylation between the exercise and control arms were observed. In an exploratory analysis, we found that the prospective change in estimated VO2max was negatively associated with RASSF1 methylation in a dose-response manner (p-trend = 0.04). A year-long aerobic exercise intervention does not affect LINE-1, Alu, APC, BRCA1, RASSF1, or hTERT methylation in healthy, inactive, postmenopausal women. Changes in DNA methylation within these genomic regions may not mediate the association between physical activity and cancer in healthy postmenopausal women. Additional research is needed to validate our findings with RASSF1 methylation.
TRIAL REGISTRATION: ClinicalTrials.gov NCT00522262.
BACKGROUND: Lung cancer is the major cause of cancer-related deaths worldwide. Differential diagnosis can be difficult, especially when only small samples are available. Epigenetic changes are frequently tissue-specific events in carcinogenesis and hence may serve as diagnostic biomarkers.
MATERIAL AND METHODS: 138 representative formalin-fixed, paraffin-embedded (FFPE) tissues (116 lung cancer cases and 22 benign controls) were used for targeted DNA methylation analysis via pyrosequencing of ten literature-derived methylation markers (APC, CDH1, CDKN2A, EFEMP1, FHIT, L1RE1, MGMT, PTEN, RARB, and RASSF1). Methylation levels were analyzed with the Classification and Regression Tree Algorithm (CART), Conditional Interference Trees (ctree) and ROC. Validation was performed with additional 27 lung cancer cases and 38 benign controls. TCGA data for 282 lung cancer cases was included in the analysis.
RESULTS: CART and ctree analysis identified the combination of L1RE1 and RARB as well as L1RE1 and RASSF1 as independent methylation markers with high discriminative power between tumor and benign tissue (for each combination, 91% specificity and 100% sensitivity). L1RE1 methylation associated significantly with tumor type and grade (p<0.001) with highest methylation in the control group. The opposite was found for RARB (p<0.001). RASSF1 methylation increased with tumor type and grade (p<0.001) with strongest methylation in neuroendocrine tumors (NET).
CONCLUSION: Hypomethylation of L1RE1 is frequent in tumors compared to benign controls and associates with higher grade, whereas increasing methylation of RARB is an independent marker for tumors and higher grade. RASSF1 hypermethylation was frequent in tumors and most prominent in NET making it an auxiliary marker for separation of NSCLC and NET. L1RE1 in combination with either RARB or RASSF1 could function as biomarkers for separating lung cancer and non-cancerous tissue and could be useful for samples of limited size such as biopsies.
Background & objectives: Invasive cervical cancer patients are primarily treated with chemoradiation therapy. The overall and disease-free survival in these patients is variable and depends on the tumoral response apart from the tumour stage. This study was undertaken to assess whether in vivo changes in gene promoter methylation and transcript expression in invasive cervical cancer were induced by chemoradiation. Hence, paired pre- and post-treatment biopsy samples were evaluated for in vivo changes in promoter methylation and transcript expression of 10 genes (ESR1, BRCA1, RASSF1A, MYOD1, MLH1, hTERT, MGMT, DAPK1, BAX and BCL2L1) in response to chemoradiation therapy.
Methods: In patients with locally advanced invasive cervical cancer, paired pre- and post-treatment biopsies after 10 Gy chemoradiation were obtained. DNA/RNA was extracted and gene promoter methylation status was evaluated by custom-synthesized methylation PCR arrays, and the corresponding gene transcript expression was determined by absolute quantification method using quantitative reverse transcription PCR.
Results: Changes in the gene promoter methylation as well as gene expression following chemoradiation therapy were observed. BAX promoter methylation showed a significant increase (P< 0.01) following treatment. There was a significant increase in the gene transcript expression of BRCA1 (P< 0.01), DAPK1 and ESR1 (P< 0.05), whereas MYOD1 and MLH1 gene transcript expression was significantly decreased (P< 0.05) following treatment.
Interpretation & conclusions: The findings of our study show that chemoradiation therapy can induce epigenetic alterations as well as affect gene expression in tissues of invasive cervical cancer which may have implications in determining radiation response.
Strzelczyk JK, Krakowczyk Ł, Gołąbek K, Owczarek AJExpression profiles of selected genes in tumors and matched surgical margins in oral cavity cancer: Do we have to pay attention to the molecular analysis of the surgical margins?
Adv Clin Exp Med. 2018; 27(6):833-840 [PubMed
] Related Publications
BACKGROUND: Head and neck squamous cell carcinomas (HNSCCs) are associated with an interplay between genetics and the environment; they account for 3% of all diagnosed malignant tumors in men and 2% of those in women.
OBJECTIVES: The aim of the study was to analyze the significance of TIMP3, SFRP1, SFRP2, CDH1, RASSF1, RORA, and DAPK1 gene expression in head and neck squamous cell carcinoma tumors, and in matching surgical margin samples. We also analyzed the association between clinical parameters and the expression of the selected genes.
MATERIAL AND METHODS: Following surgical resection, 56 primary HNSCC tumors and matching surgical margin samples were collected from patients at the Clinic of Oncological and Reconstructive Surgery of Maria Skłodowska-Curie Memorial Cancer Center and the Institute of Oncology in Gliwice, Poland. The gene expression levels were analyzed by quantitative reverse transcription (qRT)-PCR.
RESULTS: SFRP1 gene expression was statistically significantly lower in the tumor samples than in the surgical margins (0.30 ±0.36 vs 0.62 ±0.36; p < 0.01). No correlation was found between gene expression and clinical parameters, except DAPK1, where low expression correlated with alcohol abuse (0.85 ±1.19 vs 1.97 ±3.22; p = 0.074). Moreover, patients with G3 grade tumors, i.e., poorly differentiated tumors, had significantly higher values of DAPK1 gene expression than the G1 (well-differentiated tumors) and G2 (moderately differentiated) groups.
CONCLUSIONS: There are many different reasons and concepts for altered gene expression in tumors and surgical margin tissue. Tumor heterogeneity and its microenvironment are undoubtedly linked to the biology of HNSCC. In order to understand specific tumor behavior and the microenvironment, further studies are needed. To find markers connected with cancer development and to provide insight into the earliest stages of cancer development, attention should also be focused on molecular analysis of the surgical margins.