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Validation of Illumina 450 K DNA methylation bead array by Pyrosequencing. Correlations between Illumina 450 K array data and pyrosequence analysis. Representative data for CpG sites in cg27182070 (RPA2), cg16624210 (TPPP), cg19761014 (LRRC37B2), cg00481259 (DECR2), cg07271186 (TRY2P), cg01252526 (WDR9), and genes is shown
Source publication
Background:
Immune surveillance acts as a defense mechanism in cancer, and its disruption is involved in cancer progression. DNA methylation reflects the phenotypic identity of cells and recent data suggested that DNA methylation profiles of T cells and peripheral blood mononuclear cells (PBMC) are altered in cancer progression.
Methods:
We enro...
Contexts in source publication
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... determined a correlation between DNA methylation levels obtained from Illumina 450 K and pyrosequenc- ing. Pyrosequencing analysis was limited by the remaining amount of T-cell DNA. A few representative samples were used for validation purposes. Nine normal and five breast cancer T-cell DNA samples were sub- jected to bisulphite conversion and pyrosequencing ana- lysis. Seven CG probes that varied across the samples were randomly selected. Figure 3 shows the correlation between values obtained by Illumina analysis and pyro- sequencing for these CGs. Correlations were significant for all probes with r values between 0.5 and 0.8 ( Fig. ...
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... determined a correlation between DNA methylation levels obtained from Illumina 450 K and pyrosequenc- ing. Pyrosequencing analysis was limited by the remaining amount of T-cell DNA. A few representative samples were used for validation purposes. Nine normal and five breast cancer T-cell DNA samples were sub- jected to bisulphite conversion and pyrosequencing ana- lysis. Seven CG probes that varied across the samples were randomly selected. Figure 3 shows the correlation between values obtained by Illumina analysis and pyro- sequencing for these CGs. Correlations were significant for all probes with r values between 0.5 and 0.8 ( Fig. ...
Context 3
... gene panel shown in Fig. 3 was further ana- lyzed using the KM plotter (Kaplan-Meier plotter) data- base of breast cancer patients in order to investigate the prognostic significance of the genes in breast cancer relapse-free survival. The KM-plotter database was gen- erated by Gyorffy et al. using the NCBI Gene Expression Omnibus (GEO) repository of gene expression and pa- tient survival information [21], and is often used to investigate the clinical significance of particular gene(s) in several common cancers. Using gene expression data from 1764 breast cancer patients, an association between the decreased expression of genes identified and vali- dated in Fig. 3 was observed with a lower incidence of relapse-free or disease-free survival (Fig. 4). The high or low expression groups were classified according to whether the combined expression of the genes was greater than their median ...
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... gene panel shown in Fig. 3 was further ana- lyzed using the KM plotter (Kaplan-Meier plotter) data- base of breast cancer patients in order to investigate the prognostic significance of the genes in breast cancer relapse-free survival. The KM-plotter database was gen- erated by Gyorffy et al. using the NCBI Gene Expression Omnibus (GEO) repository of gene expression and pa- tient survival information [21], and is often used to investigate the clinical significance of particular gene(s) in several common cancers. Using gene expression data from 1764 breast cancer patients, an association between the decreased expression of genes identified and vali- dated in Fig. 3 was observed with a lower incidence of relapse-free or disease-free survival (Fig. 4). The high or low expression groups were classified according to whether the combined expression of the genes was greater than their median ...
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... list of 89 CGs clusters all individuals by their cancer stage, pointing to the possibility of using a combination of CG methylation states to detect and stage breast cancer early noninvasively. It is important to note that these CGs detect early stages of breast cancer and no association was found with allergy, immune mediated disorders or inflam- mation [30,31]. Genes validated in Fig. 3 are involved in DNA replication and repair, cell cycle, mitosis, oligo- dendrocyte differentiation, tubulin polymerization, signal transduction, transcription regulation, autophagy, apop- tosis and regulation of lipid metabolism, which collectively play an important role in several malignancies including breast cancer. The uniqueness of the identified signatures was further confirmed by the survival-curve generated from their gene expression profile in breast cancer (Fig. 4). Future prospective clinical studies are required to determine whether they can detect breast cancer earlier than currently available imaging methods. In these cases, loss of DNA methylation associated with advanced breast cancer stage may in fact reflect the demethylation of pro-metastatic genes as previously described by us [32][33][34][35]. Results from these studies are in line with identi- fied CpG probes which showed a change in DNA methy- lation and correlation with disease progression in liver cancer patients [16]. We also observed overlap of probes among DCIS, mixed and invasive breast cancer, their asso- ciation with canonical pathways and upstream regulators of gene expression (Additional file 1: Tables ...
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DNA methylation may be involved in development of type 1 diabetes (T1D), but previous epigenome-wide association studies were conducted among cases with clinically diagnosed diabetes. Using multiple pre-disease peripheral blood samples on the Illumina 450 K and EPIC platforms, we investigated longitudinal methylation differences between 87 T1D case...
Citations
... Previous studies have found alterations in DNA methylation patterns of peripheral blood leukocytes in the liver [8], prostate [9], and breast cancers [10]. Due to the excellent stability of DNA compared to other biological materials, such as RNAs or proteins, DNA methylation can be easily detected in small specimens, making it suitable for largescale epidemiological studies. ...
Background
Gastric adenocarcinoma is a prevalent form of cancer that often remains undetected in its early stages due to the lack of specific symptoms. This delayed diagnosis leads to poor clinical outcomes, underscoring the need for an effective and non-invasive method for early detection. Recent advances in cancer epigenetics have led to the identification of biomarkers that have the potential to revolutionize the early detection and monitoring of this disease. One such promising biomarker is the methylation of the FGFR2 promoter. This study aims to measure the methylation levels of a specific CpG site in the FGFR2 promoter gene in DNA extracted from blood leukocytes from patients with intestinal metaplasia, gastric cancer, and healthy control.
Material and methods
The CpG site of the FGFR2 gene promoter was identified in its control region. Methylation alteration of the selected FGFR2 CpG site was determined through the (methylation-sensitive restriction enzyme) MSRE-qPCR. Genomic DNA was extracted from one hundred twenty-five participants.
Results
The normal group had mean methylation levels of 93.23 ± 4.929%, while the IM group had a level of 69.85 ± 27.15%. In GC patients, the levels varied, with 25.96 ± 18.98% in the intestinal type and 28.30 ± 16.07% in the diffuse type. The methylation levels in the IM and GC patients were significantly lower than those in the normal control group. However, no significant difference was observed between the methylation status of the intestinal type of GC and the diffuse type. The Receiver operating characteristic (ROC) curve analysis showed that FGFR2 CpG methylation levels in GC patients compared to normal controls had a high sensitivity of 100% and specificity of 100%, with a cut-off of < 74.25%; when GC patients were compared to IM patients, the sensitivity was 85%, and the specificity was 80%, with a cut-off < 44.45%.
Conclusions
The potential of the FGFR2 methylation status as a non-invasive biomarker lies in its ability to be detected in blood leukocytes, which makes it a promising tool for the early detection of intestinal metaplasia and gastric cancer. This could significantly improve the detection and management of these gastric conditions.
... Furthermore, the transcription start sites (TSS) of BC patients were hypomethylated compared with normal controls (Fig. 2c), which may be related to the immune activation in BC patients. This finding is consistent with the results from ref. 30. The differentially methylated genes were then subjected to pathway analysis. ...
The immune system can monitor tumor development, and DNA methylation is involved in the body’s immune response to tumors. In this work, we investigate whether DNA methylation alterations in peripheral blood mononuclear cells (PBMCs) could be used as markers for early detection of breast cancer (BC) from the perspective of tumor immune alterations. We identify four BC-specific methylation markers by combining Infinium 850 K BeadChips, pyrosequencing and targeted bisulfite sequencing. Based on the four methylation markers in PBMCs of BC, we develop an efficient and convenient multiplex methylation-specific quantitative PCR assay for the detection of BC and validate its diagnostic performance in a multicenter cohort. This assay was able to distinguish early-stage BC patients from normal controls, with an AUC of 0.940, sensitivity of 93.2%, and specificity of 90.4%. More importantly, this assay outperformed existing clinical diagnostic methods, especially in the detection of early-stage and minimal tumors.
... In several studies, blood-based DNA methylation has been profiled to develop a robust diagnostic marker for cancer. The blood-based methylation studies are broadly divided into investigating global DNA methylation [29] and gene-specific targeted DNA methylation [18]. In addition, according to the source of DNA, these studies mainly targeted circulating tumor cells (CTCs) and cell-free DNA in serum or plasma [30]. ...
Background:
Genome-wide dysregulation of CpG methylation accompanies tumor progression and characteristic states of cancer cells, prompting a rationale for biomarker development. Understanding how the archetypic epigenetic modification determines systemic contributions of immune cell types is the key to further clinical benefits.
Results:
In this study, we characterized the differential DNA methylome landscapes of peripheral blood mononuclear cells (PBMCs) from 76 canines using methylated CpG-binding domain sequencing (MBD-seq). Through gene set enrichment analysis, we discovered that genes involved in the growth and differentiation of T- and B-cells are highly methylated in tumor PBMCs. We also revealed the increased methylation at single CpG resolution and reversed expression in representative marker genes regulating immune cell proliferation (BACH2, SH2D1A, TXK, UHRF1). Furthermore, we utilized the PBMC methylome to effectively differentiate between benign and malignant tumors and the presence of mammary gland tumors through a machine-learning approach.
Conclusions:
This research contributes to a better knowledge of the comprehensive epigenetic regulation of circulating immune cells responding to tumors and suggests a new framework for identifying benign and malignant cancers using genome-wide methylome.
... выявил в Т-клетках ~10 000 сайтов, ассоциированных с прогрессированием РМЖ, из них 89 сайтов CpG также сильно коррелировали с прогрессированием заболевания (p <0,01; r >0,7; r <-0,7). Подавляющее большинство этих гипометилированных сайтов имеют прямое отношение к генам, белки которых участвуют в функционировании иммунной системы [16]. ...
Introduction . The growth of primary breast tumor morbidity in the last ten years and increased number of patients with disseminated breast cancer in the Republic of Kazakhstan require the search for methods of early diagnosis of malignant tumors. Determination of breast cancer markers in epigenetic studies allows to use them as diagnostic signs of the presence of malignant tumor and as predictors of treatment effectiveness in patients with this pathology.
Aim . To perform a search for therapeutic and prognostic breast cancer markers.
Materials and methods . The study included samples of biological material (peripheral blood) of 50 deemed healthy individuals and 103 patients with locally advanced and disseminated breast cancer receiving special therapy. The following methods were used: blood collection, DNA extraction, creation of DNA methylation profiles, sequencing, statistical data analysis.
Results . The results of search for epigenetic mutations in peripheral blood of patients with breast cancer showed their role as specific diagnostic, therapeutic and prognostic markers with specificity 0.91 % and sensitivity 0.94 %. The hypothesis on therapeutic significance of identified earlier diagnostic markers in patients with breast cancer, namely hypermethylation of CpG islands associated with genes JAM3, C17orf64, MSC, C7orf51 and CpG island associated with intragene part of chromosome 5 (chr5: 77,208,034–77,329,434) was tested and confirmed.
Conclusion . A correlation between DNA methylation characteristics and disease progression during treatment was shown. The study results can be used in clinical practice: epigenetic markers, such as methylation in the CpG islets associated with the JAM3, C17orf64, MSC, C7orf51 genes, and in the CpG islet associated with the intragenic site of chromosome 5 (chr5: 77,208,034–77,329,434) can be used as prognostic markers and therapeutic predictors of breast cancer.
... Furthermore, patterns of blood DNA methylation have been recently connected to prognosis of OC patients. Taken together, methylation markers in OC might be helpful for evaluation of therapeutic effects and recognition of chemoresistance-related pathways [68][69][70][71]. ...
Advanced and recurrent gynecological cancers lack effective treatment and have poor prognosis. Besides, there is urgent need for conservative treatment for fertility protection of young patients. Therefore, continued efforts are needed to further define underlying therapeutic targets and explore novel targeted strategies. Considerable advancements have been made with new insights into molecular mechanisms on cancer progression and breakthroughs in novel treatment strategies. Herein, we review the research that holds unique novelty and potential translational power to alter the current landscape of gynecological cancers and improve effective treatments. We outline the advent of promising therapies with their targeted biomolecules, including hormone receptor-targeted agents, inhibitors targeting epigenetic regulators, antiangiogenic agents, inhibitors of abnormal signaling pathways, poly (ADP-ribose) polymerase (PARP) inhibitors, agents targeting immune-suppressive regulators, and repurposed existing drugs. We particularly highlight clinical evidence and trace the ongoing clinical trials to investigate the translational value. Taken together, we conduct a thorough review on emerging agents for gynecological cancer treatment and further discuss their potential challenges and future opportunities.
... Parashar et al. found a correlation between T-cell DNA methylation levels and TNM staging of breast cancer, a finding that could lead to the development of molecular markers for immune staging. In this study, T cell DNA was isolated and treated with Illumina Indinium methylated EPIC arrays and validated using Illumina amplicon sequencing and pyrophosphate sequencing platforms [64]. Specific classification of DNA methylation has also been found to be a biomarker for prognosis and clinical stratification of patients with different BC subtypes, which offers the prospect of future development of individualized treatment [65]. ...
DNA methylation is a common epigenetic modification, and the current commonly used methods for DNA methylation detection include methylation-specific PCR, methylation-sensitive restriction endonuclease-PCR, and methylation-specific sequencing. DNA methylation plays an important role in genomic and epigenomic studies, and combining DNA methylation with other epigenetic modifications, such as histone modifications, may lead to better DNA methylation. DNA methylation also plays an important role in the development of disease, and analyzing changes in individual DNA methylation patterns can provide individualized diagnostic and therapeutic solutions. Liquid biopsy techniques are also increasingly well established in clinical practice and may provide new methods for early cancer screening. It is important to find new screening methods that are easy to perform, minimally invasive, patient-friendly, and affordable. DNA methylation mechanisms are thought to have an important role in cancer and have potential applications in the diagnosis and treatment of female tumors. This review discussed early detection targets and screening methods for common female tumors such as breast, ovarian, and cervical cancers and discussed advances in the study of DNA methylation in these tumors. Although existing screening, diagnostic, and treatment modalities exist, the high morbidity and mortality rates of these tumors remain challenging.
... As a consequence, abnormal hypermethylation in the genes promoter regions is recognized as an important and primary event during the tumorigenesis process (Bosviel et al., 2012;Wong et al., 2011). Although the abnormal DNA methylation has been reported as one of the hallmarks of cancerous tissues, several studies have previously suggested that the methylation status of a gene in blood-derived DNA might be associated with a risk of developing breast cancer (Gupta et al., 2014;Hosny et al., 2016;Niwa et al., 2000;Parashar et al., 2018;Severi et al., 2014). According to the high readability and non-invasiveness characteristics of peripheral blood DNA, the assessment of white blood cells DNA, as a biomarker of breast cancers risk, is of particular interest (Brennan et al., 2012;Li et al., 2012). ...
... Accordingly, these changes are induced in blood cells and other tissues as well (Gupta et al., 2014;Khakpour et al., 2015;. It has also been suggested that the DNA methylation profile of T cells and mononuclear cells in peripheral blood, are changed while the cancer progresses (Parashar et al., 2018). It is worth mentioning that using the MSP qualitative method and sample size can affect the difference between the results of our study and those of some other studies. ...
Objective:
Similar to other types of cancer, the development of breast cancer is a multi-stage process, consisting of various mutations and epigenetic changes in many genes. Mutations in the BRCA1 gene, which is a tumor suppressor gene, are considered as the most important types of mutations. The pivotal role of epigenetics is currently considered as the primary key to carcinogenesis. Several studies have previously reported the BRCA1 epigenetic silencing through promoter methylation in the pathophysiology of breast cancer cells. This study aimed to investigate whether the BRCA1 gene promoter methylation in peripheral blood cells is correlated with the risk of breast cancer.
Methods:
In the current study, DNA samples were extracted from blood cells belonged to 74 patients with breast cancer as well as 30 healthy individuals, and the BRCA1 gene promoter methylation status in these two groups was examined using Methylation Specific PCR (MSP).
Result:
out of 74 patients, 2 cases demonstrated methylation in their BRCA1 gene promoter; however, none of the healthy controls demonstrated methylation status. Among these 74 patients, 13 cases were at the early stages (stage I), and two patients who had BRCA1 gene methylation (15.4%), were in this group (p=0.02). While 34 and 27 patients were at stages II and III, respectively, showing a negative state of BRCA1 gene methylation.
Conclusion:
Although 2 out of 74 patients resulted positive for methylation status, the healthy controls demonstrated no methylation. Consequently, there was inadequate evidence to confirm the association between BRCA1 gene promoter methylation in blood and the risk of developing breast cancer.
... In several studies, blood-based DNA methylation has been pro led to develop a robust diagnostic marker for cancer. The blood-based methylation studies are broadly divided into investigating global DNA methylation [29] and gene-speci c targeted DNA methylation [18]. In addition, according to the source of DNA, these studies mainly targeted circulating tumor cells (CTCs) and cell-free DNA in serum or plasma [30]. ...
Background
Genome-wide dysregulation of CpG methylation accompanies tumor progression and characteristic states of cancer cells, prompting a rationale for biomarker development. Understanding how the archetypic epigenetic modification determines systemic contributions of immune cell types is the key to further clinical benefits.
Results
In this study, we characterized the differential DNA methylome landscapes of peripheral blood mononuclear cells (PBMCs) from 76 canines using methylated CpG-binding domain sequencing (MBD-seq). Through gene set enrichment analysis, we discovered that genes involved in the growth and differentiation of T- and B-cells are highly methylated in tumor PBMCs. We also revealed the increased methylation at single CpG resolution and reversed expression in representative marker genes regulating immune cell proliferation (BACH2, SH2D1A, TXK, UHRF1). Furthermore, we utilized the PBMC methylome to effectively differentiate between benign and malignant tumors and the presence of mammary gland tumors through a machine-learning approach.
Conclusions
This research contributes to a better knowledge of the comprehensive epigenetic regulation of circulating immune cells responding to tumors and suggests a new framework for identifying benign and malignant cancers using genome-wide methylome.
... As we know, the methylation status is dynamic. External factors, such as viral infection, burn, and tobacco, could alter the methylation status (26)(27)(28)(29). Furthermore, several studies have shown early changes of Alu element methylation in cancer (28,30). ...
... External factors, such as viral infection, burn, and tobacco, could alter the methylation status (26)(27)(28)(29). Furthermore, several studies have shown early changes of Alu element methylation in cancer (28,30). Moreover, this alteration in methylation has also been found preclinically (22). ...
Background/aim:
During metastatic disease development, the cancer-immune system crosstalk induces epigenetic modifications to immune cells, impairing their functions. Recently, Alu elements methylation changes were widely studied in terms of early cancer detection. This study aimed to demonstrate in vitro Alu element methylation changes in peripheral immune cells in a metastatic setting and examine their prognostic values in metastatic breast cancer.
Materials and methods:
Sera from sixteen metastatic cancer patients and sixteen healthy participants were obtained and used to culture normal peripheral immune cells. After 48 h of incubation, the percentage and pattern of Alu element methylation were examined for clinical relevance.
Results:
We found that the Alu element hypomethylation was affected by age in the cancer group. Intriguingly, a decrease in Alu element methylation was found in patients with early progressive disease. Moreover, an increase in unmethylated cytosine (mCuC) loci was related to the poorer prognosis group. Accordingly, the decrease in Alu element methylation and the increase in mCuC loci pattern in peripheral immune cells correlated with poorer prognosis and early progression in metastatic breast cancer.
Conclusion:
Alu element hypomethylation in immune cells and their increased mCuC foci were related to the early progression of breast cancer. These warrant the use of Alu element methylation changes for diagnostic and therapeutic purposes in breast cancer.
... In addition to cfDNA gene sequence and mutation, cfDNA can be further analyzed for epigenetic alterations, such as DNA methylation, histone modifications, and expression of long and micro non-coding RNAs [51,52]. Methylation changes in DNA contribute to gene expression regulation and play a significant role in the etiology of early breast cancer [53,54]. The DNA methylation pattern is retained in the cfDNA released from its tissue origins of tumor cells [55,56]. ...
Simple Summary
Breast cancer is the most common cancer and leading cause of death worldwide. Therefore, it is important to diagnose and treat breast cancer early. Current diagnostic methods include mammography and tissue biopsy; however, they have limitations. Liquid biopsy is a less invasive tool for diagnosis. In this review, we summarize and focus on the recent discoveries on liquid biopsy and development of detection techniques.
Abstract
Breast cancer is the most commonly diagnosed cancer and leading cause of cancer mortality among woman worldwide. The techniques of diagnosis, prognosis, and therapy monitoring of breast cancer are critical. Current diagnostic techniques are mammography and tissue biopsy; however, they have limitations. With the development of novel techniques, such as personalized medicine and genetic profiling, liquid biopsy is emerging as the less invasive tool for diagnosing and monitoring breast cancer. Liquid biopsy is performed by sampling biofluids and extracting tumor components, such as circulating tumor cells (CTCs), circulating tumor DNA (ctDNA), cell-free mRNA (cfRNA) and microRNA (miRNA), proteins, and extracellular vehicles (EVs). In this review, we summarize and focus on the recent discoveries of tumor components and biomarkers applied in liquid biopsy and novel development of detection techniques, such as surface-enhanced Raman spectroscopy (SERS) and microfluidic devices.
