Single-cell multimodal glioma analyses identify epigenetic regulators of cellular plasticity and environmental stress response.

Glioma intratumoral heterogeneity enables adaptation to challenging microenvironments and contributes to therapeutic resistance. We integrated 914 single-cell DNA methylomes, 55,284 single-cell transcriptomes and bulk multi-omic profiles across 11 adult IDH mutant or IDH wild-type gliomas to delineate sources of intratumoral heterogeneity. We showed that local DNA methylation disorder is associated with cell-cell DNA methylation differences, is elevated in more aggressive tumors, links with transcriptional disruption and is altered during the environmental stress response.

Differentially methylated genes and androgen receptor re-expression in small cell prostate carcinomas.

Small cell prostate carcinoma (SCPC) morphology is rare at initial diagnosis but often emerges during prostate cancer progression and portends a dismal prognosis. It does not express androgen receptor (AR) or respond to hormonal therapies. Clinically applicable markers for its early detection and treatment with effective chemotherapy are needed.

Defects of Lipid Synthesis Are Linked to the Age-Dependent Demyelination Caused by Lamin B1 Overexpression.

Unlabelled: Lamin B1 is a component of the nuclear lamina and plays a critical role in maintaining nuclear architecture, regulating gene expression and modulating chromatin positioning. We have previously shown that LMNB1 gene duplications cause autosomal dominant leukodystrophy (ADLD), a fatal adult onset demyelinating disease. The mechanisms by which increased LMNB1 levels cause ADLD are unclear.

Epigenetic mechanisms of induced pluripotency.

Reprogramming of somatic cells to induced pluripotent stem cells (iPSCs) requires profound alterations in the epigenetic landscape. During reprogramming, a change in chromatin structure resets the gene expression and stabilises self-renewal. Reprogramming is a highly inefficient process, in part due to multiple epigenetic barriers.

RUNX3 promoter hypermethylation is frequent in leukaemia cell lines and associated with acute myeloid leukaemia inv(16) subtype.

Correlative and functional studies support the involvement of the RUNX gene family in haematological malignancies. To elucidate the role of epigenetics in RUNX inactivation, we evaluated promoter DNA methylation of RUNX1, 2, and 3 in 23 leukaemia cell lines and samples from acute myeloid leukaemia (AML), acute lymphocytic leukaemia (ALL) and myelodysplatic syndromes (MDS) patients. RUNX1 and RUNX2 gene promoters were mostly unmethylated in cell lines and clinical samples.

TET1 is a maintenance DNA demethylase that prevents methylation spreading in differentiated cells.

TET1 is a 5-methylcytosine dioxygenase and its DNA demethylating activity has been implicated in pluripotency and reprogramming. However, the precise role of TET1 in DNA methylation regulation outside of developmental reprogramming is still unclear. Here, we show that overexpression of the TET1 catalytic domain but not full length TET1 (TET1-FL) induces massive global DNA demethylation in differentiated cells.

Age-related epigenetic drift in the pathogenesis of MDS and AML.

The myelodysplastic syndrome (MDS) is a clonal hematologic disorder that frequently evolves to acute myeloid leukemia (AML). Its pathogenesis remains unclear, but mutations in epigenetic modifiers are common and the disease often responds to DNA methylation inhibitors. We analyzed DNA methylation in the bone marrow and spleen in two mouse models of MDS/AML, the NUP98-HOXD13 (NHD13) mouse and the RUNX1 mutant mouse model.

Fusobacterium in colonic flora and molecular features of colorectal carcinoma.

Fusobacterium species are part of the gut microbiome in humans. Recent studies have identified overrepresentation of Fusobacterium in colorectal cancer tissues, but it is not yet clear whether this is pathogenic or simply an epiphenomenon. In this study, we evaluated the relationship between Fusobacterium status and molecular features in colorectal cancers through quantitative real-time PCR in 149 colorectal cancer tissues, 89 adjacent normal appearing mucosae and 72 colonic mucosae from cancer-free individuals.

Colorectal carcinomas with CpG island methylator phenotype 1 frequently contain mutations in chromatin regulators.

Background & Aims: Subgroups of colorectal carcinomas (CRCs) characterized by DNA methylation anomalies are termed CpG island methylator phenotype (CIMP)1, CIMP2, or CIMP-negative. The pathogenesis of CIMP1 colorectal carcinomas, and their effects on patients' prognoses and responses to treatment, differ from those of other CRCs. We sought to identify genetic somatic alterations associated with CIMP1 CRCs.

Examination of whole blood DNA methylation as a potential risk marker for gastric cancer.

Whole blood DNA methylation analysis has been proposed to be a risk marker for cancer that can be used to target patients for preventive interventions. To test this, we examined whole blood DNA methylation of 16 CpG island promoters and LINE1 repetitive element in patients with gastric cancer and control subjects. Bisulfite pyrosequencing was used to quantify the methylation of 14 CpG island promoters (MINT25, RORA, GDNF, CDH1, RARAB2, ER, CDH13, MYOD1, SFRP1, P2RX7, SLC16A12, IGF2, DPYS, and N33) and LINE1 from 72 patients with gastric cancer, 67 control, and 52 healthy young individuals.

Conserved DNA methylation patterns in healthy blood cells and extensive changes in leukemia measured by a new quantitative technique.

Genome wide analysis of DNA methylation provides important information in a variety of diseases, including cancer. Here, we describe a simple method, Digital Restriction Enzyme Analysis of Methylation (DREAM), based on next generation sequencing analysis of methylation-specific signatures created by sequential digestion of genomic DNA with SmaI and XmaI enzymes. DREAM provides information on 150,000 unique CpG sites, of which 39,000 are in CpG islands and 30,000 are at transcription start sites of 13,000 RefSeq genes.

SINE retrotransposons cause epigenetic reprogramming of adjacent gene promoters.

Almost half of the human genome and as much as 40% of the mouse genome is composed of repetitive DNA sequences. The majority of these repeats are retrotransposons of the SINE and LINE families, and such repeats are generally repressed by epigenetic mechanisms. It has been proposed that these elements can act as methylation centers from which DNA methylation spreads into gene promoters in cancer.

Repetitive elements and enforced transcriptional repression co-operate to enhance DNA methylation spreading into a promoter CpG-island.

Repression of many tumor suppressor genes in cancer is concurrent with aberrantly increased DNA methylation levels at promoter CpG islands (CGIs). About one-fourth of empirically defined human promoters are surrounded by or contain clustered repetitive elements. It was previously observed that a sharp transition of methylation exists between highly methylated repetitive elements and unmethylated promoter-CGIs in normal tissues.

DNA methylation does not stably lock gene expression but instead serves as a molecular mark for gene silencing memory.

DNA methylation is commonly thought of as a "molecular lock" that leads to permanent gene silencing. To investigate this notion, we tested 24 different histone deacetylase inhibitors (HDACi) on colon cancer cells that harbor a GFP locus stably integrated and silenced by a hypermethylated cytomegalovirus (CMV) promoter. We found that HDACi efficiently reactivated expression of GFP and many other endogenous genes silenced by DNA hypermethylation.

Frequent alteration of MLL3 frameshift mutations in microsatellite deficient colorectal cancer.

Background: MLL3 is a histone 3-lysine 4 methyltransferase with tumor-suppressor properties that belongs to a family of chromatin regulator genes potentially altered in neoplasia. Mutations in MLL3 were found in a whole genome analysis of colorectal cancer but have not been confirmed by a separate study.

Methods And Results: We analyzed mutations of coding region and promoter methylation in MLL3 using 126 cases of colorectal cancer.

Aberrant DNA methylation is associated with disease progression, resistance to imatinib and shortened survival in chronic myelogenous leukemia.

The epigenetic impact of DNA methylation in chronic myelogenous leukemia (CML) is not completely understood. To elucidate its role we analyzed 120 patients with CML for methylation of promoter-associated CpG islands of 10 genes. Five genes were identified by DNA methylation screening in the K562 cell line and 3 genes in patients with myeloproliferative neoplasms.

Dissecting DNA hypermethylation in cancer.

There is compelling evidence to support the importance of DNA methylation alterations in cancer development. Both losses and gains of DNA methylation are observed, thought to contribute pathophysiologically by inactivating tumor suppressor genes, inducing chromosomal instability and ectopically activating gene expression. Lesser known are the causes of aberrant DNA methylation.

CpG island methylation profiling in human melanoma cell lines.

A better understanding of key molecular changes during the pathogenesis of melanoma could impact strategies to reduce mortality from this cancer. Two epigenetic events involved in the pathogenesis of cancer are hypermethylation of tumor-suppressor gene promoters associated with transcriptional repression and hypomethylation associated with gene reexpression and genomic instability. We analyzed 16 melanoma cell lines for aberrant hypermethylation of 15 cancer-linked genes (ER alpha, MGMT, RAR beta 2, RIL, RASSF1A, PAX7, PGR beta, PAX2, NKX2-3, OLIG2, HAND1, ECAD, CDH13, MLH1, and p16) and hypomethylation of two genes (MAGEA1, maspin) and two repetitive sequences (LINE-1 and Alu) using pyrosequencing.

Sensitive and specific detection of early gastric cancer with DNA methylation analysis of gastric washes.

Background & Aims: Aberrant DNA methylation is an early and frequent process in gastric carcinogenesis and could be useful for detection of gastric neoplasia. We hypothesized that methylation analysis of DNA recovered from gastric washes could be used to detect gastric cancer.

Methods: We studied 51 candidate genes in 7 gastric cancer cell lines and 24 samples (training set) and identified 6 for further studies.

An Sp1/Sp3 binding polymorphism confers methylation protection.

Hundreds of genes show aberrant DNA hypermethylation in cancer, yet little is known about the causes of this hypermethylation. We identified RIL as a frequent methylation target in cancer. In search for factors that influence RIL hypermethylation, we found a 12-bp polymorphic sequence around its transcription start site that creates a long allele.

Aberrant CpG island methylation in acute myeloid leukemia is accentuated at relapse.

DNA methylation of CpG islands around gene transcription start sites results in gene silencing and plays a role in leukemia pathophysiology. Its impact in leukemia progression is not fully understood. We performed genomewide screening for methylated CpG islands and identified 8 genes frequently methylated in leukemia cell lines and in patients with acute myeloid leukemia (AML): NOR1, CDH13, p15, NPM2, OLIG2, PGR, HIN1, and SLC26A4.

Methylated CpG Island Amplification and Microarray (MCAM) for High-Throughput Analysis of DNA Methylation.

INTRODUCTIONThis protocol describes the use of methylated CpG island amplification (MCA) in combination with a microarray platform to analyze genome-wide DNA methylation in a high-throughput fashion. In this approach, termed MCAM, methylated CpG islands are selectively targeted using oligonucleotide adaptors after two rounds of digestion with a combination of methylation-sensitive and methylation-insensitive nucleases. They are then amplified using PCR.

High-throughput methylation profiling by MCA coupled to CpG island microarray.

An abnormal pattern of DNA methylation occurs at specific genes in almost all neoplasms. The lack of high-throughput methods with high specificity and sensitivity to detect changes in DNA methylation has limited its application for clinical profiling. Here we overcome this limitation and present an improved method to identify methylated genes genome-wide by hybridizing a CpG island microarray with amplicons obtained by the methylated CpG island amplification technique (MCAM).

Hypomethylation of LINE-1 and Alu in well-differentiated neuroendocrine tumors (pancreatic endocrine tumors and carcinoid tumors).

Neuroendocrine tumors including carcinoid tumors and pancreatic endocrine tumors are uncommon, and the genetic alterations in these indolent tumors are not well characterized. We studied global hypomethylation by analyzing long interspersed nucleotide elements (LINE)-1 and Alu methylation using pyrosequencing in 35 neuroendocrine tumors and corresponding normal tissue. The tumor samples were less methylated than normal tissue at LINE-1 (P=0.