5-hydroxymethylcytosine in cancer: significance in diagnosis and therapy

doi:10.1016/j.cancergen.2015.02.009

Cancer Genetics

Volume 208, Issue 5, May 2015, Pages 167-177

https://doi.org/10.1016/j.cancergen.2015.02.009 Get rights and content

Emerging data have demonstrated that 5-methylcytosine (5-mC) and its oxidized products 5-hydroxymethylcytosine (5-hmC), 5-formylcytosine (5-fC), and 5-carboxylcytosine (5-CaC) play unique roles in several biological processes, including the control of gene expression and in the pathogenesis of cancer. In this review, we focus on 5-hmC and the disruption of its distribution in several cancers, including hematological malignancies and solid tumors. We present an outline of how 5-hmC is closely associated with metabolic pathways and may be the missing link connecting epigenetics with metabolism in the context of cancer cells. Finally, we discuss the diagnostic and prognostic importance of 5-mC and 5-hmC patterning, and how we may be able to establish new paradigms in cancer therapy based on these alterations.

Section snippets

Detection of 5-hmC

Commonly used techniques for the detection of methylated cytosines, such as bisulfite sequencing and methods that employ methyl-sensitive restriction enzymes, cannot distinguish between 5-mC and 5-hmC (21). However, these modified cytosines can be discriminated using several techniques, including 1) thin layer chromatography, 2) mass spectrometry, 3) antibodies, and 4) chemical conjugation.

Thin layer chromatography (TLC) and liquid chromatography-electron spray ionization tandem mass

Potential molecular mechanisms of modified cytosine bases

First identified in T-even bacteriophages approximately 60 years ago, 5-hmC was hypothesized to play a role in nucleic acid metabolism during viral replication. This base was found in mammalian cells in the 1970s (36), but its functional importance was not appreciated fully. In 2009, the catalytic function of the TETs was identified as the conversion of 5-mC to 5-hmC, which is found at reasonably high levels in Purkinje cells in the murine brain (19) and in undifferentiated embryonic stem (ES)

Disruption of 5-hmC levels in hematological malignancies

Perturbation of global 5-mC levels is one of the hallmarks of human cancers, and hypomethylation, and in some cases hypermethylation, has been observed in various hematological malignancies. More recently, several whole genome sequencing studies from myeloid malignancies have identified mutations in genes that encode DNA cytosine modifiers, including the DNMT3A, TET2, and IDH1/2 genes (3). Specifically, the disruption of TET2 and IDH1/2 gene function by mutations perturbs 5-hmC levels in

Solid tumors are associated with low 5-hmC levels

A broad spectrum of solid tumors, including gliomas, colorectal, breast, and prostate cancers, reportedly have low levels of 5-hmC associated with down-regulation of TET1, TET2, or TET3 mRNA transcription 12, 110, 111, 112, 113, suggesting that the loss of TET-mediated regulation of DNA demethylation contributes to cellular transformation. The significant loss of 5-hmC that is found in several types of solid tumors indicates a general epigenetic event in malignant transformation, correlating

Clinical implications of low 5-hmC in cancer prognosis and therapy

Many studies have sought to evaluate the impact of mutations in epigenetic modifiers on patient response to therapy. Mutations in TET2 and a favorable cytogenetic risk were both independent predictors of a higher response rate to therapy (133). A recent study correlated the levels of 5-hmC with clinical and molecular parameters in adult AML patients and found that whereas TET2 and IDH1/2-mutated cases had low levels of 5-hmC, higher levels of 5-hmC associated positively with high blast count at

Conclusions

The role of 5-hmC in cancer has been widely studied ever since its rediscovery in 2009, and these studies have revealed a loss of 5-hmC in hematological as well as solid tumors. However, the mechanisms by which 5-hmC levels are reduced are unclear. The down-regulation of 5-hmC in different malignancies appears to depend on the specific tumor type. For example, TET2 and IDH1/2 mutations in hematological malignancies lead to low hydroxymethylation, whereas down-regulation of TET transcription and

Acknowledgments

This work was supported by F32-DK092030 (A.V.) and National Institutes of Health CA129831 (L.A.G.).

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Citation Excerpt :
Studies show the emerging role of 5hmC as a prominent epigenetic marker, and it has been found to be associated with tumor progression. It is also found to be enriched in enhancers, promoters and changes in 5hmC level are linked to changes in gene expression levels as well [42–44]. A variety of techniques have been developed such as 5hmC-Seal [45], hmC–CATCH [46], oxBS-seq [47], TAB-seq [48] and hMeDIP-seq [49] etc. which makes use of 5 hydroxymethylation profiling techniques.
Cell-free DNA(cfDNA) methylation profiling is considered promising and potentially reliable for liquid biopsy to study progress of diseases and develop reliable and consistent diagnostic and prognostic biomarkers. There are several different mechanisms responsible for the release of cfDNA in blood plasma, and henceforth it can provide information regarding dynamic changes in the human body. Due to the fragmented nature, low concentration of cfDNA, and high background noise, there are several challenges in its analysis for regular use in diagnosis of cancer. Such challenges in the analysis of the methylation profile of cfDNA are further aggravated due to heterogeneity, biomarker sensitivity, platform biases, and batch effects. This review delineates the origin of cfDNA methylation, its profiling, and associated computational problems in analysis for diagnosis. Here we also contemplate upon the multi-marker approach to handle the scenario of cancer heterogeneity and explore the utility of markers for 5hmC based cfDNA methylation pattern. Further, we provide a critical overview of deconvolution and machine learning methods for cfDNA methylation analysis. Our review of current methods reveals the potential for further improvement in analysis strategies for detecting early cancer using cfDNA methylation.
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Contributions to Progress and Promise of Epigenetics for Diagnosis and Therapy in Cancer5-hydroxymethylcytosine in cancer: significance in diagnosis and therapy

Section snippets

Detection of 5-hmC

Potential molecular mechanisms of modified cytosine bases

Disruption of 5-hmC levels in hematological malignancies

Solid tumors are associated with low 5-hmC levels

Clinical implications of low 5-hmC in cancer prognosis and therapy

Conclusions

Acknowledgments

Biochem Biophys Res Commun

Blood

Biochem Biophys Res Commun

Blood

Blood

Cell

J Mol Biol

Cancer Cell

Blood

Cell Rep

Cancer Cell

Cell

Cell Rep

J Biol Chem

Cell Rep

Cell Stem Cell

Cell Stem Cell

Blood

Cancer Cell

Dev Cell

Cell

Cell

J Biol Chem

Mol Cell

J Biol Chem

J Biol Chem

Cancer Cell

Blood

Blood

Semin Hematol

Blood

Blood

J Thromb Haemost

Blood

Blood

Leuk Res

Blood

Cancer Cell

Transcriptional silencing by polycomb-group proteins

Cold Spring Harb Perspect Biol

Histone deacetylases and their inhibitors in cancer, neurological diseases and immune disorders

Nat Rev Drug Discov

Histones and their modifications in ovarian cancer - drivers of disease and therapeutic targets

Front Oncol

The emerging roles of DOT1L in leukemia and normal development

Leukemia

Histone variants: dynamic punctuation in transcription

Genes & Development

Impaired hydroxylation of 5-methylcytosine in myeloid cancers with mutant TET2

Nature

Decreased 5-hydroxymethylcytosine is associated with neural progenitor phenotype in normal brain and shorter survival in malignant glioma

PLoS ONE

Cloning and characterization of a family of novel mammalian DNA (cytosine-5) methyltransferases

Nat Genet

Conversion of 5-methylcytosine to 5-hydroxymethylcytosine in mammalian DNA by MLL partner TET1

Science

Tet proteins can convert 5-methylcytosine to 5-formylcytosine and 5-carboxylcytosine

Science

Tet-mediated formation of 5-carboxylcytosine and its excision by TDG in mammalian DNA

Science

The nuclear DNA base 5-hydroxymethylcytosine is present in Purkinje neurons and the brain

Science

Contributions to Progress and Promise of Epigenetics for Diagnosis and Therapy in Cancer
5-hydroxymethylcytosine in cancer: significance in diagnosis and therapy