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Cancer diagnostic classifiers based on quantitative DNA methylation.

Lorincz AT - Expert Rev. Mol. Diagn. (2014)

Bottom Line: Reversible methylation of cytosines is noteworthy because it can be measured accurately and easily by various molecular methods and DNA methylation patterns are linked to important tumourigenic pathways.Differential methylation may have a central role in the development and outcome of most if not all human malignancies.Also discussed is differential methylation of specific human and viral DNA targets and laboratory methods for measuring methylation biomarkers.

View Article: PubMed Central - PubMed

Affiliation: Centre for Cancer Prevention, Wolfson Institute of Preventive Medicine, Queen Mary University of London, London, EC1M 6BQ, UK.

ABSTRACT
Epigenetic change is part of the carcinogenic process and a deep reservoir for biomarker discovery. Reversible methylation of cytosines is noteworthy because it can be measured accurately and easily by various molecular methods and DNA methylation patterns are linked to important tumourigenic pathways. Clinically relevant methylation changes are known in common human cancers such as cervix, prostate, breast, colon, bladder, stomach and lung. Differential methylation may have a central role in the development and outcome of most if not all human malignancies. The advent of deep sequencing holds great promise for epigenomics, with bioinformatics tools ready to reveal large numbers of new targets for prognosis and therapeutic intervention. This review focuses on two selected cancers, namely cervix and prostate, which illustrate the more general themes of epigenetic diagnostics in cancer. Also discussed is differential methylation of specific human and viral DNA targets and laboratory methods for measuring methylation biomarkers.

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Related in: MedlinePlus

Examples of DNA methylation modifications on cytosines (C) of CpG dyads and the effects of these changes at the level of the gene and relationship to carcinogenesis. (A) Nucleotide C is shown on the left, this base can be methylated at position 5 of the aromatic ring by one of several DNMT to become 5-methyl cytosine shown on the right. The methylation is reversible and methyl modifications can be changed to hydroxymethyl or other groups or removed by various enzymes. (B) The gross effect of DNA hypermethylation in a gene promoter region, is to turn off transcription of the gene (shown by the curved arrow). Many tumor suppressor genes, such as APC and BCL2 are controlled by methylation, and if the genes are deactivated, then critical checks and balances in cells are removed, which can lead to apoptosis or to carcinogenesis. Similarly, oncogenes such as CCND2 can be activated by removal of methylation marks from important regulatory regions [73,89]. (C) The methylation levels of a DNA region can be measured by several methods. One of the more comprehensive and convenient methods involves bisulfite conversion, which chemically changes nucleotide C into uracil (U). The U then pairs with adenine (A) and upon replication of DNA by PCR the nucleotide U is converted into thymine (T). This C to T change can be measured by sequencing and forms the basis of many DNA methylation quantitation tests. The affected C and T bases are shown in underlined boldface and a methylated C is shown as C*. The upper strand represents the native DNA sequence and the lower strand the same sequence replicated after PCR amplification.
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Figure 1: Examples of DNA methylation modifications on cytosines (C) of CpG dyads and the effects of these changes at the level of the gene and relationship to carcinogenesis. (A) Nucleotide C is shown on the left, this base can be methylated at position 5 of the aromatic ring by one of several DNMT to become 5-methyl cytosine shown on the right. The methylation is reversible and methyl modifications can be changed to hydroxymethyl or other groups or removed by various enzymes. (B) The gross effect of DNA hypermethylation in a gene promoter region, is to turn off transcription of the gene (shown by the curved arrow). Many tumor suppressor genes, such as APC and BCL2 are controlled by methylation, and if the genes are deactivated, then critical checks and balances in cells are removed, which can lead to apoptosis or to carcinogenesis. Similarly, oncogenes such as CCND2 can be activated by removal of methylation marks from important regulatory regions [73,89]. (C) The methylation levels of a DNA region can be measured by several methods. One of the more comprehensive and convenient methods involves bisulfite conversion, which chemically changes nucleotide C into uracil (U). The U then pairs with adenine (A) and upon replication of DNA by PCR the nucleotide U is converted into thymine (T). This C to T change can be measured by sequencing and forms the basis of many DNA methylation quantitation tests. The affected C and T bases are shown in underlined boldface and a methylated C is shown as C*. The upper strand represents the native DNA sequence and the lower strand the same sequence replicated after PCR amplification.

Mentions: DNA methylation is a very important information storage element of the cellular epigenetic machinery and is essential for normal development [1–3]. Aberrant methylation is a central feature of carcinogenesis; it causes defective gene expression, faulty condensation and chromosomal instability, and is a hallmark of cellular defenses acting to silence foreign DNA. Methylation at position 5 of the cytosine ring in a CpG dyad is initiated and maintained by a set of DNA methyl transferases (DNMT), some of which are de-novo (DNMT3A, DNMT3B), while another (DNMT1) is a maintenance enzyme [1]. Methylation is a noteworthy event because 5-methyl cytosine (5-mC) represents a 5th base, and levels of this molecule (and related molecules) in tissue can be measured quite accurately and easily (Figure 1)[2,4]. There are several other cytosine modifications that participate in DNA methylation and demethylation events; one of the more notable ones is 5-hydroxymethyl cytosine (5-HmC), which is generated by the Ten-eleven translocation complex of enzymes. 5-HmC appears to be important in certain organs such as the central nervous system and is also an intermediate on the active demethylation pathway for 5-mC [5]. Bisulfite conversion of DNA followed by PCR does not distinguish between 5-mC and 5-HmC. In this review, methylated DNA refers to any region of DNA with one or more modified cytosines that behave like 5-mC after bisulfite conversion.


Cancer diagnostic classifiers based on quantitative DNA methylation.

Lorincz AT - Expert Rev. Mol. Diagn. (2014)

Examples of DNA methylation modifications on cytosines (C) of CpG dyads and the effects of these changes at the level of the gene and relationship to carcinogenesis. (A) Nucleotide C is shown on the left, this base can be methylated at position 5 of the aromatic ring by one of several DNMT to become 5-methyl cytosine shown on the right. The methylation is reversible and methyl modifications can be changed to hydroxymethyl or other groups or removed by various enzymes. (B) The gross effect of DNA hypermethylation in a gene promoter region, is to turn off transcription of the gene (shown by the curved arrow). Many tumor suppressor genes, such as APC and BCL2 are controlled by methylation, and if the genes are deactivated, then critical checks and balances in cells are removed, which can lead to apoptosis or to carcinogenesis. Similarly, oncogenes such as CCND2 can be activated by removal of methylation marks from important regulatory regions [73,89]. (C) The methylation levels of a DNA region can be measured by several methods. One of the more comprehensive and convenient methods involves bisulfite conversion, which chemically changes nucleotide C into uracil (U). The U then pairs with adenine (A) and upon replication of DNA by PCR the nucleotide U is converted into thymine (T). This C to T change can be measured by sequencing and forms the basis of many DNA methylation quantitation tests. The affected C and T bases are shown in underlined boldface and a methylated C is shown as C*. The upper strand represents the native DNA sequence and the lower strand the same sequence replicated after PCR amplification.
© Copyright Policy - open-access
Related In: Results  -  Collection

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Show All Figures
getmorefigures.php?uid=PMC4225655&req=5

Figure 1: Examples of DNA methylation modifications on cytosines (C) of CpG dyads and the effects of these changes at the level of the gene and relationship to carcinogenesis. (A) Nucleotide C is shown on the left, this base can be methylated at position 5 of the aromatic ring by one of several DNMT to become 5-methyl cytosine shown on the right. The methylation is reversible and methyl modifications can be changed to hydroxymethyl or other groups or removed by various enzymes. (B) The gross effect of DNA hypermethylation in a gene promoter region, is to turn off transcription of the gene (shown by the curved arrow). Many tumor suppressor genes, such as APC and BCL2 are controlled by methylation, and if the genes are deactivated, then critical checks and balances in cells are removed, which can lead to apoptosis or to carcinogenesis. Similarly, oncogenes such as CCND2 can be activated by removal of methylation marks from important regulatory regions [73,89]. (C) The methylation levels of a DNA region can be measured by several methods. One of the more comprehensive and convenient methods involves bisulfite conversion, which chemically changes nucleotide C into uracil (U). The U then pairs with adenine (A) and upon replication of DNA by PCR the nucleotide U is converted into thymine (T). This C to T change can be measured by sequencing and forms the basis of many DNA methylation quantitation tests. The affected C and T bases are shown in underlined boldface and a methylated C is shown as C*. The upper strand represents the native DNA sequence and the lower strand the same sequence replicated after PCR amplification.
Mentions: DNA methylation is a very important information storage element of the cellular epigenetic machinery and is essential for normal development [1–3]. Aberrant methylation is a central feature of carcinogenesis; it causes defective gene expression, faulty condensation and chromosomal instability, and is a hallmark of cellular defenses acting to silence foreign DNA. Methylation at position 5 of the cytosine ring in a CpG dyad is initiated and maintained by a set of DNA methyl transferases (DNMT), some of which are de-novo (DNMT3A, DNMT3B), while another (DNMT1) is a maintenance enzyme [1]. Methylation is a noteworthy event because 5-methyl cytosine (5-mC) represents a 5th base, and levels of this molecule (and related molecules) in tissue can be measured quite accurately and easily (Figure 1)[2,4]. There are several other cytosine modifications that participate in DNA methylation and demethylation events; one of the more notable ones is 5-hydroxymethyl cytosine (5-HmC), which is generated by the Ten-eleven translocation complex of enzymes. 5-HmC appears to be important in certain organs such as the central nervous system and is also an intermediate on the active demethylation pathway for 5-mC [5]. Bisulfite conversion of DNA followed by PCR does not distinguish between 5-mC and 5-HmC. In this review, methylated DNA refers to any region of DNA with one or more modified cytosines that behave like 5-mC after bisulfite conversion.

Bottom Line: Reversible methylation of cytosines is noteworthy because it can be measured accurately and easily by various molecular methods and DNA methylation patterns are linked to important tumourigenic pathways.Differential methylation may have a central role in the development and outcome of most if not all human malignancies.Also discussed is differential methylation of specific human and viral DNA targets and laboratory methods for measuring methylation biomarkers.

View Article: PubMed Central - PubMed

Affiliation: Centre for Cancer Prevention, Wolfson Institute of Preventive Medicine, Queen Mary University of London, London, EC1M 6BQ, UK.

ABSTRACT
Epigenetic change is part of the carcinogenic process and a deep reservoir for biomarker discovery. Reversible methylation of cytosines is noteworthy because it can be measured accurately and easily by various molecular methods and DNA methylation patterns are linked to important tumourigenic pathways. Clinically relevant methylation changes are known in common human cancers such as cervix, prostate, breast, colon, bladder, stomach and lung. Differential methylation may have a central role in the development and outcome of most if not all human malignancies. The advent of deep sequencing holds great promise for epigenomics, with bioinformatics tools ready to reveal large numbers of new targets for prognosis and therapeutic intervention. This review focuses on two selected cancers, namely cervix and prostate, which illustrate the more general themes of epigenetic diagnostics in cancer. Also discussed is differential methylation of specific human and viral DNA targets and laboratory methods for measuring methylation biomarkers.

Show MeSH
Related in: MedlinePlus