Cancer Epigenetics

Cancer Epigenetics
Dr Adnan Arshad
MBBS, MS (Oncology)
PhD (Radio-Oncology)
DNA (gene)
Transcription
RNA processing (splicing etc)
mRNA
Translation
Protein
Folding
Post translational modifications
Structural or Functional Activity

2
Promoters, enhancers, silencers etc.
3
Histone deacetylases and their inhibitors in cancer, neurological diseases and immune disorders
Nature Reviews Drug Discovery 13, 673–691 (2014)
Epigenetic Chromatin Regulation
A. Modification at the DNA level

1. Cytosine methylation
B. Histone modification - the histone code

1. Histone acetylation
2. Histone methylation
3. Histone phosphorylation
4. Histone ubiquitination
9
The five nucleotides that make up the DNA
DNA Methylation
• Covalent addition of methyl group to 5th position of cytosine with

CpG dinucleotides located in the promoter region of genes
DNA Methylation
CpG – Cytosine phosphate Guanine
• Strongly represented in repetitive sequence associated with

retroviral-derived sequence
• Can be methylated to generate 5-methylcytosine
• Spontaneously deaminates to form thymine
• Poorly recognized by DNA repair systems thus:
• CG→TG mutation is propagated

• CpG levels are less frequent than predicted 1/16
• May contribute to relative inactivity of retro-elements
CpG dinucleotides are

palindromic 5’ CpG 3’
3’ GpC 5’
DNA Methylation
3 human DNA methyltransferases
• DNMT1 maintenance methyltransferases

• DNMT3A de novo methyltransferases – highly expressed at embryo implantation
• DNMT3B when waves of de novo methylation are occurring in the genome
DNMT1 has high affinity for

hemimethylated DNA, hence
helps in maintenance of
methylation after replication
◄daughter strand
of DNA
◄daughter strand
DNA methylation
CpG–island methylation – how does it affect transcription?
methylated-DNA binding proteins (MECP2, methyl CpG binding protein 2 ) bind to DNA
this recruits a complex of histone deacetylases and SIN3A
induces a closed chromatin structure → gene silencing
in contrast to usual deacetylation-related silencing, when methylation is involved, it’s (almost) irreversible
Transcription factors sensitive to methylation:
✓ E2F
✓ CREB
✓ AP2
✓ NF-KB
✓ c -myc
Roles of DNA methylation
• Transcriptional silencing
• Protecting the genome from transposition
• Genomic imprinting
• X inactivation
• Tissue specific gene expression
Genomic Imprinting
“Difference in gene expression that depends on whether the gene allele originated from
the mother or the father”
Imprinting is maintained by DNA methylation
❑ Methylation causes the nucleosomes to be

packed close together, thus hindering
transcription.
❑ On the other hand, acetylation of the histone

tails increases nucleosome distances, thus
promoting transcription.
Neoplastic Transformation
It is a complex multi-event and multi-stage process
The process can be divided into two requisite sequences:
1- Neoplastic conversion
2- Neoplastic development
Neoplastic Conversion
Chemical Carcinogen Progression
-DNA Reactive Promotion

-Epigenetic effect
-DNA methylation
-Histone deacetylation Neoplastic cell
DNA alteration Neoplastic

Development
Genetic and epigenetic Levels
DNA methylation in Normal and Cancer cells
DNA methylation in Normal and Cancer cells
Genes promote hypermethylation in human cancers
The list of genes that are found to be inactivated by DNA

methylation events includes genes involved in:
A- Signal transduction pathways
B- Cell cycle regulation C-Angiogenesis
D-Apoptosis E- DNA repair

P 15/P16 Methylation in cancer
❑ Aberrant methylation of cyclin dependent kinase inhibitor

P16INK4a has been frequently detected in many human cancers
❑ Hematological malignancies and head and neck squamous cell

carcinoma
❑ The differential levels of methylated P16 and P15 in plasma might

be useful markers in screening high risk population for an early
detection of cancer
Lung cancer
❑ It has been shown that a panel of markers, for aberrant methylation that detects lung cancer
at the early stages of development has been observed.
❑ This panel includes the following genes:

-P 16 -APC
-G-ST -E-cadherin
Breast cancer
❑ BRCA gene is a breast cancer susceptibility gene, that is tumor suppressor gene responsible
for both normal development and carcinogenesis in breast.
❑ BRCA1, reveals multi functional protein involved in DNA repair. Cell cycle regulation,
transcription and apoptosis
❑ Aberrant methylation of BRCA1 CPG island Promoter is associated with decreased BRCA1
mRNA in sporadic breast cancer cells.
Methylation based cancer screening
Epigenetic Chromatin Regulation

1. cytosine methylation

Histone modifications
Mechanism of histone acetylation
Acetylation of lysine residues of histone proteins
Removal of positive charge of the histones
Decreased affinity between histones and DNA
Easier access of transcription factor to promoter region

Histone Acetylation
• Acetylation of H3 or H4 leads to unfolding and increased accessibility of chromatin to

enable transcription.
• Histones are acetylated by HAT (histone acetylases) which are parts of many chromatin
remodeling and transcription complexes.
Role of Histone De-acetylation
❑ Deacetylated histones are tightly packed and less accessible to

transcription factors
❑ Histones are deacetylated by HDAC (histone de-acetylase) proteins

Histone Phosphorylation (H3)
1. Histones are phosphorylated during mitosis.
2. Histones are also phosphorylated by signal transduction pathways like

the ERK pathway in response to external signals. It is not known how
(and if) this phosphorylation contributes to gene expression.
Epigenetic chromatin regulation
1. cytosine methylation
Histone Ubiquitylation
❑ Addition and removal of Ub (a LARGE moiety) to histone tails – Functions largely

unknown in vertebrates
ubiquitylation
H2A K119: repression
H2B K120: activation
H3 and H4: DNA repair (CUL4)
de-ubiquitylation – Recrutiment of other proteins in yeast

H2A Dub (PCAF)
H2B Ubp8 (SAGA)
Functions: transcription elongation, polycomb repression

Epigenetic Diseases
Methods to Study The Epigenome
• Bisulphite sequencing
• Methylation sensitive-High resolution melting (MS-HRM)
• Microarray-based genome-wide analysis
• Chromatin immunoprecipitation (ChIP) on Chip assays

Methylation analysis study - Bisulphite sequencing
Methylation sensitive-High resolution melting (MS-HRM)
Microarray-based genome-wide analysis
Methylated DNA immunoprecipitation

(MeDIP)
- requires immunoprecipitation of
DNA using antimethylcytosine
antibody followed by hybridization
to DNA microarrays.
- requires large amounts of genomic

DNA and antibody
Study of histone modifications
• Histone modifications are studied using the chromatin immunoprecipitation

(ChIP) assay
• ChIP on chip is the high throughput form of the ChIP assay wherein the
immunoprecipitated DNA, instead of being subject to the usual PCR, is
hybridized to a microarray chip with printed oligonucleotides corresponding
to various regions of the genome.
• This helps to study the localization of a specific histone modification to

various parts of the genome
Chromatin immunoprecipitation (ChIP) on Chip assays
Different Classes of Drugs
DNMT INHIBITORS HDAC INHIBITORS
1.Nucleoside analogue inhibitors 1.Hydroxamates
2.Non nucleoside analogue 2.Cyclic tetrapeptides

inhibitors
3.Aliphatic acids
3.Antisense oligonucleotides
4.Benzamides
DNMT Inhibitor: Decitabine
Decitabine
deoxycytidine kinase
Decitabine triphosphate
Incorporated into DNA
Binds with DNMT and traps the enzyme

HDAC inhibitors: Vorinostat
• Intrinsic and acquired drug resistance remain the most unpredictable
factors affecting chemotherapy.
• DNA hypermethylation has been found to be associated with drug

resistance acquired during cancer chemotherapy and therefore, re-
expression of methylation-silenced genes resulted in increased
sensitivity to existing chemotherapy

Cancer Epigenetics

Uploaded by

Copyright:

Available Formats

Cancer Epigenetics

Uploaded by

Document Information

Original Description:

Copyright

Available Formats

Share this document

Share or Embed Document

Sharing Options

Did you find this document useful?

Is this content inappropriate?

Copyright:

Available Formats

Cancer Epigenetics

Uploaded by

Copyright:

Available Formats

Cancer Epigenetics

Structural or Functional Activity

A. Modification at the DNA level

B. Histone modification - the histone code

• Covalent addition of methyl group to 5th position of cytosine with

• Strongly represented in repetitive sequence associated with

• CG→TG mutation is propagated

CpG dinucleotides are

3 human DNA methyltransferases

• DNMT1 maintenance methyltransferases

DNMT1 has high affinity for

CpG–island methylation – how does it affect transcription?

this recruits a complex of histone deacetylases and SIN3A

induces a closed chromatin structure → gene silencing

❑ Methylation causes the nucleosomes to be

❑ On the other hand, acetylation of the histone

It is a complex multi-event and multi-stage process

The process can be divided into two requisite sequences:

Chemical Carcinogen Progression

-DNA Reactive Promotion

DNA alteration Neoplastic

The list of genes that are found to be inactivated by DNA

A- Signal transduction pathways

B- Cell cycle regulation C-Angiogenesis

D-Apoptosis E- DNA repair

❑ Aberrant methylation of cyclin dependent kinase inhibitor

❑ Hematological malignancies and head and neck squamous cell

❑ The differential levels of methylated P16 and P15 in plasma might

❑ This panel includes the following genes:

A. Modification at the DNA level

B. Histone modification - the histone code

Acetylation of lysine residues of histone proteins

Removal of positive charge of the histones

Decreased affinity between histones and DNA

Easier access of transcription factor to promoter region

• Acetylation of H3 or H4 leads to unfolding and increased accessibility of chromatin to

❑ Deacetylated histones are tightly packed and less accessible to

❑ Histones are deacetylated by HDAC (histone de-acetylase) proteins

1. Histones are phosphorylated during mitosis.

2. Histones are also phosphorylated by signal transduction pathways like

❑ Addition and removal of Ub (a LARGE moiety) to histone tails – Functions largely

de-ubiquitylation – Recrutiment of other proteins in yeast

Functions: transcription elongation, polycomb repression

• Methylation sensitive-High resolution melting (MS-HRM)

• Microarray-based genome-wide analysis

• Chromatin immunoprecipitation (ChIP) on Chip assays

Methylated DNA immunoprecipitation

- requires large amounts of genomic

• Histone modifications are studied using the chromatin immunoprecipitation

• This helps to study the localization of a specific histone modification to

DNMT INHIBITORS HDAC INHIBITORS

1.Nucleoside analogue inhibitors 1.Hydroxamates

2.Non nucleoside analogue 2.Cyclic tetrapeptides

Incorporated into DNA

Binds with DNMT and traps the enzyme

• DNA hypermethylation has been found to be associated with drug

You might also like