2014 11 03_bioinformatics_case_studies

Bioinformatics case studies
Translating epigenetics, Personal genomics
Wim Van Criekinge
Amsterdam, 3rd November 2014
wvcrieki

Overview
Epigenetics
– Introduction
– DNA Methylation & Oncology
Translating Epigenetics
– NEXT-GENeration (Epi)genetic biomarkers
for Clinical and Prognostic Use
Personal/Recreational Genomics

Overview
Epigenetics
– Introduction
Personal Genomics

Defining Epigenetics
 Reversible changes in gene
expression/function
 Without changes in DNA
sequence
 Can be inherited from
precursor cells
 Allows to integrate intrinsic
with environmental signals
(including diet)
Genome
DNA
Epigenome
Gene Expression
Chromatin
Phenotype

Chromatin,
a Key Component of Epigenetic Regulation
Cellular DNA is packaged into a structure called
chromatin
The unit of chromatin is the nucleosome, a complex of a
histone tetramer with approx. 147 bp of DNA wound
around it
histone nucleosome
DNA
Epigenetics I Intoduction | Oncology | Biomarker
chromatin

DNA Methylation
Prevents Gene Expression
Me Me Me
 DNA methylation involves the transfer of methyl groups to cytosine residues in DNA by DNA
methyltransferases (DNMTs)
 Hypo <-> Hyper
Me
Me
Me
Me
Ac
Ac
Ac
Ac
Ac
Ac
Ac
Ac
Ac
Ac
Ac
Ac
Ac
Ac
Ac
Ac
Ac
Ac
DNMT DNMT
DNMT DNMT
Gene
expression
Gene
expression
TF
TF

Historically,
Cancer Was Considered
to be Driven Mostly by Genetic Changes
GENETIC
Example:
Replication errors
Altered
DNA sequence
X X
Altered
DNA/mRNA/proteins
Oncogenesis
Tumor
 Mutations in p53
 Activating mutations in RAS
 Mutations or amplifications of the
HER-2 gene
 Chromosomal translocations in
myeloid cells and the generation of
the BCR-ABL fusion protein

Epigenetic Changes are
Important in Causing Cancer
GENETIC EPIGENETIC
Example:
Replication errors
Example:
Chromatin modification errors
Altered
DNA sequence
Altered
DNA/mRNA/proteins
Altered
chromatin structure
Altered levels of
mRNA/proteins
X X
Oncogenesis
Tumor

Source: Schuebel et al 2007
120
100
80
60
40
20
0
Methylated Mutated
76-100 51-75 21-50 1-20
Dx
CDx
Example of Methylation
vs Mutation: Colon & Breast Cancer

Cellular programming
Evolutionary Perspective
epigenetic (meta)information = stem cells

Overview
Epigenetics
– Introduction
– Implementation

MGMT Biology
O6 Methyl-Guanine
Methyl Transferase
Essential DNA Repair Enzyme
Removes alkyl groups from damaged guanine
bases
Healthy individual:
- MGMT is an essential DNA repair enzyme
Loss of MGMT activity makes individuals susceptible
to DNA damage and prone to tumor development
Glioblastoma patient on alkylator chemotherapy:
- Patients with MGMT promoter methylation show
have longer PFS and OS with the use of alkylating
agents as chemotherapy

Enrichment Sequencing (RUO) Targeted Sequencing (IVD)
# samples
# markers
Discovery Verification Validation
3 000 000
6 000
50
5
<50
only models
and fresh frozen
> 50
All sample types
Incl. FFPE
Next Generation
Epigenetic Profiling

# samples
# markers
MethylCap_Seq
3 000 000
6 000
50
5
<50
only models
and fresh frozen
> 50
All sample types
Incl. FFPE
Next Generation

MethylCap_Seq
DNA Sheared
Immobilized
Methyl Binding Domain
Condensed Chromatin
DNA Sheared

Immobilized
Methyl binding domain
MgCl2
Next Gen Sequencing
GA Illumina: 100 million reads
MethylCap_Seq

Quality evaluation of Methyl Binding Domain based
kits for enrichment DNA-methylation sequencing
Confidential Information | ©2013 MDxHealth Inc. All rights reserved.

Data integration
Correlation tracks
expression expression
Corr =-1 Corr = 1
methylation methylation

Correlation track
in GBM @ MGMT
+1
-1

# samples
# markers
MethylCap_Seq
3 000 000
6 000
50
5
EpiHealth
<50
only models
and fresh frozen
> 50
All sample types
Incl. FFPE
Next Generation

440 cancer-related genes genes are known to
be epigenetically altered in human solid
cancers based on recent scientific and clinical
literature.

Coverage and allelic strans specific methylation signals

Dual strands accounts for genetic variant identification

# samples
# markers
MethylCap_Seq
Deep_Seq
3 000 000
6 000
50
5
EpiHealth
<50
only models
and fresh frozen
> 50
All sample types
Incl. FFPE
Methylation Specific Seq
Next Generation

GCATCGTGACTAGCGACTGATCGATGGATGCTAGCAT
25% 50% 25%
Dense methylated needed for transcriptional silencing
Are there alleles with all three positions methylated ?

unmethylated alleles
methylated alleles less methylation
more methylation
Deep Sequencing
GCATCGTGACTTACGACTGATCGATGGATGCTAGCAT

Deep Sequencing MGMT Heterogenic complexity

Data integration with
DEEP Sequencing, Infinium, Reactivation, (directional) Expression …

Whole-genome
Bisulphite seq
Enrichment
Targeted Panels
Full genome bp
RUO Clinical
Deep
Seq
Molecular Unification
genetic + epigenetic testing
109 108 107 106 105 104 103 102 101 1
E
P
I
G
E
N
E
T
I
C
Whole-genome
sequencing
Enrichment seq
(MBD, RRBS)
Enrichment seq
(Exome)
Probes
(450-27K)
Enrichment
Targeted Panels
Ultra
Deep
Sequencing

Lab for Bioinformatics and computational genomics
Instrument and Assay providers
Full genome bp
109 108 107 106 105 104 103 102 101 1
G
E
N
E
T
I
C
Whole-genome
sequencing
Enrichment seq
(Exome)
PCR
Enrichment
Targeted Panels
CLIA Lab service providers

The genome fits as an e-mail attachment

2014 11 03_bioinformatics_case_studies

The Technical Feasibility Argument
The Quality Argument
The Logistics Argument
The Price Argument

Recreational genomics

• Experimental designs are outdated by technological advances
• Genetic background (reference genome) as a concept will need to be
updated
• Traits dependent on multiple loci are “complicated”: educate and
provide tools to deal with it

• Eye color … why not the ear wax/asparagus or unibrown example
• … metabolize nutrients (newborns ?)
• … metabolize drugs in case you need it urgently ?

“several 23andMe users have reported taking the FDA’s
advice of reviewing their genetic results with their
physicians, only to find the doctors unprepared, unwilling,
or downright hostile to helping interpret the data”

my genome is too important (for me)
to leave it (only) to doctors

NXTGNT biohackerspace …

PGMv2: Personal Genomics Manifesto

Everyone should have the power and legitimacy to
be able to discover, develop and find new things
about their own genome data.
Intelligent exploration, experimentation and trial to
push the boundaries of knowledge are a basic
human right.

Personal genome data access should be
affordable to all irrespective of nationality, gender,
social background or any other circumstance.
Not having access to a personal genetic test is in
itself a new kind of discrimination.

Whether one wants to share genome data or keep it
private should be a matter of personal choice.
Whatever attitude a person has towards personal
genome privacy, it should be utterly respected.
Corporate interest can never compromise any human
right. Laws must fully protect individual human rights of
equality for every person, irrespective of predicted risks
from genetic data.

Stating that genetic tests merely provide non-clinical
information misses the point of what
personal genomics is all about.
Most genomic information is uninterpretable and
may well be meaningless. But those are not
reasons to deny it to people.
Genetic test results are not unrelated to
someone’s health, one’s ability to respond to
certain drugs and one’s ethnic ancestry.

Education in risks and opportunities for personal
genetic testing should be the primary aim of
policy makers.
Restricting access to interested people makes
no sense and it is virtually impossible to ensure.
Access to personal genomics data and tools for
its interpretation should become accessible to
everyone.

2014 11 03_bioinformatics_case_studies

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2014 11 03_bioinformatics_case_studies