Download Molecular Diagnostics

Molecular Diagnostics :
Hype or Hope ?
Patrick Willems
GENDIA, Antwerp, Belgium
We now know how God
wrote the book of life
Bill Clinton
But do we know
how to read the book ?
Genetic Diagnostics
• Cytogenetic tests
• FISH
• Molecular tests
Molecular Diagnostics
-
Diagnosis of infectious diseases
-
Genetic identification
-
Diagnosis of genetic diseases
Diagnosis of infectious diseases
HPV
Chlamydia
Hepatitis
HIV
Toxoplasmosis
Genetic Identification
- Paternity Testing
- Forensics
Paternity Testing
Forensic testing
Diagnosis of genetic diseases
- Somatic rearrangements in cancer
- Genetic risk factors
-
Pharmacogenetics
- Mutations in monogenic diseases
Rearrangements in Cancer Cells
Chromosomal breaks produce fusion genes
These cause leukemias and lymphomas
Diagnosis determines treatment and prognosis
Rearrangements in Cancer Cells
Lymphocytic Leukemia
t(9;22)
t(12;21)
t(1;19)
t(4;11):
:
BCR - ABL
:
TEL - AML1
:
E2A - PBX1
MLL - AF4
Myeloid Leukemia
Inv(16)
t(8;22)
t(9;22)
:
:
:
CBF - MYH11
AML - ETO
BCR - ABL
Diagnosis of genetic diseases
- Somatic rearrangements in cancer
- Genetic risk factors
-
Pharmacogenetics
- Mutations in monogenic diseases
Genetic Risk Factors
Monogenic diseases are caused by a
deleterious mutation in a single gene:
Disease-causing mutations
Multifactorial diseases are caused by a
combination of variations in multiple genes:
Genetic Risk Factors
Genetic Risk Factors
Deep venous thrombosis
Cardiovascular disease
Alzheimer disease
Osteoporosis
Genetic Risk Factors
Most single risk factors
have NO clinical significance
in individual patients
Genetic Risk Factors
Deep venous thrombosis
Factor V
Factor II
MTHFR
Diagnosis of genetic diseases
- Somatic rearrangements in cancer
- Genetic risk factors
-
Pharmacogenetics
- Mutations in monogenic diseases
Pharmacogenetic tests
• Drug specificity
• Drug efficacity - toxicity
Drug specificity
Herceptin : HER2
Tyrosine kinase inhibitors
BCR / ABL
KIT
PDGFR A/B
EGFR
Drug efficacity / toxicity
Cytochromes
CYP2D6
CYP2C9
CYP2C19
Diagnosis of genetic diseases
- Somatic rearrangements in cancer
- Genetic risk factors
-
Pharmacogenetics
- Mutations in monogenic diseases
Diagnostic bottle necks
•
Number of diseases
•
Nature of disease mutation
•
Technology
•
Cost
•
Number of samples
•
Organisation
Monogenic Diseases
> 4.000 monogenic diseases
> 2.000 disease genes isolated
Gene testing
• Most countries : limited number
(< 50 genes)
• Few countries
: large number
(300-500 genes)
• Nowhere
: network complete availability
(> 1000 genes)
Diagnostic bottle necks
•
Number of diseases
•
Nature of disease mutation
•
Technology
•
Cost
•
Number of samples
•
Organisation
Disease Mutations
Easy tests : Single - common mutations
Difficult tests : Private mutations
Disease Mutations
Single mutations
Fragile X
Sickle Cell Anemia
Common mutations
Deafness
Hemochromatosis
Panel of mutations
Cystic Fibrosis
Private mutations
Breast Cancer
Colorectal cancer
Easy tests
Disease
Gene
Mutation
Fragile X
FMR1
Repeat
FRAXE
FMR2
Repeat
Friedreich ataxia
FRDA
Repeat
Haw River
DRPLA
Repeat
Huntington type 1
HD
Repeat
Kennedy
AR
Repeat
Myotonic dystrophy type 1
DMPK
Repeat
Spinocerebellar ataxia
SCA1,2, 3, 6, 7, 8,10, 12,17
Repeat
Alpha 1 antitrypsin
PI
2 common mutations
Charcot-Marie-Tooth Type 1A
PMP22
1 common mutation
Cystic fibrosis
CFTR
Common mutations
Deafness
GJB2
1 common mutation
Hemochromatosis type1
HFE
2 common mutations
Hereditary neuropathy (HNPP)
PMP22
1 common mutation
Sickle cell anemia
HBB
1 common mutation
Spinal muscular atrophy
SMN1
1 common mutation
Beta thalassemia
HBB
1 exon
Difficult tests
Disease
Gene
Mutation
Breast cancer
BRCA1
Private
BRCA2
Private
MLH1
Private
MSH2
Private
MSH6
Private
Colon cancer
BRCA
testing
BRCA1 : 23 exonen, 1863 AA, 6.200 bp
BRCA2 : 28 exonen, 3418 AA, 10.300 bp
Totaal : > 17.000 bp sequence
Diagnostic bottle necks
•
Number of diseases
•
Nature of disease mutation
•
Technology
•
Cost
•
Number of samples
•
Organisation
Mutation Detection
1. Point mutations, frame shifts :
A. Sequencing
B. WAVE
2. Deletions : MLPA
Diagnostic bottle necks
•
Number of diseases
•
Nature of disease mutation
•
Technology
•
Cost
•
Number of samples
•
Organisation
Cost
Single mutations
: cheap (200 E)
Prevalent mutations : cheap (300 E)
Panel of mutations
: moderate (300 E)
Private mutations
: expensive (1000 E)
Cost
• Socioeconomic situation
• Social security
• Reimbursement by insurance
Diagnostic bottle necks
•
Number of diseases
•
Nature of disease mutation
•
Technology
•
Cost
•
Number of samples
•
Organisation
Common
Genetic Diseases
?
Common genetic diseases
Disease
Frequency Mutation Genes
Mutations
Conclusion
Hemochromatosis
1 / 600
1 / 400
HFE
2 common mutations
Easy / cheap
Breast Cancer
1 / 20
1 / 500
BRCA1
BRCA2
Mutations in 23 exons
Mutations in 28 exons
Complicated /
expensive
Hypercholesterolemia
1 / 500
1 / 750
LDLR
Mutations in 16 exons
Complicated /
expensive
Colorectal Cancer
1 / 25
1 / 1.000
MLH1
MSH2
MSH6
APC
MUTYH
Mutations in 19 exons
Mutations in 16 exons
Mutations in 10 exons
Mutations in 15 exons
Mutations in 16 exons
Complicated /
expensive
Cystic fibrosis
1 / 2.500
1 / 2.500
CFTR
Common mutations
Easy / cheap
Prelingual deafness
1 / 1.500
1 / 4.000
GJB2
1 common mutation
Easy / cheap
Fragile X syndrome
1 / 5.000
1 / 5.000
FMR1
Only 1 mutation
Easy / cheap
SMA
1 / 10.000
1 / 10.000
SMN1
Only 1 mutation
Easy / cheap
Beta Thalassemia
variable
variable
HBB
Only 1 exon
Easy / cheap
Most frequent DNA tests
•
Thalassemia
•
Cystic fibrosis
•
•
Breast cancer
Colorectal cancer
•
•
•
FRAXE
SCA
F5 Leiden
Usual portfolio of DNA tests
• Easy tests
• Common tests
• Research tests
Genetic testing in Europe
• inhabitants per country
: 10 million
• births per year
: 100.000
• disease frequency
: 1 on 10.000
• new patients per year
: 10
• genetic labs
: 10
New patients per lab per year: 1
Diagnostic bottle necks
•
Number of diseases
•
Nature of disease mutation
•
Technology
•
Cost
•
Number of samples
•
Organisation
Current Organisation
• Small local labs : small portfolio’s ( < 50 tests )
• Same spectrum of tests : common + easy tests
• Majority academic labs : research -diagnostic setting
• Many academic labs give up diagnostic testing
• No (inter)national network
Diagnostic bottle necks
•
Number of diseases
•
Nature of disease mutation
•
Technology
•
Cost
•
Number of samples
•
Organisation
Gene testing
• Unreliable
• Expensive
• Slow
Unreliable
10 % mistakes in easy tests such as CF
Nature Genetics 2000; 25: 259 - 260
Expensive
RESEARCH
DIAGNOSTICS
1 genome
< 1000 USD
1 gene
200 – 5.000 USD
Ratio : 25.000
Slow
RESEARCH
DIAGNOSTICS
100 genomes
in 10 days
1 gene
in 100 days
Ratio 25 million
Message in a bottle
• Many different tests
• Many uncommon tests
• Many esoteric tests
• Many expensive tests
• International network needed
Mission
A global network of diagnostic labs
• Large portfolio
• Reliable
• Fast
• Affordable
GENDIA
www.GENDIA.net
GENetic DIAgnostic Network
The GENDIA network
GENDIA Network
1000 Referral labs
1 Central lab
100
Test labs
Advantages GENDIA
• 1 lab to send samples to
• 1 lab to get results from
• > 2.000 genetic tests
• Large portfolio
• Best first selection of test
• Best Reflex testing
Looking into the future
Next generation sequencing
Sequencing power : billion bp / day
Will rapidly multiply
Cost : 100.000 Euros
Will rapidly decrease to 1000 Euro
Whole genome sequencing of Watson and Venter
Sequencing all patients
DNA Sequencing
1980-1990
Radio - gel
Thousand bp / day
1990-2005
Fluorescent - capillary
Million bp / day
> 2005
Next generation
Billion bp / day