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Discovering Disease Genes-Discovering Disease Genes- The Example of Schizophrenia The Example of Schizophrenia
Pippa Thomson, Medical Genetics Section, Dept of Medical Sciences, MMC, University of Edinburgh.
Importance of the illnessImportance of the illness
• Severe psychiatric
• Affects ~1% of the population
• One of the top 10 causes of disability worldwide
• Economic cost (23% Drug, 14% Hospital)
• Pharmacological rationale for treatment weak or absent
• 1/3rd patients unresponsive or experience unacceptable side effects
• Strong genetic component– concordance rate between identical twins of 60%
High Heels Cause High Heels Cause Schizophrenia Schizophrenia and 6 Other and 6 Other Outlandish Medical Theories Outlandish Medical Theories
2. High-heeled shoes cause schizophrenia. You have to wonder where some medical theories originate. Why did Swedish scientist Jarl Flensmark decide to study a connection between heeled shoes and the incidence of schizophrenia? The world may never know. But his initial research seems sound, and he has connected certain brain activity with stimulation of certain points on the feet. The spread of schizophrenia around the globe has closely followed the spread of availability of heeled shoes. Is it an eerie coincidence or a real cause for concern? Look out, men - this theory applies not only to stilettos, but to any shoe with a heel.
remedicated.com
Relative risk of developing Relative risk of developing SchizophreniaSchizophrenia
1st degree relatives
MZ~50% 50
40
30
20
10
0
no
aff
ecte
d r
elat
ive
~10-15%
~1%
general population
cou
sin
aun
t o
r u
ncl
e
gra
nd
par
ent
bro
ther
or
sis
ter
par
ent
no
n-i
den
tica
l tw
in
iden
tic
al t
win
1st degree relatives
MZ~50% 50
40
30
20
10
0
no
aff
ecte
d r
elat
ive
~10-15%
~1%
general population
cou
sin
aun
t o
r u
ncl
e
gra
nd
par
ent
bro
ther
or
sis
ter
par
ent
no
n-i
den
tica
l tw
in
iden
tic
al t
win
Environment !
Kraepelin, 1896 “As we do not know what causes the illness there cannot be a rational treatment”
Benefits of gene identificationBenefits of gene identification
• Understand aetiology
• Improved drug development & testing
• Development of definitive diagnostic tests
• Understanding of interaction with non-genetic risk factors
• Insight into normal brain development & function
Allelic architecture and Allelic architecture and mapping strategymapping strategy
Mag
nitu
de o
f ef
fect
Frequency in population
Family-based linkage studies
Association studies in populations
Unlikely to exist
Fnct. Studies
Locus Identification-problemsLocus Identification-problems
• Uncertainty in diagnostic boundaries• Non-Mendelian inheritance• Variable age of onset • Genetic heterogeneity
– Many different genes can cause the illness>1% risk world wide>phenotypic variation
• Oligogenic/polygenic causation– More than one mutant gene required to produce
phenotype
Locus identification- reducing Locus identification- reducing the problemsthe problems
• Single large families
• Avoid bilineal descent • rigorous interviews
• family history
Reduce genetic heterogeneity Significant LOD score = gene of major effect
• Reduce uncertainty of diagnosis – classify minor diagnoses as unaffected
– >1 category of affected phenotype
• Marker analysis in multiply affected family or families
• Look for co-segregation of a particular allele with phenotype
• Results expressed as a LOD score (Significant at > 3)
= log (likelihood of data, if locus & disease are linked) ---------------------------------------------------------------- (likelihood of data, if locus & disease are not linked)
• Generally a large region is identified
Linkage AnalysisLinkage Analysis
der1
der11
11
1
1
1q42
11q14
11
A balanced t(1;11)(q42;q14) A balanced t(1;11)(q42;q14) translocationtranslocation
?
recurrent major depression
minor diagnosis
unaffectedschizophrenia
bipolar affective disorder
(1;11)(q42;q14) translocation
translocation increases risk by
50-fold
t(1;11) co-segregates with t(1;11) co-segregates with major mental illnessmajor mental illness
• Genetic association studies seek to relate variation in human DNA sequence with a disease or trait
• Association method provides greater power to detect common genetic variants conferring susceptibility to complex phenotypes
• Estimates population attributable risk (effect size)
• Controls should match cases and be a representative sample of the population.
100’s Individuals = 1% Schizophrenia 100’s Individuals = 100% Schizophrenia
Case-control association Case-control association studiesstudies
• Comparison of frequencies of polymorphisms between populations of cases and controls (usually a simple chi-square test or logistic regression)
• Polymorphism studied can be directly responsible for the defect frequency of cases >>> controls
• Polymorphism studied can be in linkage disequilibrium with the mutation responsible for the disease %T cases >> controls
• Association studies can be conducted for candidate genes, or through a whole region or across the whole genome (WTCCC)
p
Mb
T
A
G
C
SNPs are genotyped in parent-offspring trios, initially in CEPH trios.This can be used to identify SNPs that co-segregate (i.e. are in linkage disequilibrium) versus those that segregate independently.
A subset of SNPs can therefore be chosen that best represent the genetic diversity in a region/gene, reducing the costs of genotyping.
Summary of genotyped SNPs: Populations CEU CHB+JPT YRITotal Non-Redundant 3,204,709 3,244,897 3,150,433
International HapMap projectInternational HapMap project
http://www.hapmap.org/
Region of interest
HapMap genotyped SNPs
Known SNPs*
Known genes in the regions
Linkage Disequilibrium(LD)
*http://www.ncbi.nlm.nih.gov/SNP/
Tagging SNP selection Tagging SNP selection
Proportion of haplotype diversity explained : SNPs 1-23 - 97% SNPs 24-46 - 98%
DISC2
2 3 4 5 6 7 8 9 10 1112 13
DISC1
TRANSLOCATION
LOD=7.1, SCOTLAND
Genetic evidence implicatingGenetic evidence implicating DISC1DISC1 in psychiatric illness in psychiatric illness
1
LOD=2, BRITAIN & ICELAND (Curtis et al 2003)
D1S251
LOD=1, TAIWAN (Hwu et al 2003)
SCZ
BPAD
HAPLOTYPE
p=0.00024, FINLAND (Hennah et al 2003)
SCZ & SCZAFF
SCZ & BPAD & MDD
LOD=3.21, FINLAND (Ekelund et al 2001)
SCZD1S2709
p=0.000027, North-America (Hodgkinson et al 2004)
rs6675281
SCZAFF
SCOTLAND
SCZ, BPADHAPLOTYPE
p=0.0044, p=0.0016
Table 1. Summary of current evidence supporting several of the more promising genes implicated in schizophrenia, bipolar disorder, and mixed bipolar-psychosis phenotypes
Craddock et al., SCZ Bulletin, 2006
>130 genes implicated
DISC1 interactomeDISC1 interactome protein-protein interactionsprotein-protein interactions
Chris Carter, http://www.polygenicpathways.co.uk/disc11_vml.htm
• ENU generated mouse mutants
• Two independent lines with missense mutations in DISC1 exon 2
– Q31L (Glutamine-Leucine) > Q- hydrophillic; L – hydrophobic
– L100P (Leucine-Proline)> Predicted to cause transition in polypeptide chain direction
• Normal levels of DISC1 protein in brain
• L100P line models schizophrenia; Q31L, depression
Effects of altered DISC1 on Effects of altered DISC1 on gene expressiongene expression
Clapcote et al., Neuron. 2007 May 3;54(3):387-402.
Phenotype 31L/31L 100P/100PAnxiety (elevated plus-maze) = =Horizontal activity = >>Vertical activity = >Prepulse inhibition (PPI) < <<Acoustic startle response = <Startle reactivity = <Latent inhibition (LI) << <<Working memory (T maze) < <<Spatial learning and memory (Morris water maze) = =Forced swim immobility (FST) > =Sociability and social novelty < =Sucrose consumption < =Brain volume < <<PDE4B activity << =
Drug Treatment 31L/31L 100P/100PPPI Clozapine = +
Bupropion +++ =
Schizophrenia
Depression
Effects of Altered DISC1 on Effects of Altered DISC1 on Behaviour (Behaviour (How do you know How do you know if a mouse is schizophrenic?if a mouse is schizophrenic?))
Effects of altered DISC1 on Effects of altered DISC1 on gene expressiongene expression
•Samples collected and microarray study ongoing –Mutated lines vs background strain (C57/BL6)
–47,000 transcripts
–Hippocampus
–Adult and embryonic stage- Microarray
–Confirmation/Investigation of changes
–Series of embryonic; postnatal and adult stages
–Drug treated adult mice
Detect disrupted pathways
Resequencing, SNP detection, genome comparisons, gene expression, transcription factor studies, small RNA analysis
Individual genomes – All SNPs in each individual
Currently :
Using the Illumina 1G to sequence genes in the DISC1 pathway. Total sequence read 3.5megabases in 1200 individuals
• Identify coding and non-coding polymorphisms• Mutation detection• Detection of variants in conserved regions• Detection of variants affecting binding of transcription factors
Whole genome sequencingWhole genome sequencing
Psychiatric Genetics-Psychiatric Genetics-Unanswered QuestionsUnanswered Questions
How many susceptibility genes are there? What is their function? Is function conserved across species? Can we relate gene (dys)function to mental (dis)order? Do gene variants predict risk, course, outcome and response to treatment? Will gene discovery lead to drug discovery? How do genes and environment interact? How and when will the patient benefit?
DISC1
Kirsty MillarShaun MackieFumiaki OgawaJennifer ChubbBecky CarlyleNick BradshawSheila Christie
Steve ClapcoteKathy EvansSarah Brown
William Hennah
Medical Genetics
Prof David PorteousProf Douglas BlackwoodWalter MuirBen Pickard
Other collaborators
DISC1 ConsortiumWellcome Trust CRFIllumina, San DiegoCold Spring Harbor laboratories
AcknowledgementsAcknowledgements