HRC2009, Birmingham, October 20th 2009

8/14/2019 HRC2009, Birmingham, October 20th 2009

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Genet ic Bas is o f Cl in ic a l Ar rhyt hm ias

Andrew Grac ePapw or t h Hospi t a l and Cam br idge Univers i t y

HRC2009, Bi rm ingham , Oc t ober 20 t h 2009


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The Human as an Experimental System

in Molecular Genetics

The core of genetics is to screen for mutations that causedetectable, heritable changes in biological form orfunction

Ray White, Science1988


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The Human as an Experimental System

in Molecular Genetics

The core of genetics is to screen for mutations that causedetectable, heritable changes in biological form orfunction

Human genes have historically been identified whenaffected by mutations producing genetic disease



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Long QT Syndrome


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Long QT Syndrome


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Monogenic DisordersSimple Mendelian disorders caused by ~1000 Disease-causing genes

e.g. Hypertrophic cardiomyopathy, Long-QT syndrome, Brugada

syndrome etc.


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Characterisation monogenic diseases

1. Careful phenotyping

2. Genotyping: positional cloning, candidate gene

3. Genetic modifiers

Monogenic DisordersSimple Mendelian disorders caused by ~1000 Disease-causing genes

e.g. Hypertrophic cardiomyopathy, Long-QT syndrome, Brugada

syndrome etc.


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Gene Mutation Arrhythmia Substrate

ARRHYTHMIAS AND GENETIC HEART DISEASE


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Ion channel

MutationTorsade de Pointes

Direct Relationship: Long QT SyndromeSarcomere

MutationVentricularFibrillation

Indirect Relationship e.g. Hypertrophic Cardiomyopathy


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Ion channel





StructuralDisorganization


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Ion channel





StructuralDisorganization

Ion channelMutation

Ventricular Fibrillation

Putative Direct Relationship e.g. Brugada Syndrome


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Mutations in Cardiac Ion Channel Genes responsiblefor Cardiac Arrhythmias

Priori and Napolitano Ann NY Acad Sci 2004; 1015:96-110


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Considerations specific to humans Highest order experimental system for genetics

Powerful documentation of phenotypes

Expert phenotypers (doctors), records over time- room for improvement



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Considerations specific to humans Highest order experimental system for genetics

Powerful documentation of phenotypes

Expert phenotypers (doctors), records over time- room for improvement

BUT not the best experimental system for intervention

- overlong generation time, social proscriptions tocontrolled crosses etc.



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GM mice as the functional bridge:

technical issues

Modification technique

GM > Dominant Negative


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technical issues



Choice of gene

Scn5a, RyR2> K channels


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technical issues



Choice of gene

Scn5a, RyR2> K channels

Method of Interrogation

Physiological range (HR >550) Translational (mouse/human) Comprehensive physiological,structural and molecular assays


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GM Mouse Models of Cardiac Arrhythmias(Cambridge Group)

Scn3b

B d S d ECG d V t i l A h th i


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50

100

012 36 72 96

ICD

Drugs

No Rx

Time [months]

% SurvivalLogrank = 0.0005

Survival according to treatment (n=63)

Brugada et al. Circulation 1998;97:457-60

Brugada Syndrome: ECG and Ventricular Arrhythmias

V1

V2

07.1998

01.2000


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Brugada Syndrome: SCN5A Mutations

Association first described 1998

SCN5A: TTX-insensitive sodium channel, 28 exons, 80 kb

Four homologous domains, each 6 membrane spanning segmen

Brugada mutations(> 120) are loss-of-function


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a

bprobe

E P P HBH 32

GFP NEO

c

B 3H

TK

B

a

probe

E P P HBH 3GFP NEO

c

HB

8.0 kb8.8 kb

+/+ +/- -/-Ac t i

Scn5

GFP

Proc Natl Acad Sci USA. 2002;99:6210

GM MOUSE MODEL


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Wild-type (n=37 9 pA/pF

Scn5a+/-(n=21 5 pA/pF

HALF THE SODIUM CHANNELS

Proc Natl Acad Sci USA. 2002;99:6210-and Leoni et al. submitted 2009


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S1

S1 S2

S

V V

V V

*

*

+/25 ms 25 m

WILD-TYPE MUTANT (Scn5a+/-)

SLOW CONDUCTION


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S1 S2

* * * ** *** ** * ** * S1 S2S1 S2

50

S1 S1 S1S1S1 S1 S1S2 S1S1 S1S2S1 S1S1 S1S1S1 S1 S2

50 ms

Wild-type

Scn5a+/-


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GENES AND FUNCTION (Brugada Syndrome)

SCN5AMutations

SodiumChannels

Ventricular

Arrhythmias

Slowed

Conduction


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GM Mouse Scn5aphenotypes

1. Long QT syndrome (2005-2007) - KPQ 1505-15072. Brugada/idiopathic VF (2002-2007)

3. Lev-Lengre disease/atrioventricular block (2002-2007)

4. Sinus node dysfunction (2002, 2005)5. Atrial arrhythmias (2007)

6. Dilated Cardiomyopathy - but structural change (2005-200

7. Population based risk: SCD, SIDS, drugs (2007)

Interstitial fibrosis in old Scn5a+/ mutants


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OLD WT

OLD HZ

Interstitial fibrosis in old Scn5a+/-mutants

Van Rijen et al. Circulation 2005; 112:1927-193

Intrauterine lethality in Scn5a / mutants


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EC 10.5

Intrauterine lethality in Scn5a -/-mutantsFailure of ventricular development

Scn5a +/-

Scn5a -/-

Intrauterine lethality in Scn5a / mutants


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Scn5a -/-Scn5a +/-

Proc Natl Acad Sci USA. 2002;99:6

Intrauterine lethality in Scn5a -/-mutantsFailure of ventricular development

C l Di S d


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Complex Disease Syndromes

Phenotypes not exhibiting classic Mendelianrecessive or dominant inheritance attributable to a

single gene locus

Practical difficulties

Incomplete penetrance and phenocopy

Genetic heterogeneityPolygenic inheritance

C l Di S d


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Complex Disease Syndromes

Phenotypes not exhibiting classic Mendelianrecessive or dominant inheritance attributable to a

single gene locus

Practical difficulties

Incomplete penetrance and phenocopy

Genetic heterogeneityPolygenic inheritance


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SUBSTRATE FOR ATRIAL FIBRILLATION

MUSCULATURE OF THE PULMONARY VEINS

Nathan and Eliakim; Circulation 1966; 34:412 Ho et al; Heart 2001; 265-270

ASSOCIATION STUDIES


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ASSOCIATION STUDIES

Indirect

Direct

** *

An analogy may help to explain thisphenomenon. In the mid-1960s, George Harrison,Paul McCartney, John Lennon, and Ringo Starr wereoften found together. If you looked for Harrison, there

was a high likelihood, but not complete certainty, thatyou would find the other members of the Beatles ..

SINGLE NUCLEOTIDE POLYMORPHISMS SNPs


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SINGLE NUCLEOTIDE POLYMORPHISMS - SNPs

IN THE GENOME:

~ 90% of all variants

~ 1 SNP / 1.25 kb between any two genomes

~ 2.5 million variants between two genomes

...GCTCCGTTT......GCTCTGTTT...

IN THE EXONS:

~ 60 - 120,000 variants / person

~ 20 - 40,000 non-conservative 1- 2 / gene / perso


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Genome-wide association scan with replication in other populations

Gudbjartsson et al., Nature 2007; 448:353-357


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Genome-wide association scan with replication in other populations

Gudbjartsson et al., Nature 2007; 448:353-357


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Genome-wide association scan with replication in other populations Variants adjacent to PITX2- critical function in left-right asymmetry in heart

Pitx2cplays key role in determination pulmonary myocardiumGudbjartsson et al., Nature 2007; 448:353-357


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SUBSTRATE FOR ATRIAL FIBRILLATION

MUSCULATURE OF THE PULMONARY VEINS

Nathan and Eliakim; Circulation 1966; 34:412 Ho et al; Heart 2001; 265-270



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Putative

Mutation Ventricular Arrhythmia



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Putative

Mutation Ventricular Arrhythmia

GENETIC/ENVIRONMENTAL MODIFIERS


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Founder Effect (1730) KCNQ1 (A341V)

Variable significant risk

Non-carrier(n = 154)

A341V-Carrier(n = 166)

430

msec

680msec

Circulation 2005;112:2602-1

Genetic Basis of Clinical Arrhythmias: where nex


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Genotype (high-throughput) Complex disease

Genetic modifiers



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y

Genotype (high-throughput) Complex disease

Genetic modifiers

Phenotype

Mechanisms (mice) Mechanisms (Stem Cell Models)

iPS Cells linking genes to function (2008- )


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CARDIAC

PHENOTYPING CLINICAL MUTATIONDETECTION

PHYSIOLOGY(mapping, confocal

EM, patch clamp etc.)

iPS Cells

iPS Cells linking genes to function (2008- )


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CARDIAC

PHENOTYPING CLINICAL MUTATIONDETECTION

PHYSIOLOGY(mapping, confocal

EM, patch clamp etc.)..human modelsof human disease..

Chien, Nature 2008 453:30

iPS Cells

P di t d li i l b fit f ti


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Predicted clinical benefits of a genetic

description of disease

New taxonomy of disease: mechanisms > phenotype

Predisposition phenotypes e.g. pharmacogenomics

P di t d li i l b fit f ti


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Rational drug development

Rational targeting of therapies: drugs, surgery, device



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Rational drug development

Rational targeting of therapies: drugs, surgery, device

Implementation will be problematic

Documents

HRC2009, Birmingham, October 20th 2009