Recent Genetic Advances in Cardiovascular Disease Linnea M. Baudhuin, Ph.D. Mayo Clinic Dec. 6,...

Recent Genetic Advances Recent Genetic Advances in Cardiovascular Diseasein Cardiovascular Disease

Linnea M. Baudhuin, Ph.D.Linnea M. Baudhuin, Ph.D.

Mayo ClinicMayo Clinic

Dec. 6, 2008Dec. 6, 2008

DISCLOSUREDISCLOSUREInformationInformation

Relevant Financial RelationshipsNone

Off Label UsageNone

Mendelian genetics Autosomal Dominant Hypercholesterolemia

LDLR, APOB, PCSK9

Familial Thoracic Aortic Aneurysm and Dissection syndromes:

FBN1, TGFBR1, TGFBR2, ACTA2, MYH11, NOTCH

Genetic markers for CAD Chromosome 9p21/ANRIL KIF6

Pharmacogenetics Warfarin sensitivity Statin efficacy

Recent Developments in CV Recent Developments in CV GeneticsGenetics

Unable to predict significant portion of CAD 35% of CAD occurs in population with TC<200 mg/dL

“CAD could be eliminated in 21st century by treating all risk factors” R. Roberts, Genetics of Premature Myocardial

Infarction, Curr Ather Reports, 2008, 10: 186-193

Atherosclerotic CVD has strong heritable component

Estimates as high as >50%

Families with early CAD (14% of population) account for 72% of early CAD cases

Most of the risk factors have predominant genetic component Need for identifying genetic modifiers and their

function

Coronary Artery DiseaseCoronary Artery Disease

Complex Genetics of CADComplex Genetics of CAD

Minority are single-gene disorders FH, FDB, ARH, Tangier,

Sitosterolemia, etc. What about the majority of CAD? What’s going on genetically?

Early stages of Early stages of atherosclerosisatherosclerosis

Late stages of Late stages of atherosclerosisatherosclerosis

Lusis et al., Ann Rev Gen Hum Gen, 2004, 5:189-218

Lusis A, Trends Cardio

Med, 2003; 13:309-16.

Genes Genes contributincontributin

g to CAD g to CAD susceptibilisusceptibili

tyty

How can we identify genetic How can we identify genetic markers for complex disease?markers for complex disease?

Family studies Genetic variants that segregate with disease in affected

vs. non-affected family members Linkage analysis

Population studies Genetic variants that segregate with disease in cases

vs. controls Candidate genes Localized genomic regions Genome-wide

Association studies

CAD Linkage StudiesCAD Linkage Studies At least 7 family studiesAt least 7 family studies

Occurrence of MI or subclinical Occurrence of MI or subclinical atherosclerosisatherosclerosis

Linkage: Chr 1, 2, 3, 13, 14, 15, 16, XLinkage: Chr 1, 2, 3, 13, 14, 15, 16, X Lack of replicationLack of replication

Genes identified:Genes identified: MEF2A (Chr 15q26)MEF2A (Chr 15q26) ALOX5APALOX5AP LTA4LTA4

Generally single gene/single polymorphismGenerally single gene/single polymorphism

Small cohortsSmall cohorts

Inability to replicate findings in additional Inability to replicate findings in additional studiesstudies

Genetic analysis of 103 candidate genes Genetic analysis of 103 candidate genes (n=1400, with follow-up n= 806) unable to (n=1400, with follow-up n= 806) unable to replicate findings (Pare et al, Am Hum Genet, replicate findings (Pare et al, Am Hum Genet, 2007, 80:673-682) 2007, 80:673-682)

Similar results in n=811 ACS patients, 70 genes Similar results in n=811 ACS patients, 70 genes (Morgan et al, JAMA, 2007, 297: 1551-1561(Morgan et al, JAMA, 2007, 297: 1551-1561

Candidate Gene Association Candidate Gene Association Analyses in CADAnalyses in CAD

Inadequately poweredInadequately powered

Phenotypic heterogeneity and imprecise definition of phenotypesPhenotypic heterogeneity and imprecise definition of phenotypes

Bias (e.g. population stratification)Bias (e.g. population stratification)

Multiple genes each contribute to small percentage of phenotype Multiple genes each contribute to small percentage of phenotype (<5-10%)(<5-10%) A single gene is insufficient to induce diseaseA single gene is insufficient to induce disease

Information from multiple polymorphisms should be integrated to Information from multiple polymorphisms should be integrated to become clinically usefulbecome clinically useful Multiple possible combinations of genotypesMultiple possible combinations of genotypes

Population and disease heterogeneityPopulation and disease heterogeneity

Why are the results of Linkage and Why are the results of Linkage and Candidate Gene studies so Candidate Gene studies so

disappointing?disappointing?

What about What about Genome Wide Genome Wide Association Association

Studies?Studies?

Genome-wide association Genome-wide association studiesstudies

Previously: single SNP/single cohort Previously: single SNP/single cohort associations with little replicationassociations with little replication

Now: large international consortium Now: large international consortium studies of tens of thousands of studies of tens of thousands of individuals and hundreds of thousands individuals and hundreds of thousands of SNPsof SNPs

TechnologyTechnology Ultra high-throughput genotyping with Ultra high-throughput genotyping with

ability to analyze 500K to >1M SNPs in ability to analyze 500K to >1M SNPs in each individualeach individual

Topol, EJ et al., JAMA, 2007; 298:218-221

Multiple GWASs confirming Multiple GWASs confirming 9p21 in Caucasian populations9p21 in Caucasian populations

McPherson et al, Science (316), June 8 2007, 1488-91McPherson et al, Science (316), June 8 2007, 1488-91 Initial study, 322 cases vs 312 controls; 73K SNPsInitial study, 322 cases vs 312 controls; 73K SNPs

Confirmed in 5 independent Caucasian populations Confirmed in 5 independent Caucasian populations (n=23,000)(n=23,000)

Helgadottir et al, Science (316), June 8 2007, 1491-3Helgadottir et al, Science (316), June 8 2007, 1491-3 Initial study, 1607 cases vs 6728 controls; 306K SNPsInitial study, 1607 cases vs 6728 controls; 306K SNPs

Replicated in Replicated in 665 cases and 3533 controls665 cases and 3533 controls

Three other case control populations Three other case control populations Philadelphia, Atlanta, & Durham, NCPhiladelphia, Atlanta, & Durham, NC

4587 cases, 12,767 controls4587 cases, 12,767 controls

Wellcome Trust Case Consortium, Nature, 447:661-Wellcome Trust Case Consortium, Nature, 447:661-678, 2007678, 2007

Samani et al, NEJM (357), Aug 2 2007, 443-53Samani et al, NEJM (357), Aug 2 2007, 443-53

PROCARDIS Consortium (Broadbent et al, Hum Mol PROCARDIS Consortium (Broadbent et al, Hum Mol Gen, 2008, 17)Gen, 2008, 17)

Chromosome 9p21Chromosome 9p21 9p21 confined to 58K bp region9p21 confined to 58K bp region

Risk associated with MI and CAD independent of Risk associated with MI and CAD independent of known risk factorsknown risk factors

50% of individuals heterozygous for allele50% of individuals heterozygous for allele 15-20% increased risk of CAD15-20% increased risk of CAD

25% of individuals homozygous for allele25% of individuals homozygous for allele 30-40% increased risk of CAD30-40% increased risk of CAD

Odds ratio Odds ratio Heterozygous: 1.3-1.5Heterozygous: 1.3-1.5

Homozygous: 1.8-2.1Homozygous: 1.8-2.1

9p21 and ANRIL9p21 and ANRIL High-risk CAD 9p21 haplotype overlaps with exons High-risk CAD 9p21 haplotype overlaps with exons

13-19 of ANRIL13-19 of ANRIL

Newly annotated gene, encodes large antisense non-Newly annotated gene, encodes large antisense non-coding RNA (ncRNA)coding RNA (ncRNA)

Identified through deletion analysis of French family Identified through deletion analysis of French family with hereditary melanoma-neural system tumorswith hereditary melanoma-neural system tumors

Expressed in atheromatous human vesselsExpressed in atheromatous human vessels Comparable cell type profiles to atherosclerotic arteriesComparable cell type profiles to atherosclerotic arteries

Expressed in vascular endothelial cells, monocyte-Expressed in vascular endothelial cells, monocyte-derived macrophages, and coronary smooth muscle derived macrophages, and coronary smooth muscle cellscells

Very little known about functionVery little known about function

Gene class is thought to be involved in transcriptional Gene class is thought to be involved in transcriptional controlcontrol Broadbent, et al, Human Mol Gen, 2008, 17: 806-14

KIF6KIF6 Variant and CAD Variant and CAD

CARECARE: Cholesterol and Recurrent Events : Cholesterol and Recurrent Events

WOSCOPSWOSCOPS: West of Scotland Coronary Prevention Study: West of Scotland Coronary Prevention Study

Association of Association of KIF6KIF6 with with CADCAD

2 Prospective Trials of 2 Prospective Trials of PravastatinPravastatin

CARE: Cholesterol and Recurrent EventsCARE: Cholesterol and Recurrent Events Secondary prevention in patients with a prior MI Secondary prevention in patients with a prior MI 40 mg pravastatin vs placebo40 mg pravastatin vs placebo 87% men and 13% women87% men and 13% women LDL-C 115 to 174 mg/dL at baselineLDL-C 115 to 174 mg/dL at baseline 2,697 participants studied2,697 participants studied

WOSCOPS: West of Scotland Coronary Prevention StudyWOSCOPS: West of Scotland Coronary Prevention Study Primary preventionPrimary prevention 40 mg pravastatin vs placebo40 mg pravastatin vs placebo Men aged 45 to 64Men aged 45 to 64 LDL-C 174 to 232 mg/dL at baselineLDL-C 174 to 232 mg/dL at baseline 1,527 participants studied1,527 participants studied

Risk of CAD in CARE Placebo Risk of CAD in CARE Placebo ArmArm

KIF6 Variant and Traditional Risk KIF6 Variant and Traditional Risk FactorsFactors

KIF6 Variant

Age ≥ 55

Smoking

LDL-C ≥130

HDL-C < 40

Hypertension

Diabetes

Magnitude of risk predicted by KIF6 variant was: Similar to that of traditional risk factors

Independent of traditional risk factors

Adjusted Hazard Ratio0.5 21 1.5 2.5 3

Risk of CAD in WOSCOPS Placebo Risk of CAD in WOSCOPS Placebo ArmArm

KIF6 Variant and Traditional Risk FactorsKIF6 Variant and Traditional Risk Factors

KIF6 Variant

LDL-C ≥ 189*

HDL-C < 40

Hypertension

Diabetes

Magnitude of risk predicted by KIF6 variant was: Similar to that of traditional risk factors

Independent of traditional risk factors

Adjusted Risk Ratio

Case and control patients were matched for age and smoking; all were men

* Median level in placebo arm

0.5 21 1.5 2.5 3

KIF6KIF6 variant predicts risk of CAD variant predicts risk of CAD Up to 55% increased risk in untreated populationsUp to 55% increased risk in untreated populations

Hazard Ratio*

*Adjusted for traditional risk factors

Untreated patients

Some treatment

KIF6KIF6 Variant is Associated Variant is Associated with CAD with CAD

5 Prospective Studies5 Prospective Studies

ARIC

WHS

CHS

WOSCOPS

CARE

0.5 21 1.5 2.5


LDLR, APOB, PCSK9



Genetic markers for CAD Chromosome 9p21/ANRIL KIF6

Pharmacogenetics Warfarin sensitivity Statin efficacy


PROVE IT - TIMI 22 studyPROVE IT - TIMI 22 study Pravastatin or Atorvastatin Evaluation and Infection Pravastatin or Atorvastatin Evaluation and Infection

Therapy, Thrombolysis in MI Therapy, Thrombolysis in MI

Patients hospitalized within 10 days after an acute Patients hospitalized within 10 days after an acute coronary syndromecoronary syndrome

0

2

4

6

8

10

12

14

Prevention of CHD Events by Prevention of CHD Events by Statins Statins

Carriers of the KIF6 Variant Benefit the Carriers of the KIF6 Variant Benefit the MostMost

KIF6KIF6 carriers received significant risk reduction from atorvastatin in carriers received significant risk reduction from atorvastatin in PROVE IT, suggesting PROVE IT, suggesting KIF6 may alsoKIF6 may also predict response with other predict response with other statinsstatins

KIF6KIF6 carriers also received significantly greater benefit than carriers also received significantly greater benefit than noncarriers (p=0.003 for WOSCOPS and 0.018 for PROVE IT)noncarriers (p=0.003 for WOSCOPS and 0.018 for PROVE IT)

3.5% 10.0%6.2%0.1% 0.8%5.5%

WOSCOPS PROVE ITPravastatin 40 mg

All CarriersNon-

carriers

Atorvastatin 80 mg

All CarriersNon-

carriers

Absolute Risk Reduction

(%)

Dea

th o

r m

ajo

r C

V e

ven

tsStatin Intensity and CHD Event ReductionAccording to KIF6 719Arg Carrier Status

KIF6 carriers received greater benefit from 80 mg atorvastatin, compared with 40 mg pravastatin, than did noncarriers

Number Needed to Treat with atorvastatin for 2 years to prevent 1 event was 10 for KIF6 carriers and 125 for noncarriers

KIF6 Carriers Noncarriers

Months of follow-up Months of follow-up0 3 6 9 12 15 18 21 24 27 30

0

10

20

30

40

0 3 6 9 12 15 18 21 24 27 300

10

20

30

40

p≤0.001P=1.0

Pravastatin

Atorvastatin

Pravastatin

Atorvastatin

KIF6KIF6 Summary SummaryRisk for CAD and Risk Reduction from Risk for CAD and Risk Reduction from

StatinsStatins

KIF6 carriers, ~60% of the population, have greater risk for CAD Up to 55% increased risk of CAD

Independent of traditional risk factors, but similar in magnitude of risk

Associated with CAD risk in men (WOSCOPS), women (WHS), the middle aged (ARIC) and the elderly (CHS)

KIF6 carriers treated with pravastatin or atorvastatin had a greater reduction of coronary events than noncarriers

Warfarin PGxWarfarin PGx Warfarin is commonly prescribed drug Variants in CYP2C9 and VKORC1 account

for high percentage of variability of warfarin response

FDA relabeled warfarin in Aug. 2007 to encourage pharmacogenetic testing

Utility still remains low Controversial topic Unanswered questions

Warfarin PGx: Ready for Prime Warfarin PGx: Ready for Prime Time?Time?

How is the information applied to clinical practice? Can genetic testing be used to determine the right

warfarin dosage? Does it add significantly to the information discerned

by careful INR monitoring and other factors (e.g. age, underlying disease state, drug-drug interactions)?

Will it really reduce clinical complications? Will it result in shorter time to stable INR? Is it cost-effective?

Should genotyping be ordered on all patients taking Warfarin?

Can the information be obtained in a timely fashion?

WarfarinWarfarin Widely prescribed anticoagulantWidely prescribed anticoagulant 1212thth most commonly prescribed drug most commonly prescribed drug

Wysowski, D. K. et al. Arch Intern Med 2007;167:1414-1419.

0

5000000

10000000

15000000

20000000

25000000

30000000

35000000

Annual Number of

Prescriptions

1998 1999 2000 2001 2002 2003 2004

Year

Annual Number of Outpatient Warfarin Prescriptions, 1998-2004 45% increase from

1998 to 2004

WarfarinWarfarin Challenges in regulating warfarin dosing Prothrombin time (INR) must remain in

narrow therapeutic range Elevated INR: risk for major bleeding

complications Risk for serious bleeding increases with INR > 4.0 Most likely to occur within first few weeks of

initiating treatment Subtherapeutic INR: thrombotic

complications

Budnitz et al, Ann Intern Med 2007;147:755-765.

Warfarin is the Most Commonly Warfarin is the Most Commonly Implicated Medication in U.S. ED VisitsImplicated Medication in U.S. ED Visits

0 5 10 15 20

WarfarinInsulinAspirin

ClopodigrelDigoxin

MetforminGlyburide

VicodinPhenytoinGlipizide

LevofloxacinLisinopril

BactrimFurosemide

Frequency

180

750780

390

180

7030 25 120

100

200

300

400

500

600

700

800

Nu

mb

er o

f P

atie

nts

0.5

to 2

3 to

4

5 to

6

7 to

8

9 to

10

11 to

12

13 to

14

15 to

16

17 to

20

Daily Dose, mg

Variability in Warfarin Dose Requirements, N = 2305

James AH. J Clin Path 1992;45:704-06

Wide Range in Warfarin Dose Wide Range in Warfarin Dose RequirementsRequirements

Warfarin Resistance

Warfarin Sensitivity

Sources of variability in Sources of variability in Warfarin dosingWarfarin dosing

VKORC1 25%

CYP2C9 17%

CYP4F2 2%

Weight 9%

Age 7%

Other 28%

Drugs 12%

Warfarin PharmacologyWarfarin Pharmacology Racemic mixture of R- and S-enantiomersRacemic mixture of R- and S-enantiomers

S-Warfarin approx. 7-10X as potent as R-S-Warfarin approx. 7-10X as potent as R- Majority of Majority of in vivoin vivo activity of warfarin resides activity of warfarin resides

in S-warfarinin S-warfarin Metabolized mainly through CYP2C9Metabolized mainly through CYP2C9

Targets VKORC1 (Vitamin K epoxide Targets VKORC1 (Vitamin K epoxide reductase complex subunit 1)reductase complex subunit 1) Interferes with recycling of Vit K in the liverInterferes with recycling of Vit K in the liver Reduced activation of clotting factorsReduced activation of clotting factors

Pharmacokinetics Pharmacodynamics

CYP2C9

CYP2C19CYP3A4CYP1A1CYP1A2CYP2C8CYP2C9

4-hydroxywarfarin

6-hydroxywarfarin

7-hydroxywarfarin

8-hydroxywarfarin

10-hydroxywarfarin

VKOR

(hepatocyte) (hepatocyte)

S-WarfarinR-Warfarin

6-hydroxywarfarin

7-hydroxywarfarinVitamin K (oxidized)

Inactive Vitamin K-dependent clotting

factors

Active Vitamin K-dependent clotting

factors

Vitamin K (reduced)

S-WarfarinR-Warfarin

inactive

GGCX

Elimination

Warfarin PGxWarfarin PGx CYP2C9CYP2C9

CYP2C9*2 variantCYP2C9*2 variant Arg144CysArg144Cys 30% decrease in enzymatic activity30% decrease in enzymatic activity

CYP2C9*3 variantCYP2C9*3 variant Ile359LeuIle359Leu 70%-95% decreased enzymatic activity70%-95% decreased enzymatic activity

VKORC1VKORC1 Promoter SNP -1639G>A Promoter SNP -1639G>A 44% decrease in promoter activity44% decrease in promoter activity

Allele FrequenciesAllele Frequencies

CYP2CCYP2C9 *29 *2

CYP2CCYP2C9*39*3

VKORC1VKORC1

--1639G>1639G>

AA

CaucasianCaucasian 8-13%8-13% 6-10%6-10% 42%42%

AsianAsian 2-5%2-5% <1%<1% 89%89%

African African AmericanAmerican <1%<1% 1-4%1-4% 8%8%

Genotype-guided Warfarin Genotype-guided Warfarin DosingDosingCYP2C9 GenotypeCYP2C9 Genotype

VKORC1VKORC1 RapidRapid IntermediIntermediateate

SlowSlow Very Very SlowSlow

GenotypGenotypee

*1/*1*1/*1 *1/*2*1/*2 *1/*3*1/*3 *2/*2*2/*2 *2/*3*2/*3 *3/*3*3/*3

Low Low SensitivitySensitivity

(GG)(GG)140%140% 113%113% 94%94% 92%92% 75%75% 63%63%

Med Med SensitivitySensitivity

(AG)(AG)100%100% 81%81% 67%67% 67%67% 54%54% 46%46%

High High SensitivitySensitivity

(AA)(AA)73%73% 58%58% 48%48% 48%48% 40%40% 33%33%McClain, et al., Gen Med, 2008, 10(2): 89-98

www.warfarindosing.org

Can Pharmacogenomics Can Pharmacogenomics Assist Warfarin Dosing?Assist Warfarin Dosing?

Caraco, Clin Pharm Ther, 2008

Warfarin DosingInitiation Stabilization

185 Patients

PGx-Informed(CYP2C9 only)

Algorithm Guided(Ageno et al, 2000)

Time to First Therapeutic INR

PGx-informedControls

P<0.001*

Pro

port

ion

of

pati

en

ts

wit

h I

NR

>2

Days


0 5 10 15 20 25

0.0

0.2

0.4

0.6

0.8

1.0

CYP2C9 Genotype-guided CYP2C9 Genotype-guided Warfarin Prescribing Warfarin Prescribing Efficacy and SafetyEfficacy and Safety

*Time to therapeutic INR 2.73 days earlier


Time to First Stable Anticoagulation

PGx-informedControls

Pro

port

ion

of

pati

en

ts

at

stab

le a

nti

coag

ula

tion

0.0

0.2

0.4

0.6

0.8

1.0

DaysCaraco, Clin Pharm Ther, 2008

*Time to stable anticoagulation 18 days earlier(22+7 vs 40+21 days)

P<0.001*

0 25 50 75 100 125


Time spent in the therapeutic range Time spent in the therapeutic range was higherwas higher (80.4 vs. 63.4%)(80.4 vs. 63.4%)

Bleeding incidence was lower (3.2 Bleeding incidence was lower (3.2 vs. 12.5%)vs. 12.5%)

Patients with 1 or 2 variant alleles Patients with 1 or 2 variant alleles required 77% and 52% of dose used required 77% and 52% of dose used by *1/*1, respectivelyby *1/*1, respectively


Using genotyping results Using genotyping results to make Warfarin dosing to make Warfarin dosing

decisionsdecisions No clinically validated algorithmNo clinically validated algorithm A “work in progress”A “work in progress”

Several large multi-center studies being Several large multi-center studies being undertaken to address this questionundertaken to address this question

www.warfarindosing.orgwww.warfarindosing.org

Warfarin PGx: SummaryWarfarin PGx: Summary Some studies suggest that warfarin PGx testing is beneficialSome studies suggest that warfarin PGx testing is beneficial

Reduced time to stable INRReduced time to stable INR Reduced adverse eventsReduced adverse events

Large multi-center studies currently underwayLarge multi-center studies currently underway Clinical outcomesClinical outcomes Validated dosing algorithmsValidated dosing algorithms

New technology allowing for more rapid resultsNew technology allowing for more rapid results Genetic variants explain 40-45% of variability in response to Genetic variants explain 40-45% of variability in response to

warfarinwarfarin Important to realize the impact of the compound effects of Important to realize the impact of the compound effects of

polymorphisms in polymorphisms in CYP2C9CYP2C9 AND AND VKORC1VKORC1 Drug-drug interactions, co-morbidities, age, BMIDrug-drug interactions, co-morbidities, age, BMI


LDLR, APOB, PCSK9



Genetic markers for CAD Multiple genes contribute to small percentage of

phenotype Chromosome 9p21/ANRIL KIF6

Pharmacogenetics Controversial, but gathering speed

Warfarin sensitivity Statin efficacy


Thank You!!Thank You!!

Recent Genetic Advances in Cardiovascular Disease Linnea M. Baudhuin, Ph.D. Mayo Clinic Dec. 6,...

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Transcript of Recent Genetic Advances in Cardiovascular Disease Linnea M. Baudhuin, Ph.D. Mayo Clinic Dec. 6,...