NOVEL BIOMARKERS and CARDIOVASCULAR DISEASE

NOVEL BIOMARKERS and NOVEL BIOMARKERS and CARDIOVASCULAR DISEASECARDIOVASCULAR DISEASE

Nathan D Wong, PhD, FACCNathan D Wong, PhD, FACCProfessor and DirectorProfessor and Director

Heart Disease Prevention ProgramHeart Disease Prevention ProgramUniversity of California, IrvineUniversity of California, Irvine

ATP III Assessment of CHD RiskATP III Assessment of CHD Risk

For persons For persons withoutwithout known CHD, other forms known CHD, other forms of atherosclerotic disease, or diabetes:of atherosclerotic disease, or diabetes:

Count the number of risk factors:Count the number of risk factors:—Cigarette smokingCigarette smoking—Hypertension (BP Hypertension (BP 140/90 mmHg or on 140/90 mmHg or on

antihypertensive medication)antihypertensive medication)—Low HDL cholesterol (<40 mg/dL)Low HDL cholesterol (<40 mg/dL)†† —Family history of premature CHDFamily history of premature CHD

CHD in male first degree relative <55 yearsCHD in male first degree relative <55 years CHD in female first degree relative <65 yearsCHD in female first degree relative <65 years

—Age (men Age (men 45 years; women 45 years; women 55 years)55 years) Use Framingham scoring for persons with Use Framingham scoring for persons with 2 risk 2 risk

factors* factors* (or with metabolic syndrome)(or with metabolic syndrome) to to determine the absolute 10-year CHD risk. determine the absolute 10-year CHD risk. (downloadable risk algorithms at (downloadable risk algorithms at www.nhlbi.nih.gov)www.nhlbi.nih.gov)

Expert Panel on Detection, Evaluation, and Treatment ofHigh Blood Cholesterol in Adults. JAMA. 2001;285:2486-2497.

© 2001, Professional Postgraduate Services®

www.lipidhealth.org

Note: Risk estimates were derived from the experience of the Framingham Heart Study, a predominantly Caucasian population in Massachusetts, USA.

Expert Panel on Detection, Evaluation, and Treatment of High Blood Cholesterol in Adults. JAMA. 2001;285:2486-2497.

Assessing CHD Risk in MenAssessing CHD Risk in MenStep 1: Age

YearsPoints

20-34 -935-39 -440-44 045-49 350-54 655-59 860-64 1065-69 1170-74 1275-79 13

Step 2: Total CholesterolTC Points at Points at Points at Points at

Points at(mg/dL) Age 20-39 Age 40-49 Age 50-59 Age 60-69

Age 70-79 <160 0 0 0 0

0160-199 4 3 2 1

0200-239 7 5 3 1

0240-279 9 6 4 2

1280 11 8 5 3

1

HDL-C(mg/dL) Points

60 -1

50-59 0

40-49 1

<40 2

Step 3: HDL-Cholesterol

Systolic BP PointsPoints

(mm Hg) if Untreated if Treated

<120 0 0120-129 0 1130-139 1 2140-159 1 2160 2 3

Step 4: Systolic Blood Pressure

Step 5: Smoking Status

Points at Points at Points at Points atPoints at

Age 20-39 Age 40-49 Age 50-59 Age 60-69Age 70-79

Nonsmoker 0 0 0 00

Smoker 8 5 3 11

Age

Total cholesterol

HDL-cholesterol

Systolic blood pressure

Smoking status

Point total

Step 6: Adding Up the Points

Point Total 10-Year Risk Point Total 10-Year Risk

<0 <1% 118%

0 1% 1210%

1 1% 1312%

2 1% 1416%

3 1% 1520%

4 1% 1625%

5 2% 1730%

6 2%7 3%8 4%9 5%

10 6%

Step 7: CHD Risk

ATP III Framingham Risk Scoring http://www.nhlbi.nih.gov/guidelines/cholesterol/index.htm

© 2001, Professional Postgraduate Services®

www.lipidhealth.org

Modified approach to CHD risk assessmentModified approach to CHD risk assessment

LOW RISKLOW RISK designated as <0.6% CHD risk per designated as <0.6% CHD risk per year (<6% in 10 years)year (<6% in 10 years)

INTERMEDIATE RISKINTERMEDIATE RISK designated as a CHD designated as a CHD risk of 0.6%-2.0% per year (6-20% over 10 risk of 0.6%-2.0% per year (6-20% over 10 years)years)

HIGH RISKHIGH RISK designated as a CHD risk of >2% designated as a CHD risk of >2% per year (20% in 10 years) (CHD risk per year (20% in 10 years) (CHD risk equivalent), including those with CVD, equivalent), including those with CVD, diabetes, and PADdiabetes, and PAD

Greenland P et al. Circulation 2001; 104: 1863-7Greenland P et al. Circulation 2001; 104: 1863-7

PresentationPresentation Examination:Examination:

— Height: 6 ft 2 inHeight: 6 ft 2 in— Weight: 220 lb (BMI Weight: 220 lb (BMI

28 kg/m28 kg/m22))— Waist Waist

circumference: 41 circumference: 41 inin

— BP: 150/88 mm HgBP: 150/88 mm Hg— P: 64 bpm P: 64 bpm — RR: 12 breaths/minRR: 12 breaths/min

Cardiopulmonary Cardiopulmonary exam:exam: normal normal

Laboratory results:Laboratory results: — TC: 220 mg/dLTC: 220 mg/dL— HDL-C: 36 mg/dLHDL-C: 36 mg/dL— LDL-C: 140 mg/dLLDL-C: 140 mg/dL— TG: 220 mg/dLTG: 220 mg/dL— FBS: 120 mg/dLFBS: 120 mg/dL

What is WJC’s 10-year absolute riskWhat is WJC’s 10-year absolute riskof fatal/nonfatal MI?of fatal/nonfatal MI?

A 12% absolute risk is derived from points A 12% absolute risk is derived from points assigned in Framingham Risk Scoring to:assigned in Framingham Risk Scoring to:— Age: Age: 66— TC: TC: 33— HDL-C: HDL-C: 22— SBP: SBP: 22— Total: 13 pointsTotal: 13 points

In 1992 he exercised 14 minutes in a Bruce protocol exercise stress test to 91% of his maximum predicted heart rate without any abnormal ECG changes. He started on a statin in 2001. But in Sept 2004, he needed urgent coronary bypass surgery.

Not all individuals with coronary Not all individuals with coronary heart disease have traditional risk heart disease have traditional risk

factorsfactors

Khot et al. JAMA 2003

The Detection Gap in CHDThe Detection Gap in CHD

““Despite many available risk assessment Despite many available risk assessment approaches, a substantial gap remains in approaches, a substantial gap remains in the detection of asymptomatic individuals the detection of asymptomatic individuals who ultimately develop CHD”who ultimately develop CHD”

““The Framingham and European risk scores… The Framingham and European risk scores… emphasize the classic CHD risk factors…. is emphasize the classic CHD risk factors…. is only moderately accurate for the prediction only moderately accurate for the prediction of short- and long-term risk of manifesting a of short- and long-term risk of manifesting a major coronary artery event…”major coronary artery event…”

Pasternak and Abrams et al. 34Pasternak and Abrams et al. 34thth Bethesda conf. JACC 2003; 41: Bethesda conf. JACC 2003; 41: 1855-19171855-1917

Is there clinical evidence that novel risk Is there clinical evidence that novel risk

markers predict future coronary events markers predict future coronary events

and provide additional predictive and provide additional predictive

information beyond traditional risk information beyond traditional risk

factors?factors?

Fibrinogen and AtherosclerosisFibrinogen and Atherosclerosis

Promotes atherosclerosisPromotes atherosclerosis Essential component of platelet aggregationEssential component of platelet aggregation Relates to fibrin deposited and the size of the Relates to fibrin deposited and the size of the

clotclot Increases plasma viscosityIncreases plasma viscosity May also have a proinflammatory roleMay also have a proinflammatory role Measurement of fibrinogen, incl. Test Measurement of fibrinogen, incl. Test

variability, remains difficult. variability, remains difficult. No known therapies to selectively lower No known therapies to selectively lower

fibrinogen levels in order to test efficacy in fibrinogen levels in order to test efficacy in CHD risk reduction via clinical trials.CHD risk reduction via clinical trials.

Fibrinogen and CHD Risk: Fibrinogen and CHD Risk: Epidemiologic StudiesEpidemiologic Studies

Recent meta-analysis of 18 studies involving 4018 Recent meta-analysis of 18 studies involving 4018 CHD cases showed a relative risk of CHD of 1.8 CHD cases showed a relative risk of CHD of 1.8 (95% CI 1.6-2.0) comparing the highest vs lowest (95% CI 1.6-2.0) comparing the highest vs lowest tertile of fibrinogen levels (mean .35 vs. .25 g/dL)tertile of fibrinogen levels (mean .35 vs. .25 g/dL)

ARIC study in 14,477 adults aged 45-64 showed ARIC study in 14,477 adults aged 45-64 showed relative risks of 1.8 in men and 1.5 in women, relative risks of 1.8 in men and 1.5 in women, attenuated to 1.5 and 1.2 after risk factor attenuated to 1.5 and 1.2 after risk factor adjustment.adjustment.

Scottish Heart Health Study of 5095 men and 4860 Scottish Heart Health Study of 5095 men and 4860 women showed fibrinogen to be an independent women showed fibrinogen to be an independent risk factor for new events--RRs 2.2-3.4 for coronary risk factor for new events--RRs 2.2-3.4 for coronary death and all-cause mortality.death and all-cause mortality.

Fibrinogen and CHD Risk FactorsFibrinogen and CHD Risk Factors

Fibrinogen levels increase with age and body Fibrinogen levels increase with age and body mass index, and higher cholesterol levelsmass index, and higher cholesterol levels

Smoking can reversibly elevated fibrinogen Smoking can reversibly elevated fibrinogen levels, and cessation of smoking can lower levels, and cessation of smoking can lower fibrinogen.fibrinogen.

Those who exercise, eat vegetarian diets, and Those who exercise, eat vegetarian diets, and consume alcohol have lower levels. Exercise consume alcohol have lower levels. Exercise may also lower fibrinogen and plasma viscosity.may also lower fibrinogen and plasma viscosity.

Studies also show statin-fibrate combinations Studies also show statin-fibrate combinations (simvastatin-ciprofibrate) and estrogen therapy (simvastatin-ciprofibrate) and estrogen therapy to lower fibrinogen.to lower fibrinogen.

P. Ridker

CRP vs hs-CRPCRP vs hs-CRP

CRP is an acute-phase protein produced by the liver CRP is an acute-phase protein produced by the liver in response to cytokine production (IL-6, IL-1, tumor in response to cytokine production (IL-6, IL-1, tumor necrosis factor) during tissue injury, inflammation, necrosis factor) during tissue injury, inflammation, or infection.or infection.

Standard CRPStandard CRP tests determine levels which are tests determine levels which are increased up to 1,000-fold in response to infection increased up to 1,000-fold in response to infection or tissue destruction, but cannot adequately assess or tissue destruction, but cannot adequately assess the normal rangethe normal range

High-sensitivity CRPHigh-sensitivity CRP (hs-CRP) assays (i.e. Dade (hs-CRP) assays (i.e. Dade Behring) detect levels of CRP within the normal Behring) detect levels of CRP within the normal range, levels proven to predict future cardiovascular range, levels proven to predict future cardiovascular events.events.

C-Reactive Protein:C-Reactive Protein:Risk Factor or Risk Marker?Risk Factor or Risk Marker?

CRP previously known to be a marker of CRP previously known to be a marker of high risk in cardiovascular diseasehigh risk in cardiovascular disease

More recent data may implicate CRP as More recent data may implicate CRP as an actual mediator of atherogenesisan actual mediator of atherogenesis

Multiple hypotheses for the mechanism Multiple hypotheses for the mechanism of CRP-mediated atherogenesis:of CRP-mediated atherogenesis:—Endothelial dysfunction via Endothelial dysfunction via ↑ NO ↑ NO

synthesissynthesis—↑↑LDL deposition in plaque by CRP-LDL deposition in plaque by CRP-

stimulated macrophagesstimulated macrophages

hs-CRP as a Risk Factor For Future CVD : hs-CRP as a Risk Factor For Future CVD : Primary Prevention CohortsPrimary Prevention Cohorts

0 1.0 2.0 3.0 4.0 5.0 6.0

Kuller MRFIT 1996 CHD DeathRidker PHS 1997 MI

Ridker PHS 1997 Stroke

Tracy CHS/RHPP 1997 CHD

Ridker PHS 1998,2001 PAD

Ridker WHS 1998,2000,2002 CVD

Koenig MONICA 1999 CHD

Roivainen HELSINKI 2000 CHD

Mendall CAERPHILLY 2000 CHD

Danesh BRHS 2000 CHD

Gussekloo LEIDEN 2001 Fatal Stroke

Lowe SPEEDWELL 2001 CHD

Packard WOSCOPS 2001 CV Events*

Ridker AFCAPS 2001 CV Events*

Rost FHS2001 Stroke

Pradhan WHI 2002MI,CVD death

Albert PHS 2002 Sudden Death

Sakkinen HHS 2002 MIRelative Risk (upper vs lower quartile)Ridker PM. Circulation 2003;107:363-9

hs-CRP Adds to Predictive Value of TC:HDL hs-CRP Adds to Predictive Value of TC:HDL Ratio in Determining Risk of First MIRatio in Determining Risk of First MI

0.0

1.0

2.0

3.0

4.0

5.0

High Medium Low Low

Medium

High

Total Cholesterol:HDL RatioTotal Cholesterol:HDL Ratio

Ridker et al, Circulation. 1998;97:2007–2011.

hs-CRP

hs-CRP

Rel

ativ

e R

isk

Rel

ativ

e R

isk

Risk Factors for Future Cardiovascular Risk Factors for Future Cardiovascular Events: WHSEvents: WHS

0 1.0 2.0 4.0 6.0

Lipoprotein(a)

Homocysteine

IL-6

TC

LDLC

sICAM-1

SAA

Apo B

TC: HDLC

hs-CRP

hs-CRP + TC: HDLC

Relative Risk of Future Cardiovascular Events

Ridker et al, N Engl J Med. 2000;342:836-43

Is there clinical evidence that Is there clinical evidence that

inflammation can be modified by inflammation can be modified by

preventive therapies?preventive therapies?

0

5

10

15

20

25

Elevated CRP Levels in Obesity: Elevated CRP Levels in Obesity: NHANES 1988-1994NHANES 1988-1994

Visser M et al. JAMA 1999;282:2131-2135.

Normal

Perc

en

t w

ith C

RP

0.2

2

mg/d

L

Overweight Obese

Effects of Weight Loss on CRPEffects of Weight Loss on CRPConcentrations in Obese Healthy Concentrations in Obese Healthy

WomenWomen 83 women (mean BMI 33.8, range 28.2-43.8 kg/m2) placed

on very low fat, energy-restricted diet (6.0 MJ, 15% fat) for 12 weeks

Baseline CRP positively associated with BMI (r=0.281, p=0.01)

CRP reduced by 26% (p<0.001)

Average weight loss 7.9 kg, associated with change in CRP

Change in CRP correlated with change in TC (r=0.240, p=0.03) but not changes in LDL-C, HDL-C, or glucose

At 12 weeks, CRP concentration highly correlated with TG (r=0.287, p=0.009), but not with other lipids or glucose

Heilbronn LK et al. Arterioscler Thromb Vasc Biol 2001;21:968-970.

Effect of HRT on hs-CRP: Effect of HRT on hs-CRP: the PEPI the PEPI StudyStudy

3.0

2.0

1.0hs-

CR

P (

mg/d

L)

Months

0 12 36

Cushman M et al. Circulation 1999;100:717-722.1999 Lippincott Williams & Wilkins.

CEE + MPA cyclicCEE + MPA continuousCEE + MP

CEE

Placebo

Long-Term Effect of Statin Therapy on Long-Term Effect of Statin Therapy on hs-CRP: Placebo and Pravastatin Groupshs-CRP: Placebo and Pravastatin Groups

PravastatinPravastatin

PlaceboPlacebo

Me

dia

n h

s-C

RP

Me

dia

n h

s-C

RP

Co

nc

en

tra

tio

nC

on

ce

ntr

ati

on

(mg

/dL

)(m

g/d

L) -21.6%-21.6%

((PP=0.004)=0.004)

0.18

0.19

0.20

0.21

0.22

0.23

0.24

0.25

Baseline 5 Years

Ridker et al, Circulation. 1999;100:230-235.

hs-

CR

P (

mg/L

)Effect of Statin Therapy on hs-CRP Levels Effect of Statin Therapy on hs-CRP Levels

at 6 Weeksat 6 Weeks

Jialal I et al. Circulation 2001;103:1933-1935.2001 Lippincott Williams & Wilkins.

6

5

4

3

2

1

0 Baseline

* * *

Prava(40 mg/d)

Simva(20 mg/d)

Atorva(10 mg/d)

*p<0.025 vs. Baseline

However, while intriguing and of potential public health importance, the observation in AFCAPS/TexCAPS that statin therapy might be effective among those with elevated hsCRP but low cholesterol was made on a post hoc basis. Thus, a large-scale randomized trial of statin therapy was needed to directly test this hypotheses.

Ridker et al, New Engl J Med 2001;344:1959-65

Low LDL, Low hsCRP

Low LDL, High hsCRP

Statin Effective Statin Not Effective

1.0 2.00.5

[A]

[B]

Low LDL, Low hsCRP

Low LDL, High hsCRP

Statin Effective Statin Not Effective

1.0 2.00.5

AFCAPS/TexCAPS Low LDL Subgroups

RR

AFCAPS/TEXCAPS showed statins to be effective in lowering risk in the setting of normal LDL-C, but only when inflammation

was present

A Randomized Trial of Rosuvastatin in the Preventionof Cardiovascular Events Among 17,802 Apparently Healthy

Men and Women With Elevated Levels of C-Reactive Protein (hsCRP):

The JUPITER Trial

Paul Ridker*, Eleanor Danielson, Francisco Fonseca*, Jacques Genest*,Antonio Gotto*, John Kastelein*, Wolfgang Koenig*, Peter Libby*,

Alberto Lorenzatti*, Jean MacFadyen, Borge Nordestgaard*, James Shepherd*, James Willerson, and Robert Glynn*

on behalf of the JUPITER Trial Study Group

An Investigator Initiated Trial Funded by AstraZeneca, USA

* These authors have received research grant support and/or consultation fees from one or morestatin manufacturers, including Astra-Zeneca. Dr Ridker is a co-inventor on patents held by the

Brigham and Women’s Hospital that relate to the use of inflammatory biomarkers in cardiovascular disease that have been licensed to Dade-Behring and AstraZeneca.

To investigate whether rosuvastatin 20 mg compared to placebo would decrease the rate of first major cardiovascularevents among apparently healthy men and women with LDL < 130 mg/dL (3.36 mmol/L) who are nonethelessat increased vascular risk on the basis of an enhanced inflammatory response, as determined by hsCRP > 2 mg/L.

To enroll large numbers of women and individuals of Black or Hispanic ethnicity, groups for whom little data on primaryprevention with statin therapy exists.

Justification for the Use of statins in Prevention: an Intervention Trial Evaluating Rosuvastatin

Ridker et al NEJM 2008

Rosuvastatin 20 mg (N=8901)Rosuvastatin 20 mg (N=8901) MIMIStrokeStroke

UnstableUnstable AnginaAngina

CVD DeathCVD DeathCABG/PTCACABG/PTCA

JUPITERJUPITERMulti-National Randomized Double Blind Placebo Controlled Trial of Multi-National Randomized Double Blind Placebo Controlled Trial of

Rosuvastatin in the Prevention of Cardiovascular EventsRosuvastatin in the Prevention of Cardiovascular EventsAmong Individuals With Low LDL and Elevated hsCRPAmong Individuals With Low LDL and Elevated hsCRP

4-week 4-week run-inrun-in

Ridker et al, Circulation 2003;108:2292-2297.

No Prior CVD or DMNo Prior CVD or DMMen Men >>50, Women 50, Women >>6060

LDL <130 mg/dL hsCRP >2 mg/L

JUPITERTrial Design

Placebo (N=8901)Placebo (N=8901)

Argentina, Belgium, Brazil, Bulgaria, Canada, Chile, Colombia, Costa Rica, Denmark, El Salvador, Estonia, Germany, Israel, Mexico, Netherlands, Norway, Panama, Poland, Romania, Russia, South Africa, Switzerland,

United Kingdom, Uruguay, United States, Venezuela

JUPITERBaseline Blood Levels (median, interquartile range)

Rosuvastatin Placebo(N = 8901) (n = 8901)

hsCRP, mg/L 4.2 (2.8 - 7.1) 4.3 (2.8 - 7.2) LDL, mg/dL 108 (94 - 119) 108 (94 - 119)

HDL, mg/dL 49 (40 – 60) 49 (40 – 60)

Triglycerides, mg/L 118 (85 - 169) 118 (86 - 169)

Total Cholesterol, mg/dL 186 (168 - 200) 185 (169 - 199)

Glucose, mg/dL 94 (87 – 102) 94 (88 – 102)

HbA1c, % 5.7 (5.4 – 5.9) 5.7 (5.5 – 5.9)

All values are median (interquartile range). [ Mean LDL = 104 mg/dL ]


0

1

2

3

4

5

hsC

RP

(m

g/L

)

0

20

40

60

80

100

120

140

LD

L (

mg

/dL

)

Months0 12 24 36 48

0

10

20

30

40

50

60

0

20

40

60

80

100

120

140

0 12 24 36 48

TG

(m

g/d

L)

HD

L (

mg

/dL

)

Months

JUPITEREffects of rosuvastatin 20 mg on LDL, HDL, TG, and hsCRP

LDL decrease 50 percent at 12 months

hsCRP decrease 37 percent at 12 months

HDL increase 4 percent at 12 months

TG decrease 17 percent at 12 months


JUPITERPrimary Trial Endpoint : MI, Stroke, UA/Revascularization, CV Death

Placebo 251 / 8901

Rosuvastatin 142 / 8901

HR 0.56, 95% CI 0.46-0.69P < 0.00001

Number Needed to Treat (NNT5) = 25

- 44 %

0 1 2 3 4

0.0

00

.02

0.0

40

.06

0.0

8

Cu

mu

lati

ve In

cid

ence

Number at Risk Follow-up (years)

Rosuvastatin

Placebo

8,901 8,631 8,412 6,540 3,893 1,958 1,353 983 544 157

8,901 8,621 8,353 6,508 3,872 1,963 1,333 955 534 174


JUPITERSecondary Endpoint – All Cause Mortality

Placebo 247 / 8901

Rosuvastatin 198 / 8901

HR 0.80, 95%CI 0.67-0.97P= 0.02

- 20 %

0 1 2 3 4

0.00

0.01

0.02

0.03

0.04

0.05

0.06

Cu

mu

lati

ve I

nci

den

ce

Number at Risk Follow-up (years)

RosuvastatinPlacebo

8,901 8,847 8,787 6,999 4,312 2,268 1,602 1,192 683 2278,901 8,852 8,775 6,987 4,319 2,295 1,614 1,196 684 246


JUPITERImplications for Primary Prevention

Among men and women age 50 or over :

If diabetic, treatIf LDLC > 160 mg/dL, treat

If hsCRP > 2 mg/L, treat

A simple evidence based approach to statin therapy for primary prevention.


AHA / CDC Scientific StatementAHA / CDC Scientific StatementMarkers of Inflammation and Cardiovascular Disease:Markers of Inflammation and Cardiovascular Disease:

Applications to Clinical and Public Health PracticeApplications to Clinical and Public Health Practice

Circulation January 28, 2003Circulation January 28, 2003

“Measurement of hs-CRP is an independent marker of risk“Measurement of hs-CRP is an independent marker of riskand may be used at the discretion of the physician as partand may be used at the discretion of the physician as part

of global coronary risk assessment in adults without knownof global coronary risk assessment in adults without knowncardiovascular disease. Weight of evidence favors use cardiovascular disease. Weight of evidence favors use particularly among those judged at intermediate risk byparticularly among those judged at intermediate risk by

global risk assessment”.global risk assessment”.

1 mg/L 3 mg/L 10 mg/L

LowRisk

ModerateRisk

HighRisk

Acute Phase ResponseIgnore Value, Repeat Test in 3

weeks

>100 mg/L

Ridker PM. Circulation 2003;107:363-9

Clinical Application of hs-CRP forClinical Application of hs-CRP forCardiovascular Risk Prediction Cardiovascular Risk Prediction

HomocysteineHomocysteine

Intermediary amino acid formed by the Intermediary amino acid formed by the conversion of methionine to cysteineconversion of methionine to cysteine

Moderate hyperhomocysteinemia occurs in 5-Moderate hyperhomocysteinemia occurs in 5-7% of the population7% of the population

Recognized as an independent risk factor for Recognized as an independent risk factor for the development of atherosclerotic vascular the development of atherosclerotic vascular disease and venous thrombosisdisease and venous thrombosis

Can result from genetic defects, drugs, vitamin Can result from genetic defects, drugs, vitamin deficiencies, or smoking deficiencies, or smoking


Homocysteine implicated directly in vascular Homocysteine implicated directly in vascular injury including:injury including:—Intimal thickeningIntimal thickening—Disruption of elastic laminaDisruption of elastic lamina—Smooth muscle hypertrophySmooth muscle hypertrophy—Platelet aggregationPlatelet aggregation

Vascular injury induced by leukocyte Vascular injury induced by leukocyte recruitment, foam cell formation, and recruitment, foam cell formation, and inhibition of NO synthesisinhibition of NO synthesis


Elevated levels appear to be an independent Elevated levels appear to be an independent risk factor, though less important than the risk factor, though less important than the classic CV risk factorsclassic CV risk factors

Screening recommended in patients with Screening recommended in patients with premature CV disease (or unexplained DVT) premature CV disease (or unexplained DVT) and absence of other risk factorsand absence of other risk factors

Treatment includes supplementation with Treatment includes supplementation with folate, B6 and B12folate, B6 and B12

The Future of Cardiac BiomarkersThe Future of Cardiac Biomarkers

Many experts are advocating the Many experts are advocating the move towards a move towards a multimarker multimarker strategystrategy for the purposes of for the purposes of diagnosis, prognosis, and diagnosis, prognosis, and treatment designtreatment design

As the pathophysiology of ACS is As the pathophysiology of ACS is heterogeneous, so must be the heterogeneous, so must be the diagnostic strategiesdiagnostic strategies

Multiple Biomarkers for the Prediction of Multiple Biomarkers for the Prediction of First CVD Events and Death (Wang TJ et al., First CVD Events and Death (Wang TJ et al.,

NEJM 2006; 355: 2631-9)NEJM 2006; 355: 2631-9) 10 biomarkers examined in 3209 pts of the Framingham 10 biomarkers examined in 3209 pts of the Framingham

Heart StudyHeart Study CRP, BNP, N-T pro-ANP, aldosterone, renin, fibrinogen, d-CRP, BNP, N-T pro-ANP, aldosterone, renin, fibrinogen, d-

dimer, PAI-1, homocysteine, and urine albumin/creatinine dimer, PAI-1, homocysteine, and urine albumin/creatinine ratio.ratio.

7.4 years medial follow-up7.4 years medial follow-up Adjusted HR’s per SD: BNP 1.4, CRP 1.4, Adjusted HR’s per SD: BNP 1.4, CRP 1.4,

albumin/creatinine 1.2, homocysteine 1.2, renin 1.5 for albumin/creatinine 1.2, homocysteine 1.2, renin 1.5 for death, and BNP 1.25, albumin/creatinine 1.2 for CVD death, and BNP 1.25, albumin/creatinine 1.2 for CVD eventsevents

Multimarker scores in highest quintile vs. lowest two Multimarker scores in highest quintile vs. lowest two quintiles had adjusted HR for death of 4.1, p<0.001 and quintiles had adjusted HR for death of 4.1, p<0.001 and CVD events of 1.8, p=0.02CVD events of 1.8, p=0.02

Only moderate increases in C-statistic seen from Only moderate increases in C-statistic seen from biomarkers over standard risk factorsbiomarkers over standard risk factors

Multiple biomarkers and C-statistics Multiple biomarkers and C-statistics (discrimination)(discrimination)

DeathDeath First CVDFirst CVD

Age, sexAge, sex 0.750.75 0.680.68

Risk factors aloneRisk factors alone 0.800.80 0.760.76

RF plus RF plus biomarkersbiomarkers

0.820.82 0.770.77

Multiple biomarkers and Multiple biomarkers and reclassificationreclassification

StandardStandard

risk factors risk factors alonealone

Standard risk factorsStandard risk factors

plus multimarker scoreplus multimarker score

<10%<10% 10-20%10-20% >20%>20%

<10%<10% 79%79% 3%3% 0%0%

10-20%10-20% 3%3% 9%9% 1%1%

>20%>20% 0%0% 1%1% 3%3%

Use of Multiple Biomarkers to Improve Use of Multiple Biomarkers to Improve Prediction of CVD Death (Zethelius B et al., Prediction of CVD Death (Zethelius B et al.,

NEJM 2008; 358: 2107-16)NEJM 2008; 358: 2107-16)

1135 elderly men from the Uppsala Longitudinal Study 1135 elderly men from the Uppsala Longitudinal Study of Adult Men, mean age 71 years at baseline, 10 years of Adult Men, mean age 71 years at baseline, 10 years median follow-upmedian follow-up

Examined role of multiple markers reflecting myocardial Examined role of multiple markers reflecting myocardial cell damage—troponin I, LV dysfunction– N-T pro BNP, cell damage—troponin I, LV dysfunction– N-T pro BNP, renal failure—cystatin C, and inflammation – CRPrenal failure—cystatin C, and inflammation – CRP

C-statistic increased significantly when the four C-statistic increased significantly when the four biomarkers were put in a model with established risk biomarkers were put in a model with established risk factors (0.77 vs. 0.66, p<0.0001) in the whole cohort factors (0.77 vs. 0.66, p<0.0001) in the whole cohort and in those without CVD at baseline (0.748 vs. 0.688, and in those without CVD at baseline (0.748 vs. 0.688, p=0.03).p=0.03).

Among elderly men, multiple biomarkers may Among elderly men, multiple biomarkers may significantly improve risk for death from CVD causes significantly improve risk for death from CVD causes beyond standard risk factors.beyond standard risk factors.

Current Biomarkers for ACSCurrent Biomarkers for ACS

Biomarker assessment of high risk patients may Biomarker assessment of high risk patients may include:include:— Inflammatory cytokines Inflammatory cytokines — Cellular adhesion molecules Cellular adhesion molecules — Acute-phase reactantsAcute-phase reactants— Plaque destabilization and rupture biomarkersPlaque destabilization and rupture biomarkers— Biomarkers of ischemia Biomarkers of ischemia — Biomarkers of myocardial stretch (BNP)Biomarkers of myocardial stretch (BNP)— Biomarkers of myocardial necrosis (Troponin, Biomarkers of myocardial necrosis (Troponin,

CK-MB, Myoglobin)CK-MB, Myoglobin)

Apple Clinical Chemistry March 2005

Progression of Biomarkers in ACSProgression of Biomarkers in ACS

ACS, acute coronary syndrome; UA, unstable angina; NSTEMI, non–ST-segment elevation myocardial infarction; STEMI, ST-segment elevation myocardial infarction

Adapted from: Apple Clinical Chemistry March 2005

STEMIUA/NSTEMISTABLE CAD PLAQUE RUPTURE

MPOCRPIL-6

MPO ICAMsCD40LPAPP-A

MPOD-dimerIMAFABP

TnITnTMyoglobinCKMB

Inflammation has been linked to the development of vulnerable plaque and to plaque rupture

Stefan Blankenberg, MD; Renate Schnabel, MD; Edith Lubos, MD, et al., Myeloperoxidase Early Indicator of Acute Coronary Syndrome and Predictor of Future Cardiovascular Events 2005

History: TroponinHistory: Troponin

Troponin I first described as a biomarker specific Troponin I first described as a biomarker specific for AMI in 1987for AMI in 198711; Troponin T in 1989; Troponin T in 198922

Now the biochemical “gold standard” for the Now the biochemical “gold standard” for the diagnosis of acute myocardial infarction via diagnosis of acute myocardial infarction via consensus of ESC/ACCconsensus of ESC/ACC

1 1 Am Heart J 113: 1333-44Am Heart J 113: 1333-4422 J Mol Cell Cardiol 21: 1349-53 J Mol Cell Cardiol 21: 1349-53

TroponinsTroponins

Elevated serum levels are an independent Elevated serum levels are an independent predictor of prognosis, morbidity and mortalitypredictor of prognosis, morbidity and mortality

Meta-analysis of 21 studies involving ~20,000 Meta-analysis of 21 studies involving ~20,000 patients with ACS revealed that those with patients with ACS revealed that those with elevated serum troponin had 3x risk of cardiac elevated serum troponin had 3x risk of cardiac death or reinfarction at 30 daysdeath or reinfarction at 30 days11

11 Am J Heart (140): 917 Am J Heart (140): 917

All-Cause Mortality by Cardiac Troponin All-Cause Mortality by Cardiac Troponin TT (n=733) (n=733)

CP1090800-14Circulation 106:2944, 2002Circulation 106:2944, 2002

Time since blood draw (years)Time since blood draw (years)

Cumulative survival

(%)

Cumulative survival

(%)

Patients at risk (no.) Baseline 1 yr 2 yr 2.5 yr

cTnT <0.01 g/L 132 106 25 12

cTnT 0.01 to <0.04 g/L 214 166 41 15

cTnT 0.04 to <0.10 g/L 239 180 63 18

cTnT 0.10 g/L 148 93 20 8

Patients at risk (no.) Baseline 1 yr 2 yr 2.5 yr

cTnT <0.01 g/L 132 106 25 12

cTnT 0.01 to <0.04 g/L 214 166 41 15

cTnT 0.04 to <0.10 g/L 239 180 63 18

cTnT 0.10 g/L 148 93 20 8

0

20

40

60

80

100

0.0 0.5 1.0 1.5 2.0 2.5 3.0

cTnT <0.01 g/LcTnT <0.01 g/L

cTnT 0.04 g/LcTnT 0.04 g/L

cTnT 0.10 g/LcTnT 0.10 g/L

cTnT cTnT 0.040.04to 0.10 to 0.10 g/Lg/L

cTnT and Survival (Rancho Bernardo)cTnT and Survival (Rancho Bernardo)

Daniels et al: JACC 52:450, 2008Daniels et al: JACC 52:450, 2008

CP1322078-9

20

40

60

80

100

0 2 4 6 8

Su

rviv

al (

%)

Su

rviv

al (

%)

YearsYears

All SubjectsAll Subjects

20

40

60

80

100

0 2 4 6 8

Su

rviv

al (

%)

Su

rviv

al (

%)

YearsYears

Subjects WithoutBaseline CHD


P<0.001P<0.001TnT 0.01 ng/mLTnT 0.01 ng/mL

TnT undetectableTnT undetectable

P<0.001P<0.001

TnT undetectableTnT undetectable

TnT 0.01 ng/mLTnT 0.01 ng/mL

BNPBNP

BNP has also shown utility as a prognostic BNP has also shown utility as a prognostic marker in acute coronary syndromemarker in acute coronary syndrome

It is associated with increased risk of death at It is associated with increased risk of death at 10 months as concentration at 40 hours post-10 months as concentration at 40 hours post-infarct increasedinfarct increased

Also associated with increased risk for new or Also associated with increased risk for new or recurrent MIrecurrent MI

BNP as a Predictor of Risk in Asymptomatic BNP as a Predictor of Risk in Asymptomatic Adults: The Framingham Heart StudyAdults: The Framingham Heart Study

Wang et al., NEJM 2004

Association of increasing BNP Association of increasing BNP levels and outcomeslevels and outcomes

End point End point Hazard ratio for Hazard ratio for 1 SD increment 1 SD increment in log BNP value in log BNP value

p p

Death Death 1.27 1.27 0.009 0.009

First major First major CV event CV event

1.28 1.28 0.03 0.03

HF HF 1.77 1.77 <0.001 <0.001

Atrial Atrial fibrillation fibrillation

1.66 1.66 <0.001 <0.001

Stroke or Stroke or TIA TIA

1.53 1.53 0.002 0.002

CHD event CHD event 1.1 1.1 0.37 0.37 SD=standard deviation

Wang TJ et al. N Engl J Med 2004; 350:655-63.

Conjoint Effects of cTnT and NT-proBNP on Conjoint Effects of cTnT and NT-proBNP on Prognosis (Rancho Bernardo)Prognosis (Rancho Bernardo)

Daniels et al: JACC 52:450, 2008Daniels et al: JACC 52:450, 2008

CP1322078-12

20

40

60

80

100

0 2 4 6 8

Su

rviv

al (

%)

Su

rviv

al (

%)

YearsYears

All SubjectsAll Subjects

P<0.001 for all comparisonsP<0.001 for all comparisons20

40

60

80

100

0 2 4 6 8

Su

rviv

al (

%)

Su

rviv

al (

%)

YearsYears

P<0.001 for all comparisonsP<0.001 for all comparisons



Low NT-proBNP(n=758)


High NT-proBNP,low TnT (n=171)


High NT-proBNP,high TnT (n=27)








MyeloperoxidaseMyeloperoxidase

MPO is an enzyme that aids white blood cells in MPO is an enzyme that aids white blood cells in destroying bacteria and viral particlesdestroying bacteria and viral particles

MPO catalyzes the conversion of hydrogen peroxide and MPO catalyzes the conversion of hydrogen peroxide and chloride ions (Cl-) into hypochlorous acidchloride ions (Cl-) into hypochlorous acid

Hypochlorous acid is 50 times more potent in microbial Hypochlorous acid is 50 times more potent in microbial killing than hydrogen peroxidekilling than hydrogen peroxide

MPO is released in response to infection and MPO is released in response to infection and inflammationinflammation

EPIC Norfolk Study showed its predictive value for future EPIC Norfolk Study showed its predictive value for future cardiovascular disease events in asymptomatic adults.cardiovascular disease events in asymptomatic adults.

Sugiyama Am J Pathology 2001

Summary of MPO and ACSSummary of MPO and ACS

MPO leads to oxidized LDL cholesterol MPO leads to oxidized LDL cholesterol — Oxidized LDL is phagocytosed by macrophages Oxidized LDL is phagocytosed by macrophages

producing foam cells*producing foam cells* MPO leads to the consumption of nitric oxideMPO leads to the consumption of nitric oxide

— Vasoconstriction and endothelial dysfunctionVasoconstriction and endothelial dysfunction MPO can cause endothelial denuding and MPO can cause endothelial denuding and

superficial platelet aggregationsuperficial platelet aggregation MPO indicates activated immune cellsMPO indicates activated immune cells

— Activated immune cells and inflammation lead to Activated immune cells and inflammation lead to unstable plaque*unstable plaque*

Inflammatory plaque is inherently less stableInflammatory plaque is inherently less stable— Thin fibrous cap/fissured/denudedThin fibrous cap/fissured/denuded

Brennan, NEJM 2003*Hansson, NEJM 2005

MPO and MI in Asymptomatic MPO and MI in Asymptomatic Subjects: EPIC-NORFOLKSubjects: EPIC-NORFOLK

0

5

10

15

20

24 hours 72 hours 30 days 6 months

1st tertile2nd tertile3rd tertile

Tertile 1 MPO Tertile 1 MPO << 222 ug/L 222 ug/L

Tertile 2 MPO 222 – 350 ug/LTertile 2 MPO 222 – 350 ug/L

Tertile 3 MPO Tertile 3 MPO >> 350 ug/L 350 ug/L

Dea

th o

r M

I (%

)

Baldus, et al. Circulation 2003;108: 1440-5.

MPO and CVD Event Risk (%)MPO and CVD Event Risk (%)

2.22.5

4.9

4.3

0.0

0.5

1.0

1.5

2.0

2.5

3.0

3.5

4.0

4.5

5.0

MPO Quartile

1st 2nd 3rd 4th

P-trend = 0.05 (Wong et al. JACC Cardiovasc Img 2009 )

Figure 2

Combined MPO-CAC Groups and CVD Event Combined MPO-CAC Groups and CVD Event Risk (%)Risk (%)

7.1%14.0%

3.2% 3.9%

0.3%0.9%0.0%

2.0%4.0%6.0%8.0%

10.0%12.0%14.0%

CV

D E

ven

ts (

%)

MPO<257pm MPO>=257pm

CAC 0-9

CAC 10-99

CAC>=100

Log-rank test for trend P<0.0001; Wong et al., JACC Cardiovasc Img 2009

Figure 3

NOVEL BIOMARKERS and CARDIOVASCULAR DISEASE

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