Beyond Cholesterol: Predicting Cardiovascular Risk In the 21 st Century

1. Inflammation, Thrombosis, Infection, and CARDIOVASCULAR DISEASENathan D Wong, PhD, FACCProfessor and DirectorHeart Disease Prevention ProgramUniversity of California, Irvine

2. LipidsHTNDiabetes Behavioral HemostaticThrombotic Inflammatory Genetic Beyond Cholesterol: Predicting Cardiovascular Risk In the 21st Century Cardiovascular Risk

3. Inflammation and Atherosclerosis • Inflammation may determine plaque stability - Unstable plaques have increased leukocytic infiltrates - T cells, macrophages predominate rupture sites - Cytokines and metalloproteinases influence both stability and degradation of the fibrous cap • Lipid lowering may reduce plaque inflammation - Decreased macrophage number - Decreased expression of collagenolytic enzymes (MMP-1) - Increased interstitial collagen - Decreased expression of E-selectin - Reduced calcium deposition Libby P. Circulation 1995;91:2844-2850. Ross R. N Engl J Med 1999;340:115-126.

4. Is there clinical evidence that inflammatory markers predict future coronary events and provide additional predictive information beyond traditional risk factors?

5. Evaluating Novel Risk Factors for CAD • Low variability • High reproducibility • Biologic plausibility • Low cost • Modifiable • Consistency of prospective data • Strength of association • Independence of association • Improve predictive value • Standardized measure

6. Parameter Venous Arterial Fibrinogen – +++ Factor VII – + vWF: ag – ++ tPA: ag – +++ PAI-1: ag – ++ Platelet function – ++ Lp(a) – + hs-CRP / SAA / IL-6 / TNF – +++ Biomarkers for Venous and Arterial Thrombosis

7. Parameter Venous Arterial Factor V Leiden +++ – Prothrombin mutation ++ – Prothrombin + – Factor VIII ++ – Anti-thrombin III ++ – Protein C + – Protein S + – Homocysteine ++ ++ D-dimer ++ ++ Biomarkers for Venous and Arterial Thrombosis (cont’d)

8. Thrombosis and Cardiovascular Risk • Thrombus formation is a crucial factor in the precipitation of unstable angina or myocardial infarction, as well as occlusion during or following angioplasty. • Often preceded by platelet aggregation and activation of the coagulation system. • A thrombus may develop at sites of only mild to moderate coronary stenosis. The majority of coronary events occur where there is less than 70% stenosis. • Occlusive coronary thrombosis plays a role in over 80% of myocardial infarctions and about 95% of sudden death victims.

9. Fibrinogen and Atherosclerosis • Promotes atherosclerosis • Essential component of platelet aggregation • Relates to fibrin deposited and the size of the clot • Increases plasma viscosity • May also have a proinflammatory role • Measurement of fibrinogen, incl. Test variability, remains difficult. • No known therapies to selectively lower fibrinogen levels in order to test efficacy in CHD risk reduction via clinical trials.

10. Fibrinogen and CHD Risk: Epidemiologic Studies • Recent meta-analysis of 18 studies involving 4018 CHD cases showed a relative risk of CHD of 1.8 (95% CI 1.6-2.0) comparing the highest vs lowest tertile of fibrinogen levels (mean .35 vs. .25 g/dL) • ARIC study in 14,477 adults aged 45-64 showed relative risks of 1.8 in men and 1.5 in women, attenuated to 1.5 and 1.2 after risk factor adjustment. • Scottish Heart Health Study of 5095 men and 4860 women showed fibrinogen to be an independent risk factor for new events--RRs 2.2-3.4 for coronary death and all-cause mortality.

11. Fibrinogen and CHD Risk Factors • Fibrinogen levels increase with age and body mass index, and higher cholesterol levels • Smoking can reversibly elevated fibrinogen levels, and cessation of smoking can lower fibrinogen. • Those who exercise, eat vegetarian diets, and consume alcohol have lower levels. Exercise may also lower fibrinogen and plasma viscosity. • Studies also show statin-fibrate combinations (simvastatin-ciprofibrate) and estrogen therapy to lower fibrinogen.

12. Other Thrombotic Factors and CHD • Mixed reports of coagulation factor VIIc in cardiovascular disease. PROCAM study showed no association with CHD events, CHS also showed no relation to subclinical CVD. • Endogenous tissue-type plasminogen activator (tPA) shown in some studies to relate to increased cardiovascular risk--Physician’s Health Study showed RR for MI 2.8, stroke 3.5 in those in 5th vs. 1st quintile of tPA. • Plasminogen activitor inhibitor type 1 (PAI-1) shown associated with increased cardiovascular risk, esp in diabetic patients.

13. Aspirin and Cardiovascular Risk: Clinical Trial Evidence for Primary Prevention • US Physician’s Health Study- 22,071 male physicians - 44% reduction in MI risk, 13% nonsignificant increase in risk of stroke • British Doctor’s Study of 5139 male physicians showed nonsignificant 3% reduction in MI risk,13% nonsignificant increase in stroke • Hypertension Optimal Treatment (HOT) study among 18,790 pts w/htn showed 15% reduction in CVD events, 36% reduction in MI • Women’s Health Study (n=39,876 women aged 45+) randomized to 100 mg asprin/day vs placebo, 10 years follow-up – results recently released and asprin preventive only for stroke (17% reduction overall, p=0.04; 24% ischemic stroke, p<.001); nonfatal MI RR=1.02, CVD death 0.95, ns) (NEJM 2005; 352: 1366-8).

14. Aspirin and Cardiovascular Risk: Clinical Trial Evidence for Secondary Prevention • Antiplatelet Trialists Collaboration of 54,000 patients with cardiovascular disease (10 trials post-MI) showed 31% reduction in MI, 42% reduction in stroke, 13% reduction in total vascular mortality • International Study of Infarct Survival of 17,187 pts w/evolving MI showed 49% reduction in reinfarction, 26% reduction in nonfatal stroke, and 23% reduction in total vascular mortality

15. Antiplatelet Therapy: Targets dipyridamole clopidogrel bisulfate ticlopidine hydrochloride phosphodiesterase ADP ADP Gp 2b/3a Inhibitors CollagenThrombinTXA2 Gp IIb/IIIa Activation (Fibrinogen Receptor) COX TXA2 aspirin ADP = adenosine diphosphate, TXA2 = thromboxane A2, COX = cyclooxygenase Schafer AI. Am J Med 1996;101:199–209

16. Antiplatelet Therapy: Common Oral Agents 1Topol EJ et al. Circulation. 2003;108:399-406 2Diener H-C et al. Lancet 2004;364;331-7 3Plavix® package insert. www.sanofi-synthelabo.us 4Peters RJ et al. Circulation 2003;108:1682-7 5Hass WK. NEJM 1989;321:501-7 6Urban P. Circulation. 1998;98:2126-32 7Ticlid® package insert. www.rocheusa.com *Clopidogrel is generally given preference over Ticlopidine because of a superior safety profile

17. Aspirin: Mechanism of Action Membrane Phospholipids ARACHIDONIC ACID Aspirin COX-1 Prostaglandin H2 Thromboxane A2  Platelet Aggregation Vasoconstriction Prostacyclin  Platelet Aggregation Vasodilitation

18. IIa IIa IIa IIa IIb IIb IIb IIb III III III III I I I IIa IIa IIa IIa IIb IIb IIb IIb III III III III I I I IIa IIa IIa IIa IIb IIb IIb IIb III III III III I I I B Aspirin Recommendations Primary Prevention Aspirin (75-162 mg daily) for intermediate risk men with a 10 year risk of CHD >10%. Aspirin (75-162 mg daily) for intermediate risk women with a 10 year risk of CHD >10%. Aspirin for low risk women with a 10 year risk of CHD<10%. Aspirin (75-325 mg daily) for those with known CHD. Secondary Prevention

19. Rader, NEJM 2000; 343: 1181.

20. P. Ridker

21. CRP vs hs-CRP • CRP is an acute-phase protein produced by the liver in response to cytokine production (IL-6, IL-1, tumor necrosis factor) during tissue injury, inflammation, or infection. • Standard CRP tests determine levels which are increased up to 1,000-fold in response to infection or tissue destruction, but cannot adequately assess the normal range • High-sensitivity CRP (hs-CRP) assays (i.e. Dade Behring) detect levels of CRP within the normal range, levels proven to predict future cardiovascular events.

22. Potential Mechanisms Linking CRP to Atherothrombosis • Marker for subclinical atherosclerosis- EBCT / IMT / ABI • Marker for insulin resistance/ obesity • Marker for endothelial dysfunction • Marker for dysmetabolic syndrome • Marker for plaque vulnerability • Confounding by cigarette consumption • Innocent bystander- Acute phase response • Cytokine surrogate- IL-6, TNF-, IL-1 • Direct effects of CRP- Innate immunity- Complement activation- CAM induction • Prior infection- Chlamydia, H pylori, CMV

23. 3 2 1 0 hs-CRP and Risk of Future MI in Apparently Healthy Men P Trend <0.001 P<0.001 P<0.001 P=0.03 Relative Risk of MI 1 2 3 4 Quartile of hs-CRP Ridker et al, N Engl J Med. 1997;336:973–979.

24. hs-CRP and Risk of Future Stroke in Apparently Healthy Men P Trend <0.03 P<0.02 P=0.02 2 Relative Risk of Ischemic Stroke 1 0 1 2 3 4 Quartile of hs-CRP Ridker et al, N Engl J Med. 1997;336:973–979.

25. hs-CRP as a Risk Factor For Future CVD : Primary Prevention Cohorts Kuller MRFIT 1996 CHD Death Ridker 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 FHS 2001 Stroke Pradhan WHI 2002 MI,CVD death Albert PHS 2002 Sudden Death Sakkinen HHS 2002 MI 0 1.0 2.0 3.0 4.0 5.0 6.0 Relative Risk (upper vs lower quartile) Ridker PM. Circulation 2003;107:363-9

26. hs-CRP Adds to Predictive Value of TC:HDL Ratio in Determining Risk of First MI Relative Risk hs-CRP Total Cholesterol:HDL Ratio Ridker et al, Circulation. 1998;97:2007–2011.

27. Risk Factors for Future Cardiovascular Events: WHS Lipoprotein(a) Homocysteine IL-6 TC LDLC sICAM-1 SAA Apo B TC: HDLC hs-CRP hs-CRP + TC: HDLC 0 1.0 2.0 4.0 6.0 Relative Risk of Future Cardiovascular Events Ridker et al, N Engl J Med. 2000;342:836-43

28. Is there clinical evidence that inflammation can be modified by preventive therapies?

29. Elevated CRP Levels in Obesity: NHANES 1988-1994 Percent with CRP 0.22 mg/dL Normal Overweight Obese Visser M et al. JAMA 1999;282:2131-2135.

30. Effects of Weight Loss on CRPConcentrations in Obese Healthy Women • 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.

31. Effect of HRT on hs-CRP: the PEPI Study 3.0 2.0 1.0 CEE + MPA cyclic CEE + MPA continuous CEE + MP CEE hs-CRP (mg/dL) Placebo 0 12 36 Months Cushman M et al. Circulation 1999;100:717-722. 1999 Lippincott Williams & Wilkins.

32. Long-Term Effect of Statin Therapy on hs-CRP: Placebo and Pravastatin Groups Placebo 0.25 0.24 0.23 -21.6% (P=0.004) Median hs-CRP Concentration (mg/dL) 0.22 0.21 0.20 Pravastatin 0.19 0.18 Baseline 5 Years Ridker et al, Circulation. 1999;100:230-235.

33. * * * Effect of Statin Therapy on hs-CRP Levels at 6 Weeks 6 5 4 3 2 1 0 *p<0.025 vs. Baseline hs-CRP (mg/L) Baseline Prava(40 mg/d) Simva(20 mg/d) Atorva(10 mg/d) Jialal I et al. Circulation 2001;103:1933-1935. 2001 Lippincott Williams & Wilkins.

34. Effect of Bezafibrate with and without Fluvastatin on Plasma Fibrinogen, PAI-1, and CRP in Patients with CAD and Mixed Hyperlipidemia Fibrinogen PAI-1 CRP n: 81 80 74 Change at 24 weeks, % 70 72 63 83 80 75 * P<0.05 vs. baseline * * Beza 400 mg/d Beza 400 mg/d+ fluva 20 mg/d Beza 400 mg/d+ fluva 40 mg/d Cortellaro M et al. Thromb Haemost 2000;83:549-553.

35. JUPITERRandomized Trial of Rosuvastatin in the Primary Prevention of Cardiovascular Events Among Individuals with Low Levels of LDL-Cand Elevated Levels of hs-CRP MI Stroke Unstable Angina CVD Death CABG/PTCA Rosuvastatin (N=7500) Placebo (N =7500) No History of CAD Men > 55, Women > 65 LDL-C <130 mg/dL hs-CRP >2 mg/L 4 weekRun-in Randomization Visit End of Study Visit Screening Visit 1 Screening Visit 2 Safety Visit Bi-Annual Follow-Up Visits hs-CRP Lipids hs-CRP LFTs HbA1C hs-CRP LFTs UA hs-CRP LFTs UA Lipids HbA1C

36. N Engl J Med. 2002;347:1157-1165

37. Event-Free Survival According to Baseline Quintiles of C-Reactive Protein and LDL Cholesterol Quintiles of CRP Quintiles of LDL 1.00 1.00 1 0.99 0.99 1 2 2 3 0.98 0.98 3 CVD Event-Free Survival Probability 4 4 0.97 0.97 5 5 0.96 0.96 0 2 4 6 8 0 2 4 6 8 Years of Follow-Up Years of Follow-Up Ridker et al, N Engl J Med. 2002;347:1157-1165.

38. CV Event-Free Survival Using Combined hs-CRP and LDL-C Measurements Median LDL 124 mg/dl Median CRP 1.5mg/l 1.00 Low CRP-low LDL 0.99 Low CRP-high LDL 0.98 Probability of Event-free Survival High CRP-low LDL 0.97 0.96 High CRP-high LDL 0.00 4 0 2 6 8 Years of Follow-up Ridker et al, N Engl J Med. 2002;347:1157-1165.

39. hs-CRP Adds Prognostic Information at all Levels of LDL-C and at all Levels of the Framingham Risk Score <1.0 1.0-3.0 >3.0 <1.0 1.0-3.0 >3.0 C-Reactive Protein (mg/L) C-Reactive Protein (mg/L) 3 25 20 2 15 Relative risk Multivariable relative risk 10 1 5 0 0 0-1 2-4 5-9 10-20 <130 130-160 >160 Framingham estimate of 10-year risk (%) LDL cholesterol (mg/dL) Ridker et al, N Engl J Med. 2002;347:1557.

40. What is the role of hs-CRP with regard to diabetes and the metabolic syndrome?

41. Circulation. 2003;107:391-397.

42. Plasma hs-CRP Levels According to Severity of the Metabolic Syndrome 8 6 C-reactive protein (mg/L) 4 2 0 0 1 2 3 4 5 Number of Components of the Metabolic Syndrome Ridker et al, Circulation 2003;107:391-7

43. Event Free Survival According to hs-CRP Levels: Analysis Limited to Participants withMetabolic Syndrome at Baseline 1.00 0.99 0.98 CVD Event-Free Survival Probability CRP <1 mg/L 0.97 CRP 1-3 mg/L 0.96 CRP >3 mg/L 0.95 0 2 4 6 8 Years of Follow-Up Ridker et al, Circulation 2003;107:391-7

45. AHA / CDC Scientific StatementMarkers of Inflammation and Cardiovascular Disease: Applications to Clinical and Public Health PracticeCirculation January 28, 2003“Measurement of hs-CRP is an independent marker of risk and may be used at the discretion of the physician as part of global coronary risk assessment in adults without known cardiovascular disease. Weight of evidence favors use particularly among those judged at intermediate risk by global risk assessment”.

46. Clinical Application of hs-CRP forCardiovascular Risk Prediction >100 mg/L 3 mg/L 10 mg/L 1 mg/L Low Risk Moderate Risk Acute Phase Response Ignore Value, Repeat Test in 3 weeks High Risk Ridker PM. Circulation 2003;107:363-9

47. Inflammatory and Infections Agents in CHD • Belgian epidemiologic study included 446 of 16307 male workers aged 35-39 who had evidence of CHD vs. 892 controls. • CRP, but none of the infectious agents (H. pylori, C. pneumoniae, CMV, and EBV) were associated with CHD, even after adjustment for other risk factors. De Backer et al. Atherosclerosis 2002; 160: 457-63.

48. Infection and CHD - is there a connection? • Local or systemic infections resulting from gram negative bacteria such as Chlamydia pneumoniae and Helicobacter pylori, including cytomegalovirus (CMV) have been implicated in atheroscelosis • While several case control studies have shown increased titers of C.pneumoniae and H. Pylori in those with vs. without CHD, convincing evidence from prospective studies is lacking.

49. Prospective Studies of CHD and Infectious Pathogens • Physician’s Health Study (nested case-control) shows RR 1.1 (0.8-1.5) for C. Pneumoniae, 0.94 (0.7-1.2) for cytomegalovirus, and 0.72 (0.6-0.9) for Herpes simplex virus. • H. pylori also shows mixed results. Whincup showed a nonsignificant 1.3 OR when adjusted for other risk factors, the large ARIC study showed no relation, and the Caerphilly Prospective study showed RR=1.05 in 1796 men followed 14 years.

50. Other Studies of Infectious Agents • In South Asian persons with CHD vs. controls, C. pneumoniae specific IgG antibody was seropositive in similar proportions; risk factors appeared to mediate any relations (Mendis et al. Int J Cardiol 2001; 79: 191-6). • Cross-sectional survey of 704 individuals of C. pneumoniae and CMV with risk factors did nto show significant associations (Danesh et al., J Cardiovasc Risk 1999; 6: 387-90). • Meta-analysis of 24 articles involving H. pylori infection and CHD showed a pooled odds ratio of 1.55 (95% CI: 1.38-1.74) (p<0.001), suggested a weak relation, but high hetrogeneity between studies precludes clear demonostration (Pellicano et al., Eur J Epidemiol 1999; 15: 611-9). • ARIC Study failed to show clear relation between IgG antibodies for C. pneumoniae and incident CHD occurring over average 3.3 years. (Nieto et al. Am J Epidemiol 1999; 150: 149-56).