An Aggregate of Biomarkers Predict Very High Risk of AMI and Death Stephen E. Epstein Danny J. Eapen Christina Wassel Dylan Malayter Sergey Sikora Arshed A. Quyyumi Cardiovascular Research Institute, MedStar Health Research Institute Emory University University of California, San Diego GenWay Biotech, Inc.

Stephen E. Epstein, MD Intellectual Property Rights / Patent Holder: GenWay

The Courage Trial Randomly assigned 2287 patients with stable coronary disease to PCI plus optimal medical therapy or to optimal medical therapy alone. 4

Kaplan–Meier Survival Curves. The Courage Trial Figure 2. Kaplan–Meier Survival Curves. In Panel A, the estimated 4.6-year rate of the composite primary outcome of death from any cause and nonfatal myocardial infarction was 19.0% in the PCI group and 18.5% in the medical-therapy group. In Panel B, the estimated 4.6-year rate of death from any cause was 7.6% in the PCI group and 8.3% in the medical-therapy group. In Panel C, the estimated 4.6-year rate of hospitalization for acute coronary syndrome (ACS) was 12.4% in the PCI group and 11.8% in the medical-therapy group. In Panel D, the estimated 4.6-year rate of acute myocardial infarction was 13.2% in the PCI group and 12.3% in the medical-therapy group. Conclusion: As an initial management strategy there is no difference in outcomes between PCI plus optimal medical management vs. optimal medical management alone.

The Courage Trial Implication for standard of care for the patient with possible CAD: Since optimal medical therapy is just as effective as PCI as an initial approach to the patient with suspected CAD: PCI as an initial management strategy is not indicated; It could therefore also be argued that angiography is not indicated as an initial approach to such patients. Figure 2. Kaplan–Meier Survival Curves. In Panel A, the estimated 4.6-year rate of the composite primary outcome of death from any cause and nonfatal myocardial infarction was 19.0% in the PCI group and 18.5% in the medical-therapy group. In Panel B, the estimated 4.6-year rate of death from any cause was 7.6% in the PCI group and 8.3% in the medical-therapy group. In Panel C, the estimated 4.6-year rate of hospitalization for acute coronary syndrome (ACS) was 12.4% in the PCI group and 11.8% in the medical-therapy group. In Panel D, the estimated 4.6-year rate of acute myocardial infarction was 13.2% in the PCI group and 12.3% in the medical-therapy group.

Kaplan–Meier Survival Curves. Patients with stable CAD The Courage Trial Figure 2. Kaplan–Meier Survival Curves. In Panel A, the estimated 4.6-year rate of the composite primary outcome of death from any cause and nonfatal myocardial infarction was 19.0% in the PCI group and 18.5% in the medical-therapy group. In Panel B, the estimated 4.6-year rate of death from any cause was 7.6% in the PCI group and 8.3% in the medical-therapy group. In Panel C, the estimated 4.6-year rate of hospitalization for acute coronary syndrome (ACS) was 12.4% in the PCI group and 11.8% in the medical-therapy group. In Panel D, the estimated 4.6-year rate of acute myocardial infarction was 13.2% in the PCI group and 12.3% in the medical-therapy group. End point: Death or AMI A concern with this strategy is that: All potential patients for Courage had angiography, and many were eliminated from consideration because of angiographic findings; Despite this, there was still appreciable risk of AMI and death in the Courage cohorts.

Attempt to identify a high risk subgroup in the Courage cohort

Rates of Death or MI Related to Residual Ischemia [COURAGE substudy of 314 pts: 5 yr follow-up] Death or MI Rate (%) 0% (n=23) 1 - 4.9% (n=141) 5 - 9.9% (n=88) > 10% (n=62) Shaw LA. AHA 2007

Therefore, the ISCHEMIA Trial is about to be initiated. Purpose: To attempt to identify a high risk subgroup of pts with stable CAD and to determine the best management strategy for them. Multicenter randomized trial of patients with at least moderate ischemia on stress imaging. Patients randomized to: a routine invasive strategy with cardiac catheterization followed by revascularization plus optimal medical therapy, or to… a conservative strategy of optical medical therapy--with cath and revascularization reserved for those with an acute coronary syndrome, ischemic heart failure, resuscitated cardiac arrest or refractory symptoms. What about those patients who experience rupture of a plaque that prior to rupture produced <50% stenosis—ie, no induced ischemia prior to AMI?

What does kill is plaque rupture. Key Concept: Coronary plaques can, if severe enough, cause angina-- but they don’t usually kill. What does kill is plaque rupture. Since plaques producing <50% stenosis can rupture, the demonstration of ischemia is not necessarily the best way to identify the patient at risk of plaque rupture. 11

The clinical question we attempted to address in our study was: Is there a subgroup of patients within the Courage-like cohort, independent of the presence or absence of ischemia, that is at particularly high risk of AMI and death—a risk high enough to warrant consideration for: a more aggressive diagnostic approach (coronary angiography). more aggressive therapeutic intervention; i.e., with more intense medical therapy or even, if risk is sufficiently high, consideration for PCI or bypass surgery? 12

The strategy we used to identify the patient at risk of AMI is based on the following concepts… 13

Induction of plaques vulnerable to rupture requires activation of multiple intraplaque “injury” processes; each contributes to plaque vulnerability. Hypothesis: The greater the number of abnormal pathways the greater the likelihood of ... Hansson GK, Libby P. Nat Rev Immunol. 2006;6:508-19 14

…plaque rupture Hansson GK, Libby P. Nat Rev Immunol. 2006;6:508-19 15

Aggregate of Biomarkers We therefore hypothesized that an aggregate of pathway-related biomarkers comprised of: C-reactive protein (CRP)—inflammatory pathway; Fibrin degradation products (FDP)—thrombotic pathway; Heat Shock Protein 70 (HSP70)—immune/inflammatory/cell stress pathways. …by identifying individuals with abnormalities in multiple pathways that are associated with plaque vulnerability, would improve our capacity to identify individuals at low risk, and at high risk, of future AMI or death.

Design: Case-Control Subjects: 3800 individuals admitted to Emory Cardiac Catheterization Laboratory; Blood drawn prior to catheterization. 2004-present with mean follow-up of 2.75 years.

Aggregate of Pathway–Specific Biomarker Risk Score Patients with Significant CAD

Aggregate of Pathway–Specific Biomarker Risk Score Patients with Non-significant CAD

Hazard Ratios All Participants* HR(95% CI); p-value Significant CAD **   All Participants* HR(95% CI); p-value Significant CAD ** Non-significant CAD† Biomarkers CRP at 3.0 mg/L HSP70 at 0 ng/mL FDP at 1.0 ug/mL 1.64 (1.31, 2.07); <0.0001 2.39 (1.87, 3.04); <0.0001 1.69 (1.34, 2.12); <0.0001 1.60 (1.20, 2.12); 0.0011 2.37 (1.75, 3.22); <0.0001 1.53 (1.15, 2.04); 0.0040 2.08 (1.40, 3.08); 0.0003 1.82 (1.25, 2.65); 0.0018 2.02 (1.38, 2.96); 0.0003 Continuous Biomarker Risk Score 1.84 (1.64, 2.07); <0.0001 1.74 (1.50, 2.02); <0.0001 1.95 (1.61, 2.36); <0.0001 Categorical Biomarker Risk Score 1 vs 0 markers 2 vs 0 markers 3 vs 0 markers 1.85 (1.35, 2.54); 0.0001 3.73 (2.71, 5.13); <0.0001 5.66 (3.78, 8.49); <0.0001 2.08 (1.42, 3.06); 0.0002 3.38 (2.28, 5.03); <0.0001 5.37 (3.17, 9.09); <0.0001 1.40 (0.82, 2.38); 0.22 4.11 (2.42, 6.98); <0.0001 5.50 (2.91, 10.38); <0.0001 * Adjustment for age, race, gender, BMI, ever smoking, HTN, diabetes, aspirin use, statin use, acute MI, previous MI, Gensini score, hyperlipidemia and eGFR ** Adjustment for age, race, gender, BMI, ever smoking, HTN, diabetes, aspirin use, plavix use, statin use, PCI history, acute MI, previous MI, Gensini score, hyperlipidemia, and eGFR † Adjustment for age, race, gender, BMI, ever smoking, HTN, diabetes, aspirin use, statin use, Gensini score, hyperlipidemia, and eGFR Note: these models adjust for AMI

Annual event rate (AMI or death)   10 20 30 Non-significant CAD CAD patients AMI or Death/year = 1 biomarker = 0 biomarkers = 2 biomarkers = 3 biomarkers 1.8 2.5 8.4 14.2 2.4 5.0 9.3 18.2 37% 44% 14% 5% 40% 40% 16% 4%

Results of adding the “Aggregate of Pathway-Specific Biomarkers” to standard risk factors on the C-statistic. 22

Observed AUC or C-statistic for Framingham-based models with and without the addition of CRP 0.5 0.65 0.75 0.9 No discrimination Framingham Framingham + CRP Shah et al. Int. J. Epidemiology. 2009 23

(Lower Bound, Upper Bound) +CRP, FDP, HSP70 in one model† C-statistics for our study: all participants C-statistic (Lower Bound, Upper Bound) p-value* Base Model** 0.695 (0.65, 0.725) -- +CRP, FDP, HSP70 in one model† 0.750 (0.728, 0.776) 7.1311 x 10-8 *The p-values refer to comparing the C-statistics for models containing biomarkers with the baseline model. **Baseline model includes age, sex, race, BMI, ever smoking, diabetes, HTN, aspirin use, Plavix use, statin use, previous AMI, Gensini Score, lipid Composite, and GFR. †CRP cut at 3.0, HSP70 cut at 0, and FDP cut at 1.0 are added to the baseline model 24

Conclusions: Individuals with all 3 biomarkers abnormal (CRP, HSP70, and FDP), whether they fall into the category of non-significant (<50% stenosis) or significant CAD, are at about 5.5 times greater risk of AMI/death over the next two years—despite having optimal medical management. Their event rates after one year of follow-up are 14% and 18%, respectively. Implications: It is unknown whether clinical outcomes of these high risk individuals will improve with more aggressive therapeutic approaches—such as more intensive medical therapy, PCI, or bypass surgery. However, if these biomarker/risk findings are confirmed, it could be argued that more aggressive therapeutic approaches to these very high risk patients should be at least considered, until prospective trials are carried out.