Use of Biomarkers for Detection of Acute Myocardial Infarction

Use of Biomarkers for Detection of Acute Myocardial Infarction Allan S. Jaffe, MD.* Consultant - Cardiology & Laboratory Medicine Professor of Medicine Chair, CCLS Division, Department of Laboratory Medicine and Pathology Mayo Clinic and Medical School Rochester, Minnesota *Dr. Jaffe is or has been a consultant to most of the major diagnostic companies as well as Pfizer and Amgen. CP887679-0

Biomarkers for Detection of Myocardial Infarction Preferably Detection of rise and/or fall of Troponin (I or T) with at least one value above the 99 th percentile of the upper reference limit measured with a coefficient of variation 10%. CP887679-1

Biomarkers for Detection of Myocardial Infarction Preferably Detection of rise and/or fall of Troponin (I or T) with at least one value above the 99 th percentile of the upper reference limit measured with a coefficient of variation 10%. CP887679-2

GUSTO-IV (Death and MI) % 8 7 6 5 4 3 2 1 0 Death Myocardial infarction 22 50 20 0.01 0.01-0.03 >0.03-0.1 >0.1 99 % 10%CV ROC Evaluated cutoff points for troponin T (µg/l) James et al: Am J Med 115:182, 2003 31 33 66 201 235 CP1142356-16 CP887679-3

Table 1. Analytical characteristics of commercial and research cardiac troponin I and T assays declared by the manufacturer. Commercially available assays - Company/ platform(s)/ assay LoB # (ng/l) LoD* (ng/l) 99 th % (ng/l) %CV at 99 th % 10% CV (ng/l) Risk Stratificati on Epitopes recognised by Antibodies Abbott AxSYM ADV 20 40 14.0 160 Yes C 87-91, 41-49; D 24-40 ALP Detection Antibody Tag Abbott ARCHITECT <10 28 14.0 32 Yes (No in US) C 87-91, 24-40; D: 41-49 Acridinium Abbott i-stat 20 80 16.5 100 Yes C: 41-49, 88-91; D: 28-39, 62-78 ALP Alere Triage SOB 50 NAD NA NA No C: NA; D: 27-40 Fluorophor Alere Triage Cardio 3 (r) 10 NAD 17.0 (at 20) NA No NA Fluorophor Beckman Coulter Access Accu 10 40 14.0 60 Yes C: 41-49; D: 24-40 ALP biomerieux Vidas Ultra 10 10** 27.7 110 No C: 41-49, 22-29; D: 87-91, 7B9 ALP Mitsubishi Chemical PATHFAST 8 29 5.0 14 No C: 41-49; D: 71-116, 163-209 ALP Ortho Vitros ECi ES 12 34 10.0 34 Yes C: 24-40, 41-49; D: 87-91 HRP Radiometer AQT90 FLEX TnI 9.5 23 17.7 39 NA C: 41-49, 190-196; D: 137-149 Europium Radiometer AQT90 FLEX TnT 10 17 15.2 25 NA C: 125-131; D: 136-147 Europium Response Biomedical RAMP 30 NAD 18.5 (at 50) 210 No C: 85-92; D: 26-38 Fluorophor CP887679-4

http://ifcc.org/pdf/scientificactivities/ IFCC_Troponin_Table_vDec_2010_ FINAL_ng_L_28Jan11.pdf CP887679-5

Table 1. Analytical characteristics of commercial and research cardiac troponin I and T assays declared by the manufacturer. Commercially available assays - Company/ platform(s)/ assay Abbott AxSYM ADV 20 LoB # (ng/l) Lo D* (ng /L) 40 99 th % (ng/l) %CV at 99 th % 10% CV (ng/l) Risk Stratificati on Epitopes recognised by Antibodies 14.0 160 Yes C 87-91, 41-49; D 24-40 ALP Detection Antibody Tag Abbott ARCHITECT <10 28 14.0 32 Yes (No in US) C 87-91, 24-40; D: 41-49 Acridinium Abbott i-stat 20 Alere Triage SOB 50 80 NAD 16.5 100 Yes C: 41-49, 88-91; D: 28-39, 62-78 ALP NA NA No C: NA; D: 27-40 Fluorophor Alere Triage Cardio 3 (r) 10 NAD 17.0 (at 20) NA No NA Fluorophor Beckman Coulter Access Accu 10 biomerieux Vidas Ultra 10 Mitsubishi Chemical PATHFAST 8 Ortho Vitros ECi ES 12 40 10** 29 34 14.0 60 Yes C: 41-49; D: 24-40 ALP 27.7 110 No C: 41-49, 22-29; D: 87-91, 7B9 ALP 5.0 14 No C: 41-49; D: 71-116, 163-209 ALP 10.0 34 Yes C: 24-40, 41-49; D: 87-91 HRP Radiometer AQT90 FLEX TnI 9. 5 23 Radiometer AQT90 FLEX TnT 1 0 17 17.7 39 NA C: 41-49, 190-196; D: 137-149 Europium 15.2 25 NA C: 125-131; D: 136-147 Europium Response Biomedical RAMP 30 NAD 18.5 (at 50) 210 No C: 85-92; D: 26-38 Fluorophor http://ifcc.org/pdf/scientificactivities/ifcc_troponin_table_vdec_2010_final_ng_l_28jan11.pdf CP887679-6

AUC More Diagnoses with Higher-Sensitivity Assays 1.0 Sensitive troponin I 99 th % value 0.8 0.6 0.4 Troponin T 10%CV value 0.2 0.0 NEJM 361:868, 2009 0 to <3 3 to <6 6 to <9 9 to <12 12-15 All Time of chest-pain onset (hr) Diagnosis of AMI (no.) 227 97 44 29 17 528 Single determination (no.) Sensitive troponin I 184 100 38 32 25 534 >0.04 ng/ml Troponin T >0.03 ng/ml 81 56 19 21 12 305 CP887679-7

Area under the ROC curve Accuracy by Time of Admission 1.000 0.950 0.900 99 th % value 0.850 0.000 NEJM 361:858, 2009 Mostly same assays but < 99 th % value cut off used 0.800 0.750 0.700 Abbott-architect troponin I Roche high-sensitive troponin T Roche troponin I Siemens troponin I ultra Standard assay 2 4 6 10 All patients Hours since onset of symptoms CP887679-8

Sensitivity Sensitivity Diagnostic Accuracy of Biomarkers at the Time of Admission 1.0 All Patients 1.0 Chest-Pain Onset <3 Hr 0.8 0.6 0.4 0.2 0.0 1.0 0.8 Troponin I (0.96) Troponin T (0.85) Myoglobin (0.82) Creatine kinase MB (0.73) Creatine kinase (0.70) 0.0 0.2 0.4 0.6 0.8 1.0 Chest-Pain Onset <6 Hr 0.8 0.6 0.4 0.2 0.0 1.0 0.8 Troponin I (0.95) Troponin T (0.76) Myoglobin (0.83) Creatine kinase MB (0.64) Creatine kinase (0.62) 0.0 0.2 0.4 0.6 0.8 1.0 Chest-Pain Onset <12 Hr 0.6 0.4 0.2 0.0 0.0 0.2 0.4 0.6 0.8 1.0 NEJM 361:868, 2009 1-specificity Troponin I (0.95) Troponin T (0.79) Myoglobin (0.83) Creatine kinase MB (0.67) Creatine kinase (0.63) 0.6 0.4 0.2 0.0 0.0 0.2 0.4 0.6 0.8 1.0 1-specificity Troponin I (0.96) Troponin T (0.81) Myoglobin (0.83) Creatine kinase MB (0.70) Creatine kinase (0.65) CP887679-9

Reduction in Time with a 3 Compared to a 6-9 Hour Sampling Schedule 60 50 Relative frequency (%) 40 30 20 10 0 <1 1-3 4-6 7-9 >9 Reduction in time to diagnosis (hr) Clin Chem 52(5):812, 2006 CP887679- CP1232829-4 10

Biomarkers for Detection of Myocardial Infarction Preferably Detection of rise and/or fall of Troponin (I or T) with at least one value above the 99 th percentile of the upper reference limit measured with a coefficient of variation 10%. CP887679-11

All-Cause Mortality by Cardiac Troponin T (n=733) Cumulative survival (%) 100 80 60 40 20 0 Time since blood draw (years) ctnt <0.01 g/l ctnt 0.04 g/l ctnt 0.04 to 0.10 g/l ctnt 0.10 g/l 0.0 0.5 1.0 1.5 2.0 2.5 3.0 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 Circulation 106:2944, 2002 CP1090800-14 CP887679-12

ctnt Determinants of Elevation in the Community* 40 P<0.0001 30 % with elevated ctnt 20 10 0 0 1 2 >3 Risk determinants present (no.) No. 2,087 478 120 22 *Circulation, 113: 1958 1965, 2006 CP887679- CP1219620-13 13

CV (%) Elecsys Troponin T: Functional sensitivity (trend analysis) 50,0 40,0 20% CV: 0.015 ng/ml 10% CV: 0.035 ng/ml 5% CV: 0.080 ng/ml 30,0 20,0 10,0 0,0 0,00 0,02 0,04 0,06 0,08 0,10 0,12 0,14 0,16 0,18 Concentration (µg/l) Data from the multicenter evaluation; each dot corresponds to a day to day imprecision data set of at least 11 runs (August 1998 - March 1999 in 8 labs on 2010 and 1010) CP887679-14

DETERMINING ASSAY VALUES ARE DIFFERENT CP887679-15

Baseline Elevations in Patients Without AMI 2 Roche high-sensitive troponin T Siemens troponin I ultra Abbott-Architect troponin I 1 0 >70 years 70 years Reiter M et al: EHJ 32:1379-1389, 2011 CP887679-16

Short- and Long-term Bio variation in hsctnt Analytical variation Short Term Long term CVA, % 53.5 98 Biological Variation CVI, % 48.2 CVG, % 85.9 Index of Individuality 84.5 RCV, log-normal increase, % 84.6 94 94 19.6 315 CP887679-17

Use of high sensitivity troponin T to diagnose myocardial infarction Clinical setting consistent with myocardial ischemia Baseline < 14 ng/l 14-52 ng/l 53 ng/l Retest hstnt 3 hours after symptom onset or if timing of symptom onset is unclear at 6 hours after presentation Retest hstnt 3 hours later Change < 50% Change 50% Retest hstnt 3 hours later Change < 20% Change 20% 14ng/L rules out MI with >90% probability If 14ng/L then proceed to middle part of algorithm. Adverse Prognosis Retest hstnt at 6,12 hr Evidence based treatments Myocardial infarction CP887679-18 White HD; AHJ 2010

ug/l Absolute and Relative Changes of ctn Over 1- and 2-hours 0.400 0.300 Hs-cTnT 1h absolute changes Hs-cTnT 2h absolute changes ctnl Ultra 1h absolute changes ctnl Ultra 2h absolute changes 300 Hs-cTnT 1h relative changes Hs-cTnT 2h relative changes ctnl Ultra 1h relative changes ctnl Ultra 2h relative changes 0.300 0.200 200 0.200 % 100 0.100 0.100 0 0.000 0.000 AMI Unstable angina Cardiac but not CAD Non- Unknown cardiac chest pain -100 AMI Unstable angina Cardiac but not CAD Non- Unknown cardiac chest pain Reichlin et al: Circulation 124:135, 2011 CP887679-19

Myocardial Infarction Type 2 Myocardial infarction secondary to ischemia due to either increased oxygen demand or decreased supply in a setting where coronary supply is at least some part of the problem, e.g. spasm, fixed coronary artery disease, endothelial dysfunction. CP887679-20

ctnt and Angiographic Measures Stenosis (%) Thrombus (%) TFG 0/1 (%) TMPG 0/1 (%) 74 P<0.001 50 P=0.02 20 P=0.03 70 P<0.001 72 70 72.0 40 42.5 15 15.6 60 50 58.1 68 66 64.8 30 20 29.3 10 7.0 40 30 42.1 64 62 10 5 20 10 60 0 0 0 Prog Cardiovasc Dis 47(3), 2004 TnT (<0.01) TnT (>0.01) CP887679- CP1179389-5 21

Subsets of the Population with ACS Studied 200 150 Culprit lesion No Yes P<0.001 168 Pt (no.) 100 106 108 76 50 0 2 28 STEMI (n=108) NSTEMI (n=196) UAP (n=184) Ong et al: JACC 52:523, 2008 CP887679- CP1322205-2

Response to Acetylcholine 50 40 Reproduction of symptoms No Yes 42 Pt (no.) 30 20 15 10 4 7 10 8 0 Ong et al: JACC 52:523, 2008 0-25 25-75 >75 Vasoconstriction to acetylcholine (%) CP887679- CP1322205-3 23

Cum survival All-Cause Mortality 1.0 0.8 0.6 No Yes 0.4 0.2 0.0 P<0.0005 0 200 400 600 800 1,000 1,200 Culprit lesion 76 76 76 76 75 75 Culprit lesion + 270 240 231 229 224 219 Ong et al: JACC 57(2):147, 2011 CP887679-24

Myocardial Infarction Type 4a PCI-related increase of biomarkers (assuming a normal troponin baseline) > 3 X 99th percentile of the upper reference limit (URL) is by convention defined as myocardial infarction. R CP887679-25

Values ctn CK-MB Upper reference value Upper reference value 0 2 4 6 8 10 12 Time (hr) 0 2 4 6 8 10 12 Time (hr) CP887679-26

CK-MB (ng/ml) Magnitude of CK-MB Elevations at Each Time Point Baseline ctnt Elevation 200 150 100 50 0-50 31.3 65.8 25.3-100 None 8-hour peak 16-hour peak Miller, Jaffe, EHJ 27:1061, 2006 CP887679- CP1217284-8 27

CK-MB (ng/ml) Magnitude of CK-MB Elevations at Each Time Point No Baseline ctnt Elevation 50 30 10-10 2.25 7.42 9.79-30 -50 None 8-hour peak 16-hour peak Miller, Jaffe, EHJ 27:1061, 2006 CP887679- CP1217284-6 28

Death or MI (%) Pre-PCI Troponin and Long-Term Mortality 40 35 30 25 20 15 10 5 0 <0.01 0.01-0.03 >0.03-0.1 >0.1-0.5 >0.5-1.0 >1.0 P<0.001 0 12 24 36 48 60 Months from PCI Prasad et al: Circ Cardiovasc Intervent 1:10, 2008 CP887679- CP1331879-12 29

Mortality (%) 30-Day Mortality According to Periprocedural ctnt Levels 2.5 2.0 P<0.001 Kaplan-Meier Estimates Pre 0.01 2.3% 1.5 1.0 0.5 0.0 8/13 events non cardiac Pre <0.01, post >0.01 0.6% Pre <0.01, post <0.01 0.1% 0 5 10 15 20 25 30 Days from PCI Prasad, Jaffe et al: Circ Cardiovasc Intervent 1:10, 2008 CP887679- CP1331879-11 30

Mortality (%) Long-Term Mortality According to Periprocedural ctnt Levels 30 25 20 15 10 5 P<0.001 Pre 0.01 Pre <0.01, post >0.01 Pre <0.01, post <0.01 0 0 12 24 36 48 60 Months from PCI Prasad, Jaffe et al: Circ Cardiovasc Intervent 1:10, 2008 CP887679- CP1331879-14 31

Multivariable Cox Model for Long-Term Risk of Death 95% hazard Variable 2 Hazard ratio ratio CL P Pre-PCI ctnt elevation 15.9 1.79 1.35, 2.39 <0.001 Age 83.0 * <0.001 Congestive heart failure 42.8 2.11 1.69, 2.65 <0.001 Body mass index 25.5 * <0.001 Chronic renal failure 22.5 2.256 1.61, 3.16 <0.001 Diabetes 19.4 1.60 1.30, 1.97 <0.001 Ejection fraction 11.3 * 0.010 Cerebrovascular disease 9.3 1.43 1.14, 1.80 0.002 Smoking 5.3 1.28 1.04, 1.60 0.021 Isolated post-pci ctnt elevation 3.4 1.31 0.98, 1.75 0.065 Prasad, Jaffe et al: Circ Cardiovasc Intervent 1:10, 2008 CP887679- CP1331879-15 32

Event-free survival 1.0 0.8 0.6 Survival with by ctn post PCI 1.00 Troponin 0.45 ng/ml Troponin >0.45 ng/ml P=0.04 0.4 0.2 0.0 After covariate correction, p = ns 0 100 200 300 400 500 600 700 Follow-up (days) Cavallini et al: Circ Cardio Interv 3:431, 2010 0.97 0.94 0.91 0 100 200 300 400 500 600 700 CP887679-33

Criteria for Type 4A when baseline values are elevated but stable Use reinfarction criteria 20% rise from the baseline value CP887679-34