1. INCITE Study Co-PI: Frederick Welt, M.D. Co-PI: Daniel I. Simon, M.D. Brigham and Women’s Hospital West Roxbury VA Medical Center Harvard Medical School Boston, MA INflammatory Cessation with abcIximab or InTEgrilin INCITE

2. InflammationInflammationAtherosclerosisAtherosclerosis ThrombosisThrombosis Thrombus Quiescent plaque Platelets and thrombin Plaque rupture Acute Coronary Syndromes Evolving Understanding of Pathophysiology CD40L

3. CD40L in Immunity: Isotype Switching TM TNF Homology Helper T cell B cell CD40LCD40L sCD40LsCD40L NNCC CD40LCD40L CD40CD40

4. CD40L Distribution 300 200 100 0 CD40L Units/mL blood BloodPlateletsLymphocytes CD40L Units/1x10 6 cells P=0.016 Cells PlateletsLymphocytes 3 2 1 0 Henn et al., Blood, 2001, 98, 1047 Lindmark et al. ATVB, 2000, 20, 2322 Andre et al., Nature Med., 2002, 8, 247

5. CD40L in Vascular Disease CD40L antibody inhibits atherosclerotic lesion progression in LDLR -/- mice CD40L gene deletion inhibits lesion progression in Apo E-/-mice Soluble form detected in ACS and PCI Elevated plasma levels of sCD40L identify healthy women at risk for CV events. CD40L antibody inhibits atherosclerotic lesion progression in LDLR -/- mice CD40L gene deletion inhibits lesion progression in Apo E-/-mice Soluble form detected in ACS and PCI Elevated plasma levels of sCD40L identify healthy women at risk for CV events. Nature 1998;391:591 PNAS 2000;97:7458 Nature Med 1999;5:1313 Circulation 1999;100:614 Circulation 2001;104:2266

6. Platelet CD40L-Induced Inflammation thrombin CD40L CD40L Cultured HUVECs IL-8 MCP-1 ICAM-1 VCAM-1 TF Modified from: Nature 1998;391:591 sCD40L

7. (min) WT CD40L-/- CD40 -/- 0 10 20 30 40 * CD40L-/- + rsCD40L Time to reach occlusion sCD40L Promotes Thrombosis: Delayed Arterial Occlusion in CD40L-/- Mice 30 min after FeCl 3 WT CD40L-/- * P < 0.0005 † P < 0.02 Nature Med, 2002, v.8, 247-52

8. GP IIb-IIIa Antagonism: sCD40L Release and Aggregation 012345 125 100 75 50 25 0 Eptifibatide (  M) 125 100 75 50 25 0 00.20.40.60.81 Tirofiban (  M) Abciximab (  M) 00.20.40.60.81 % sCD40L Release % aggregation Circulation, Supplement. 2001;104:II-318, 1533

9. sCD40LsCD40L Inflammation Thrombosis Functions of Released sCD40L sCD40LsCD40L GP IIb/IIIa

10. Abciximab: Anti-Inflammatory Action AM Lincoff at al. Circ 2001;104:163

11. GP IIb-IIIa Inhibitors: Specificity/Selectivity IIb/IIIaxxxxxxxxx  V  3 xxx?x Mac-1x AbciximabEptifibatideTirofiban Charo IF et al. Proc Natl Acad Sci. 1986;83:835 Coller BS et al. Blood. 1991;77:75 Altieri DC et al. J Immunol. 1988;141:2656 Simon DI et al. Arterioscler Thromb Vasc Biol 1997;17:528 Lele M et al. Circ 2001;104:582

12. INCITE: Hypothesis  Our central hypothesis is that GP IIb-IIIa inhibitors will modulate peri-PCI inflammation by virtue of their effect on platelet-induced leukocyte activation. INCITE

13. INCITE: Study Design Elective PCI ASA, heparin (50-70 U/kg) Elective PCI ASA, heparin (50-70 U/kg) RPFA baseline and 10 min post-bolus ELISA: sCD40L, RANTES, IL-6, sICAM-1 Timepoints: Baseline, 30 min, 2 hrs, 24 hrs RPFA baseline and 10 min post-bolus ELISA: sCD40L, RANTES, IL-6, sICAM-1 Timepoints: Baseline, 30 min, 2 hrs, 24 hrs No GP IIb-IIIa (n=15) No GP IIb-IIIa (n=15) Abciximab 0.25 mg/kg 0.1  g/kg/min (n=15) Abciximab 0.25 mg/kg 0.1  g/kg/min (n=15) Eptifibatide 180/180  g/kg 2  g/kg/min (n=15) Eptifibatide 180/180  g/kg 2  g/kg/min (n=15) INCITE

14. Exclusion Criteria -Patients already on GP IIb/IIIa inhibitor -Patients with clear indications for GP IIb/IIIa: -Acute ST elevation MI or non q-wave MI with CPK elevation -Pain at rest with EKG changes < 24 hours -Visible Thrombus -Pain refractory to heparin -Lesions located in vascular grafts or at sites of prior intervention -Bleeding disorder -Thrombocytopenia (plts< 100) -History of GI bleeding -History of stroke or intracranial bleed -Uncontrolled hypertension (SBP > 200 mm Hg, DBP> 100 mm Hg) -Condition requiring oral anticoagulation -History of allergy or contraindication for aspirin or heparin

15. NoneAbciximabEptifibatidep-value n15 Age70.3±11.464.3±9.761.6±10.8NS Diabetes Mellitus, %20.046.726.7NS Hypertension, %66.780.073.3NS Elevated Cholesterol, %66.793.380.0NS Current Smoker, %26.740.0 NS Prior MI, %26.753.3 NS Prior PCI, %26.753.320.0NS Prior CABG, %33.346.7 NS Unstable Angina, %20.06.720.0NS WBC6.6±1.87.1±1.97.3±2.0NS Platelet223±80227±61246±67NS Hematocrit37.1±3.337.4±4.141.1±3.6NS Creatinine1.1±0.31.0±0.2 NS Baseline Characteristics

16. Procedural Characteristics NoneAbciximabEptifibatidep-value n15 Activated Clotting Time267±49287±68317±55NS Platelet Inhibition, % (RPFA) N/A93±498±3P<0.01 Multivessel PCI, %26.733.340.0NS

17. INCITE Plasma sCD40L Post-PCI 30 min2 hour 24 hour ANOVA P=0.018 P<0.03 None vs Eptifibatide -0.1 -0.05 0 0.05 0.1 Delta Plasma Concentration (ng/ml) None Eptifibatide Abciximab

18. Plasma RANTES Post-PCI (Pro-inflammatory Platelet derived chemokine) INCITE 30 min2 hour 24 hour ANOVA P=0.006 P<0.005 None vs Eptifibatide -150 -100 -50 0 50 100 Delta Plasma Concentration (ng/ml) None Eptifibatide Abciximab

19. INCITE Plasma sICAM-1 Post-PCI 30 min2 hour 24 hour ANOVA P=0.23 -100 -50 0 50 100 Delta Plasma Concentration (ng/ml) None Eptifibatide Abciximab

20. INCITE Plasma IL-6 Post-PCI 30 min2 hour 24 hour ANOVA P=0.1 0 1 2 3 4 5 6 Delta Plasma Concentration (ng/ml) None Eptifibatide Abciximab

21. Summary INCITE  GP IIb/IIIa inhibition lowers levels of sCD40L and RANTES post stenting, possibly conferring anti-inflammatory as well as anti-thrombotic effects.

22. Conclusion INCITE  These findings may have significant implications for long term benefits associated with GP IIb/IIIa inhibitor use.  The findings of this pilot study need to be examined in a larger clinical trial that confirms these findings and correlates reduction in inflammation with improved clinical outcome.  These findings may have significant implications for long term benefits associated with GP IIb/IIIa inhibitor use.  The findings of this pilot study need to be examined in a larger clinical trial that confirms these findings and correlates reduction in inflammation with improved clinical outcome.

23. Procedural Characteristics NoneAbciximabEptifibatidep-value n15 Activated Clotting Time267±49287±68317±55NS Platelet Inhibition, % (RPFA) N/A93±498±3P<0.01 Multivessel PCI, %26.733.340.0NS

24. Platelet aggregation inhibition with abciximab, eptifibatide, and tirofiban RIAR Study (Keriakes. Am J Cardiol 1999) tirofiban 0.4/0.1 (n=9) eptifibatide 180/2 (n=10) abciximab 0.25/0.125 (n=10) 0.7261224 0.7261824 0.7261824 Time from bolus (h)

25. CRP INCITE Preliminary Results