Update on Anacetrapib, a Novel CETP Inhibitor Douglas Johns, PhD, FAHA Basic Science Lead, CETP/Anacetrapib Merck Research Laboratories Rahway, NJ 1

Douglas Johns, PhD Salary: Merck and Company, Inc. Ownership Interest (Stocks, Stock Options, or other Ownership Interest):

Outline Review/update of Anacetrapib clinical programs DEFINE Hps3-TIMI55-REVEAL Basic science/mechanism-of-action studies Reverse cholesterol transport Bulk cholesterol excretion HDL function: Cholesterol efflux HDL function: Anti-inflammatory effects of HDL

Anacetrapib clinical program

Cannon CP et al. N Engl J Med 2010;363 (19) online 11/17/2010 Anacetrapib treatment had robust effects on HDL-C, LDL-C, non HDL-C and Lp(a) with sustained effects over 18 months. Anacetrapib had an acceptable side-effect profile with no effects on blood pressure, electrolytes or aldosterone. Within the power of the study, anacetrapib did not exhibit adverse cardiovascular effects seen with a prior CETP inhibitor The long term safety and efficacy of anacetrapib will now be tested in a large clinical outcomes trial.

Worldwide cardiovascular outcome study N=30,000, anacetrapib 100mg po or placebo Sponsor: University of Oxford Clinical Trial Service Unit Population: Stable cardiovascular disease with high risk of recurrent events Primary outcomes: Coronary mortality, myocardial infarction, coronary revascularization Follow-up: minimum median follow up of at least 4 years Progress: First patient enrolled May 2011, first patient randomized August 2011, currently enrolling in North America, Europe and Asia, enrollment is on schedule

Anacetrapib: basic science/ mechanism-of action studies

How does CETP inhibition affect HDL function? Cholesterol efflux and RCT Anti-oxidant Anti-inflammatory Endothelial function Improve β-cell function Anti-thrombotic Anti-apoptotic

How does CETP inhibition affect HDL function? Cholesterol efflux and RCT Anti-oxidant Anti-inflammatory Endothelial function Improve β-cell function Anti-thrombotic Anti-apoptotic

Comprehensive study of CETP inhibitors and cholesterol metabolism requires more than a single approach Study cholesterol movement with CETP inhibition using “classical approaches” Macrophage-to-feces RCT CETP-containing rodent species (e.g hamsters) Monitor changes in bulk cholesterol pools High-resolution mass spectrometry Assess HDL functionality with ex vivo methods Macrophage to feces RCT Bile acids

Macrophage to feces reverse cholesterol transport

Anacetrapib increases macrophage to feces reverse cholesterol transport in hamsters Increase in macrophage-derived 3H-tracer in HDL, suggesting increased cholesterol efflux from macrophages Increase in macrophage-derived 3H-chol and 3H-bile acids in the feces indicates net increase in macrophage-to-feces RCT 3H content in HDL 2.0 Control *** Anacetrapib (60mg/kg) 1.5 % of injected dose in HDL 1.0 0.5 0.0 24 48 72 Time (hours) 3H content feces 0.8 72 hours collection * 0.6 * % injected dose in feces 0.4 0.2 0.0 Control Ana Control Ana J Lipid Res. 2011, 52:1965-73 Cholesterol Bile acids

Cholesterol excretion Anacetrapib increases cholesterol in large HDL and promotes excretion of bulk cholesterol (no tracers) Cholesterol in HDL subfractions Increase in cholesterol is greatest in large HDL (HDL2) Beyond macrophage-to-feces RCT, ANA treatment increase fecal cholesterol and bile acid concentration These results indicate that anacetrapib is promoting movement of cholesterol out of the system From periphery to feces (RCT) Bulk cholesterol excretion 150 150 *** Vehicle Vehicle ** ** Ana 100 100 Cholesterol (mg/dL) 50 50 HDL3 HDL2 Day 0 Day 14 Ana (60mg/kg) Fecal cholesterol Fecal cholic acid 50 100 150 200 * ­ 29.4% Cholesterol excretion rate (ug/day) 5 10 19.3% CA ( m g/g feces) J Lipid Res. 2011, 52:1965-73

HDL functionality: Cholesterol efflux In addition to aqueous passive diffusion, cholesterol effluxes to HDL via protein-facilitated pathways ABCG1 and SR-BI facilitate cholesterol efflux to large/alpha HDL ABCA1 transports cholesterol to pre-beta HDL Ex vivo methods Apply serum from treated animals/humans and assess functional properties Macrophage ABCG1 α cholesterol SR-BI α ABCA1 Lipid-poor apoA-I Pre-β-HDL

Plasma from Anacetrapib-treated hamsters displays improved cholesterol efflux 14 ABCA1 Both major pathways of cholesterol efflux (alpha-HDL, pre-beta HDL) are increased with anacetrapib treatment ↑ABCA1-mediated efflux suggests increased pre-beta HDL component ↑ABCG1, ↑SR-BI mediated efflux indicates increased efflux to αHDL Similar observations with HDL from ANA-treated humans * 12 (% efflux) 10 8 6 Veh Ana SR-B1 ABCG1 5 2.0 ** *** 4 1.5 3 1.0 (% efflux) (% efflux) 2 0.5 1 0.0 Veh Ana Veh Ana J Lipid Res. 2011, 52:1965-73

Plasma from Anacetrapib-treated humans displays improved cholesterol efflux Total Cholesterol Free Cholesterol Cholesterol Ester *P<0,05, significant difference vs control PEG-HDL Increase in efflux at the same HDL-c concentration suggests an improvement in HDL “quality” 1Arterioscl Thromb Vasc Biol. 2010 30(7):1430-8

HDL from ANA-treated hamsters maintains ability to inhibit TNFα-induced inflammation in endothelial cells VCAM-1 expression 20 40 60 80 100 120 140 160 180 ** VCAM-1 mRNA Control TNF a Veh-HDL Ana-HDL HDL suppresses TNF-induced adhesion molecule expression1 VCAM, ICAM, E-selectin mRNA and protein HDL from ANA-treated hamsters shows no reduction in this effect (vs vehicle) Similar to observations with HDL from ANA-treated human1 Monocyte adhesion to endothelial cells suppressed by HDL No reduction in this effect with ANA-treated hamster HDL vs vehicle These results suggest that HDL functionality is improved, or not reduced with Anacetrapib treatment. ** p<0.01 vs. TNFα * p<0.05 vs. TNFα Monocyte adhesion 10 20 30 40 50 60 70 80 90 100 110 * % Adherent monocyte Control TNF a Veh-HDL Ana-HDL + 1Deuel Lipid Conference poster, Napa, CA 2011 2Arterioscl Thromb Vasc Biol. 2010 30(7):1430-8

Summary Multiple lines of evidence suggest that HDL functionality is improved or “maintained” with anacetrapib inhibition of CETP Enhanced macrophage-to-feces reverse cholesterol transport in hamsters Increased cholesteryl ester content in hamster HDL (enhanced maturation from HDL3 to HDL2) Improved excretion of bulk cholesterol/bile acids in hamsters Improvement in plasma cholesterol efflux capacity via ABCA1 (pre-beta-dependent), and via SR-BI/ABCG1 No reduction in anti-inflammatory effects of HDL