H. Bryan Brewer, Jr., MD Medstar Heart Institute Update on HDL Infusions And Atherosclerosis Regression in the ACS Patients H. Bryan Brewer, Jr., MD Medstar Heart Institute Washington, DC

H. Bryan Brewer, MD Consulting: Merck and Company, Inc., Roche, Sanofi-Aventis, Genentech, Pfizer, Inc., AstraZeneca Honoraria: Merck and Company, Inc., Roche, Sanofi-Aventis, Eli Lilly and Company, and Pfizer, Inc. Stocks, Stock Options, other ownership interest: The Medicines Company, InfraReDx, Inc, HDL Therapeutics

Acute HDL Therapy: Focus on Infusion of Preβ-HDL Liver Intestine LDLr CETP LDL CD36 SR-A ABCA1 SR-BI LCAT A-I αHDL A-I αHDL Preβ-HDL Lipid Filled Macrophage in the Heart Acute HDL Therapy: Key - Infusion of Preβ-HDL A-I

Acute HDL Infusion Therapy 2 Step Process for Production of Recombined HDL (rHDL) Phospholipids Synthesized ApoA-I 1 243 aa rHDL A-I Clinical Trials rHDL

HDL Infusions in ACS Patients I. ApoA-I Milano

ApoA-I Milano Clinical Trial: Evaluation of Coronary Atherosclerosis by IVUS Infusion of ApoA-I Milano Resulted in Regression of Atherosclerosis in Patients with the Acute Coronary Syndrome Subjects: 46 Acute Coronary Patients Infusions: 5 Weekly Infusions 33 ApoA-I Milano Infusion (15 mg/kg, n=12; 45 mg/kg, n=24; 11 Saline Infusions) Results: Regression – 14.1% Decrease in Total Atheroma Volume Nissen et al JAMA 2003:290,2292-2300. Nissen et al JAMA 2003:290,2292-2300.

Reduction in Atheroma Volume in Acute Coronary Syndrome Patients Following HDL Infusions 46 Acute Coronary Patients: 5 Weekly Infusions 33 ApoA-I Milano Infusion; 11 Saline Infusions 2 4 6 8 10 14 16 - + A-I Milano* -14.10 - - Change in Total Atheroma Volume (mm)3 - 12 -5.34 - *Nissen S et al JAMA 2003;290:2292-2300.

ApoA-I Milano acquired by Pfizer Medicines Company licensed ApoA-I Milano and is currently developing MDC0-216 in Phase I Trial

HDL Infusions in ACS Patients I. ApoA-I Milano II. CSL

Acute HDL Infusion Therapy:CSL111 2 Step Process for Production of CSL111 (ApoA-I/PL,rHDL) using Cholate Detergent Phospholipids Isolated ApoA-I from plasma 1 243 aa Cholate Detergent Clinical Trials CSL111 CSL111 (αHDL) A-I Cholate

Gel Electrophoresis of CSL111 (rHDL) Patel, S. et al. J Am Coll Cardiol 2009;53:962-971

Reduction in Atheroma Volume in Acute Coronary Syndrome Patients Following HDL Infusions 183 Acute Coronary Patients: 4 Weekly Infusions, 123 Patients Received CSL111 rHDL Infusion; 60 Patients Control Infusions 2 4 6 8 10 14 16 - + A-I Milano* -14.10 -5.34 ERASE** **Tardif JC et al. JAMA 2007;297:1675-82 - - Change in Total Atheroma Volume (mm)3 - 12 -5.34 - *Nissen S et al JAMA 2003;290:2292-2300.

CSL has reformulated CSL111 and are currently developing CSL112

Phase I CSL112 (αHDL3) Infusions in Humans 4 8 12 16 20 24 Hours Cholesterol (mg/dl) Free Cholesterol Cholesteryl esters Diditchenko S et al Presented at ISA 2012 Australia

HDL Infusions in the ACS Patient I. ApoA-I Milano II. CSL III. Cerenis

Acute HDL Infusion Therapy:CER-001 ApoA-I/PL/ Sphingomyelin 2 Step Process for Production of CER-001 (ApoA-I,PL/Sphingomyelin) (αHDL) Synthesized ApoA-I 1 243 aa Phospholipids + Sphingomyelin A-I Clinical Trials Cerenis-001 ApoA-I/PL/ Sphingomyelin (αHDL)

Acute HDL Infusion Therapy Effect of Cerenis CER-001 Infusion on Plasma Lipoproteins 4 8 12 16 20 24 Hours Cholesterol (mg/dl) Free Cholesterol Cholesteryl esters

CHI SQUARE Clinical Trail IVUS Analysis of the Effect of Cerenis CER-001 on Coronary Atherosclerosis Control Arm 6 Infusions ? mg/kg n =500+ 18

HDL Infusions in ACS Patients I. ApoA-I Milano II. CSL III. Cerenis IV. HDL Therapeutics

Synthesis of Ligands for the ABCA1 Transporter 2 Step Process for Production of rHDL 1 Step Process for Synthesis of Preβ-HDL αHDL Limitations of 2 Step Procedure 1. Low Yield 2. Safety 4. Cost 3. Preβ-HDL binds to the the ABCA1 Transporter, αHDL does not bind. 5. Regulatory Hurdles 1 243 aa Synthesized ApoA-I Phospholipids Selective HDL Delipidation A-I Preβ-HDL A-I Clinical Trials Selective HDL Delipidation Clinical Trials rHDL rHDL

HDL Selective Delipidation Increase Preβ Levels by Converting αHDL to Preβ-HDL A-I A-I A-I αHDL Preβ-HDL ”Loaded Dump Truck” “Unloaded Dump Truck” 21 21

Preβ Enriched Plasma Obtained by HDL Selective Delipidation Plasma Bag 1 Plasma Collected Preβ Enriched Plasma Obtained By Selective HDL Delipidation A-I αHDL Preβ-HDL 20% 80% Plasma Bag 2 Preβ Enriched Plasma A-I A-I αHDL Preβ-HDL 95% 5% Preβ-HDL

2-D Gel Electrophoresis of Control Plasma, Sham Treated Plasma and Selective HDL Delipidated Plasma Demonstrating a Shift from aHDL to preb-HDL with Selective HDL Delipidation a HDL a HDL a HDL Preb-1 Preb-1 Preb-1 Sham Treated Plasma Selective HDL Delipidation Control Plasma Analysis performed by Dr. B. Asztalos, Tufts University Boston, MA

% Efflux Untreated Control) ABCA1 AND SR-BI MEDIATED EFFLUX OF CONTROL PLASMA, SHAM TREATED PLASMA AND SELECTIVE HDL DELIPIDATED PLASMA (% Efflux Treated/% Efflux Untreated Control) ABCA1 Mediated Efflux SR-BI Mediated Efflux 25.0 Control Serum Sham Treated Selective HDL Delipidated Serum 20.0 15.0 % Efflux Untreated Control) (% Efflux Treated/ 10.0 5.0 1.0 0.0 Analysis performed by Dr. G. Rothblat

Step 1 Collected ~1 Liter of Patients Plasma HDL Therapeutics Human Trial: Conducted Using Autologous Preβ-HDL Enriched Plasma Step 1 Collected ~1 Liter of Patients Plasma Step 2 Cholesterol removed from αHDL to yield preβ-HDL Step 3 Re-infused Preβ Enriched Plasma 25

HDL Selective Delipidation*** Reduction in Atheroma Volume in Acute Coronary Syndrome Patients Following HDL Infusions 28 Acute Coronary Patients: 7 Weekly Infusions, 14 Received Delipidated HDL and 14 Control Plasma Infusions 2 4 6 8 10 14 16 - + HDL Selective Delipidation*** ***Waksman R et al J. Am. Coll Card 2010;55:2727-2735C -12.10 A-I Milano* -14.10 -5.34 ERASE** **Tardif JC et al. JAMA 2007;297:1675-82 - - Change in Total Atheroma Volume (mm)3 - 12 -5.34 - *Nissen S et al JAMA 2003;290:2292-2300.

Acute HDL Infusion Therapy Recombined Particle (rHDL) Stage of Development Recombined Particle (rHDL) ClinicalData Regulatory Pathway Safety ApoA-I Milano MDC-216 Phase I ?OK αHDL ApoA-I Milano IVUS decreases Athero Biologic CSL111/112 Phase I ? OK αHDL CSL-111 IVUS Decreases Athero Biologic Phase II ? OK apoA-I/ PL/SPM ? Cerenis CER-001 Drug Phase III Delipidated HDL Infusions OK Preβ-HDL IVUS Decreases Athero Device

HDL Selective Delipidation** Comparison of Change in Total Atheroma Volume in the HDL Infusion And Statin Saturn Trials 2 4 6 8 10 14 16 - + HDL Infusion therapy - 6.4 - 4.4 Atorvastatin SATURN Trial*** Rosuvastatin 2.2 Years Statin therapy -12.10 HDL Selective Delipidation** 5 - 7 Weeks -14.10 A-I Milano* - - Change in Total Atheroma Volume (mm)3 - 12 -5.34 - *Nissen S et al JAMA 2003;290:2292-2300. **Waksman R et al J. Am. Coll Card 2010;55:2727-2735C ***Nicholls SJ et al, N Eng J Med 2011;365:2078-2087

Take Home Messages HDL infusions in ACS patients were associated with marked regression of atherosclerosis. HDL infusions for 5 - 7 weeks had 2 - 3 greater regression of atherosclerosis than 2.2 years of statin therapy in the Saturn Trial. Clinical trials will be required to definitively establish if HDL infusions in ACS patients will decrease clinical cardiovascular events.