Acute HDL Therapy Update on HDL Infusion Therapy

Acute HDL Therapy: Update on HDL Infusion Therapy in the ACS Patient H. Bryan Brewer, Jr. MD Washington Cardiovascular Associates Medstar Research Institute Washington, DC 20010

DISCLOSURES H. Bryan Brewer, MD Consulting Fees – Schering-Plough Corp. / Merck & Co. , Inc. , F. Hoffmann-La Roche Ltd. , sanofi aventis US, LLC Honoraria – Schering-Plough Corp. / Merck & Co. , Inc. , Abbott Vascular Ownership Interest (Stocks, Stock Options or Other Ownership Interest) – The Medicines Company, Infra. Re. Dx

NORMAL HDL METABOLISM Intestine Modification Liver LD Lr LDL CETP -I ABCG 1 Apo. A ABC A 1 LCAT -I o. A Ap αHDL Arterial Wall Macrophage A-I Lipid Poor Apo. A-I/Preβ-HDL

HDL THERAPY IN HIGH RISK PATIENTS WITH CARDIOVASCULAR DISEASE I. Acute HDL Infusion Therapy – ACS Patient 1. Apo. A-I Infusions A. Apo. A-I Milano - Pfizer/Medicines Company Slide 4

ACUTE HDL THERAPY – APOA-I INFUSIONS Intestine Modification Liver LD Lr LDL CETP -I ABCG 1 Apo. A ABC A 1 LCAT Acute HDL Infusions -I o. A Ap Apo. A-I Milano; Delipidated HDL; Apo. A-I Mimetic Peptides αHDL Arterial Wall Macrophage A-I A-IA-I Lipid Poor Apo. A-I/Preβ-HDL

Apo. A-I Milano Limone Sul Garde “Preβ-HDL in a Bottle” Cys Apo. A-I Milano 6

INFUSION OF Apo. A-I Milano RESULTED IN REGRESSION OF ATHEROSCLEROSIS IN PATIENTS WITH THE ACUTE CORONARY SYNDROME Subjects: 46 Acute Coronary Patients Infusions: 5 Weekly Infusions 33 Apo. A-I Milano Infusion (15 mg/kg, n=12; 45 mg/kg, n=24; 11 Saline Infusions) Results: Regression – 4. 2% Decrease in Atheroma Volume Nissen et al JAMA 2003: 290, 2292 -2300.

HDL THERAPY IN HIGH RISK PATIENTS WITH CARDIOVASCULAR DISEASE I. Acute HDL Infusion Therapy – ACS Patient 1. Apo. A-I Infusions A. Apo. A-I Milano - Pfizer/Medicines Company B. Selective HDL Delipidation - Lipid Sciences 2. HDL Mimetic Peptides Slide 8

Selective HDL Delipidation Results in an Increase in Plasma Preβ-HDL Liver Selective HDL 1 B SRDelipidation Modification Lr Liver LD Intestine LDL CETP A-I LC αHDL Selective HDL Delipidation Infusions of Selectively Delipidated HDL LCA T AT A-I A-I Preβ-αHDL ABCA 1 Macrophage Slide 9

Method for the Selective Delipidation of HDL in Plasma Slide 10

A-I A-I 95% 5% A-I A-I 80% 20% A-I Slide 11

Human Clinical Trial

Schematic of Selective HDL Delipidation HDL Clinical Trial Plasma HDL Delipidation Process • Uses patient’s own HDL • Cholesterol removed from αHDL to yield preβ HDL • Delipidated plasma is reinfused into patient Delipidated Plasma

Selective HDL Delipidation Clinical Trial Patient Selection Acute Coronary Syndrome Patients Clinical Protocol Baseline Initial IVUS 7 Weekly Infusions of Either Selectively Delipidated HDL Autologous Plasma (n=15) or Untreated Autologous Plasma ( n=15). Repeat IVUS Clinical Trial Site P. I. IVUS Lab - Dr. Ron Waksman Dr. Neil Weissman Dr. Peter Fitzgerald (Stanford) Medstar Research Institute Washington Hospital Center

Selective HDL Delipidation Clinical Trial Delipidated Plasma (n=15) Stable dose of Lipid Drugs (N=30) 1: 1 Randomization Control Undelipidated Plasma (n=15) Day 0 IVUS 1 2 3 4 Week 5 6 7 Delipidated or Control Plasma Infusion 8 IVUS

Phase II Results: Comparison of Plaque Reduction with Selective Delipidation vs Apo. A-I Milano Selective HDL Apo. A-I Milano Delipidation Trial** n=14 N=36 Variable (mean ± SD) Change in Total Atheroma Volume (mm 3) -12. 18 ± 36. 75 -14. 10 ± 39. 50 Change in Percent Atheroma Volume - Plaque Burden (%) -1. 0% ± 4. 0% -1. 1% ± 3. 2% Change in Most Diseased 10 mm Segment Atheroma Volume (mm 3) -6. 24 ± 17. 94 -7. 20 ± 12. 60 *Waksman et al JACC in press **Nissen et al JAMA 2003: 290, 2292 -2300. Slide 16

Infusion of Selective Delipidated HDL (7 weeks ) Had Greater Reduction in Plaque than 80 mg Lipitor Therapy (18 months) Selective HDL Delipidation Trial* n=14 Reversal Trial** N=253 Change in Total Atheroma Volume (mm 3) -12. 18 ± 36. 75 -0. 4 ± 31. 8 Change in Percent Atheroma Volume - Plaque Burden (%) -1. 0% ± 4. 0% -0. 6% ± 5. 1% Change in Most Diseased 10 mm Segment Atheroma Volume (mm 3) -6. 24 ± 17. 94 -4. 2 ± 12. 8 Variable (mean ± SD) *Waksman et al JACC in press *Nissen et al JAMA. 2004; 291: 1071 -1080 Slide 17

HDL THERAPY IN HIGH RISK PATIENTS WITH CARDIOVASCULAR DISEASE I. Acute HDL Infusion Therapy – ACS Patient 1. Apo. A-I Infusions A. Apo. A-I Milano - Pfizer/Medicines Company B. Selective HDL Delipidation - Lipid Sciences C. Apo. A-I - CSL 2. HDL Mimetic Peptides Slide 18

Apo. A-I Mimetic Peptide A-I Lipid Poor Apo. A-I 1 Apo. A-I 243 Synthesis Apo. A-I Mimetic Peptide Slide 19

Apo. A-I Mimetic Peptides Bruin – D 4 -F 18 Amino Acid Amphipathic Helical Peptide Designed to be an Anti-inflammatory Peptide Decreases aortic atherosclerosis in LDLr-KO Slide 20

Apo. A-I Mimetic Peptides Bruin – D 4 -F Pfizer - Esperion ETC 642 22 Amino Acid Amphipathic Helical Peptide Designed to Increase the Activation of the LCAT Enzyme and Promote Reverse Cholesterol Transport. Slide 21

Apo. A-I Mimetic Peptides Bruin – D 4 -F Pfizer - Esperion ETC 642 NIH – 5 A 37 Amino Acid Amphipathic Helical Peptide Designed to Selectively Increase Cholesterol Efflux From the ABCA 1 Transporter Slide 22

LSI 518 P Apo. A-I Mimetic Peptide Ideal Apo. A-I Mimetic Peptide Increase Plasma Total and HDL Cholesterol Designed to Selectively Increase Cholesterol Efflux from the ABCA 1 Transporter Decrease Cellular Adhesion Molecules and Inflammation Decrease Atherosclerosis in the Apo. E Knockout Mouse Model. Slide 23

Effect of IV Injection of LSI 518 P on Plasma Total and HDL Cholesterol In Control Mice HDL Cholesterol % Change Total Cholesterol (mg/dl) % Change HDL Cholesterol (mg/dl) Total Cholesterol Time After Injection (min. ) Slide 24

LSI 518 P Apo. A-I Mimetic Peptide Ideal Apo. A-I Mimetic Peptide √ Increase Plasma Total and HDL Cholesterol Designed to Selectively Increase Cholesterol Efflux from the ABCA 1 Transporter Decrease Cellular Adhesion Molecules and Inflammation Decrease Atherosclerosis in the Apo. E Knockout Mouse Model. Slide 25

Effect of Apo. A-I and LSI 518 P on Cholesterol Efflux From ABCA 1 Positive and ABCA 1 Negative Expressing Cells ABCA 1 Negative Cells Percent Cholesterol Efflux (4 Hours) ABCA 1 Positive Cells Apo. A-I 20 ug/ml LSI 518 P 20 ug/ml George Rothblat University of Pennsylvania Apo. A-I 20 ug/ml LSI 518 P 20 ug/ml Slide 26

LSI 518 P Apo. A-I Mimetic Peptide Ideal Apo. A-I Mimetic Peptide √ Increase Plasma Total and HDL Cholesterol √ Designed to Selectively Increase Cholesterol Efflux from the ABCA 1 Transporter Decrease Cellular Adhesion Molecules and Inflammation Decrease Atherosclerosis in the Apo. E Knockout Mouse Model. Slide 27

CD 11 b Expression (Fluorescence, % of control) Effect of Apo. A-I and LSI 518 P on the Level of Expression of CD 11 b on Human Monocytes 70% Percent Decrease of Control Value Unstimulated PMA 1 u. M + Apo. A-I 20 ug/ml PMA 1 u. M + LSI 520 P 20 ug/ml Slide 28

LSI 518 P Apo. A-I Mimetic Peptide Ideal Apo. A-I Mimetic Peptide √ √ √ Increase Plasma Total and HDL Cholesterol Designed to Selectively Increase Cholesterol Efflux from the ABCA 1 Transporter Decrease Cellular Adhesion Molecules and Inflammation Decrease Atherosclerosis in the Apo. E Knockout Mouse Model. Slide 29

Comparison of the Atherosclerosis in Control Apo. E Knockout Mice and Apo. E Knockout Mice Injected with LSI Peptide 518 P p =. 01 Lesion area mm 3 32% n = 19 Peptide Control Slide 30

LSI 518 P Apo. A-I Mimetic Peptide √ √ Ideal Apo. A-I Mimetic Peptide Increase Plasma Total and HDL Cholesterol Designed to Selectively Increase Cholesterol Efflux from the ABCA 1 Transporter Decrease Cellular Adhesion Molecules and Inflammation Decrease Atherosclerosis in the Apo. E Knockout Mouse Model. Slide 31

HDL THERAPY IN HIGH RISK PATIENTS WITH CARDIOVASCULAR DISEASE I. Acute HDL Infusion Therapy – ACS Patient 1. Apo. A-I Infusions A. Apo. A-I Milano - Pfizer/Medicines Company B. Selective HDL Delipidation - Lipid Sciences C. Apo. A-I - CSL 2. HDL Mimetic Peptides A. B. C. D. D 4 -F Peptide 642 Peptide 5 A Peptide 518 P - Novartis - Pfizer - NIH - Lipid Sciences Slide 32

FUTURE THERAPY FOR CARDIOVASCULAR DISEASE Combination of Statin and HDL Therapy for the Chronic Protection Against Cardiovascular Disease Ischemic Stroke Myocardial Infarction Transient Ischemic Attack Vulnerable Plaque Peripheral Arterial Disease: