SHAPE Scientific Symposium, Dallas, TX 2013

Concept of a Vaccine for Vascular DiseaseModulating Autoimmunity in Atherosclerosis

Using a Vaccine

Kuang-Yuh Chyu, MD, PhDAssociate Director, CICU

Cedars Sinai Heart InstituteCedars-Sinai Medical Center

Disclosures: Co-Inventor of ApoB-100 Peptide Vaccines for Atherosclerosis. Patent rights are assigned to Cedars-Sinai Medical Center.

Disclosures: Co-Inventor of ApoB-100 Peptide Vaccines for Atherosclerosis. Patent rights are assigned to Cedars-Sinai Medical Center.

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Atherosclerosis is an Auto-Immune Mediated Inflammatory Disease

Pro-atherogenic Adaptive Immunity

Th1

Th 17

Inflammatory Cytokines

IFN- γ and TNF-α

Inflammatory Cytokine

IL 17

Atherosclerosis

Activated CD4+ T-cell

Naïve

T cell

APC

Auto

antigen

MHC/CD1NKT

2

Athero-protective Adaptive Immune Response

Antigen

APC

NaiveT cell

Regulatory T cellCD25+, Foxp3+

Secrete anti-inflammatory cytokines

IL-10/TGF-β

Suppression ofAutoimmune Response

Suppression ofAutoimmune Response

Treg

TolerogenicDendritic Cell

Athero-protectiveEffect

Th1

3

Oxidized LDL in Human Coronary Plaques

Colocalizing with Macrophages

OxLDL

Macrophages

Auto-antigens in

Atherosclerosis

LDL/Ox-LDLHsp-602GP1

4

Would Immunization with LDL/oxLDL

Worsen or ReduceAtherosclerosis?

5

Immunization of Cholesterol-fed Rabbits with Homologous LDL Reduces Aortic Atherosclerosis Despite Hypercholesterolemia

Ameli, Shah, Nillson et al: ATVB 1996Nilsson, Ameli, Shah et al: JACC 1997

ImmunizedN=9

Ext

ent o

f P

laqu

e (m

m2 )

Cholesterol 1259 mg/dl 1181 mg/dl

ControlN=7

0

0.5

1

1.5

2

2.5

3

3.5

4

4.5

5

P<0.05

6

Immunization of LDL-R null mice with homologous native or MDA-LDL reduces atherosclerosis (Freigang et al ATVB 1998)

Immunization of apo E null mice with homologous MDA-LDL reduces atherosclerosis (George et al : Atherosclerosis 1998)

Timing affects the athero-protective effects of LDL-immunization (Chyu, Shah et al. Atherosclerosis 2004)

Effect of Immunization with Native and Oxidized Homologous LDL On Experimental

Atherosclerosis in Hypercholesterolemic Mice

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Apo B-100 550kD4536 AA

Johs, A. et al. JBC 2006;281:19732-19739

302 Peptides, 20 amino acids long with 5 amino acid overlap simulating the entire amino acid sequence of human Apo B 100 were synthesized.

Using an ELISA with peptides sequences as antigens, antibodies to 101 of these peptide sequences were identified in pooled human sera

Several peptide sequences were then used to create vaccines for Immunization in apo E null mice fed a high cholesterol diet

KTTKQ SFDLS VKAQY KKNKH

Amino acid 3136-3155

p210

Apo B100 Related Peptide (p210) Vaccine Reduces Atherosclerosis in Hyperlipidemic Mice

61%

P210: apo B-100 amino acids 3136-3155 Chyu KY, Shah PK et al: PLoS One 2012Chyu KY, Shah PK et al: PLoS One 2012 9

PBS Adjuvant p210 Control Control Vaccine PBS Adjuvant p210 Control Control Vaccine

Apo B100 Related Peptide (p210) Vaccine Reduces Plaque Macrophages in ApoE-/- Mice

Apo B100 Related Peptide (p210) Vaccine Reduces Plaque Macrophages in ApoE-/- Mice

* p<0.05

Chyu KY, Shah PK et al: PLoS One 2012Chyu KY, Shah PK et al: PLoS One 2012

Apo B100 Related Peptide (p210) Vaccine Reduces Plaque Dendritic Cells in ApoE-/- Mice

Apo B100 Related Peptide (p210) Vaccine Reduces Plaque Dendritic Cells in ApoE-/- Mice

PBS Adjuvant p210 Control Control Vaccine PBS Adjuvant p210 Control Control Vaccine

p < 0.05

Chyu KY, Shah PK et al: PLoS One 2012Chyu KY, Shah PK et al: PLoS One 2012

Adoptive Transfer of CD8 T-cells from p210 Immunized Mice Conveys Athero-protection

to Unimmunized Mice

Chyu KY et al : PLoS One 2012Chyu KY et al : PLoS One 2012

p < 0.05

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Adoptive Transfer of p210 loaded Dendritic Cells reduces Atherosclerosis in Apo E-/- Mice

Adoptive Transfer of p210 loaded Dendritic Cells reduces Atherosclerosis in Apo E-/- Mice

Nasal delivery of p210-CTB Fusion Construct reduces Atherosclerosis in Apo E-/- Mice

Nasal delivery of p210-CTB Fusion Construct reduces Atherosclerosis in Apo E-/- Mice

Prolonged subcutaneous delivery of p210 reduces Atherosclerosis in Apo E-/- Mice

Prolonged subcutaneous delivery of p210 reduces Atherosclerosis in Apo E-/- Mice

Daugherty et al Br J Pharmacol. 2001 October; 134(4): 865–870.

Abdominal aortic aneurysm formation after AngII infusion in apoE-/- mice

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% Total Aneurysmal Segment number of aneurysmal segment/number of total segment (%)

Number of aneurysmal segment (severity of disease) was significantly reduced in p210 immunized mice.

(Aneurysmal segment = diameter>1.35mm)

Honjo T: Circulation (abst) 2011Honjo T: Circulation (abst) 2011

1 2

3

4

5

68

7

p<0.05

p210 Vaccine Reduces Mortality in AngiotensinII Induced Aneurysm Model in ApoE-/- Mice

Survival Curve

p<0.05 vs. PBSp<0.05 vs. cBSA by Logrank test

Mortality

P210 = 6.2%

PBS = 32.1%

cBSA = 29.7%

Honjo T: Circulation (abst) 2011Honjo T: Circulation (abst) 2011 16

p210 Vaccine Reduces Pro-inflammatory IL-17 Positive CD4 T cells

* *

CD4+IL-17A+

N: PBS=8, cBSA=6, p210=6Honjo T et al : unpublished data 2013

p < 0.05

P<0.05

P<0.05

AngII

AngII

Honjo T: Circulation (abst) 2011Honjo T: Circulation (abst) 2011 18

N=6, P<0.05

IB; AT1R

IB; GAPDH

PBS cBSA p210

p210 vaccine reduced the expression of AT1R in whole aorta.

P210 Vaccine Reduces AT1R Expression in Aorta

Honjo T: Circulation (abst) 2011Honjo T: Circulation (abst) 2011 19

Conclusions• Immunization with selected ApoB-100 peptides

reduces atherosclerosis and favorably modifies plaque phenotype in hypercholesterolemic mice.

• Immunization with selected ApoB-100 peptides also reduced AngII induced hypertension and aneurysm formation and mortality due to rupture.

• These experimental observations provide proof of concept regarding the feasibility and efficacy of apoB-100 peptide based vaccine for vascular diseases.

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Lund University, Sweden

Oppenheimer Atherosclerosis Research Center

P.K.Shah, MDKuang-Yuh Chyu, MD, PhD

Paul Dimayuga,PhDTomoyuki Honjo, MD,PhD

Xiaoning Zhao,PhDJianchang Zhou, PhD

Nicole LioBojan Cercek, MD, PhD

Cheryl ChanJuliana Yano

Jan Nilsson, MD, PhDGN Fredriksson,PhD

CardiovaxFundingFunding

NIH-NHLBIEisner Foundation

CardiovaxThe HeartFoundationSpielberg FoundationCorday Foundation

NIH-NHLBIEisner Foundation

CardiovaxThe HeartFoundationSpielberg FoundationCorday Foundation

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p210

APC

CD8+T cell CD8+ Regulatory T cellCD25+, IL-10+

Suppressionof

Antigen Specificadverse

Autoimmune Response

Suppressionof

Antigen Specificadverse

Autoimmune Response

Increased Lysis of APC

Potential Mechanisms of Action of p210 Vaccine

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Incidence

46.7%

Survival

0%

IL-17 Deletion Protects Mice from AngiotensinII Induced Aortic Aneurysm Formation

Honjo T et al : unpublished data 2013

Abdominal Aortic Aneurysm

Pathophysiology

Inflammation

Immune activation

Oxidative stress

Matrix proteolysis

SMC death