modifying atherosclerosis in obesity is an inﬂammatory ... · page 1! modifying atherosclerosis...

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Modifying atherosclerosis in cardiometabolic disease: targeting

inflammation!Peter Libby!

Brigham & Women�s Hospital!Harvard Medical School!

Primary Care Congress for Cardiometabolic Health

Boston April 24,, 2013!

Visc

eral

Adi

pose

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sue

(cm

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st g

irth

(cm

)

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120

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220

1 1

1,3 1,3

90

94

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102

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1 1

1,2 1,2,3

C-Reactive Protein Quintiles

(1) (2) (3) (4) (5) (1) (2) (3) (4) (5)

C-Reactive Protein Quintiles

Lemieux I. et al. Arterioscler Thromb Vasc Biol (2001) 21:961-967

Obesity is An Inflammatory Stimulus!

ATP III Metabolic Syndrome Criteria!

0 1 2 3 4 5 0

2

4

6

8 C

-rea

ctiv

e pr

otei

n (m

g/L)

Number of Components of the Metabolic Syndrome

Inflammation in the Metabolic Syndrome!Women�s Health Study: n=14,719!

Ridk

er C

ircul

atio

n 10

03;1

07-3

91-3

97!

*Adjusted for BMI, family history of diabetes, smoking, physical activity, alcohol consumption, hormone therapy.!Pradhan et al. JAMA. 2001;286:327-334.!

Inflammation Drives Diabetes Elevated Levels of CRP Predict Development of Type 2 Diabetes in the Women�s Health Study!

0

1

2

3

4

5

1 2 3 4 Quartile of C-reactive protein (CRP)

Adj

uste

d R

elat

ive

R

isk

for D

iabe

tes*

P=0.001

!

P. Libby Inflammation in atherosclerosis!Nature 2002;420:868-874.!

Monocyte/Macrophage

Heterogeneity in Atherosclerosis!

Siamon Gordon, J. Clin. Invest. 2007;117:89-93

Mononuclear Phagocyte Heterogeneity in Atherosclerosis!

Hyperlipidemia markedly expands circulating inflammatory monocytes in mice

Swirski, F et al.!J Clin Invest 2007;117:195-205. !

Myeloid cells accumulate in the splenic red pulp in hypercholesterolemic mice!

red pulp

white pulp

wt spleen

red pulp

white pulp

CD11b (myeloid cells)

apoE–/– spleen

Swirski!2011!

!

Hematopoietic stem cell

differentiation in the spleen shapes the evolution of atheromata!

Robbins et al. Circulation

2012;125:364-374

Do new insights into plaque

inflammation have clinical

implications?!

Hypothesis:An echo in the arterial

wall of the inflammation due to acute myocardial

infarction aggravates atherosclerosis!

Clinton SK, et al.!Am. J. Pathol. 1991;138:1005-14.!

Interleukin-1 gene expression in rabbit vascular

tissue in vivo!!Induction of IL-1α and β mRNA in aortic tissue 2 hours after intravenous injection of 10 pg/kg of E. coli endotoxin in rabbits fed control and atherogenic diets for 10 weeks. Three centimeters of thoracic aorta from rabbits provided RNA for Northern analysis for rabbit IL-1α and IL-1β. The data presented represent one rabbit per treatment group. Similar results were observed for three additional rabbits per treatment.!

!

Atherogenic diets enhance endotoxin-stimulated interleukin-1 and tumor necrosis factor gene expression in rabbit aortae!

Fleet JC, Clinton SK, Salomon RN, Loppnow H, Libby P. J Nutr 1992;122:294-305. !

The “echo” phenomenon:!

A systemic inflammatory stimulus, Intravenous endotoxin, evokes a local cytokine response in arteries dependent on the amount of pre-existing atherosclerosis!J Nutr 1992;122:294-305. !!

Infectious agents by systemic and local effects can activate artery wall cells

Libb

y P

et a

l. C

ircul

atio

n 19

97;9

6:40

95-4

103

19 JULY 2012 | VOL 487 | NATURE | 325 !

Molecular Biology of the High-Risk Plaque!

After Libby P. Circulation 1995!

+! +! +! +!

+!+!

–!

Synthesis! Breakdown!

Lipid core!

IL-1"TNF-α"MCP-1"M-CSF!

Fibrous "cap!IFN-γ!

CD-40L!

Collagen-degrading!Proteinases!

Tissue Factor!Procoagulant!

No MI MI

0

15

30

Incr

ease

in p

rote

ase

activ

ity

(pm

ol)

*!

MI

No

MI

Week 0 Week 3

MI enhances protease activity in plaque!

No MI MI

4!

7!

Ex v

ivo

TBR

FR

I

1!

*

Week 3

Dutta et al. Nature 487:325; 2012!

!

Age (in weeks)!

0

3

6

20! 30!Ly-6

Chi

gh m

onoc

ytes

(103 )

/ ao

rta

MI!

No MI!

MI enhances pro-inflammatory monocytes in plaque!

No MI MI

Fibr

ous

cap

thic

knes

s (µ

m)! *

12

0

8

4

0.15

0

0.05 Necr

otic

co

re a

rea !

(mm

2 ) ! *

No M

I!M

I!

Post MI plaque necrotic cores enlarge and fibrous caps thin!

Increased progenitor numbers in spleen after MI!

No MI! MI!

No MI! MI!MDPs

GMPs

GM

Ps

(106 )

/ spl

een

0.0

0.7

1.4 **! ***! **! *!

MD

Ps

(106 )

/ spl

een

0.0

0.6

0.3

**! *! *! *!

c-kit

Line

age+

IL7R!

Sca-1 SS

C-A

c-kit

CD

115

CD34 CD

16/3

2

Week after MI!1 3 6 12


0.0

1.5

3.0 *! *! *!

LSKs

(106 )/

spl

een!


LSKs

Long-term hematopoietic

stem cell!

Short-term hematopoietic

stem cell!

Granulocyte Macrophage progenitor!

Macrophage !dendritic cell!progenitor!

Increased splenic progenitor proliferation in patients after MI !

c-kit DAPI Ki-67 c-kit DAPI Ki-67

c-ki

t+ c

ells

/ 5

HPF

0

1

2 *

No MI (n=29)

MI (n=13)

Human splenic red pulp!

MI enhances atherosclerosis!

Nature. 2012;487:325-9!

!

Innate Immunity in Atherosclerosis!

Initiation!

Complication!

Progression!

Libby, Ridker, MaseriCirculation 2002!

Key Collaborators!

Filip Swirski! Matthias Nahrendorf!

≠!

Can we translate inflammation biology in atherosclerosis to the clinic?!

Can Targeted Anti-Inflammatory Therapy Reduce Cardiovascular Event Rates and Prolong Life?!

Ridker, Thromb Haemost 2009

How to define?

Cardiovascular Inflammation Reduction Trial (CIRT)

What agent

to study?

Stable CAD (post MI) On Statin, ACE/ARB, BB, ASA

Persistent Evidence of Inflammation

Anti-Inflammatory Intervention

Placebo

Nonfatal MI, Nonfatal Stroke, Cardiovascular Death

TC LDL HDL TG Chylo CRP / IL-6

Statins

Issues in the Selection of Anti-inflammatory Agents for Trials of Cardiovascular Inflammation Inhibition

!


Statins TNF inhibition

IL-6 Inhibition




IL-6 Inhibition


Low dose methotrexate

Cardiovascular Inflammation Reduction Trial (CIRT) BWH/NHLBI U01 HL101422 (Ridker) UO1 HL101389 (Glynn, DCC)

A randomized, double-blind, placebo-controlled, event-driven trial of weekly low-dose

methotrexate (LDM) in the prevention of recurrent cardiovascular events among stable post-myocardial infarction patients with type 2

diabetes or metabolic syndrome

BWH/HMS - P. Ridker (PI), R. Glynn (DCC)

NHLBI – A. Hasan, D. Gordon

Methotrexate (MTX) Inhibits Atherogenesis in Cholesterol-Fed Rabbits !

Bulgarelli et al, J Cardiovasc Pharmacol 2012;59:308-14!

MTX ! !Control!

Hematoxylin-eosin!

VSMC!

Macrophages!

MMP-9!

MTX Inhibits Atherogenesis in Cholesterol-Fed Rabbits !

MTX! ! !Control!Bulgarelli et al, J Cardiovasc Pharmacol 2012;59:308-14!

No change!!TC!HDLC !Non-HDLC!TG!

Cohort Group HR* (95 % CI) Endpoint Exposure Wichita RA 0.4 (0.2 - 0.8) Total Mortality LDM Choi 2002 0.3 (0.2 - 0.7) CV Mortality LDM

0.4 (0.3 – 0.8) CV Mortality LDM < 15 mg/wk Netherlands RA 0.3 (0.1 – 0.7) CVD LDM only van Helm 2006 0.2 (0.1 – 0.5) CVD LDM + SSZ

0.2 (0.1 – 1.2) CVD LDM + HCQ 0.2 (0.1 – 0.5) CVD LDM + SSZ + HCQ

Miami VA PsA 0.7 (0.6 – 0.9) CVD LDM Pradanovich 2005 0.5 (0.3 – 0.8) CVD LDM < 15 mg/wk

RA 0.8 (0.7 – 1.0) CVD LDM 0.6 (0.5 – 0.8) CVD LDM < 15 mg/wk

CORRONA RA 0.6 (0.3 – 1.2) CVD LDM Solomon 2008 0.4 (0.2 – 0.8) CVD TNF-inhibitor QUEST-RA RA 0.85 (0.8 – 0.9) CVD LDM Narango 2008 0.82 (0.7 – 0.9) MI LDM

0.89 (0.8 - 1.0) Stroke LDM UK Norfolk RA, PsA 0.6 (0.4 – 1.0) Total Mortality LDM 2008 0.5 (0.3 – 1.1) CV Mortality LDM

LDM and CVD: Observational Evidence

!

� To test directly the inflammatory hypothesis of atherothrombosis by evaluating in a randomized, double-blind, placebo-controlled trial whether LDM given at a target dose of 15 mg po weekly over a three to four year period will reduce rates of recurrent myocardial infarction, stroke, or cardiovascular death among patients with previous myocardial infarction and either type 2 diabetes or metabolic syndrome.!

Cardiovascular Inflammation Reduction Trial (CIRT)!Primary Aims!

N = 7,000 NHLBI-Sponsored!Enrollment to Start March 2013!

350 US and Canadian Sites!

� To determine in a randomized, double-blind, placebo-controlled setting whether LDM will reduce the rate of new onset type 2 diabetes among those with metabolic syndrome but not diabetes at study entry.!

Cardiovascular Inflammation Reduction Trial (CIRT)!Primary Aims!

N = 7,000 NHLBI-Sponsored!Enrollment to Start March 2013!

350 US and Canadian Sites!



IL-6 Inhibition

Issues in the Selection of Anti-inflammatory Agents for!Trials of Cardiovascular Inflammation Inhibition!

LDM IL-1β$inhibition

Human endothelial cells express the interleukin-1β gene inducibly!

Endotoxin and tumor necrosis factor induce interleukin-1 gene expression in adult human vascular endothelial cellsLibby et al. Am J Path 124:179-186 (1986) !

Interleukin-1 Induces Interleukin-1: an auto-amplification loop!

IL-1! IL-1!

Interleukin-1 Induces Interleukin-1!Warner SJC, Auger KR, Libby P. Human interleukin-1 induces interleukin-1 gene expression in human vascular smooth muscle cells. J Exp Med 1987;165:1316-1331.!

Dinarello CA, Ikejima T, Warner SJC, Orencole SF, Lonnemann G, Cannon JG, Libby P. Interleukin-1 induces interleukin-1. I. Induction of circulating interleukin-1 in rabbits in vivo and in human mononuclear cells in vitro. J Immunol 1987;139:1902-1910.!

!

!

Pro-inflammatory actions of IL-1 β$

♥  IL-1 induces IL-6 production, a key mediator of the acute phase response !

Interleukin-1 Induces Interleukin-6:another amplification loop!

IL-1! IL-6!Loppnow H, Libby P. J Clin Invest 1990;85:731-738.!

Proliferating or interleukin-1 activated human vascular smooth muscle cells secrete copious interleukin-6.!Loppnow H, Libby P. J Clin Invest 1990;85:731-738.!

Circulation!

Primary Pro-Inflammatory Cytokines !( e.g., IL-1, TNF-α )! IL-6!

“Messenger” Cytokine!ICAM-1 Selectins, HSPs, etc.

Liver Endothelium and other cells

after Libby, Ridker.!Circulation 1999;100:1148–1150.!

Inflammatory Pathways in Atherogenesis!

Pro-Inflammatory Risk Factors!

CRP!SAA!

�  Fibrinogen!� Plasminogen

activator inhibitor-1!

Interleukin-1: A Signature Cytokine of Innate Immunity!

♥ IL-1 β is synthesized as inactive precursor!

♥ Limited proteolysis by IL-1 β converting enzyme (caspase 1) activates IL-1 β !

Caspase 1 activates IL-1 β !

Caspase 1!(IL-1 β converting enzyme, ICE)!

!

Caspase 1 in Human Atheromata!

Geng%Y'J,%Libby%P.%Evidence%for%apoptosis%in%advanced%human%atheroma:%colocalizaAon%with%interleukin'1β'converAng%enzyme.%Am%J%Pathol%1995;%147(2):%251'266.%

Caspase 1 !(IL-1β

converting enzyme, ICE) in

Human Plaque!

The T-cell-Derived Cytokine CD-40L Stimulates Vascular Cells to Produce Active IL-1 β$

Inactive IL-1β precursor!

Active, mature IL-1β!

©1997 by American Society for Biochemistry and Molecular Biology

IL-1 β processing by extracts of human SMC stimulated by CD40L!

NLRP3 Cryopyrin Inflammasome, Caspase-1, and IL-1β Maturation Endogenous Danger Signals in Vascular Biology?

Drenth JPH, et al, NEJM 2006; 355:730-732

CR-59

NLRP3: a Genetic Determinant of Plasma CRP Level!

Dehgman et al, Circulation 2011;123:731-8

The inflammasome activates caspase-1 and

hence formation of mature IL-1β !

The Inflammasome!

!

Duewell, P, et al, Nature 2010; 464:1357-1362!

Rajamaki K et al, PLoS One 2010;5:e11765!

Geng%YJ,%Phillips%JE,%Mason%RP,%Casscells%SW.%Cholesterol%crystalliza=on%and%macrophage%apoptosis:%implica=on%for%atherosclero=c%plaque%instability%and%rupture.%Biochem%Pharmacol%2003;66(8):1485Q92.%%

Human atheromata

contain cholesterol

crystals!

Copyright ©2010 American Heart Association

Cholesterol Crystals in Atheromata Kim, S. H. et al. Circ Res 2010;106:1332-1341


Cholesterol%crystalliza=on%in%human%THPQ1%lipidQrich%foamy%macrophages%exposed%to%7Qketocholesterol.%Human%THPQ1%macrophages%transformed%by%phorbol%ester%(PMA)%s=mula=on%were%loaded%with%acetylated%LDL%and%then%treated%with%7Qketocholesterol.%Polarized%microscopy%was%performed%to%image%cholesterol%crystals%(arrows).%(a)%Foamy%macrophages%with%cholesterol%crystals%at%37°;%(b%and%c)%7QketocholesterolQtreated%foamy%macrophages%with%crystals;%and%(d)%floa=ng%crystals.%%%

Macrophages engulf cholesterol crystals!



Macrophages engulf cholesterol crystals! IL-1: Potential Roles in Atherogenesis and Methods of Inhibition!

IL-1 type 1!Receptor!

Adapted from Fearon W, Fearon D. Circulation 2008;117:2577-9!

iNOS!!Endothelin-1!!Chemokines/cytokines!!Adhesion molecules!!Endothelial & Smooth !Muscle Proliferation!!Macrophage Activation!!!Endothelial Dysfunction!!Athero-progression!

IL-1α,β$!

IL-1 type II!Decoy Receptor!

Anti IL-1β antibody!Canakinumab!

IL-1 Receptor!Antagonist!

Endogenous!Exogenous !

IL-1r/IL-1r accessory protein!IL-1 trap!

!

Canakinumab (Ilaris, Novartis)!•  high-affinity human monoclonal anti-human

interleukin-1β (IL-1β) antibody currently indicated for the treatment of IL-1β driven inflammatory diseases (Cryopyrin-Associated Period Syndrome [CAPS], Muckle-Wells Syndrome)

•  designed to bind to human IL-1β and functionally neutralize the bioactivity of this pro-inflammatory cytokine

•  long half-life (4-8 weeks) with CRP and IL-6 reduction for up to 3 months

Dose Response of Canakinumab on Plasma IL-6, hsCRP, and fibrinogen: Phase II data

IL-6 hsCRP Fibrinogen

Effects of Interleukin-1β Inhibition With Canakinumab on Hemoglobin A1c, Lipids, C-Reactive Protein, Interleukin-6, and Fibrinogen: A Phase IIb Randomized, Placebo-Controlled Trial. Ridker, Howard, Walter, Everett, Libby, Hensen and Thuren Circulation. 2012;126:2739-2748 !!

CANTOS: a 17,200 person phase III Trial!

Am Heart J 2011;162:597-605 !

•  To test directly the inflammatory hypothesis of atherothrombosis

•  To determine whether long-term inhibition of interleukin-1β with canakinumab (50 mg,150 mg or 300 mg SQ every three months) as compared to placebo will reduce rates of recurrent cardiovascular events among stable post-myocardial infarction patients who remain at elevated vascular risk due to increased levels of hsCRP (> 2 mg/l) despite usual care, including statin therapy.

Canakinumab Anti-inflammatory Thrombosis Outcomes Study (CANTOS) Stable CAD (post MI)

On Statin, ACE/ARB, BB, ASA Persistent Elevation of hsCRP (> 2 mg/L)

Randomized Canakinumab 150 mg

SC q 3 months

Randomized Placebo

SC q 3 months

Primary Endpoint: Nonfatal MI, Nonfatal Stroke, Cardiovascular Death


SC q 3 months

Canakinumab Anti-inflammatory Thrombosis Outcomes Study (CANTOS) (PM Ridker PI)

Secondary Endpoints: Total Mortality, New Onset Diabetes, Other Vascular Events

Exploratory Endpoints: DVT/PE; SVT; hospitalizations for CHF; PCI/CABG; biomarkers


SC q 3 months

N = 17,200!Novartis!

(>5000 currently)!

!

Argentina Australia Austria Belgium Brazil Bulgaria Canada China Colombia Czech Republic

Estonia Germany Greece Guatemala Hungary Iceland India Italy Japan Korea Latvia Lithuania Netherlands Norway Peru Poland Russia Slovakia South Africa Sweden Taiwan Turkey

United Kingdom USA Venezuela

CANTOS Trial Update – 2013! Potential Strengths of IL-1β Antagonism in Atherosclerosis!

♥ Selective: an important pro-inflammatory mediator, but not essential for host defenses!

♥ Leaves IL-1α signaling intact!♥  Implicated in atherogenesis by several

pre-clinical studies!♥ Potential beneficial metabolic effects!

IL-1 beta Antagonism!♥ CANTOS provides an

exciting opportunity to!♥  Test the inflammatory hypothesis

of atherosclerosis!♥  Provide a novel therapy to address

residual risk in secondary prevention!

Inflammation, Atherothrombosis, and Cardiovascular Prevention:

Three Key Translational Questions Do individuals with elevated levels of inflammatory biomarkers have high cardiovascular risk even when other risk factors are acceptable? Yes!Do individuals identified at increased risk due to inflammation benefit from a therapy they otherwise would not have received? JUPITER – Yes!Is there evidence that reducing inflammation per se will reduce vascular events and slow progression of diabetes? CIRT, CANTOS – Let’s find out! P. Ridker!

For More Information : 1-855-437-9330 theCIRT.org theCANTOS.org!

With Appreciation National Heart Lung and Blood Institute National Cancer Institute American Heart Association Doris Duke Charitable Foundation Foundation Leducq Donald W Reynolds Foundation Dade Behring / Seimens Bristol Myers Squibb Amgen / Illumina AstraZeneca Novartis

!

Thanks! Filip Swirski Matthias Nahrendorf Mikael Pittet Elena Aikawa Bill Lucinskas Farouc Jaffer Ralph Wiessleder

Eugenia Schvarz!Galina Sukhova!Eduardo Folco!!Paul M Ridker!Brendan Everett!

D.W. Reynolds Foundation!

modifying atherosclerosis in obesity is an inﬂammatory ... · page 1! modifying atherosclerosis...

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