The Science and Medicine of Sepsis Management The Role of Inflammation, Signaling Cascades, and Immune Modulation on the Natural History and Treatment

The Science The Science andand Medicine Medicine ofof

Sepsis ManagementSepsis Management

The Role of Inflammation, Signaling Cascades, and The Role of Inflammation, Signaling Cascades, and Immune Modulation on the Natural History and Immune Modulation on the Natural History and

Treatment of the Sepsis SyndromeTreatment of the Sepsis Syndrome

New Frontiers New Frontiers andand Evolving Perspectives Evolving Perspectives

Dr. Steven Opal, MDDr. Steven Opal, MDProgram Chair and ModeratorProgram Chair and Moderator

Chairman Elect, International Sepsis ForumChairman Elect, International Sepsis ForumProfessor of Medicine at Brown Medical SchoolProfessor of Medicine at Brown Medical School

Director of the Infectious Disease DivisionDirector of the Infectious Disease DivisionMemorial Hospital of Rhode IslandMemorial Hospital of Rhode Island

Pawtucket, Rhode Island, USAPawtucket, Rhode Island, USA

CME-certified symposium CME-certified symposium jointly jointly sponsored by the Postgraduate sponsored by the Postgraduate Institute of Medicine and Institute of Medicine and CMEducation Resources, LLCCMEducation Resources, LLC

Commercial Support: Commercial Support: Sponsored Sponsored by an independent educational by an independent educational grant from Eisai, Inc.grant from Eisai, Inc.

Faculty disclosures: Faculty disclosures: Listed in Listed in program syllabusprogram syllabus

Welcome and Program Overview Welcome and Program Overview Welcome and Program Overview Welcome and Program Overview

Distinguished Program FacultyDistinguished Program Faculty

Program Chair and ModeratorProgram Chair and Moderator

Dr. Steven Opal, MDDr. Steven Opal, MDChairman Elect, International Sepsis ForumChairman Elect, International Sepsis ForumProfessor of Medicine at Brown Medical SchoolProfessor of Medicine at Brown Medical SchoolDirector of the Infectious Disease DivisionDirector of the Infectious Disease DivisionMemorial Hospital of Rhode IslandMemorial Hospital of Rhode IslandPawtucket, Rhode Island, USAPawtucket, Rhode Island, USA

Dr. Jean-Paul Mira, MD, Dr. Jean-Paul Mira, MD, PhDPhD

Professor of Critical Care Professor of Critical Care MedicineMedicine

Chair, Medical Intensive Care Chair, Medical Intensive Care UnitUnit

Head, Variability of Innate Head, Variability of Innate ImmunityImmunity

Research LaboratoryResearch Laboratory Cochin-St. Vincent de Paul Cochin-St. Vincent de Paul

University University Hospital Hospital Paris, FranceParis, France

Dr. Pierre-Francois LaterreDr. Pierre-Francois LaterreProfessor of Critical CareProfessor of Critical CareSt Luc University HospitalSt Luc University HospitalUniversite Catholique de LouvainUniversite Catholique de LouvainBrussels, BelgiumBrussels, Belgium

Evolving Perspectives in Sepsis Evolving Perspectives in Sepsis Research: The Pivotal Role of Immune Research: The Pivotal Role of Immune

Modulation and the Unregulated Modulation and the Unregulated Inflammatory CascadeInflammatory Cascade

Dr. Steven Opal, MDDr. Steven Opal, MDProgram Chair and ModeratorProgram Chair and Moderator

Chairman Elect, International Sepsis ForumChairman Elect, International Sepsis ForumProfessor of Medicine at Brown Medical SchoolProfessor of Medicine at Brown Medical School

Director of the Infectious Disease DivisionDirector of the Infectious Disease DivisionMemorial Hospital of Rhode IslandMemorial Hospital of Rhode Island

Pawtucket, Rhode Island, USAPawtucket, Rhode Island, USA

The Evolving Science The Evolving Science andand Medicine Medicine ofofSepsis ManagementSepsis Management

SIRS = systemic inflammatory response SIRS = systemic inflammatory response syndrome.syndrome.

Infection/TraumaInfection/Trauma SIRSSIRS SepsisSepsis Severe SepsisSevere Sepsis

► A clinical response arisingfrom a nonspecific insult, including ³2 of the following:

► Temperature ≥38oC or ≤36oC► HR ≥90 beats/min► Respirations ≥20/min► WBC count ≥12,000/mm3 or

≤4,000/mm3 or >10% immature neutrophils

► SIRS with a presumed or confirmed infectious process

Bone et al. Bone et al. Chest.Chest. 1992;101:1644 1992;101:1644

Sepsis: Defining a Disease ContinuumSepsis: Defining a Disease Continuum

Bone et al. Bone et al. Chest.Chest. 1992;101:1644; Wheeler and Bernard. 1992;101:1644; Wheeler and Bernard. N Engl J MedN Engl J Med. 1999;340:207. . 1999;340:207.


Sepsis with ≥1 sign of organ failure

– Cardiovascular (refractory hypotension)

– Renal– Respiratory– Hepatic– Hematologic– CNS– Unexplained metabolic acidosis

ShockShock

Sepsis: Defining a Disease ContinuumSepsis: Defining a Disease Continuum

► High and variable mortality rate (20-60%)High and variable mortality rate (20-60%)► Heterogeneous patient populationHeterogeneous patient population► Unpredictable disease progressionUnpredictable disease progression► Unclear etiology and pathogenesisUnclear etiology and pathogenesis► Based on inflammation as cause, regardless of source Based on inflammation as cause, regardless of source

► Knowledge base has progressed since 1992Knowledge base has progressed since 1992


Challenges to Research and StudyChallenges to Research and Study

1.1. Serum lactate measuredSerum lactate measured2.2. Blood cultures obtained prior to antibiotic administrationBlood cultures obtained prior to antibiotic administration3.3. From the time of presentation, broad-spectrum antibiotics From the time of presentation, broad-spectrum antibiotics

administered within 3 hours for ED admissions and 1 hour for administered within 3 hours for ED admissions and 1 hour for non-ED ICU admissionsnon-ED ICU admissions

4.4. In the event of hypotension and/or lactate > 4 mmol/L (36 In the event of hypotension and/or lactate > 4 mmol/L (36 mg/dl):mg/dl):

a)a) Deliver an initial minimum of 20 ml/kg of crystalloid (or colloid Deliver an initial minimum of 20 ml/kg of crystalloid (or colloid equivalent)equivalent)

b)b) Apply vasopressors for hypotension not responding to initial fluid Apply vasopressors for hypotension not responding to initial fluid resuscitation to maintain mean arterial pressure (MAP) > 65 mm Hgresuscitation to maintain mean arterial pressure (MAP) > 65 mm Hg

5.5. In the event of persistent hypotension despite fluid In the event of persistent hypotension despite fluid resuscitation (septic shock) and/or lactate > 4 mmol/L (36 resuscitation (septic shock) and/or lactate > 4 mmol/L (36 mg/dl):mg/dl):

a)a) Achieve ventral venous pressure (CVP) of > 8 mm HgAchieve ventral venous pressure (CVP) of > 8 mm Hgb)b) Achieve central venous oxygen saturation (ScvO2) of > 70%*Achieve central venous oxygen saturation (ScvO2) of > 70%*

* Achieving a mixed venous oxygen saturation (ScvO2) of 65% is an acceptable * Achieving a mixed venous oxygen saturation (ScvO2) of 65% is an acceptable alternativealternativeCopyright 2007 by SCCM, ESICM, and the International Sepsis ForumCopyright 2007 by SCCM, ESICM, and the International Sepsis Forum

Sepsis Resuscitation Bundle (6 Sepsis Resuscitation Bundle (6 Hours)Hours)

1.1. Low-dose steroids administered for septic shock in Low-dose steroids administered for septic shock in accordance with a standardized ICU policyaccordance with a standardized ICU policy

2.2. Recombinant Activated protein C administered in Recombinant Activated protein C administered in accordance with a standardized ICU policyaccordance with a standardized ICU policy

3.3. Glucose control maintained > lower limit of Glucose control maintained > lower limit of normal, but < 150 mg/dl (8.3 mmol/L)normal, but < 150 mg/dl (8.3 mmol/L)

4.4. Inspiratory plateau pressures maintained < 30 cm Inspiratory plateau pressures maintained < 30 cm H2) for mechanically ventilated patientsH2) for mechanically ventilated patients

Copyright 2007 by SCCM, ESICM, and the International Sepsis ForumCopyright 2007 by SCCM, ESICM, and the International Sepsis Forum

Sepsis Management Bundle (24 Sepsis Management Bundle (24 Hours)Hours)

Riedemann NC et al. Nature Medicine 2003;9:517-524

Pathophysiology of SepsisPathophysiology of SepsisPathophysiology of SepsisPathophysiology of Sepsis

The Apoptosis Theory of SepsisThe Apoptosis Theory of Sepsis

► Massive apoptosis of lymphocytes is Massive apoptosis of lymphocytes is seen in lymphoid tissues of animals seen in lymphoid tissues of animals and humans with sepsisand humans with sepsis

► Later phase of immunosuppression Later phase of immunosuppression may in part be due to apoptosismay in part be due to apoptosis

► Secondary nosocomial infection Secondary nosocomial infection and/or viral re-activationand/or viral re-activation

Hotchkiss RS et al. 1999-2009Hotchkiss RS et al. 1999-2009

Rittirsch D et al. 2008Rittirsch D et al. 2008

Schemata for Pathophysiology of SepsisSchemata for Pathophysiology of SepsisSchemata for Pathophysiology of SepsisSchemata for Pathophysiology of Sepsis

What do we know about the immunopathophysiology of What do we know about the immunopathophysiology of sepsis?sepsis?

► All encompassing single mechanism still All encompassing single mechanism still elusiveelusive

► Host inflammation hypothesized to be the Host inflammation hypothesized to be the cause of the syndrome cause of the syndrome

► Natural experiment with mice suggests that Natural experiment with mice suggests that we may be able to reduce inflammation we may be able to reduce inflammation without compromising host defense, perhaps without compromising host defense, perhaps through manipulation of TLRsthrough manipulation of TLRs

The Science and Medicine of SepsisThe Science and Medicine of SepsisThe Science and Medicine of SepsisThe Science and Medicine of Sepsis

Microorganisms

Immune cells

Host-derived mediators

PRRPRRss

PAMPsPAMPs DAMPsDAMPsHSP

Heparan Sulfate

Hyaluronic acid

FibrinogenBiglycan

Surfactant AHMGB-1

HemeMRP8/14

Caspase-1 & 5ASC

NALP1 & 3Pyrin

ASC

NF-κB

TLRs NOD-LRRs RLHs

Cinel& Opal CCM 2009;37:291Cinel& Opal CCM 2009;37:291

SEPSIS: A Dynamic, Complex Host Response to PAMP/DAMPsSEPSIS: A Dynamic, Complex Host Response to PAMP/DAMPs

Incidence of Severe Sepsis by AgeIncidence of Severe Sepsis by AgeC

ase

sC

ase

s

0

20,000

40,000

60,000

80,000

100,000

120,000

<1<1 11 55 1010 1515 2020 2525 3030 3535 4040 4545 5050 5555 6060 6565 7070 7575 8080 85+85+

Age/YearsAge/Years

0

5

10

15

20

25

30

Number of casesNumber of cases

Incidence rateIncidence rate

Inci

den

ce/1

,00

0 P

opu

latio

nIn

cide

nce

/1,0

00

Po

pula

tion

Angus DC, at al. Crit Care Med 2001; 29:1303 10.Angus DC, at al. Crit Care Med 2001; 29:1303 10.

Hotchkiss and Karl NEJM 2003;348:138

Receptor downregulation (TLR 4, TNFR, HLA-DR)

Soluble (sIL1,6,TNFr) and decoy receptors (IL-1R2)

Receptor antagonists (IL-1ra)

Anti-inflammatory cytokines (IL-4,10,13)

Intracellular inhibitors (SOCS, IkB, Tollip, MyD88s)

Cellular apoptosis of B cells CD4 T cells and FDCs

Sepsis-induced immunosuppression

Sepsis-targeting the host response

Sepsis-targeting microbial mediators-LPS

4 million LPS molecules/cell - 75% of outer membrane

Increasing Levels of EndotoxemiaIncreasing Levels of EndotoxemiaAggravates Severity of IllnessAggravates Severity of Illness

Marshall et al. the Medic trial Marshall et al. the Medic trial J Infect DisJ Infect Dis 2004;190:527 2004;190:527

<0.4<0.4(n=367)(n=367)

0.4-0.60.4-0.6(n=228)(n=228)

>0.6>0.6(n=262)(n=262)

P*P*

ICU LOS (days)ICU LOS (days) 55 55 77 P=0.04P=0.04

APACHE IIAPACHE II 13.313.3 15.315.3 17.617.6 P<0.001P<0.001

% Hospital Death% Hospital Death 1616 2323 2323 P=0.05P=0.05

% ICU Death% ICU Death 1111 1313 1717 P=0.04P=0.04

WBC (% abnormal)WBC (% abnormal) 4242 4848 5656 P<0.001P<0.001

% with Shock% with Shock 1212 2121 2323 P<0.001P<0.001

% Hypoxemic% Hypoxemic 4242 5252 5656 P=0.005P=0.005

*p values compare < 0.4 vs > 0.6*p values compare < 0.4 vs > 0.6

nuclear localization sequence

3’5’ 5’

3’

LPS M

DNA NFB

Early Signaling Events of Innate Immunity are now increasingly understood

Host response-antimicrobial defense programs

LPS M

DNA NFB

MONOCYTE-MACROPHAGE

CytokinesChemokinesNitric oxide

Acute phase proteinsPro-coagulants

LPS

LBP

CD 14

TLR4

MD-2

Signal transduction

TIR

NFkB

tk, mapk

3714 genes (12% of the 3714 genes (12% of the human transcriptome) is human transcriptome) is

altered over 24 hours altered over 24 hours upon exposure to LPS upon exposure to LPS

Lowry et al. Nature 2005;437:1032-7

LPS-mediated gene induction

BS Park BS Park et al.et al. NatureNature 2009; 7830:1-5 2009; 7830:1-5

The Hexameric TLR4–MD-2–LPS The Hexameric TLR4–MD-2–LPS Signalling ComplexSignalling Complex

Lipid A

BS Park BS Park et al.et al. Nature Nature (2009);7830(2009);7830

The main The main trimerization trimerization

interfaceinterface of the TLR4–MD-of the TLR4–MD-2–LPS complex2–LPS complex

BS Park et al. Nature (2009); 7830:1-5

Structural Comparison of LPS with Structural Comparison of LPS with AntagonistsAntagonists

TLR4 binding

site

Lipid A

O-side chain

Inner core Outer core

oligosaccharide

Current Anti-LPS projects for severe infection

• Currently in use-PMX B hemoperfusion columns

• Phase 3 trials-E5564

• Phase 2 trials-recombinant Lactoferrin

• Preclinical/early clinical study: AOAH transgenes, rAlkaline phosphatase, small molecule inhibitors, cationic peptides, heme absorption columns, monoclonal antibodies, receptor fusion constructs

E. coli Lipopolysaccharide

Prophylactic and Salvage Rx with anti-TLR4 antibodies protect mice Prophylactic and Salvage Rx with anti-TLR4 antibodies protect mice from lethal Gram-negative bacterial sepsis (from lethal Gram-negative bacterial sepsis (E. coliE. coli 018 given i.p.) 018 given i.p.)

Roger T et al. PNAS 2009;106:2348-2352

-15 min

109cfu

+13 hr

105cfu

With gentamicin and ceftriaxoneWith gentamicin and ceftriaxone

-15 min

109cfu

+13 hr

109cfu

-15 min

109cfu

+13 hr

105cfu

Packed Crystal Structure of Human Packed Crystal Structure of Human LactoferrinLactoferrin

Suzuki et al J Mol Biol 2003; 331:485Suzuki et al J Mol Biol 2003; 331:485

Japelj et al. J. Biol. Chem. 2005;280:16955

Lactoferrin LF11 Peptide Bound to Lactoferrin LF11 Peptide Bound to LPSLPS

LPS

Cationic region

Phosphoryl Phosphoryl groupgroup

Lipid Lipid A

Lactoferrin

Cationic LPS binding protein-competes for LPS with LBP, CD14

Fe++ chelator, limits oxidant tissue injury

Bacteristatic

Promotes neutrophil binding and activity

Promotes efficient antigen presentation and clearance by GALT

Phase 2 clinical trials in prevention of neonatal sepsis positive, adult study results completed

Phase II Results with Oral Talactoferrin Phase II Results with Oral Talactoferrin in Severe Sepsisin Severe Sepsis

Total (n=190)Total (n=190) No cardiovascular dysfunction No cardiovascular dysfunction (n=69)(n=69)

pp<0.04<0.04

pp=0.06=0.06

Agennix AG Press Release Dec. 1, 2009Agennix AG Press Release Dec. 1, 2009

Approved for use in Japan for many years and available on a limited Approved for use in Japan for many years and available on a limited basis in several European countriesbasis in several European countries

PMX Cartridge for Clinical ApplicationPMX Cartridge for Clinical Application

Cruz, D. N. et al. JAMA 2009;301:2445Cruz, D. N. et al. JAMA 2009;301:2445

Estimation of Survival Rate According to Estimation of Survival Rate According to Treatment GroupTreatment Group

Patients with septic shock secondary to PeritonitisPatients with septic shock secondary to Peritonitis

Polymyxin B hemoperfusion therapyPolymyxin B hemoperfusion therapy

Conventional therapyConventional therapy

Sur

viva

l Pro

port

ion

Sur

viva

l Pro

port

ion

Log-rank Log-rank PP=.03=.03

0 5 10 15 20 25 300 5 10 15 20 25 30

Time, dTime, d

1.001.00

0.750.75

0.500.50

0.250.25

No. at riskNo. at risk Polymyxin B hemoperfusion therapy 34 34 32 30 27 22 18Polymyxin B hemoperfusion therapy 34 34 32 30 27 22 18 Conventional therapy 30 22 19 15 15 12 11Conventional therapy 30 22 19 15 15 12 11

Cruz, D. N. et al. JAMA 2009;301:2445Cruz, D. N. et al. JAMA 2009;301:2445

Physiological End Points by Treatment Physiological End Points by Treatment Group at Baseline and 72 HoursGroup at Baseline and 72 Hours

Severely septic patients with peritonitisSeverely septic patients with peritonitis

Jung et al. PLoS One Jung et al. PLoS One 2009;4(1):e7042009;4(1):e704

Jung et al. PLoS One Jung et al. PLoS One 2009;4(1):e7042009;4(1):e704

ConclusionsConclusions

► The discovery of the TLRs and other pattern recognition The discovery of the TLRs and other pattern recognition receptors of the innate immune system offers new receptors of the innate immune system offers new treatment options to the initiating events in severe sepsistreatment options to the initiating events in severe sepsis

► Targeting microbial mediators and their signaling Targeting microbial mediators and their signaling receptors is a rational and probably safe approach to treat receptors is a rational and probably safe approach to treat sepsissepsis

► The results of current ongoing clinical trials targeting The results of current ongoing clinical trials targeting microbial ligands and their receptors will answer microbial ligands and their receptors will answer longstanding questions about adjuvant therapies to longstanding questions about adjuvant therapies to improve the outcome in sepsisimprove the outcome in sepsis

► We will next hear from two experts in sepsis research:We will next hear from two experts in sepsis research:● Jean-Paul Mira (Toll like receptors)Jean-Paul Mira (Toll like receptors)

● Pierre-Francois Laterre (Clinical development of TLR4 inhibitors) Pierre-Francois Laterre (Clinical development of TLR4 inhibitors)

New Approaches for Treating Sepsis Using New Approaches for Treating Sepsis Using Novel Interventions Against Old TargetsNovel Interventions Against Old Targets

Toll-Like Receptors in SepsisToll-Like Receptors in SepsisEmerging Implications for Critical Care ManagementEmerging Implications for Critical Care Management

Dr. Jean-Paul Mira, MD, PhDDr. Jean-Paul Mira, MD, PhDProfessor of Critical Care MedicineProfessor of Critical Care MedicineChair, Medical Intensive Care UnitChair, Medical Intensive Care Unit

Head, Variability of Innate ImmunityHead, Variability of Innate ImmunityResearch LaboratoryResearch Laboratory

Cochin-St. Vincent de Paul University HospitalCochin-St. Vincent de Paul University HospitalParis, FranceParis, France

The Science The Science andand Medicine Medicine ofofSepsis ManagementSepsis Management

Recognition of PAMPs from Different Recognition of PAMPs from Different Classes of Microbial Pathogens Classes of Microbial Pathogens

Mogensen TH. Clin. Microbiol. Rev.2009; 22 : 240-273 Mogensen TH. Clin. Microbiol. Rev.2009; 22 : 240-273

Kumar H. Biochem Biophys Res Comm. 2009;388:621Kumar H. Biochem Biophys Res Comm. 2009;388:621

Akira S, 2009

EMBO reports 2006;7:775EMBO reports 2006;7:775

TLRs: Receptors of AlarminsTLRs: Receptors of Alarmins

Known Endogenous TLR LigandsKnown Endogenous TLR Ligands

Bianchi M. J Leuk Biol 2007Bianchi M. J Leuk Biol 2007

TLR Recognition of Exogenous TLR Recognition of Exogenous versus versus

Endogenous LigandsEndogenous Ligands

Iwasaki A. Science 2010; 327:291Iwasaki A. Science 2010; 327:291

Canonical model Canonical model of sepsisof sepsis

New model of sepsisNew model of sepsis

Animal Models of SepsisAnimal Models of Sepsis

TLR2 and TLR2 and Streptococcus pneumoniaeStreptococcus pneumoniae MeningitisMeningitis

Echchannaoui H et al. JID 2002;186:798

WT

TLR2 -/-

Tissieres P. Curr Opinion Infect Dis 2009;22:285

Nagai H. Nat Immunol 2002;3:667Nagai H. Nat Immunol 2002;3:667

LPS IP + D-galactosamine IPLPS IP + D-galactosamine IP LPS IP LPS IP

Essential Role of MD2 in LPS Essential Role of MD2 in LPS ResponsivenessResponsiveness

0 10 20 30 40 500 10 20 30 40 50

1010

88

66

44

22

00Sur

iviv

al (

# of

mic

e)S

uriv

ival

(#

of m

ice) MD-2 MD-2 -/--/-

WTWT

HoursHours

88

44

44

22

000 1 2 3 4 5 6 70 1 2 3 4 5 6 7

DaysDaysS

uriv

ival

(#

of m

ice)

Sur

iviv

al (

# of

mic

e)

MD-2 MD-2 -/--/-

WTWT

Nagai H. Nat Immunol 2002;3:667Nagai H. Nat Immunol 2002;3:667

Essential Role of MD2 in Essential Role of MD2 in Gram Negative InfectionGram Negative Infection

WTWT

MD-2 MD-2 -/--/-

DaysDays

Sur

viva

l (#

of m

ice)

Sur

viva

l (#

of m

ice)

0 5 10 150 5 10 15

1010

88

66

44

22

00

Salmonella Peritonitis

Relevance Relevance of Polymorphisms in of Polymorphisms in TLR and TLR TLR and TLR Adapters for SepsisAdapters for Sepsis

Understanding the Role of TLR Signaling in Understanding the Role of TLR Signaling in Susceptibility to Human InfectionsSusceptibility to Human Infections

Clinical Clinical immunodeficiencyimmunodeficiency

Mutated Mutated genegene

Mutated Mutated proteinprotein

Immunological Immunological pathways pathways affectedaffected

Infectious susceptibilityInfectious susceptibility

X-lined recessive X-lined recessive EDA-IDEDA-ID IKBKGIKBKG

IKKIKKƴƴ(NE(NE

MO)MO)

Multiple innate Multiple innate and adaptive and adaptive

pathwayspathways

Pyogenic bacterial Pyogenic bacterial infections infections ((Streptococcus Streptococcus

pneumoniae, pneumoniae, Staphylococcus aureus, Staphylococcus aureus,

Haemophilus influenzaeHaemophilus influenzae) )

Atypical mycobacteria Atypical mycobacteria ((Mycobacterium avium Mycobacterium avium

intracellulareintracellulare))

Autosomal Autosomal dominant EDA-IDdominant EDA-ID NFKBIANFKBIA IIκκBBαα

Multiple innate Multiple innate and adaptive and adaptive

pathwayspathwaysAs aboveAs above

IRAK4 deficiencyIRAK4 deficiency IRAK4IRAK4 IRAK4IRAK4 TLR signalingTLR signaling

Pyogenic bacterial Pyogenic bacterial infections infections ((Streptococcus Streptococcus

pneumoniae, pneumoniae, Staphylococcus aureus)Staphylococcus aureus)

Human monogenic immunodeficiencies affecting Toll-like receptor functionHuman monogenic immunodeficiencies affecting Toll-like receptor function

Pyogenic Bacterial Infections in Humans Pyogenic Bacterial Infections in Humans with IRAK-4 Deficiency with IRAK-4 Deficiency

Pyogenic Bacterial Infections in Pyogenic Bacterial Infections in Humans with IRAK-4 Deficiency Humans with IRAK-4 Deficiency

40

20

IL-1

(pg/

ml)

WT/WT WT/Asp299Gly&

Thr399Ile

TLR4 Polymorphisms and Septic TLR4 Polymorphisms and Septic ShockShock

ControlControl

% T

LR4

mut

ated

pat

ient

s%

TLR

4 m

utat

ed p

atie

nts 30

25

20

15

10

5

0

Gram negativeGram negativeSeptic shockSeptic shock

Lorenz , Arch. Intern. Med. 2002 162:1028Lorenz , Arch. Intern. Med. 2002 162:1028

Bochud PY. N Engl J Med 2008; 359:1766Bochud PY. N Engl J Med 2008; 359:1766

Hypothesis : Hypothesis : polymorphisms in polymorphisms in TLR TLR genes from the donor and genes from the donor and the recipient may influence susceptibility to invasive the recipient may influence susceptibility to invasive aspergillosis in stem cell transplantationaspergillosis in stem cell transplantation

Discovery study: Discovery study: DNA from 336 patients and their unrelated DNA from 336 patients and their unrelated donors donors (1995 – 2003)(1995 – 2003)33 cases of aspergillosis33 cases of aspergillosis

Validation study: Validation study: matched case-control study with recipients of matched case-control study with recipients of donorsdonors103 patients with invasive aspergillosis and 263 recipients without 103 patients with invasive aspergillosis and 263 recipients without aspergillosisaspergillosis. .

Genes: Genes: 20 SNPs in 20 SNPs in TLR2TLR2, , TLR3TLR3, , TLR4TLR4 and and TLR9TLR9

Toll-like Receptor 4 Polymorphisms and Toll-like Receptor 4 Polymorphisms and Aspergillosis in Stem-Cell TransplantationAspergillosis in Stem-Cell Transplantation

Bochud PY. N Engl J Med 2008; 359:1766Bochud PY. N Engl J Med 2008; 359:1766

Toll-like Receptor 4 Polymorphisms and Toll-like Receptor 4 Polymorphisms and Aspergillosis in Stem-Cell TransplantationAspergillosis in Stem-Cell Transplantation

Legionnaire’s Disease and TLR5Legionnaire’s Disease and TLR5

Hawn TR. J Exp Med 2003; 198: 1563 Hawn TR. J Exp Med 2003; 198: 1563

Common TLR5 Stop Codon PolymorphismCommon TLR5 Stop Codon Polymorphism

1174 1775

CC

TT

AG

AG

TLR5 Polymorphisms and Legionnaire’s TLR5 Polymorphisms and Legionnaire’s DiseaseDisease

0

2

4

6

8

10

12

14

16

18

CG TA

Cas

CTL

CTL appariés

1174 1775

CC

TT

AG

AG

P P = 0.02= 0.02

P P = 0.03= 0.03

Cases n=109CTL n=508Paired CTL n=89

Hawn TR. J Exp Med 2003; 198: 1563 Hawn TR. J Exp Med 2003; 198: 1563

SNP Smokers Nonsmokers P

No. BPNo. BP Allele OR (95% CI) P OR (95% CI)

Haplotype 1174-1775Haplotype 1174-1775

CACA 00 100 1 1

CGCG 01 1.33 (0.55, 3.25)01 1.33 (0.55, 3.25) 0.530.53 1.99 (1.04, 3.80)1.99 (1.04, 3.80) 0.040.04

TATA 10 0.76 (0.19, 3.05)10 0.76 (0.19, 3.05) 0.700.70 2.43 (1.00, 5.89)2.43 (1.00, 5.89) 0.0050.005

IRAK-1 Haplotype Increases IRAK-1 Haplotype Increases NF-kB ActivationNF-kB Activation

Arcoli J. Am J Respir Crit Care Med 2006;175:1335Arcoli J. Am J Respir Crit Care Med 2006;175:1335

IRAK-1 gene located on X chromosomeIRAK-1 gene located on X chromosome

2 haplotypes: htSNP = IRAK-1 532L2 haplotypes: htSNP = IRAK-1 532LSS

IRAK-1 HaplotypeIRAK-1 HaplotypeWildtypeWildtype

AU

CA

UC

150

100

50

0VariantVariant

P=0.0009P=0.0009

IRAK-1 Haplotype Increases IRAK-1 Haplotype Increases Morbidity of SepsisMorbidity of Sepsis


155 septic Caucasians patients

OR:2.6 (95% CI, 1.1-7.7)OR:2.6 (95% CI, 1.1-7.7)

WildtypeWildtype

%%

70

60

50

40

30

20

10

0VariantVariant

P=0.047P=0.047

IRAK-1 HaplotypeIRAK-1 Haplotype

ShockShock

Pulmonary InfectionPulmonary Infection

WildtypeWildtype

VF

DV

FD

20

10

0

VariantVariant

P=0.03P=0.03


WildtypeWildtype

VF

DV

FD

20

10

0

VariantVariant

P=0.02P=0.02


IRAK-1 Haplotype Increases IRAK-1 Haplotype Increases Mortality of SepsisMortality of Sepsis


WildtypeWildtype

%%

70

60

50

40

30

20

10

0VariantVariant

P=0.03P=0.03


60-Day Mortality60-Day MortalityPulmonary InfectionPulmonary Infection

WildtypeWildtype

%%

70

60

50

40

30

20

10

0VariantVariant

P=0.05P=0.05


60-Day Mortality60-Day Mortality

CContribution of Toll-like receptor-ontribution of Toll-like receptor-mediated responses to sepsis mediated responses to sepsis

pathogenesis pathogenesis

Salamao R. Crit Care Med 2009; 37:132Salamao R. Crit Care Med 2009; 37:132

TLR signaling pathway: 84 genes expression analyzedTLR signaling pathway: 84 genes expression analyzed

Mononuclear cells: down regulation in septic shock patients

Neutrophils: up-regulation throughout the stages of sepsis

Tissieres P. Curr Opinion Infect Dis 2009;22:285 Tissieres P. Curr Opinion Infect Dis 2009;22:285

Jia HP. Am J Physiol Lung Cell Mol Physiol 2004;287:L428Jia HP. Am J Physiol Lung Cell Mol Physiol 2004;287:L428

Endotoxin Responsiveness of Human Airway Endotoxin Responsiveness of Human Airway Epithelia is Limited by Low Expression of MD-Epithelia is Limited by Low Expression of MD-

22

Pugin J. Blood 2004; 104:4071Pugin J. Blood 2004; 104:4071

Soluble MD-2 Activity in Plasma from Patients Soluble MD-2 Activity in Plasma from Patients with Severe Sepsis and Septic Shockwith Severe Sepsis and Septic Shock

Wolfs TG. Mol Immunol 2008;45:3268Wolfs TG. Mol Immunol 2008;45:3268

WB anti-MD2

Healthy Septic

Increased Release of sMD-2 During Human Increased Release of sMD-2 During Human Endotoxemia and Sepsis: A Role for Endothelial CellsEndotoxemia and Sepsis: A Role for Endothelial Cells

Wolfs TG. Mol Immunol 2008;45:3268Wolfs TG. Mol Immunol 2008;45:3268

Increased Release of sMD-2 During Human Increased Release of sMD-2 During Human Endotoxemia and Sepsis: A Role for Endothelial Endotoxemia and Sepsis: A Role for Endothelial

CellsCells

Endotoxemia in human volunteersEndotoxemia in human volunteers

Time (hours)Time (hours)

sMD

-2 (

mg/

ml)

sMD

-2 (

mg/

ml) * *

* *

Lauer S. Cell Immunol 2009;255:8 Lauer S. Cell Immunol 2009;255:8

LPS decreases TLR4 cell expressionLPS decreases TLR4 cell expression

Soluble MD2 Increases TLR4 Levels on Soluble MD2 Increases TLR4 Levels on the Epithelial Cell Surfacethe Epithelial Cell Surface

TLR

4 ce

ll su

rfac

e ex

pres

sion

TLR

4 ce

ll su

rfac

e ex

pres

sion

Mea

n flu

ores

cenc

eM

ean

fluor

esce

nce

TL

R4

/ve

cto

rT

LR

4/v

ect

or

TL

R4

/ve

cto

r(n

oL

PS

)T

LR

4/v

ect

or(

no

LP

S)

0 0.2 0.4 0.6 0.8 1.00 0.2 0.4 0.6 0.8 1.0

LPS (ug/ml)LPS (ug/ml)

1.21.2

11

0.80.8

0.60.6

0.40.4

0.20.2

00

Jia HP. Am J Physiol Lung Cell Mol Physiol 2004;287:L428Jia HP. Am J Physiol Lung Cell Mol Physiol 2004;287:L428

sMD2 may prime epithelial cells for enhanced immunoresponsive function

Endotoxin Responsiveness of Human Airway Endotoxin Responsiveness of Human Airway Epithelia is Limited by Low Expression of MD-Epithelia is Limited by Low Expression of MD-

22


► Cell response to pathogen stimulation is a complex Cell response to pathogen stimulation is a complex phenomenon! phenomenon!

► During sepsis, both exogenous and endogenous During sepsis, both exogenous and endogenous ligands stimulate TLRsligands stimulate TLRs

► Effects of TLRs stimulation are cell-dependent Effects of TLRs stimulation are cell-dependent

► Consequences of TLRs stimulation are host-Consequences of TLRs stimulation are host-dependent dependent

► TLRs coreceptors and signaling molecules play a TLRs coreceptors and signaling molecules play a pivotal role in the regulation of the inflammatory pivotal role in the regulation of the inflammatory responseresponse

► Toll Like Receptors and TLR adaptors: attractive Toll Like Receptors and TLR adaptors: attractive drug targetsdrug targets

Clinical Trials with Clinical Trials with TLR inhibition in SepsisTLR inhibition in Sepsis

The Journey from the Bench to the The Journey from the Bench to the BedsideBedside

Emerging Perspectives inEmerging Perspectives in Sepsis ManagementSepsis Management

Dr. Pierre-Francois LaterreDr. Pierre-Francois LaterreProfessor of Critical CareProfessor of Critical CareSt Luc University HospitalSt Luc University Hospital

Universite Catholique de LouvainUniversite Catholique de LouvainBrussels, BelgiumBrussels, Belgium

TLR4 LigandsTLR4 Ligands

► Pathogen-associated pattern Pathogen-associated pattern moleculesmolecules● LPSLPS● Mannan Mannan

► Danger-associated pattern Danger-associated pattern moleculesmolecules● HMGB1HMGB1● Heat shock proteinsHeat shock proteins● HyaluronanHyaluronan● BiglycansBiglycans● FibronectinFibronectin

Clinical Trials of Anti-TLR4 AgentsClinical Trials of Anti-TLR4 Agents

► Agents targeting LPSAgents targeting LPS● Antibodies against LPSAntibodies against LPS● Polymyxin BPolymyxin B● Bactericidal/Permeability-Increasing ProteinBactericidal/Permeability-Increasing Protein

► Agents targeting TLR4 or the TLR Agents targeting TLR4 or the TLR SignalsomeSignalsome● TAK-242TAK-242● EritoranEritoran

Lipid A

O-side chain

Inner core Outer core

Oligosaccharide

Eschericia coli Lipopolysaccharide

Clinical Trials of Antibodies against LPS Clinical Trials of Antibodies against LPS Core Region EpitopesCore Region Epitopes

Ann Intern Med 1994;120:771-78

P1 Proof-of-Concept Trial of Eritoran in Normal P1 Proof-of-Concept Trial of Eritoran in Normal Human Volunteers Challenged with LPSHuman Volunteers Challenged with LPS

► 100 and 250 mg doses 100 and 250 mg doses of eritoran completely of eritoran completely blocked all clinical signs blocked all clinical signs and symptoms of LPS and symptoms of LPS toxicity toxicity

● ChillsChills● FeverFever● HeadacheHeadache● MyalgiaMyalgia● TachycardiaTachycardia

► 100 and 250 mg doses 100 and 250 mg doses of eritoran completely of eritoran completely blocked all biochemical blocked all biochemical effects of LPS challenge effects of LPS challenge

Lynn M, et al. J Infect Dis. 2003 Feb 15;187(4):631-9Lynn M, et al. J Infect Dis. 2003 Feb 15;187(4):631-9

PlaceboPlacebo

60, 100, 260 ug E668460, 100, 260 ug E6684

Time after LPS infusion (hrs)Time after LPS infusion (hrs)

Pla

sma

TN

F

Pla

sma

TN

F αα

(pg

/mL)

(pg

/mL)

0 2 4 6 8 100 2 4 6 8 10

400400

350350

300300

250250

200200

150150

100100

5050

00

Phase 2 Study of EritoranPhase 2 Study of Eritoran

► Multicentric, Multicentric, randomized, randomized, double-blind trialdouble-blind trial

Tidswell et al. Tidswell et al. Crit Care Med Crit Care Med 2010; 38:72-83 2010; 38:72-83

Baseline Characteristics: P2 EritoranBaseline Characteristics: P2 Eritoran


Characteristics of Infection: P2 EritoranCharacteristics of Infection: P2 Eritoran


CharacteristicCharacteristic PlaceboPlacebo(n=96)(n=96)

Eritoran Eritoran tetrasodium tetrasodium 45mg (n=103)45mg (n=103)

Eritoran Eritoran tetrasodium tetrasodium 105 mg (n=94)105 mg (n=94)

Overall Overall p Valuep Value

Primary focus of infection, n (%)Primary focus of infection, n (%) .5054.5054

PulmonaryPulmonary 38 (39.6)38 (39.6) 39 (37.9)39 (37.9) 29 (31.2)29 (31.2)

Intra-abdominal/gynecologicIntra-abdominal/gynecologic 20 (20.8)20 (20.8) 14 (13.6)14 (13.6) 15 (16.1)15 (16.1)

Urinary tractUrinary tract 13 (13.5)13 (13.5) 13 (12.6)13 (12.6) 19 (19.4)19 (19.4)

Skin/soft tissueSkin/soft tissue 5 (5.2)5 (5.2) 7 (6.8)7 (6.8) 7 (7.5)7 (7.5)

Indwelling catheterIndwelling catheter 2 92.1)2 92.1) 8 (7.8)8 (7.8) 5 (5.4)5 (5.4)

UnknownUnknown 3 (3.1)3 (3.1) 8 (7.8)8 (7.8) 9 (9.7)9 (9.7)

OtherOther 7 (7.2)7 (7.2) 4 (3.9)4 (3.9) 5 (5.4)5 (5.4)

No evidence of infectionNo evidence of infection 8 (8.3)8 (8.3) 10 (9.7)10 (9.7) 5 (5.4)5 (5.4)

Characteristics of Infection: P2 EritoranCharacteristics of Infection: P2 Eritoran(continued)(continued)


CharacteristicCharacteristic PlaceboPlacebo(n=96)(n=96)

Eritoran Eritoran tetrasodium tetrasodium 45mg (n=103)45mg (n=103)

Eritoran Eritoran tetrasodium tetrasodium 105 mg (n=94)105 mg (n=94)

Overall Overall p Valuep Value

Infection type, m (%)Infection type, m (%) .6944.6944

Gram-negativeGram-negative 26 (27.1)26 (27.1) 23 (22.3)23 (22.3) 29 (31.2)29 (31.2)

Gram-positiveGram-positive 30 (31.3)30 (31.3) 38 (36.9)38 (36.9) 29 (31.2)29 (31.2)

Mixed bacterialMixed bacterial 10 (10.4)10 (10.4) 7 (6.8)7 (6.8) 13 (14.0)13 (14.0)

FungalFungal 1 (1.0)1 (1.0) 4 (3.9)4 (3.9) 1 (1.1)1 (1.1)

ViralViral 1 (1.0)1 (1.0) 2 (1.9)2 (1.9) 0 (0)0 (0)

UnkownUnkown 17 (17.7)17 (17.7) 18 (17.5)18 (17.5) 15 (16.1)15 (16.1)

Bacteremia, without focal Bacteremia, without focal infection, n (%)infection, n (%) 3 (3.1)3 (3.1) 10 (9.7)10 (9.7) 8 (8.5)8 (8.5)

Bacteremia, Bacteremia, with focal infection, n )%)with focal infection, n )%) 25 (26.1)25 (26.1) 29 (28.1)29 (28.1) 26 (27.7)26 (27.7)

Adequate antimicrobial Adequate antimicrobial therapy, n (%)therapy, n (%) 87 (91)87 (91) 91 (88)91 (88) 85 (90)85 (90)

Endotoxin in Critically IllEndotoxin in Critically Ill

J Marshall JID 2004J Marshall JID 2004

Endotoxin in Critically IllEndotoxin in Critically Ill

J Marshall JID 2004J Marshall JID 2004

EA LevelEA Level

Gram-negative infectionGram-negative infection Gram-positive infectionGram-positive infection All infectionsAll infections

Prevalence, Prevalence, % (no./total)% (no./total)

OR OR (95% CI)(95% CI)


OR OR (95% CI)(95% CI)


OR OR (95% CI)(95% CI)

Low (<0.40)Low (<0.40) 1.4 (5/367)1.4 (5/367) ---- 3.8 (14/367)3.8 (14/367) ---- 5.2 (19/367)5.2 (19/367) ----

Intermediate Intermediate (0.40-0.60)(0.40-0.60) 4.8 (11/228)4.8 (11/228)

3/7 3/7

(1.3-10.7)(1.3-10.7)7.9 (18/228)7.9 (18/228)

2.2 2.2

(1.1-4.4)(1.1-4.4)11.4 (26/228)11.4 (26/228)

2.4 2.4

(1.3-4.4)(1.3-4.4)

High (>0.60)High (>0.60) 6.9 (18/262)6.9 (18/262)5.3 5.3

(2.0-14.6)(2.0-14.6)5.7 (15/262)5.7 (15/262)

1.5 1.5

(0.7-3.2)(0.7-3.2)10.7 (28/262)10.7 (28/262)

2.2 2.2

(1.2-4.0)(1.2-4.0)

Phase 2 Study of EritoranPhase 2 Study of Eritoran

► Multicentric, randomized, Multicentric, randomized, double-blind trialdouble-blind trial

► Placebo versus two eritoran Placebo versus two eritoran dosing regimens (45 mg/6 dosing regimens (45 mg/6 d or 105 mg/6 d)d or 105 mg/6 d)

► Patients with severe sepsis Patients with severe sepsis and predicted risk of and predicted risk of mortality (PROM) of 20-mortality (PROM) of 20-80% based on APACHE II 80% based on APACHE II score score

► Study drug started within Study drug started within 12 h of recognition of 12 h of recognition of severe sepsissevere sepsis

► 300 patients randomized; 300 patients randomized; 293 included in the intent 293 included in the intent to treat (ITT) analysisto treat (ITT) analysis

Tidswell et al. Tidswell et al. Crit Care Med Crit Care Med 2010; 38:72-83 2010; 38:72-83Treatment GroupTreatment Group

Per

cent

28-

day

Mor

talit

yP

erce

nt 2

8-da

y M

orta

lity

Mortality inMortality inmodified intent-to-treat populationmodified intent-to-treat population

(n=293)(n=293)

pp = 0.846 = 0.846

pp = 0.335 = 0.335

Eritoran P2 Clinical Trial Eritoran P2 Clinical Trial Prospectively Defined Subgroups Prospectively Defined Subgroups


APACHE II QuartileAPACHE II Quartile

Per

cent

28-

day

Mor

talit

yP

erce

nt 2

8-da

y M

orta

lity

Mortality by APACHE II QuartileMortality by APACHE II Quartile

Presence of Shock at BaselinePresence of Shock at Baseline

Mortality by Presence of ShockMortality by Presence of Shock

Per

cent

28-

day

Mor

talit

yP

erce

nt 2

8-da

y M

orta

lity

P=0.598P=0.598P=0.434P=0.434

P-0.105P-0.105

P-0.083P-0.083

P-0.503P-0.503

P-0.913P-0.913

Mortality in Important Subpopulations: P2Mortality in Important Subpopulations: P2


Treatment GroupTreatment Group

Per

cent

28-

day

Mor

talit

yP

erce

nt 2

8-da

y M

orta

lity

Clinically Evaluable PopulationClinically Evaluable Population(n=235)(n=235)

pp = 0.094 = 0.094

N DAA (Xigris) PopulationN DAA (Xigris) Population(n=225)(n=225)

pp = 0.036 = 0.036

Kaplan-Meier Survival-Time Curves: P2Kaplan-Meier Survival-Time Curves: P2


Relative Reduction in Risk of Death Relative Reduction in Risk of Death at 28 Days and 95% CI: P2 Eritoranat 28 Days and 95% CI: P2 Eritoran


MITT populationMITT population

APACHE II predicted mortalityAPACHE II predicted mortality Low (20-50%)Low (20-50%) High (51-80%)High (51-80%)

Type of pathogenType of pathogen Gran negGran neg Gram posGram pos Mixed bacterialMixed bacterial Other/unknownOther/unknown

AgeAge 65 and younger65 and younger 66 and older66 and older

Stage of studyStage of study Stage IIStage II Stage IIIStage III

0.100.10 1.0 1.0 10 10

Relative Reduction in Risk of Death Relative Reduction in Risk of Death at 28 Days and 95% CI: P2 Eritoran at 28 Days and 95% CI: P2 Eritoran (cont.)(cont.)


Xigra UsedXigra Used YesYes NoNo

Baseline EndotoxinBaseline Endotoxin DetectableDetectable Elevated Elevated >> 0.2 endotoxin status 0.2 endotoxin status

Baseline HDLBaseline HDL < 25 mg/dL< 25 mg/dL > 25 mg/dL> 25 mg/dL

Time to drug infusionTime to drug infusion 8 hours or less8 hours or less More than 8 hoursMore than 8 hours

0.100.10 1.0 1.0 10 10

Infectious Adverse Events: P2 EritoranInfectious Adverse Events: P2 Eritoran


Infectious Infectious Adverse EventsAdverse Events

PlaceboPlacebo(n=96)(n=96)

Eritoran Eritoran TertrasodiumTertrasodium

45 mg45 mg(n=103)(n=103)

Eritoran Eritoran Tetrasodium Tetrasodium

105 mg105 mg (n=94)(n=94)

Overall Overall p valuep value

Investigator-Investigator-reported infectious reported infectious complicationcomplication

50.550.5 45.645.6 37.037.0 .2001.2001

Clinical evaluation Clinical evaluation committee committee determination of determination of infectious infectious complicationcomplication

36.836.8 35.035.0 38.038.0 .8967.8967

Infectious adverse events were defined as either a) recurrent infection Infectious adverse events were defined as either a) recurrent infection at the same site as the sepsis-initiating infection, either relapse of the at the same site as the sepsis-initiating infection, either relapse of the same organism, or superinfection by a different organism; or b) new same organism, or superinfection by a different organism; or b) new infection occurring at a different site that the sepsis-initiating infection.infection occurring at a different site that the sepsis-initiating infection.

Eritoran P2 Clinical Trial: Eritoran P2 Clinical Trial: No Effect on Circulating IL-6 No Effect on Circulating IL-6

ConcentrationConcentration


PlaceboPlacebo 45 mg45 mg 105 mg105 mg

0 12 hr 48 hr0 12 hr 48 hr 0 12 hr 48 hr0 12 hr 48 hr 0 12 hr 48 hr0 12 hr 48 hr

IL-6

lvel

(pg

/mL)

IL-6

lvel

(pg

/mL)

100000100000

1000010000

10001000

100100

1010

11

ACCESS Trial: P3 Eritoran TrialACCESS Trial: P3 Eritoran Trial

A Phase 3, Multicenter, Randomized, A Phase 3, Multicenter, Randomized, Double-Blind, Placebo-Controlled Study Double-Blind, Placebo-Controlled Study

Evaluating Eritoran Tetrasodium in Evaluating Eritoran Tetrasodium in Patients with Severe Sepsis: Can Patients with Severe Sepsis: Can

Inhibition of TL-4 Improve All-Cause Inhibition of TL-4 Improve All-Cause Mortality in Patients with Severe SepsisMortality in Patients with Severe Sepsis

• • 159 worldwide study locations159 worldwide study locations• • 2000 patients enrolled in trial2000 patients enrolled in trial

Controlled Comparison of Eritoran Tetrasodium and Controlled Comparison of Eritoran Tetrasodium and Placebo in Patients with Severe SepsisPlacebo in Patients with Severe Sepsis

ACCESS Trial Protocol Overview: Study ObjectivesACCESS Trial Protocol Overview: Study ObjectivesAnalysis of Efficacy VariablesAnalysis of Efficacy Variables

Primary Efficacy EndpointPrimary Efficacy Endpoint: : All-cause mortality at Day 28. The All-cause mortality at Day 28. The difference in difference in mortality rates between treatment mortality rates between treatment groups will be groups will be tested by chi-squared testtested by chi-squared test

Key Secondary Endpoint:Key Secondary Endpoint: Mortality at 1 yearMortality at 1 year

Other Endpoints of Interest:Other Endpoints of Interest:1.1. Length of ICU stay within 28 daysLength of ICU stay within 28 days

2.2. Length of Hospital stay within 28 daysLength of Hospital stay within 28 days

3.3. Duration of dialysis within 28 daysDuration of dialysis within 28 days

4.4. Duration of mechanical ventilation within 28 daysDuration of mechanical ventilation within 28 days

5.5. Duration of use of vasopressors within 28 daysDuration of use of vasopressors within 28 days

6.6. Measurement of cytokine inflammatory panel and procalcitoninMeasurement of cytokine inflammatory panel and procalcitonin

7.7. Quality of LifeQuality of Life

8.8. SOFA assessmentSOFA assessment

9.9. Incidence of infection episodes subsequent to randomizationIncidence of infection episodes subsequent to randomization

10.10. PharmacoeconomicsPharmacoeconomics

11.11. Mortality at 3 and 6 monthsMortality at 3 and 6 months

I. I. Age > 18 years; no upper age limitAge > 18 years; no upper age limit

II. II. Confirmed early onset of severe sepsis defined as:Confirmed early onset of severe sepsis defined as:

• • Objective evidence of infection – likely bacterial or fungal Objective evidence of infection – likely bacterial or fungal pathogenpathogen

Examples of objective evidence:Examples of objective evidence:

Clinical findings (i.e. cellulitis or abscesses)Clinical findings (i.e. cellulitis or abscesses)

CulturesCultures

Gram stainsGram stains

X-raysX-rays

Surgical pathology specimensSurgical pathology specimens

** ** Note: A positive culture is not a requirement for entry into Note: A positive culture is not a requirement for entry into the trialthe trial

ACCESS Trial Entry Criteria ACCESS Trial Entry Criteria Inclusion CriteriaInclusion Criteria


• • Presence of at least 3 of 4 SIRS criteria:Presence of at least 3 of 4 SIRS criteria:

— — Core Temperature Core Temperature >> 38 38C or C or << 36 36C C

— — Heart Rate Heart Rate >> 90 beats/min 90 beats/min

**Note: Patients who cannot be assessed for sepsis-induced **Note: Patients who cannot be assessed for sepsis-induced tachycardia due to another medical condition known to tachycardia due to another medical condition known to increase heart rate, or those receiving treatment that increase heart rate, or those receiving treatment that prevents tachycardia, must have 2 of the remaining 3 SIRS prevents tachycardia, must have 2 of the remaining 3 SIRS criteria.criteria.

— — Respiratory Rate > 20 breaths/min OR a PaCORespiratory Rate > 20 breaths/min OR a PaCO22 < 32 < 32 mmHg, or mechanical ventilationmmHg, or mechanical ventilation

— — WBC Count WBC Count >> 12,000 cells/ 12,000 cells/L, L, << 4,000 cells/ 4,000 cells/L, OR > 10% L, OR > 10% band forms.band forms.

III.III. At least 1 of the following organ dysfunctions:At least 1 of the following organ dysfunctions:

A. Acute Lung Injury (ALI)/Acute Respiratory Distress A. Acute Lung Injury (ALI)/Acute Respiratory Distress

Syndrome (ARDS)Syndrome (ARDS)

— — Acute Onset of the following:Acute Onset of the following:

1. PaO1. PaO22/FiO/FiO2 2 << 300 (<200 in pts. with pneumonia). If 300 (<200 in pts. with pneumonia). If altitude > 1000m, then PaOaltitude > 1000m, then PaO22/FiO/FiO2 2 << 300 x (PB/760) 300 x (PB/760)

2. Bilateral infiltrates consistent with pulmonary edema on 2. Bilateral infiltrates consistent with pulmonary edema on frontal chest x-ray. (Infiltrates may be patchy, diffuse, frontal chest x-ray. (Infiltrates may be patchy, diffuse, homogeneous, or asymmetric)homogeneous, or asymmetric)

3. Requirement for positive pressure ventilation via 3. Requirement for positive pressure ventilation via endotracheal tubeendotracheal tube

4. No clinical evidence of left atrial hypertension4. No clinical evidence of left atrial hypertension

Criteria 1-4 must occur together within 24-hour intervalCriteria 1-4 must occur together within 24-hour interval


B. ThrombocytopeniaB. Thrombocytopenia

Acute onset of platelet count <100,000 or a reduction of Acute onset of platelet count <100,000 or a reduction of 50% or more from prior known levels, without past history 50% or more from prior known levels, without past history of thrombocytopenia, and without attributable cause other of thrombocytopenia, and without attributable cause other than infectionthan infection

C. Lactic AcidemiaC. Lactic Acidemia

Acute onset of serum lactate level > 4mmol/L (36 mg/dL)Acute onset of serum lactate level > 4mmol/L (36 mg/dL)

(Protocol amendment-lactic acidosis >2.2mmol/L (19.8 (Protocol amendment-lactic acidosis >2.2mmol/L (19.8 mg/dL) mg/dL) and and evidence of metabolic acidosis: pH<7.30 or evidence of metabolic acidosis: pH<7.30 or base deficit>5.0 mmol/L)base deficit>5.0 mmol/L)

D. Acute Renal FailureD. Acute Renal Failure

Urine output < 0.5 mL/kg/hr for at least 2 hrs, despite Urine output < 0.5 mL/kg/hr for at least 2 hrs, despite administration of at least 500 mL of crystalloid or 200 administration of at least 500 mL of crystalloid or 200 mL of colloid over a 30 minute periodmL of colloid over a 30 minute period


E. ShockE. Shock

Acute onset of systolic BP < 90 mmHg or MAP of < 65 mmHg. Acute onset of systolic BP < 90 mmHg or MAP of < 65 mmHg. BP is poorly responsive to initial aggressive fluid resuscitation BP is poorly responsive to initial aggressive fluid resuscitation with a crystalloid or colloid, and vasopressors are required to with a crystalloid or colloid, and vasopressors are required to maintain MAP > 65 mmHg despite initial fluid resuscitation for a maintain MAP > 65 mmHg despite initial fluid resuscitation for a least 1 hour.least 1 hour.

Mechanically ventilated patients must exhibit hypotension due Mechanically ventilated patients must exhibit hypotension due to sepsis before the institution of mechanical ventilation or be to sepsis before the institution of mechanical ventilation or be hypotensive for at least 60 min following intubation to qualify hypotensive for at least 60 min following intubation to qualify for the study on basis of shock.for the study on basis of shock.


IV. IV. A reasonable likelihood that administration of A reasonable likelihood that administration of study drug can be started within 12 hours of study drug can be started within 12 hours of the time of recognition of organ dysfunction. the time of recognition of organ dysfunction.

V. V. Must be a commitment to full patient supportMust be a commitment to full patient support

**Note: If a patient’s family has not committed to **Note: If a patient’s family has not committed to aggressive management of patient’s condition or has aggressive management of patient’s condition or has requested the patient be requested the patient be classified as “Do not classified as “Do not resuscitate” or “Do not treat”, the patient is excluded. resuscitate” or “Do not treat”, the patient is excluded. If a family directive allows all resuscitative efforts other If a family directive allows all resuscitative efforts other than chest compressions, the patient may be enrolled.than chest compressions, the patient may be enrolled.


VI. APACHE II ScoreVI. APACHE II Score

Baseline APACHE II Score of 21-37, inclusiveBaseline APACHE II Score of 21-37, inclusive

The Clinical Coordinating Centers will be The Clinical Coordinating Centers will be responsible for calculating the APACHE II responsible for calculating the APACHE II Scores and enrolment approvalScores and enrolment approval

OSCCC (RI) and SLUCCC (Brussels). OSCCC (RI) and SLUCCC (Brussels).


12 hrs12 hrsOnset of Onset of

1st Organ 1st Organ FailureFailure

Study Drug Infusion

Onset of Organ FailureOnset of Organ Failure<< 12 hrs between documentation of the 1 12 hrs between documentation of the 1stst qualifying qualifying organ dysfunction and administration of study drugorgan dysfunction and administration of study drug

ACCESS Trial Entry Criteria: ACCESS Trial Entry Criteria: Time WindowTime Window

ACCESS Trial Protocol Overview ACCESS Trial Protocol Overview Prior & Concomitant TherapyPrior & Concomitant Therapy

In addition to the appropriate antibiotic therapy, it is In addition to the appropriate antibiotic therapy, it is expected that all patients will receive evidence-based expected that all patients will receive evidence-based appropriate treatment of their severe sepsis.appropriate treatment of their severe sepsis.

Appropriate treatment modalities include, but are not Appropriate treatment modalities include, but are not limited to:limited to:

Initial resuscitation goalsInitial resuscitation goals

Use of low tidal volumes for mechanical ventilationUse of low tidal volumes for mechanical ventilation

Control of blood glucose levelsControl of blood glucose levels

Maintenance of target hemoglobin levelsMaintenance of target hemoglobin levels

Source controlSource control

In countries where recombinant human activated Protein C In countries where recombinant human activated Protein C is approved for use, careful consideration should be given is approved for use, careful consideration should be given to contraindications and locally approved indications for to contraindications and locally approved indications for use. use.

► Over 1600 subjects randomizedOver 1600 subjects randomized

► Enrolling ~40-60 subjects/month in recent Enrolling ~40-60 subjects/month in recent monthsmonths

ACCESS Trial: Current StatusACCESS Trial: Current Status

At 1400 SubjectsAt 1400 Subjects

► Mean APACHE II score ~27 in all regions of Mean APACHE II score ~27 in all regions of the world the world

► Mean age= 65Mean age= 65► Median time to treatment = 9.3 hrsMedian time to treatment = 9.3 hrs

50% Shock50% Shock25% Lactic Acidosis25% Lactic Acidosis

Organ Dysfunctions:Organ Dysfunctions: ~33% 1~33% 1 ~33% 2~33% 2

~24% 3~24% 3

InfectionInfection

► Site of Infection?Site of Infection?● ~50% lung~50% lung● ~20% genitourinary~20% genitourinary● ~20% abdominal~20% abdominal

► Incidence of new infection etc. (after 48 Incidence of new infection etc. (after 48 hours)hours)● ~43% …..similar to that reported by ~43% …..similar to that reported by

investigators in the Phase II study investigators in the Phase II study (~47%)(~47%)

Data Monitoring Reviews of StudyData Monitoring Reviews of Study

► DMC reviews at 375 and 750 and 1100 DMC reviews at 375 and 750 and 1100 subjects resulted in recommendations to subjects resulted in recommendations to continue the studycontinue the study

Summary and ConclusionsSummary and Conclusions

► TLR-4 inhibition represents a potentially promising TLR-4 inhibition represents a potentially promising strategy for treatment of severe sepsisstrategy for treatment of severe sepsis

► Phase II Study with eritoran is completed, peer-Phase II Study with eritoran is completed, peer-reviewed, and published in JCCMreviewed, and published in JCCM

► There are signals in the Phase II eritoran study of There are signals in the Phase II eritoran study of improved mortality outcomes in high-risk subgroups; improved mortality outcomes in high-risk subgroups; and safety profiles are acceptableand safety profiles are acceptable

► Phase III ACCESS eritoran study is completing Phase III ACCESS eritoran study is completing enrollmentenrollment

* Three interim analyses have been conducted for the Phase III * Three interim analyses have been conducted for the Phase III

ACCESS study and the DSMB has given authorization to ACCESS study and the DSMB has given authorization to complete the studycomplete the study

Summary and Summary and Take Home MessageTake Home Message

► Despite its complexity and the multitude of Despite its complexity and the multitude of pathogens that can cause sepsis, a limited pathogens that can cause sepsis, a limited number of pattern recognition receptors of number of pattern recognition receptors of the innate immune system activate the the innate immune system activate the systemic host response in sepsis systemic host response in sepsis

► The Toll like receptors are now major The Toll like receptors are now major targets for therapeutic intervention in targets for therapeutic intervention in sepsis.sepsis.

► Polymorphisms of TLRs and related adaptor Polymorphisms of TLRs and related adaptor and signaling molecules are associated with and signaling molecules are associated with susceptibility or protection from a number susceptibility or protection from a number of infectious diseasesof infectious diseases


► TLR4 is the signal receptor for LPS along TLR4 is the signal receptor for LPS along with MD2 and CD 14 with MD2 and CD 14

► A number of TLR 4 inhibitors show promise A number of TLR 4 inhibitors show promise as adjuvant therapies for sepsis; one TLR4 as adjuvant therapies for sepsis; one TLR4 inhibitor, known as eritoran or E5564, is inhibitor, known as eritoran or E5564, is now in late stage clinical development.now in late stage clinical development.

► If TLR inhibitors can improve survival in If TLR inhibitors can improve survival in human sepsis, they will likely be most human sepsis, they will likely be most effective given early in the septic process. effective given early in the septic process. This is a major challenge in clinical trial This is a major challenge in clinical trial design and implementationdesign and implementation


Documents

The Science and Medicine of Sepsis Management The Role of Inflammation, Signaling Cascades, and Immune Modulation on the Natural History and Treatment