Disseminated intravascular coagulation (DIC)

1
Disseminated intravascular coagulation(DIC)

• Jørn Dalsgaard Nielsen
• Thrombosis Centre
• Gentofte Hospital
• Denmark

2
Characteristics of DIC
DIC
Arterial thrombosis
Venous thrombosis
3
Introduction

• Thrombotic microangiopathy (TMA) and disseminated
  intravascular coagulation (DIC) are disorders
  causing obstruction of the microvascular
  circulation

4
Thrombotic microangiopathy
TTP Thrombotic Thrombocytopenic
Purpura HELLP Haemolysis, Elevated Liver
enzymes, Low Platelets HUS Haemolytic Uremic
Syndrome
5
Thrombotic microangiopathy
Type Symp. Etiology TTP CNS vW cleaving factor
deficiency, cong/aquired HELLP Liver Pregnant
women HUS Kidney /- diarrhoea associated (E
coli 0157)
6
Thrombotic microangiopathy

• Treatment TTP HELLP HUS
• Eliminate the causal factor
• Plasmapheresis (or FFP or cryosupernatant
  transfusion)
• Corticosteroids () ()
• IV gamma globulin ()
  ()
• Rituximab ()
• Avoid platelet transfusions
  

7
DIC is a complication
Banal Serious illness
Critical illness
8
(No Transcript)
9
Causes of DIC
Other causes
Sepsis
10
The Vicious Cycle of Inflammation and Coagulation
Septic shock
Sepsis
11
The Vicious Cycle of Inflammation and Coagulation
Infection
Inflammation
Organ Failure
Coagulation
Ischemia
Death
Endothelial Dysfunction
Inflammation
Coagulation
Inflammation
Inflammation
Coagulation
Esmon. Immunologist. 1998684.
12
Progression of SEPSIS
Platelets
Monocytes
Endothelial cells
Endothelial cells
Non-adhesive surface
Adhesive surface
Leuko- cytes
Activation of coagulation ? Thrombin ?
Fibrin
13
THE CLASSIC COAGULATION SYSTEM
APTT
Prothrombin time
VIIa VII
Ca
Phospholipid, Ca, VIII
X Xa X
Phospholipid, Ca, V
14
EXPRESSION OF TISSUE FACTOR
CONSTITUTIVEe.g.epithelial cellsglial cells
INDUCEDe.g.monocytic cellsendothelial cells
PROHIBITEDe.g.lymphocyteserythrocytes
15
SUBENDOTHELIAL TISSUE
ENDOTHELIAL CELLS
NORMAL HAEMOSTASIS
Haemostatic mechanisms
Intravascular clot formation
Activated monocyte
Monocyte
Activation of monocytes
SYSTEMIC INFLAMMATION
16
Causes of DIC (mechanisms)

• Extensive release of tissue factor
• Increased formation of tissue factor
• Abnormal activators of coagulation
• Contact activation ? hypotension

17
Causes of DIC (Clinical conditions, I)

• Infections
• Sepsis
• Gram negative (endotoxin)
• Gram positive (polysaccharides, peptides)
• Viremias
• Varicella
• Hepatitis
• Cytomegalovirus
• HIV

18
Causes of DIC (Clinical conditions, II)

• Trauma
• Crush injuries
• Other trauma with tissue necrosis
• Severe burns
• Extensive surgery
• Obstetric complications
• Amniotic fluid embolism
• Placental abruption
• (Pre)eclampsia
• Dead fetus syndrome

19
Causes of DIC (Clinical conditions, III)

• Hemolysis
• Hemolytic transfusion reactions
• Massive transfusions
• Malaria
• Other severe hemolysis
• Malignant disorders
• Metastatic malignancy
• Tumors producing cancer procoagulant
• Tumor with tissue necrosis

20
Causes of DIC (Clinical conditions, IV)

• Vascular abnormalities
• Giant hemangioma
• Heriditary teleangiectasis
• Prosthetic devices
• Aortic balloon assist devices
• Denver shunts
• Other conditions
• Pancreatitis
• Acute liver necrosis
• Transplant rejection
• Heat stroke

21
Course of DIC- Accelerating factors -

• Shock
• Acidosis
• Hypoxaemia
• Stasis
• Dehydration

• Fever
• Stress
• Renal insufficiency
• Liver disease
• Malnutrition

• Deficient antithrombotic mechamisms
  Deficient antiplatelet mechanisms Deficient
  anticoagulant mechanisms Decreased
  fibrinolytic activity Dysfunction of the
  reticulo-endotelial system

22
Trauma-induced DIC Phases
Activation phase
No DIC-symptomsShort APTT and PTHigh
fibrinolytic activity
ClinicalParaclinical
Early consumption phase
Recovery
Death
ClinicalParaclinical
MicrothrombosisThrombocytopenia, highD-dimer
and soluble fibrin
Regression of clinical and paraclinical changes
Late consumption phase
Microthrombosis and bleeding Severe
thrombocytopeniaProlonged APTT and PT
ClinicalParaclinical
23
Progression of DIC
Time
Onset of DIC
24
Progression of DIC
Systemic fibrin formation
Abrupt onset of DIC
Time
Onset of DIC
25
Examples of hyperfibrinolytic DIC

• DIC in women with post-partum bleeding
• DIC in patients with promyelocytic leukaemia
• Early after severe trauma
• Contact with Lonomia caterpillars

26
Fibrinolytic activity in patients with DIC
Abrupt onset of DIC
Plasminogen
Plasmin
Fibrin
Fibrin degradation products
27
Hyper- and non-hyperfibrinolytic DIC

• Hyperfibrinolytic DIC
• Main problem Severe bleeding
• Non-hyperfibrinolytic DIC
• Main problem Microvascular occlusion

• DIC in septic patients is a
• non-hyperfibrinolytic type of DIC

28
TF and LPS induced DIC

• DIC was induced in rats by infusion of TF or LPS
• TF LPS p
• Platelets 109/l 204 177 ns
• Fibrinogen mg/dl lt50 lt50 ns
• TAT ng/ml 162 170 ns
• D-dimer ?g/ml 12,4 1,2 0,001
• PAI U/ml 22 245 0,001
• Glomerular fibrin 12 73 0,001

Asakura et al. Crit Care Med 2002 30 161-4
29
Clinical manifestations of DIC
30
Symptoms of DIC

• Dysfunction of multiple organs
• The pulmonary microembolism syndrome
• Acute vascular and bronchial constriction
• Late ARDS
• Acute renal failure
• Oliguria, increasing serum creatinine, haematuria
• Cerebral dysfunction
• Confusion, blurred consciousness, coma
• Cutane haemorrhagic necroses
• Failure of liver, endocrine glands etc.

31
Diagnostic criteria of DIC
32
Local haemostatic response to an injury
Local immunological response to an injury
Modification (Amplification)
SIRS
DIC
Ischaemia
Destruction
MODS
33
SIRSDIC hyperproteolysis
Coagulation
Fibrinolysis
Complement
Kinines
Cytokines
34
BLOOD TESTS WHEN DIC IS SUSPECTED
Simple screening
Extended screening
Supplementary tests
Further evidence for activation of
coagulation and fibrinolysis
35
Activation of coagulation
D fragmentsE fragments
Fibrinogen
Prothrombin
Fragment 12
THROMBIN
Fibrino-peptideA B
Antithrombin
Fibrin
Plasmin
FXIII FXIIIa
Thrombin-Antithrombincomplex(TAT)
Cross-linkedfibrin
D dimerE fragments
36
Soluble fibrin monomer as predictor for DIC in
neonatal sepsis

• Healthy neonates 24,5 6,09 mg/l
• Sepsis, no DIC 33,7 11,9 mg/l
• Sepsis DIC 73,2 31,6 mg/l

ISTH DIC score ?5
Critical level 48,5 mg/l Sensitivity
100 Specificity 93 Overall accuracy 97,5
Selim et al. Haematologica 200590419-20
37
BLOOD TESTS WHEN DIC IS SUSPECTED
Simple screening
Extended screening
Supplementary tests
Prothrombin fragment 1.2
Thrombin-antithrombin complexes (TAT)
Fibrinopeptid A (FPA) or soluble fibrin
Protein C
Fibrinolytic activity (Clot lysis time)
Thrombin time
Plasmin-antiplasmin complexes (PAP)
38
Activation of coagulation
D fragmentsE fragments
Fibrinogen
Prothrombin
Fragment 12
THROMBIN
Fibrino-peptideA B
Antithrombin
Fibrin
Plasmin
Anti-plasmin
FXIII FXIIIa
Thrombin-Antithrombincomplex(TAT)
PAP complex
Cross-linkedfibrin
D dimerE fragments
39
Procalcitonin (PCT) is a marker for activation of
mononuclear cells Oberhoffer et al. J Lab Clin
Med 199913449-55

• Oberhoffer et al. Clin Chem Lab Med 199937363-8
• In septic shock PCT is a better prognostic
  marker than IL-6, TNF and CRP
• Schroder et al. Langenbecks Arch Surg
  199938433-8
• PCT might be a better marker than the classic
  criteria of inflammation, CRP, leukocyte count,
  and body temperature to identify patients
  endangered by severe infection or sepsis.

40
PCT and sepsis
41
Septic ARDS
Nonseptic ARDS
Crit Care Med 1999272172-6
42
SCHISTOCYTES
Longitudinally cut blood vessel
43
Definition of disseminated intravascular
coagulation
DIC is an acquired syndrome characterized by the
intravascular activation of coagulation with loss
of localization arising from different causes. It
can originate from and cause damage to the
microvasculature, which if sufficiently severe,
can produce organ dysfunction
ISTHs Scientific Subcommittee on DIC, July 2001
44
Considerations in practical diagnostic approach
to DIC

• Presence of an underlying disorder
• The severity of haemostatic changes
• Decompensated haemostatic system Overt DIC
• Compensated haemostatic system Non-overt DIC
• The duration of activation
• Temporary e.g. Abruptio placentae, transfusion
  reaction
• Prolonged e.g. Sepsis, malignancy, polytrauma
• Laboratory tests
• Global tests / Molecular markers
• Diagnostic value / Prognostic value
• Use of scoring systems
• DIC scoring system
• Other scoring systems

ISTHs Scientific Subcommittee on DIC, July 2001
45
Scoring system for overt DIC

• Underlying disorder known to be associated with
  overt DIC

YES
NO
continue
stop

• Platelet count
• (gt1000, lt1001, lt502) ..........................
  ....
• Soluble fibrin/D-dimer
• (normal0, ?2, ??3) ............................
  .
• Prolongation of PT
• (lt3s0, 3-6s1, gt6s2) ...........................
  .....
• Fibrinogen
• (gt1g/l0, lt1g/l1) ...............................
  ...........
• Calculate sum ....................................
  ....

ISTHs Scientific Subcommittee on DIC, July 2001
46
Scoring system for overt DIC
- Example -

• Underlying disorder known to be associated with
  overt DIC

YES
NO
Polytrauma
continue
stop

• Platelet count
• (gt1000, lt1001, lt502) ..........................
  ....
• Soluble fibrin/D-dimer
• (normal0, ?2, ??3) ............................
  .
• Prolongation of PT
• (lt3s0, 3-6s1, gt6s2) ...........................
  .....
• Fibrinogen
• (gt1g/l0, lt1g/l1) ...............................
  ...........
• Calculate sum ....................................
  ....

85 1
8 3
3 1
2,2 0
5
ISTHs Scientific Subcommittee on DIC, July 2001
47
Scoring system for overt DIC

• If the calculated score is
• ?5 compatible with overt
  DIC repeat scoring daily
• lt5 suggestive (not affirmative) for non-overt
  DIC repeat next 1-2 days.

ISTHs Scientific Subcommittee on DIC, July 2001
48
Scoring system for non-overt DIC

• Presence of underlying disorder
• (no0, yes2) ....................................
  ......................
• Platelet count changes
• (?1000, lt1001) (?-1, stable0, ?1) .......
• Sol.fibrin/D-dimer changes
• (normal0, ?1) (?-1, stable0, ?1) ........
• Prolongation of PT changes
• (?3s0, gt3s1) (?-1, stable0, ?1)
  ...........
• Antithrombin
• (normal-1, low1) ...............................
  ..................
• Protein C
• (normal-1, low1) ...............................
  ..................
• TAT complexes
• (normal-1, high1) ..............................
  ...................
• Calculate sum ....................................
  ............

ISTHs Scientific Subcommittee on DIC, July 2001
49
Validation of the ISTH scoring system for overt
DIC
Distribution of DIC scores in 217 patients in
intensive care unit
Bakhtiari et al. Crit Care Med 2004 322416 2421
50
Validation of the ISTH scoring system for overt
DIC
Bakhtiari et al. Crit Care Med 2004 322416 2421
51
Validation of the ISTH scoring system for
non-overt DIC
Survivors
Deaths
Non-overt DIC scores in 490 patients in intensive
care unit
Toh Downey. Blood Coagul Fibrinolysis
2005166974
52
Validation of the ISTH scoring system for
non-overt DIC
Toh Downey. Blood Coagul Fibrinolysis
2005166974
53
Validation of the ISTH scoring system for
non-overt DIC

• The mortality rate for non-overt DIC was
• 29 (105 of 360) for scores below 5
• 78 (70 of 90) for scores of 5 or above
• The mortality rate for overt DIC was also 78 (38
  of 49).
• The non-overt DIC scoring template is workable
  and has prognostic relevance.
• A score of 5 and greater is recommended as
  diagnostic of non-overt DIC.

Toh Downey. Blood Coagul Fibrinolysis
2005166974
54
Scoring system for overt DIC

• Underlying disorder known to be associated with
  overt DIC

YES
NO
continue
stop

• Platelet count
• (gt1000, lt1001, lt502) ..........................
  ....
• Soluble fibrin/D-dimer
• (normal0, ?2, ??3) ............................
  .
• Prolongation of PT
• (lt3s0, 3-6s1, gt6s2) ...........................
  .....
• Fibrinogen
• (gt1g/l0, lt1g/l1) ...............................
  ...........
• Calculate sum ....................................
  ....

ISTHs Scientific Subcommittee on DIC, July 2001
55
D-dimer- low cutoff -

• A low cutoff of 1 mg/l was used by
• Dempfle et al. Thromb Haemost 2004 91 8128
• Toh et al. Blood Coagul Fibrinolysis 2005 16
  69-74
• Angstwurm et al. Crit Care Med 2006 34 31420

56
Interassay variation of D-dimer
D-dimer was determined in 39 plasma samples with
23 D-dimer assays
Dempfle et al. Thromb Haemost 2001 85 6718
57
D-dimer tests giving comparable results
Suggested cutoffs Low 1 mg/l High 4 mg/l
BioMérieux Vidas D-dimer
BioMérieux MDA D-dimer
58
Cut-off levels of fibrin related markers
59
Cut-off levels of D-dimer
60
The ATryn Study
Clinical Study Protocol Exploratory efficacy and
safety, pharmacokinetics and dosefinding study of
recombinant human antithrombin in patients with
disseminated intravascular coagulation associated
with severe sepsis
61
Calculation of overt DIC score
62
Calculation of non-overt DIC score
lt
63
Treatment of DIC
64
Treatment of DICdirected against etiological
factors

• Infections
• Trauma
• Obstetriccomplications

Antibiotics
Removal of damaged tissuestabilisation of
fractures
Evacuation of the uterus
65
Treatment of DICdirected against pathogenetic
factors

• Toxic O- and OH-radicals
• Complement activation
• Cytokine formation
• Thrombus formation

66
SUBENDOTHELIAL TISSUE
ENDOTHELIAL CELLS
NORMAL HAEMOSTASIS
Haemostatic mechanisms
Intravascular clot formation
Activated monocyte
Monocyte
Activation of monocytes
SYSTEMIC INFLAMMATION
67
THE COAGULATION SYSTEM
FVII FVIIa
TF
INITIATION PHASE
68
THE COAGULATION SYSTEM
FVII FVIIa
FIX
TF
FXIa FXI
INITIATION PHASE
PROPAGATION PHASE
FIXa
FVIIIa FVIII
FX FXa
FVa FV
Thrombin
Prothrombin
Fibrin Fibrinogen
Cross-linked fibrin
FXIIIa FXIII
STABILIZATION PHASE
69
THE COAGULATION SYSTEM
NATURAL ANTICOAGULANTS TFPI AT PC/PS/TM
INITIATION PHASE
PROPAGATION PHASE
FXa
Thrombin
Prothrombin
Fibrin Fibrinogen
Cross-linked fibrin
FXIIIa FXIII
STABILIZATION PHASE
70
THE CENTRAL ROLE OF THROMBIN
FVII FVIIa
TF
INITIATION PHASE
FX FXa
Thrombin
Prothrombin
71
T
hrombin-signaling in sepsis
Endothelial cell
Binding of thrombin to Protease Activated
Receptor 1 (PAR-1) causes nuclear translocation
of Nuclear Factor Kappa B (NF?B)
TF PAI-1 PAF ICAM IL-1
NF?B
NF?B
PAR-1
T
PAR-1
NF?B is a ubiquitous transcription factor that
governs the expression of genes encoding
cytokines, chemokines, growth factors, cell
adhesion molecules, and a number of acute phase
proteins in various disease states. Tak
Firestein. J Clin Invest 2001 107 7-11
72
The way to microvascular suicide
73
T
hrombin-signaling
Endothelial cell
PC
EPCR
A
TM
A
From normal haemostasis to sepsis
PC
EPCR
A
TM
A

NF?B
PAR-1
A
TM
PC
A
EPCR
A
TM
PC
A
EPCR
74
T
hrombin-signaling in sepsis
Endothelial cell
PC
A
EPCR
T
TM
EPCR
A
T
Cytokines Adhesion molecules
NF?B
NF?B
PAR-1
PAR-1
T
A
T
EPCR
A
TM
PC
A
EPCR
75
T
hrombin-signaling in sepsis
Endothelial cell
Treatment with heparin
PC
A
EPCR
TM
T
A
EPCR
A
A
Cytokines Adhesion molecules
T
NF?B
NF?B
PAR-1
PAR-1
T
T
A
A
T
EPCR
Heparin
A
TM
Elastase
PC
A
EPCR
Pulletz et al. Crit Care Med 2000 28 2881-6.
Jordan et al. Am J Med 1989 87 Suppl 3B 1989
76
T
hrombin-signaling in sepsis
Endothelial cell
Treatment with heparin and antithrombin
PC
A
EPCR
TM
T
A
EPCR
A
T
A
A
Cytokines Adhesion molecules
T
T
A
NF?B
NF?B
PAR-1
PAR-1
T
T
A
A
A
A
T
EPCR
T
A
Heparin
A
TM
Elastase
PC
A
EPCR
77
T
hrombin-signaling in sepsis
Endothelial cell
Treatment with activated protein C /- heparin
PC
A
EPCR
TM
T
APC
A
EPCR
A
T
A
A
T
T
A
NF?B
PAR-1
PAR-1
T
T
A
A
A
A
T
APC
EPCR
T
A
Heparin
A
TM
Elastase
PC
A
EPCR
78
Role of heparin in sepsis
Heparin treatment after cecum ligation and
puncture increases mortality in rats (13 ? 75,
p lt 0.015).
Heparin treatment after cecum ligation and
puncture
CLPPBS
Echtenacher et al. Infect Immun 2001 69 3550-5
79
The role of heparin in sepsis
A DIC model 22 sheeps had intravenous infusion
of E. coli endotoxin
Schiffer et al. Crit Care Med 2002 302689 2699
80
(No Transcript)
81
The role of heparin in sepsis
A meta-analysis of animal studies
Cornet et al. Thromb Haemost 2007 98 579586
82
The role of heparin in sepsis
N2x5
Two studies showing negative effect of heparin
were not included in the meta-analysis Corrigan
et al. J Infect Dis 1975131 139143.
Echtenacher et al. Infect Immun 2001 69 3550-5
Cornet et al. Thromb Haemost 2007 98 579586
83
Role of heparin in sepsis

• Heparin seems to have
• a beneficial effect in animal models where DIC is
  induced by endotoxin probably due to the
  anti-inflammatory effect of binding and
  neutralisation of TNF
• a harmful effect when given to septic animals
  containing living bacteria.

84
Sepsis-induced capillary leak syndrome
Severe sepsis induces systemic capillary leak
85
Decreased plasma activity of AT and PC in septic
patients with DIC

• Asakura et al. Eur J Haematol 2001 67 170-5
• Among 139 septic patients the 68 patients with
  DIC had significantly higher levels of TAT and
  lower levels of AT and PC than the 71 patients
  without DIC (plt0.001 for all variables) but no
  significant correlation was observed between
  plasma levels of TAT and AT or between plasma
  levels of TAT and PC.

• When the patients were classified into three
  groups according to the albumin level, no
  significant differences in AT activity or PC
  activity were observed between the patients with
  and without DIC.

86
Decreased plasma activity of AT and PC in septic
patients with DIC

• Asakura et al. Eur J Haematol 2001 67 170-5

The results suggest that the reduced activity of
AT and PC is not due to consumption coagulopathy
but rather to capillary leak, degradation by
elastase, and/or reduced synthesis.
87
Dual-chamber system to analyze endothelial cell
layer permeability
Evans blue-labelled bovine serum albumin
Upper 500 µl chamber for addition of test material
Endothelial cell layer
Polycarbonate membrane of 3µm pore size
Lower 1500 µl chamber for collection of media to
be analyzed for Evans blue labelled bovine serum
albumin (OD at 650 nm)
Feistritzer Riewald. Blood 2005 105 3178-84
88
Endothelial barrier protection by activated
protein C through PAR1-dependentsphingosine
1phosphate receptor-1 crossactivation
PAR1-dependent signaling by the interdependent
procoagulant and anticoagulant proteases thrombin
and APC can have opposite effects on endothelial
barrier integrity. Barrier protection by APC or
low concentrations of thrombin is mediated by
sphingosine kinase-1 activity and crossactivation
of S1P1 signaling.
Feistritzer et al. Blood. 20051053178-84
89
APC reduces the mortality of LPS-induced
endotoxemia in mice
APC
Saline
Kerschen et al. J Exp Med. 20072042439-48
90
APC reduces the mortality of LPS-induced
endotoxemia in mice
- No effect in EPCR ?/? mice -
EPCR ?/? mice
APC
Saline
Kerschen et al. J Exp Med. 20072042439-48
91
APC reduces the mortality of LPS-induced
endotoxemia in mice
- Reduced effect in PAR1 -/- mice -
PAR1 -/- mice
APC
Saline
Kerschen et al. J Exp Med. 20072042439-48
92
APC reduces the mortality of LPS-induced
endotoxemia in mice
- and so does 5A-APC -
5A-APC A recombinant APC variant with normal
signaling but lt10 anticoagulant activity
5A-APC ?2, ?10 ?g
Saline
Kerschen et al. J Exp Med. 20072042439-48
93
Sepsis and DIC

• Septic patients may develope DIC as a result of
  activation of Toll-like receptors (TLRs) on
  monocytes, neutrophils and endothelial cells by
  microbial products like lipopolysaccharides,
  peptidoglycan, and lipoteichoic acid.

Zhang Ghosh. J Clin Invest 2001 107
13-19 Henneke Golenbock. Crit Care Med 2002 30
Suppl 207-13
94
TLR-mediated activation of NF?B
Zhang Ghosh. J Clin Invest 2001 107 13-19
95
Role of NF?B in sepsis
NF?B in nuclear extracts from peripheral blood
mononuclear cells is comparable to the APACHE-II
score as a predictor of outcome in septic
patients.
Böhrer et al. J Clin Invest 1997 100 972-85
96
APC inhibits activation of NF?B

• Esmon CT. J Autoimmun 2000 15 113-6
• Activated protein C reduces nuclear translocation
  of NF?B resulting in reduction of cytokine
  synthetic rates.

97
Antiinflammatory effects of APC

• APC blocks cytokine elaboration by mononuclear
  cells
• Grey et al, J Immunol 19941533664 68
• Endothelial Protein C Receptor
• Fukudome Esmon, J Biol Chem 199426926486-91
• Mononuclear Phagocyte Protein C Receptor
• Hancock et al, Transplantation 1995601525-32

98
Anti-inflammatory effects of AT

• Binding to endothelium stimulates prostanoid
  release
• Yamauchi et al. BBRC 1989
• Horie et al. Thromb Res 1990
• Increased prostacyclin levels in animal models
• Uchiba et al. Thromb Res 1995
• Uchiba et al. Am J Physiol 1996
• Inhibition of endotoxin-induced cytokine
  synthesis, platelet aggregation, leukocyte
  differentiation and vascular permeability by
  prostacyclin
• Uchiba et al. Thromb Res 1996
• Reduction of ischemia-reperfusion injury
• Ostrovsky et al. Circulation 1997
• Harada et al. Blood 1999

99
Anti-inflammatory effects of AT

• Inhibition of IL-6 and tissue factor release from
  endothelial cells and monocytes
• Souter et al. Crit Care med 2001
• Inhibition of endothelial cell proliferation
• Larson et al. J Biol Chem 2001
• ICAM-1-dependent inhibition of adhesion of
  neutrophils to endothelial cells
• Kaneider et al. Ann Hematol 2003
• Inhibition of NF?B activation in endothelial
  cells and monocytes
• Mansell et al. FEBS Lett 2001
• Oelschlager et al Blood 2002
• Effect on endothelial-leukocyte interactions in
  endotoxemia exerted predominantly via the
  endothelium
• Mizutani et al. Blood 2003

100
AT inhibits LPS-induced activation of NF?B

• Mansell et al. FEBS Letters 2001 508 313-7
• LPS-induced, TLR4-mediated activation of NF?B in
  human mononuclear cells was inhibited
  dose-dependently by AT.
• Modified AT without serpin activity had no
  effect.
• The effect was not increased by addition of
  heparin.

101
AT inhibits LPS-induced TF and IL-6 production by
mononuclear cells, endothelial cells, and whole
blood

• TF and IL-6 were determined after stimulation of
  whole blood, HUVEC, and MNC with LPS for 4-6
  hours.
• Production of TF and IL-6 was reduced in the
  presence of varying concentrations of AT.

Souter et al. Crit Care Med 2001 29 134-9
102
AT prevents LPS-induced pulmonary vascular injury

• Uchiba Okajima. Semin Thromb Haemost 1997
  23583-90
• Intravenous administration of AT (250 U/Kg) to
  rats prevented LPS-induced accumulation of
  leukocytes and increases in pulmonary vascular
  permeability.
• Trp49-modified AT, which lacks affinity for
  heparin, had no effect.
• AT had no effect in animals pretreated with
  indomethacin, suggesting that the protective
  effect was a result of endothelial release of
  prostacyclin.
• Inhibition of thrombin formation by inactive FXa
  did not prevent pulmonary vascular injury

103
Prevention of vascular leakage during sublethal
porcine sepsis by antithrombinDickneite, G
Kroez, M

• Sepsis was induced in 18 pigs by a 3 hours
  infusion of LPS 0,25 ?g/kg/h.
• 90 minutes after start of LPS infusion the
  animals were randomised to
• AT 120 IU/kg iv bolus 5 IU/kg/h iv infusion
• AT 250 IU/kg iv bolus 10 IU/kg/h iv infusion
• Placebo (HSA)

Presented at the 13th Annual Congress of ESICM in
Rome, 1-4 October 2000
104
Prevention of vascular leakage during sublethal
porcine sepsis by antithrombinDickneite, G
Kroez, M
Conclusion

• The broadening and dispersal of interlobular
  connective tissue was decreased significantly and
  dose-dependently by AT.
• Also the periportal edematisation and
  accumulation of leukocytes could be reduced.

Presented at the 13th Annual Congress of ESICM in
Rome, 1-4 October 2000
105
Effect of AT on DIC in man- the first randomised
study -
Blauhut et al. Thromb Res 1985 39 81-9

• 51 patients with DIC were randomised to
  substitution with
• AT infusion, target 100
• Heparin infusion, 3000 IU as bolus 300 IU/h
• AT reduced dose of heparin (1000 IU 100 IU/h)
• Conclusion
• AT reduced duration of DIC
• Concomitant heparin caused
• Acccelerated reduction of platelet count
• Increased consumption of AT
• Increased need of blood transfusion

106
Randomised trials on the use of AT in patients
with sepsis or evidence of DIC
107
Randomised trials on the use of AT in patients
with sepsis or evidence of DIC
Mortality odds ratio
0,37 (0,15-0,88) 0,43 (0,20-0,92) 0,63
(0,28-1,39)
AT vs. Hep (N 133) AT vs. Placebo (N
130) ATHep vs. Hep (N 150)
0,0
1,0
2,0
AT reduces mortality significantly when used
alone but not when accompanied by heparin
treatment
108
Randomised trials on AT replacement in intensive
care management of patients with an antithrombin
activity below 70
Mortality
Odds ratio
Harper (1991)
Albert (1992)
Diaz-Cremadez (1994)
Baudo (1998)
Waydhas (1998)
Total
0,85 (0,51-1,73)
0,0
1,0
2,0
3,0
4,0
109
KyberSept Study

• A phase 3 trial of antithrombin versus placebo in
  patients with severe sepsis or septic shock.
• The trial was powered to detect a 15 reduction
  of an expected placebo 28-day mortality of 45.
• 2339 patients were randomised. In the treatment
  group the mean increase in plasma antithrombin at
  24 hours of inclusion was 130.
• Mortality was lower than expected and almost
  identical in the two groups (Antithrombin 38,9
  and placebo 38,7).

Warren et al. JAMA 2001 286 1869-78
110
KyberSept Study

• A shortcoming of the study was that use of
  unfractionated or LMW heparin ?10.000 IU was
  allowed without randomisation.
• Less than a quarter of the patients in the study
  had no heparin.
• Heparin increased the risk of poor outcome and
  adverse bleeding.

Warren et al. JAMA 2001 286 1869-78
111
KyberSept Study

• 28-day mortality

Warren et al. JAMA 2001 286 1869-78
112
Randomised trials on the use of AT in patients
with sepsis or evidence of DIC
KyberSept results included
Mortality odds ratio
0,37 (0,15-0,88) 0,72 (0,55-0,96) 1,09
(0,89-1,32)
AT vs. Hep (N 133) AT vs. Placebo (N
828) ATHep vs. Hep (N 1766)
0,0
1,0
2,0
AT reduces mortality significantly when used
alone but not when accompanied by heparin
treatment
113
Effects of activated protein C

• inhibits leukocyte-endothelial cell interaction
• inhibits thrombin formation
• increases fibrinolytic activity

Bernard GR et al. N Engl J Med 2001 344 699-709
114
Activation of protein C
Activated protein C
Thrombomodulin
Thrombomodulin
PAI-1 ? TAFI ?
Protein C
Thrombin
EPCR
Endothelial cell
115
Protein C deficiency

• Hereditary PC deficiency was first described by
  Griffin et al. in 1981
• Hereditary PC deficiency occurs in 0,3 of the
  population and in 4-8 of patients with venous
  thromboembolism
• Homozygous PC deficiency causes thromboembolic
  disease and often DIC early in the neonatal
  period
• Aquired PC deficiency is common in patients with
  sepsis and DIC

116
Protein C in septic patients

• The plasma concentration of PC is reduced in
  septic patients due to
• Capillary leak
• Degradation by elastase
• Decreased synthesis
• Increased consumption during DIC
• Activation of PC is compromised in septic
  patients because the endothelial expression of
  thrombomodulin and endothelial protein C receptor
  is suppressed

117
rhAPC Phase 2 Trial

• Placebo-controlled, double-blind, dose-finding
• study
• 131 patients with severe sepsis were
• randomised to
• continuous infusion of 1 of 4 doses of
  recombinant
• human activated protein C for 48-96 hours
• placebo for 48-96 hours.

As presented at the 13th Annual Congress of
European Society of Intensive Care Medicine,
Rome, 1-4 October 2000
118
rhAPC Phase 2 Trial

• The study population was divided into 3 groups
• Placebo
• Low-dose (12 or 18 ?g/kg/hr)
• High dose (24 or 30 ?g/kg/hr)
• A statistically significant reduction in D-dimer
  (p lt0,01)
• and plasma IL-6 (p lt0,05) was observed in the
• high-dose group compared with the placebo group
• A non-statistically significant 15 relative risk
  reduction
• in the 28-day all-cause mortality was observed
• (mortality was not a primary endpoint for this
  study).

As presented at the 13th Annual Congress of
European Society of Intensive Care Medicine,
Rome, 1-4 October 2000
119
Activated protein C in the treatment of DIC

• randomised, double-blind trial of APC versus
  heparin in the treatment of 104 patients with DIC
• Treatment
• plasma-derived APC (300 U/kg/day) or
• heparin (192 IU/kg/day) for 6 days
• Mortality
• APC group 20,4
• Heparin group 40,0

Kenji Okajjima and the APC-IM Clinical Research
Group Presented at the ASH meeting in San
Francisco, USA, Dec. 2000
120
The PROWESS Study

• Protein C Worldwide Evaluation in Sepsis
• Clinical controlled phase III multicenter
  double-
• blind placebo-controlled trial in severe sepsis

Bernard GR et al. N Engl J Med 2001 344 699-709
121
PROWESS Study Description

• Design
• Randomized, double-blind, placebo-controlled
  trial
• 11 countries at 164 sites
• Population - Severe Sepsis
• Presence of a known or suspected infection
• Evidence of a systemic response to the infection
  (3 SIRS criteria)
• At least one sepsis-associated organ dysfunction
  of no greater than 24 hours duration
• Treatment Arms
• 11 randomization to Drotrecogin Alfa (activated)
  24 µg/kg/hr or placebo for 96 hours

Data from the PROWESS Trial
122
PROWESS Study Design
Infection with organ failure
Assessment of 28-day all-cause mortality Alive or
Dead?
Start of study drug infusion
48 Hours Maximum - Consent - Start Drug
Routine Patient Care
End of 96 hour infusion of study drug
Data from the PROWESS Trial
123
Results

• Drotrecogin Alfa (activated) in patients with
  severe sepsis
• Acceptable safety profile
• Significantly reduces mortality
• 6.1 absolute, 19.4 relative risk reduction
• Lower mortality observed across most subgroups
• Number needed to treat to save one additional
  life equals 16

124
Treatment of sepsis and DIC

• Probably the primary goal of DIC treatment should
  not be inhibition of coagulation but protection
  of endothelial cells from being activated to
  produce inflammatory cytokines and adhesion
  molecules.

• Increased leukocyte-endothelial cell interaction
  may be the key mechanism in the development of
  organ dysfunction.
• Neutropenic patients are unlikely to develope
  septic ARDS.

Okajima et al. Am J Hematol 1991 36 265-71
125
Treatment of sepsis and DIC

• Activated protein C (APC) has in addition to its
  anticoagulant effect been shown to inhibit
  leukocyte-endothelial cell interaction
• In the PROWESS trial (N 1690) 28-days mortality
  was 6.1 lower in the APC group than in the
  control group (p lt 0.01)
• AT has EC protective properties when heparin is
  avoided.
• In the non-heparin groups of the KyberSept trial
  (N 698) 28-days mortality was 5.8 lower in the
  AT group than in the control group (n.s.)

Bernard GR et al. N Engl J Med 2001 344
699-709 Warren et al. JAMA 2001 286 1869-78
126
Treatment of sepsis and DIC
AT
aPC

• Natural anticoagulant
• Present on EC surface
• EC protective properties
• Inhibits activation of NF?B
• Reduces effects of LPS
• .

• Natural anticoagulant
• Present on EC surface
• EC protective properties
• Inhibits activation of NF?B
• Reduces effects of LPS
• .
• .

127
Treatment of sepsis and DIC
AT
aPC

• Natural anticoagulant
• Present on EC surface
• EC protective properties
• Inhibits activation of NF?B
• Reduces effects of LPS
• Displaced from EC surface by heparin
• Reduction of mortality in patients with
  sepsis/DIC not proven in phase III trial

• Natural anticoagulant
• Present on EC surface
• EC protective properties
• Inhibits activation of NF?B
• Reduces effects of LPS
• Not displaced from EC surface by heparin
• Reduction of mortality in patients with
  sepsis/DIC proven in phase III trial

128
Treatment of sepsis and DIC
Conclusion
Meta-analysis suggests that antithrombin without
concomitant heparin treatment may reduce
mortality in patients with sepsis and DIC. Until
now, this has, however, not been proven in a
sufficiently powered phase III trial. Further
studies are warranted to define the role of AT
replacement in the treatment of sepsis and DIC.
129
The face of sepsis