Paroxysmal Nocturnal Hemoglobinuria (PNH) - Enerca. Notaro.pdf · Paroxysmal Nocturnal...
Transcript of Paroxysmal Nocturnal Hemoglobinuria (PNH) - Enerca. Notaro.pdf · Paroxysmal Nocturnal...
6th EUROPEAN SYMPOSIUM ON RARE ANAEMIAS 1st Dutch-Belgian meeting for patients and health professionals
21st - 22nd November 2015
Amsterdam - The Netherlands
Paroxysmal Nocturnal Hemoglobinuria (PNH)
Introduction to PNH Rosario Notaro
Cancer Genetics and Gene Transfer
Core Research Laboratory
Istituto Toscano Tumori
Florence, Italy
6th European Symposium on Rare Anaemias - 1st Dutch-Belgian meeting for patients and health professionals
Disclosures Company name Research
support Employee
Consultant Stockholder Speakers bureau Advisory board Other
No conflicts to declare
6th European Symposium on Rare Anaemias - 1st Dutch-Belgian meeting for patients and health professionals
This talk is applicable for:
Definite Probable
Thalassemia’s
Sickle cell disease
Membrane disorders (e.g. sferocytosis)
Enzym defects (e.g. PKD, G6PD)
PNH Yes
Other forms of hemolytic disease Yes
6th European Symposium on Rare Anaemias - 1st Dutch-Belgian meeting for patients and health professionals
Paroxysmal Nocturnal Hemoglobinuria
Acquired disease (it is not transmitted to offspring)
Prevalence: 5-10 cases per million
Geographic distribution: worldwide
Age: any
Gender: same frequency in men and in women
6th European Symposium on Rare Anaemias - 1st Dutch-Belgian meeting for patients and health professionals
Chronic intravascular hemolysis:
with crisis
Thrombosis: Often Multiple, Venous, Abdominal
PNH is a rare clonal acquired blood disorder
characterized by the clinical triads:
Cytopenias (Bone marrow failure): Common, sometimes severe
WBC /µl Plts /µl
Paroxysmal Nocturnal Hemoglobinuria
6th European Symposium on Rare Anaemias - 1st Dutch-Belgian meeting for patients and health professionals
Transmembrane Proteins and GPI-linked Proteins
Cell Membrane
Cytoplasm
Proteins
glycophosphatidylinositol- GPI Anchor
CD59, CD90, CD109
CD55
CD58*
CD59
CD48
CDw52
PrPc
CD16*
CD24 CD55
CD58* CD59
CD48 PrPC
CD73 CDw108
CD55
CD58*
CD59
CD109
PrPC
GP500
Gova/b
CD55
CD58*
CD59
PrPC
AChE
JMH Ag
Dombroch
HG Ag
CD55 CD58*
CD59 CD14
CD16 CD24
CD48 CD66b
CD66c CD87
CD109 CD157
LAPNB1 PrPC
p50-80 GPI-80
ADP-RT
NA1/N
A2
CD14 CD55
CD58* CD59
CD48 CDw52
CD87 CD109
CD157 Group 8
PrPC GPI-80
CD16*
CD55 CD58*
CD59 CD48
CDw52 CD87
CDw108 PrPc
ADP-RT CD73
CD90 CD109
CD16*
GPI-linked Proteins Deficient on PNH Blood Cells
Hematopoietic
Stem Cell
Platelets
RBC
PMN
B cells
Monocytes
T cells
NK cells
nucleus
ER
X chr.
PIGA
+ GPI protein
Etn-P GlcN
Inositol-P
glycan
PIGA
GPI anchor
nucleus
ER
X chr.
PIGA
+ GPI protein
PIGA
GPI anchor
Etn-P GlcN
Inositol-P
glycan
Hematopoietic Stem Cell
Normal cell Normal cell PNH cell
Paroxysmal Nocturnal Hemoglobinuria (PNH)
Pathogenesis
6th European Symposium on Rare Anaemias - 1st Dutch-Belgian meeting for patients and health professionals
• Intravascular hemolysis
• Thrombosis
• Bone marrow failure
Paroxysmal Nocturnal Hemoglobinuria
6th European Symposium on Rare Anaemias - 1st Dutch-Belgian meeting for patients and health professionals
• Intravascular hemolysis
• Thrombosis
• Bone marrow failure
Paroxysmal Nocturnal Hemoglobinuria
C5
C3
CD55
CD59
Factor H
CD46
CR1
Factor I
C5
C3
C4bp
Complement Inhibitors
Membrane Soluble
Complement System Microorganisms Antigen-Antibody
Constitutive/
Microorganisms
Classical pathway
Lectin pathway
Alternative pathway
modified from Brown et al, Nat Clin Pract Nephrol 2007
Clusterin
Protein S
C1 Inh
Factor I
Membrane
Attack Complex
CD55
CD59
in PNH
6th European Symposium on Rare Anaemias - 1st Dutch-Belgian meeting for patients and health professionals
Effects of intravascular hemolysis
Hemosiderinuria
Iron loss
ANEMIA (low Hgb)
Nitric oxide
depletion
smooth muscle
dystonia
• Abdominal pain
• Fatigue
• Dysphagia
• Erectile disfunction
• (Renal failure)
• (Pulmonar hypertension)
Intravascular hemolysis
Hemoglobinemia
Hemoglobinuria
High LDH
High Retics
6th European Symposium on Rare Anaemias - 1st Dutch-Belgian meeting for patients and health professionals
• Intravascular hemolysis
• Thrombosis
• Bone marrow failure
Paroxysmal Nocturnal Hemoglobinuria
6th European Symposium on Rare Anaemias - 1st Dutch-Belgian meeting for patients and health professionals
• 30-40% of PNH patients suffer from at least a thrombotic event
• 40–67% of deaths in PNH results from thromboembolisms
• More frequent in hemolytic than in aplastic PNH patients
• There is a rough correlation with the size of PNH population - but thrombosis can occur also in patients with small PNH population
• More frequent in patients with congenital and acquired thrombophilic conditions
• Substantial increased risk after the first thrombotic event
PNH is a severe acquired thrombophilic state - thrombosis in PNH -
6th European Symposium on Rare Anaemias - 1st Dutch-Belgian meeting for patients and health professionals
Hillmen et al, Blood 2007
Ictus/transient ischaemic attack (TIA)
Myocardial infarction/unstable angina
Arterial Thrombosis 15.3%
Deep vein thrombosis
- lower extremity
- other
Mesenteric/splenic vein thrombosis
Hepatic/portal vein thrombosis
Pulmonary embolus
Cerebral/internal jugular thrombosis
Superficial vein thrombosis
Venous Thrombosis 84.7%
% of Total
33.1
18.5
14.5
18.5
16.9
6.5
5.6
4.0
13.7
1.6
Sites of thrombosis IN PNH
6th European Symposium on Rare Anaemias - 1st Dutch-Belgian meeting for patients and health professionals
Coagulation Complement
Mo
difie
d fro
m M
ark
iew
ski e
t a
l, T
ren
ds Im
mu
no
l 2
00
7
Coagulation, Complement and PNH
Partners in Crime?
Coagulation Complement
6th European Symposium on Rare Anaemias - 1st Dutch-Belgian meeting for patients and health professionals
• Intravascular hemolysis
• Thrombosis
• Bone marrow failure
Paroxysmal Nocturnal Hemoglobinuria
6th European Symposium on Rare Anaemias - 1st Dutch-Belgian meeting for patients and health professionals
Bone Marrow Failure and PNH Aplastic Anemia and PNH relationship
Bone Marrow Failure and PNH
Dual Pathogenesis of PNH
Hemolysis
Cytopenia
PNH
AA
PNH/AA
Bone Marrow Failure targeting GPI+ cells (?)
Complement Susceptibility targeting GPI– cells
Rotoli & Luzzatto, 1989
PIG-A+
cells
PIG-A null
cells
crippled
PIG-A+ cells
Noxious agent
Time
Paradoxical Expansion of PNH Clone (Escape Theory)
Y
Y
Y
Y
Y
T Lymphocytes autoreactive T Lymphocytes Hematopoietic Stem cell
?? IFN
IFN
IFN
IFN
GPI protein
GPI protein
ID of Lymphocytes that
damage normal cells and spare PNH cells Which is the target?
Skewed TCR repertoire (Karadimitris et al, Blood 2000)
Similar TCR sequences in oligoclonal T cells (Gargiulo et al, Blood 2007)
GPI specific and CD1d restricted T cells (Gargiulo et al, Blood 2013)
Normal Stem Cell (GPI+) PNH Stem cell (GPI-)
A Unified Pathogenetic Model for PNH and AA CD1d restricted autoreactive T cells target the GPI anchor
anti-GPI TCR
CD1d
6th European Symposium on Rare Anaemias - 1st Dutch-Belgian meeting for patients and health professionals
Hematopoietic stem cells GPI-targeted
noxious agent
Healthy
Hematopoietic System
Damaged
Hematopoietic stem cells
PNH clone(s)
(GPI-
deficient)
No clinical
consequences
PIG-A
mutation
Selective
expansion
of PNH clone(s)
PNH Intravascular
haemolysis Thrombosis
CD59(-)
red cells
Abnormal
platelets
Pancytopenia
Aplastic
Anaemia
A Unified Pathogenetic Model for PNH and AA
6th European Symposium on Rare Anaemias - 1st Dutch-Belgian meeting for patients and health professionals
Clinical Management
Paroxysmal Nocturnal Hemoglobinuria
6th European Symposium on Rare Anaemias - 1st Dutch-Belgian meeting for patients and health professionals
Support treatments • Folic Acid • Iron supply • Red blood cell transfusions • Anti-coagulant Profilaxis • (Thrombolysis)
Phatogenetic treatments • Allogeneic HSC Transplantation • Immuno-suppression (ALG/CSA) • (Androgens)
Paroxysmal Nocturnal Hemoglobinuria therapeutic options in pre-eculizumab era
6th European Symposium on Rare Anaemias - 1st Dutch-Belgian meeting for patients and health professionals
Support treatments • Folic Acid • Iron supply • Red blood cell transfusions • Anti-coagulant Profilaxis • (Thrombolysis)
Phatogenetic treatments • Allogeneic HSC Transplantation • Immuno-suppression (ALG/CSA) • (Androgens)
Paroxysmal Nocturnal Hemoglobinuria therapeutic options in pre-eculizumab era
6th European Symposium on Rare Anaemias - 1st Dutch-Belgian meeting for patients and health professionals
Support treatments • Folic Acid • Iron supply • Red blood cell transfusions • Anti-coagulant Profilaxis • (Thrombolysis)
Phatogenetic treatments • Allogeneic HSC Transplantation • Immuno-suppression (ALG/CSA) • (Androgens)
Paroxysmal Nocturnal Hemoglobinuria therapeutic options in pre-eculizumab era
6th European Symposium on Rare Anaemias - 1st Dutch-Belgian meeting for patients and health professionals
Support treatments • Folic Acid • Iron supply • Red blood cell transfusions • Anti-coagulant Profilaxis • (Thrombolysis)
Phatogenetic treatments • Allogeneic HSC Transplantation • Immuno-suppression (ALG/CSA) • (Androgens)
Paroxysmal Nocturnal Hemoglobinuria therapeutic options in pre-eculizumab era
6th European Symposium on Rare Anaemias - 1st Dutch-Belgian meeting for patients and health professionals
Support treatments • Folic Acid • Iron supply • Red blood cell transfusions • Anti-coagulant Profilaxis • (Thrombolysis)
Phatogenetic treatments • Allogeneic HSC Transplantation • Immuno-suppression (ALG/CSA) • (Androgens)
Paroxysmal Nocturnal Hemoglobinuria therapeutic options in pre-eculizumab era
6th European Symposium on Rare Anaemias - 1st Dutch-Belgian meeting for patients and health professionals
Support treatments • Folic Acid • Iron supply • Red blood cell transfusions • Anti-coagulant Profilaxis • (Thrombolysis)
Phatogenetic treatments • Allogeneic HSC Transplantation • Immuno-suppression (ALG/CSA) • (Androgens)
Paroxysmal Nocturnal Hemoglobinuria therapeutic options in pre-eculizumab era
6th European Symposium on Rare Anaemias - 1st Dutch-Belgian meeting for patients and health professionals
Complement Blockade
- eculizumab -
Paroxysmal Nocturnal Hemoglobinuria
6th European Symposium on Rare Anaemias - 1st Dutch-Belgian meeting for patients and health professionals
CH
3
CH
2
Rother et al, Nature Biotech 2007
Eculizumab
Anti-C5 humanized monoclonal antibody
Variable
heavy chain
Variable light
chain
Human constant
light chain Ck
Human constant
heavy chain
IgG4 CH2 and CH3
Human constant
heavy chain
IgG2 CH1 and hinge
Human framework
regions
Murine
complementarity
determining regions
6th European Symposium on Rare Anaemias - 1st Dutch-Belgian meeting for patients and health professionals
in Healthy Subjects in PNH Patients
C5 C3
CD
55
CD
59
CD
46
CR
1
Complement Inhibitors
C5 C3
Regulation of Complement Cascade
CD
55
CD
59
C5b-C9
Facto
r H
Facto
r I
C4b
p
Classical
pathway
Lectin
pathway
Alternative
pathway
6th European Symposium on Rare Anaemias - 1st Dutch-Belgian meeting for patients and health professionals
in PNH Patients Treated with Eculizumab
C5 C3
CD
55
CD
59
CD
46
CR
1
Complement Inhibitors
C5 C3
Regulation of Complement Cascade
CD
55
CD
59
Eculizumab Facto
r H
Facto
r I
C4b
p
Classical
pathway
Lectin
pathway
Alternative
pathway
Effect of eculizumab on intravascular haemolysis LDH levels
2500
LD
H
2000
1500
1000
500
TRIUMPH – placebo
0
weeks
10 20 30 40 50
Pilot
TRIUMPH – eculizumab
SHEPHERD
Eculizumab
Hillmen, N Eng J Med 2004; Hillmen, N Eng J Med 2006; Brodsky, Blood 2008
Eculizumab
Placebo
Eculizumab
Hillmen et al, Brit J Haematol 2013
Eculizumab abrogates the need of blood transfusion in most PNH patients
Patients
without
blo
od tra
nsfu
sio
n
du
rin
g th
e p
revio
us 6
mo
nth
s
35%
18%
92%
PRE
Eculizumab
6th European Symposium on Rare Anaemias - 1st Dutch-Belgian meeting for patients and health professionals
(P<0,001)
Patients = 103 10.61
0.62
0
4
8
12
Pre eculizumab On eculizumab
TE
/ 1
00
pa
tie
nt
ye
ars
Hillmen et al, Blood 2007
Eculizumab and Thrombosis thromboembolism in patients on prophylaxis
6th European Symposium on Rare Anaemias - 1st Dutch-Belgian meeting for patients and health professionals
100
80
60
40
20
0
0 10 20
Years since diagnosis
Su
rviv
al
(%
)
PNH (n 80; Hillmen 1995)
30 40
PNH (n 424; De Latour 2008)
Overall Survival of PNH Patients in pre-eculizumab era
Hillmen et al, Brit J Haematol 2013
in eculizumab era
Mortality by: Thrombosis
Bone marrow failure
(MDS/leukemia?)
6th European Symposium on Rare Anaemias - 1st Dutch-Belgian meeting for patients and health professionals
Extravascular Hemolysis
- a pattern change in PNH -
Paroxysmal Nocturnal Hemoglobinuria
6th European Symposium on Rare Anaemias - 1st Dutch-Belgian meeting for patients and health professionals
C5 blockade is associated with C3-binding on GPI- RBC
CD
59
-PE
CD
59-P
E
C3-FITC
CD
59-P
E
CD
59-P
E
Untreated PNH patient
PNH patients during eculizumab treatment
Hgb≥12 8.5≤ Hgb <10 Transfused
C3-FITC C3-FITC
2% of
PNH RBC
45% of
PNH RBC 67% of
PNH RBC
Risitano AM, Notaro R et al, Blood 2009
C3-FITC
6th European Symposium on Rare Anaemias - 1st Dutch-Belgian meeting for patients and health professionals
Eculizumab treatment is associated
with C3-binding on PNH red cells
CD
59-P
E
C3-FITC
CD
59-P
E
Patient not on eculizumab
Patient on eculizumab
45% of
PNH RBC
C3-FITC
Risitano, Notaro et al, Blood 2009
C5 C3 Alternative
pathway C5 C3
C3
Eculizumab
C5 C3 Alternative
pathway C3
C3
6th European Symposium on Rare Anaemias - 1st Dutch-Belgian meeting for patients and health professionals
A clinical case
Paroxysmal Nocturnal Hemoglobinuria
years
0
1
2
3
4
5
6
7
8
9
10
11
12
pre-ecu
0 1 2 3 4 5 6 7 8 9
Hb (g/dl)
Neutrophils *103/µl
LDH (*UNL)
Retics *105/µl
Plts *105/µl
13 Packed RBC
2 U/mo
years from starting eculizumab
Eculizumab (900 mg/14 days)
Heavily transfused PNH patients suffering from recurrent thrombosis
no more blood transfusions and no more thrombosis
(despite the discontinuation of anti-coagulant prophylaxis)
-2 -1
recurrent
thrombosis
(Budd Chiari)
Hb (g/dl)
LDH (*UNL)
Retics *105/µl
Plts *105/µl
eculizumab starts and the story changes ……………
Blood
Transfusions
Eculizumab stops intravascular hemolysis and related symptoms
abrogates or reduce transfusion need
reduces thrombotic risk
Hemoglobinuria
Anemia
Hemoglobinemia
• Fatigue
• Dysphagia
• Erectil Dysfunction
•Abdominal Pain
is safe (also during the pregnancy)
However, it is not curative
extravascular hemolysis may result in transfusional need
it is not available to all patients (even in EU)
Thrombosis
Future Developments
Paroxysmal Nocturnal Hemoglobinuria
6th European Symposium on Rare Anaemias - 1st Dutch-Belgian meeting for patients and health professionals
C5
C3
CD55
CD59
Factor H
CD46
CR1
Factor I
C5
C3
C4bp
CD55
CD59
Complement Inhibitors Membrane Soluble
New complement inhibitors
Eculizumab
MoAb AntiC3
Compstatin
TT30 (FH-CR2 hybrid)
CR1-like (no EPN)
Fattore H-like
MoAb AntiCFD (no EPN)
MoAb AntiCFB
siRNA anti CFB
Small molecules
Aptamers
Coversin
MoAb AntiC5 (no EPN)
Cyclic Peptids
siRNA
CRL- ITT, Florence Genetics and Gene Transfer
Rosario Notaro
Michela Sica, Lucia Gargiulo, Tommaso Rondelli, Margherita Berardi, Giulia Talini, Annamaria Pellecchia,
Emanuele Di Lorenzo, Benedetta Peruzzi, Chiara Pescucci,
Maria De Angioletti
Imperial College - London (UK) Anastassios Karadimitris, Lucia Gargiulo
Federico II University - Naples (Italy) Antonio M. Risitano, Giuseppina Ruggiero, Fiorella Alfinito,
New York University - New York (USA) David Araten
Hopital Saint-Louis, Paris (France) Gerard Sociè, Regis Peffault de Latour
Kanazawa University, Kanazawa (Japan) Shinji Nakao, Yoshitaka Zaimoku
Sapienza University - Rome (Italy) Anna Paola Iori
Ospedale Maggiore - Milan (Italy) Carla Boschetti, Wilma Barcellini
AOU Careggi - Florence (Italy) Roberto Caporale, Luigi Rigacci, Renato Alterini, Valentina
Carrai, Giacomo Gianfaldoni,
Barbara Scappini, Giovanni Longo, Grazia Gelli,
Francesco Attanasio,
University of Florence (Italy) Alberto Bosi, Valeria Santini, Francesco Mannelli,
Ilaria Cutini, Cristina Nativi, Barbara Richichi
Nurse Staff Careggi Hospital
Istituto Toscano Tumori
Lucio Luzzatto
Core Research
Laboratory
6th European Symposium on Rare Anaemias - 1st Dutch-Belgian meeting for patients and health professionals
Dank aan de Patiënten en hun Families
Merci aux patients et à leurs familles
Thanks to Patients and their Families
Grazie ai Pazienti ed alle loro Famiglie
6th European Symposium on Rare Anaemias - 1st Dutch-Belgian meeting for patients and health professionals
There is a soul of goodness in things evil, Would men observingly distil it out.
William Shakespeare, King Henry V, IV, i