Transcript GENE THERAPY IN HEMOPHILIA

Rare Bleeding Disorders (RBDs)
Flora Peyvandi
Hemophilia and Thrombosis Center,
University of Milan
ISTH Educational Course: Advanced
Course in Haemostasis and
Thrombosis
Moscow, September 17-19, 2014
Bleeding disorders
Inherited bleeding deficiencies affect 65,000 people in Europe
•
Hemophilia A and B (prevalence 1:5,000 and 1:30,000)
•
von Willebrand disease (VWD) (prevalence 1:100)
•
Platelet disorders (prevalence 1:1,000,000)
•
Rare bleeding disorders (deficiency of fibrinogen, FII, FV, FV+FVIII, FVII,
FX, FXI and FXIII) (prevalence ranging from 1:500,000 for FVII deficiency to
1:2 million for FII and FXIII deficiencies)
•
•
•
RBDs represent 3-5% of all inherited coagulation
deficiencies
Usually transmitted as autosomal recessive traits
In some countries, prevalence is higher due to
consanguineous marriage
Erhardtsen http://www.springerreference.com/index/chapterdbid/33723 2012
Poon M-C et al. Vasc Health Risk Manag 2007;3:655–64
Peyvandi F et al. Semin Thromb Hemost 2009;35:349–355
Bolton Maggs P Pediatr Blood Cancer 2013;60(Suppl 1):S37-40
Outlines of discussion on RBDs
•
Epidemiology
•
Data collection: registers
•
Clinical manifestations
•
Phenotype diagnosis
•
Association between laboratory phenotype and
clinical severity
•
Genotype characterization
•
Treatments and available products
Epidemiology
Funded by the European Community - action programme for public
health (DG SANCO)
Survey updated to December 2013
(http://www1.wfh.org/publications/files/pdf-1574.pdf)
WFH
WORLD
EU
109
28
172,373
44,494
N° of people with
VWD
66,144
34,518
N° of people with rare
coagulation disorders
17,519
8,959
N° of people with
platelet disorders
11,430
2,716
4,192
1,763
271,658
92,450
N° of countries
N° of people identified
with HA and B
N° of people with
unknown type of
hemophilia or
coagulation disorder
Total
World map of bleeding disorders other
than hemophilia and VWD
Bleeding disorders
in Russia
Uncertain diagnosis
Data collection: registers
•
Improving knowledge on prevalence
•
Observing course of disease
•
Understanding variations in symptoms
•
Associating the laboratory phenotype and clinical picture
•
Monitoring treatments
•
Assessing effectiveness of existing treatments
F. Peyvandi, M. Spreafico. Blood Trand 2008; 6(Suppl. 3):s34-8.
Soucie JM et al. Am J Prev Med 2010; 38:S475-81.
Orphanet Report Series – Rare Disease collection. Rare Disease Registries in Europe. Jan 2014
National registries in EU
UK - NHD: national
haemophilia
database
Belgian patient
registry for rare
bleeding disorders
FranceCoag: French
prospective cohort
of patients affected
with haemophilia or
severe form of
other hereditary
hemorrhagic
diseases except
platelet disorders
European network of Rare Bleeding Disorders (EN-RBD)
PRO-RBDD : Prospective data collection on patients with
coagulation fibrinogen and factor XIII deficiencies
DHR: German
Haemophilia Registry
Austrian Haemophilia
Registry
Slovenian registry of
patients with inherited
bleeding disorder
Serbian registry of
patients with rare
bleeding disorders
EMOWEB: Italian
registry of
coagulopathies
Registry of inherited
bleeding disorders in
Emilia Romagna region
Clinical manifestations
Highly variable bleeding tendency
•
The most typical symptoms are
-
mucosal tract bleedings
-
excessive bleeding at the time of invasive procedures,
-
Bleeding at labour and delivery in women, bleeding after circumcision in
affected boys
•
CNS bleeding, umbilical cord bleeding, hemarthroses and soft tissue haematomas
are frequent in fibrinogen, FVII, FX and FXIII deficiencies
•
GI bleeding occurs mainly in FX deficiency
•
Heterozygous patients (coagulant activity level >30%):
– 20% have mucocutaneous bleedings and bleeding during surgery
– Post-traumatic hemarthrosis and hematomas are rarely reported in FVII and FX
deficiencies
– Menorrhagia, spontaneous abortion and bleeding during vaginal delivery were
reported in roughly 20% of women with all type of deficiencies
•Peyvandi F et al. Semin Thromb Hemost 2013;39:579-84; Peyvandi F et al. Hematology Education Book 2010;4(1):6368; Karimi M et al. Haematologica 2008;93:934-938; Mariani G et al. Thromb Haemost 2005;93:481-7
Women with RBDs: clinical problems
Menstruation
Age
• Menorrhagia
Pregnancy
•
Bleeding during pregnancy
•
Miscarriage
Delivery
• Post-partum hemorrhage
Kadir R and James A WFH Reproductive health in women with bleeding disorders 2008
Prevalence of obstetric-gynecological symptoms
Bleeding
disorder
Menorrhagia
35-78%
Rare bleeding
disorders
Haemophilia A
and B carriers
(Lak 1999, Siboni
2009, James 2010,
Khair 2013, Mariani
2013, Shetty 2014,
Napolitano 2014)
10-67%
(Siboni 2009, James
2010, Khair 2013)
Abortion
Post partum haemorrhage
Afibrinogenemia 30-50%
Hypofibrinogenemia 29%
Dysfibrinogenemia 38%
FXI 10%
FXIII 50-63%
Hypofibrinogenemia 45%
FV 76%
FVII,FX,FXI,FXIII high rates
(Goodwin 1989, Haverkate 1995,
Burrows 2000, Lak 2003, Myers 2007,
Mensah 2011)
14-20%
(Chi 2008, Knol 2010)
(Goodwin 1989, Noia 1997, Kadir 1998, Burrows
2000, Kulkarni 2006, Teixeira 2012, Baumann
Kreuziger 2013)
the most significant cases in
women with FVIII <50% and no
hemostatic coverage during labor
and postpartum
(Kadir et 1997)
32-100%
VWD
(Lak 2000, Siboni 2009,
James 2010 and 2011,
Khair 2013)
Glanzmann’s
thrombasthenia
Full references in slide notes
50-98%
(James 2010, Khair
2013)
15-25%
(Kadir 1998, Kirtava 2003)
11%
(Siddiq 2011)
16-29% primary
20-29% secondary
(Ramsahoye 1995, Kadir 1998)
34% primary
24% secondary
(Siddiq 2011)
12
Complications in women
•
Anemia due to iron deficiency
•
Dysmenorrhea
•
Hysterectomy: effective and definitive treatment
-
•
However, it is a major surgical procedure with significant physical and
emotional complications and social and economic costs
Maternal death (post-partum bleeding)
Peyvandi F Expert Opinion 2014
Geographical variation in causes of
maternal death
*Represents HIV/AIDS. †Represents embolism.
‡Represents ectopic pregnancy. §Represents anaemia.
Khan K et al. Lancet 2006;367:1066–74
Phenotype diagnosis
Kallikrein + Kininogen +
calcium
(XII)
Tissue factor + calcium
XI
Intrinsic pathway
IX
aPTT
VII
VIII
X
V
II
Trombin
TT
Fibrinogen
PT
Extrinsic pathway
TR
Reptilase
D-Dimer
Fibrinolysis
Fibrin
XIII
Screening tests
aPTT and PT tests:
• Usually sensitive enough to detect abnormal coagulant factor actiivity:
fibrinogen, FII, FV, FV+FVIII, FVII, FX and FXI
• May be normal in mild deficiencies, especially fibrinogen and FII
• Normal in FXIII deficiency
Mixing tests:
• Correction of the abnormality indicates that a factor deficiency
• No correction indicates presence of an inhibitor
TT test:
• Required for the diagnosis of fibrinogen deficiency
Interpretation in relation to the personal and family history
Kitchen S, McCraw A, Echenagucia A. Diagnosis of Hemophilia and Other Bleeding Disorders. A LABORATORY MANUAL. Second Edition. WFH 2010
Specific factor assays
Factor activity level
• One-stage coagulation assays:
•
•
-
aPTT based – FXI deficiency
-
PT based – FII, FV, FVII, FX deficiencies
Fibrinogen assays
FXIII assays
Factor antigen level
• Immunochemical methods
Not mandatory except for fibrinogen and FII deficiencies.
Kitchen S, McCraw A, Echenagucia A. Diagnosis of Hemophilia and Other Bleeding Disorders. A LABORATORY MANUAL. Second Edition. WFH 2010
Peyvandi F et al. Semin Thromb Hemost 2009; 35(4): 349-55.
Fibrinogen activity assays
•
•
Clauss fibrinogen assay
PT-derived Fg assay
Levels measured by different assays may
vary significantly
Uncertainty in assigning the severity of the
hypofibrinogenemia
Recommendations
•
Clauss fibrinogen assay remains the reference method
•
PT-Fg assays are not recommended for routine clinical use
Mackie IJ et al. Guidelines on fibrinogen assays. Br J Haematol 2003; 121: 396-404.
FXIII activity assays
•
•
Clot solubility assay
Functional FXIII activity assays:
Recommendations by the FXIII and Fibrinogen SSC
subcommittee of the ISTH
-
Photometric assays
1. First-line screening test
Quantitative functional FXIII activity assay
-
Fluorimetric assays
If FXIII activity is decreased
-
Putrescine incorporation assays
2. Establish subtype of FXIII deficiency
Measurement of antigen concentration
(FXIII-A2B2, FXIII-A, FXIII-B)
Note
•
Association in patients with FXIII < 5% is
difficult to interpret and may not be accurate
•
Clot solubility assay is no longer
recommended since it is qualitative, poorly
standardized, and detects only severe FXIII
deficiency
Only in case of suspected acquired FXIII deficiency:
3. Detection of autoantibodies
Mixing study (detection of neutralizing antibodies
against FXIII-A)
Binding assay (detection of nonneutralizing antibodies)
For research purpose only:
Characterization of molecular genetic defect
Schroeder V et al. FXIII deficiency: An Update. Semin Thromb Hemost 2013; 39: 632-41
Phenotype diagnosis: a summary
Deficiency
TT
First screening
PT
APTT
Second screening
Notes
activity antigen



no
no
Low erythrocyte sedimentation rates
Hypofibrinogenemia





TT is the most sensitive
Dysfibrinogenemia



normal
normal
FII, FV, FV+FVIII, FX
normal



FVII
normal

normal

FXI
normal
normal


FXIII
normal
normal
normal
Afibrinogenemia

When activity is normal, gnetic mutations is nedeed
normal PT performed with rTF is extremely sensitive to tiny amount
normal of FVII, attention needs to be payd to level <1%
normal If plasmatic FXIII-A2B2 :
FXIII-A in plasma and platelet lysate could be or normal (FXIII-B >30%)
FXIII-B in plasma and in platelet lysate 
Problems with clotting assays
Despite the existence of recommendations, a number of problems still
remain, particularly for some deficiencies (Fibrinogen and FXIII):
─
─
Assay accuracy at low factor levels
Lack of standardization – high inter-laboratory variability
Laboratory phenotype and clinical severity
•
•
•
3 years of data collection
13 European centers from 11 countries
489 patients registered in the database
Assigned categories of clinical bleeding severity
•Peyvandi F et al J Thromb Haemost2012;10:615-621
Results
Linear regression analysis: adjusted for age, gender, and country where diagnosis was made
Factor activity (Y) = Beta* Bleeding severity (X) + Constant
Excellent correlation
Good correlation
No correlation
SSC – ISTH recommendations on RBDS
Laboratory phenotype
Coagulant factor
deficiency
Severe
Moderate
Mild
(coagulant activity associated
with spontaneous major
bleeding)
(coagulant activity associated
with minor spontaneous or
triggered bleeding)
(coagulant activity associated
with a mostly asymptomatic
disease course)
Coagulant activity
Fibrinogen
undetectable clot
≤0,1 g/L
>0,1 g/L
FII
undetectable activity
≤10%
>10%
FV
undetectable activity
10%
≥10%
FV+FVIII
<20%
20-40%
>40%
FVII
<10%
10-20%
>20%
FX
<10%
10-40%
>40%
undetectable activity
30%
≥30%
FXIII
Do clotting factor levels predict severity?
• The relationship between coagulation factor activity level and
clinical bleeding severity is heterogeneous
• A strong association exists for fibrinogen, FX and FXIII deficiencies
• No association for FXI deficiency
• The minimum factor levels to prevent spontaneous major
bleeding is different in various deficiencies
It is not appropriate to use a single criterion of classification
for all types of RBDs
Global hemostasis assays
Tests investigating the global hemostatic capacity could help:
- to predict clinical phenotype
- to determine the effectiveness of therapies
- to monitor treatment
particularly FXI deficiency where standard assays fail to correlate
with bleeding risk
TGT
TEG
•
Recently, these tests have been used to evaluate hemostasis in patients with
RBDs, specifically FV and FXI deficiency
•
However test standardization to reproduce reliable measurements facilitated
by standardized pre-analytical and analytical procedures is required before
widespread clinical use.
•
In global hemostasis assays, a number of barriers may prevent implementation
and quality assurance. Quality assurance studies have observed that regular
proficiency testing is needed to ensure accuracy of such methods
Al Dieri R et al, Thromb Haemost. 2002; Strey RF et al, Pathophysiol Haemost Thromb. 2005; Schols SE et al, Thromb Haemost 2008; Rugeri L et al,
Haemophilia 2010; Guegen P et al, Br J Haematol 2011; Riddell A et al, Thromb Haemost 2011: Van Geffen M et al, Haemophilia. 2012; Spiezia L et
al, Haemophilia. 2012; Livnat T et al. Blood Coag Fibrynol 2012
Genotype characterization
• Currently based on the mutation search in the genes that encode each
corresponding coagulation factor
• Exceptions:
- combined deficiency of coagulation FV and FVIII (mutations in genes for
FV and FVIII intracellular transport – MCFD2 and LMAN1)
- combined deficiency of the vitamin-K-dependent proteins (mutations in
genes for post-translational modifications and vitamin K metabolism
(GGCX and VKORC1)
• Pattern of inheritance is autosomal recessive for all RBDs, except for some
cases of FXI, where some missense mutations were shown to exert a
dominant negative effect, and of dysfibrinogenemia
Mutations causing RBDs
Type of deficiency
Gene
Indels (%)
Fibrinogen
FGA
FGB
FGG
20,9
57,3
11,5
7,5
2,8
FII
F2
13
77,8
5,5
3,7
0
FV
F5
27,3
48,5
12,9
11,3
0
LMAN1
50
8,8
20,6
20,6
0
MCFD2
27,3
50
4,5
18,2
0
FVII
F7
12,3
62,2
7,8
11,5
6,2
FX
F10
10,5
80
1,9
7,6
0
FXI
F11
10,4
70
10,5
8,2
0,9
FXIII
F13
29
47,1
9,1
14
0,8
GGCX
10
60
0
30
0
VKORC1
0
100
0
0
0
FV+VIII
Vit-K factors
Missense (%) Nonsense (%) Splicing (%) 3'-5' UTR (%)
•
5-10% of patients remain with no identified genetic defect
•
The use of next generation sequencing (NGS) might help to identify
novel pathways in coagulation disorders
•
While the potential of these genome-wide strategies is indisputable,
these approaches have yet to be utilized in the analysis of RBDs for
which causative mutations remain elusive
•Peyvandi F et al. Blood 2013;122:3423-31.
Treatment of RBDs
Non-transfusional treatment:
•
antifibrinolytic amino acids
•
fibrin glue
•
oestrogen-progesterone preparations
•
desmopressin (DDAVP)
Replacement therapy:
•
FFP
•
Cryoprecipitate (also virus-inactivated)
•
prothrombin complex concentrates (PCC)
•
single factor concentrates (plasma-derived or recombinant)
Flora Peyvandi
Registered RBDs concentrates
39
40
35
30
25
20
18
15
15
10
5
4
4
0
1
2
2
0
Registry of Clotting Factor Concentrate, Ninth Edition
Mark Brooker, WFH
Novel PD concentrates: ongoing studies
• Fibrinogen
─
NCT01575756 – Octapharma (safety/efficacy, phase 2)
─
NCT02065882 – Biotest (PK/safety/efficacy, phase 1-2)
─
NCT00916656 – CSL (Phase 3b non inferiority safety/efficacy - withdrawn prior
to enrollment)
─
NCT02094430 – LFB (Phase 2-3 - clinical pharmacology/efficacy/safety in
paediatric patients)
─
FIBGrifols
• FX (high purity)
─
NCT01721681 – BPL (prophylaxis in <12 years, phase 3)
• FV
─
Only recently a concentrate has been developed. Preclinical studies are
currently being performed for the orphan drug designation application (EMA
and FDA)
On-demand treatment
Deficient
factor
Plasma half-life
Recommended
trough levels
10–20%
Cryoprecipitate (15-20 mL/kg)
SD-treated plasma (15–30 mL/kg)
Fibrinogen concentrate (50–100 mg/kg)
SD-treated plasma (15–25 mL/kg)
FIX concentrate and PCC (20–40 units/kg)
SD-treated plasma (15–25 mL/kg)
10–15%
As for FV
Fibrinogen
2–4 days
0.5-1g/L
Prothrombin
3–4 days
20–30%
Factor V
36 hours
Factor V and
FV 36 hours
factor VIII
FVIII 10–14 hours
Factor VII
4–6 hours
10–15%
Factor X
40–60 hours
10–20%
Factor XI
50 hours
15–20%
Factor XIII
9–12 days
2–5%
Vitamin K
dependent
On demand dosages
FVII concentrate (30–40 mL/kg)
PCC (20–30 units/kg)
rFVIIa (15–30 μg/kg every 4–6 hours)
SD-treated plasma (10–20 mL/kg)
PCC (20–30 units/kg)
FX/FIX concentrate (10–20 units/kg)
SD-treated plasma (15–20 mL/kg)
FXI concentrate (15–20 units/kg)
Cryoprecipitate (2–3 bags)
SD-treated plasma (3 mL/kg)
FXIII concentrate (till 50 units/kg for high
hemorrhagic events)
rFXIII-A (35 units/kg)
Vitamin K (10 mg) IV. or SC. for minor bleeding
PCC (20–30 units/kg) with vitamin
K (5–20 mg) for severe bleeding or major surgery
FFP 15–25 ml/kg is an alternative to PCC
Recommended trough levels to
maintain asymptomatic state after
publication of the EN-RBD results
1 g/L
>10 %
10%
40%
>20%
>40%
15-20%
30%
No data available
Peyvandi F. Haemophilia 2002; Mannucci PM. Blood. 2004; Peyvandi F et al J Thromb Haemost2012;10:615-621; Mumford AD, Br J Haematol 2014
Prophylaxis
The choice of prophylaxis is related to:
– the frequency of bleeding
– the risk of severe spontaneous bleeding
– the risk of long-term disabilities associated with the occurrence of
bleeding in a particular region of the body despite on-demand
treatment (e.g., CNS, GI and joint bleedings)
Prophylactic treatment
Deficient factor
Recommended
trough levels
Reported dose schedule for successful long-term prophylaxis
Notes
Products
Cryoprecipitate
Fibrinogen
0.5-1g/L
Prothrombin
20–30%
Fibrinogen
concentrate
PCC
Factor V
10–20%
SD-treated plasma
Factor V and
Factor VIII
10–15%
Factor VII
10–15%
Factor X
10–20%
SD-treated plasma
pdFVII
rFVIIa
PCC
15–20%
Cryoprecipitate
Factor XIII
2–5%
SD-treated plasma
FXIII concentrate
rFXIII-A
Vitamin K
dependent
Frequency
1 unit
3 units
3 times/week
Every 7–10 days
30–100 mg/kg
Every week
20–40 units/kg
1/ week
20-30 mL/kg
2/week
No data
FX/FIX concentrate
Factor XI
Dose
Vitamin K
10–15 mL/kg
10–40 units/kg
20–40 µg/kg
20–40 units/kg
20–40 units/kg
No data
2 units
Afibrinogenemic patients with
recurrent life-threatening bleedings
or undergoing surgeries
—
Only in patients with lifethreatening bleedings, as CNS
—
2/week
3 times/week
2-3 times/week
2-3 times/week
1-2 times/week
Prevention of bleeding during
surgery or in children with recurrent
hemarthrosis or CNS
Patients with recurrent lifethreatening bleedings or
undergoing surgeries
—
Every 3 weeks
15–20 mL/kg
Every 4–6 weeks
10-40 units/kg
35 units/kg
Every 4–6 weeks
Every 4
5-20 mg
1/week orally
Mannucci PM. Blood. 2004; Castaman G. Blood Transfusion 2008; Mumford AD, Br J Haematol 2014
Highly recommended in severe
patients
—
Conclusion
Due to the RBDs rarity, little information is available on the optimal
management of patients with these disorders, particularly on longterm prophylaxis
• In addition, the technical limitations of laboratory testing,
particularly in patients with factor activity <5%, make difficult to
adopt the appropriate prophylactic treatment
• To overcome these limitations, new strategies are required:
•
–
–
–
–
–
creation of global partnerships
networking between treatment centers
increasing the support provided by public health organizations
Prospective data collection
Better and more sensitive assays
The needs
Patients association
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ACKNOWLEDGMENTS
All patients and clinicians who are collaborating with us
•
•
European Commission
•
•
Executive Agency for Health and Consumers
(EAHC)
Fondazione IRCCS
Maggiore Policlinico
•
University of Milan
•
Luigi Villa Foundation
The European Hemophilia Network
(EUHANET)
project
•
Cà
Granda
Ospedale
Novo Nordisk Health Care AG (for Extra-EU
support