Download Molecular and Clinical Markers for the Diagnosis and Treatment of

Molecular Genetics in the Von
Willebrand disease
Ghasem Rastegarlari
VON WILLEBRAND DISEASE
VWD General definitions
• The most frequent congenital bleeding
disorder caused by defects of VWF:
- quantitative = Types 1 & 3 VWD
- qualitative = Type 2 VWD
• Autosomal dominant/recessive pattern
• Women are more symptomatic
Von Willebrand disease
Genetic aspect
Human VWF Gene
Chromosome 12
1
p
q
7
14
VWF
(12p13.3)
178 kb, 52 exons
mRNA 8.7 kb
52
ENCODING REGIONS OF VWF
AND FUNCTIONAL DOMAINS
VWF gene in Chromosome 12
5’
bp 0
3’
1000
2000
1-23
H2N
3000
D2
5000
6000
7000
8000
Multimers
S-S
163
D1
4000
D’
FVIII
D3
Dimers
S-S
A1
A2
A3
Platelets Endothelial
cells
D4
9000
2813
C1 C2 CK COOH
B1
B2
B3
RGD
IIb3
ADHESION ACTIVITIES OF VWF
PLATELET
GPIb
Collagen
Heparin
Sulphatide
VWF:RCo
A1
A2
CC
CC
VWF:CP
A3
A3
VWF:CB
SUBENDOTHELIUM COLLAGEN
Single Platelets Adherent to Endothelial Monolayer
6
A.J. Reininger
Platelet Aggregate Adherent to Endothelial Monolayer
A.J. Reininger
7
MOLECULAR MARKERS
of Type 3 VWD
• Complete deficiency of VWF (VWF:Ag < 1)
• Autosomal recessive pattern of inheritance
• Rare (1- 5 per million) but severe disorder
•Caused by several defects: gene deletions,
frameshift-nonsense-missense-splite site
mutations, defects of mRNA expression
CLINICAL & MOLECULAR MARKERS
of Type 2 VWD
• Type 2A, 2B and 2M variants with decreased
platelet-dependent function
Autosomal dominant
• Can be caused mainly by missense mutations
(small deletions or frameshift mutations also)
CLINICAL & MOLECULAR MARKERS
of Type 2N VWD
• Variants
with markedly decreased
affinity for factor VIII (FVIII)
• Inherited by recessive patterns
•Caused by missense mutations
CLINICAL & MOLECULAR MARKERS
Type 1 VWD
• Inheritance (autosomal dominant)
•BUT: Factors which can modify VWF levels:
• Sex (females may exhibit greater variability)
• Age (VWF higher in older individuals)
• Exercise and stress (VWF increases)
• Blood Group O (lower VWF levels)
Von Willebrand disease
Genetic aspect
It is important to determine the causative defect of VWF
gene:
to prove phenotypic diagnosis or to make a definite
diagnosis of VWD when the phenotypic diagnosis is
uncertain,
Prenatal diagnosis
Direct sequencing of the VWF gene
Conclusion (1)
We have investigated 121 unrelated VWD patients
66 unrelated type 2 VWD patients
50 unrelated type 3 VWD B patients
The molecular defects have been found in 109 patients with a
detection rate of ~ 90%
Nineteen novel mutations (not previously reported, in the
International VWD mutation databases).
Conclusion (2)
Identified mutations in VWD patients allowed direct
carrier diagnosis and prenatal diagnosis
Mutation analysis is now routinely carried out and is
used as a first line method for carrier detection and will
be used for prenatal diagnosis.
All molecular analysis from the DNA extraction to
sequencing were done in our Iranian Comprehensive
Hemophilia Treatment Center