Belgian-Czech cooperation in the Brno-VWD study

Transcription

1 Inge Vangenechten I Vangenechten, P Smejkal, O Zapletal, F Bouddount, J Zavrelova, J Blatny, M Penka, JJ Michiels, A Gadisseur.

2 Aim of the study Characterisation of VWD in South Moravia (Czech Republic) Brno University Hospital Brno, population South Moravia, Czech Republic, population 1.1 million Antwerp University Hospital, Belgium

3 Brno University Hospital Patients with suspected or known VWD based on laboratory analysis (FVIII:c, VWF:Ag, VWF:RCo, Platelet aggregation) Grouped into families Inclusion of proband and affected sibling or parent Collection/shipping of samples Platelet Aggregometry (PFA) Antwerp University Hospital Full laboratory analysis (FVIII:c, VWF:Ag, VWF:RCo, VWF:CB, VWFpp, and VWF-FVIII binding where indicated) VWF multimers (cfr Budde, Hamburg) Molecular analysis

4 patients suspected of having VWD (proband): Type 3: VWF:Ag and VWF:RCo <5% Type 2: Decreased Ristocetin Induced Platelet Aggregometry RIPA (concentration mg/dl) and/or VWF:RCo/VWF:Ag < 0.7 (type 2A/2M) or unexplained thrombocytopenia where there is a suspicion of VWD and/or positive low concentration RIPA (0.8mg/ml) (type 2B) Or all patients with VWF:CB/VWF:Ag ratio < 0.7 (type 2A, Collagen type) Or FVIII:c/VWF:Ag <0.5 (type 2N) Type 1: VWF:Ag< 35% type 1 patients with VWF:Ag levels >35% can only be included with severe phenotype

5 205 patients in 95 families Median CI95 Range FVIII:c (%) VWF:Ag (%) VWF:RCo (%) VWF:CB (%) Current status Full laboratory analysis completed Multimeric analysis completed in all samples Molecular work completed in 34 families, rest partially Sheffield Algorithm for VWD analysis

6 Patients VWD diagnosis confirmed in 163 samples (79.5%), 89 families (93.7%) VWD awaiting confirmation in 42 samples (20.5%), 6 families (6.3%) Type Families (%) Samples (%) remarks Type 1 60 (63.2) 90 (55.2) 1 Type 1 Vicenza Type 3-6 (3.7) (all in type 1 families) Type 2A 17 (17.9) 43 (26.4) 2A/IIA A/IIC 1 3 2A/IIE Type 2B 4 (4.2) 9 (5.5) Type 2M 8 (8.4) 15 (9.2) Type 2N - - Carriers in 6 families, 8 pts

7 Type 2M Type 2M Type 2B Type 2B Type 2A Type 1 Type 2A Type 1 Type 3

8 Mutations (preliminary data) 25 mutations in 51 families Exon

9 p.n166i / c.497a>t p.r416w / c.1246c>t (new) p.p812rfs(x31) / c.2435delc p.r854q / c.2561g>a p.r924q / c.2771g>a p.s979n / c. 2936G>A p.c1130g / c.3388t>g p.w1144g / c.3430t>g p.y1146c / c.3437a>g p.r1205h / c.3614g>a p.p1266l / c.3797c>t p.v1279i / c.3835g>a p.r1306w / c.3916c>t p.r1308c / c.3922c>t p.r1341q / c.4022g>a p.r1341w / c.4021c>t p.e1359k / c.4075g>a p.g1415d / c.4244g>a p.g1579r / c.4735g>a p.y1584c / c.4751a>g p.g1609r / c.4825g>a p.v1760i / c.5278g>a p.r1830c / c.5488c>t (new) p.p2063s / c.6187c/t p.s2179f / c.6536c>t * D1 D2 D D3 A1 A2 A3 D4 B C1 C2 CK exon SP Propeptide mature VWF

10 Mutation Fam Pts Mutation Fam Pts p.p812rfs(x31) / c.2435delc 9 14 p.s979n / c. 2936G>A 1 3 p.g1579r / c.4735g>a 5 17 p.c1130g / c.3388t>g 1 1 p.r854q / c.2561g>a (2N) 4 6 p.y1146c / c.3437a>g 1 2 p.y1584c / c.4751a>g 4 8 p.r1205h / c.3614g>a 1 1 p.n166i / c.497a>t 3 4 p.r1306w / c.3916c>t 1 3 p.w1144g / c.3430t>g 3 6 p.r1308c / c.3922c>t 1 2 p.r924q / c.2771g>a (2N) 2 2 p.r1341q / c.4022g>a 1 1 p.p1266l / c.3797c>t 2* 2* p.r1341w / c.4021c>t 1 3 p.v1279i / c.3835g>a 2* 2* p.g1415d / c.4244g>a 1 2 p.e1359k / c.4075g>a 2 5 p.g1609r / c.4825g>a 1 1 p.r416w / c.1246c>t (new) 1 2 p.v1760i / c.5278g>a 1 1 p.p2063s / c.6187c/t 1 1 p.s2179f / c.6536c>t 1 2

11 p.p812rfs(x31) / c.2435delc (type 1, ex 18)

12 New mutations Exon 11: p.r416w / c.1246c>t VWD type 2A/IIE Exon 32: p.r1830c / c.5488c>t VWD type 1 Awaiting expression studies Linkage (preliminary data) Type 1 34/96 pts (35.4%) Type 2 45/67 pts (67.2%) Type 2A 29/43 (67.4%) Type 2B 9/9 (100.0%) Type 2M 7/15 (46.7%)

13 VWFpp VWD type VWFpp/VWF:Ag mean VWFpp/VWF:Ag CI A IIA IIC IIE B M p<0.001

14 VWFpp/VWF:Ag Samples (ranged according to family number)

15 VWFpp/VWF:Ag (mean) Mutation Type Family A/IIE (4.87) p.r1205h / c.3614g>a 1 Vicenza M (3.61) p.y1146c / c.3437a>g 2A/IIE (3.68) p.s2179f / c.6536c>t 2M (3.68) p.s2179f / c.6536c>t 2M (2.48) p.w1144g / c.3430t>g 2A/IIE (2.48) p.w1144g / c.3430t>g 2A/IIE (2.63) p.r1306w / c.3916c>t 2B A/IIE (1.42) p.y1584c / c.4751a>g 1 9

16 Families with Y1584C mutation Family (Y1584C) VWFpp/VWF:Ag 1 VWFpp/VWF:Ag 2 VWFpp/VWF:Ag ( A) 3.0 (A) (A) 0.88 (A) 1.54 (O) (A) 1.23 (A) (O) 0.90 (A) Presence of second mutation is predicted in families 9 and 95

17 Conclusion: VWFpp/VWF:Ag ratio = tool in the diagnostic work-up of VWD patients ð differentiation between type 1 and type 2 VWD ð no differentiation type 2 subtypes.

18 FVIIIc ð no type 2NvWD patients VWD type FVIIIc/VWF:Ag mean FVIIIc/VWF:Ag CI95 All types

19 FVIIIc/vWF:Ag 4,00 3,50 3,00 2,50 2,00 1,50 1,00 0,50 0,00 Samples (ranged according to family number)

20 FVIIIc/VWF:Ag Families (%) Samples (%) Type vwd < (13.7) 19 (9.3) Type 2A (II), 2M (3 pts type 1; p.p812rfs + p.r854q) (57.9) 146 (71.2) > (28.4) 40 (19.5) Type 1

21 FVIIIc/VWF:Ag <0.5 mutation Fam pts Type vwd p.g1579r / c.4735g>a 3 4 2A p.c1130r / c.3388t>c (2A) p.e1359k / c.405g>a 1 1 2M p.g1609r / c.4825g>a 1 1 2A p.r854q / c.2561g>a + p.p812rfs/c.2435delc 2 3 2N + 1

22 FVIIIc/VWF:Ag >1.5 mutations Fam pts Type vwd p.p812rfs/c.2435delc p.n166i / c.497a>t p.y1584c / c.4751a>g p.s979n / c.2936g>a 1 1 2A p.c1130g / c.3388t>g (2A) p.y1146c / c.3437a>g p.p1266l / c.3797c>t + p.v1279i / c.3835g>a /2M p.c996s / c.2987g>c p.d47v / c.140a>t (new) 1 2 (1) p.r416w / c.1246c>t (new) 1 2 (2A)

23 Conclusion: majority of VWD patients FVIII:c/VWF:Ag ratio ±1/3: very low / high ratio. FVIII:c/VWF:Ag ratio <0.5 ð not always type 2N VWD ð present in other type 2 mutations. High ratios are mostly seen in type 1 VWD.

24 We learned a lot from this study How to perform a study vwfpp/vwf:ag FVIIIc/vWF:Ag Brno-vWD set-up ð multicentric for the next study (B-Will study)

25 Antwerp, Belgium A Gadisseur I Vangenechten F Bouddount JJ Michiels Brno, Czech Republic P Smejkal O Zapletal J Zavrelova J Blatny M Penka Supported by an unrestricted grant from