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Genetics in Coagulation Disorders: Venous Thromboembolism (VTE)
Roche Symposium Molecular Diagnostics Forum St Peter Zurich, March 3+4, 2016 PD Dr Lars M. Asmis Zürich
Genetics in VTE
Conflict of Interest Statement In the last 5 years I have received honoraria for presentations
and/or advisory boards or research support from the following companies: Axon Lab, Bayer, CSL Bering, Dade Behring/Siemens, Glaxo Smith Kline, Pfizer, Roche and Sanofi Aventis
I was free to chose the content of this presentation I follow the guidelines of the swiss academy of medical sciences
(www.samw.ch) and the foederatio medicorum helveticorum (www.FMH.ch)
Genetics in VTE
Overview
1. Introduction to VTE & coagulation
2. Hypothesis
3. Three case descriptions
4. Summary & Conclusions
5. Acknowledgements
6. Discussion
Thrombosis and embolism may have similar causes and treatments thromboembolism (TE)
vascular bed specificty gross simplification: venous & arterial TE
multimodal disease risk factors - acquired RF - congenital RF (thrombophilia)
1. Introduction: venous thromboembolism
>50% (GTH2016)
prevalent disease: 1/1000 year (incidence 100/100’000 and year) modified by acquired and congenital risk factors
disease burden: acute event postthrombotic syndrome mortality: case fatality rate 1-15% recurrent VTE
treatment: anticoagulants vitamin K antagonists direct oral anticoagulants
- efficiency: 96-98%, - safety: 1-4% major bleeds (≈ every 6th MB is fatal)
1. Introduction: venous thromboembolism (VTE)
1. Introduction: risk factors
relative risk RR cohort studies: exposed to non-exposed/controls RR = 1 no increase in RR RR = 1.1 10% increased risk RR = 2.0 100% increased risk, 2x
absolute risk AR incidence, prevalence, frequency number of cases/100’000 and year
balance between activation and inhibition
too much activation too little inhibtion
4 (5) principal inhibitors factor V Leiden TFPI deficiency prothrombin G20210A antithrombin (III) deficency (FVIII) protein C deficiency (FXI) protein S deficiency (antithrombin I “deficiency”)
1. Introduction: coagulation
1. Introduction: coagulation
FX
FVa
Fibrinogen Fibrins Fibrini
Prothrombin Thrombin (FIIa)
FXIII
HK
FVII
Tissue factor (FIII)
FIX
FVIIIa
FXI
FXII
PK
PL
PL
Ca++
Ca++
AT
AT
AT
AT
TFPI
Inhibitors
activated Protein C (aPC) Protein S (PS= cofaktor) Antithrombin (AT) Tissue Factor Pathway Inh. (TFPI)
aPC
aPC
PS
PS
1. VTE recurrence – J.A. Heit
Heit JA. Am J Hematol 2012; 87: S63-67
If one anticoagulates all patients with idiopathic VTE lifelong, then after 10 years 30% of patients (who would have recurred) are «protected» BUT the other 70% were anticoagulated (bleeding risk) «without benefit»
during the first 3-6 months of anticoagulation following the acute VTE event, the risk of recurrent VTE is 2-4%
in the 12 months after stopping anticoagulant therapy the recurrence risk is approximately 10% (5-15)
in the next years the risk remains elevated: approximately 5% (2-8%)
much of the disease burden is due to recurrence
… one can think of a scenario in which genetic testing is used to identify those at high risk of recurrence and to treat those longer or at higher dosages with anticoagulants
1. Genetic testing – P.H. Reitsma
Reitsma PH. Hämostaseologie 2015; 1: 1-5.
Gene sequencing of patients/kindreds can identify mutations that are associated with a high risk of VTE recurrence.
Ideally these mutations predict VTE recurrence in in symptomatic patients 1st VTE in asymptomatic individuals
2. Hypothesis
3.1 Antithrombin (AT-3) Deficiency
Antithrombin (AT3) 464 aa serine protease inhibitor «serpin» binds to active center IIa, Xa (IXa, XIa, XIIa) RR 5-10x
Congenital Acquired
heparin ttt necrosis nephrotic syndrome loss via urine anasarca loss via ascites
SERPINC1 chromosome 1 1 q23-25.1 13.4 kB – 7 exons autosomal dominant homozyg = lethal 0.02% of healthy ind 1% pts with 1st VTE
1.) JBC 1998; 273: 7478-87. 2.) Huntington JA. JBC 2000; 275:15377-83. 3.)
FIIa or FXa AT-FIIi or AT-FXi
+
UFH or LMWH or PS
PS
3.1 Antithrombin - mechanism of action
Patient with a idiopathic1/provoked 2pulmonary embolism 2007 Sy: dyspnea on exertion, and shortness of breath RF: prolonged febrile illness
Hospital A1 (2007): AT-3 deficiency: ATact: 72% (ref range 80-120%) long term oral anticoagulation (LT oAc)
Hospital B2 (5/2015): ATag: 79% (80-120%); ATact: 82% (80-120%) “wurde (extern) übereifrig ein AT Mangel diagnostiziert” hiermit kann die Dg AT Mangel ausgeschlossen werden stop LT oAC
ZPTH “third” opinion
3.1. Antithrombin Deficiency „excluded“
Antithrombin:
pPE, LT oAC
Antithrombin:
ATact 77% (anti FXa: 70-130) ATact 69% (anti FIIa: 70-130) ATag 87% (70-130%)
Antithrombin: SERPINC1 c.[391C>T]; [=] p.Leu131Phe
3.2 Protein C Deficiency
Protein C (PrC) 461 aa, 62 kDa serine protease 3-5ug/ml t½ 6-8h RR 3-6x
Congenital Acquired
vitamin K depedent synthesis (liver) childhood 40% pregancy by 20%
PROC chromosome 2 2 q13-14 10 kB – 9 exons autosomal dominant homozygous= cave purpura fulminans 0.3% of healthy ind 3% pts with 1st VTE
1st description: J YStdenflo 1978; C as in 3rd protein eluted by DEAE sepahrose
3.2 Protein C - mechanism of action
TM
TM thrombomodulin, PC protein C, PS protein S, FVa: activated, FVi: inactivated
FIIa = Thrombin procoagulant
FIIa = Thrombin anticoagulant PS
PC FVa FVIIIa
FVi FVIIIi
Protein C
pTVT, 42y iLE, 43y
pTVT, 43y (BMI)
Rec PE, LT oA
«VTE»
Protein C:
PrCag 48% (65-140) PrCakt 43% (65-140) PrCag 46%
PrCakt 35%
PrCakt 72% PrCakt 79%
? ? ? ? ?
Protein C: PROC: c.[989T>C];[=], p.Leu330Pro
Protein C: conclusions
General
inhibitor deficiency: likely high risk thrombophilia
frequency: 0.3% of general population
Our kindred
PROC: c.[989T>C];[=], p.Leu330Pro is a known mutation, however due to insufficient evidence it is not (yet) officially OMIM registered
autosomal dominant
in our kindred probably type I deficiency
family history suggests that it is predictive
3.3 Protein S Deficiency
Protein S (PrS) 672 aa, 71 kDa non-enzymatic cofactor vitamin K dependent 20-25ug/ml t½ 42h PrSfree : active PrStotal : inactive bound toC4bBP
Congenital Acquired
vitamin K depedent synthesis (liver) estrogen reduces PrS synthesis
PROS1 /(PROS2) chromosome 3 3 p11.1 (3q.11.2) 80 kB – 15 exons autosomal dominant homozygous = cave purpura fulminans 0.2% of healthy ind 1-13% pts with 1st VTE
1st description: diScipio 1977; S as in Seattle, where it was discovered
TM
TM thrombomodulin, PC protein C, PS protein S, FVa: activated, FVi: inactivated
FIIa = Thrombin procoagulant
FIIa = Thrombin anticoagulant PS
PC FVa FVIIIa
FVi FVIIIi
3.2 Protein S mechanism of action
Protein S:
30y, no VTE E2 pill
3 VTE PrS ↓
No VTE PrS nor
No VTE
2 VTE: 26y and 34y LT oAC; PrS ↓
Protein S:
PrS act 14% (60-130) PrS agfr 17% (60-130) Prs agto 32% (60-130)
Strong desire for children!
Protein S: PROS1 c.[1454A>G]; [=] p.Tyr485Cys
4. Summary
Caspers M. Thromb Haemost 2012; 108:247-57.
PrC PrC AT
4. Summary
Patient with AT deficiency with low penetrance (?) (type ?, Heparin binding site, novel mutation) clinical consequence: anticoagulation restarted daughters to be rechecked
Family with PrC deficiency with high penetrance (type: 1, one previous case report, unknown significance) clinical consequence anticoagulation in both brothers risk of coumarin skin necrosis (DOAC) 2 children with normal PrC, 4 to be checked
Family with PrS deficiency (type 2, known muttaion) clinical consequence medical VTE prophylaxis!!! in our asymptomatic pt Pregnancy risk+++
4. Summary
I hope to have introduced you to coagulation and VTE presented interesting cases
of antithrombin, PrC, PrS deficiency contributed to efforts towards better defining patients
at increased risk of recurrent VTE shown that these genetic tests have relevant clinical implications maybe motivated you to add to these experiences,
as there is still quite a way to go…
5. Acknowledgements
Many thanks to Prof Christine Mannhalter
University of Vienna for a brilliant talk at GTH 2016 and kind provision of key references
Dr Pierre Alain Menoud & Team Unilabs Molecular Genetics Lab in Lausanne for Gene sequencing, fruitful discussions and images
Barbara, Ilona, Jasmin and Sara the best team in the world!
Indirect inhibition mediated by
γ-COOHase Antithrombin Gla domain active center
Heparins, Heparinoids (iv, sc) Ximelagatran (po) Lepirudin (iv) Dabigatran (po)
Direct inhibition mediated by
active center
IIa IIa
Xa Xa
II, VII IX, X
Vitamin K Antagonists (po)
Rivaroxaban (po) Apixaban (po)
Suppl.: Mechanisms of action
PD Lars Asmis
Xa
IIa
Xa
Xa +
No reaction
No reaction
Suppl.: anti FXa activity
AntiXa
AntiXa activity
AntiXa
Suppl.: anti FXa activity
Cellular model