1. Involvement of genetics in poor outcomes in anticoagulation therapy Farhad Kamali Thrombosis & Anticoagulation Research Group Institute of Cellular Medicine

2. Introduction n Warfarin a widely used oral anticoagulant for treatment and prophylaxis of thromboembolism (e.g. DVT) n Warfarin therapy decreases risk of stroke by 68% in patients with non-valvular atrial fibrillation n In the West about 1% of the population are on warfarin (about 2 million people in the US start taking warfarin each year)

3. Stroke prevention in AF Warfarin v placebo AFASAK-1 (671) SPAF (421) BAATAF (420) CAFA (378) SPINAF (571) EAFT (439) All trials (n=6) 0% -50% -100% 100% 50% Hart RG, et al. Ann Intern Med 2007; 146: 857-67

4. Dosing of warfarin is complex Narrow therapeutic index Small separation between dose-response curves for preventing emboli and excessive anticoagulation Nonlinear dose-response Small changes in dose may cause large changes anticoagulation response with a time lag Between-patient variability in dose requirement Wide range (50x) in dose requirement (2-112 mg/week) to achieve target INR of 2-3 (difficulty with attaining a stable control of anticoagulation during initiation of therapy)

5. Safety of warfarin 10-24 episodes of haemorrhage per 100 patients. Account for 3.6% of all drug- induced AEs; 3rd ranked drug in AEs Major risk is bleeding: frequent and severe; 1.2 – 7 major bleeding episodes per 100 patients; Relative risk of fatal extracranial bleeds 0 - 4.8% Responsible for 1 in 10 hospital admissions Schulman, N Engl J Med 349:675-683, 2003 Pirmohamed, British Med J 329:15-19, 2004 Kamali & Pirmohamed, Br J Clin Pharmacol 61: 746-751, 2006 Evans, Annals of Pharmaco 39:1161-1168, 2005 Wadelius, The Pharmacogenomics J, 5:262-270, 2005

6. Anticoagulation Status Determined by INR

7. Benefit: INR and Stroke Prevention

8. Risk: INR and intracranial hemorrhage

9. Induction therapy- Patients dosed on a trial and error basis Days/weeks dose (INR<2.0) or dose (INR>3.0) Fixed dose Maintenance dose

10. Frequency of major bleeds following initiation of warfarin dosing Landefeld, Am J Med 87:144-152, 1989

11. O R O O VII VIIa X Xa II IIa Thrombin (carboxylated) Prothrombin Vit K (epoxide) R O O Vit K (quinone) warfarin Vit K reductase _ warfarin _ Vit KH 2 OH R Carboxylase Vit K reductase

12. Factors Contributing to Inter-Individual variability in dose requirement  Disease  Drug interactions  Age -ve correlation between age and liver volume +ve correlation between liver volume and dose Wynne, et al. Br J Clin Pharmacol (1995) 40: 203-207

13. Factors contributing to inter-individual variability in dose requirement  Dietary vitamin K Changes in dietary vitamin K affect anticoagulation response Khan et al. BJH (2004) 124, 348-354 Sconce et al. Thromb Haemost (2005) 93, 872-875 -responsible for inter-individual variability in dose requirement?  Disease  Drug interactions  Age

14. Factors contributing to inter-individual variability  Disease  Drug interactions  Age  Dietary Vitamin K  Genetics

15. Warfarin chemical structure O O CH 3 R Vitamin K (natural vitamin) ONa O O CHCH 2 COH 2 C6H5C6H5 Warfarin (vitamin K antagonist) Warfarin is a coumarin derivative

16. CHCH 2 COH 2 C6H5C6H5 ONa O O Warfarin [(R)- & (S)-enantiomers)] CYP2C9*1 CYP2C9*2 (12% activity) CYP2C9*3 (5% activity) Furuya, et al. Pharmacogenetics (1995) 5: 389-392 Steward, et al. Pharmacogenetics (1997) 7: 361-367 Warfarin a 50:50 racemic mixture of R & S enantiomers

17. CYP2C9 Polymorphisms CYP2C9 *1/*1 (wild-type) CYP2C9 *1/*2 CYP2C9 *2/*2 CYP2C9 *2/*3 CYP2C9 *1/*3 CYP2C9 *3/*3 Decreasing enzyme activity

18. Genetic polymorphism for VKORC1 Several non-coding polymorphisms for VKOR influence coumarin dose requirements. D'Andrea et al. Blood. (2005) 105:645-649. Bodin et al. Blood. (2005) 106:135-40.

19. VKORC1 Polymorphisms GGGAAADecreasing dose requirement -1639G>A

20. Distribution of warfarin dose by CYP2C9 and VKORC1 genotype Sconce et al. Blood. 2005; 106: 2329-2333.

21. Estimated warfarin daily dose requirements (mg) (95% confidence interval) based on patient age, genotype and height  Predicted dose will vary by varying height and VKOR genotype. Individuals with VKOR AA genotype will require lower doses of warfarin than those with AG or GG genotypes. AA 1700.75 (0.50-1.05) 1.08 (0.83-1.36) 1.47 (1.23-1.74) 1.93 (1.68-2.20) 2.45 (2.15-2.76) 80 AA 1701.29 (0.97-1.66) 1.72 (1.42-2.05) 2.21 (1.94-2.51) 2.76 (2.49-3.05) 3.38 (3.06-3.71) 60 AA 1701.99 (1.55-2.49) 2.52 (2.09-2.98) 3.10 (2.69-3.54) 3.75 (3.33-4.19) 4.45 (3.97-4.96) 40 AA 170 2.84 (2.20-3.56) 3.46 (2.82-4.16) 4.14 (3.49-4.84) 4.88 (4.20-5.61) 5.68 (4.93-6.49) 20 VKOR genotype  Height  (cm) *2*3 (*3*3) *2*2*1*3*1*2*1*1Age (years)

22. IWPC algorithm Comparisons of Clinical and Pharmacogenetic algorithms based on genotype and use of amiodarone Genotype can markedly change the recommended dose from more than 45 mg per week to less than 10 mg per week when all other factors are the same. (NEJM, 2009; 19; 360(8):753-64.)

23. Genotype-guided dosing: translation of research data to practice Can pharmacogenetic-guided algorithms improve the accuracy of warfarin dosing during the initiation phase and reduce the incidence of warfarin-related adverse events, i.e., unintentional bleeding (over-dosing) and embolisms (under-dosing)?

24. Prospective pharmacogenetic-guided dosing studies COAG study across USA- funded by NIHLB involving patients starting warfarin -Primary end point: %TIR in the first 1 month of therapy ‘Genotype-guided warfarin doing was no better than a clinical algorithm’ EU-PACT study across Europe- funded by EC-FP7 programme involving patients for each of warfarin, acenocoumarol and phenprocoumon -Primary end point : %TIR in the first 3 months of therapy ‘Genotype-guided warfarin dosing was superior to fixed-dose regimens’ Pirmohammed et al. NEJM, 2013, 12;369(24):2294-303 Kimmel SE et al. NEJM, 2013, 12;369(24):2283-93

25. Further analyses of the EU-PACT and COAG trial data  Analysis of the EU-PACT data-  Analysis of the EU-PACT data- genotype-guided dosing caused greatest improvement in %TTR compared to the control arm in individuals with two or more CYP2C9 / VKORC1 variants (11.05% difference in TTR; P<0.009)  EU-PACT dosing-  EU-PACT dosing- more accurate in predicting the maintenance dose to within 1mg/day than COAG for both the dose initiation (62% vs 53%) and the dose revision algorithms (80% vs 62%) Unpublished data

26. PK/PD pharmacometric modelling patients homozygous with CYP2C9 variants benefitted the most from genotype-guided dosing, consistent with the EU-PACT findings. Unpublished data

27. Conclusions Current warfarin fixed dose regimens are inadequate Genetic polymorphisms in CYP2C9 and VKORC1, age, body size influence warfarin dose requirement Genotype-guided dosing regimens for initiation of warfarin therapy -more ‘individualised’ warfarin therapy Improved safety, reduced monitoring/management and better patient satisfaction Improved safety, reduced monitoring/management and better patient satisfaction

28. Acknowledgements Colleagues: Hilary Wynne Judith Coulson Maggie Fearby Jo Wincup Jill Henderson Liz Sconce Ellen Hatch Tayyaba Khan Peter Avery Patrick Kesteven John Hanley Peter Wood Sponsors: Baxter Healthcare