# Presentation: 369994

## New Insights on Warfarin: How CYP 2C9 & VKORC1 Information May Improve Benefit-Risk Ratio 

- Brian F. Gage, MD, MSc
- Associate Professor of Medicine,
- Washington University in St. Louis
- Blood Thinner Service Medical Director,
- Barnes-Jewish Hospital

## Overview

- Cytochrome P450 (CYP) 2C9
- Vitamin K Epoxide Reductase, Complex 1 (VKORC1)
- Derivation of pharmacogenetics-based warfarin dosing
- Validation of pharmacogenetics-based warfarin dosing

## Slide 3

## Slide 4

## Effect of CYP2C9*2 on Warfarin Dose

- S. Sanderson et al.
- Genet Med. 2005 Feb;7

## Effect of CYP2C9*3 on Warfarin Dose

- S. Sanderson et al.
- Genet Med. 2005 Feb;7

## Vitamin K epoxide reductase, complex 1 (VKORC1)

**Mutations in *****VKORC1***** Cause Warfarin Resistance and Multiple Coagulation Factor Deficiency Type 2**
***VKORC1***** synthesizes vitamin K epoxide reductase (VKOR), which resides in the endoplasmic reticulum of the hepatocyte and other cells**
**VKOR is inhibited by warfarin, especially S-warfarin**
**S-warfarin is metabolized by CYP2C9**
**VKOR activity is required for post-translational modification (****γ****-glutamyl carboxylation) of Glu residues on clotting factors II, VII, IX, X and proteins C, S, and Z**
**VKORC1 may or may not be part of a complex**

## VKORC1 SNPs and Warfarin Dose 

- Hypothesis: informative SNPs in VKORC1 correlate with warfarin dose
- We collaborated with Mark Rieder and Allan Rettie at University of Washington
- To sequence this gene in archived DNA (N = 47)  from CEPH families and from the Coriell depository
- To correlate informative SNPs (inferred haplotypes) in 186 patients
- To correlate the 4 tagSNPs and inferred haplotypes in a larger cross-sectional study

## Effect of VKORC1 Inferred Haplotype on Warfarin Dose

- Rieder MJ, Reiner AP, Gage BF et al.
- Effect of VKORC1 haplotypes on transcriptional regulation and warfarin dose.
*N Engl J Med. *Jun 2005.

**3673 = rs9923231 aka -1639, a promoter SNP (M. Wadelius;** **H. Yuan; E. Sconce)**
**6484 = rs9934438 aka C1173T, (M. Wadelius; H. Yuan; L. Bodin; D’Andrea); **
**6853 = rs17886369**
**7566 = rs2359612**

## VKORC1 Distributions Stratified by Race

- Group B haplotypes had larger doses, more mRNA transcript for VKORC1, and were more frequent in white and African-American pts.

- Group A haplotypes had smaller doses and were more frequent in Asian pts.

## Pyrosequencing in Dr. Eby’s and McLeod’s Labs: Thanks to Sharon, Christi, Rhonda

**Pyrogram of VKORC1 6853 heterozygote subject.  **
**The sequence for nucleotides is:   G/C G A G C G.**

**Frequency of VKORC1-6853C allele: 37% in white and 24% in black pts.**

## Effect of VKORC1 Haplotype on Warfarin Dose

**Effect of *****VKORC1***** Haplotype on Warfarin Dose**
- Stratified by VKORC1 haplotype and CYP2C9 status
- Primary cohort: UW (N=185);
- Replication cohort: Wash U (N=368).
- All participants were Caucasian.

## D’Andrea et al. and VKORC1, Blood

- Genotyped 147 warfarin-treated patients for common SNPs in *VKORC1*
- Found that 1173 did not affect mRNA processing.

| VKORC1
1173
Genotype | Warfarin Dose (mg/d) |
| --- | --- |
| CC (37%) | 6.2 |
| CT (47%) | 4.8 |
| TT (16%) | 3.5 |

**P **_**<**_** 0.002**

## M. Wadelius et al. The Pharmacogenomics Journal (2005)

- Genotyped 200 warfarin-treated patients for common SNPs in *VKORC1*.
- found *VKORC1* C1173T, which explained 29% of the variability in warfarin dose
- Combined *VKORC1* SNP, the CYP2C9*2 and CYP2C9*3 SNPs, and clinical factors, to derive a regression model that accounted for 56% of the variability in the warfarin dose.

## M. Wadelius et al. The Pharmacogenomics Journal (‘05)

- rs2359612

## L. Bodin et al. Measured FVII & INRs in 222 Patients after 1 dose of Acenocoumarol: 

## E. Sconce et al. ‘05 Derived (N = 297) & Validated (N=38) a Dosing Algorithm using VKORC1, CYP2C9, Age, Height

- R2 = 54%-64%

## Development and Validation of a Warfarin Dosing Algorithm at Washington University

**Development and Validation of a Warfarin Dosing Algorithm at Washington University**

- Gage BF, Eby C, Johnson JA, Rieder MJ, Ridker PM...McLeod H. [ASH abstract]

## Warfarin Dosing Equation in Derivation Cohort, N= 900 

## Accuracy in Validation Cohort (N = 100)

- R2 = 26% for clinical model.

**Accuracy in Validation Cohort (N = 100)**

- (P<0.0001 vs. standard dose; P = 0.02 vs. clinical dose)

- R2 = 56% for pharmacogenetics model, but lower in African-Americans

- By reducing the dosing error from 1.79 to 1.31 mg, pharmacogenetics should be able to increase the time in range in month one by 2%-4% and should decrease adverse events in patients w/ usual genotypes

## Future Studies

- SNP discovery in targeted genes: APOE, calumenin, Factors II, VII, IX, X, γ-GCx
- Quantifying the relationship between new SNPs and warfarin dose
- VKORC1 SNPs specific to African-American populations.
- Factor VII  G-402A & G-401T
- Factor II 165Thr>Met
- APOE
- γ-GCx
- Prospective validation of a pharmacogenetics- dosing model
- Would benefit from a platform that could quickly and economically genotype individuals

## Conclusions

- The maintenance warfarin dose can be estimated from clinical and pharmacogenetic factors that can be obtained at the time of warfarin initiation
- _>_ 50% of the variability in the warfarin dose can be predicted from regression model using 2 genes: CYP2C9 and VKORC1
- Although no dosing algorithm has been prospectively validated, the relationship between SNPs in these genes and the therapeutic warfarin dose is biologically and statistically compelling
- For patients initiating warfarin therapy, we estimate a 2%-4% increase in time in the therapeutic INR range in month one with less benefit thereafter
- After month 1, pharmacogenetic knowledge may allow for more cautious dose escalation patients with CYP2C9*2 or CYP2C9*3

## Methods for this Prospective, Open-labeled Study 

**Patients are scheduled for elective orthopedic surgery**
**Screen for exclusion criteria, obtain consent for clinical trial, gather sample for genotyping, and obtain non genetic data**
**Determine CYP2C9 genotype (in commercial lab run by Mark Linder) and estimate pharmacogenetic dose**
**5- or 10-mg dose of warfarin taken the day before surgery**
** **
**      ****To OR**
**Inpatient (daily INR) and outpatient pharmacogenetics-based warfarin doses (2-3x/week INR) to be adjusted and monitored by anticoagulation service for 6 weeks**

- Time

- D. Voora et al. **Thromb Haemost** 2005

## Results

## Time Until INR > 4 or Major Bleed