Identifying patients who may require warfarin dosing adjustments

(2,3) including:

- Patients who have previously been prescribed warfarin

  and have required multiple dosing adjustments to

  maintain the INR in the target range

- Patients with a history of thrombosis or bleeding when

  previously taking warfarin

- Patients being started on a first prescription for warfarin

Warfarin is a coumarin-based drug commonly utilized in

anticoagulation therapy to prevent thrombosis due to inherited

and acquired hemostatic disorders. The drug is also used in a

number of other medical conditions and treatments including

atrial fibrillation and hip replacement surgery. Warfarin acts by

interfacing with the metabolism of vitamin K, which is necessary

for production of key coagulation factors. Warfarin inhibits vitamin

K recycling by blocking its metabolism at the vitamin K-epoxide

intermediate and thereby decreasing the amount of available

vitamin K. Warfarin has a narrow therapeutic window;

undermedicating increases the risk for thrombosis and

overmedicating increases the risk for cerebrovascular

accidents. Warfarin therapy has one of the highest rates

of severe adverse drug reactions.

Warfarin is dosed using nongenetic factors including gender,

weight, and age, and is monitored by coagulation testing in

order to maintain the international normalized ratio (INR)

within specific limits. However, warfarin metabolism is highly

variable and dependent upon genetic factors. Polymorphisms

within 2 genes are known to affect the metabolism of warfarin

and the dose needed to maintain the correct serum drug level

and degree of anticoagulation, as measured by the INR. The

cytochrome P450 2C9 gene (CYP2C9) encodes an enzyme

that metabolizes the more active isomer of warfarin (S-warfarin)

to inactive products. Polymorphisms in this gene decrease the

activity of the enzyme and may result in increases in serum

warfarin and overmedicating, resulting in increases in the INR

above the therapeutic target level. The second gene (VKORC1)

encodes vitamin K epoxide reductase complex subunit-1

(VKORC1), a small transmembrane protein of the endoplasmic

reticulum that is part of the vitamin K cycle and the target of

warfarin therapy.(1) VKORC1 is primarily transcribed in the liver,

although it is present in smaller amounts in the heart and

pancreas. Vitamin K epoxide, a by-product of the carboxylation

of blood coagulation factors, is reduced to vitamin K by VKORC1.

A polymorphism within the promoter of VKORC1 decreases

expression of the gene, decreasing the availability of vitamin K.

Thus, a reduced warfarin dose is needed to compensate for

the effects of the VKORC1 promoter polymorphism in order

to maintain the target INR.

CYP2C9

CYP2C9 metabolizes a wide variety of drugs including warfarin

and many nonsteroidal anti-inflammatory drugs. It is also partially

responsible for metabolizing other drugs such as fluoxetine,

fluvastatin, oral hypoglycemic drugs, and phenytoin.

A number of specific polymorphisms have been found in the

CYP2C9 gene that result in enzymatic deficiencies.

The following information outlines the relationship between the

polymorphisms detected in this assay and the effect on the

activity of the enzyme encoded by that allele:

CYP2C9        Nucleotide        Effect on Enzyme

Allele            Change             Metabolism

*1                         None (wild type)   Extensive metabolizer (normal)

*2                         430C->T                   Reduced activity

*3                         1075A->C                Minimal activity

*4                         1076T->C                Reduced activity

*5                         1080C->G                Reduced activity

*6                         818delA                   No activity

Dosing of warfarin, which is metabolized through CYP2C9, may

require adjustment for the individual patient. Patients who are poor

metabolizers (reduced activity) may benefit by dose reductions

or by being switched to other comparable drugs that are not

metabolized primarily by CYP2C9. The following is a partial

listing of drugs known to affect CYP2C9 activity as of the date

of this report.

Drugs that undergo metabolism by CYP2C9:

       - Angiotensin II blockers: irbesartan, losartan

       - Anticoagulants: warfarin (more active S-isomer)

       - Antidepressants: amitriptyline (minor), fluoxetine

       - Nonsteroidal anti-inflammatory drugs (NSAIDS):

             celecoxib, diclofenac, ibuprofen, naproxen, piroxicam

       - Oral hypoglycemic agents: glipizide, glimepiride, glyburide/

             glibenclamide, rosiglitazone (minor), tolbutamide

       - Miscellaneous drugs: fluvastatin, phenytoin, sulfamethoxazole,

             tamoxifen, torsemide

Coadministration of these drugs may decrease the rate of elimination of

other drugs metabolized by CYP2C9

Drugs known to increase CYP2C9 activity:

-        Rifampin, secobarbital

Coadministration of these drugs increase the synthesis

of CYP2C9, resulting in increased CYP2C9 activity

and metabolism of warfarin. A dose increase may

be needed to maintain the INR in the target range.

 Drugs known to decrease CYP2C9 activity:

      - Amiodarone, fluconazole, fluvastatin, isoniazid,

        lovastatin, ticlopidine

Coadministration of these drugs may decrease the rate of metabolism of

CYP2C9-metabolized drugs, including warfarin, increasing

the possibility of toxicity

VKORC1

The -1639 promoter polymorphism is located in the second

nucleotide of an E-Box (CANNTG) and its presence disrupts

the consensus sequence, reducing promoter activity. In vitro

experiments show a 44% higher transcription level of the G

versus the A allele.(1)

VKORC1 allele    Nucleotide Change    Effect of Enzyme Metabolism

   -1639 G->A                -1639 G->A                     Reduced activity

An interpretive report will be provided.

An interpretive report will be provided.

The normal genotype (wild-type) for CYP2C9 is termed CYP2C9*1.

Other genotypes that lead to inactive or reduced activity alleles

include CYP2C9*2, CYP2C9*3, CYP2C9*4, CYP2C9*5, and

CYP2C9*6. An individual who has homozygous wild-type,

CYP2C9*1/CYP2C9*1, is considered an extensive metabolizer.

The normal genotype for VKORC1 is -1639G. A polymorphism

at -1639A reduces VKORC1 expression. The VKORC1 GA or

AA genotype leads to a significant decrease in mRNA

expression in the liver compared with individuals with the GG

genotype.

Individuals who have polymorphisms in both the VKORC1

promoter (GA or AA) and also in CYP2C9 should receive a reduced dose

of warfarin to reduce and maintain the INR in the target range;

dosing adjustments are required when polymorphisms in both

genes are present.

Drug-drug interactions and drug-metabolite inhibition must be

considered when dealing with heterozygous individuals.

Drug-metabolite inhibition can occur, resulting in inhibition of

residual functional CYP2C9 or VKORC1 catalytic activity. A

clinical pharmacologist should be consulted for assessing

the potential for drug interactions.

Patients may also develop toxicity problems if liver and kidney

function are impaired.

This test does not detect polymorphisms other than those listed.

Mutations in the primer-binding regions can affect testing and,

ultimately, the genotyping results. Warfarin metabolism may

be inhibited through drug-drug interactions, including amiodarone

and statins.

Genotyping patients using DNA obtained from leukocytes may

not provide useful information in patients who have had a bone

marrow or liver transplant or a recent transfusion. To obtain an

accurate genotype on a bone marrow transplant recipient, buccal

cells should be provided. To obtain an accurate genotype for a

patient who has received a donor liver, testing must be done on

donor cells. If the patient has been transfused, wait 4 to 6 weeks until

transfused cells have left the circulation.

1.   Oldenburg J, Bevens C, Muller C, Watzka M:  Vitamin K

      epoxide reductase complex subunit I (VKORC1):  

      the key protein of the vitamin K cycle. Antioxid Redox Signal

      2006;8(3-4):347-353

2.   Yuan H, Chen J, Lee M, et al:  A novel functional VKORC1

      promoter polymorphism is associated with inter-individual

      and inter-ethnic differences in warfarin sensitivity. Hum Mol Genet

      2005;14:1745-1751

3.   Sconce E, Khan T, Wynne H, et al:  The impact of CYP2C9

      and VKORC1 genetic polymorphism and patient

      characteristics upon warfarin dose requirements

      proposal for a new dosing regimen. Blood 2005;106:2329-2333