**4.1. Warfarin**

Warfarin is a widely used anticoagulant in the treatment and prevention of thrombosis. It was initially marketed as a pesticide against rats and mice and is still used for this purpose. It was approved for use as a medication in the early 1950s and is widely prescribed. Despite its common use, warfarin therapy can be associated with significant bleeding complications. Achieving a safe therapeutic response can be difficult because of warfarin's narrow thera‐ peutic index and great individual variability in the dose required, which is mostly a conse‐ quence of individual genetic variants. This fact is well known among clinicians and the wide range, from 1 mg/day to 20 mg/day, of warfarin maintenance doses are observed across the population. To maintain a therapeutic level of anti-thrombosis and to minimise the risk of bleeding complications, warfarin therapy requires intensive monitoring via the International Normalized Ratio (INR) to guide its dosing. The INR is used to monitor the effectiveness of warfarin and measures the pathway of blood coagulation. It is used to standardize the re‐ sults for a prothrombin time. INR is the ratio of a patient's prothrombin time to a control sample, raised to the power of the index value for the analytical system used.

Several factors increase the risk of over-anticoagulation: genetic polymorphisms affecting the metabolising enzymes, impaired liver function, drug interactions, congestive heart fail‐ ure, diarrhoea, fever, and diets rich in vitamin K [61] [62]. Nevertheless, genetic factors and drug interactions mostly account for the risk of over-anticoagulation. Warfarin metabolism involves primarily the cytochrome P450 (CYP) enzymes. Some loss-of-function CYP2C9 and vitamin K epoxide reductase complex subunit 1 (VKORC1) polymorphisms are known to be associated with decreased enzymatic activity and as a result, with an increased risk of hae‐ morhage. These are CYP2C9\*2 (Cysl44/Ile359), CYP2C9\*3 (Argl44/Leu359) and VKORC1 (-1639G>A) [63-65].

Warfarin-induced haemorrhage is an important complication of anticoagulation therapy. A review of many studies shows average yearly rates of warfarin-related bleeding as high as 0.8%, 4.9%, and 15%, for fatal, major and minor bleeding complications respectively [66].

Vitamin K is required by proteins C and S, together with clotting factors II, VII, IX, and X, to allow assembly of the procoagulant enzyme complexes necessary to generate fibrin. Warfar‐ in as an anticoagulant agent has the ability to interfere with the recycling of vitamin K in the liver. The pharmacologic effect of warfarin is mediated by the inhibition of vitamin K epox‐ ide reductase complex subunit 1 (EC 1.1.4.1) [67].

mines, which are selective for the myocardium; dihydropyridines which mostly affecting

A few studies describe some association; three SNPs in CACNA1C had significant associa‐ tions with treatment in a study of BP lowering with calcium channel blockers [59]; between CYP3A5\*3 and \*6 variants and verapamil treatment for BP and hypertension risk outcomes in blacks and Hispanics [60]; individuals that are homozygous for the T allele of NPPA T2238C had more favourable clinical outcomes when treated with a calcium channel blocker whereas C carriers responded better to a diuretic [32]. Beta Adrenergic Receptor 1 (BAR1) Ser49-Arg389 haplotype carriers had higher death rates than those with other haplotypes

Warfarin is a widely used anticoagulant in the treatment and prevention of thrombosis. It was initially marketed as a pesticide against rats and mice and is still used for this purpose. It was approved for use as a medication in the early 1950s and is widely prescribed. Despite its common use, warfarin therapy can be associated with significant bleeding complications. Achieving a safe therapeutic response can be difficult because of warfarin's narrow thera‐ peutic index and great individual variability in the dose required, which is mostly a conse‐ quence of individual genetic variants. This fact is well known among clinicians and the wide range, from 1 mg/day to 20 mg/day, of warfarin maintenance doses are observed across the population. To maintain a therapeutic level of anti-thrombosis and to minimise the risk of bleeding complications, warfarin therapy requires intensive monitoring via the International Normalized Ratio (INR) to guide its dosing. The INR is used to monitor the effectiveness of warfarin and measures the pathway of blood coagulation. It is used to standardize the re‐ sults for a prothrombin time. INR is the ratio of a patient's prothrombin time to a control

sample, raised to the power of the index value for the analytical system used.

Several factors increase the risk of over-anticoagulation: genetic polymorphisms affecting the metabolising enzymes, impaired liver function, drug interactions, congestive heart fail‐ ure, diarrhoea, fever, and diets rich in vitamin K [61] [62]. Nevertheless, genetic factors and drug interactions mostly account for the risk of over-anticoagulation. Warfarin metabolism involves primarily the cytochrome P450 (CYP) enzymes. Some loss-of-function CYP2C9 and vitamin K epoxide reductase complex subunit 1 (VKORC1) polymorphisms are known to be associated with decreased enzymatic activity and as a result, with an increased risk of hae‐ morhage. These are CYP2C9\*2 (Cysl44/Ile359), CYP2C9\*3 (Argl44/Leu359) and VKORC1

Warfarin-induced haemorrhage is an important complication of anticoagulation therapy. A review of many studies shows average yearly rates of warfarin-related bleeding as high as 0.8%, 4.9%, and 15%, for fatal, major and minor bleeding complications respectively [66].

smooth muscle and benzothiazepines with a broad range.

when treated with verapamil [15].

**4. Anticoagulants**

(-1639G>A) [63-65].

**4.1. Warfarin**

82 Drug Discovery

Warfarin consists of (R)- and (S)-warfarin enantiomers. (R)- and (S)-warfarins differ in their relative plasma concentrations, in their antithrombotic potency and in the specific isoen‐ zymes responsible for their metabolism. (S)-warfarin has a 3 to 5 times greater anticoagulant effect than the (R)-enantiomer and accounts for 60% to 70% of warfarin's overall anticoagu‐ lant activity. (S)-warfarin is metabolised almost exclusively by CYP2C9 [68-70].

The activity of the CYP2C9 enzyme has a significant impact on the clearance of (S)-warfarin and as a consequence on anticoagulant effect. In the presence of genetic variations where the activity of CYP2C9 is reduced, clearance of (S)-warfarin is also reduced. Activity of CYP2C9 between individuals can vary by more than 20-fold. (R)-warfarin is metabilised by multiple different CYP enzymes [71].

While several single-nucleotide polymorphisms of CYP2C9 have been reported, the CYP2C9\*2 (Cysl44/Ile359) and CYP2C9\*3 (Argl44/Leu359) polymorphisms have been identi‐ fied as clinically relevant [72]. Both of these variants are associated with decreased enzymat‐ ic activity [24, 73-78].

Homozygous CYP2C9\*3 variant genotypes have only 5% to 10% metabolic efficiency com‐ pared to the wild-type genotype. As a result, compared to wild-type CYP2C9\*1\*1 controls, enzyme activity and the median maintenance warfarin dose for CYP2C9\*3\*1 heterozygotes was reduced by 40%, and by approximately 90% for CYP2C9\*3\*3 homozygotes [72-74].

Furuya [79] and Steward [75] showed that the CYP2C9\*2 variant is also associated with re‐ duced warfarin elimination. Heterozygotes demonstrate 40% and homozygotes 15% of the wild-type enzyme activity, causing dose adjustment for heterozygote CYP2C9\*2 individuals down to 20% less than the standard dose.

Margaglione [76] has also demonstrated bleeding rates as high as 27.9 per 100 patient-years in carriers of CYP variants. In this study, findings were adjusted for other common variables associated with increased bleeding risk, such as increased age, drug interactions and abnor‐ mal liver function.

Several studies of the \*2 and \*3 CYP2C9 polymorphisms consistently show that patients with at least one CYP2C9 allele polymorphism have reduced warfarin requirements [76, 80-84]. Freeman [85] reported reduced warfarin weekly dosages for carriers of CYP2C9\*2 or CYP2C9\*3 alleles compared with patients who were homozygous for the wild-type allele (0.307 mg/kg/wk and 0.397 mg/kg/wk, respectively). Taube [83] compared warfarin mainte‐ nance dosages in 683 patients carrying different CYP2C9 genotypes. Mean warfarin mainte‐ nance dosages were 86% in patients with CYP2C9\*1\*2, 79% in patients with CYP2C9\*1\*3, 82% in compound heterozygotes CYP2C9\*2/\*3, and 61% in patients homozygous for CYP2C9\*2. Furthermore, Aithal [80] warns that even when warfarin dosages are decreased, carriers of CYP2C9 poor metaboliser alleles experience a rate of major bleeding that is 3.68 fold higher than the rate seen in patients with the wild type genotype.

One study demonstrated that acetaminophen, at 2 g/day or 3 g/day, enhanced the anticoa‐ gulant effect of warfarin in stable patients, thus requiring close INR monitoring in the clini‐

Drug Interactions, Pharmacogenomics and Cardiovascular Complication

http://dx.doi.org/10.5772/48423

85

One of the preventative treatments of thromboembolic disease in patients is a prescription of heparin. However, heparin induced thrombocytopenia (HIT) is one of the most serious adverse reactions. HIT consequences can include thromboembolic complications and death.

An association between the Fc receptor gene and the risk for HIT has been found in some studies and it was demonstrated that the homozygous 131Arg/Arg genotype occurred sig‐ nificantly more often in patients with HIT than in the healthy volunteers' group [102] [103] ; however, another group have found no association [104]. Results are very preliminary and more evidence are needed before it may be possible to genotype candidates for heparin ther‐

Hydroxymethylglutaryl-coenzyme A reductase inhibitors (statins) have reduced coronary and cerebrovascular events and overall mortality when used for both primary and secon‐ dary prevention of ischemic heart disease [105]. Several known gene polymorphisms are as‐

Some studies examined polymorphism in the gene encoding cholesteryl ester transfer pro‐ tein (CETP), which is involved in the metabolism of high-density lipoprotein (HDL). Pravas‐ tatin-treated patients with either the B1/B1 or B1/B2 genotype (B1 presence and B2 absence of polymorphism) had significantly less atherosclerotic progression than patients receiving a placebo. Placebo-treated patients with the B2/B2 genotype had the least progression. How‐ ever, pravastatin-treated patients with the B2/B2 genotype (16% of the study population) de‐

The substitution (-455G/A) of the fibrinogen gene was found to be associated with an in‐ creased risk of myocardial infarction and stroke. During follow-up, placebo-treated pa‐ tients homozygous for the -455A genotype had the greatest disease progression; although, no association was found with benefit in disease progression in patients on

A five year study of pravastatin therapy in patients with a history of myocardial infarction and hypercholesterolemia showed that the largest benefit of pravastatin treatment in reduc‐ ing these events occurred in patients with the platelet GP IIIa PlA1/A2 genotype who also

An effect of polymorphism in the alloprotein gene was found on simvastatin therapy in a Scandinavian study. Among patients who received the placebo and had at least one apolipo‐

apy to identify those at risk for drug-induced thromboembolic complications.

sociated with the treatment progress [106, 107].

rived no benefit from pravastatin [108, 109].

carried at least one D allele of the ACE gene [111, 112].

pravastatin therapy [110].

cal setting [101].

**4.2. Heparin**

**5. Statins**

The frequency of CYP2C9 alleles is ethnically related [82, 86]. Approximately 20% of the Caucasian population carries one of the loss-of-function CYP2C9 alleles, and it is estimated that 1% of Caucasian carry two such alleles [71]. The frequency of the CYP2C9\*2 allele re‐ portedly ranges from 8-13% in different Caucasian populations. CYP2C9\*2 is present in 4% of African-Americans and is rare among Japanese individuals [87, 88]. The frequency of CYP2C9\*3 is 6-10% among Caucasian populations and 3.8% in Japanese populations [88, 89]. This data suggests that a substantial fraction of the Caucasian patient population may carry at least one defective CYP2C9 allele. In this group, the usual prescription dosage of warfarin may lead to major or even life-threatening haemorrhage.

Warfarin is commonly prescribed in combination with selective serotonin reuptake inhibi‐ tors (SSRIs), as depression often coexists with cardiovascular disease. Case reports suggest that some SSRIs can interact with warfarin to increase the likelihood of bleeding [90]. SSRIs cause adverse effects in isolation [91, 92] and can interact with other medications by inhibit‐ ing various isoenzymes of the CYP450 enzyme group [93, 94]. It has been shown that metro‐ nidazole and cimetadine increase the prothrombin time in patients on warfarin therapy. Chloramphenicol enhances warfarin's effect by inhibiting the action of the hepatic P450 sys‐ tem [71]. Some authors [95], [96] have warned that antidepressants with a known or predict‐ able interaction with warfarin, such as fluoxetine and fluvoxamine, should be avoided in patients receiving warfarin because of the risk of adverse outcomes.

Drug-drug interaction is a main concern in adverse drug reactions. The primary complica‐ tion occurring with warfarin treatment is bleeding. SSRIs may increase the risk of bleeding during warfarin therapy by hindering platelet aggregation through depletion of platelet se‐ rotonin levels [97-99]. Some SSRIs may also inhibit the oxidative metabolism of warfarin by CYP 2C9 [95].

It has been shown that concurrent use of selective serotonin reuptake inhibitors and warfar‐ in increases the risk of hospitalisation due to haemorrhage [90, 98]. Drugs which affect sero‐ tonin may have a detrimental effect on platelet function, as drugs which inhibit the reuptake of serotonin may decrease platelet serotonin levels leading to a reduction in serotonin-medi‐ ated platelet aggregation. Potential drug interactions can involve modification in either of these mechanisms and may result in pharmacodynamic interference or enhancement of war‐ farin's action.

It was shown that major and moderate drug-drug interactions with warfarin are very common in inpatients and are associated with INR results outside the therapeutic range. The most common drugs involved in the increase of anticoagulation effect were enoxaparin, simvastatin, omeprazole and tramadol. Multivariate analysis showed that age, length of hospital stay, exposure to >/=4 major or moderate drug interactions, and refusal of pharmacist recommendations contribute significantly to the patient's INR re‐ sult >5 [100].

One study demonstrated that acetaminophen, at 2 g/day or 3 g/day, enhanced the anticoa‐ gulant effect of warfarin in stable patients, thus requiring close INR monitoring in the clini‐ cal setting [101].
