13. Pharmacogenomics
13.6. Examples of pharmacogenetic studies
13.6.1. Pharmacogenetics of statins
Statins (or HMG-CoA reductase inhibitors) are a class of drugs used to lower cholesterol levels by inhibiting the enzyme HMG-CoA reductase, which plays a central role in the production of cholesterol in the liver. Increased cholesterol levels have been associated with cardiovascular diseases, and statins are therefore used in the prevention of these diseases. Statins have rare but severe adverse effects, particularly muscle damage, and some doctors believe they are overprescribed. The best-selling statin is atorvastatin, marketed as Lipitor (manufactured by Pfizer) and Torvast. By 2003 atorvastatin became the best-selling pharmaceutical in history.
Because of their widespread use and rare but severe adverse effects, a lot of pharmacogenetic studies have been carried out. The CYP enzymes play an important role in the metabolism of statins. CYP3A4 metabolize the lovastatin, simvastatin and the atorvastatin. The level of CYP3A4 can show a 10-fold difference between people indicating genetic variants. In one study, a -290A/G promoter SNP influenced significantly the LDL-C level after atorvastatin treatment, while the M445T variant influenced the LDL-C level both before and after treatment (Kajinami K et al., 2004).
Individuals carrying rs35599367, C>T SNP in intron 6 of the gene needed 0.2-0.6-fold simvastatin dose reduction for the optimal lipid level.
CYP3A5 contribute to biotransformation of some statin. There is a 6986 G/A SNP in intron 3 of the gene, which influence significantly the expression of the gene. Only 10%
of the European populations show high CYP3A5 expression, and in these people the lovastatin, simvastatin and the atorvastatin treatment is significantly less effective (Kivistö KT et al., 2004).
The multidrog resistance-1 (MDR-1, ABCB1) significantly influences the transport and localization of the statins. In a study the previously mentioned ABCB1 C3435 SNP influenced the LDL-C level in atorvastatin therapy (Becker ML et al., 2010).
Polymorphisms in the HMGCR gene encoding HMG-CoA reductase, the main target of statins, also influence the response to the drugs.
Mutations in the LDLR gene cause familiar hypercholesterolemia (FH). The responds to statins of the mutation carriers depend on the types of the mutations. People with null mutations respond worse than people with mutations influencing only the functions of the receptor.
GWAS were also carried out in this topic. The Study of the Effectiveness of Additional Reductions in Cholesterol and Homocysteine (SEARCH) identified a SNP in the SLC01B1 gene (SLCO1B1*5) which is associated with statin-induced myopathy in simvastatin (Zocor) treated patients with cardiovascular diseases.
It must be noted, however, that the pharmacogenetic results in connection with the statins are rather controversial, and thus in the FDA-approved list there are only two items of statins with pharmacogenomic drug labels.
13.6.2. Clopidogrel
Clopidogrel is an oral, thienopyridine class antiplatelet agent used to inhibit blood clots in coronary artery disease, peripheral vascular disease, and cerebrovascular disease. It is marketed by Bristol-Myers Squibb and Sanofi under the trade name Plavix. The drug works by irreversibly inhibiting a receptor called P2Y12, an adenosine diphosphate (ADP) chemoreceptor on platelet cell membranes. Adverse effects include haemorrhage, severe neutropenia, and thrombotic thrombocytopenic purpura (TTP). It is prescribed for 40 million patients annually.
Clopidogrel is a pro-drug activated in the liver by cytochrome P450 enzymes, including CYP2C19. Three-four percent of the Caucasian population homozygote, while 24% heterozygote for the inactive variants of the gene associating with higher rate of cardiovascular complications.
GWAS was carried out in an Amish population, and a SNP in the CYP2C19 gene was identified, which was associated with reduced drug response, and this was responsible for 12% of the drug response variations. The traditional factors (BMI, age, cholesterol level) were responsible for only 10% of the variations. This was later confirmed in another study and in a 12-year follow-up study the CYP2C19 status was the only independent risk factor, when cardiovascular death, non-fatal myocardial infarction or coronary revascularization were applied as target values. In another study two variants of the ABCB1 were shown to be associated with adverse drug response. The product of this gene plays a role in the absorption of the drug. CYP2C19 has a gain of function allele (CYP2C19*17) which codes for an ultra-fast metabolizing form of the enzyme. Carriers of this allele respond better to the drug (Myburgh R et al., 2012). Presently, FDA recommends alternative therapies for poor responders, and in March 2010 the warnings about CYP2C19 genotypes were put into the drug label.
13.6.3. Pharmacotherapy of asthma
There are four major classes of asthma pharmacotherapy currently in widespread use:
(1) β2-agonists used by inhalation for the relief of airway obstruction (e.g. albuterol, salmeterol, fenoterol); (2) glucocorticosteroids for both inhaled and systemic use (e.g., beclomethasone, triamcinolone, prednisone); (3) theophylline and its derivatives, used for both the relief of bronchospasm and the control of inflammation; and (4) inhibitors and receptor antagonists of the cysteinyl-leukotriene pathway (e.g. montelukast, pranlukast, zafirlukast, zileuton).
Variability in individual asthma treatment response may be due to many factors, including the severity and type of disease, treatment compliance, intercurrent illness, other medication taken (drug–drug interaction), environmental exposures, and age.
However, there is reason to believe that genetic factors underlie much of the observed treatment response variance. A study of treatment response to glucocorticosteroids, a beta-2 adrenergic agonist, and an experimental leukotriene inhibitor has found that up
178 Genetics and genomics
to 60–80% of the variance in drug response may be due to differences between individuals. This value corresponds to the maximum limit of genetic variance, and indicates that a clinically relevant part of the response to the main classes of asthma drugs may be due to genetic determinants (Szalai et al., 2008).
To date, investigations in the field of asthma pharmacogenomics have focused on three classes of asthma therapies: β2-agonists, leukotriene antagonists and glucocorticosteroids. Below examples are shown of pharmacogenetic studies about β2-agonists and leukotriene antβ2-agonists in asthma.
13.6.4. Interaction between genetic variations and β2-agonists
The 5q31-33 is an important pharmacogenomic region for asthma. β2-agonists are used widely by inhalation for the relief of airway obstruction. These drugs act via binding to the β2 adrenergic receptor (ADRB2), a cell surface G protein-coupled receptor located on 5q32. Responses to this drug are currently the most investigated pharmacogenomic pathway in asthma. Two coding variants (at positions 16 and 27) within the ADRB2 gene have been shown in vitro to be functionally important. The Gly16 receptor exhibits enhanced downregulation in vitro after agonist exposure. In contrast, Arg16 receptors are more resistant to downregulation. Because of linkage disequilibrium, individuals who are Arg/Arg at position 16 are much more likely to be Glu/Glu at position 27;
individuals who are Gly/Gly at position 16 are much more likely to be Gln/Gln at position 27. The position 27 genotypes influence but do not abolish the effect of the position 16 polymorphisms with regard to downregulation of phenotypes in vitro.
Retrospective studies and prospective clinical trials have suggested that adverse effects occur in patients homozygous for arginine (Arg/Arg), rather than glycine (Gly/Gly), at position 16. Bronchodilator treatments avoiding β2-agonist may be appropriate for patients with the Arg/Arg genotype.
13.6.5. Interaction between genetic variations and leukotriene antagonists
Leukotrienes, released by eosinophils, mast cells and alveolar macrophages, are among the main mediators in asthma, inducing airway obstruction, migration of eosinophils and proliferation of smooth muscle. Of the three enzymes exclusively involved in the formation of the leukotrienes (5-lipoxygenase (ALOX5), leudotriene C4 (LTC4) synthase, and LTA4 epoxide hydrolase), ALOX5 is the enzyme required for the production of both the cysteinyl-leukotrienes (LTC4, LTD4, and LTE4 ) and LTB4. ALOX5 activity in part determines the level of bronchoconstrictor leukotrienes present in the airways, and pharmacological inhibition of the action of ALOX5 or antagonism of the action of the cysteinyl-leukotrienes at their receptor is associated with an amelioration of asthma. A polymorphism located in the promoter of the ALOX5 gene decreases gene transcription, and less enzyme is produced when the number of repeats of an Sp1 binding motif GGGCGG, which acts as a transcription modulating site, is different from the usual number of 5 (Kalayci et al., 2006). In a study in the United States approximately 6% of asthma patients did not carry a wild-type allele at the ALOX5 core promoter locus (Drazen et al., 1999). It was hypothesized that patients possessing the altered promoter might be less responsive to a leukotriene modifier. In randomized, double-blind, placebo-controlled trials of ABT-761, an ALOX5 inhibitor, which is a derivative of the antileukotriene drug Zileuton this hypothesis was investigated. The
primary outcome of the clinical study was improvement in FEV1 (forced expiratory volume in 1 second). In the unstratified population, the inhibitor produced a 12% to 14% improvement in FEV1. Patients homozygous for the wild-type promoter had a 15%
improvement in FEV1. In contrast, those patients homozygous for the mutant version of the promoter had a significantly decreased FEV1 response. Otherwise the ALOX5 core promoter locus does not account for all patients who did not respond to ALOX5 inhibition, which suggests that there may be other gene defects in the pathway leading to a lack of response to this form of treatment. It was suggested that patients who fail to respond to ALOX5 inhibition are those in whom other mechanisms are responsible for asthmatic airway obstruction.
LTC4 synthase is a membrane-bound glutathione transferase expressed only by cells of hematopoietic origin and is a key enzyme in the synthesis of cys-LTs, converting LTA4 to LTC4. The gene encoding LTC4 synthase is located on 5q35. An adenine to cytosine transversion has been found 444 bp upstream (-444) of the translation start site of the LTC4 synthase gene and reported that the polymorphic C -444 allele occurred more commonly in patients with aspirin intolerant asthma (AIA) (Sanak et al., 1997 and 2000). A 5-fold greater expression of LTC4 synthase has been demonstrated in individuals with AIA when compared with patients with aspirin-tolerant asthma;
furthermore, the expression of LTC4 synthase mRNA has also been shown to be higher in blood eosinophils from asthmatic subjects compared with control subjects and was particularly increased in eosinophils from patients with AIA. In addition, it was found that, among subjects with asthma treated with zafirlukast (a leukotriene receptor antagonist), those homozygous for the A allele at the -444 locus had a lower FEV1 response than those with the C/C or C/A genotype (Palmer et al., 2002).
It must be noted that these examples are researches, and the results have not got into clinical practice yet.