1 Department of Anaesthesia and Intensive Care Medicine, Beaumont Hospital, Dublin 9, Ireland. 2 Royal College of Surgeons in Ireland, Dublin 2, Ireland
* Corresponding author. E-mail: anthonyc{at}rcsi.ie
Keywords: anaesthesia, general ; genetic factors, polymorphism ; metabolism, poor metabolizers ; metabolism, ultrarapid metabolizers ; pharmacogenetics
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Introduction |
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Pharmacogenetics emerged as a discipline that attempted to understand the hereditary basis for differences in responsiveness or inter-individual variation to therapeutic agents.85 Variation in a drug effect may vary from 2- to 10-fold or 100-fold, even among members of the same family.35 90 Similar inter-patient variability is observed in the risk of adverse effects of a drug or a chemical.85
Pharmacogenetics has been defined as the study of variability in drug response as a result of heredity factors.52 More recently, the term pharmacogenomics has been introduced. While the former term is largely used in relation to variants in genes that influence drug response, the latter refers to changes in gene expression as a consequence of drug exposure.14 The value of an understanding of pharmacogenetics for the clinician is to enable optimum therapeutic efficacy; to avoid toxicity of those drugs whose metabolism is catalysed by polymorphic isoenzymes; and to contribute to the rational design of new drugs. Pharmacogenetics and pharmacogenomics cannot be understood without a grasp of basic medical genetics.
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Medical genetics |
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Genes and alleles
A gene has its specific locus (from Latin, place) on a given chromosome. The gene for eye colour thus has its position in the human DNA determined by chromosome, and position on that chromosome. The gene, however, comes in various forms or alleles. For eye colour, the allele for blue eyes and the allele for brown eyes are two different alleles for the same gene. Although the expression is sometimes used, strictly speaking there is no such thing as a disease gene or a disease locus, only disease alleles.
Phenotype and genotype
The classical definition of phenotype is the way you look. If you have genes for blue eyes, that is your genotype. Your phenotype is being a blue-eyed person. A phenotype could also be an enzyme activity above or below a certain value, or being able to metabolize a certain substance.
Markers
Genetic research necessitates distinction between individuals at the DNA level. Different as we may be, we are identical for long stretches of DNA. Researchers usually attempt to identify gene markers, a short piece of DNA that can easily be detected. Two separate forms of markers exist, and the different forms can be used to tell the difference between individuals (or chromosomes, or parts of DNA). Finding a marker is like spotting the lanterns of a ship in the night. If you see one of the lanterns, you know you are not actually seeing the ship, but you can have a good guess as to where the ship is. When investigators find a marker that may be located close to a gene of interest, while the gene per se may not be located, substantial progress has been made. Some markers are single base mutations, others consist of repeats of short sequences where individuals differ in how many repeats they have, termed microsatellite markers.
Polymorphisms
The word polymorphism comes from the Greek poly, several, and morphe, form. Polymorphism, thus, is something that can take one of several forms. A DNA polymorphism exists when individuals differ in their DNA sequence at a certain point in their genome. A normal form and a mutated form may represent such a difference. The mutated form can be a single base mutation, or variation over a short stretch of DNA. The term polymorphism is a general one. It is most often used to describe a marker that occurs in several formsa marker polymorphism. Only 3% of DNA consists of coding sequences and in most regions of the genome, a polymorphism is of no clinical consequence. However, the term polymorphism is also used about a mutation inside a coding sequence, where the mutation might be causing disease.
Single nucleotide polymorphisms (SNPs) are changes in a single base at a specific position in the genome, in most cases with two alleles. SNPs are found at a frequency of about 1:1000 bases in humans. By definition, the more rare allele should be more abundant than 1% in the general population.40 The relative simplicity of SNP genotyping technologies and the abundance of SNPs in the human genome have made them very popular in recent years.21 Yet, there still is some debate about the usefulness of SNP markers compared with microsatellite markers for linkage studies, and how many SNP markers will have to be analysed for meaningful association studies.41
Genotyping
Several genotyping technologies have reached maturity in the last few years and are being integrated into large sale genotyping operations supported by automation. The choice of a technology for genotyping depends on whether a few different SNPs are to be genotyped in many individuals, or many different SNPs are to be genotyped in a few individuals.64 Although genotyping methods are very diverse, broadly, each method can be separated into two elements. The first element is a method for interrogating an SNP. This is a sequence of molecular biological, physical, and chemical procedures for the distinction of the alleles of an SNP, that is hybridization,79 primer extension,72 oligonucleotide ligation,54 and nuclease cleavage.55 The second element is the actual analysis or measurement of the allele-specific products, that is by gel separation,63 microarrays,7 mass spectrometry,22 flow cytometry,9 etc. Often, very different methods share elements, like reading out a fluorescent tag in a plate reader, or the method of generating allele-specific products (i.e. by primer extension or oligonucleotide ligation), which can be analysed in different analysis formats.21
Linkage studies
A gene and a marker are said to be linked if they reside close to each other on the same chromosome. In linkage studies, a set of markers, with a known location on the genetic map, is used to track down a gene of unknown location. In a linkage study, the disease allele is not known, but merely manifests itself as disease in the person who carries it. The basic assumption is that if a certain disease allele and a certain marker allele are found together in a family, the two are physically close on the same chromosome. The statistical methods of linkage analysis calculate just how unlikely it would be to consistently find a marker allele and a disease allele together. If this turns out to be very unlikely to have occurred by chance, the alternative hypothesis of linkage of marker and disease gene is accepted. If a marker allele and a disease allele occur together consistently in a population, they are said to be in linkage disequilibrium. Genetic linkage studies have identified various loci on causative genes for malignant hyperthermia susceptibility (MHS).49
Association studies
Association studies exploit the fact that there may be linkage disequilibrium in the population. A study may start out with a large number of markers with a known location. A number of patients with a disease are examined for these markers. If a substantial majority of patients have the same markers, it is likely that the gene responsible for the disease is located close to the markers that the patients have in common. The apolipoprotein (apo) E4 genotype, for example, is strongly associated with Alzheimer's disease, representing a susceptibility gene, but is not necessarily causative of the disease, meaning that having this genotype is not generally sufficient to cause Alzheimer's.57
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Pharmacogeneticsfocusing on drug targets |
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Enzymes
Genetic determinants of drug response can be divided into two types: (i) those characterized by alteration in drug metabolism, such as those as a result of differences in levels of N-acetyltransferase (NAT) or atypical plasma cholinesterase and (ii) those characterized by alteration in pharmacodynamics. Inter-individual variation in therapeutic drug response and toxicity is most often a result of variability in drug metabolism rather than pharmacodynamics.34
Observed genetic variability in drug metabolism can be either monogenic or polygenic. When the variability is a result of single genes, the term monogenic is used. When genes, which individually produce small effects but collectively lead to significant effects are involved, the term polygenic is used.30 Much pharmacogenetic research focuses on the monogenic variants of drug-metabolizing enzymes and on polymorphisms (i.e. variants that exist in at least 1% of the population).1 Such genes generally affect drug biotransformation by altering the amount or function of an enzyme. The existence of such polymorphisms explains why drug metabolism shows a polymodal distribution. In other words, patient populations can be divided into two groups (or phenotypes) according to their abilities to metabolize specific probe drugs. Poor or slow metabolizers have deficient metabolizing ability; in contrast, extensive metabolizers metabolize drugs more rapidly and may need higher doses to produce a therapeutic response.18
Drug metabolism is divided into phase I and phase II reactions. Phase I reactions, including oxidation, reduction, and hydrolysis, introduce a polar group into the molecule, whereas phase II reactions conjugate an endogenous hydrophilic substance with the drug, resulting in more water-soluble compounds.
Phase I enzymes
Oxidation, a major route of metabolism for many drugs, is catalysed by the mixed function oxidase system, which comprises cytochrome P450 (CYP) enzymes.
P450 enzymes are found in virtually all tissues with the highest concentration in the endoplasmic reticulum of the liver.66 73 The recommended nomenclature of cytochrome P450 isoenzymes is based upon grouping enzymes and genes into families and subfamilies with the prefix CYP denoting cytochrome P450. Families are characterized by an Arabic number (i.e. CYP2) and subfamilies are indicated by a letter (i.e. CYP2D). The individual genes coding for one specific isoenzyme are denoted by a second Arabic number after the letter in italics, that is CYP2D6. Members of the same enzyme/gene family may exhibit more than 40% identity in amino acid sequences, while a subfamily consists of those sharing greater than 55% sequence identity (Fig. 1).67
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The activity of cytochrome P450 enzymes can be measured by administration of a probe drug, known to be selectively metabolized by the CYP enzyme under study, followed by measurement of the metabolic ratio (the ratio of the drug dosage or unchanged drug to metabolite in serum or urine). However, such phenotyping takes into account all factors influencing the activity of the enzyme, such as the presence of a competing substrate, and is sensitive to the overall process of drug metabolism.
Genotyping involves identification of defined genetic mutations on the CYP genes that give rise to the specific drug metabolism phenotype. These mutations include genetic alterations that lead to over-expression (gene duplication), absence of an active protein product (null allele), or production of a mutant protein with diminished catalytic capacity (inactivating allele). Genotyping methods require small amounts of blood or tissue, are not affected by underlying disease or by drugs taken by the patient, and need to be done only once in a lifetime. By screening for genetic variants, an individual's drug metabolism phenotype can be characterized.84
Besides the P450 genes, other phase I enzymes are polymorphic, such as alcohol dehydrogenases (ADH) and acetaldehyde dehydrogenase (ALDH), as well as dihydropyrimidine dehydrogenase (DPD). With respect to the first two enzymes, the clearance of ethanol is significantly affected, ADHB2 giving a higher rate of ethanol metabolism and ALDH2 polymorphism influencing acetaldehyde metabolism. Poor metabolizers for ALDH2 develop flush reactions and anti-abuse like side-effects when drinking ethanol and the number of alcoholics with this genotype is lower. A polymorphism relevant to treatment with anticancer drugs is present in DPD. 5-Fluorouracil is metabolized by this enzyme. Subjects with impaired enzyme activity caused by inactivating gene mutations suffer from a severely increased risk of adverse reactions, including myelotoxicity and neurotoxicity following 5-fluorouracil administration.31
Phase II enzymes
Several enzyme families directly conjugate drugs or their oxidative metabolites. There are 15 human uridine diphosphate glucuronosyltransferases (UGTs), broadly classified into the UGT1 (phenol/bilirubin) and UGT2 (steroid/bile) families.78 87 Considerable polymorphism in glutathione S-transferase (GST) expression has been described and associated with susceptibility to disease, particularly cancer and asthma (both as disease-causing and disease-modifying factors).13 23 NAT was the first drug metabolizing enzyme for which a genetic polymorphism was discovered (slow and fast acetylators). Slow acetylators show a greater therapeutic response than fast acetylators to several drugs (i.e. isoniazid, hydralazine) but may be more susceptible to side-effects. There are two human NATs, NAT1 and NAT2, with discrete but overlapping substrate specificities. Although there are polymorphisms in NAT1, it is the genetic variability in NAT2 that is responsible for the slow-acetylator phenotype.24 Sulfotransferases (STs) catalyse the elimination of acetaminophen and morphine in neonates.53
The clinical implications of polymorphism of drug metabolizing enzymes are drug toxicity and therapeutic failure. The clinical relevance, however, depends on the therapeutic ratio of the drug.77
Transporter proteins
The influence of the genetic make-up of an individual is not limited to drug metabolism. Genetic variability influences drug absorption and this forms the basis for slow and rapid drug absorption.
Most drugs or drug metabolites enter the cells by passive diffusion. Some drugs are actively transported by transporter proteins, of which membrane transporters may play a key role. These transmembrane transporters are members of the large protein family known as ABC (adenosine triphosphate binding cassette) proteins.39 Although they do not catalyse biotransformation per se, they nonetheless markedly affect drug bioavailability and can act in conjunction with intracellular drug metabolizing enzymes.
P-Glycoprotein (also called multi-drug resistant P-glycoprotein, MDR1) is the first cloned and best-characterized ABC protein.61 92 At the bloodbrain barrier, P-glycoprotein may influence the uptake of substrates into the brain: high P-glycoprotein levels may limit the uptake of sufficient amounts of the desired drug into the brain, and reduced P-glycoprotein activity could lead to abnormally increased accumulation in the brain and undesired side-effects of a drug.6
A second subfamily of ABC proteins is the multi-drug resistance-associated proteins, also known as the multi-specific organic anion transporter.5 The first protein to be discovered in this category was MRP1, whose over-expression is responsible for the majority of non-P-glycoprotein-mediated multi-drug resistance. There are seven currently known MRPs with uncertain clinical significance. Rifampicin is known to induce human MRP2.17
Receptors
When examining the response to a drug, the most obvious target for genetic studies of drug response is the receptor. Genetic variability influences interactions with receptors and this forms the basis for poor or efficient receptor interactions. The polymorphisms in genes encoding receptors relevant to drug treatment of different diseases cause widespread variation in sensitivity to many drugs. For example, individuals with a mutation in the gene encoding prothrombin may have increased risk of cerebral vein thrombosis when using oral contraceptives. Other examples of the impact of genetic polymorphisms include: angiotensin converting enzyme (ACE) and its sensitivity to ACE inhibitors; ß-adrenergic receptors and their sensitivity to ß-agonists in asthmatics; and 5 hydroxytryptamine receptors and the response to certain neuroleptics.15
Mutations in cardiac potassium channel genes such as HERG (human ether-a-go-go-related gene) and KvLQT1 (chromosome 11 linked LQT gene) may both give susceptibility to drug-induced long QT syndrome, or KCNE2 (a potassium gene encoding MinK-related peptide-1, MiRP1) may give susceptibility to drug-induced arrhythmias. All are of clinical relevance to an anaesthetist.15
Pharmacogenomics is the research activity, which at the genome level, aims to identify disease genes and new drug response markers.31 The role of pharmacogenetics is increasingly recognized by the pharmaceutical industry with research programmes directed at drug discovery and development.93 Candidate drugs whose metabolism may involve polymorphic pathways may be screened out. Pharmacogenetic data may be used either to design better compounds or to help plan clinical studies. Screening volunteers and patients included in clinical trials may become necessary to minimize adverse events and optimize efficacy. Clinical investigations in various populations will help clarify inter-ethnic differences in drug disposition and response to a given drug. Knowledge of pharmacogenetics should help reduce the time and cost associated with new drug development.
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Anaesthetic implications |
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Recovery from general anaesthesia is dependent on factors governing drug sensitivity and drug disposition. Recovery from a single dose of i.v. anaesthetic agent is dependent on redistribution, whereas recovery after a prolonged infusion is progressively more dependent on metabolism and elimination of the drugs.28 Aging as well as environmental factors may influence drug dynamics. Both alcohol and tobacco play an important role in determining the degree of liver enzyme induction, which determines the rate of metabolism of some medications, including volatile anaesthetic agents, thus influencing outcome from anaesthesia.71
Enzymes
Genetic polymorphisms in metabolizing enzymes become relevant if: they are responsible for 50% or more of the clearance of a drug; when using drugs with a steep doseresponse curve and a narrow therapeutic window; and when using drugs whose activity depends upon a metabolite formed by a polymorphic enzyme.
Plasma cholinesterase
Inherited deficiency/reduced effect of plasma cholinesterase will result in prolonged muscle relaxation after succinylcholine.68 This was the first documented example of inherited variations in anaesthetic drug effects. The level and quality of plasma cholinesterase activity (acylcholine-acylhydrolase E.C.1.1.8, butyrylcholinesterase (BChE)) in a patient determines the duration of action of succinylcholine and mivacurium.33 Genetic variation is one of several factors determining the activity of cholinesterase in plasma. The expansion from only four known forms of human serum BChE a few years ago to over 20 variants identifiable at DNA level at present has greatly increased the complexity of diagnosis and interpretation of these genetic traits. Although the presence of a single genetic variant allele does not usually cause an increased duration of action of succinylcholine or mivacurium, it may do so if it occurs in heterozygous combination with otherwise induced low BChE activity. Therefore, it is important to be able to diagnose not only the well-known atypical variant but also the low-activity variants such as the H, J, K, and S variants. Using standard enzymatic and inhibition analysis, it is not possible to distinguish between the usual genotype (UU) and genotypes in which one of the quantitative variants, H, J, K, or S, is present in heterozygous combination with the usual gene (UH, UJ, UK, or US).33 Recent advances in molecular biology techniques have allowed analysis of the detailed structure of the human BCHE gene. For qualitative variants, a portion of the structural gene responsible for the amino acid sequence of the protein (BChE) is altered. This structural gene mutation accounts for the abnormal kinetic properties of the variant BChE protein. Variants in which there is a marked quantitative reduction in the level of enzymatic activity could result from: (i) a structural modification that causes little or no active enzyme being preserved (such as a mutation to a stop codon, or the production of a very unstable enzyme), or (ii) a regulatory defect affecting primarily the rate of enzyme synthesis. The latter type mutations are much more difficult to diagnose, as they can occur over a much larger region of the BCHE gene. Structural gene differences should be within the 1722 nucleotide bases located in exons 2, 3, or 4, as these contain all the codons representing the 574 amino acids making up the monomeric BChE enzyme protein sequence.42
In addition, plasma cholinesterase availability could be decreased, and thus neuromuscular block after succinylcholine lengthened, by competitive interaction with (i) anticholinesterases, including neostigmine, edrophonium, and ecothiopate (an organophosphorus compound once used topically as a miotic in ophthalmology) and/or (ii) other drugs metabolized by plasma cholinesterase, such as etomidate, propanidid, ester local anaesthetics, methotrexate, monoamine oxidase inhibitors, and esmolol.68
CYP enzymes
CYP families 13 are responsible for phase I metabolism of most drugs. Enzymes in the CYP2C, CYP2D, and CYP3A subfamilies are most active in metabolizing clinically used drugs.18
CYP2C enzymes eliminate oral hypoglycaemics, warfarin, some antiepileptics, non-steroidal anti-inflammatory drugs, amitriptyline, barbiturates, diazepam, and omeprazole.
The most important substrates for CYP2D6 are a number of psychoactive drugs such as antidepressants and neuroleptics, and cardiovascular drugs such as beta-blockers and antiarrhythmics. Drugs in this class relating to anaesthetic practice include codeine, tramadol, ondansetron, granisteron, and metaraminol.10 It has been postulated that CYP2D6 poor metabolizers are more susceptible to pain than extensive metabolizers because of a defect in synthesizing endogenous opioids.15 Codeine is ineffective as an analgesic in 67% of a Caucasian population as a result of homozygosity for non-functional CYP2D6 mutant alleles. CYP2D6 deficient patients will not convert codeine to morphine. Postoperative pain treatment with codeine-containing drugs will therefore have limited effect in patients with this trait, whose request for larger doses of codeine could easily be misinterpreted as drug addiction. This genetic variation makes it not surprising that a standardized prescription of codeine for pain relief will result in remarkable variation in the adequacy of pain relief.
CYP2E1 is more of toxicological interest as it has been reported to have a unique capacity to activate many xenobiotics to hepatotoxic (among them acetaminophen) or carcinogenic products.47 CYP2E1 is the principal, if not sole human liver microsomal enzyme catalysing defluorination of sevoflurane. It is also the principal, but not exclusive enzyme responsible for the metabolism of methoxyflurane, and is responsible for a significant fraction of isoflurane and enflurane metabolism (Table 1). Identification of CYP2E1 as the major anaesthetic metabolizing enzyme in humans provides a mechanistic understanding of fluorinated ether anaesthetic metabolism and toxicity.37 75
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Enzymes of phase II of drug metabolism show extensive polymorphism (Table 2). Despite this, investigations of genotypephenotype correlations have been confounded by limited substrate specificity and a less pronounced hereditary contribution to differences in bioactivity.31 87 Genetic polymorphism in genes encoding enzymes such as UGTs, STs, NAT 1 and 2, and GSTs may cause significant variation in rendering drugs water soluble and then suitable for renal excretion.85
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Angiotensin converting enzyme
Most recently, Lasocki and colleagues provided the first evidence for an in vivo association between the pressureflow relationship and the insertion/deletion polymorphism of the ACE gene. Multivariate analysis showed that homozygosity for the D allele was the only predictive variable of the slope of the curve. These findings indicate a modified vascular response to flow in DD patients. The investigators also showed an increased vascular reactivity to phenylephrine associated with the D allele of the ACE gene.43
Transporter proteins
P-Glycoprotein, a member of the adenosine triphosphate-binding cassette superfamily of cellular efflux drug transporters, is expressed in the capillary endothelium of the bloodbrain barrier and in many other cell membranes such as intestinal enterocytes and biliary and renal epithelial cells.82 Block of P-glycoprotein allows enhanced central nervous system (CNS) entry of some drugs, offering new possibilities to explain CNS-related adverse effects during the administration of drugs that are substrates of P-glycoprotein and, furthermore, to manipulate the CNS entry of drugs whose target is located in the brain.89
The P-glycoprotein substrate and inhibitor cyclosporin was shown to increase fentanyl-induced analgesia in mice.11 More recently, morphine has been shown to increase analgesia in P-glycoprotein knockout mice compared with wild-type mice.74 Thus, P-glycoprotein may limit morphine entry into the brain. Loperamide, a widely used anti-diarrhoeal agent, although a potent opioid in vitro, produces only gastrointestinal opioid effects and lacks CNS effects. This apparent tissue selectivity is probably a result of loperamide being a P-glycoprotein substrate, so that P-glycoprotein in the CNS effectively prevents access of loperamide to the CNS. Supportive of this hypothesis is the finding that in mice with MDR1 gene disruption, brain loperamide concentrations were 8-fold higher than those observed in normal mice, and lethal opioid effects were produced.62 In contrast to loperamide, opioids in widespread clinical use as i.v. anaesthetic agents or adjuvants, such as fentanyl, sufentanil, and alfentanil, are not in vitro substrates of P-glycoprotein. Morphine is a P-glycoprotein substrate with clearly less clinical relevance than loperamide. Inhibitory effects of the opioids fentanyl, sufentanil, and alfentanil on P-glycoprotein activity in vitro are reached only with relatively high concentrations.82 Thus, the wide spectrum of P-glycoprotein activity may partly explain the varying CNS-related effects of opioids.
As new drugs are introduced into clinical practice, it will be important to assess whether they are P-glycoprotein substrates or inhibitors to assess their potential for drug interaction. Inter-individual variability in P-glycoprotein activity is now recognized, which may at least partially depend on genetic polymorphism. Homozygosity for an allele associated with deficient P-glycoprotein activity occurs in 24% of white people.26
Receptors
Polymorphisms in genes encoding receptors (drug targets) may explain some of the variation in sensitivity to drugs.
Ryanodine receptor (RYR1)
MH is a classic example of a dramatic interaction between a drug and a mutated receptor. In patients with a mutation of the skeletal muscle RYR1 gene, exposure to halothane and/or succinylcholine produces uncontrolled release of calcium from skeletal muscle cells, leading to hyperthermia, myolysis, and ultimately multi-organ failure. To date, almost 50 different mutations in the RYR1 gene on chromosome 19 are known to cause susceptibility to malignant hyperthermia (MHS phenotype).49 Genetic linkage studies indicate that the RYR1 locus (MHS1) on chromosome 19q13.1 accounts for at least 50% of MH families. A second locus, MHS2, was assigned tentatively to chromosome 17q in North American families,45 but could not be replicated in European families despite extensive efforts. However, it is quite possible that the differences in the protocols result in detection of different phenotypes and are weighted differently with respect to identification of modulating gene effects.16 Markers linked to the CACNL2A gene on chromosome 7q have been tentatively linked to MHS in a single European family (MHS3).29 A systematic linkage study using a set of polymorphic microsatellite markers covering the entire human genome in a small number of large, apparently non-chromosome 19 linked European MH families, identified a locus on chromosome 3q13.1 (MHS4),70 a locus on chromosome 1q (MHS5),50 59 and a tentative locus on chromosome 5q (MHS6).59 A causative gene at the MHS3 and MHS4 locus has yet to be identified. The CACNL1A3 gene encodes the 1-subunit of the DHP receptor maps to the MHS5 locus and functions as a voltage gated channel. Evidence for the MHS6 locus is weak and its validity remains to be confirmed.49
µ-Opioid receptor (MOR)
Variation in the expression of this receptor determines the analgesic potency of morphine. It also explains the difference in the response to painful stimuli and response to opioid drugs, probably a result of a genetic polymorphism in the transcription-regulating region of this gene.77 This makes the MOR gene a candidate for susceptibility or resistance to pain. A SNP in the human MOR gene at position 118 (A118G transition) results in a receptor variant that binds ß-endorphin nearly three times more tightly than the most common allelic form of the receptor.4 This makes ß-endorphin nearly three times more potent than in individuals without the mutation. It is unclear whether this variant has direct or indirect implications for opioid addiction.15
More recently, three SNPs in the hMOR gene that cause amino acid substitutions in the third intracellular (i3) loop of MOR have been identified (R260H, R265H, and S268P). Each of the three SNPs caused substantial changes in basal G protein coupling, calmodulin binding, or both. Carriers of the mutant alleles might display altered responses to narcotic analgesics.83
GABAA receptor
A new class of human GABAA receptor subunit (), that confers insensitivity to the potentiating effects of i.v. anaesthetic agents on gabaminergic transmission, has been identified. Wilke and colleagues identified the gene, symbolized GABRE, coding for class epsilon of the GABAA receptor (gene map locus Xq28).88 Genetic variation in the gene encoding for this subunit of a GABA-receptor may be of importance for the sensitivity to diazepam, barbiturates and propofol, or susceptibility to alcohol addiction.8 32 Volatile anaesthetics act through a different site on the GABAA receptor molecule from the i.v. anaesthetics, although the nature and location of that site remains unclear (Fig. 2). Volatile anaesthetics and propofol show no significant selectivity between any receptor subtype. In contrast, etomidate acts preferentially through receptors containing ß2 or ß3 subunits. This selectivity is determined by a single amino acid (an asparagine at amino acid number 265 in ß2 and ß3 subunits; a serine at the equivalent position in ß1.2 Extrapolating the results of animal studies to humans, it could be predicted that an anaesthetic that is selective for ß3 containing GABAA receptors would enable faster recovery, perhaps without the hangover effect.86 Gene targeting in mice may be valuable for elucidating the mechanism of action for some drugs. A variety of manipulations are possible, including introducing a gene not present normally (transgenic mice), removing an endogenous gene (knockout mice), or replacing an endogenous gene with an altered copy (knockin mice).27 Knockout of the GABAA
2 receptor subunit gene resulted in mice that were insensitive to the sedative/hypnotic actions of benzodiazepines.20
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An understanding of the CYP system and its substrates is also a key factor in the prevention of important drugdrug interactions, either as a result of enzyme induction or inhibition. The former may take some time to develop and usually reduces the effect of the drug involved, while the latter takes place instantly with side-effects as a common result.12 The number of possibilities are overwhelming, but may be reduced somewhat by a few rules of thumb. It is usually required that these interactions occur when the involved drugs are substrates for the same CYPs. Susceptibility to, for instance, inhibition interactions usually requires that one metabolic product accounts for 3040% of the effect of a drug, and that its metabolic pathway is catalysed by a single enzyme.46
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Future developments |
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Routine screening of patients before starting pharmacotherapy would have significant cost implications. Cost savings associated with toxic episodes or therapeutic failure and subsequent intervention could be expected in most specialties. But in anaesthesia, we administer drugs to a large number of patients, often once only, and frequently only briefly after the patient has presented for treatment. In this setting, a genetic screening programme is unlikely to represent a cost-effective method for reducing morbidity.
However, once conditions such as MH are documented, family screening becomes a logical follow-up. The quest for a simple non-invasive diagnostic test for MH susceptibility has moved forward dramatically since the first descriptions of linkage of the RYR1 gene to MHS. However, resolution of issues confounding the genetics of MH and its associated disorders will be necessary before genetic diagnosis can widely replace the in vitro contraction test.49
Similarly, screening for abnormal BChE genotypes is neither practical nor cost effective in daily clinical practice, but it could be used to supplement equivocal results obtained by biochemical methods or in situations where establishing the genotype may be clinically important.81
Furthermore, recent findings in molecular research suggest that the outcome of cardiovascular surgery is at least partly determined by the individual patient's genetic predisposition to react to surgical trauma and extracorporeal circulation. Genomic variations may prove to serve as future diagnostic tools for the risk stratification of patients undergoing cardiovascular surgery. The evaluation of possible genomic markers for risk stratification of patients at high risk of developing adverse outcomes has begun.69
Pharmacogenetics will have its application in clinical research.76 When designing clinical trials, genotype can be used a priori, as an exclusion criterion. With this methodology, the study group can be smaller and more homogeneous, though less representative. Phase I trials can thus be designed for representative populations of the principal metabolic patterns.51 Alternatively, the genotype can be used a posteriori, as a stratification factor. It should also be recalled that ethnic origin has an effect on allelic polymorphism, and that trials conducted on Caucasians may not necessarily be extrapolated to other ethnic groups.58
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Conclusions |
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Although pharmacogenetics is unlikely to change the way anaesthesia is practised today it may help to elucidate inter-patient variability in drug response. We will, undoubtedly, see its impact on other specialties, on new drug development, and in drug delivery systems.
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Acknowledgments |
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References |
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