�14.1. OVERVIEW OF FACTORSGOVERNING INTERINDIVIDUALVARIATIONS
Patients on drug therapy manifest differentresponses to the same regimen.A drug mayhave little or no therapeutic effects in somepatients but elicit adverse effects at low
doses in others. Interindividual variation indrug efficacy and safety has resulted in thefailure of drug candidates in clinical trialsand the withdrawal of other drugs from themarket.
Intersubject variation in drug responseis thought to cost lives and dramaticallyincrease health care costs [1,2]. While
14.6. CURRENT AND POTENTIAL APPLICATION OFPHARMACOGENETICS
14.7. SUMMARY
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382 INDIVIDUALIZATION OF DRUG REGIMENS
matching the patient with the right drugand dosing regimen tailored to his or hertherapeutic need would overcome theseproblems, efforts to use therapeutic drugconcentration monitoring to individualizedosing have had limited clinical impact.Therapeutic drug monitoring, albeit cum-bersome and expensive, is sometimes usedas part of drug safety and efficacy man-agement for a small number of drugs withlow therapeutic indexes, such as anticon-vulsants, digoxin, some chemotherapeuticagents, and some immunosuppressants.
With advances in medical genetics andthe study of the genetic basis of variation indrug disposition and biologic activity (phar-macogenetics), some interindividual vari-ability can be predicted. In such cases theoverall response to a drug is determined byvariations in key proteins regulating dispo-sition and activity. These advances have also led to development of medicinal agentsdesigned to interfere with proteins orprocesses involved in medical conditionsand perhaps benefit only defined individu-als. In principal, genetic information—genotyping and phenotyping—can accountfor differences in drug response amongindividuals and identify individuals withdisease-causing mutations that might becountered with targeted therapies.
While an individual’s phenotype candirectly assess a clinical measureable bio-logic function of interest (e.g., whether anindividual is a fast or slow metabolizer of acertain drug), phenotyping is often expen-sive and not readily adapted for routineclinical practice. Analysis of genetic infor-mation—genotyping—might prove to bemore efficient and better able to be adaptedto the individualization of drug therapy.
�14.2. HISTORICAL PERSPECTIVE ONPHARMACOGENETICS
We have learned a great deal about the roleof genetics in determining the safety andefficacy of drug therapy and the molecular
basis of disease during the past decade.Thestudy of the association between geneticsand response to drug therapy is calledpharmacogenetics. Pharmacogenetics isdistinctly different from pharmacoge-nomics in that it is only one area of phar-macogenomic research. According to aleading pharmaceutical researcher’s defini-tion, pharmacogenetics refers to people,including gene identification and selectingthe “right medicine for the right patient.”Pharmacogenomics, on the other hand,includes other applications of genetic infor-mation related to drug response, includinggene expression, alteration of protein func-tion, and pathological consequences [3].Pharmacogenetic investigations startedmore than 150 years ago.
By 1910 biological chemists in Europediscovered that humans were capable oftransforming ingested drugs and chemicalsinto other products before excreting them.This observation, which ignited interest inlinkage to Mendel’s discovery, 44 yearsearlier, of the fundamental laws of hered-ity, invited the inference of receptors by sci-entists in Germany and France in the 1870s,and led to the suggestion that genetic mate-rial was crucial in directing chemical trans-formations in humans and other animals.These pioneering investigators envisionedenzymes as detoxifiers of drugs and otherchemicals. In this role, enzymes enabledpeople to use drugs effectively and thenexcrete them harmlessly. These visionaryscientists stressed that because of thefailure of enzymes to detoxify drugs, somepeople might experience a clinical responsevery much out of proportion to that seen inan average person.
In the first part of the twentieth century,reports appeared on differences amongWhites, Blacks, and Chinese in response todrugs and chemicals. During the 1920s and1930s, individual differences in sensory per-ception of chemicals were studied. Humanfamily studies showed that the failure toperceive bitterness after tasting phenyl-thiourea was transmitted from parents to
BIO14 5/2/03 2:47 PM Page 382
14.2. HISTORICAL PERSPECTIVE ON PHARMACOGENETICS 383
children as an autosomal recessive trait. Inthe late 1940s another major advanceemerged from studies of sickle cell anemiaand the sickle cell trait. These studies pro-vided the first irrefutable proof that theresponse of individuals to their environ-ment was linked directly to the proteinsthey synthesize. The studies proved theautosomal transmission of this disorderfrom parents to offspring and demonstratedthat a change in the globin molecule involv-ing the replacement of a single amino acidby another was the basis of the disease.
Wendell W.Weber,who has authored twobooks on pharmacogenetics, noted [4]: “In1953, the molecular basis of heredity, thedouble helix of DNA, was described. In1956, human chromosomes had been visu-alized,enumerated,and soon thereafter oneform of cancer (chronic myeloid leukemia)had been associated with an aberrant chro-mosome (the Philadelphia chromosome).. . . The development of electrophoresis andadvances in chromatography permittedcomplex proteins and smaller molecules tobe separated and analyzed.” Consequently,he noted, “Protein polymorphism, initiallyassociated with enzymes that occurred inmultiple forms, was soon recognized as aphenomenon of much broader biologicalsignificance. For the first time, new relation-ships between the metabolic fate of exoge-nous substances in humans and the geneticcontrol of human drug response werecoming into view.”
For many in the pharmaceutical sciences,the most important application of pharma-cogenetics is as a predictor of drug response.CYP2D6, one of many cytochrome P-450drug-metabolizing enzymes, typifies thepharmacogenetic diversity that has beenidentified in humans (Table 14.1) [5].CYP2D6 is involved in the metabolism of atleast 30 drugs, many of which are widelyused. At least 48 nucleotide variations thatcreate 53 CYP2D6 alleles have been identi-fied in the CYP2D6 gene. Of these, one isfound in all human populations, some occurin several, while others are limited to a
single population. Some of these variationslead to multiple copies of the enzyme, whileothers lead to the absence of the enzyme.Consequently a drug dose that produces the desired response in the average personcan be therapeutically ineffective in somepeople or unsafe in others [5]. New method-ology for gene analysis reflective ofCYP2D6 activity allows clinical decisions tobe made quickly [6].
Genetic polymorphisms in drug meta-bolism are certainly one reason for the dif-ferences in how patients respond to drugs.Another reason, of looming importance, ismutations in genes coding for a receptor or another protein that controls drugresponse; such mutations are being identi-fied with increasing frequency. The genesfor more than a dozen inherited traits orpolymorphisms—among them cystic fibro-sis, insulin receptor resistance, throm-bophilia, estrogen resistance, and HIVresistance—have been identified. In thesecases pharmacogenetics may serve as aguide to the development and applicationof drug therapy. In thrombophilia, forexample, variation in the structure of ablood-clotting protein predicts suscepti-bility to deep-vein thrombosis. Moleculartechniques have shown that the susceptiblephenotype is associated with a singlechange in factor V—the Leiden mutation—that substitutes a glutamine for an argininein the molecule. Not everyone who carriesthe variant protein experiences deep-veinthrombosis, but all carriers are lifelong can-didates for oral anticoagulation, whichrequires close monitoring to maintain effi-cacy and avoid serious bleeding.
The promise of pharmacogenetics is thatby determining a patient’s genotype, physi-cians will be able to make better prescribingdecisions. Determining genotype prior to aprescribing decision may improve care byincreasing the proportion of patients forwhom the drug is beneficial or by decreasingthe risk of adverse events. In such circum-stances the key question is: Which gene(s)can predict response to a specific drug?
BIO14 5/2/03 2:47 PM Page 383
384 INDIVIDUALIZATION OF DRUG REGIMENS
�TA
BLE
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BIO14 5/2/03 2:47 PM Page 384
14.2. HISTORICAL PERSPECTIVE ON PHARMACOGENETICS 385�
TABL
E 14
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Co
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Fun
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Gen
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Var
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BIO14 5/2/03 2:47 PM Page 385
386 INDIVIDUALIZATION OF DRUG REGIMENS
Candidate gene association studies havebeen the approach usually used to establisha genetic basis for individual drugresponses. These experiments consist ofstatistical analysis of the relationshipbetween a drug response trait (phenotype)and variants in a selected candidate gene(genotype). Research scientists can estab-lish a relationship by demonstrating thatparticular genotypes are more prevalent inindividuals who have a specific phenotypethan those who do not, or by demonstrat-ing that the mean value of a phenotypescale is dependent on genotype. Candidategenes are selected because they are knownto be in a pathway related to the phenotypeof interest or because experiments havesuggested a mechanism by which variationin the gene could account for variation inclinical outcome.
Genome-wide linkage analysis has beenused to establish the genetic causes ofmany Mendelian diseases, but because itrelies on patterns of genetic and pheno-typic changes within families, the analysis israrely applicable for drug response traits.Therefore another strategy has been pro-posed—that a genome-wide map of thou-sands of single nucleotide polymorphisms(SNPs) will be useful for pharmacogeneticdiscovery. SNPs are simply locations alongthe chromosomes where a single basevaries among different people. Where some have a guanine in a given string ofnucleotides, for example, others might havea cytosine. The premise of SNP mapping isin the assumption that common diseasessuch as cancer must be caused in part bycommon mutations.And the most commonmutations in the genome are SNPs, whichoccur about every 1000 bases.
The author of a recent review on phar-macogenetics observed: “The proposedstrategy is to determine SNP marker fre-quencies in two populations: cases (respon-ders or affected individuals) and controls(nonresponders or unaffected individuals).Since markers are closely spaced through-out the genome, there should be detectable
marker frequency differences correspond-ing to each genetic determinant” [7]. Theincreased frequency of the joint inheritanceof two genes that are closely linked on achromosome is termed linkage disequilib-rium. Association studies based on SNPscan work because of linkage disequilibrium.
Linkage disequilibrium-based associa-tion studies using candidate genes fromselected genomic regions may be an efficientstep toward a genome-wide associationstudy. Within those regions, SNPs areselected at random locations, rather thanspecifically within the genes.Those SNPs arethen used to genotype cases and controls.Using biological intuition, if successful,greatly reduces the time and cost required todetermine an association.At today’s prices itwould cost as much as $50 million to conducta genome-wide association study [7].
�14.3. PHARMACOGENETICS: DRUGMETABOLISM AND TRANSPORT
14.3.1. Cytochrome P450 3A Geneticsin Drug Metabolism
Various defense mechanisms have evolvedto protect humans and other animals fromthe thousands of chemicals present in food,drinks, and the environment. Of particularinterest are cytochrome P450 (CYP)enzymes, which catalyze the final step inthe incorporation of oxygen into organicmolecules.They frequently convert xenobi-otics, including human-made chemicals anddrugs, into less toxic products but can alsotransform nontoxic chemicals into toxic orcarcinogenic-reactive species.
There are more than 50 known humanP450 genes (Table 14.1). The most highlyexpressed subfamily is CYP3A, whichincludes the isoforms CYP3A4, CYP3A5,CYP3A7, and CYP3A43. These isoformsaccount for as much as 30% of total P450content in liver and have an important role in the oxidative metabolism of at least50% of all drugs. The most abundant
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14.3. PHARMACOGENETICS: DRUG METABOLISM AND TRANSPORT 387
CYP isoform expressed in liver and gut isCYP3A4.
Hepatic expression of CYP3A4 isknown to vary by more than 50-fold amongindividuals, and CYP3A4 enzyme functionin vivo (drug clearance) varies by at least20-fold. The level of CYP3A4 expressionmay determine which patients best respondto certain drugs and which patients experi-ence side effects or even toxicity when thesame dosage is given. The causes of thevariability in constitutive CYP3A4 areunknown, but a better understanding ofthese mechanisms may allow individualpatients to receive tailored drug dosages.Recent analyses suggest that, depending onthe drug, 60% to 90% of interpatient vari-ability in CYP3A4 function is caused bygenetic factors.
Polymorphisms in the coding regions ofCYP2C9, CYP2C19, and CYP2D6 under-lie the variability in expression of theseenzymes and have been associated withdistinct phenotypes throughout the popu-lation. The few reported mutations in thecoding region of CYP3A4, however, havenot been observed to have a profoundeffect on enzyme expression or function,but the possibility that other as-yet-unidentified polymorphisms affect itsexpression cannot be ruled out. The uni-modal distribution in drug clearance ofCYP3A4 substrates indicates that this islikely a multigenic process.
It has been proposed that the variabilityin CYP3A4 expression may be regulated at the transcriptional level, either throughpolymorphisms in upstream regulatory ele-ments or in the genes encoding the tran-scription factors themselves. Rifampin, ananti-infective agent that induces CYP3A4transcription, does so by activating amember of the nuclear receptor superfam-ily PXR (also called SXR or PAR).Rifampin is a ligand of PXR, bound at reg-ulatory regions of the CYP3A4 gene, allow-ing it to activate transcription. Targeteddisruption of mouse PXR abolishesCYP3A induction but does not affect the
level of constitutive CYP3A expression [8].
The publication of the map of humangenome sequence variation, reported tocontain 1.42 million SNPs, will allow a comprehensive search for genetic loci thatare likely to regulate expression of CYP3Aenzymes. Because DNA elements thatcontrol expression of CYP3A4 have beenlocalized to far upstream enhancers,searching for genetic polymorphism in reg-ulatory regions by traditional methods suchas sequencing is exhausting. The SNP mapwill facilitate the search for polymorphismsin distant regions [7].
“Polymorphisms in the CYP3A4 gene orother genetic loci controlling expressionand function of CYP3A4 may explain theperson-to-person variations seen in inten-sity and duration of drug action, as well asin the occurrence of side effects. Under-standing the genetic basis of differences inCYP3A function will allow us someday todetermine the proper drug dosages forindividual patients, and achieve an optimaltherapeutic response with minimal sideeffects” [8].
14.3.2. Cytochrome P450 SNP Could Be the Key to Variations inDrug Response
Researchers report that variations in thecytochrome P4503A5 gene, largely ignoredup until now, with CYP3A4 getting all the attention, determine how individualsrespond to drugs metabolized by CYPsystems in the liver and the intestine. Acommon variant leads to some individualsnot making a significant amount of theCYP3A5 protein [9].
The investigators used DNA in liversamples from diverse individuals to iden-tify the most common single nucleotidepolymorphisms in the CYP3A family ofgenes. They then looked for associationsbetween the SNPs and differences in drugmetabolism and clearance of the anestheticmidazolam by these individuals. They
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388 INDIVIDUALIZATION OF DRUG REGIMENS
found that one SNP, CYP3A5*3, results ina shorter, less active CYP3A5 protein.People with this SNP metabolize midazo-lam more slowly than do people with theCYP3A5*1 variant (which encodes a full-length protein). The researchers plan toundertake pharmacokinetic and pharma-codynamic studies of drugs metabolized bythe CYP3A family of enzymes to see howgenotype affects efficacy. The lead authorpredicted, “. . . in a reasonable amount oftime people will be doing targeted dosingbased on genotyping” [10].
14.3.3. Genotypic Differences inCYP2C19 May Modulate Dispositionof Drug Therapy for H. pyloriInfection and Cure Rates
Proton pump inhibitors (PPIs) such asomeprazole and lansoprazole are mainlymetabolized by cytochrome P450 2C19(CYP2C19), and genotype status may mod-ulate the therapeutic effects of theseagents. To explore this possibility, medicalinvestigators in Japan examined whetherCYP2C19 genotype status was related toeradication of H. pylori on treatment withtriple therapy—PPI, clarithromycin, andamoxicillin. They also attempted to estab-lish a treatment plan after failure to eradi-cate H. pylori. Eradication of H. pylori isthe most effective strategy known for thetreatment of peptic ulcer [11].
The study population consisted ofCYP2C19-genotyped patients infectedwith H. pylori who had completed initialtreatment with omeprazole 20mg or lan-soprazole 30mg twice daily, and clar-ithromycin 200mg and amoxicillin 500mgthree times a day for 1 week. Patients inwhom the infection was not eradicatedafter initial treatment were retreated withlansoprazole 30mg and amoxicillin 500mgfour times a day for 2 weeks.
The patients were classified into the fol-lowing three genotype groups: homozygousextensive metabolizers (n = 88), heterozy-
gous extensive metabolizers (n = 127),and poor metabolizers (n = 46). Overall,the infection was eliminated in 87% ofpatients. Eradication rates were 73%, 92%,and 98% in the homozygous extensive, het-erozygous extensive, and poor metabolizergroups, respectively. Thirty-four of 35patients without eradication had an exten-sive metabolizer genotype of CYP2C19,and 19 of those patients were infected with clarithromycin-resistant strains of H.pylori. The researchers found no amoxi-cillin-resistant strains. On retreatment withhigh-dose lansoprazole and high-doseamoxicillin, therapy succeeded in 30 of 31 patients with extensive metabolizergenotype of CYP2C19 who had failed torespond to initial treatment.
The investigators concluded: “Theresults of this study suggest that CYP2C19genotyping appears to be one of the pre-dictable determinants for a PPI-based H.pylori eradication therapy with the aid of bacterial sensitivity testing. . . . If theCYP2C19 genotype status is determinedbefore treatment, an optimal dose of a PPImay be prescribable on the basis of phar-macogenetic or pharmacogenomic status.This predetermined strategy should in-crease the eradication rates of H. pyloriachieved by the initial treatment . . .” [11].
14.3.4. Pharmacogenetics and Altered Drug Metabolism ofChemotherapeutic Agents for Cancer
One of the main messages of the 2001American Association of Cancer Researchmeeting was this: variability in response todrugs can lead to therapeutic failure oradverse drug reactions in individuals orsubpopulations of patients [12]. It is nowrecognized that all patients do not react inthe same way to chemotherapy, but in somecases treatment response can be predicted.Pharmacogenetics provides the opportu-nity to tailor drug treatments and decreaseuncertainties of therapy.
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14.3. PHARMACOGENETICS: DRUG METABOLISM AND TRANSPORT 389
Polymorphisms in cytochrome P450drug-metabolizing enzymes are associatedwith both efficacy and adverse drug reac-tions. One research group used a SNPscreening approach to study drug dis-position in previously untreated patientswith advanced breast cancer. The re-searchers found 22 unique SNPs in 12genes involved in drug metabolism. Theyalso found a statistically significant associ-ation of three genes with drug metabolismrates, tumor response, and patient survivalfollowing chemotherapy with high-dosecyclophosphamide, cisplatin, and carmus-tine. In particular, about one-quarter ofpatients with decreased metabolism ofcyclophosphamide correlated with a sig-nificantly shorter survival. Decreasedcyclophosphamide metabolism was linkedto a SNP in the promoter region ofCYP3A4 or CYP3A5. These findings couldlead potentially to a simple pretreatmentgenotype analysis to determine what doseof drug or even which drug combinationwould be most effective for each patient.
Oncologists are moving closer to corre-lating genetic data with phenotypic dataand using this information to help forecasta patient’s response to chemotherapy andpredict toxic effects. A medical center inAlabama is already testing patients withcolorectal cancer for polymorphisms in the gene involved in the metabolism of fluorouracil before initiating treatmentwith the drug. In particular, scientists at the center look for polymorphisms in the gene coding for dihydropyrimidine dehydrogenase.
14.3.5. Genetic Polymorphisms ofAlcohol Dehydrogenase ModulatesAlcohol Disposition and May Relate to Increased Risk ofMyocardial Infarction
Moderate alcohol consumption is consis-tently associated with a reduced risk of
myocardial infarction (MI). Studies ofalcohol metabolism have shown that theclass I alcohol dehydrogenase (ADH)isoenzymes, encoded by ADH1, ADH2,and ADH3, oxidize ethanol and other smallaliphatic alcohols. ADH2 and ADH3 havepolymorphisms that produce isoenzymeswith distinct kinetic properties. AmongWhite populations, however, variant allelesare relatively uncommon at the ADH2locus (present in less than 10% of the pop-ulation) but common at the ADH3 locus(present in 40–50%). At the ADH3 locus,the g1 allele differs from the g2 allele by two amino acids. Pharmacokinetic studiesshow a 2.5-fold difference in the maximalvelocity of ethanol oxidation between the homodimeric g1 isoenzyme (associatedwith a fast rate) and the homodimeric g2
isoenzyme (associated with a slow rate).Effects of moderate alcohol consump-
tion on the risk of MI and the role ofADH3 genotype have been evaluated.Moderate drinkers with the g2 variant werefound to have a lower risk of heart attackthan those who disposed of alcohol morequickly [13].
The investigators identified 396 patientswith newly diagnosed cases of acute MIamong men in the Physician’s HealthStudy. The patients were matched with oneor two controls. The researchers deter-mined the ADH3 genotype (g1g1, g1g2, org2g2) in all subjects.
As compared with men homozygous forthe allele associated with the fast rate ofethanol oxidation (g1), those homozygousfor the allele associated with a slow rate ofethanol oxidation (g2) had a significantlyreduced risk of acute MI (relative risk,0.65). Moderate alcohol consumption wasassociated with a lower risk of a heartattack in all three genotype groups. Amongmen who were homozygous for the g1
allele, those who consumed at least onedrink per day had a relative risk of MI of0.62, as compared with the risk among menwho consumed less than one drink per
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week. Men who consumed at least onedrink per day and were homozygous forthe g2 allele had the greatest reduction inrisk (relative risk, 0.14).These men also hadthe highest plasma HDL levels, which seemto protect against coronary heart disease.The researchers confirmed the interactionamong the ADH3 genotype, the level ofalcohol consumption, and the HDL level in an independent study of postmenopau-sal women. They concluded, “Moderatedrinkers who are homozygous for the slow-oxidizing ADH3 allele have higher HDLlevels and substantially decreased risk ofmyocardial infarction” [13].
14.3.6. Cellular Transporters and Drug Resistance
About three-quarters of cancer patients areintrinsically unresponsive to or developresistance to anticancer drugs. In epilepsy,the situation is also problematic as the entirearsenal of anticonvulsant drugs fails up to30% of patients.Fortunately solutions in thearea of cancer research are emerging.Scien-tists have developed tests that can predictwhether a patient is likely to respond to themost common chemotherapeutic agents.There is now the potential of developingnovel drugs that, given with currentchemotherapy, can overcome multidrugresistance. Researchers are hoping that,should newly emerging theories on drugresistance hold true, the progress made inthe cancer field may lead to benefits forother drug-resistant diseases.
While there seems to be little incommon between intractable epilepsy andcancer, researchers in each field havefocused on a common mechanism underly-ing multidrug resistance: a cellular pumpcalled P-glycoprotein (Pgp). Pgp protectscells from toxic substances by activelyexcreting the toxic agent.The pumps residein tissues that are extensively exposed totoxic material: liver, lungs, kidney, intestine,placenta, and blood-brain barrier. How-
ever, the pumps not only protect tissue butalso influence the uptake of drugs. Almost30% of medicinal agents are thought to besubstrates of Pgp and thereby challenged inreaching target sites.
A link between Pgp and drug-resistantepilepsy was reported in 1999, whenresearchers found high levels of the Pgpgene, MDR1, in brain lesions of a 4-month-old patient unresponsive to drug therapy.This finding may explain why anticonvul-sant drugs such as phenytoin and pheno-barbital, both of which are Pgp substrates,may not reach pharmacologically effectiveconcentrations in critical parts of the brainin drug-resistant patients. This phenome-non is well known in oncology.
Applying a pharmacogenetic approach,researchers have identified 15 polymor-phisms for MDR1, only one of which cor-relates with poor drug uptake. A test forthis polymorphism could allow physiciansto predict drug uptake from the outset and eliminate the administration of use-less therapy. There is, however, a largerproblem in that chemotherapy itselfinduces Pgp expression. Stifling the Pgptransporter with an inhibitor could resolveboth issues. A newly discovered, orallyeffective molecule, OC144-093, is enteringphase II trials. Not only is this agent apotent Pgp inhibitor, so far it also appearsto be safe.Tempering enthusiasm, however,is the understanding that Pgp expression is but one of several mechanisms of drugresistance identified by cancer researchers.
Other researchers are studying poly-morphisms and ethnic variation. In onestudy 1280 people from 10 different ethnicgroups were evaluated for the C3435Tpolymorphism in the gene coding for Pgp.The investigators found that while 20% to47% of White and Asian people werehomozygous for the polymorphism leadingto underexpression of Pgp and sensitivityto Pgp-dependent drugs, less than 6% ofAfricans had this genotype. These differ-ences may have important implications for
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use of Pgp-dependent drugs in cancerpatients of African origin [12].
14.3.7. MDR1 Expression PredictsOutcome of Liver Transplantation
Living-donor liver transplantation andimmunosuppressive therapy with tacro-limus (Prograf) are key elements in therecovery of patients from end-stage liverfailure. The optimal tacrolimus dosageregimen, however, has not been estab-lished, because in large measure thebioavailability of orally administeredtacrolimus is highly variable. In this light,researchers have examined whether barri-ers to tacrolimus absorption—multidrugresistance protein (MDR1) or cytochromeP450 3A4 (CYP3A4)—are important phar-macokinetic factors and prognostic indica-tors of liver transplantation outcome.
The investigators measured messengerribonucleic acid (mRNA) expression levelsof MDR1 and CYP3A4 in mucosal cells ofthe upper jejunum collected during living-donor liver transplantation in 48 recipients.Tacrolimus was initiated at an oral dose of0.075mg/kg every 12 hours and adjusted on the basis of trough levels in wholeblood.
The mRNA expression level of MDR1,but not the level of CYP3A4, was inverselyrelated to the concentration/dose ratio oftacrolimus. High levels of MDR1 expres-sion were associated with low bioavailabil-ity and reductions in survival rates aftertransplantation. Achieving a tacrolimustrough level of about 11ng/ml required adaily dose of 0.13mg/kg in the high-MDR1group and 0.07mg/kg in the low-MDR1group. For the overall study population,the 600-day survival rates in the high- andlow-MDR1 groups were 70.8% and 95.6%,respectively. The findings suggest thatintestinal MDR1 variability should betaken into consideration for tacrolimustherapy, especially the determination of theinitial dosage regimen, as a pharmacoki-
netic and prognostic factor for live-donorliver transplantation recipients [14].
�14.4. PHARMACOGENETICS:THERAPEUTIC RESPONSE
14.4.1. SNPs Predispose to Cancerand Alter Response to Therapy
Alteration of a single nucleotide in a genecan determine whether the gene will pre-dispose an individual to developing cancer.At the same time these SNPs can alter genefunction in ways that affect response totherapy or chemoprophylaxis.
Researchers (at an American Associa-tion for Cancer Research conference onGenetic Modification of Cancer Suscepti-bility, held in 2001) suggested that this isthe case for SRD5A2 gene variants, whichencode steroid type 2,5-alpha reductase,an enzyme that catalyzes the conversion of testosterone to dihydrotestosterone(DHT). They proposed that increases inthe activity of this enzyme, resulting inhigher levels of DHT, increase a man’s likelihood of developing benign prostatichypertrophy (BPH) and prostate cancer.Their experiments revealed that a particu-lar variant of SRD5A2 was associated with a sevenfold increased risk of prostatecancer in African-American men, a four-fold risk in Hispanic men, and a threefoldrisk in White men.
The investigators found that thedescribed 10 SNP variants of SRD5A2differ in their biologic effects, not only inhow much they affect levels of DHT,but also in their response to finasteride,a competitive inhibitor of 2,5-alpha reduc-tase. Finasteride (Proscar) is marketed for the treatment of BPH and is underinvestigation for the chemoprevention ofprostate cancer. The investigators found a200-fold range in activity of the enzymesencoded by these gene variants and as muchas a 60-fold difference in the ability of finas-teride to inhibit the enzyme’s activity.
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Information about the pharmacogeneticvariation in the SRD5A2 gene was notavailable in 1993, when the NationalCancer Institute-sponsored ProstateCancer Prevention Trial began recruiting18,000 participants to determine whetherfinasteride can prevent prostate cancer inmen 55 years and older. All of the partici-pants in the active treatment arm of theplacebo-controlled trial are receiving thesame dosage 5mg daily finasteride. Whenthe trial ends in 2003, the investigatorshope to determine whether finasteride hasa role in preventing prostate cancer.Absence of information on the partici-pants’ SRD5A2 genotype, however, maymake it more difficult to interpret the findings.
Inevitably some of the men given finas-teride will develop prostate cancer. Wouldthose who had SRD5A2 variants producinga weak response to finasteride have faredbetter if they had been given a higherdosage of the drug? As the Prostate CancerPrevention Trial draws to a close, it may befruitful to explore this possibility by geno-typing individual participants for SRD5A2and assessing whether certain genotypesare linked to a particular response to finas-teride [15].
14.4.2. Altered Response toChemotherapy in Women with BRCA1 Mutation
According to findings presented at the 2001annual meeting of the American Associa-tion for Cancer Research, women whoharbor mutations in BRCA1, a gene implicated in some types of hereditarybreast cancer, may respond differently tochemotherapy than those lacking suchmutations. The BRCA1 gene is believed to function normally as a tumor suppres-sor and transcriptional (DNA to RNA)regulator. Moreover it encodes a proteininvolved in the cellular response to DNAdamage. BRCA1 mutations increase the
susceptibility to breast, ovarian, andprostate cancer. The frequency of BRCA1mutations in the general population is 1 in833, and in Ashkenazi Jewish women 1 in107. Women with BRCA1 mutations face a50% to 80% lifetime risk of developingbreast cancer and develop it at an earlierage than those without such mutations.
Investigators found that human breastcancer cell lines with BRCA1 mutationsshowed a twofold to fourfold increase inapoptosis after treatment with ionizingradiation, cisplatin, or doxorubicin, com-pared with cells free of mutations.They alsofound that BRCA1 tumor cell lines wereresistant to other agents, such as paclitaxel(Taxol) and docetaxel (Taxotere), treat-ments used commonly in ovarian cancerand advanced-stage breast cancers.
Differences in drug sensitivity could betraced to the levels of another protein,Bcl2, which is also implicated in apoptosis.Loss of Bcl2 results in higher levels of pro-grammed cell death after DNA damage.Normal BRCA1 regulates the expressionof Bcl2. Breast tumors expressing BRCA1mutations lacked or had reduced levels ofthe Bcl2 protein and, consistently, wereresistant to chemotherapy with taxanes,which induce cell death through a Bcl2pathway. They remained susceptible tochemotherapy that interacted directly withDNA, independent of Bcl2.
Hence women with BRCA1mutationswho develop breast or ovarian cancer maynot be good candidates for treatment withcertain types of chemotherapy agents suchas Taxol. Doxorubicin, cisplatin, or othernewly discovered agents might be a betterchoice. In the future, genetic profiling may point the way to optimal treatment ofcancer.
14.4.3. Pharmacogenetics andMethotrexate Toxicity
Linking pharmacogenetics and adversedrug reactions has proved fruitful in some
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instances. Methotrexate, an antifolate oftenused to prevent graft-versus-host diseaseafter bone marrow transplantation (BMT),has several side effects such as oral muco-sitis, which can impede eating, talking, oreven breathing.
Methylenetetrahydrofolate reductase(MTHFR) is a key enzyme in folate metab-olism. Researchers have investigatedwhether the 677 CÆT polymorphism in theMTHFR gene, which leads to an amino-acid substitution and decreased enzymeactivity, modifies toxicity to methotrexatein BMT patients. Patients with lowerMTHFR activity (homozygous for thepolymorphism) had 36% higher mean oral mucositis than those with the wild-type genotype, as well as 34% slower recovery of platelet counts. Seeminglypatients homozygous for C677T showhigher methotrexate toxicity because ofdecreased nucleotide synthesis, resulting ina decreased DNA repair capacity and, con-sequently, delayed healing of the mucosa.Patients with polymorphisms in theMTHFR gene may be candidates for doseadjustment or use of alternative drugs [12].
14.4.4. b-Blocker Response in HeartFailure May Have Genetic Link
Activation of the renin-angiotensin systemand elevation of circulating catecholaminesboth contribute to heart failure progres-sion. Angiotensin converting enzyme(ACE) inhibitors and b-adrenergic recep-tor antagonists are mainstays of therapy. Aconsiderable portion of the variability inACE activity is genetically based and hasbeen linked to a common polymorphism ofthe ACE gene. The mutation is known asthe ACE deletion (ACE D) because it lacksa piece of DNA found in the normal gene. Although the mutation increasesrenin-angiotensin activation, its influenceon patient outcome has been uncertain,and its pharmacogenomic interactions withb-blockers have not been studied.
A recent study suggests that patientswith congestive heart failure who carry theACE deletion are likely to respond espe-cially well to b-blockers. The researchersfollowed 328 men and women with heartfailure (mean left ventricular ejection fraction, 0.24) to assess the impact of theACE D allele on transplant-free survival.Patients with two ACE deletion (short)versions of the ACE gene (DD) had thehighest levels of ACE activity, and patientswith two insertion (long) versions of thegene (II) had the lowest levels.
The adverse impact of the deletion onheart failure progression was dramatic inpatients not treated with b-blockers.Two-year patient survival was 81% in ACE insertion, II genotype; 48% in ACEdeletion, DD genotype; and 61% in theheterozygote, ID genotype. In contrast,patients who received b-blocker therapyshowed no influence of the ACE genotypeon transplant-free survival. Two-yearpatient survival was 70% in the II, 71% inthe ID, and 77% in the DD genotypes.
These data clearly linked the ACE Dallele with significantly poorer transplant-free survival. This effect was primarilyevident in patients not treated with b-blockers and was not seen in patientsreceiving adrenergic therapy. These find-ings suggest a potential pharmacogeneticinteraction between the ACE D/I poly-morphism and therapy with b-blockers inthe determination of heart failure survival[16].
14.4.5. Stent Restenosis, ACEInhibitors, and Genetic Polymorphism
Some patients seem to have an inherentgenetic susceptibility to restenosis follow-ing coronary stent implantation. Researchhas shown that patients homozygous forthe deletion (D) allele of angiotensin converting enzyme (ACE) gene polymor-phism lack functional enzyme and are atincreased risk. These patients have ele-
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vated concentrations of ACE that couldcontribute to an accelerated growthresponse of smooth-muscle cells, leading toloss of patency.Thus one might suspect thatACE inhibition could modify the conse-quences of these increased ACE levels inDD patients. A recent report concerns thefindings of a well-controlled pharmacoge-netic trial to establish whether blockade ofACE with high doses of the ACE inhibitorquinapril (Accupril) could limit neointimalproliferation and reduce restenosis afterstent implantation in patients carrying theDD genotype [17].
The ACE gene insertion/deletion (I/D)polymorphism was characterized in 345patients who were undergoing coronarystenting. Among them, 115 had the DDgenotype. The investigators assigned 91 ofthese patients to quinapril 40mg daily orplacebo. Treatment was initiated within 48 hours after stent implantation and con-tinued for 6 months.
Surprisingly the primary end point ofloss in minimum lumen diameter—a quantitative index of restenosis—was sig-nificantly higher in the quinapril groupthan in the placebo group. Secondary endpoints also showed consistent trendstoward increased restenosis in the treat-ment group. The authors of the report con-cluded: “Contrary to our expectations,ACE inhibitor treatment did not reducerestenosis after coronary stent implanta-tion in patients with DD genotype, but was associated with an exaggerated restenoticprocess when compared with administra-tion of placebo [17].”
ACE inhibitors are widely prescribed forpatients with atherosclerosis, hypertension,or diabetes,and after myocardial infarction,because of proven beneficial effects in eachof these groups. In light of the results in thesetting of stent restenosis, whether a patientwith hypertension or some other target dis-order who carries the DD genotype derivesless benefit from ACE inhibitor therapywarrants consideration.
14.4.6. Prothrombotic MutationsIncrease Risk of Acute MI in WomenTaking HRT
Observational studies have suggested thatthe use of hormone replacement therapy(HRT) in postmenopausal women reducesthe risk of coronary heart disease and haveprompted the prescribing of HRT for thispurpose. But estrogens are also prothrom-botic, and high doses are associated withcomplications that include myocardial in-farction, stroke, and venous thrombosis.The results of the Heart and Estrogen/Progestin Replacement Study (HERS), asecondary prevention trial that showedcombined HRT was certainly no betterthan placebo at preventing coronary eventsin postmenopausal women, have renewedinterest in the potential adverse effects ofHRT. Indeed, in post hoc analyses ofHERS data, treatment was associated with harm during the first year of follow-upand with benefits in years 4 and 5 of thestudy.
To explain this pattern, researchershypothesized an immediate prothrombotic,proarrhythmic, or proischemic effect oftreatment that is gradually outweighed bya beneficial effect on the underlying pro-gression of atherosclerosis. According tothis interpretation, a subgroup of patients,perhaps defined by a clinical characteristic,an environmental exposure, or a genetictrait, is susceptible to an early adverseeffect of estrogen, while the rest of the population benefits from estrogen therapy.
In light of earlier evidence that geneticvariations in prothrombin of human sub-jects are clearly associated with the risk ofvenous thrombosis, and before the resultsof HERS were known, investigators initi-ated a case-control study to assess whetherprothrombotic mutations modify the asso-ciation between HRT and the incidence offirst MI. Cases were 232 postmenopausalwomen aged 50 to 79 years who had theirfirst nonfatal MI between 1995 and 1998.
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14.5. INDIVIDUALIZED GENE-BASED MEDICINE: A MIXED BLESSING 395
Controls were a sample of 723 post-menopausal women without MI who werematched to cases by age, calendar year, andhypertension status. The main outcomemeasure was risk of first nonfatal MI based on current use of HRT and the pres-ence or absence of coagulation factor VLeiden and prothrombin 20210 GÆA vari-ants among cases and controls, stratified byhypertension.
In the study populations, 108 MI casesand 387 controls had hypertension. Amongwomen with hypertension, the prothrom-bin variant was a risk factor for MI (oddsratio: 4.32). Compared with nonusers ofHRT with wild-type genotype, women whowere current users and who had the pro-thrombin variant had a nearly 11-foldincrease in risk of a nonfatal MI. The interaction was absent among normo-tensive women. No interaction was foundfor factor V Leiden in either hypertensiveor normotensive women.
The authors suggest that if their findingsare confirmed, screening for the prothrom-bin 20210 GÆA variant may permit a betterassessment of the risks and benefits associ-ated with HRT in postmenopausal women.It is important to note, however, that theprothrombin variant would account foronly part of the pattern of early harm andlate benefit seen in the HERS trials [18].
�14.5. INDIVIDUALIZED GENE-BASEDMEDICINE: A MIXED BLESSING
Thanks to receptive media, scientific andmedical discoveries have tantalized us withthe prospect of individualized medicine in which drugs are prescribed based oneach person’s genetic makeup.A significantnumber of genes relating to human medicalconditions has now been identified, andtheir location in chromosomes has beenmapped. These details are now available asa part of the human genome project(Figure 14.1). This knowledge can now be
put to use in developing genetic tests aimedat improving therapeutic management andpreventing medical disorders and diseases.The technology for genetic testing is available and finding use, albeit limited. Insome quarters this development is seen asa threat to the pharmaceutical industry,because it limits the target population for adrug and makes it more difficult to launcha drug destined to have blockbuster sales.
One company, Genaissance Pharmaceu-ticals, has invested heavily in genetic testingequipment in the hope of developing bloodtests that, by providing a patient’s geneticprofile, would suggest which medicine has the best chance of working. One of its first targets is asthma medication. Glaxo-SmithKline, however, has filed for patentson genetic tests for the effectiveness of itsasthma medication fluticasone (Flovent).Observers surmise that GlaxoSmithKlinehas no intention of having the tests see thelight of day nor allowing Genaissance or any other company to develop them.
The head of genetics at Glaxo said:“They can go screw with someone else’sdrugs” [The Wall Street Journal InteractiveEdition, 18 June 2001]. GlaxoSmithKline isnot opposed to all such tests. The companyplans to use them to identify patients likelyto suffer serious side effects from certaindrugs and exclude them from clinical trials.
Despite the roadblocks, Genaissance ismoving forward. Its asthma trial willinvolve 700 patients and test five drugs:Serevent, Flovent, Pulmicort, Accolate, andSingulair. Using complex algorithms, theinvestigators will attempt to correlate testresults with how patients respond to eachdrug, taking into account variations indozen of genes. A pilot study of albuterolin 121 asthma patients showed that thedrug failed to produce meaningful bron-chodilation in 11% of patients, who hadone type of genetic variation, but signifi-cantly improved breathing in 28% of thegroup, who had other genetic variations[19].
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�14.6. CURRENT AND POTENTIALAPPLICATION OFPHARMACOGENETICS
The promise of pharmacogenetics has beenrealized for only a few drugs. One reasonfor the slow pace of development is thatmuch of the science needed to practicegene-based medicine needs to be worked
out. There is also caution because theconcept is unconventional and widely perceived as adding to the cost of healthcare. Many clinicians view individualizeddrug therapy as impractical and questionwhether such an approach may ever berealized in light of the delay in making adecision and the cost of implementinggenotyping or other tests to measure surrogate markers of gene function.
IL2RGX-linked severe combinedimmunodeficiency (SCID)TNFSF5Immunodeficiency with hyper-IgMFMR1Fragile X syndromeMeCP2Rett syndromeALDAdrenoleukodystrophyHEMAHemophillia A
Figure 14.1. Human chromosomes and locations of genes linked to medical conditions. For some ofthe genes identified, recombinant DNA encoding the protein’s RNA transcript and the amino-acidsequences, as well as the regulatory DNA sequences—including promoter binding regions, exons andintrons, and splicing information—are reported and compiled in the National Center for BiotechnologyInformation (NCBI) database. (Figure reprinted from the noncopyrighted information in the database)
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known as trastuzumab (Herceptin) for thetreatment of patients with metastaticbreast cancer whose tumors overexpresserbB-2 (HER2) protein.
Only 20% to 33% of metastatic breastcarcinomas robustly express HER2, andtumors that do not overexpress the proteinare unlikely to benefit from treatment.Hence demonstration of clinical benefit inunselected breast cancer patients was achallenging task [24]. The need to keeppatients on other chemotherapies addedto the complexity of clinical trials. Never-theless, the decision to first demonstrateoverexpression of HER2 antigen levels asone of the key clinical trial inclusion crite-ria, reached in consultation with the FDA,allowed the investigators to demonstrateefficacy (Table 14.2).
The data on Herceptin—alone or in com-bination with current chemotherapies formetastatic breast cancer—clearly demon-strate that the degree of HER2 overexpres-sion is related to treatment response.Inhibition of this tumor-transforminggrowth factor has proved to produce clini-cal effects but only in those patients withoverexpressed phenotypes (2+ and 3+).These results formed the basis for therequirement to determine levels of HER2 on tumor tissues before deciding whetherHerceptin is the best treatment strategy for an individual patient.
14.6. CURRENT AND POTENTIAL APPLICATION OF PHARMACOGENETICS 397
While no biopharmaceutical is approvedwith the requirement of genotyping as apart of its therapeutic indication, a recently approved monoclonal antibodytherapy against breast cancer, trastuzumab(Herceptin), is indicated only for thosetumors measurably expressing the proteinexpressed by the gene erbB-2. Because theoncogene product of erbB-2 is elevated
BOX 14.1. HOW HERCEPTIN WAS APPROVED AS A THERAPY THAT REQUIRESHER2 EXPRESSION TESTS ON PATIENT TUMOR TISSUES BEFORE USE
in no more than 30% of breast cancerpatients, patients with no erbB-2 (HER2)protein in metastatic cancer cells derive nobenefit from treatment with Herceptin.HER2 antigen testing was integral to thedevelopment of this product, and Her-ceptin’s label directs that its use be basedon a commercially available diagnostic test called Her2/neu (Box 14.1). Therefore
The search for genes that transform normalcells to tumor cells (oncogene research) inthe 1980s led to the discovery of tumor-causing viral proteins capable of enhancingthe growth rate of normal cells. Two viraloncogenes, erbA and erbB, identified fromavian erythroblastosis virus, were shown toderegulate cell growth through receptormodifications (glycosylation and phospho-rylation of the proteins) [20,21]. A cell homo-logue of viral erbB was later shown to be aproto-oncogene product that is a truncatedform of epidermal growth factor (EGF). Cellsthat express erbB exhibit uncontrolledgrowth due to the lack of EGF receptor-regulated functions. This is proposed to be amechanism of tumorogenesis [22]. Aboutfour years after discovery of cellular erbBexpression, antibodies were developed torecognize erbB-2. By using these antibodies,researchers learned that about 10% ofmalignant tumor tissues collected fromhuman organs overexpressed erbB-2.
By early 1990 Genentech had reportedthe development of murine monoclonalantibodies with therapeutic potentialagainst tumor cells that overexpress erbB-2. One of these clones was humanized toproduce a mouse/human chimeric formtermed 4D5 and determined to be a suit-able candidate for preclinical and clinicalstudies [23]. In 1998 the FDA approved thehumanized monoclonal antibody, now
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identifying the right breast cancer patientfor Herceptin therapy is the first exampleof individualized drug therapy. Herceptintherapy, requiring prescreening of apatient’s genetic makeup, is a significantstep toward acceptance by clinicians of anew clinical practice paradigm.
Other examples illustrating that gene-based medicine can improve clinical out-comes are less widely used or less heraldedbut are important. Prescribing a smallerdose of azathioprine to those who lackthiopurine methyltransferase, the primarymetabolic enzyme for this immunosup-pressant, reduces the risk of toxicity inpatients undergoing renal transplantation.As the toxicity of azathioprine is severe,and sometimes fatal, pharmacogenetictesting is becoming part of the standard ofcare in azathioprine therapy. Individualswho lack glucose-6-phosphate dehydroge-nase show increased sensitivity towarddapsone-related hemolytic anemia. Somephysicians routinely screen for this charac-teristic before prescribing dapsone to treat Pneumocystis carinii pneumonia inpatients with AIDS. In each case, thebenefit-to-risk ratio is poor or unaccept-able without a test predictive for outcomes.
Despite its seeming benefit to individualpatients, pharmacogenetics has not beenwidely embraced by the pharmaceuticalindustry. Certainly pharmacogenetic principles were applied in the develop-ment of the anticancer drugs Herceptin and
Gleevec, both of which target a mutatedgene or gene product. But these are excep-tions. Pharmaceutical companies strive todevelop drugs that are effective in as largea population as possible to reap maximumprofits. Many currently available drugs areeffective in no more than half of the targetpopulation. No pharmaceutical companydesires to be told to develop a pharmaco-genetic test to determine which patient willbenefit from their product and which willnot. This would be a time-consuming taskand limit the market for their product. Thesame is true for the segment of the targetpopulation that displays adverse effects,unless they are serious ones.
Pharmaceutical companies will continueto develop targeted drugs like Herceptin asthey accept the fact that, though the userpopulation is limited, these drugs are likelyto be profitable. Routine application ofpharmacogenetics to identify patients whowill safely benefit from a drug, however, isa long way off.The regulatory environmentmust change to accommodate this para-digm shift, the linkage between geneticsand therapeutic response must be mademanifest, and the cost of this strategy mustbe commensurate with its value.
�14.7. SUMMARY
Individualization of drug therapy to en-sure optimal therapeutic response is a goal
�TABLE 14.2. Levels of HER2 antigen expression and overall response rate toHerceptin
Patient Paclitaxel Anthracycline +HER2 and Anthracycline + CyclophosphamideLevelsa Herceptin Paclitaxel Herceptin Cyclophosphamide and Herceptin
2+ 4% 16% 21% 43% 40%3+ 17% 14% 44% 35% 53%
Data source: Biologic License Application to FDA.aPatient tissues were evaluated using an immunohistochemical staining for HER2 proteins based on a 0–3scale where 0 = none detected and 3+ being strongly positive. Patient tissues with 2+ scores are consideredweakly positive for HER2 expression.
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REFERENCES 399
within reach. Application of pharma-cogenetics in clinical practice is in itsinfancy, but there is every reason to thinkit will mature. Some unbridled enthusiastsbelieve that a day will come when eachperson carries a gene chip identificationcard, analogous to electronic IDs, or evenan implanted chip, that contains completegenetic information, information that canbe used for therapeutic decision-making.The technical feasibility, however, must bematched by a paradigm shift in clinicalpractice and an acute sensitivity to ethicalconcerns before that day comes. As thestudy and application of pharmacogeneticsmatures, associations of genetic variationslinked to therapeutic response need to bevalidated, and genotyping assay must beimproved to be more “user friendly” (i.e.,rapid, sensitive, reproducible, readily avail-able, simple to use, and cost effective). Asexemplified by the use of Herceptin, newdrugs will increasingly be targeted to onlysome patients, those who display a particu-lar genetic marker that critically influencesefficacy and safety.
REFERENCES
1. Brewer, T., and G.A. Colditz, Postmarketingsurveillance and adverse drug reactions:current perspectives and future needs. Jama,1999. 281(9): 824–9.
2. Dormann, H., U. Muth-Selbach, S. Krebs, M.Criegee-Rieck, I. Tegeder, H.T. Schneider,E.G. Hahn, M. Levy, K. Brune, and G.Geisslinger, Incidence and costs of adversedrug reactions during hospitalisation:computerised monitoring versus stimulatedspontaneous reporting. Drug Saf, 2000.22(2): 161–8.
3. Roses, A.D., Pharmacogenetics and pharmacogenomics in the discovery anddevelopment of medicines. Novartis FoundSymp, 2000. 229: 63–6; discussion 66–70.
4. Weber, W.W., The history of pharmacoge-netics. Pharmaceutical News, 2000. 7(6):18.
5. Mahgoub, A., J.R. Idle, L.G. Dring, R.Lancaster, and R.L. Smith, Polymorphichydroxylation of Debrisoquine in man.Lancet, 1977. 2(8038): 584–6.
6. Scarlett, L.A., S. Madani, D.D. Shen, andR.J. Ho, Development and characterizationof a rapid and comprehensive genotypingassay to detect the most common variants incytochrome P450 2D6. Pharm Res, 2000.17(2): 242–6.
7. Katz, D., The promise of pharmacogenetics.Pharmaceutical News, 2001. 7(6): 47–52.
8. Eichelbaum, M., and O. Burk, CYP3Agenetics in drug metabolism. Nat Med, 2001.7(3): 285–7.
9. Kuehl, P., J. Zhang, Y. Lin, et al., Sequencediversity in CYP3A promoters and charac-terization of the genetic basis of polymorphicCYP3A5 expression. Nat Genet, 2001. 27(4):383–91.
10. Larkin, M., Key to drug response may be inthe SNP. Lancet, 2001. 357: 1020.
11. Klein, E., and R.J. Ho, Challenges in thedevelopment of an effective HIV vaccine:current approaches and future directions.Clin Ther, 2000. 22(3): 295–314; discussion265.
12. Hutchinson, E., Working towards tailoredtherapy for cancer. Lancet, 2001. 357(9267):1508.
13. Hines, L.M., M.J. Stampfer, J. Ma, J.M.Gaziano, P.M. Ridker, S.E. Hankinson, F.Sacks, E.B. Rimm, and D.J. Hunter, Geneticvariation in alcohol dehydrogenase and the beneficial effect of moderate alcohol consumption on myocardial infarction. NEngl J Med, 2001. 344(8): 549–55.
14. Hashida, T., S. Masuda, S. Uemoto, H. Saito,K. Tanaka, and K. Inui, Pharmacokineticand prognostic significance of intestinalMDR1 expression in recipients of living-donor liver transplantation. Clin PharmacolTher, 2001. 69(5): 308–16.
15. Stephenson, J., As genes differ, so shouldinterventions for cancer. Jama, 2001.285(14): 1829–30.
16. McNamara, D.M., R. Holubkov, K. Janosko,A. Palmer, J.J. Wang, G.A. MacGowan, S.Murali, W.D. Rosenblum, B. London, andA.M. Feldman, Pharmacogenetic interac-tions between beta-blocker therapy and the angiotensin-converting enzyme deletion
BIO14 5/2/03 2:47 PM Page 399
400 INDIVIDUALIZATION OF DRUG REGIMENS
polymorphism in patients with congestiveheart failure. Circulation, 2001. 103(12):1644–8.
17. Meurice, T., C. Bauters, X. Hermant, V.Codron, E. VanBelle, E.P. Mc Fadden, J.Lablanche, M.E. Bertrand, and P. Amouyel,Effect of ACE inhibitors on angiographicrestenosis after coronary stenting (PARIS):a randomised, double-blind, placebo-controlled trial. Lancet, 2001. 357(9265):1321–4.
18. Psaty, B.M., N.L. Smith, R.N. Lemaitre, H.L.Vos, S.R. Heckbert, A.Z. LaCroix, and F.R.Rosendaal, Hormone replacement therapy,prothrombotic mutations, and the risk ofincident nonfatal myocardial infarction inpostmenopausal women. Jama, 2001. 285(7):906–13.
19. Drysdale, C.M., D.W. McGraw, C.B. Stack,J.C. Stephens, R.S. Judson, K. Nandabalan,K. Arnold, G. Ruano, and S.B. Liggett,Complex promoter and coding region beta 2-adrenergic receptor haplotypes alterreceptor expression and predict in vivoresponsiveness. Proc Natl Acad Sci U S A,2000. 97(19): 10483–8.
20. Frykberg, L., S. Palmieri, H. Beug, T. Graf,M.J. Hayman, and B. Vennstrom, Trans-forming capacities of avian erythroblastosisvirus mutants deleted in the erbA or erbBoncogenes. Cell, 1983. 32(1): 227–38.
21. Hayman, M.J., G.M. Ramsay, K. Savin, G.Kitchener, T. Graf, and H. Beug, Identifi-cation and characterization of the avian erythroblastosis virus erbB gene product
as a membrane glycoprotein. Cell, 1983.32(2): 579–88.
22. Coussens, L., C. Van Beveren, D. Smith, E.Chen, R.L. Mitchell, C.M. Isacke, I.M.Verma, and A. Ullrich, Structural alterationof viral homologue of receptor proto-oncogene fms at carboxyl terminus. Nature,1986. 320(6059): 277–80.
23. Lewis, G.D., I. Figari, B. Fendly, W.L. Wong,P. Carter, C. Gorman, and H.M. Shepard,Differential responses of human tumor celllines to anti-p185HER2 monoclonal anti-bodies. Cancer Immunol Immunother,1993. 37(4): 255–63.
24. Pegram, M.D., G. Pauletti, and D.J. Slamon,HER-2/neu as a predictive marker ofresponse to breast cancer therapy. BreastCancer Res Treat, 1998. 52(1–3): 65–77.
25. Meyer, U.A., Pharmacogenetics and adversedrug reactions. Lancet, 2000. 356(9242):1667–71.
26. Phillips, K.A., D.L. Veenstra, E. Oren, J.K.Lee, and W. Sadee, Potential role of pharmacogenomics in reducing adversedrug reactions: a systematic review. Jama,2001. 286(18): 2270–9.
27. Ingelman-Sundberg, M., A.K. Daly, M.Oscarson, and D.W. Nebert, Humancytochrome P450 (CYP) genes: recom-mendations for the nomenclature of alleles.Pharmacogenetics, 2000. 10(1): 91–3.
28. Ingelman-Sundberg, M., Genetic susceptibil-ity to adverse effects of drugs and environ-mental toxicants. The role of the CYP familyof enzymes. Mutat Res, 2001. 482(1–2): 11–9.
