中國醫藥大學藥學系洪靚娟. Objectives Provide an overview of pharmacogenomics and its clinical relevance Discuss clinically-relevant examples of: Drug metabolism

中國醫藥大學藥學系洪靚娟

ObjectivesProvide an overview of pharmacogenomics

and its clinical relevance

Discuss clinically-relevant examples of:Drug metabolism pharmacogenomicsDrug target pharmacogenomics

Discuss the challenges facing pharmacogenomic studies and the movement of pharmacogenomics into clinical practice

Introduction and Background

Pharmacogenetics vs. Pharmacogenomics generally defined as:

the study of the relationship between genetics and drug effect

the application of genetic analysis to predict drug response, efficacy, and toxicity

Introduction and Background Pharmacogenetics vs. Pharmacogenomics

Key differences: pharmacogenetics

focused on variation in individual, specific genes that influence the response to a drug

associated with: a large clinical effect mutation in a single gene affects a relatively small number of individuals


Key differences: pharmacogenomics

focused on variation in a large collection of genes, up to the whole genome, that influence response to a drug

associated with: smaller clinical effect involves many mutations or multiple variants affects many individuals within a population



The promise is personalized medicine! Drug therapy tailored to a patients unique genetic

makeup: choice of the drug choice of the dosing regimen

Basis for Pharmacogenetics

Pharmacogenetics vs. Pharmacogenomics Concept of pharmacogenetics

based on several factors Most current medications are associated with a

significant risk for drug toxicity and drug inefficacy Variability of drug response Genetic variation

Basis for Pharmacogenetics Drug inefficacy

Response rates vary markedly across therapeutic areas

Estimated response rates

80% - analgesics 25% - cancer chemotherapy30% - Alzheimer’s disease 60% - depression (SSRIs)40% - incontinence 47% - HIV50% - rheumatoid arthritis 60% - schizophrenia50% - migraine (prophylaxis) 52% - migraine (acute)57% - diabetes 60% - asthma60% - cardiac arrhythmias

Overall, 50% of patients do not respond to drugs in the major therapeutic classes

Human Genome ProjectDetermine the sequence of the 3 billion nucleotides

that make up human DNACharacterize variability in the genomeIdentify all the genes in human DNA

The Era of Genomic Medicine: Improve prediction of drug efficacy or toxicity Improve the diagnosis of diseaseEarlier detection of genetic predisposition to disease

Clinical Relevance

Can we predict who will derive an optimal response?

Can we predict who will have a toxicity?

Host (patient) genotype determines optimal drug therapy approach

Disease (pathogen) genotype determines optimal drug therapy approach

DNA is InformationDNA

A, T, G, C

Codon

Gene

Chromosome

Genome

ENGLISH

Abcdefg….xyz

Word

Sentence

Chapter

Book

Composition of the Human Genome

Mutation/Polymorphism 1 bpUnit of genetic code 3 bpCoding sequence (exons) 3,000 bpGene (exons and introns) 50,000 bpChromosome 150,000,000

bpHuman genome 3,000,000,000 bp

The Foundation of Pharmacogenomics: Differences in the Genetic Code

Between PeopleMutation: difference in the DNA code that

occurs in less than 1% of populationOften associated with rare diseases

Cystic fibrosis, sickle cell anemia, Huntington’s disease

Polymorphism: difference in the DNA code that occurs in more than 1% of the populationA single polymorphism is less likely to be the main

cause of a diseasePolymorphisms often have no visible clinical impact

Single Nucleotide Polymorphisms (SNP)

Pronounced “snip”Single base pair difference in the DNA sequence

Over 2 million SNPs in the human genomeOther polymorphisms:

Insertion/deletion polymorphismsGene duplicationsGene deletions

Polymorphisms Two main types:

SNPs – polymorphisms that occur at a single nucleotide

Can be located in either coding regions (DNA that is transcribed; occur less frequently) or non-coding regions

Coding polymorphisms are further classified as: Non-synonymous (missense) – results in translation of a

different amino acid

Synonymous (sense) – results in translation of the same amino acid

Nonsense – results in the insertion of a stop codon

Polymorphisms SNPs

Non-coding polymorphisms when located in promoters, introns, or other

regulatory regions may alter transcription factor binding, mRNA transcript stability or RNA splicing

Polymorphisms Two main types:

Coding & Non-coding SNPs –

Polymorphisms Indels

insertion or deletion of multiple nucleotides commonly result in gene insertions, duplications or

deletions

Basis for Pharmacogenetics Polymorphisms (mainly SNPs) are used to

characterize genetic differences between individuals

However, a pharmacogenetic trait cannot be linked to just

one SNP In this case, haplotypes can be used to associate a

genotype with a phenotype

Basis for Pharmacogenetics Haplotypes

Defined as: Group of SNPs located closely together on a

chromosome are inherited together

Most genes contain between 2 and 53 haplotypes avg. – 14

Basis for Pharmacogenetics

Haplotypes Haplotypes themselves may not have a direct

effect on drug response their proximity to a causative SNP allows them to act

as a marker for a particular drug response

SNP

chromosome

Haplotype

Drug Metabolism Pharmacogenomics

Evidence of an inherited basis for drug response dates back in the literature to the 1950sSuccinylcholine: 1 in 3000 patients developed

prolonged muscle relaxationMonogenicPhenotype to genotype approach

Examples of Drug Metabolism Pharmacogenomics

NEJM 2003; 348: 529-537

Examples of Drug Metabolism Pharmacogenomics

NEJM 2003; 348: 529-537

Warfarin and CYP2C9Widely prescribed anticoagulant drug used to

prevent blood clotsNarrow range between efficacy and toxicityLarge variability in the dose required to achieve

therapeutic anticoagulationDoses vary 10-fold between people

CYP2C9 is the enzyme responsible for the metabolism of warfarin

SNPs exist in CYP2C9 gene that decrease the activity of the CYP2C9 metabolizing enzyme

Warfarin Dosing Pharmacokinetics

racemic mixture of R and S isomers S 5X more potent than R

S-warfarin is transformed by CYP2C9; R-warfarin is mainly transformed by CYP1A2

rapidly absorbed by GI tract with high bioavailability

plasma concentrations peak approximately 90 minutes after administration

half-life 36-42 hours, binds to plasma proteins (mainly albumin)

CYP2C9 Polymorphisms and Warfarin Dose

Warfarin dose is affected by CYP2C9 genotype

Gage BF et al. Thromb Haemost 2004; 91: 87-94

*2 and *3 are SNPs

CYP2C9 Genotype and Bleeding Events

Compared to wild-type, CYP2C9 variants had a higher risk of serious or life-threatening bleeds

Hazard Ratio of 3.94 during the first 3 months of follow-up

Hazard Ratio of 2.39 for the entire follow-up period

Higashi et al. JAMA 2002; 287

WT

Variant

MutationIncreased Clearance

Decreased Clearance

Dosage Adjustment

CYP2C9*1/*1 27% Dose X 1.27

CYP2C9*1/*2 20% Dose X 0.8

CYP2C9*1/*3 40% Dose X 0.6

CYP2C9*2/*2 50% Dose X 0.5

CYP2C9*2/*3 60% Dose X 0.4

CYP2C9*3/*3 85% Dose X .15

Example dosing table for warfarin based on SNP type

Challenges Facing Warfarin Pharmacogenomics

Despite the strong association between CYP2C9 genotype and warfarin dose, CYP2C9 genotype accounts for only a small portion of the total variability in warfarin doses (~10-20%)

Need to determine other genetic and non-genetic factors that contribute to interindividual variability in warfarin doses

CYP2D6 Polymorphisms CYP2D6

The gene is located on chromosome 22 Nearly 100 drugs are substrates for this enzyme

β-adrenergic blockers Antidepressants Neuroleptics

metoprolol amitriptyline haloperidol

propanolol clomipramine resperidone

desipramine thoridazine

Antiarrhythmics fluoxetine

encainide fluvoxamine Others

sparteine imipramine codeine

flecainide nortryptaline dextramethophan

propafenone paroxetine tramadol

CYP2D6 polymorphisms “poor metabolizer” (PM) phenotypes

CYP2D6*3 – A2637 del (frameshift)

CYP2D6*4 – G1934A (splicing defect) most common in Caucasian populations

CYP2D6*5 – Gene deletion (no enzyme)

CYP2D6*10 – C188T most common in Asian populations CYP2D6*4 is almost completely absent in this group

CYP2D6*17 – C1111T most common in African populations

CYP2D6 duplication “extensive metabolizer” (EM) phenotype

repetition of a 42 kb XbaI fragment containing the CYP2D6*2 gene that results in 2-13 copies of the enzyme

the frequency of individuals possessing CYP2D6 duplication suggests a geographical gradient, possibly resulting from dietary pressures

1% - Sweden 4% - Germany 7-10% - Spain 10% - Italy 30% - Ethopians

CYP2D6 PhenotypesNEJM 2003; 348:529

Roden DM et al. Ann Intern Med 2006; 145:749-57

CYP2D6 Polymorphisms and Psychiatric Drug Response

Increased rate of adverse effects in poor metabolizers due to increased plasma concentrations of drug:

Fluoxetine (Prozac) death in child attributed to CYP2D6 poor metabolizer genotype

Side effects of antipsychotic drugs occur more frequently in CYP2D6 poor metabolizers

CYP2D6 poor metabolizers with severe mental illness had more adverse drug reactions, increased cost of care, and longer hospital stays

CYP2D6 and Codeine

Codeine requires activation by CYP2D6 in order to exert its analgesic effect

Due to genetic polymorphisms, 2-10% of the population cannot metabolize codeine and are resistant to the analgesic effects

Interindividual variability exists in the adequacy of pain relief when uniform doses of codeine are given

Strattera® (Atomoxetine)

Treatment of attention deficit hyperactivity disorderCYP2D6 poor metabolizers have 10-fold higher

plasma concentrations to a given dose of STRATTERA compared with extensive metabolizers

Approximately 7% of Caucasians are poor metabolizers

Higher blood levels in poor metabolizers may lead to a higher rate of some adverse effects of STRATTERA

CYP2C19 and Proton Pump Inhibitors

Proton pump inhibitors are used to treat acid reflux and stomach ulcers

Ulcer cure rates using omeprazole and amoxicillin by CYP2C19 phenotype:

Cure RateRapid metabolizers 28.6%Intermediate metabolizers 60%Poor metabolizers 100%

Furuta, T. et. al. Ann Intern Med 1998;129:1027-1030

Thiopurine-S-Methyltransferase (TPMT)

Thiopurine drugs are used to treat cancerAcute lymphoblastic leukemia

TPMT is important for metabolizing thiopurinesazathioprine, mercaptopurine (6-MP)

Polymorphisms in the TPMT gene result in decreased TPMT enzyme activity

Decreased TPMT activity predisposes individuals to severe, life-threatening toxicities from these drugs

Variability in TPMT Activity

Enzyme Activity Levels in 300 Caucasian Patients

0102030405060708090

100

low medium high

TPMT Enzyme Activity

% o

f S

ub

ject

s

Pharmacogenomics 2002;3(1):89-98.

6-Mercaptopurine Prescribing Information

There are individuals with an inherited deficiencyof the enzyme thiopurine methyltransferase(TPMT) who may be unusually sensitive to themyelosuppressive effects of mercaptopurine andprone to developing rapid bone marrowsuppression following the initiation of treatment.

Substantial dosage reductions may be required toavoid the development of life-threatening bonemarrow suppression in these patients.

TPMT genotyping or phenotyping can be used to identify patients with absent or reduced TPMT activity.

Patients with low or absent TPMT activity are at an increased risk of developing severe, life-threatening myelotoxicity from IMURAN if conventional doses are given.

Physicians may consider alternative therapies for patients who have low or absent TPMT activity (homozygous for non-functional alleles). IMURAN should be administered with caution to patients having one non-functional allele (heterozygous) who are at risk for reduced TPMT activity that may lead to toxicity if conventional doses are given. Dosage reduction is recommended in patients with reduced TPMT activity.

TPMT and Thioguanines

Clinical implications:Genetic testing for TPMT is routine practice at

some cancer centers for protocols involving thiopurine drugs

Implications for cancer, transplant, rheumatoid arthritis, lupus, dermatology, and Crohn’s disease treatment

Drug Target PharmacogenomicsDirect protein target of drug

ReceptorEnzyme

Proteins involved in pharmacologic response Signal transduction proteins or downstream proteins

Polymorphisms associated with disease risk “Disease-modifying” polymorphisms“Treatment-modifying” polymorphisms

POLYGENIC

Assessing Phenotype in Drug Target Pharmacogenomics

Depression—Symptom rating scalesIndirect measure of drug responseInter-rater reliability

Hypertension—Blood pressureMinute to minute and diurnal variabilityInfluence of environmental factors (e.g. lack of rest

before measurement)Diabetes—Blood glucose

Diurnal variation in blood glucoseInfluence of environmental factors (e.g.

diet/exercise)

Evans WE. NEJM 2003; 348:538-48

Evans WE. NEJM 2003; 348:538-48

Beta-blockers and Hypertension (HTN)

HTN is the most prevalent chronic disease in the US and a contributor to morbidity and mortality

Beta-blockers are first-line agent in the treatment of HTN

Marked variability in response to beta-blockers 30-60% of patients fail to achieve adequate

blood pressure lowering with beta-blockersCommon beta-blockers used in HTN:

Metoprolol Atenolol

Podlowski, et al. J Mol Med 2000;78:90.

Codon 49 SerGly

Codon 389ArgGly

Beta-1 Receptor Polymorphisms and Response to Metoprolol

Johnson JA et al. Clin Pharmacol Ther 2003; 74:44-52

Beta-2 Adrenergic Receptor Polymorphisms and Response to Albuterol in Asthma

Hyperreactivity of the airways is the hallmark of asthmaAirway smooth muscle contains beta-2 receptors that

produce broncodilationAlbuterol is a beta-2 agonist that is used in the treatment

of asthmaProduces smooth muscle cell relaxation and

bronchodilationForced expiratory volume in 1 second (FEV1)

Phenotypic measure of response

Lima JJ. Clin Pharmacol Ther 1999; 65:519-25

•Single 8 mg albuterol dose

•Albuterol-evoked increases in FEV1 were higher and more rapid in Arg16 homozyotes compared with Gly carriers

• Codon 16 polymorphism is a determinant of bronchodilator response to albuterol

Lima JJ et al. Clin Pharmacol Ther 1999; 65: 519-25

VKOR and Warfarin

Warfarin works by inhibiting Vitamin K Epoxide Reductase (VKOR)

VKOR helps recycle vitamin K which is important in proper functioning of clotting factors

By inhibiting VKOR, warfarin alters the vitamin K cycle and results in the production of inactive clotting factors

Polymorphisms exist in the gene for VKOR (VKORC1)

VKORC1 POLYMORPHISM

At least 10 different single-nucleotide-polymorphisms (SNPs) were identified

Haplotype A (-1639GA, 1173CT): lower maintenance dose Haplotype B (9041GA): higher maintenance dose

VKORC1 A/A: 2.7 ± 0.2 mg/dVKORC1 A/B: 4.9 ± 0.2 mg/dVKORC1 B/B: 6.2 ± 0.3 mg/dMean maintenance dose: 5.1 ± 0.2 mg/d

Rieder MJ, Reiner AP, Gage BF, et el. N Eng J Med 2005;352:2285-93.Rieder MJ, Reiner AP, Gage BF, et el. N Eng J Med 2005;352:2285-93. Schalekamp T, Brasse BP, Roijers JF, et el. Clin Pharmacol Ther. 2006 Jul; 80(1):7-12.Schalekamp T, Brasse BP, Roijers JF, et el. Clin Pharmacol Ther. 2006 Jul; 80(1):7-12.Herman D, Peternel p, Stegnar M, et el. Thromb Haemost 2006; 95:782-7.Herman D, Peternel p, Stegnar M, et el. Thromb Haemost 2006; 95:782-7. Sconce EA, Khan TI, Wynne HA, et el. Blood Oct 2005;106(7):2329-33 Sconce EA, Khan TI, Wynne HA, et el. Blood Oct 2005;106(7):2329-33 Gage BF, MD, MSc. http://www.fda.gov/ohrms/dockets/ac/05/slides/2005-4194S1_04_Gage.pptGage BF, MD, MSc. http://www.fda.gov/ohrms/dockets/ac/05/slides/2005-4194S1_04_Gage.ppt

DOSING ALGORITHM 2005 PROPOSED

Sconce EA, Khan TI, Wynne HA, et el. Blood Oct 2005;106(7):2329-33

DOSING ALGORITHM 2006 PROPOSED

Linder MW Ph.D. DABCC, Manage the “Over-steer” in warfarin dose titration.

Warfarin Dosing In August 2007, the FDA updated the warfarin

prescribing guidelines to include genetic testing: VKROC1 and CYP2C9

Pharmacogenetics has been slow to be implemented clinically

As of 2003, 51 drugs contain pharmacogenetic-related information on their product label

Pharmacogenetic Influence in Therapeutics

Limitations Cost

Existing DNA sequencing technology makes genetic screening inherently expensive

Who is responsible for the cost burden associated with genotyping

patient, government, or insurance company? influence on drug development CYP2D6-metabolized drugs

Potential emotional and financial liability associated with genetic information

Availability and timeliness of genetic testing


However: Recent study suggests a tremendous interest

among patients and clinicians for pharmacogenetics to be more involved in therapeutic decision making

March 2005, FDA officially encouraged pharmacogenomic data to be submitted with drug approval application materials


Documents

中國醫藥大學 藥學系 洪靚娟. Objectives Provide an overview of pharmacogenomics and its clinical relevance Discuss clinically-relevant examples of: Drug metabolism

中國醫藥大學藥學系洪靚娟. Objectives Provide an overview of pharmacogenomics and its clinical relevance Discuss clinically-relevant examples of: Drug metabolism