Pharmacogenetics

Presented by:

Dr.Adithi S Raghavan

Moderated by:

Dr.Anuradha H.V

Pharmacogenetics

1. Definition – Pharmacogenetics

2. Variation in drug response

3. Pharmacogenetic importance

4. Elementary genetics

5. Single gene PK disorders

6. Therapeutic drugs & clinically available PG tests

7. Conclusion

Outline

Pharmacogenetics is the study of the genetic basis

for variation in drug response.

Encompasses Pharmacogenomics which employs

tools for surveying the entire genome to assess

multigenic determinants in drug response.

Introduction

Interindividual variation in response to drugs –Serious problem

Results in Lack of efficacy/Unexpected side effects

Variation:

Pharmacokinetic

Pharmacodynamic

Idiosyncratic

Variation In drug response

• Too much/not enough drug @site of action

• Genes influence PK by altering expression

of Proteins involved in ADME

Pharmacokinetic variation

PD->/<effect of a drug @ a given conc @ site

of action

Interindividual variation in

• Drug targets

• G-proteins

• Other downstream events

Pharmacodynamic variation

Qualitatively abnormal reaction that occurs

only in a few exposed individuals

Results from differences in enzymes or

immune mechanisms

Idiosyncratic

Importance of Pharmacogenetics to variability in Drug response

May be due to :

A.Single mutant gene-Genetic polymorphism

B.Polygenic influences

Polygenic influences & environmental factors

are responsible for normal biological

variations.

Clinical practice-not significant

Pharmacogenetic variations

Classical family studies provide information

on drug response & genetics

Data limited

Twin studies show that drug metabolism is

highly heritable, with genetic factors

accounting for variation

Figure 7–2.

Pharmacogenetic contribution to pharmacokinetic parameters. t1/2 of antipyrine is more concordant in identical in comparison to fraternal twin pairs. Bars show the t1/2 of antipyrine in identical (monozygotic) and fraternal (dizygotic) twin pairs. (Redrawn from data in Vesell and Page, 1968.)

Variation in Drug

responses

Fraternal twins-Wide

Genetic

Environmental

Identical Only Environmental

Comparison of Intra twin vs. Inter pair variability

75-80% variability in PK t1/2 s of drugs eliminated

by metabolism is heritable

Heritability estimated by

Comparing intra-subject vs. inter subject

variability in drug response

or disposition in unrelated individuals

Assuming high intra subject reproducibility

translates into high heritably.

Elementary genetics

Fundamental units

of heredity

Consist of ordered

sequence of

nucleotides(Adenin

e,

Guanine ,Thymidin

e & Cytosine-

A,G,T,C.

Genes

Written in italics

• CYP2D6-protein

• CYP2D6-Gene

Most DNA-Chromosome

Small amount-

Mitochondria.(Maternal

Ovum)

DNA TranscribedComplementary

mRNA

Translated in Rough

Endoplasmic reticulum

Sequence of Amino

Acids

Post translational modification

Protein product

Rate of transcription -

Promoter region

RNA polymerase binds to

initiate transcription

Redundant Polymorphism

Confers advantage

Persist in several

generations

Silent mutation

Eliminated by natural selection

Heritable changes in

base sequence of DNA

Mutations

Different alternative sequences at a locus within

DNA strand(alleles) that persist in a population

through several generations.

Arise due to mutation.

Stable-non functional.- Die out-disadvantageous

Increase in frequency over generations-selective

advantage

Polymorphisms

Balanced polymorphism

Situation where several

functionally distinct

forms of a gene are

common in a population

Ambiguity-preserves the

geneSusceptibility to Hemolysis

Linked gene for G6PD

Deficiency

Partial resistance to

Malaria

SNPs are DNA sequence variations that occur

when a single nucleotide in the genome

sequence is altered.

May entail substitution of one nucleotide to

another(C for T)

Result in ‘frame shift’ in translation

Single Nucleotide Polymorphism

Result can be loss of protein synthesis, abnormal protein

synthesis or an abnormal rate of protein synthesis.

Individuals differ from each other approx. every 300-

1000 nucleotides with an estimated total of 30 million

SNP.

Can occur in coding & non coding regions

Important determinant of disease-e.g. Inherited

Thrombophilia

SNPs

SNP in Factor V Leiden

Prolonged immobility

Increased risk of

Venous Thrombosis

Advantage?

In Case of Hemorrhage

than thrombosis

Inherited Thrombophilia

Combination of SNPs

• In or near a gene

Known as Haplotype

• Inherited from each parent

Predisposition to a Disease

Figure 7–10.Types of genetic variants that have been significantly associated with complex human traits and disease in 208 genome-wide association studies. See www.genome.gov/gwastudies/.

Single gene pharmacokinetic disorders

Inherted;Mendelian fashion

‘Single gene disorder’

Disrupts Gene function

Mutation

Walter Kalow ;Suxamethonium sensitivity ;rate

of metabolism

Mendelian Autosomal Recessive trait

Short acting NM blocker

Plasma cholinesterase

Atypical Plasma cholinesterase

1:3000 fail to

inactivate SXM

rapidly

Recessive gene

Prolonged NM Block

Abnormal plasma

cholinesterase

DibucaineHeterozygotes-

Intermediate

Inhibits Abnormal enzyme less

Blood test

Homozygotes-when exposed

Autosomal dominant

inherited.

Idiosyncratic ADR due to

SXM on Ryanodine receptor

Also due to halogenated

inhalational agents

(Halothane)

Incidence 1:20000

Rapid rise of body

temperature,

muscle rigidity,

tachycardia & cyanosis.

Malignant hyperthermia

Mechanism: Sudden rise in

release Ca2+ from sarcoplasmic

stores leading to muscle

contraction & hyper metabolic

rate.

Potentially fatal .

Important test family members

of affected.

Impractical to screen routinely

Treatment:

Dantrolene 1 mg/kg i.v repeated up to 10mg/kg.

(prevents release of Ca2+ from sarcoplasmic

reticulum)

Symptomatic Rx of Hyperthermia

Rx of Cardiac arrhythmias

Commonest & most severe form of hepatic

porphyria

Autosomal dominant

Mutation in gene coding Porphobilinogen

deaminase(PBGD)

Acute intermittent porphyria

PBGD key enzyme

Haem Biosynthesis

Red cell precursors

Hepatocytes & other cells

Mutation

Reduces activity

PBGD Build up

Haem precursors

(incl.Porphyrin)

Strong interplay with environment through

exposure to drugs ,hormones& other chemicals

Use of sedative, anticonvulsant or other drugs in

patients undiagnosed-can be Lethal

Most drugs(not just CYP inducers) can precipitate

acute attacks in susceptible individuals

ALA synthase

in liver

Induced by drugs like

Barbiturates

ALA(delta amino laevulanic acid)

Increased ALA production

Porphyrins

Acute Attack

Frank disease 5 times more common in Women

Hormonal Fluctuations

Precipitate Acute Attacks

Drug acetylation status

Acetyltransferase

• Single recessive gene associated with low enzyme activity

Acetylation

Elimination of Isoniazid

American population; equal no's ‘fast’

&’slow’ acetylators

Other ethnic groups ;different proportions

Slow>Egyptians,british swedish

Rapid> Canadians,Asians,Latin americans

Peripheral Neuropathy

Isoniazid

Slow acetylators

Hepatotoxicity

Acetylhydrazine

Fast acetylators

Isoniazid;2 different forms toxicity

AcetyltransferaseMetabolism

of

Hydralazine

Procainamide

Dapsone

Other sulfonamides

Idiopathic ADR

Caused By many agents

Drug induced lupusAutoimmune Skin, joints &

kidneys

Acetylator status Influences

Click icon to add picture

• Mitochondrial gene

• Most common predisposing

mutation

m.1555A>G,mitochondrial

DNA mutation

• 30-60% ototoxicity in

china(Aminoglycosides-cheap)

Aminoglycoside ototoxicity

Bind to Bacterial ribosomes

Mutation of human mitochondrial ribosomes

is similar

For a single dose in susceptible individuals.

Screening for this variant appropriate in children

Increased affinity to ribosomes in hair cells in ear for several months

Aminoglycosides

• Rate of Metabolism differs with

race

• Oriental races- accumulation

of acetaldehyde.

• Due to slower rate of oxidation

of acetaldehyde as a result of

genetic polymorphismEspecially in Japanese

Defect in Ethanol metabolism

Around 80% of Asians have a variant gene ADH1B

Almost all Chinese and Koreans- ADH1C

coding alcohol dehydrogenase -toxic acetaldehyde at a much higher efficiency

50% of Asians, the increased acetaldehyde accumulation, the mitochondrial ALDH2 allele,

less functional acetaldehyde dehydrogenase enzyme,

Therapeutic Drugs and Clinically available Pharmacogenetic tests

Understanding human genome

Simpler methods identify genetic

information

Genetic information specific to individual

Preselect effective drug

Personalized Medicine

No toxicity

No trial & error

US FDA has approved PG labeling info to package inserts of over 50 drugs

Use patchy

Anticipated to be one of the first applications of

human genome sequencing.

Development slowed by various scientific ,

commercial, political and educational barriers.

Cost effectiveness?

Evidence in support of a test is less convincing than

the ideal of an RCT of PG informed prescribing

strategy versus current best practice

Clinical Pharmacogenetic tests

1.Variants of different HLA strongly linked to susceptibility to severe idiosyncratic reactions

2.Genes controlling aspects of drug metabolism

3.Genes encoding drug targets

Tests increasingly used

Methodology

Mutations in Germline

All cells of the Next generation

Venous blood samples(Chromosomal &

Mitochondrial DNA in WBCs

Tumours

Pathogenesis

Somatic cell mutationsPresence or absence guides drug selection

• Genomic tests done on DNA

from samples of tumor

obtained surgically.

• Tests involve amplification of

relevant sequences and

molecular biological methods

often utilizing chip technology

to identify various

polymorphisms

HLA Gene tests

Abacavir-Reverse transcriptase inhibitor

Highly effective - HIV Infection

Severe Rashes

Susceptibility linked to HLA variant

HLAB*5701

ABACAVIR & HLA-B*5701

Severe life threatening rashes

Stevens Johnson Syndrome

Toxic epidermal Necrolysis

Almost only in Asians

FDA recommends Chinese patients to be screened for this

allele

Similar problem with Phenytoin for same allele

Anticonvulsants & HLAB*1502 Carbamazepine

Effective antipsychotic drug

Agranulocytosis 1% of patients

Studies-small

Specificity and sensitivity yet to be

established

Clozapine and HLA-DQB1*0201

Drug metabolism and related gene tests

Thioguanine,Mercaptopurine & its

prodrug Azathioprine

Treat Leukemia's(ALL),Inflammatory

Bowel disease & Immunosuppression

Cause Bone marrow & Liver toxicity

Detoxified by Thiopurine S

methyltransferase(TPMT) present in

blood cells & by Xanthine oxidase

Low TPMT activity

High TPMT

Reduced efficacy

Lower conc TGN

Bone marrow Toxicity

High Conc of active TGN in

blood

Thiopurines and TPMT

Phenotyping (by a blood test for TPMT activity)

Genotyping TPMT Alleles

TPMT*3A,TPMT*3C,TPMT*2 is recommended.

Careful monitoring of WBC count & drug

interaction with allopurinol(due its effect on

Xanthine Oxidase)

Before treatment

Extensively used to treat solid Tumours.

Unpredictable mucocutaneous toxicity.

Detoxified by dihydropyrimidine

dehydrogenase(DPYD)-clinically identifiable

multiple genetic variants

FDA recommends no to be given to those

with DPYD deficiency

5-FLUOROURACIL(5-FU) &DPYD

TAMOXIFEN

TAMOXIFEN & CYP2D6CYP2D6

Estrogen antagonist

ENDOXIFENPolymorphic variation

Suggested link between CYP2D6 genotype&

efficacy.

Genotyping tests available.

Tetrabenzaine used to Huntington's disease may

also be influenced by cyp2d6

Topoisomerase I inhibitor.

Marked activity against colorectal & lung

cancers(minority)

Toxicity(Diarrhoea & BM suppression very severe

Active metabolite SN-38

UDP glucuronyltransferase

Reduced activity Hyberbilirubinemia Gilberts

syndrome

UGT1A1 Testing clinically available

Uncertain

IRINOTECAN & UGTA1*28

Drug target related gene tests

• Herceptin is mAB that antagonizes

epidermal growth factor(EGF) by binding to

one of its receptors(human epidermal

growth factor receptor 2-HER2)

• Somatic mutation HER2 in tumour

tissue

• Used in BrCA where HER2 is

overexpressed.

TRASTUZUMAB & HER2

o DASATINIB –dual BCR/ABL & Src tyrosine

kinase inhibitor

o Used in hematological

malignancies(Philadelphia chromosome)

o CML ALL

o Mutation (T3151) in BCR/ABL confers

resistance to inhibitory effect of dasatinib.

DASATINIB,IMATINIB & BCR-ABL1

o IMATINIB-TYROSINE kinase inhibitor

o CML & other myelodysplastic disorders.

Combined(Metabolism and Target ) Gene tests

WARFARIN

Dosing individualized by measuring

INR(International normalized ratio)

Thrombotic effects(lack of efficacy)

Adverse effects(bleeding) common

PG testing proposed based on polymorphism in its

key target, vitamin K epoxide reductase(VKOR)

&CYP2C9 GENOTYPE involved in its metabolism

WARFARIN & CYP2C9 +VKORC1 GENOTYPING

Pharmacogenetics

proves that concept of susceptibility to ADR can be

genetically determined

Offers possibility of a more precise ‘Personalised ‘

Medicine for several drugs & disorders.

Field of intense research, rapid progress.

Challenge remains about its feasibility in Clinical

setup

Conclusion