Genetic polymorphisms can affect how individuals metabolize and respond to drugs. Variations in genes encoding drug metabolizing enzymes, like CYP450 isoforms and dihydropyrimidine dehydrogenase, have been shown to result in decreased, increased, or absent enzyme expression/activity. This can lead to inter-individual differences in drug effects, like a higher risk of toxicity from drugs metabolized by the affected enzymes. Single nucleotide polymorphisms in these genes have been linked to variability in drug dosing requirements, interactions, and treatment outcomes.

1. POLYMORPHISM AFFECTING DRUG
METABOLISM
PRESENTED BY,
RAMESHA.S
Ist M pharm
Department of pharmacology
Gcp,Bangalore

2. Definition: Genetic polymorphism is a term used in genetics to describe multiple
forms of a single gene that exist in an individual or among a group of individuals.
Causes of genetic polymorphism
Deletion and duplication of millions of base pairs of DNA.
Changes in one or a few bases in the DNA located between genes or within
exons.
Sequence changes may also be located in the coding sequence of genes
themselves and result in different protein variants that may lead in turn to different
phenotypes.

3. Genetic Polymorphism & Drug Metabolism
• inter-individual variation of drug effects
• Genetic polymorphisms of drug-metabolizing enzymes give rise to distinct
subgroups in the population that differ in their ability to perform certain drug
biotransformation reactions.
• Polymorphisms are generated by mutations in the genes for these enzymes,
which cause decreased, increased or absent enzyme expression or activity by
multiple molecular mechanisms.

4. Drug Metabolism
• The metabolism of drugs and other xenobiotics into more hydrophilic
metabolites is essential for their elimination from the body, as well as for
termination of their biological and pharmacological activity.
• Drug metabolism or biotransformation reactions are classified as,
 Phase I - functionalization reactions.
 Phase II - biosynthetic (conjugation reactions).

5. Drug Metabolism (cont.)
• The enzyme systems involved in the biotransformation of drugs are localized
primarily in the liver.
• Other organs with significant metabolic capacity include the GI tract, kidneys, and
lungs.
• These biotransformation reactions are carried out by CYPs (cytochrome CYP450
isoforms) and by a variety of transferases.

6. Drug Metabolism (cont.)
• Pathways of drug metabolism are classified as either:
• Phase I reactions: oxidation, reduction, hydrolysis
• Phase II reactions: acetylation, glucuronidation, sulfation, methylation
• Both types of reactions convert relatively lipid soluble drugs into relatively
inactive and more water soluble metabolites, allowing for more efficient
systemic elimination.

7. Gene Polymorphisms
• Genetic differences in drug metabolism are the result of genetically based
variation in alleles for genes that code for enzymes responsible for the metabolism
of drugs.
• In polymorphisms, the genes contain abnormal pairs or multiples or abnormal
alleles leading to altered enzyme function.
• Differences in enzyme activity occur at different rates according to racial group.

8. Single Nucleotide Polymorphisms (SNPs)
Single changes in one allele of a gene responsible for a variety of metabolic
processes including enzymatic metabolism.
• The combination of alleles encoding the gene, determines the activity and
effectiveness of the enzyme.
• The overall function of the enzyme is the phenotype of enzyme function.
Phenotype : the observable physical or biochemical characteristics determined
by both genetic makeup and environmental influences

9. 9
Human cytochrome P450 family
Of the super-family of all cytochromes, the following families were confirmed in humans:
CYP 1-5, 7, 8, 11, 17, 19, 21, 24, 26, 27, 39, 46, 51
CYP 1, 2A, 2B, 2C, 2D, 2E, 3 –metabolism of xenobiotica.
CYP 2G1, 7, 8B1, 11, 17, 19, 21, 27A1, 46, 51 - steroid metabolism.
CYP 2J2, 4, 5, 8A1 -fatty acid metabolism.
CYP 24 (vitamine D), 26 (retinoic acid), 27B1 (vitamine D), ...

11. Inhibitors & Inducers
Polymorphisms affect drug interactions by altering the effect of inhibitors and
inducers on the enzyme.
results in an exaggerated effect or minimal effect on the substrate
Inhibitor: An enzyme inhibitor is a molecule, which binds to enzymes and
decreases their activity.
Inducer : An enzyme inducer is a type of drug that increases the metabolic activity
of an enzyme either by binding to the enzyme and activating it, or by increasing
the expression of the gene coding for the enzyme.

12. Extensive Metabolizers - Inhibitors
• Extensive metabolizer - level of substrate drug is normally low due to rapid
metabolism by the enzyme.
• An inhibitor to the enzyme will inhibit the extensive metabolism and cause
significant elevations in the substrate drug.
• Effect of inhibitors is much greater in an extensive metabolizer  inc. level of
substrate levels

13. Poor Metabolizers - Inhibitors
• In a poor metabolizer, the level of substrate drug remains high because the
metabolism of the substrate is much less than normal.
• When an inhibitor is added, the additional inhibition of metabolism is not much
greater than is already occurring in the PM.
• The effect of inhibitor is less in a PM than in normal metabolizers.
• The drug interaction might not occur.

14. Extensive Metabolizers - Inducers
• Level of substrate drug is lower than in a normal metabolizer due to rapid
metabolism.
• The addition of an inducer does not cause a greater difference in the level of
substrate because the metabolism is already increased greatly.
• The drug interaction might not occur.

15. Poor Metabolizers - Inducers
• Level of substrate drug is higher than expected in normal metabolizer because of
the lower metabolism of substrate.
• The addition of inducer will cause a signification increase in the metabolism of the
substrate  much lower level of substrate than expected in a normal metabolizer.
• Drug interaction may occur to a greater extent.
• Drug interaction may result in substrate levels similar to those of normal
metabolizers.

16. NOTE:
• The effect of inhibitor is great in EMs than in PMs.
• The effect of inducer is greater in PMs than in EMs.

17. Complex Drug Interactions
• Can be seen when a substrate is metabolized through more than one enzyme
systems where one or more enzymes are affected by polymorphism.
Substrate is metabolized through a
polymorphic enzyme
Substrate becomes active metabolite
This active metabolite acts as an
inhibitor or inducer in second system

18. Phase I Enzymes
Enzyme Substrate Clinical Consequence
CYP1A1 Benzopyrine, phenacetin Inc. or dec. cancer risk
CYP1A2 Acetaminophen, amonafide, caffeine, paraxanthine, ethoxyresorufin,
propranolol, fluvoxamine
Decreased theophylline metabolism
CYP1B1 Estrogen metabolites Possible inc. cancer risk
CYP2A6 Coumarin, nicotine, halothane Dec. nicotine metabolism and cigarette
addiction
CYP2B6 Cyclophosphamide, aflatozin, mephenytoin Significance unknown
CYP2C8 Retinoic acid, paclitaxel Significance uknown
CYP2C9 Tolbutamide, warfarin, phenytoin, NSAIDS Anticoagulant effect on warfarin
CYP2C19 Mephenytoin, omeprazole, hexobarbital, mephobartibal, propranolol,
proquanil, phenytoin
Peptic ulcer response to omeprazole
CYP2D6 Betablockers, antidepressants, antipsychotics, codeine, debrisoquin,
dextromethorphan, encainide, flecanide, fluoxetine, guanoxan,
methxyamphetamine, phenacetin, propafenone, sparteine
Tardive dyskinesia from
antipsychotics; narcotic side effects,
efficacy and dependency, imipramine
dose requirement; beta blocker effects
Genetic Polymorphisms in Genes that Can Influence Drug Metabolism – CYP450
Isoforms

19. CYP2E1 Acetaminophen, ethanol Possible effects on alc consumption
Possible inc cancer risk
CYP3A4/3A7/3A7 Macrolides, cyclosporine, tacrolimus, calcium
channel blockers, midazolam, tefrenadie, lidocaine,
dapsone, quinidine, triazolam, etoposide, teniposide,
loastatian, alfentanil, tamoxifen, steroids,
benzo(a)pyrene
Tacrolimus dose requirement in pediatric cancer
patients
Aldehyde
dehydrogenase
Cyclophosphamide, vinyl chloride SCE frequency in lymphocytes
Alcohol
dehydrogenase
Ethanol Inc. alc consumption and dependence
Dihydrodyrimidine
dehydrogenase
(DPD)
5-fluorouracil Inc. 5-flurorouracil toxicity
NQO1 Ubiquinone, menadione, mitomycin C Menadione-associated orlithiasis, dec tumor
sensitivity to mitomycin-C; possible inc. cancer
risk

20. P450 Enzymes in Drug Metabolism
• The polymorphic P450 (CYP) enzyme superfamily is the most important system
involved in the biotransformation of many endogenous and exogenous
substances including drugs, toxins, and carcinogens.
• Genotyping for CYP polymorphisms provides important genetic information that
help to understand the effects of xenobiotics on human body.
• For drug metabolism, the most important polymorphisms are those of the genes
coding for CYP2C9, CYP2C19, CYP2D6, and CYP3A4/5, which can result in
therapeutic failure or severe adverse reactions.

21. CYTOCHROME P4502C SUBFAMILY
• Accounts for approximately 18% of the CYP content in the liver
• Catalyzes roughly 20% of the CYP-mediated metabolism of drugs
CYP2C19
• Study using mephenytoin as probe drug determined that individuals can be
segregated into EMs and PMs.
• PM trait is autosomal recessive – present in 3-5% of Caucasians & 12-23% of
Asian populations

22. CYTOCHROME P450246
• Most extensively studied polymorphic drug metabolizing enzyme
• Impaired ability to hydroxylate, and therefore, inactivate debrisoquin
• 5-10% of white subjects have relative deficiency in ability to oxidize debrisoquin.
Also have impaired ability to metabolize the antiarrhythmic and oxytocic drug
sparteine
• PM  lower urinary concentration, higher plasma concentrations
• Subjects inherited two copies of a gene or genes that encoded an enzyme with
either decreased CYP2D6 activity or no activity at all prominent in East African
population – frequency as high as 29%.

23. CYP2C19 (cont.)
• Also catalyzes the metabolism of several proton pump inhibitors (i.e. omeprazole),
diazepam, thalidomide, and some barbiturates.
• Responsible for inactivation or propranolol and metabolic activation of
antimalarial drug proquanil.

24. CYP2C19 & Diazepam
• Diazepam is demethylated by CYP2C19
• Plasma diazepam half-life is longer in individuals who are homozygous for the
defective CYP2C19*2 allele compared to those who are homozygous for the wild
type allele.
• Half-life of the desmethyl diazepam metabolite is also longer in CYP2C19 poor
metabolizers.
• High frequency in Asian population.
• Diazepam induced toxicity may occur as a result of slower metabolism – careful
dosing in Asian population.

25. CYP2C9
• Major CYP2C subfamily member in the liver
• Primarily responsible for the oxidative metabolism of important compounds –
warfarin, phenytoin, tolbutamide, glipizide, losartan, etc.
• 6 different polymorphisms – CYP2C9*1, *2, *3, *4, *5, *6
• CYP2C9*1 – wild type allele, CYP2C9*2-*6 – variants
• Variants *2 and *3 alleles are common in Caucasians (≈35%)
• CYP2C9*2 and *3 alleles associated with significant reduction in the metabolism
and clearance of selected CYP2C9 substrates

26. CYP2C9 & Warfarin
• Polymorphisms linked to both toxicity and dosage requirements for optimal
anticoagulation with warfarin.
• *2 and *3 variants – higher risk of acute bleeding complications than patients with
*1 wild type genotype.
• Require 15-30% lower maintenance dose of warfarin to achieve target INR
• Patients with variant CYP2C9 genotype take a median of 95 days longer to
achieve stable dosing compared to wild-type group

27. CYTOCHROME P4503A SUBFAMILY
• CYP3A subfamily plays a critical role in the metabolism of more drugs than any
other phase I enzyme.
• Expressed in liver and small intestine
• Contribute to oral absorption, first-pass, and systemic metabolism
• Expression is highly inducible – enzyme activity influence by factors such as
variable homeostatic control mechanisms, up- or down- regulation by environment
factors, and polymorphisms.

28. CYP3A4
• More than 30 SNPs have been identified for CYP3A4 gene
• Unlike other P450s, there is no evidence for deleted or null allele for CYP3A4.
• The most common variant in CYP3A4, CYP3A4*1B is an A392G transition in the
promoter region referred to as the nifedipine response element.
• One study shows that this variant may be associated with a slower clearance
of cyclosporine.
• This is a rather controversial finding.

29. • PXR signaling serves as a central regulator of inducible CYP3A4 expression as
well as several other genes involved in drug detoxification.
• Polymorphisms in PXR suggest observed variability in CYP3A4 enzymatic
activity may be due to, in part, inherited differences in the upstream signaling
proteins that control induction of gene expression.

30. CYP3A5
• Polymorphically expressed in adults in about 10-20% in Caucasians, 33% in
Japanese, and 55% in African Americans.
• The variable CYP3A5*3 is a result of improper mRNA splicing and reduced
translation of functional protein.
• CYP3A5 is the primary extra-hepatic CYP3A isoform, its polymorphic expression
has been implicated in disease risk and the metabolism of endogenous steroids or
drug in tissues other than liver.
• CYP3A5 has been linked to tacrolimus dose requirements to maintain adequate
immunosuppression in solid organ transplant patients.

31. CYP3A7
• Expressed in fetal liver during development
• Hepatic expression is generally down-regulated after birth, but the CYP3A7
protein has been detected in some adults
• Increased CYP3A7 expression has been associated with the replacement of 60
nucleotide fragment of the CYP3A7 promoter with the corresponding region form
of the CYP3A4 promoter (CYP3A7*1C allele.)
• This promoter swap results in increased gene expression of the pregnane X
receptor response element.

32. Dihydropyrimide Dehydrogenase
• Metabolism of antineoplastic agent fluorouracil.
• In the 1980s, fatal CNS toxicity developed in several patients after treatment with
standard doses fluorouracil.Patients had inherited deficiency of dihyropyrimidine
dehydrogenase.
• DPD metabolizes fluorouracil and endogenous pyrimidines.
• Severe fluorouracil toxicity occurs when DPD activity < 100 pmol/min/mg
protein.3% of population carries heterozygous mutations that inactivate DPD and
1% are homozygous for the inactivating mutations.Heterozygous individuals do
not exhibit no phenotype until challenged with fluorouracil.

33. Phase 2 Enzymes
Enzyme Substrate Clinical Consequence
N-acetyltransferase (NAT1) Aminosalicylic acids, aminobenzoic
acid, sulfamethoxazole
Possible increased cancer risk
Hypersensitivity to sulfonamides;
amonafide toxicity; hydralazine-
induced lupus, isoniazid neurotoxicity
and hepatitis
N-acetyltransferase (NAT2) Isoniazid, hydralazine, sulfonamides,
amonifidide, procainamine, dapsone,
caffeine
Glutathione transferase (GSTM1, M3,
T1)
Busulfan, aminochrome, dopachrome,
adrenochrome, noradrenochrome
Possible inc cancer risk; cisplatin
induced ototoxicity
Glutathione transferase (GSTP1) 13-cis retinoic acid, busulfan,
ethacrynic acid, epirubicin
Possible inc cancer risk
Sulfotransferases Steroids, acetaminophen, tamoxifen,
estrogens, dopamine
Possible inc or dec cancer risk; clinical
outcomes in women receiving
tamoxifen for breast cancer
Catechol-O-methyltransferases Estrogens, levodopa, ascorbic acid Decreased response to amphetamine,
substance abuse, levodopa response

34. Thiopurine methyltransferase Mercatopurine, thioguanine,
azathioprine
Thiopurine toxicity and efficacy, risk
of second cancers
UDP-glucuronosyl-transferase
(UGT1A1)
Irinotecan, troglitazone, bilirubin Irinotecan glucuronidation and
toxicity, hyperbilirubinemia (Crigler-
Najjar syndrome, Gilbert’s syndrome)
UDP-glucuronosyl-transferase
(UGT2B)
Opioids, morphine, naproxen,
ibuprofen, epirubicin
Significance unknown

35. N-ACETYLTRANSFERASE
• N-acetylation of isoniazid to acetylisoniazid
• Individuals are slow or rapid acetylators
• Ethnic variation is seen
• Slow acetylation: Japanese (10%), Chinese (20%), Caucasians (60%)
• NAT2 protein is the specific protein isoform that acetylates isoniazid.
• 27 unique NAT2 alleles identified
• NAT2*4 is the wild type allele
• NAT2 alleles containing the G191A, T341C, A434C, G590A, and/or G857A
missense associated substitutions are associated with slow acetylator phenotype.

36. REFERENCE:
• Shargel, Leon. Chapter 12 – Pharmacogenetics. Applied Biopharmaceutics and
Pharmacokinetics, 5th edition. E-book.
• Shargel, Leon. Comprehensive Pharmacy Review, 7th Edition. Philadelphia:
Lipincott- William & Wilkins, 2010. Print. Pages 430-433.
• David B. Troy, Paul Beringer. Remington: The Science and Practice of
Pharmacy, 21st Edition. Pages 1230 – 1239.
• Brunton, Laurence. Chabner, Bruce. Knollman, Bjorn. Goodman & Gilman’s
The pharmacological basis of therapeutics, 12th edition. Pages 124-130.

37. • http://dmd.aspetjournals.org/content/29/4/570.full
•http://www.biologyonline.org/dictionary/Genetic_polymorphism
•http://www.ncbi.nlm.nih.gov/pmc/articles/PMC1934960/
•http://www.medscape.com/viewarticle/444804_5
•http://en.wikipedia.org/wiki/Drug_metabolism