In this presentation I have tried to explain in brief about Genetic polymorphism in drug metabolism.

1. By: Dr. Ankit Gaur
M.Sc, Pharm. D, R.Ph

2.  Genetic polymorphism is the variations in
DNA sequences
 This explain some of the variability in drug-
metabolizing enzyme activities which
contribute to
 alterations in drug clearance and
 impact patients' response to drug therapy

3.  Several extensively studied SNPs (single
nucleotide polymorphism) include the genes
encoding for :
1. glucose-6-phosphate dehydrogenase
2. N-acetyltransferase and
3. The superfamily of cytochrome P-450 (CYP)
isoenzymes.

4.  Because CYP isoenzymes metabolize a large
number of structurally diverse drugs and chemicals,
most of the variant genotypes of the CYP2D6,
CYP2C9, CYP2C19, and CYP3A families have been
identified and studied.
 Individuals with aberrant genes for these enzymes
may experience diminished efficacy or increased
toxicity in response to certain drugs because of the
different levels of activities associated with variant
genotypes.

5.  The frequency of variant alleles for drug
metabolizing enzymes often differs among ethnic
groups.
 The application of this knowledge will ultimately
help in
▪ individualize drug dosing
▪ drug therapy selection
▪ predict toxicity or therapeutic failure
▪ and improve clinical outcomes.

6.  Pharmacogenetics has elucidated the genetic basis
for interindividual variability in drug response and
will continue to play a key role in defining strategies
to optimize drug therapy.
 The pharmacogenetics of drug-metabolizing
enzymes is a prominent focus of this field, because
genetic makeup is responsible for a significant
portion of drug-induced toxicity; many drugs are
metabolized by enzymes that are encoded by
polymorphically expressed genes

7.  Genotype analysis can be used to identify DNA
changes in specific metabolic pathways that
produce aberrant phenotypes.
 Hence, patients can be classified according to their
ability to metabolize certain drugs as
 extensive
 intermediate
 poor metabolizers
 This classification can differentiate interpatient and
intrapatient pharmacokinetic and
pharmacodynamic variability.

8.  DNA consists of four bases: adenine, guanine,
thymine, and cytosine.
 Any combination of three nucleic acids can
form a codon, which is transcribed into
mRNA and translated into a particular amino
acid (e.g., ACG encodes for threonine).

9.  The stop codon,TAG, terminates protein
synthesis. An incorrectly placed stop codon in
a gene caused by mutation, prematurely
truncates an amino acid chain and may form
a nonfunctional protein.
 Many of the variations in the human genome
are single base changes, termed SNPs

12. A mutation of one nucleotide of a codon may
result in either :
 change in the coded amino acid
(nonsynonymous SNP) or
 no change in the coded amino acid (silent
polymorphism or synonymous SNP).

14.  One or more genes that code for a particular
protein, such as an enzyme or a receptor, may be
expressed in different amounts in different tissues.
 All individual alleles (or genes) are referenced by
their gene name (e.g., CYP2D6), followed by an
asterisk and an Arabic number (e.g., CYP2D6*1
designates the wild-type allele and CYP2D6*3 is a
mutant allele)

15.  Glucose-6-phosphate dehydrogenase (G6PD or G6PDH) is a
cytosolic enzyme in the pentose phosphate pathway
 G6PD also protects red blood cells from potentially harmful
byproducts that can accumulate when a person takes certain
medications or when the body is fighting an infection.
 Cell growth and proliferation are affected by G6PD
 This deficiency can cause hemolytic anemia, usually after
exposure to certain medications, foods, or even infections.

16.  G6PD deficiency is passed along in genes from one
or both parents to a child.The gene responsible for
this deficiency is on the X chromosome.
 G6PD deficiency is most common in African-
American males.
 Many African-American females are carriers of
G6PD deficiency, meaning they can pass the gene
for the deficiency to their children but do not have
symptoms; only a few are actually affected by G6PD
deficiency.

17. G6PD Deficiency SymptomTriggers
 illness, such as bacterial and viral infections
 certain painkillers and fever-reducing drugs
 certain antibiotics (especially those that have "sulf"
in their names)
 certain antimalarial drugs (especially those that
have "quine" in their names)

18.  Only 30 different functional mutations in the gene
have been reported, virtually all of which are found in
the region of the gene that codes for the protein.
 All but one are point mutations, with more than 50%
being nucleotide conversions from cytosine to
guanine.
 The consequence of these genetic polymorphisms is
low G6PD activity, resulting in reduced glutathione
concentrations in erythrocytes and subsequently
clinical manifestation of hemolytic anemia following
the ingestion of certain drugs.

19.  The prevalence of G6PD deficiency differs
among ethnic groups. For instance, males of
African and Mediterranean descent more
frequently express the trait.Two types of
mutations are commonly found in Africans,
G6PD A and G6PDA(-).

20.  There are three different G6PDA(-) variants in
one allele.The A376G mutation occurs in all
people, but the enzyme deficiency is caused
by a second amino acid substitution, usually a
G202A mutation, resulting in a valine-to-
methionine substitution at codon 68
(Val68Met)

21.  primaquine,sulfones, sulfonamides,
nitrofurans, vitamin K analogues, cefotetan,
and chloramphenicol, are known to cause
hemolytic anemia in G6PD-deficient patients.

22.  The acetylation polymorphism illustrates another genetic
polymorphism of a drug-metabolizing enzyme.
 N-acetyltransferase (gene, NAT), a phase-II conjugating liver
enzyme, catalyzes the N-acetylation(usually deactivation) and
O-acetylation (usually activation) of arylamine carcinogens
and heterocyclic amines
 NAT is controlled by 2 genes NAT1 and NAT2 of which
NAT2A & B are responsible for clinically significant
metabolic polymorphism.

23.  If any drug known to be a ‘marker’ for certain enzyme is
given and plasma or urine drug conc measured after a std time
interval, then it is possible to separate individuals into 2
groups - fast acetylators and slow acetylators
 The slow acetylator phenotype often experiences toxicity from
drugs such as isoniazid, sulfonamides, procainamide, and
hydralazine, whereas the fast acetylator phenotype may not
respond to isoniazid and hydralazine in the management of
tuberculosis and hypertension, respectively.

24.  During the development of isoniazid, isoniazid plasma
concentrations were observed in a distinct bimodal population
after a standard dose.
 Patients with the highest plasma isoniazid levels were
generally slow acetylators and they suffered from peripheral
nerve damage, while fast acetylators were not affected.
 Slow acetylators are also at risk for sulfonamide-induced
toxicity and can suffer from idiopathic lupus erythematosus
while taking procainamide

25.  The slow acetylator phenotype is an autosomal
recessive trait.
 Studies have shown large variations of the slow
acetylator phenotype among ethnic groups: 40- 70%
of Caucasians and African-Americans, 10-20% of
Japanese and Canadian Eskimo, more than 80% of
Egyptians, and certain Jewish populations are slow
acetylators

26.  Allelic variation at the NAT2 gene locus accounts for
the polymorphism seen with acetylation of substrate
drugs. There are 27 NAT2 alleles that have been
reported.

27.  NAT2*5B and *6A account for 72-75% of all the variant NAT2
alleles, which includes at least 94% of all variant alleles in
Caucasians, Japanese, and Hispanics and 83% of the NAT2 alleles
in African-Americans.
 NAT2*5B is the most common allele in Caucasians (40-46%), but
occurs at a very low frequency in Japanese (0.5%).
 Genotype NAT2 mutations could be an addition to the traditional
therapeutic drug monitoring for isoniazid in the near future.

28.  The CYP isoenzyme superfamily comprises over 50 heme-
containing proteins that catalyze the oxidative metabolism of
many structurally diverse drugs and chemicals.
 But 6 of them metabolize 90 of drugs – CYP1A2, CYP2C9,
CYP2C19, CYP2D6, CYP3A4, CYP3A5
 Among which 2 most significant enzymes are CYP3A4,
CYP2D6

29.  CYP2D6 isoenzyme metabolizes 25-30% of all clinically used
medications, including
 Dextromethorphan,
 ß-blockers(e.g., metoprolol),
 Antiarrhythmics,
 Anti-depressants (e.g., fluvoxamine, fluoxetine, imipramine,
nortriptyline),
 Antipsychotics (e.g., haloperidol, risperidone),
 Morphine derivatives, and many other drugs

30.  Drug metabolism via CYP450 enzymes
exhibit genetic variability(polymorphism) that
influences a patients response to a particular
drug.
 For ex 1 out of 15 whites or blacks may have
exaggerated response to std.doses of beta-
blockers or no response to analgesic-tramadol.

31.  The gene encoding CYP2D6 isoenzyme, has the most
variations of all genes for CYP isoenzymes, with more than
75 allelic variants identified to date, resulting from point
mutations, single base-pair deletions or additions, gene
rearrangements, and deletion of the entire gene.
 These mutations result in either a reduction or complete loss
of activity

32.  A specific gene encodes each cyp450 enzyme
 Every person inherits one genetic allele from each parent
 Alleles are referred to as “wild-type” or “variant” with wild type
occuring most commonly in the general population.
 An “Extensive”(i.e normal) metabolizers receives 2 copies of wild-
type alleles (i.e) EMs carry an autosomal dominant wild type gene
and may be homozygous or heterozygous for this allele.
 Polymorphism occurs when a variant allele repalces 1 or both wild-
type alleles .

33.  Persons with 2 copies of variant alleles are “Poor
metabolizers” and those with 1 wild-type and 1 variant allele
have reduced enzyme activity.
 Genotype-phenotype studies have revealed that poor
metabolizers possess two nonfunctional alleles and that the
phenotype is an autosomal recessive trait.
 Some persons inherit multiple copies of wild-type alleles,
which result in excess enzyme activity. This phenotype is
termed as “Ultra-rapid”metabolizers.

34.  An ultra-rapid metabolizer phenotype has been identified and
found to result from gene duplication (upto 13 copies of CYP2D6).
 Poor metabolizers are more likely to have adverse effects from
drugs that are substrates of the isoenzyme and decreased efficacy
from drugs requiring CYP2D6-mediated activation
 (e.g., codeine is converted into morphine by CYP2D6), while
extensive and ultra-rapid metabolizers may have therapeutic failure
with drugs activated by CYP2D6
 (e.g., standard antidepressant doses)

35.  The frequency of the phenotype of poor metabolizers differs
among ethnic groups. Less than 1% of Asians, 2-5% of
African-Americans, and 6- 10% of Caucasians are poor
metabolizers of CYP2D6.[4] The most common variant alleles
in Caucasians are CYP2D6*3, *4, *5, and *6, which account
for about 98% of poor metabolizers

36.  Genotyping CYP2D6 has been shown to successfully predict
the clearance of fluoxetine, fluvoxamine, desipramine,and
mexiletine.
 In some instances, the genotype for CYP2D6 has been useful
in predicting adverse effects associated with antidepressants
and neuroleptics.

37.  Impaired metabolism of drugs metabolized
by the CYP2C9 isoenzyme, such as
phenytoin, S-warfarin, tolbutamide,losartan,
and nonsteroidal antiinflammatory drugs
(NSAIDs) (e.g., ibuprofen, diclofenac,
piroxicam, tenoxicam, mefenamic acid) has
been noted.

38.  Three allelic variants of the CYP2C9 gene have been
identified that are associated with decreased enzyme
activity.
 The normal, or wild-type, variant is referred to as *1
("star 1"), the two polymorphic versions are *2 ("star
2") and *3 ("star 3"), and each person can carry any
two versions of the SNP. For example, a person with
two normal copies would be *1/*1, a person with only
one polymorphism could be *1/*2, and a person with
both polymorphisms could be *2/*3.The prevalence
of each variant varies by race;
 10% and 6% of Caucasians carry the *2 and *3
variants, respectively, but both variants are rare (<
2%) in those of African orAsian descent.

39.  CYP2C9*2 and *3 were associated with a 5.5-
and 27.0-fold decrease in the intrinsic
clearance of S-warfarin, respectively,
compared with the wild-type allele.
 Homozygous CYP2C9*3 alleles were found in
poor metabolizers of phenytoin, glipizide,
tolbutamide, and Losartan.

40.  Phenytoin is a substrate of both CYP2C9 and
CYP2C19 isoenzymes, but CYP2C9 is responsible
for its metabolism to a greater extent; thus,
mutant alleles encoding the CYP2CP9gene have
a greater effect on the clinical toxicity of
phenytoin.
 The CYP2C9*3 mutant allele occurs in
approximately 6-9% of Caucasians and Asians.
 CYP2C9*2 occurs in approximately 8-20% of
Caucasians and less frequently in African-
Americans and is virtually absent in Asians

41.  CYP2C9 genotyping may affect the clinical
use of warfarin because of the relatively high
prevalence of poor metabolizers, severe
outcomes as a consequence of drug
overdose, and frequency with which it is
prescribed.

42.  Warfarin is metabolized primarily via
oxidation in the liver by CYP2C9, and exerts
its anticoagulant effect by inhibiting the
protein vitamin K epoxide reductase
complex, subunit 1 (VKORC1).Three single
nucleotide polymorphisms (SNPs), two in
the CYP2C9 gene and one in
the VKORC1 gene, have been found to play
key roles in determining the effect of warfarin
therapy on coagulation.

43.  CYP2C9*1 metabolizes warfarin normally,
CYP2C9*2 reduces warfarin metabolism by
30%, and CYP2C9*3 reduces warfarin
metabolism by 90%. Because warfarin given
to patients with *2 or *3 variants will be
metabolized less efficiently, the drug will
remain in circulation longer, so lower warfarin
doses will be needed to achieve
anticoagulation

44.  In patients where genetic information is not
available but are at increased risk of bleeding
with standard dosing algorithms, guidelines
suggest starting with a reduced (2-5 mg)
initial dose and basing the frequency of
monitoring on the INR response.

45.  CYP2C19 isoenzyme metabolizes several
pharmacologically important therapeutic
agents.
 Extensive and poor metabolizers exist for S-
mephenytoin, omeprazole and other proton-
pump inhibitors, diazepam, propranolol,
imipramine, and amitriptyline

46.  Genetic polymorphism (mainly CYP2C19*2,
CYP2C19*3 and CYP2C19*17) exists for
CYP2C19 expression, with approximately 3–
5% of Caucasian and 15–20%
of Asian populations being poor
metaboliserswith no CYP2C19 function.

47.  pronounced pharmacodynamic effects tend
to be seen in Asians treated with omeprazole
because of the higher frequency of poor
metabolizers.

48.  CYP2C19 genotype might influence the cure rates
for Helicobacter pylori infection in patients with
peptic ulcers.
 The cure rate was 100% in poor metabolizers,
60% in patients with heterozygous genotypes,
and 29% in patients with homozygous wild-
types.
 This may be explained by the higher
accumulation of plasma omeprazole
concentrations in poor metabolizers, resulting in
a greater degree of gastric acid suppression.

49.  CYP3A iso-enzymes are the predominant
subfamily of CYP enzymes, making it one of the
most important drug metabolizing enzymes.
 The genes for CYP3A iso-enzymes are
expressed primarily in the liver and small
intestines.
 Hepatic CYP3A4 iso-enzyme has been estimated
to metabolize almost 50% of currently used
drugs as well as endogenous and exogenous
corticosteroids.

50.  IntestinalCYP3A4 isoenzyme contributes
significantly to the first-pass metabolism of
orally administered drugs.
 CYP3A activities are the sum of the activities
of at least three CYP3A isoenzymes: CYP3A4,
CYP3A5, and CYP3A7. Interindividual
variability due to CYP3A4 activity alone may
vary by up to 50-fold.

51.  It is estimated that CYP3A5 isoenzyme is only
present in 10-30% of liver samples tested.
 CYP3A5 isoenzyme was found to account for
at least 50% of the total CYP3A content in
people who carry the wild-type CYP3A5*1
allele.
 people with at least one wild-type CYP3A5*1
allele express large amounts of CYP3A5 iso-
enzyme, resulting in a 2.5-fold increase in
clearance of the probe drug (midazolam).

52.  The most common cause of loss of the
expression of the CYP3A5 gene in the liver is
an SNP at nucleotide 22,893 in intron 3
(CYP3A5*3), which causes alternative splicing
(exon 3B) and protein truncation.

53.  CYP3A5*1 is more frequently expressed in non-
Caucasian populations (30% of Caucasians,
Japanese, and Mexicans; 40% of Chinese; and
60% of African- Americans, Southeast Asians,
Pacific Islanders, and Southwestern American
Indians); thus, these populations may
metabolize CYP3A substrates more rapidly.
 CYP3A5 appears to be an important genetic
contributor to inter-individual and interracial
differences in CYP3A-dependent drug
metabolism.

54.  Prediction of drug interactions involving the
inhibition and induction of CYP3A will
continue to be a challenge and clinically
important because of the diverse role CYP3A
plays in the metabolism of currently available
and future drugs.

55.  Reference:
 Genetic Basis of Drug Metabolism. Margaret K.
Ma, Michael H.Woo, Howard L. Mcleod. American
Journal of Health-System Pharmacy. 2002;59(21)