This document discusses genetic polymorphisms in drug transporters and drug targets. It defines genetic polymorphisms as variations in gene sequences that occur in at least 1% of the general population. The most common type is a single nucleotide polymorphism (SNP) resulting from a change in a single nucleotide base pair. SNPs can be synonymous or non-synonymous, with non-synonymous SNPs potentially altering the protein's structure and function. The document outlines various drug transporters including P-glycoprotein and discusses genetic polymorphisms that can affect their expression and activity levels. It also discusses how genetic polymorphisms in drug metabolizing enzymes and drug receptors can influence drug response and side effects.

1. CLINICAL PHARMACOKINETICS AND
PHARMACOTHERAPEUTIC DRUG
MONITORING
GENETIC POLYMORPHISM IN DRUG
TRANSPORT AND DRUG TARGETS
SUBMITTED BY
PAVITHRA.V
V-PHARMD

2. GENETIC POLYMORPHISMS:
DEFINITION: Geneticpolymorphisms are variations in gene
sequences that occur in at least 1% of the general population,
resulting in multiple alleles or variants of a gene sequence.
Polymorphisms are distinct from mutations that occur in less
than 1% of the population.
INTRODUCTION:
The most commonly occurring form of genetic variability
is the single nucleotide polymorphism (SNP, often called
“snip”), resulting from a change in a single nucleotide base pair
within the gene sequence.
Synonymous SNPs in the coding region of a gene generally
result in no change in the amino acid sequence of the eventual
protein product. Non- synonymous SNPs in the coding region
will result in a change in the amino acid sequence of the protein.
In some cases, this alteration may have little effect on the
protein’s structure and function,
example, if one acidic amino acid is replaced by another.
However, non-synonymous SNPs have the potential to
drastically alter the function of .An example of such an effect
occurs if nucleotideposition 2935 of the CYP2D6 gene has a C
instead of an A (c.2935A>C).
During translation this results in the insertion of a proline
instead of histidine at amino acid position 324 gen- erating the
CYP2D6*7 allele, with no drug metabo- lizing activity Genetic
variants that result from the insertion or deletion of a nucleotide
in the coding region are also classified as SNPs. Since the
mRNAs from genes are translated to protein in 3-nucleotide

3. codons, such insertions or deletions can have a significant
effect on the eventual protein product. An example of such a
polymorphism is the CYP2D6*3 allele where a sin- gle
nucleotide deletion(A2637)resultsin a frame shift in translation
that produces an enzyme with no cata- lytic activity
Each variant of a gene is represented by the star designation
(*) followed by a number, and each gene could potentially
containmul-tiple variants. A groupingof select variants is called
a haplotype and results in unique combinations of poly-
morphisms with potentially novel phenotypes.
Single nucleotidepolymorphisms outside the coding region
of the gene can result in altered levels of protein activity as well.
Polymorphisms in the promoter sequence of a gene can
influence gene transcription rates resulting in greater or lesser
amounts of mRNA, and consequently protein expres- sion.
Alternatively, SNPs in a splicing control region of the gene
can result in the production of a unique protein often missing
one or more exons and result- ing in a unique(often truncatedor
inactive) protein.
In some cases, multiple copies of a gene on a chromosome
can result in increased levels of protein being expressed, and
once again the CYP2D6 gene serves as a relevant example.
However, it isn’t difficult to see that a mixed form of CYP
gene expression due to genetics and drug induction could
increase metabolic activity to an even greater extent.
Deletion or inversion of entire genes on the chromosome
would obviously have the opposite effect on enzyme activity
and drug metabolism.

4. DRUG TRANSPORTERS
DEFINITION:
•Transportersare those proteins that carry either endogenous
compoundsor xenobioticsacross biological membranes.
INTRODUCTION:
•They can be classified into either efflux or uptakeproteins,
dependingon the direction of transport.
•The extent of expression of genes coding for transport proteins
can have a profound effect on the bioavailability and
pharmacokinetics of various drugs.
•Additionally, genetic variation such as single-nucleotide
polymorphisms (SNPs) of the transportproteins can cause
differences in the uptake or efflux of drugs.
•In terms of cancer chemotherapy, tumor cells expressing these
proteins can have either enhancedsensitivity or resistance to
various anticancerdrugs.
•Transportersthat serve as efflux pumps on a cell membrane can
remove drugs from the cell before they can act.
•Transport proteinsthat are responsiblefor the vital influx of
ions and nutrientssuch as glucose can promotegrowth of tumor
cells if overexpressed, or lead to increased susceptibility for a
drug if the transportercarries that drug into the cell.
TYPES OF DRUG TRANSPORTER
Two types of transporter:

5. •ATP binding Cassette(ABC) – Found in ABCB, ABCD and
ABCG family. Associated with multidrug resistance (MDR) of
tumor cells causing treatment failure in cancer.
•SoluteCarrier(SLC) – Transport varieties of soluteinclude
both charged or uncharged
P-glycoprotein
• ATP binding cassette sub family B member- 1 (ABCB 1)
• Multidrugresistance protein 1 (MDR1)
• Transport various molecules, including xenobiotic, across cell
membrane
• Extensively distributed and expressed throughout the body
Mechanism of Pglycoprotein
 Substratebind to P-gp form the inner leaflet of the
membrane
 ATP binds at the inner side of the protein
 ATP is hydrolyzed to produceADP and energy

6.  Substrateis excreted outside the cell
TRANSPORT
ERS
GENETIC POLYMORPHISM

7. MDR1 (P-
Glycoprotein)
 The MDR1 or ABCB1 gene codes for the
efflux protein P-glycoprotein (P-gp) that is
frequently asso- ciated with drug resistance to
antineoplastic agents including vincristine and
doxorubicin.
 There are many PGP substrates and
inhibitors as outlinedin Chapter11. At least 66
SNPs in the ABCB1 gene have been reported,
and the three most studied SNPs include two
synonymous and one non-synonymous
variants.
 The synonymous SNPs are reported to
result in decreased expression of PGP due to
decreased mRNA expression, unstable mRNA,
or alterations in protein folding.
ABC
Transporters
 T
he multidrug resistance-associated proteins
(MRPs) are members of the ATP-binding
cassette (ABC) superfamily with six members
currently, of which MRP1 (ABCC1), MRP2
(ABCC2), and MRP3
 (
ABCC3) are commonly known to effect drug
dispo- sition. Like MDR, these transporters
can also be expressed in cancer cells, which
confer resistance to the chemotherapeutic
agent tamoxifen. It appears that
polymorphisms in this family are rare and
occur at different frequencies among different
populations.
 F

8. uture studies with specific substrates and
polymorphisms may ultimately provide
additional information on the variable
responses or adverse effects of drugs.
Solute Carrier
Transporters
 Another important class of drug
transporters is the solute carriers (SLCs) such
as the organic anion transporter protein
(OATP) and organic cation trans- porter
(OCT).
 These transportersare located throughout
the body and have various roles in the
transport of many different drugs. OATP1B1
(coded by the SLCO1B1 gene) is a hepatic
influx trans- porter with at least 40 non-
synonymous SNPs identi- fied that result in
either an altered expression or activity of
OATP1B1
 While the clinical consequences of all of
these SNPs are unknown, one SNP
(c.521T>C) has been associated with an
increased risk of simvastatin-induced myop-
athy
 This non-synonymous SNP is associated
with a lower plasma clearance of simvastatin
and is found in the SLCLO1B1*5, *15, and
*17 alleles.
 These alleles are present in most
populations with a frequency between 5%
and 20% and warrant the avoidance of high-
dose simvastatin (>40 mg) or treatment with
another statin to decrease the risk of

9. simvastatin- induced myopathies .
GENETIC POLYMORPHISM IN DRUG
TARGETS:
DEFINITION:
Drug Target is an biological agent in which the drug is directed
and or binds to it resulting in a change in its behaviour or
function . examples : proteins and nucleic acids
A) GENETIC POLYMORPHISM IN DRUG
METABOLISING ENZYMES:
 Polymorphisms have been reported in both phases of drug-
metabolizing enzymes and can affect the phar- macokinetic
profile of a drug for a given patient.
 Understandinga patient’s genetic determinantsof drug
metabolism and the consequencesof these polymorphisms could
be used to design optimum, personalized dosing regimens in the
clinic that would avoid adverse reactions or treatment failures
due to subtherapeuticdoses.
 While this may appear perfectly logical, the redundancyof
drug metabo- lism and potential contributionfrom numerous
otherfactors (such as diet, other drugs, age, weight, etc) make it
difficult to translateenzyme status data to a clinical decision.
For example, warfarin ther-apy is complicated by a
combination of metabolic (CYP2C9 polymorphisms contribute
2%–10%), pharmacodynamic (VKORC1 polymorphisms con-

10. tribute 10%–25%), and environmental factors (20%–25%
contribution).
 Several algorithms that take into account genetic
information have been developed for warfarin dosing and some
are avail- ableonline
 While these appear to be useful tools to account for genetic
differences, the reported effectiveness of achieving an optimal
anti- coagulant dose of warfarin using algorithms is vari- able
 These confoundingresults demonstratethe need for more
investigation into the factors (including pharmacokineticand
pharmacodynamic factors) that contributeto variable responses,
as well as robust clinical investigations to validate these
observations.
 There are 70 drugs that includephar- macogenetic
information related to polymorphisms in drug-metabolizing
enzymes that contributeto variabledrugresponse
 Drugs that are thought to be affected by the polymorphisms,
the consequence,and label information are included in Table
Clinically Important Genetic Polymorphisms of Drug
Metabolism and Transporters That Influence Drug
Response
Enzy
me
Drug Drug
Effect/
Side
Effect
(Pharmacoge
netics )
CYP2
C9
Warfarin Hemorr
hage
Actionable
Tolbutami Hypogl -

11. de ycemia
Phenytoin Phenyto
in
toxicity
-
Glipizide Hypogl
ycemia
-
Losartan Decreas
ed
antihyp
ertensiv
e effect
-
CYP2
D6
Antiarrhyt
hmics
Proarrh
ythmic
and
other
toxic
effects
in poor
metabol
izers
-
Antidepre
ssants
Ineffica
cy in
ultrarap
id
metabol
izers
Actionable/Inf
ormation
Antipsych
otics
Tardive
dyskine
sia
Actionable/Inf
ormation
Eliglustat Ineffica
cy in
ultrarap
Testing
recommended

12. id
metabol
izers
Opioids Inefficac
y of
codeine
as
analgesi
c,
narcotic
side
effects,
depende
nce
Actionable
Pimozide Toxicit
y with
high
dosein
poor
metabol
izers
Testing
recommended
Tetrabena
zine
Toxicit
y with
high
dose in
poor
metabol
izers or
ineffica
cy in
ultrar-
apid
metabol
Testing
recommended

13. izers
Warfarin Higher
risk of
hemorr
hage
-
-
Adrenocep
tor
antagonist
s
Increase
d
blockad
e
Actionable/Inf
ormation
CYP2
C19
Omeprazo
le
Diazepam
Clopidogre
l
Higher
cure
rates
when
given
with
clarithr
omycin
Prolong
ed
sedation
Ineffica
cy in
poor
metabol
izers
Information
Actionable
Testing
recommended
Dihyd
ropyri
midin
e
dehyd
Fluoroura
cil
Myelot
oxicity,
neuroto
xicity
Actionable

14. rogen
ase
Plasm
a
pseud
o-
cholin
estera
se
Succinylc
holine
Prolong
ed
apnea
-
N-
acetyl
transf
erase
Sulfonami
des
Hyperse
nsitivity
-
Amonafid
e
Myelot
oxicity
-
Procaina
mide
Drug-
induced
lupus
erythem
atosus
-
Hydralazi
ne
Drug-
induced
lupus
erythem
atosus
Information
Isoniazid Drug-
induced
lupus
erythem
atosus
Information

15. GENETIC POLYMORPHISM IN DRUG
RECEPTORS:
Genetic polymorphismin drug receptorshas leads to various
effects from severe to null reaction due to drugs.
IMPORTANCE:
This has been reported with the breast cancer agent
tamoxifen .
Tamoxifen has an active metabolite(endoxifen) pro- duced
by CYP2D6 that is thought to be responsible for much of
its antiestrogenic activities. The patient with the PM
phenotype would not metabolize tamoxifen to the active
metabolite and, therefore, does not benefit from clinically
relevant endoxifen concentrations .
Genotypically, PM have two null alleles, which do not
code for functional CYP2D6 due to a frame shift
(CYP2D6*3 and *6), a splicing defect (CYP2D6*4), or a
gene deletion (CYP2D6*5).

16. The UM have very high rates of CYP2D6 enzy- matic
activity resulting in low plasma concentrations of drugs
with consequent lower efficacy. Active drugs like the
tricyclic antidepressant amitriptyline may require doses
several-fold higher than standarddoses to achieve
therapeutic activity when the patient is a UM. On the other
hand, drugs that require metab- olism to an active
metabolite are extremely active, with potentially serious
consequences.
Understandingthis complex interplaybetween all the
different allelesof CYP2D6 and the many drugs that it
metabolizes provides a great opportunityfor accurate
genotyping to provide for sound clinical decisions to
prevent adverse events and prevent therapeuticfailures
Examples of Polymorphisms Affecting Drug Receptors and
Enzymes Showing Frequencyof Occurrence
Enzyme/Re
ceptor
Frequency of
Polymorphism Drug Drug
Effect/Side
Effect
CYP2C9 14%–28%
(heterozygotes)
Warfarin Hemorrhage
Tolbutamide Hypoglycemia
0.2%–1%
(homozygotes)
Phenytoin Phenytoin
toxicity
Glipizide Hypoglycemia
Losartan Decreased
antihypertensive
effect
CYP2D6 5%–10%
(poor
Antiarrhythmic
s
Proarrhythmic
and other toxic

17. metabolizers) effects
Toxicity in poor
metabolizers
1%–10%
(ultrarapid
metabolizers)
Antidepressants Inefficacy
in
ultrarapid
metabolize
rs
Antipsychotics Tardive
dyskinesia
Opioids Inefficacy of
codeine as
analgesic,
narcotic side
effects,
dependence
Warfarin Higher risk of
hemorrhage
-
Adrenoceptor
antagonists
Increased—
blockade
CYP2C19 3%–6%
(whites)
8%–23%
(Asians)
Omeprazole
Diazepam
Higher cure
rateswhen
given with
clarithromycin
Prolonged
sedation
Dihydropyri
midine
dehydrogena
se
0.1% Fluorouracil Myelotoxicity,
Neurotoxicity
Plasma 1.5% Succinylcholine Prolonged apnea

18. pseudo-
cholinesteras
e
N-
acetyltransfe
rase
40%–70%
(whites)
10%–20%
(Asians)
Sulphonamides
Amonafide
Procainamide,
hydralazine,
Isoniazid
Hypersensitivity
Myelotoxicity
(rapid
acetylators)
Drug-
induced
lupus
erythemat
osus
Thiopurine
methyltransf
erase
0.3% Mercaptopurin
e, thioguanine,
azothioprine
Myelotoxicity
UDP-
glucuronosyl
transferase
10%–15% Irinotecan Diarrhea,
myelosuppressio
n
ACE Enalapril,
lisinapril
captopril
Renoprotective
effect, cardiac
indexes, blood
pressure
Potassium
channels
Quinidine Drug-induced
QT syndrome
HERG Cisapride Drug-induced
torsade de
pointes
KvLQT1 Terfenadine
disopyramide
Drug-induced
long-QT
syndrome
VKORC Warfarin Over-
anticoagulation

19. Epiderma
l growth
factor
receptor
(EGFR)
Gefitinib Certain
polymorphs
susceptible
HKCNE2 Meflaquine
clarithromycin
Drug-induced
arrhythmia
CYP1A2
CYP1A2 activity varies widely with genetic poly- morphisms
contributing to observed differences in levels of gene
expression. CYP1A2 is responsible for the metabolism of
about 5% of marketed drugs including fluvoxamine, clozapine,
olanzapine, and theophylline. Approximately 15% of the
Japanese, 5% of the Chinese, and 5% of the Australian popula-
tions are classified as CYP1A2 poor metabolizers.
CYP2C9
CYP2C9 has at least 30 different allelic variants with the two
most common being CYP2C9*2 and *3. Both of these
variants result in reduced CYP2C9 activity and are carried by
about 35% of the Caucasian population. CYP2C9 is a major
contributor to themetabolism of the narrow therapeutic index
blood thinner warfarin.
CYP2C19
CYP2C19 is a highly polymorphic drug-metabolizing enzyme
with at least 30 variants reported (The Human Cytochrome
P450 Allele Nomenclature Database, 2013). Polymorphisms
in CYP2C19 result in vari- able drug response to clopidogrel
and several antide- pressants.

20. CYP3A4
CYP3A4 is the most abundant CYP450 in the liver and
metabolizes over 50% of the clinically used drugs (Fig. 13-2).
In addition, the liver expression of CYP3A4 is variable
between individuals. To date, over 20 allelic variants of
CYP3A4 have been iden- tified (The Human Cytochrome
P450 Allele Nomenclature Database, 2013).
Other PhaseI Enzymes
While the CYP450s are the most abundant and exten- sively
studied phase I drug-metabolizing enzymes, others have
polymorphisms that have an effect on the clearance (or
activation) ofdrugs and, therefore, affect the clinical outcomes
of patients secondary to, at least partially, changes in
pharmacokinetics.
PLASMA PSEUDOCHOLINESTERASE OR SERUM
BUTYRYLCHOLINESTERASE
Plasma pseudocholinesterase is responsible for the
inactivation through ester hydrolysis of the neuro- muscular
blockers succinylcholineand mivacurium. While mivacurium
is no longer marketed in the US market, succinylcholine is
used to provide skeletal muscle relaxation or paralysis for
surgery or mechan- ical ventilation. There are at least 65
allelic variants of pseudocholinesterase that have been
identified in approximately1.5% of the population that result
in various levels of pseudocholinesterase deficiencies
(Soliday et al, 2010). These allelic variants include non-
synonymous point mutations or frame shift mutations that
result in a PM phenotype for succi- nylcholine..

21. DIHYDROPYRIMIDINEDEHYDROGENASE(DPD)
DPD is the first reduction and rate-limited step in breakdown
of the pyrimidine nucleic acids and their analogs.
Polymorphisms in DPD result in a loss of enzymatic activity
leading to the accumulationof the chemotherapeutic agent 5-
flourouracil (5-FU), which leads to significant toxicity
includingleukopenia, thrombocytopenia, and stomatitis. It is
estimated that approximately 3%–5% of population has low
or defi- cient DPD activity .There are three alleles, each with
low fre- quency, that appearto account for the majority of the
deficient DPD activity observed and more than 20% of the
serious toxicity observed with 5-FU adminis- tration.
DPYD*2A is the most common allelic vari- ant, although the
exact frequency is not clear.
PHASE II ENZYMES
As discussed in the previous chapter (drug metabo- lism),
phase II drug-metabolizing enzymes are com- monly referred
to as transferases and perform conjugation reactions that add
a biochemical com- pound to a xenobiotic to facilitate its
elimination. Just like the phase I reactions, there are genetic
variations in the several phase II enzymes that influ- ence the
pharmacokinetics of drugs.
THIOPURINE S-METHYLTRANSFERASE
Thiopurine drugs including 6-mercaptopurine (MP) and
azathioprine are used for their anticancer and
immunosuppressive properties but can have signifi- cant
adverse effects including myelosuppression. The phase II
metabolizing enzyme thiopurine S-methyltransferase
(TPMT) is involved in the deg- radation of thiopurine drugs

22. and TPMT polymor- phisms account for about one-third of
the variable responses to MP and azathioprine. While TPMT
alone only explains one-third of the variability, other factors
are known to contribute, which highlights the challenge and
multifactorial nature of personalized medicine to account for
intraindividual differences. At least twenty-eight allelic
variants in the coding and splic- ing region of TPMT have
been identified with most of the null phenotypes being
associated with TPMT*2, TPMT*3A, and TPMT*3B alleles
result- ing in non-synonymous mutations that lead to the
production of an unstable enzyme and reduced activ- ity
overall. The loss of TPMT function is present in about 5% of
the Caucasian population and results in accumulation of MP
leading to an increased risk for adverse effects like
leukopenia
URIDINE DIPHOSPHATE (UDP)-
GLUCURONOSYLTRANSFERASE
UDP-glucuronosyltransferase (UGT) is a super- family of
phase II drug-metabolizing enzymes that produce
glucuronidation metabolites through conju- gation reactions
(see Chapter 12). Like the CYP450s, the UGTs are divided
into families identified with numbers (UGT1, UGT2, etc) and
subfamilies identi- fied with letters (UGT1A, UGT2B, etc)
based on amino acid similarities. Drug metabolism is cata-
lyzed almost exclusively by UGT1 and UGT2 (Meech et al,
2012). At least 200 alleles for UGT1 and UGT2 gene families
have been reported causing changes in enzymatic activity or
expression levels that may contribute to individual variations
in drug response (UGT Alleles Nomenclature Home Page,
June 2005).

23. N-ACETYLTRANSFERASE
N-acetyltransferase (NAT) was identified as a poly- morphic
enzyme through phenotypic observations of fast or slow
acetylatorsof the anti-tuberculosis drug, isoniazid (Evans and
White, 1964). There are two different human genes, NAT1
and NAT2, that code for functional NAT activity. While both
NAT1 and NAT2 are polymorphic, the fast and slow
acetylator phenotype is associated with the NAT2 gene. The
slow acetylator phenotype is found in about 50% of
Caucasians, 90% of Arabs, and 10% of Japanese populations

24. REFERENCE:
1)Shargeland yu’s “APPLIED BIOPHARMACEUTICS
AND PHARMACOKINETICS” – Seventh Edition by leon
shargeland AndrewB.C . YU