Pharmacogenomics mine

• Pharmacogenomics is the study of inherited differences in
interindividual drug disposition and effects, with the goal of
selecting the optimal drug therapy and dosage for each
patient.
i.e how genetic inheritance influences response to drugs.

…..based on a patient’s genetic profile.

• Using the individual genetic characteristics, such as gene
polymorphisms, may help to predict drug efficacy and toxicity.
• Gene polymorphisms consist of minor changes in DNA
sequence: substitutions, deletions and insertions, repeats,
gene copy number variations and sometimes more important
rearrangements.

Gene polymorphisms
Errors occurring during DNA replication or lesions induced by
mutagenic agents may lead to the
• Substitution : replacement of a nucleotide by another one
• Deletion : loss of a nucleotide
• Insertion : addition of a nucleotide
When they occur within the coding sequence of a gene,
the protein originating from this gene may bear a structural
alteration which may lead to its instability, a decrease or increase
in its activity or the loss of its functionality.
• Insertions and deletions alter the reading frame, therefore
change the complete downstream sequence of the protein,
and are generally non-sense mutations.

Mutations :
• Synonymous or silent : when there is no change of the amino-
acid encoded
• Missense : when there is a replacement of the amino-acid by
another one
• Non-sense : when they lead to a truncated protein, because
of the occurrence of a stop codon or of the alteration of a
splicing site, giving rise to a truncated protein.
When mutations occur within the gene regulatory
regions, especially in the 30 and 50 untranslated regions (UTR),
or in intronic sequences, they can lead to alterations in protein
expression, which are generally less deleterious than mutations
affecting protein sequence .

Single nucleotide polymorphisms
(SNPs)
• Numerous polymorphisms which involve only one nucleotide,
either by substitution, insertion or deletion.
Some polymorphisms are more complex:
• VNTR (Variable number of tandem repeats : they concern the
number of minisatellite or microsatellite repeats
• CNV (Copy number variations) : number of copies of a gene ,
which play a major role in the level of the mRNA and the
protein produced.

Harnessing and understanding the genetic code of both
the patient (germline) and the tumor (somatic)
provides the opportunity for personalized dose and
therapy selection for cancer patients.

BIOMARKERS
• It is a characteristic that is objectively measured and
evaluated as an indicator of normal biological processes,
pathogenic processes, or pharmacologic responses to a
therapeutic intervention.
• A pharmacogenetic or pharmacogenomic biomarker is
defined as any “genetic or genomic marker that is associated
with drug response

TYPES
• A prognostic biomarker is a marker, or measurable trait, that
provides information on the likely course of cancer, including
aggressiveness of disease, regardless of treatment.
• Eg :
1. 70-gene profile MammaPrintTM10 or 21-gene profile
Oncotype Dx11 for estrogen/ progesterone receptor-positive,
lymph node-negative breast cancer.
2. Microsatellite instability (MSI) in colorectal cancer patients.
3. BRAF mutations are considered a poor prognostic biomarker
conferring aggressive disease in colorectal cancer

• A predictive biomarker is a marker, or measurable trait, that
can be used to identify patients most likely to benefit from
treatment and/or those predisposed to toxicity.
1. Somatic mutations : Genetic variations found within the
tumor DNA ,but not DNA from the normal (germ line)
tissues,which also have functional consequences that
influence disease outcomes and/or response to certain
therapies.
1. Germline mutations :These are heritable variations found
within the individual and, in practical terms, are focused on
DNA markers predictive for toxicity or therapeutic outcomes
of a particular therapy as well as inheritable risk of certain
cancers

PHARMACOGENOMICS OF TUMOR
RESPONSE

EGFR
• Activating mutations in the EGFR gene ~20% of NSCLC cases
and results in constitutive signaling via the PI3K-AKT and
MAPK pathways.
• Deletions in exon 19 and a missense mutation at exon 21
(arginine to leucine substitution [L858R]) account for >90% of
all EGFR mutations and are considered a positive predictive
biomarker for small-molecule TKIs targeting the EGFR domain
• Overexpression of EGFR or KRAS determines the response of
tumor to TKI’s .

EGFR mutations- 1st Gen TKI’S
EGFR
L858R or
delL747-S752
T790M ,
T854A, D761Y, L747S
Enhanced advantages to ATP
competitive inhibitors by
creating a site with increased
affinity to the compounds
results in stronger affinity for ATP,
reducing the ability of erlotinib,,
to bind to the EGFR tyrosine
kinase domain.

• Osimertinib is an irreversible inhibitor with over 600 times the
killing capacity of Gefitinib (and over 40 times that of Afatinib)
against T790M/L858R cancer cells .
• Osimertinib received accelerated FDA approval in 2015, while
another T790M inhibitor, rociletinib, is also currently being
evaluated by the FDA for approval.
• Osimertinib resistance : Mutations around the gatekeeper
site (such as the L792 position, G796D, C797S, etc.) causes
resistance to Osimertinib
• Non-EGFR gene-derived instances of resistance include Met
amplification and the notorious V600E mutation in the BRAF
proto-oncogene (which is downstream of EGFR)

ALK
• The abnormal fusion gene, EML4-ALK, resulting in constitutive
activation of anaplastic lymphoma kinase (ALK), occurs in ~5%
of all non-small-cell lung cancer (NSCLC) cases.
• Normally, they function towards proper neuronal
development and microtubule formation, respectively,
however when fused, they can cause development of lung
cancer by irregular activation of their downstream targets
• The translocation activates various growth stimulatory
signaling pathways, including RAS-MEK-ERK, janus kinase 3
(JAK3)-STAT3, and PI3K-AKT pathways

Crizotinib :
• Competitive inhibitor at the
ATP binding pocket to shut
down carcinogenic kinase
activity
• Also has anti-ROS1 and C-
Met activities
Resistance to Crizotinib :
• L1196M- prevents the
interaction between Crizotinib and
the ATP-binding pocket.
• G1202R
• S1206Y
• C1156Y - conformational
changes to the entire binding
cavity
reduce affinity to
Crizotinib by changing the
solvent-exposed region

Resistance to Crizotinib
• Activation of other
pathways EGFR, ROS1,
KIT, MET
• Secondary resistant
ALK translocations
Crizotinib -- overall response rate (ORR) of 60.8% in ALK positive
cases
2ND GENERATION ALK
inhibitors :
1. Ceritinb
2. Alectinib
3. Brigatinib - ALK
L1196M, EGFR
T790M, and
Osimertinib-
resistant EGFR
C797S also

BRAF
• 50% of cutaneous metastatic melanoma cases carry mutations
in BRAF.
• V600E-- 90% of cases , causes change in protein amino acid
sequence, Valine at amino acid 600 to GlutaminE
• Next common : V600K – valine to lysine
• Activating BRAF mutations result in constitutive activation and
downstream signaling of the MAPK pathway, promoting cell
proliferation and tumor growth

COBAS 4800 BRAF V600 Mutation Test

Vemurafenib
• Potent inhibitor of BRAF V600E,
Phase III trial,
• 675 treatment-naïve patients
• Dacarbazine vs vemurafenib.
• 63% reduced risk of death (HR 0.37; P,0.001) and a 74% reduced risk of
disease progression in the vemurafenib arm compared to dacarbazine (HR
0.26; P,0.001).
Resistance to BRAF inhibtors :
• resistance eventually starts in most patients with a mean time to diagnosis
of first lesion at ∼3 months and a median PFS of 6–8 months.
• Resistance mechanisms:
RAS and MEK mutations resulting in paradoxical activation of the MAPK
pathway.

• Combining a BRAF inhibitor with a MEK inhibitor has been a
logical approach to combat BRAF inhibitor resistance.
• 2015 : These drugs received FDA approval to be used in
combination to treat patients with BRAF-positive unresectable
or metastatic melanoma.

BCR-ABL
• The BCR-ABL translocation occurs in 95% of CML cases and
results in constitutive activation of signal transduction
pathways associated with cell proliferation and tumor growth

• Imatinib, nilotinib, and ponatinib all bind to the inactive
conformation of the c-ABL kinase, preventing the
conformational switch to the active form,
• While dasatinib inhibits both the active and inactive
conformations.
• Nilotinib has a higher affinity for c-ABL kinase than imatinib,
resulting in greater selectivity and potency.
• Approximately, 10%–15% of CML patients will present with
resistance to imatinib and ∼20%–25% will develop resistance
over time

• The T315I point mutation – 315th amino acid ,threonine
changes to isoleucine
• Current TKIs require threonine at position 315 in order to bind
to their targets.
• Ponatinib features a carbon–carbon triple bond, which limits
steric hindrance, ultimately overcoming initial resistance.

• The HER2 gene is a member of the type 1 tyrosine kinase
growth factor receptor family that is found on the long arm of
chromosome 17.
• The biology of the HER2 receptor is complex, but it is
involved in both cell differentiation and proliferation.
• HER2 protein is over-expressed in 25-30% of human breast
cancers; in 90-95% of these cases over-expression is a direct
result of gene amplification.
• HER2 protein over-expression correlates with poor clinical
prognosis.

• HER2 testing is carried out principally by two methods in the
UK:
a. IHC identifies HER2 receptor over-expression
b. FISH identifies HER2 gene amplification.
• Current UK recommendations for HER2 testing -Two phase
testing is recommended,involving both IHC and FISH.

• Trastuzumab is a humanized monoclonal antibody that binds
specifically to the HER2 receptor and suppresses cell
proliferation that is driven by overexpression of the HER2
protein
• However, only 25% to 30% of patients with HER-2/neu–
positive breast cancers will respond to this mAb.

• Metastatic colorectal disease has achieved a certain
improvement in terms of survival and outcome benefit with
the introduction of novel targeted molecules to current
practice.
• Yet, a considerable proportion of patients still experience
disease progression after a short treatment period and seem
to not respond to the therapies, probably due to the onset of
resistance mechanisms and the overlap of alternative
pathways downstream of the blockade point

• RAS mutations have an adverse effect on the prognosis .
• RAS mutation status remains the most relevant biological
marker of response to anti-EGFR treatment.
• All patients with mCRC who are candidates for anti-EGFR
antibody therapy should have their tumor tested for mutations
in RAS.
• Constitutive mutations of RAS predict resistance to anti-EGFR
MoAbs in CRC.

• However, many patients (~40%) with a wild-type RAS status
do not respond to anti-EGFR mAbs.
• This occurs due to mutations in other downstream
components of the EGFR signalling pathway (BRAF, NRAS, and
PIK3CA).
• BRAF V600E mutations are detected in 8–10% early stage and
4–5% late stage CRC, being a strong indicator of very poor
prognosis.
• These mutations have not shown a clear predictive effect
related to anti-EGRF therapies in randomized clinical trials
,thus no changes in the label by any regulatory authority have
been made.

PHARMACOGENOMICS OF
CHEMOTHERAPY
DRUG TOXICITY

• Individual’s genetic constitution is an important regulator of
variability in drug effect.
• Differences in drug effects are more pronounced between
individuals compared to within an individual.
• The majority of pharmacogenomic examples affecting adverse
events or efficacy from cytotoxic drugs involve hepatic
metabolizing enzymes that detoxify or biotransform
xenobiotics .

THIOPURINES
Thiopurines are a family of drugs that includes
• Mercaptopurine (childhood ALL treatment )
• Thioguanine (acute myeloblastic leukemias)
• Azathioprine (solid organ transplants, rheumatic disease, and
dermatologic disorders)

Thiopurines are inactive prodrugs that require metabolism to
TGN to exert cytotoxicity.
• Activation :
Hypoxanthine phosphoribosyl transferase (HPRT).
• Inactivation :
a. Oxidation by xanthine oxidase (XO)
b. Methylation by Thiopurine methyltransferase (TPMT)

• TPMT is a cytosolic enzyme that catalyzes S-methylation of
thiopurine agents, resulting in an inactive metabolite, thereby
shunting drug away from TGN formation
• TPMT polymorphisms have been associated with the
therapeutic efficacy and toxicity of mercaptopurine.
• TPMT activity is highly variable and polymorphic in all large
populations
• Approximately 90% of individuals have high activity, 10% have
intermediate activity, and 0.3% have low or no detectable
enzyme activity

• The variable activity results from polymorphism in the TPMT
gene, located on chromosome locus 6p22.3
• While eight TPMT alleles have been identified, three alleles
(TPMT*2, TPMT*3A, TPMT*3C) account for about 95% of
intermediate or low enzyme activity cases
• All three alleles are associated with lower enzyme activity
due to enhanced rates of proteolysis of the mutant proteins

• TPMT deficiency results in higher intracellular activation of
6MP to form thioguanine nucleotides, resulting in severe or
fatal hematologic toxicity from standard doses of therapy .
• The presence of TPMT*2, TPMT*3A, or TPMT*3C is predictive
of TPMT activity.
• Patients heterozygous for these alleles all have intermediate
activity and subjects homozygous for these alleles are TPMT
deficient .
• Compound heterozygotes (TPMT*2/3A, TPMT*2/TPMT*3C,
TPMT*3A/3C) are also TPMT deficient .

• TPMT genotype tests are now available commercially in a
Clinical Laboratory Improvement Amendments (CLIA)-certified
environment.
• To date, patients homozygous for TPMT variant alleles appear
to tolerate 10%, and heterozygotes appear to tolerate 65% of
the recommended doses of 6MP, with no apparent decrease
in clinical efficacy

Clinical Pharmacogenomics
Implementation Consortium (CPIC)
Guidelines
• Homozygous wild-type patients be started at the full standard
dose.
• Heterozygous patients should start with reduced doses at 30%
to 70% of the full dose with adjustments made after 2 to 4
weeks based on myelosuppression and disease-specific
guidelines.
• Homozygous variant patients should start with 10% of the full
dose. Adjustments should be made after 4 to 6 weeks based
on myelosuppression and disease-specific guidelines

5-FU
• 5-FU is a prodrug that requires activation to 5-fluoro-2-
deoxyuridine monophosphate (5-FdUMP), which inhibits
thymidylate synthase
• TS inhibition results in impaired de novo pyrimidine synthesis
and suppression of DNA synthesis
• Approximately 85% of a 5FU dose is catabolized by
dihydropyrimidine dehydrogenase (DPD) to inactive
metabolites.
• Therefore, DPD is a primary regulator of 5FU activity .

• DPD enzyme, exhibits up to 20-fold variation in activity among
individuals
• Patients with low DPD activity cannot effectively inactivate 5-FU,
leading to excessive amounts of 5-FdUMP, causing gastrointestinal,
hematopoietic, and neurological toxicities.
• The molecular basis of genetic DPD deficiency is complex. To date,
at least 20 mutations associated with reduced DPD activity are
found.
• General population :
a. 3 – 5% - heterozygous carriers of mutations that inactivate DPD
b. 0.1% of individuals are homozygous for mutations that inactivate
DPD

• Most common inactivating allele of DPD accounts for about
50% of known nonfunctional alleles, and is caused by a G 
A transition at the invariant GT splice donor site, flanking
exon 14 of the DPYD gene .
• This allele (known as DPYD*2A) causes the skipping of exon
14, and leads to the production of a nonfunctional protein

• DPYD*2A is not the only mechanism for severe 5-FU toxicity.
• There are additional DPD mutations that have been
associated with impaired enzyme activity, including DPYD *3
and DPYD*13
• Many patients with severe 5-FU toxicity have no detected
mutations in the coding region of the DPYD gene .
• The complex nature of the molecular mechanisms controlling
DPD activity in vivo complicate the application of DPD
pharmacogenetics for the prospective identification of
patients likely to experience severe 5-FU toxicity.

CPIC guidelines
• Standard dosing for homozygous wild-type patients.
• Reduce the dose by at least 50% in heterozygous patients
(*1/*2A) is recommended, followed by dose adjustment
based on toxicity and/or pharmacokinetic testing.
• The use of an alternative agent is recommended in
homozygous-variant patients (*2A/*2A).
There are many patients with severe 5FU toxicity that
have normal DPD activity. This highlights that many factors,
including multiple genes, are potential causes of 5FU toxicity,
and there will not be one simple test to avoid this important
clinical problem.

Thymidylate Synthase
• Genetic polymorphisms in the gene encoding thymidylate
synthase have also been shown to influence response to 5-FU
therapy .
• Both TS mRNA and protein levels are inversely related to
clinical antitumor response
• Survival of patients with advanced colorectal cancer is inferior
if high TS expression is present

• The expression of TS is controlled in part by a polymorphism
characterized by a multiple number of tandem repeats of a
28-bp sequence in the 5V-promoter enhancer region (TSER) of
the gene.
• Alleles containing two, three, four, five, and nine copies of the
repeated sequence have been described (TSER*2, TSER*3,
TSER*4, TSER*5, and TSER*9), with TSER*2 and TSER*3 being
the predominant alleles in all populations studied to date
• Increasing the number of repeats leads to an increase in TS
mRNA levels and protein expression .

Patients homozygous for TSER*3 have a higher TS activity and
a poorer response to 5-FU therapy than patients homozygous
for TSER*2

• These studies suggest that combined genotyping of DPYD and
TSER functional variants might be useful in selecting patients
who are likely to tolerate and respond to 5-FU therapy.
• This is particularly important because of the availability of
other active antitumor agents (i.e., irinotecan, oxaliplatin),
which can be used in combination with or in place of 5-FU if
high TS or deficient DYPD are detected.

• Several SNPs in folate pathway enzymes are
associated with toxicity to MTX.

• MTHFR is a central regulatory enzyme in folate metabolism
that diverts folate metabolites from pyrimidine synthesis
towards methionine synthesis.
• A common functional polymorphism of the gene encoding this
enzyme occurs at C677T (alanine222valine).
• The 677TT genotype produces an enzyme with only 30% of the
activity of the wild-type (677CC) enzyme in vitro and is
associated with increased plasma concentrations of
homocysteine.
• A second common polymorphism in the MTHFR gene
(occuring at A1298C; Glut429Ala) also results in somewhat
decreased MTHFR activity.

• When MTX was given intravenously for the treatment of leukaemia,
the TT genotype was associated with increased incidence of
mucositis and slowed recovery of platelet counts.
• The wild‐type (CC) polymorphism in the C677T genotype is
associated with normal activity, whereas the CT heterozygote, found
in 40% of the population, is associated with a 40% decrease in the
ability to regenerate 5‐methyl‐tetrahydrofolate.
• The homozygotic TT genotype is found in 8–12% of the population
and is associated with a 70% decrease in the ability to regenerate
5‐methyl‐tetrahydrofolate

IRINOTECAN
• Irinotecan is a topoisomerase I inhibitor
• Irinotecan is a prodrug, which requires activation by
carboxylesterase to its active metabolite, SN-38.
• Hepatic UDP-glucuronosyl- transferase 1A1 (UGT1A1)
glucuronidates SN-38 to form the more polar and inactive SN-
38 glucuronide, which is eliminated in the bile and urine

• Diarrhea and leukopenia are the dose-limiting toxicities of
irinotecan, which are associated with increased levels of SN-
38 .
• The UGT1A1 promoter region contains a number of TA
repeats, with between five and eight repeats observed in the
general population.
• An inverse relationship exists between the number of repeats
and the expression of UGT1A1 .
• The wild-type allele UGT1A1*1 contains six TA repeats in the
promoter, leading to normal UGT1A1 expression and normal
toxicity risk.

• UGT1A1 expression is highly variable, and studies have shown
an interpatient variability in the rate of SN-38 glucuronidation
of up to 50-fold.
• The presence of seven repeats, instead of the wild-type
number of six, results in the variant allele UGT1A1*28.
• The UGT1A1*28 allele is associated with reduced UGT1A1
expression, and leads to reduced SN-38 glucuronidation .

• Two recent papers have shown that the UGT1A1*28 allele
leads to significantly increased amounts of the active
metabolite SN-38, and an increased chance of developing
diarrhea and leukopenia during irinotecan therapy .
• This study also showed that UGT1A1 promoter genotype is
significantly correlated with absolute neutrophil count at
nadir .
• These studies suggest that determination of the UGT1A1
genotypes may be clinically useful for predicting severe
toxicity to irinotecan.

PLATINUM AGENTS
• Platinum agents (cisplatin, carboplatin, oxaliplatin) inhibit
cellular replication by forming inter and intrastrand helix-
deforming DNA adducts.
• Resistance to platinum agents can occur through decreased
drug accumulation, detoxification through conjugation,
enhanced tolerance to platinum-induced DNA adducts, or
enhanced DNA repair
• Polymorphisms in DNA excision repair genes thus influence
the response to chemotherapy with platinum agents.

Polymorphisms in the XPD gene (official HUGO name ERCC2) :
• A nonsynonomous SNP, altering a lysine to glutamine at codon
751 of the XPD protein, was significantly associated with
treatment outcome.
• Patients with the lysine/lysine genotype had a median survival
of 17.4 months, compared to 12.8 months for
lysine/glutamine heterozygotes and 3.3 months for
glutamine/glutamine homozygotes

Polymorphisms in the XRCC1 protein :
• Polymorphism in XRCC1, a SNP that encodes either an
arginine or a glutamine at codon 399 of the protein, was
significantly associated with treatment response.
• Seventy-three percent of responders had the
arginine/arginine genotype, whereas 66% of nonresponders
had either the glutamine/glutamine or arginine/glutamine
genotypes ( P = 0.038).
• The functional consequence of this arginine to glutamine
change has been established-- the glutamine 399 form of the
XRCC1 protein results in a decreased DNA repair capacity.

Polymorphisms in glutathione (GSH)-dependent enzymes have
also been shown to influence response to platinum
chemotherapy agents.
• Glutathione-S-transferases (GSTs) catalyze the conjugation of
GSH to a wide variety of toxic compounds, including platinum
agents, forming less toxic and more water-soluble conjugates
that are exported out of target cells .
• There are five subclasses of the GST family (GSTA1, GSTP1,
GSTM1, GSTT1, and GSTZ1) that influence cytotoxicity to a
variety of chemotherapeutic agents

• One SNP in GSTP1 was associated with overall survival.
• This SNP results in the replacement of isoleucine with valine
at amino acid position 105 of the protein, which is known to
substantially diminish enzyme activity
• In this study, The valine homozygotes survived for a median of
24.9 months, the heterozygotes survived for a median of 13.3
months, and the isoleucine homozygotes survived for a
median of 7.9 months ( P < 0.001).
• Interestingly, GSTM1 and GSTT1 mutations that abolish
enzyme activity had no predictive power for patient outcome

TAMOXIFEN
• Tamoxifen is a selective estrogen-receptor modulator
• Endoxifen concentration α antiestrogen effects

• CYP2D6 is highly polymorphic, with more than 80 allelic
CYP2D6 variants described.
• These alleles vary in enzyme activity and prevalence with
respect to race and ethnicity
• Based on genotype :
1. Ultrarapid metabolizers (UM; approximately 1% to 2% of
patients) who carry more than two functional allele copies.
2. Extensive metabolizers (EM; 77% to 92% )
3. Intermediate metabolizers (IM; 2% to 11% )
4. Poor metabolizers (PM; 5% to 10%)

• Concentrations of endoxifen
EM >IM >PM patients
• Up to a sixfold variation in endoxifen levels may be seen
between homozygous PM and homozygous EM patients.

Trial results
• CYP2D6 PM phenotypes were associated with decreased DFS
• A statistically significant increased risk of disease recurrence
was seen in the IM and PM patients compared with the EM
patients
• Not all trial results have been consistent.
• Dosing guidelines for genotype-guided therapy do not yet
exist.
• IM patients who received an increased dose of 40 mg daily
instead of the standard 20 mg were shown to have endoxifen
concentrations similar to that of EM patients (p = 0.25).

• FDA advisory panel suggested that the tamoxifen package
insert should alert physicians of the following concerns:
1. CYP2D6 poor metabolizer patients are at increased risk for
recurrence of their breast cancers if treated with tamoxifen,
2. Co-administration of certain selective serotonin reuptake
inhibitors known to inhibit CYP2D6 can affect the
metabolism of tamoxifen
• For breast cancer prevention, raloxifene appears to be a viable
alternative for CYP2D6 poor metabolizer;
• Raloxifene and tamoxifen have been shown to be equally
effective in reducing breast cancer incidence in high-risk
postmenopausal women

Polymorphisms in Drug Transporters
MDR1 (P-glycoprotein, ABCB1) :
• P-glycoprotein (PGP), encoded by the MDR1 gene (ABCB1), is
the best-characterized ATP-binding cassette (ABC) transporter.
• PGP was initially found to be overexpressed in multidrug
resistant cancers.
• PGP is involved in the transport of a large range of
hydrophobic drugs, including cytotoxic chemotherapeutic
agents (e.g., doxorubicin), hormones, and carcinogens as well
as an array of structurally divergent drugs.
• In addition to its wide tissue distribution, PGP expression
varies markedly among individuals.

• Multiple MDR1 polymorphisms have been described to occur
in various allelic combinations. Grouping these
polymorphisms into haplotypes may serve as a useful
predictor of the functional consequences of MDR1
polymorphisms.
• The frequencies of variant MDR1 alleles differ, depending on
racial background, raising the possibility of racial differences
in chemotherapy pharmacokinetics and response.
• For example, the C3435T polymorphism (exon 26) has a
frequency of 73%–84% in individuals of African origin and
frequencies of 34%–59% in individuals of European and Asian
origin.
• A recent report also indicated that MDR1 polymorphisms may
play a role in determining the pharmacokinetic and clinical
toxicity profile of irinotecan

APPLYING
PHARMACOGENOMICS
---CHALLENGES

1. Frequency of mutations and cost-
effectiveness :
• While some mutations occur at a relatively high frequency (ie,
BCR-ABL in 95% of CML cases,C-KIT in 85% of
gastrointestinal stromal tumors, BRAF in 50% of metastatic
melanomas), others occur at very low frequencies (ie, ALK in
5% of NSCLCs, DPYD in 1% of the general population).
• The higher the NNT, the more difficult it is to prove cost-
effectiveness of testing .
• Single gene testing is likely to be substituted by multiplex
genomic characterization using next-generation sequencing
technology

2. Identifying an actionable target
• Distinguishing between functional “driver mutations” and
random, nonfunctional “passenger mutations”.
• Whether the mutation is silent or non-silent, the location or
frequency of the mutation, and the depth of sequencing
required to detect the mutation.
• Multi-pathway inhibition is likely necessary in a variety of
tumors prone to resistance or feedback upregulation.

3.Intra-tumoral heterogeneity
• Cancers are rarely homogenous and contain large areas with a
variable ratio of normal and cancerous cells.
Methods to overcome :
• deeper sequencing (of up to 1,000-fold), measuring the effect
of treatment at the biopsy site (ie, pretreatment and on-
treatment biopsy to assess target inhibition),
• determine the disease prognosis at multiple biopsy sites or
new lesions
• determine the biopsy sites that most likely account for disease
prognosis, profiling of circulating tumor cells (CTCs), or re-
biopsy the primary tumor at disease progression or at the
metastatic site.

Circulating free DNA
• CTCs and associated circulating free DNA (cfDNA) provide a
unique opportunity to identify driver mutations from
noninvasive blood specimens.
• Tumors shed CTCs and cfDNA into the bloodstream and it is
thought that these cells contribute to tumor metastases.
• cfDNA is typically easier to isolate and more abundant that
CTCs themselves, and they tend to represent all tumors in the
body and all cells in the tumor, helping to alleviate the issue of
tumor heterogeneity

4. Big data and integration into health
care systems
• Challenges to integrating genomics-based medicine into
health care systems include tumor and biologic heterogeneity,
limited access to novel targeted therapies, limited eligibility
criteria for enrollment of heterogenous populations in clinical
trials, and the paucity of evidence-based medicine to guide
• The Electronic Medical Records and Genomics (eMERGE)
Network was created to develop, disseminate, and apply
research that combines DNA biobanks with EMRs for large-
scale, high-throughput genetic research

5. Patient-focused perspectives
• The integration of pharmacogenomic testing into routine
clinical practice will depend not only on clinician acceptance
but also on patient adoption and understanding of the risks
and benefits.

Pharmacogenomics mine

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