Pharmacogenetics - PGx

Dx
Screen
Monitor
Background
Lab Tests

Diagnosis

Indications for testing

Provide pretherapeutic guidance for drug and dose selection
Provide post-therapeutic monitoring
- Evaluate the cause of post-therapeutic adverse drug reactions or therapeutic failure
- Optimize dose with pharmacokinetic or pharmacodynamic monitoring
Drug-gene associations (also see FDA-approved drugs with pharmacogenomic information in their labels)

Laboratory testing

Germ-line genetic tests (whole blood)
- 5-Fluorouracil and DPYD, TYMS
- Antidepressants and CYP2D6, CYP2C19
- Clopidogrel and CYP2C19
  - American Heart Association and American College of Cardiology recommend testing for patients at moderate to high risk of cardiovascular events who are treated with clopidogrel
- Codeine and CYP2D6
- Irinotecan and UGT1A1
- Tamoxifen and CYP2D6
  - Most important therapeutic agent for treatment of estrogen receptor (ER)-positive breast cancer for the past three decades
  - Metabolites are ~100-fold stronger than parent compound
  - CYP2D6 is responsible for activity
- Warfarin and CYP2C9, VKORC1
  - Narrow therapeutic index (causal agent for 10% of all adverse drug reactions)
  - Recommended by FDA
- Interferon-α and ILB-28
  - Inosine triphosphatase (ITPA)
- HCV therapy and interleukin 28 B (IL28B), inosine triphosphatase (ITPA)
- Abacavir and HLA-B*5701
  - 5-8% of Caucasians develop hypersensitivity reactions within 6 weeks of therapy
  - FDA considers test mandatory prior to therapy
- Carbamazepine or lamotrigine and HLA-B*1502
- Thiopurines and thiopurine methyltransferase (TPMT)
  - Inactivates toxic metabolites of thiopurine prodrugs
  - Heterozygous individuals (7-15% of Caucasians) have intermediate TPMT activity
  - 1/300 individuals are homozygous with low to negligible detoxification activity
  - Genetic and phenotype testing is available
Monoclonal antibody therapy and molecular testing
- Cetuximab or panitumumab and KRAS/NRAS
- Trastuzumab and HER2/neu
- Tyrosine kinase inhibitors and EGFR testing
Phenotype testing
- Azathioprine/6-mercaptopurine and TPMT activity
- Succinylcholine and pseudocholinesterase activity
Pharmacokinetic testing
- Therapeutic drug monitoring (TDM) is available for many drugs and drug metabolites and can be used to assess phenotype of a patient post-therapeutically, as well as to optimize dose
  - Examples include tricyclic antidepressants and codeine

Screening

Pretherapeutic drug selection/avoidance
- Predict risk of toxicity and likelihood of response that is dose-independent
Pretherapeutic dose selection
- Predict risk of toxicity and likelihood of response that is dose-dependent
- Select dose (eg, dose-escalate a patient that is predicted to have poor response or reduce dose for a patient that is predicted to be very sensitive to a drug)
- Predict pharmacokinetics of a drug in order to optimize dosing frequency as well as determine best time to evaluate response to therapy (ie, estimate time to achieve steady state)
Familial testing may be appropriate
- Testing options should be discussed with laboratory or genetic counselor

Monitoring

Post-therapeutic evaluation of adverse drug reactions or failure to respond
- Dependent upon the following
  - Clinical factors
  - Clinical scenario (eg, whether a reaction is likely to be related to the drug and/or dose administered)
  - Compliance
  - Drug
  - Drug formulation

Clinical Background

Genetic variations associated with drug response or drug disposition may predispose a patient to risk of drug-related toxicity or lack of therapeutic benefit and are referred to as pharmacogenetic variants. Pharmacogenetics can explain and predict variations in both pharmacokinetic and pharmacodynamic processes.

As such, pretherapeutic pharmacogenetic testing to identify people who have inherited clinically significant variants may guide drug and dose selection to promote personalized therapeutics. Pharmacogenetic testing may be designed to detect human germ-line variants, somatic variants (eg, tumor tissue), or genomic variants of an infectious organism (eg, virus). The goals of pharmacogenetic testing are to reduce the high number of nonresponders (averaging 30-60% of patients) and to reduce adverse drug reactions.

Pharmacogenetics Definitions

Definitions of common pharmacogenetics terms

Definitions of Pharmacogenetics Terms
Allele	Alternate form of a gene located on a specific chromosome
Chromosome	Linear bodies in the cell nucleus containing most or all genes of the organism
Deletion	Absence of a section of genetic material from a gene or absence of one or more entire genes from a chromosome
Duplication	Part of a chromosome in which the genetic material, including an entire gene or genes, is repeated; process of forming a duplication
Expression	Detectable effect of a gene, usually manifested by the amount and/or type of protein
Gene	Functional unit of heredity occupying a specific locus on a chromosome; capable of reproducing itself exactly at each cell division; directs formation of an enzyme or other protein
Genotype	Genetic constitution of an individual gene; may reflect a single nucleotide polymorphism, mutation, or series of variants
Haplotype	Group of genetic variants from one or more genes (eg, of the major histocompatibility complex) located on a single chromosome that are closely enough linked to usually be inherited as a unit or characteristic pattern
Heterozygote (HET)	Two genes at corresponding loci on homologous chromosomes different for one or more loci; two different copies
Homozygote	Two genes at corresponding loci on homologous chromosomes identical for one or more loci; two identical copies
Insertion	Section of genetic material inserted into an existing gene sequence
Linkage	Relationship between genes on the same chromosome that causes them to be inherited together
Metabolizers	Poor metabolizer – lacking or near total lacking capacity to metabolize a substrate through a specific pathway
	Intermediate metabolizer – less than normal capacity to metabolize a substrate through a specific pathway
	Ultrarapid metabolizer – enhanced capacity to metabolize a substrate through a specific pathway
	Extensive metabolizer – normal population-based capacity to metabolize a substrate through a specific pathway
Mutant (MUT)	Change in hereditary material involving either a physical change in chromosome relations or a biochemical change in the codons that make up genes that are associated with a phenotype
Phenotype	Observable properties of an organism that are produced by the interaction of the genotype and the environment
Single nucleotide polymorphism	Naturally occurring substitution of a single nucleotide at a given location in genome of an organism, often resulting in phenotypic variability
Variant	Exhibiting variation or diversity, either genotypically or phenotypically
Wild-type	Phenotype, genotype, or gene that predominates in a natural population of organisms or strain of organisms in contrast to that of mutant forms

Pathophysiology

Strengths and limitations of pharmacogenetic results are based on the following
- Actual result (heterozygote vs mutant)
- Allele frequency
- Assay content
- Clinically accepted guidelines
- Drug choices available
- Genotype-phenotype relationship
- Methodology
- Other factors that impact phenotype
  - Comedications
  - Clinical status
  - Age/sex
  - Alternate metabolic pathways
  - Other genes

Clinical Presentation

Pharmacogenetic variations

Adverse drug reactions with pharmacogenetic associations

Examples of Adverse Drug Reactions With Pharmacogenetic Associations
Drug	Pharmacogenetic variation(s)	Effect
Warfarin	CYP2C9, VKORC1	Bleeding
Azathioprine/6-Mercaptopurine/6-Thioguanine	Thiopurine methyltransferase (TPMT)	Neutropenia due to increased myelosuppression
Irinotecan	UGT1A1	Severe diarrhea or neutropenia due to decreased metabolism
Abacavir	HLA-B*5701	Hypersensitivity reaction (rash, fever, gastrointestinal symptoms) in Caucasian populations
Carbamazepine	HLA-B*1502	Hypersensitivity reaction (rash, fever, gastrointestinal symptoms) in Asian populations
Codeine, antidepressants, several other drugs	CYP2D6	Drug-specific
Ribavirin	Inosine triphosphatase (ITPA)	Hemolytic anemia

Therapeutic failures

Examples of Therapeutic Failures
Drug	Pharmacogenetic variation(s)	Effect
Antiestrogens (tamoxifen), alpha-blockers, analgesics, anticonvulsives, antidepressants (amitriptyline), antidiabetics, antihypertensives, antipsychotics, beta blockers, norepinephrine reuptake inhibitors	CYP2D6	Drug specific
Warfarin	CYP2C9, VKORC1	Bleeding/thrombosis
Clopidogrel	CYP2C19	New arterial blood clots
Interferon	IL28B	Disease progression
Cetuximab panitumumab	KRAS/NRAS	Lack of therapeutic benefit Prescribed only in nonmutated cases
Tyrosine kinase inhibitors	BCR/ABL, c-Kit	Presence necessary for therapeutic effect Characteristic mutations imply resistance to some tyrosine kinase inhibitors
Trastuzumab	HER2	Presence of over expression necessary for therapeutic effort

Indications for Laboratory Testing

Tests generally appear in the order most useful for common clinical situations
Click on number for test-specific information in the ARUP Laboratory Test Directory

Test Name and Number	Recommended Use	Limitations	Follow Up
5-Fluorouracil (5-FU) Toxicity and Chemotherapeutic Response, 7 Mutations 2007228 Method: Polymerase Chain Reaction/Single Nucleotide Extensions/Fragment Analysis	Use to detect genetic variants that influence metabolism of 5-FU Only the targeted DPYD and TYMS mutations will be evaluated	Genotype should be interpreted with clinical information; consultation with clinical pharmacist recommended Rare diagnostic errors can occur due to primer-site mutations
HLA-B*5701 Associated Variant Genotyping for Abacavir Sensitivity 2002429 Method: Polymerase Chain Reaction/Fluorescence Monitoring	Standard of care prior to abacavir therapy per FDA Screening test to determine susceptibility to abacavir hypersensitivity syndrome	Routinely used in many countries before introducing patient to abacavir Less predictive of hypersensitivity within black population Frequency of HLA-B*5701 is very low in Asian populations, limiting value of test in this population 80-fold increase in liver injury risk for patients on flucloxacillin
Cytochrome P450 2C9 (CYP2C9) 2 Mutations 0051103 Method: Polymerase Chain Reaction/DNA Hybridization/Electrochemical Detection	Aids in warfarin dosage planning in conjunction with VKORC1 testing Most useful for Warfarin-naïve individuals Individuals with personal or family history of difficulty with anticoagulation Individuals who are therapy adherent but difficult to treat Individuals who might be placed on warfarin prophylactically after surgery (eg, those requiring <21 mg or >49 mg per week to maintain therapeutic INR) Other uses Dosage planning for any drug that is metabolized by CYP2C9 (eg, Dilantin)	Does not provide individualized dosing recommendations Additional mutations in this or other genes will not be detected Mutation detection is not a substitute for therapeutic drug or other clinical monitoring Nongenetic factors may also affect drug metabolism	Refer to Warfarin Genotyping Plus test if individualized warfarin dosing recommendations are needed
Warfarin Genotyping Plus 2004358 Method: Polymerase Chain Reaction/DNA Hybridization/Electrochemical Detection	This is the preferred test for warfarin dosage planning Warfarin Sensitivity (CYP2C9 and VKORC1) 3 Mutations test is performed and individualized dosing recommendations are provided when specific patient parameters are submitted Clinical utility has not been established
Warfarin Sensitivity (CYP2C9 and VKORC1 ) 3 Mutations 0051370 Method: Polymerase Chain Reaction/DNA Hybridization/Electrochemical Detection	Aids in warfarin dosage planning Most useful for Warfarin-naïve individuals Individuals with personal or family history of difficulty with anticoagulation Individuals who are therapy adherent but difficult to treat Individuals who might be placed on warfarin prophylactically after surgery (eg, those requiring <21 mg or >49 mg per week to maintain therapeutic INR) Does not provide individualized dosing recommendations Clinical sensitivity – 90% of CYP2C9 and VKORC1 mutations causing warfarin sensitivity in Caucasians are detected; the detection rate in other ethnicities is reduced	Other mutations that may impact warfarin sensitivity or resistance in these genes or other genes will not be detected Rare mutations within primer or probe regions may interfere with the assay	Refer to Warfarin Genotyping Plus test if individualized warfarin dosing recommendations are needed
Cytochrome P450 2D6 (CYP2D6) 14 Mutations and Gene Duplication 0051232 Method: Polymerase Chain Reaction/Primer Extension	Order to detect CYP2D6 allelic variants associated with impaired or increased drug metabolism May aid in dose planning for drugs such as Antiestrogens (tamoxifen) Antidepressants (amitriptyline) Analgesics (codeine, oxycodone) Antipsychotics Alpha-blockers Anticonvulsives Antidiabetics Antihypertensives Beta blockers Norepinephrine reuptake inhibitors	Only the targeted CYP2D6 mutations will be detected; mutations in other genes will not be detected Additional mutations in this or other genes will not be detected Mutation detection is not a substitute for therapeutic drug or other clinical monitoring Nongenetic factors may also affect drug metabolism
Cytochrome P450 2C19 (CYP2C19) 9 Mutations 0051104 Method: Polymerase Chain Reaction/Primer Extension	Order to detect CYP2C19 allelic variants associated with impaired (2, 3) or increased (*17) drug metabolism May aid in dose planning for drugs such as Clopidogrel Diazepam Some antidepressants (eg, citalopram, amitriptyline, clomipramine) Proton pump inhibitors (eg, omeprazole, lansoprazole) Antimalarials (eg, chloroguanide) Mephenytoin	Additional mutations in this or other genes will not be detected Mutation detection is not a substitute for therapeutic drug or other clinical monitoring Nongenetic factors may also affect drug metabolism
KRAS Mutation Detection 0040248 Method: Polymerase Chain Reaction/Pyrosequencing	Order to assess resistance to anti-EGFR therapy (cetuximab and panitumumab)	40% of wild-type, KRAS-type tumors will not respond to anti-EGFR monoclonal antibodies
KRAS Mutation Detection with Reflex to BRAF Codon 600 Mutation Detection 2001932 Method: Polymerase Chain Reaction/Pyrosequencing	Order to assess response to EGFR-directed therapy Detects mutations in codons 12, 13, and 61; if KRAS is not detected, BRAF codon 600 Mutation Detection will be performed
NRAS Mutation Detection by Pyrosequencing 2003123 Method: Polymerase Chain Reaction/Pyrosequencing	Order to predict response to EGFR-directed therapy
UDP Glucuronosyltransferase 1A1 (UGT1A1) Genotyping 0051332 Method: Polymerase Chain Reaction/Fragment Analysis	May be useful for dose planning in individuals who Will receive high-dose irinotecan (doses >150 mg/m²) Have a personal or family history of irinotecan sensitivity Experience neutropenia while receiving irinotecan May be used to diagnose suspected Gilbert syndrome (benign familial hyperbilirubinemia)	No specific dosing recommendations are available for patients with decreased UGTIAI activity Clinical significance of the rare (TA)5 and (TA)8 alleles in predicting irinotecan toxicities is not well established; genetic and nongenetic factors other than UGT1A1 may contribute to irinotecan toxicity and efficacy
Pseudocholinesterase, Dibucaine Inhibition 0020159 Method: Quantitative Enzymatic	Order to detect increased sensitivity in individuals who experience prolonged paralysis following succinylcholine or mivacurium administration
Mercaptopurine Quantitation, Serum or Plasma 0091084 Method: Quantitative High Performance Liquid Chromatography-Tandem Mass Spectrometry	Compliance monitoring
Thiopurine Methyltransferase, RBC 0092066 Method: Enzymatic/Quantitative Liquid Chromatography-Tandem Mass Spectrometry	Preferred screening test to detect individuals at excess risk for myelosuppression when exposed to standard dose of thiopurines May aid in dose planning for individuals who will receive thiopurine drugs Intended for pretherapeutic evaluation only	Do not use for patients currently on thiopurine (will give falsely low results) or who have had a blood transfusion in the last 30 days
EGFR Mutation Detection by Pyrosequencing 2002440 Method: Polymerase Chain Reaction/Pyrosequencing	Order to assess response to gefitinib therapy for non-small cell lung cancer Detects mutations in EGFR exons 18, 19, 20, and 21
TPMT Genotype 2002573 Method: Qualitative Polymerase Chain Reaction	Aids in detecting individuals at high risk for myelosuppression when exposed to standard dose of thiopurines Consider if red blood cell (RBC) TPMT testing is not practical, as in the following Current treatment with thiopurine drugs (results will be falsely low) Blood transfusion within the last 30-60 days Treatment within the last 48 hours with naproxen, ibuprofen, ketoprofen, furosemide, sulfasalazine, mesalamine, olsalazine, mefenamic acid, thiazide diuretics, and benzoic acid inhibitors	TPMT polymorphism explains only 60% of the thiopurine hematotoxicity but none of the thiopurine induced liver injury Continued monitoring on hematologic and liver parameters is necessary
Thiopurine Metabolites 2002575 Method: Qualitative High Pressure Liquid Chromatography (High Performance Liquid Chromatography)	Aids in therapeutic monitoring in individuals treated with thiopurines
Interleukin 28 B (IL28B)-Associated Variants, 2 SNPs 2004680 Method: Qualitative Polymerase Chain Reaction/Qualitative Fluorescence Monitoring	Detects genetic variants associated with interleukin 28 B (IL28B) that predict Probability of treatment response to peginterferon alfa and ribavirin, as well as to protease inhibitor triple therapy Treatment duration	SNPs other than those targeted will not be detected
Inosine Triphosphatase (ITPA) and Interleukin 28 B (IL28B)-Associated Variants, 4 SNPs 2006344 Method: Polymerase Chain Reaction/Single Nucleotide Extensions	Detect genetic variants associated with interleukin 28 B (IL28B) that may aid in Predicting peginterferon alfa (PEG-INF)/ribavirin (RBV) treatment response Predicting triple therapy (PEG-INF/RBV/protease inhibitor) treatment response for HCV genotype 1 Protease inhibitors – boceprevir, telaprevir Detect genetic variants associated with inosine triphosphatase (ITPA) that may aid in Management of patients during any therapies containing RBV, particularly for patients at increased risk of anemia	SNPs other than those targeted will not be detected

Additional Tests Available

Click the plus sign to expand the table of additional tests.

Test Name and Number	Comments
Warfarin, Urine 0091230 Method: High Performance Liquid Chromatography
Warfarin, Serum or Plasma 0090805 Method: High Performance Liquid Chromatography
Pseudocholinesterase, Total 0020167 Method: Quantitative Enzymatic