1. Pharmacogenomics: The Promise of Personalized Medicine
Christina Aquilante, Pharm.D.
Assistant Professor
Department of Pharmaceutical Sciences
School of Pharmacy
University of Colorado at Denver and Health Sciences Center

2. Objectives
Provide an overview of pharmacogenomics and its clinical relevance
Discuss clinically-relevant examples of:
Drug metabolism pharmacogenomics
Drug target pharmacogenomics
Discuss the challenges facing pharmacogenomic studies and the movement of pharmacogenomics into clinical practice
Discuss pharmacogenomics from the FDA and pharmaceutical industry perspective

3. Interindividual Variability in Drug Response
Disease Drug Class Rate of Poor Response
Asthma Beta-agonists %
Hypertension Various 30%
Solid Cancers Various 70%
Depression SSRIs, tricyclics %
Diabetes Sulfonylureas, others 50%
Arthritis NSAIDs, COX-2 inhibitors 30-60%
Schizophrenia Various %

4. Factors Contributing to Interindividual Variability in Drug Disposition and Action
Age
Race/ethnicity
Weight
Gender
Concomitant Diseases
Concomitant Drugs
Social factors
GENETICS
PERSONALIZED
MEDICINE

5. “We wish to suggest a structure for the salt of [DNA].
This structure has novel features which are of considerable biological interest.”

6. Human Genome Project
Determine the sequence of the 3 billion nucleotides that make up human DNA
Characterize variability in the genome
Identify all the genes in human DNA
The Era of Genomic Medicine:
Improve prediction of drug efficacy or toxicity
Improve the diagnosis of disease
Earlier detection of genetic predisposition to disease

7. Newsweek June 25, 2001 “…pharmacogenetics promises to target
treatment to a patient’s
genetic profile…”
Quote
Mary Relling, Bill St. Jude’s hospital in Memphis (and Mayo)
Herceptin—humanized monoclonal antibody, Genentech, FDA-approved in 1998
Metastatic CA where tumors overexpress the HER-2 protein (only found in 30% of breast cancers)
US News and World Report

8. Genetics or Genomics? Pharmacogenetics Pharmacogenomics
Study of how genetic differences in a SINGLE gene influence variability in drug response (i.e., efficacy and toxicity)
Pharmacogenomics
Study of how genetic (genome) differences in MULTIPLE genes influence variability in drug response (i.e., efficacy and toxicity)

9. Current Concept of Pharmacogenomics
Roden DM et al. Ann Intern Med 2006; 145:749-57

10. Potential of Pharmacogenomics
All patients with same diagnosis 1 2
Responders and patients
not predisposed to toxicity
Non-responders
and toxic
responders
Treat with alternative
drug or dose
Treat with
conventional

11. Clinical Relevance Can we predict who will derive an optimal response?
Can we predict who will have a toxicity?
Host (patient) genotype determines optimal drug therapy approach
Disease (pathogen) genotype determines optimal drug therapy approach

12. DNA is Information DNA A, T, G, C Codon Gene Chromosome Genome ENGLISH
Abcdefg….xyz
Word
Sentence
Chapter
Book

13. Composition of the Human Genome
Mutation/Polymorphism 1 bp
Unit of genetic code 3 bp
Coding sequence (exons) 3,000 bp
Gene (exons and introns) 50,000 bp
Chromosome ,000,000 bp
Human genome 3,000,000,000 bp

14. The Foundation of Pharmacogenomics: Differences in the Genetic Code Between People
Mutation: difference in the DNA code that occurs in less than 1% of population
Often associated with rare diseases
Cystic fibrosis, sickle cell anemia, Huntington’s disease
Polymorphism: difference in the DNA code that occurs in more than 1% of the population
A single polymorphism is less likely to be the main cause of a disease
Polymorphisms often have no visible clinical impact

15. Single Nucleotide Polymorphisms (SNP)
Pronounced “snip”
Single base pair difference in the DNA sequence
Over 2 million SNPs in the human genome
Other polymorphisms:
Insertion/deletion polymorphisms
Gene duplications
Gene deletions
1.4 million
Promoter, coding, non-coding, 3’untranslated region

16. SNPs are present in every 1000 to 3000 base pairs throughout the body.
Can occur in exons, introns, and 3’ untranslated regions

17. Consequences of Polymorphisms
May result in a different amino acid or stop codon
May result in a change in protein function or quantity
May alter stability of mRNA
No consequence

18. Genetics Terminology Alleles = different DNA sequences at a locus
Codon 389 1-AR
Arg (0.75)
Gly (0.25)
Genotype = pair of alleles a person has at a region of the chromosome
Codon 389 1-AR
Arg389Arg
Arg389Gly
Gly389Gly

19. Genetics Terminology Phenotype: outward manifestation of a trait
Linkage: measure of proximity of 2 or more polymorphisms on a single chromosome
Polymorphisms in close proximity tend to be co-inherited
Regions of linked polymorphisms are known as haplotypes

20. Haplotype Map
For specific locations in the genome, a small number of SNP patterns (haplotypes) can account for 80-90% of entire human population
International HapMap Project:
Identifying common haplotypes in four populations from different parts of the world
Identifying “tag” SNPs that uniquely identify these haplotypes

21. Pharmacogenomics DRUG TARGETS DRUG TRANSPORTERS DRUG METABOLIZING
ENZYMES
PHARMACODYNAMICS
PHARMACOKINETICS
Variability in
Efficacy/Toxicity
Johnson JA. Trends in Genetics 2003:

22. Drug Metabolism Pharmacogenomics
Evidence of an inherited basis for drug response dates back in the literature to the 1950s
Succinylcholine: 1 in 3000 patients developed prolonged muscle relaxation
Monogenic
Phenotype to genotype approach

23. Drug Metabolizing Enzymes
Metabolism can take place in many organs, but the liver frequently has the greatest metabolic capacity. The liver is the major site of drug metabolism.
Most drugs undergo chemical modifications in the body so that they can eventually be eliminated.
Very generally, these chemical reactions help to increase the water solubility of the drug so that it can be eliminated from the body, in most cases through the kidneys or the bile.
Phase I: biotransformation reactions: oxidation, hydroxylation, reduction, hydrolysis—a polar functional group is added onto the substrate drug
Phase II: conjugation reactions—add small molecules onto drugs to increase their water solubility and elimination from the body. The major reactions are glucuronidation, sulation, acetylation, glutathione conjugation

24. Examples of Drug Metabolism Pharmacogenomics
NEJM 2003; 348:

25. Examples of Drug Metabolism Pharmacogenomics
NEJM 2003; 348:

26. Warfarin and CYP2C9
Widely prescribed anticoagulant drug used to prevent blood clots
Narrow range between efficacy and toxicity
Large variability in the dose required to achieve therapeutic anticoagulation
Doses vary 10-fold between people
CYP2C9 is the enzyme responsible for the metabolism of warfarin
SNPs exist in CYP2C9 gene that decrease the activity of the CYP2C9 metabolizing enzyme

27. CYP2C9 Polymorphisms and Warfarin Dose
Warfarin dose is affected by CYP2C9 genotype
*2 and *3 are SNPs
Gage BF et al. Thromb Haemost 2004; 91: 87-94

28. CYP2C9 Genotype and Bleeding Events
Compared to wild-type, CYP2C9 variants had a higher risk of serious or life-threatening bleeds
Hazard Ratio of 3.94 during the first 3 months of follow-up
Hazard Ratio of 2.39 for the entire follow-up period WT
Variant
Higashi et al. JAMA 2002; 287

29. Challenges Facing Warfarin Pharmacogenomics
Despite the strong association between CYP2C9 genotype and warfarin dose, CYP2C9 genotype accounts for only a small portion of the total variability in warfarin doses (~10-20%)
Need to determine other genetic and non-genetic factors that contribute to interindividual variability in warfarin doses

30. CYP2D6 Polymorphisms
CYP2D6 is responsible for the metabolism of a number of different drugs
Antidepressants, antipsychotics, analgesics, cardiovascular drugs
Over 100 polymorphisms in CYP2D6 have been identified
Based on these polymorphisms, patients are phenotypically classified as:
Ultrarapid metabolizers (UMs)
Extensive metabolizers (EMs)
Poor metabolizers (PMs)

31. CYP2D6 Phenotypes NEJM 2003; 348:529
Roden DM et al. Ann Intern Med 2006; 145:749-57

32. CYP2D6 Polymorphisms and Psychiatric Drug Response
Increased rate of adverse effects in poor metabolizers due to increased plasma concentrations of drug:
Fluoxetine (Prozac) death in child attributed to CYP2D6 poor metabolizer genotype
Side effects of antipsychotic drugs occur more frequently in CYP2D6 poor metabolizers
CYP2D6 poor metabolizers with severe mental illness had more adverse drug reactions, increased cost of care, and longer hospital stays

33. CYP2D6 and Codeine
Codeine requires activation by CYP2D6 in order to exert its analgesic effect
Due to genetic polymorphisms, 2-10% of the population cannot metabolize codeine and are resistant to the analgesic effects
Interindividual variability exists in the adequacy of pain relief when uniform doses of codeine are given

34. Strattera® (Atomoxetine)
Treatment of attention deficit hyperactivity disorder
CYP2D6 poor metabolizers have 10-fold higher plasma concentrations to a given dose of STRATTERA compared with extensive metabolizers
Approximately 7% of Caucasians are poor metabolizers
Higher blood levels in poor metabolizers may lead to a higher rate of some adverse effects of STRATTERA

35. CYP2C19 and Proton Pump Inhibitors
Proton pump inhibitors are used to treat acid reflux and stomach ulcers
Ulcer cure rates using omeprazole and amoxicillin by CYP2C19 phenotype:
Cure Rate
Rapid metabolizers 28.6%
Intermediate metabolizers 60%
Poor metabolizers 100%
Furuta, T. et. al. Ann Intern Med 1998;129:

36. Roche AmpliChip: FDA-Approved

37. Roche AmpliChip P450 Test
The Roche AmpliChip CYP450 Test is intended to identify a patient's CYP2D6 and CYP2C19 genotype from genomic DNA extracted from a whole blood sample.
Information about CYP2D6 and CYP2C19 genotype may be used as an aid to clinicians in determining therapeutic strategy and treatment dose for therapeutics that are metabolized by the CYP2D6 or CYP2C19 gene product.

38. Thiopurine-S-Methyltransferase (TPMT)
Thiopurine drugs are used to treat cancer
Acute lymphoblastic leukemia
TPMT is important for metabolizing thiopurines
azathioprine, mercaptopurine (6-MP)
Polymorphisms in the TPMT gene result in decreased TPMT enzyme activity
Decreased TPMT activity predisposes individuals to severe, life-threatening toxicities from these drugs

39. Variability in TPMT Activity

40. Genotype-Guided 6-MP Dosing
Childhood ALL
3 non-synonymous SNPs account for over 90% of the clinically relevant TPMT mutations and result in a trimodal distribution of TPMT activity
Pharmacogenomics 2002;3(1):89-98.

41. 6-Mercaptopurine Prescribing Information
There are individuals with an inherited deficiency
of the enzyme thiopurine methyltransferase
(TPMT) who may be unusually sensitive to the
myelosuppressive effects of mercaptopurine and
prone to developing rapid bone marrow
suppression following the initiation of treatment.
Substantial dosage reductions may be required to
avoid the development of life-threatening bone
marrow suppression in these patients.

42. Imuran Prescribing Information
TPMT genotyping or phenotyping can be used to identify patients with absent or reduced TPMT activity.
Patients with low or absent TPMT activity are at an increased risk of developing severe, life-threatening myelotoxicity from IMURAN if conventional doses are given.
Physicians may consider alternative therapies for patients who have low or absent TPMT activity (homozygous for non-functional alleles). IMURAN should be administered with caution to patients having one non-functional allele (heterozygous) who are at risk for reduced TPMT activity that may lead to toxicity if conventional doses are given. Dosage reduction is recommended in patients with reduced TPMT activity.

43. TPMT and Thioguanines Clinical implications:
Genetic testing for TPMT is routine practice at some cancer centers for protocols involving thiopurine drugs
CLIA approved test available
Implications for cancer, transplant, rheumatoid arthritis, lupus, dermatology, and Crohn’s disease treatment

44. Drug Target Pharmacogenomics

45. Drug Target Pharmacogenomics
Direct protein target of drug
Receptor
Enzyme
Proteins involved in pharmacologic response
Signal transduction proteins or downstream proteins
Polymorphisms associated with
disease risk
“Disease-modifying” polymorphisms
“Treatment-modifying” polymorphisms
POLYGENIC

46. Complexity of Drug Effect

47. Assessing Phenotype in Drug Target Pharmacogenomics
Depression—Symptom rating scales
Indirect measure of drug response
Inter-rater reliability
Hypertension—Blood pressure
Minute to minute and diurnal variability
Influence of environmental factors (e.g. lack of rest before measurement)
Diabetes—Blood glucose
Diurnal variation in blood glucose
Influence of environmental factors (e.g. diet/exercise)

48. Comparison Drug Metabolism Pgx
Polymorphisms often lead to non-functional or absent proteins
Distinct phenotypes
Bimodal/trimodal distribution
Phenotypes are easily measured
Drug concentration
In vitro catalytic activity
Drug Target Pgx
Polymorphisms usually don’t result in lack of protein function
“Subtle” effects
Differences in phenotypes are smaller
Measurement of phenotypes is difficult
Imprecise and variable
Failure to consider haplotypes
Johnson JA and Lima JJ. Pharmacogenetics 2003; 13:

49. Examples of Drug Target Pharmacogenomics
Evans WE. NEJM 2003; 348:538-48

50. Examples of Disease or Treatment Modifying Pharmacogenomics
Evans WE. NEJM 2003; 348:538-48

51. Beta-blockers and Hypertension (HTN)
HTN is the most prevalent chronic disease in the US and a contributor to morbidity and mortality
Beta-blockers are first-line agent in the treatment of HTN
Marked variability in response to beta-blockers
30-60% of patients fail to achieve adequate blood pressure lowering with beta-blockers
Common beta-blockers used in HTN:
Metoprolol
Atenolol

52. Beta-1 Adrenergic Receptor
Codon 49 SerGly
Codon 389
ArgGly
Podlowski, et al. J Mol Med 2000;78:90.

53. Beta-1 Receptor Polymorphisms and Response to Metoprolol
Arg homozygotes had a 2-fold greater reduction in 24h DBP than Gly carriers.
This was largely driven by differences in daytime DBP. Arg homozygotes had a 3-fold greater reduction in daytime DBP than Gly carriers
Johnson JA et al. Clin Pharmacol Ther 2003; 74:44-52

54. Beta-2 Adrenergic Receptor Polymorphisms and Response to Albuterol in Asthma
Hyperreactivity of the airways is the hallmark of asthma
Airway smooth muscle contains beta-2 receptors that produce broncodilation
Albuterol is a beta-2 agonist that is used in the treatment of asthma
Produces smooth muscle cell relaxation and bronchodilation
Forced expiratory volume in 1 second (FEV1)
Phenotypic measure of response

55. Beta-2 Polymorphisms and Response to Albuterol
Single 8 mg albuterol dose
Albuterol-evoked increases in FEV1 were higher and more rapid in Arg16 homozyotes compared with Gly carriers
Codon 16 polymorphism is a determinant of bronchodilator response to albuterol
Lima JJ et al. Clin Pharmacol Ther 1999; 65:
Lima JJ. Clin Pharmacol Ther 1999; 65:519-25

56. VKOR and Warfarin
Warfarin works by inhibiting Vitamin K Epoxide Reductase (VKOR)
VKOR helps recycle vitamin K which is important in proper functioning of clotting factors
By inhibiting VKOR, warfarin alters the vitamin K cycle and results in the production of inactive clotting factors
Polymorphisms exist in the gene for VKOR (VKORC1)

57. VKORC1 Genotype and Warfarin Dose Requirements
46 mg/wk
33 mg/wk
21 mg/wk
G/G G/A A/A

58. Warfarin Pharmacogenomics
CYP2C9 SNPs account for a small amount of variability in warfarin doses (~10%)
VKORC1 SNPs explain a larger portion of variability in warfarin doses (~20-25%)
Almost 50% of variability in warfarin doses can be explained by a combination of factors:
VKORC1 SNPs
CYP2C9 SNPs
Non-genetic factors (age, weight, concomitant drugs, concomitant disease states)

59. Warfarin Pharmacogenomics
Current Status:
Develop and validate dosing algorithms to that include VKORC1 genetic information, CYP2C9 genetic information, and non-genetic factors (e.g., age, weight, concomitant drugs, concomitant disease states)
Test if dosing warfarin based on genotype is better than the “usual” care approach

60. Abacavir Hypersensitivity
Antiretroviral used for treatment of HIV
5% of patients experience hypersensitivity reactions to the drug
Hypersensitivity is fatal in rare cases
Hypersensitivity reaction starts with severe GI symptoms, followed by fever and rash
Discontinuation of drug reverses symptoms
Re-challenge of abacavir in hypersensitive individuals can result in life-threatening low blood pressure or death
Lancet 2002;359:

61. Abacavir Hypersensitivity
Hypersensitivity typically believed to be an immunologic reaction
Hypersensitivity might be genetically linked, and thus predictable
Major histocompatibility proteins (MHC) investigated because of known links in other immune responses and allergic reactions

62. Genetics of Abacavir Hypersensitivity
Western HIV Australia HIV Cohort Study
Patients with the HLA-B*5701 variant were 117 times more likely to be hypersensitive to abacavir than those who did not have the variant
13 patients had 3 linked genetic variants (*5701, DR7, DQ3) and all patients were abacavir hypersensitive
All abacavir hypersensitive patients were Caucasian, therefore
studies in other racial groups are needed

63. Disease Risk Polymorphisms
Polymorphisms can predispose individuals to a disease or increase the risk for disease
If a drug with a known adverse effect is given to a person with a genetic susceptibility to that adverse effect, there is an increased likelihood for that adverse effect

64. Clotting Factor Polymorphisms, Blood Clots, and Oral Contraceptive Pills
Polymorphisms exist in clotting factor genes
Oral contraceptive pills alone are associated with an increased risk of blood clots
Women who have clotting factor polymorphisms are at an even greater risk for blood clots if they receive oral contraceptive pills

65. Oral Contraceptive Pills and Blood Clots
Heterozygotes
Patients on OCP who are homozygous for Factor V Leiden have
50 to100-fold increased risk of VTE
Martinelli I. Pharmacogenetics 2003; 13:

66. A Different Aspect of Drug Target Pharmacogenomics
In all of the examples thus far, the drug target has been a human protein
The gene encoding that human protein has generally NOT undergone mutation during the patient’s life
However, in the areas of infectious diseases and cancer, the drug target is often a non-human protein
Cancer: Tumor DNA
Infectious Diseases: Viral genotype
e.g., HIV, HBV, HCV

67. HER-2 Protein and Herceptin
Herceptin (trastuzumab):
Metastatic breast cancer
Targets tumor cells that over-express the human epidermal growth factor receptor 2 (HER2) protein
Best response attained in women who over-express the HER2 protein
HER-2 over-expression in breast cancer cells should be done before patients receive the drug

68. Herceptin: Prescribing Information
HERCEPTIN (Trastuzumab) as a single agent is indicated for the treatment of patients with metastatic breast cancer whose tumors overexpress the HER2 protein and who have received one or more chemotherapy regimens for their metastatic disease.
HERCEPTIN should be used in patients whose tumors have been evaluated with an assay validated to predict HER2 protein overexpression

69. Hepatitis C
Interferon--2b and ribavirin are used to treat patients with hepatitis C virus
Different mutations exist in the hepatitis C virus
Knowledge of a person’s hepatitis C genotype may help play a role in the therapeutic decision-making process

70. HIV and Antiretroviral Drugs
Resistance to antiretroviral agents hinders the management of HIV disease
In the virus, mutations occur which confer drug resistance
Knowledge of viral genotype (or phenotype) can help guide the selection of antiretroviral therapy

71. Challenges Facing the Field of Pharmacogenomics
Multiple studies, but literature is inconclusive in some instances
Genetics accounts for an insufficient percentage of response variability for a given drug
Few studies documenting genotype-guided therapy is better than the “usual care” approach
Few “point-of-care” tests available to determine a person’s genetic make-up or protein expression

72. Potential Reasons for Discrepancies
Inadequately powered studies
Studying different drug response phenotypes
Studying different patient populations (differences in allele frequencies)
Problems precisely measuring phenotype
Subtlety of functional effects of polymorphisms
Focus on single SNPs instead of haplotypes
Failure to consider the complexity of drug response
Johnson JA and Lima JJ. Pharmacogenetics 2003; 13:

75. Pharmacogenetics and Pharmacogenomics Knowledge Base (PharmGKB)
Publicly accessible knowledge base
Goal: establish the definitive source of information about the interaction of genetic variability and drug response
Store and organize primary genotyping data
Correlate phenotypic measures of drug response with genotypic data
Curate major findings of the published literature
Provide information about complex drug pathways
Highlight genes that are critical for understanding pharmacogenomics

76. Role of Pharmacogenomics in the Drug Development Process
80% of products that enter the development pipeline FAIL to make it to market
Pharmacogenomics may contribute to a “smarter” drug development process
Allow for the prediction of efficacy/toxicity during clinical development
Make the process more efficient by decreasing the number of patients required to show efficacy in clinical trials
Decrease costs and time to bring drug to market
Drug Safety issues
Lack of efficacy
Manufacturing problems
8-10 years to see a product to market
$800 million to develop a new product

77. Pharmacogenomic Paradigm in the Drug Development Process
Current Options Options with Pharmacogenomics
High
Continue clinical trials
to market
Abandon drug
before market
Optimize clinical trials,
making them
smaller and shorter
Continue trials safely
by excluding at-risk pts
Proportion of patients showing
poor or no response
Low

78. Personalized Medicine and the Pharmaceutical Industry
Targeted Therapies:
Herceptin: treatment of HER2 positive metastatic breast cancer
Gleevec: treatment for patients with Philadelphia chromosome-positive chronic myeloid leukemia
Erlotinib: treatment for non-small cell lung cancer
Most effective in epidermal growth factor receptor positive tumors
Maraviroc (not approved): treatment for HIV
Studies have incorporated a screening process for different receptors that HIV uses to gain access to cells
Iloperidone (not approved): schizophrenia treatment
Company identified a genetic marker that predicts a good response to the drug

79. Pharmacogenomic Paradigm in the Pharmaceutical Industry
Efficacy prediction
Common side effect
prediction
Rare side effect
prediction
Market expansion

80. FDA and Pharmacogenomics
Traditionally, industry has been hesitant to submit pharmacogenomic data due to fears of:
Delays in drug development
Request for additional clinical studies
Potentially put clinical trials on hold
FDA published: “Draft Guidance for Industry: Pharmacogenomic Data Submission” in (currently under revision)
Set criteria for Voluntary Genomic Data Submission (VGDS)

81. Pharmacogenomics Information in the Published Literature
Zineh I et al. Ann Pharmacother ; 40:

82. Pharmacogenomics Information in FDA-Approved Prescribing Information
Zineh I et al. Pharmacogenomics J; 2004: 1-5

83. Moving Pharmacogenomics to Clinical Practice
Document Pgx superiority: Pgx-guided versus usual care
Documenting sufficient variability to predict clinical utility
Studies that mimic clinical practice
Proof-of-concept clinical studies
In vitro functional studies
Identify sequence variability in candidate genes
Johnson JA. Trends in Genetics 2003; 19:

84. The Future of Pharmacogenomics
Genome wide approach versus candidate gene approach
Thousands of SNPs
Thousands of patients
Replication studies
Sophisticated databases housing pharmacogenomic information
Drug selection and dosing algorithms incorporating non-genetic and genetic information
Point of care genetic testing

85. "Here's my
sequence..."
The New Yorker

86. “Personalized medicine: elusive dream or imminent reality
“Personalized medicine: elusive dream or imminent reality? In summary: it is both.”
Larry Lesko, Director of the FDA Office of Clinical Pharmacology and Biopharmaceutics
Clin Pharmacol Ther; 2007: