Pharmacogenomics

1. Pharmacogenomics • Drug Metabolism Monitoring • Drug Sensitivity Testing

2. Market Evolution Cancer Opportunity Pharmacogenomics Genetic disorders InfectiousDiseases Time

3. Pharmacogenomics… Alleles Is the relationship between a patient’s inherited genetic Makeup and their response to pharmaceutical drugs.

4. Clinical Goals of PGx: SAM • Safety of the patients • Avoidadverse drug reactions • Maximizedrug efficacy

5. Current Drug Treatment “One Size Fits All”

6. Actual Outcomes in the Clinic Reduced dose? Different drug? Normal dose OK Different drug?

7. Drug Response Rates Low!

8. Major Drugs are Ineffective for Most

9. Adverse Drug Responses Defined • Three outcomes for therapy • It works • It doesn’t work (efficacy) } Adverse Drug Response • It is toxic (safety) } Adverse Drug Response ADR’s – Huge Healthcare Costs $177 Billion annually CEDR-http://www.fda.gov/CDER/DRUG/drugreactions/default.htm

10. Adverse Drug Reactions Cases of Adverse Drug Reactions: Over 2 Million/Yr Cause of Death: 3rd in US Cause of Illness: 5th in US Cost of Mortality from ADRs: $177 Billion Sources: Trescot, et. al. – American Society of Interventional Pain Physicians

11. Drug Treatment Paradigm Genotype Based Treatment Trial & Error Genetic Test Drug A Side effects Metabolic Character Drug B Drug D Drug C Ineffective TARGETED

12. Genetic Factors Environmental Factors Drug Response Drug Variations in Drug Response Absorption Distribution Metabolism Excretion

13. Significance of Pharmacogenomics A patient’s inherited genetic makeup and their response to pharmaceutical drugs are seen with regards to their metabolism. Ultra-rapid Metabolizer Normal Metabolizer Poor Metabolizer Over-dosed: Adverse Drug Reactions Under-dosed: Lack of Efficacy Expected Response

14. Variations in Drug Response

15. Where Does Drug Metabolism Occur • Majority of drug metabolism occurs in the intestine and liver • Many drugs metabolized by members of same enzyme family

16. Drug Metabolizing Enzymes • Phase I (oxidative) • Cytochrome P450's (CYP) • Monoamine oxidase • Alcohol dehydrogenase • Aldehyde dehydrogenase • Dopamine B-hydroxylase • Phase II (conjugative) • N-acetyl transferase (NAT) • Thiopurine methyltransferase (TPMT)

17. Cytochrome P450 Enzymes The super-family of cytochrome P450 enzymes has a crucial role in the metabolism of drugs. Approximately 85% of all drugs in use are processed by one or more of these CYP450 enzymes!

18. Cytochrome P450 Enzymes Major CYP450 Isoforms 1. 1A2 2. 2B6 3. 2C8 4. 2C19 5. 2C9 6. 2D6 7. 2E1 8. 3A4 9. 3A5 10. 3A7

19. CYP450 Testing Applications Psychiatry – Antidepressants, Antipsychotics  Anticoagulants – Plavix, Warfarin, Prasrugel  Oncology – Tamoxifen, Chemotherapy  Pain management – Codeine, Vicodin 

20. Drugs Metabolized by CYP450 CYP2C9 Warfarin, Losartan, Naproxen, Fluconazole CYP2C19 Clopidogrel, Diazepam, Citalopram, Omeprazole CYP2D6 Fluoxetine, Haloperidol, Paroxetine, Tamoxifen CYP3A4 Carbamazepine, Cyclosporine, Irinotecan, Quinidine CYP3A5 Ritonavir, Rifampin, Rifabutin, Viagra

21. Driving PGx U.S. Food & Drug Administration “It’s mission is to advance and protect public health… by helping to speed innovations that make medicines more effective, safer, and more affordable.” - Dr. Mark McLennan FDA Commissioner

22. FDA Relabeling Initiatives Major FDA relabelings since 2000: • August 2002: Herceptin (Trastuzumab) HER-2 • March 2003: Strattera (Atomoxetine) 2D6 • July 2004: Purinethol (6-mercaptopurine) TPMT • November 2004: Irinotecan (Camptosar) UGT1A1 • August 2007: Warfarin (Coumadin) 2C9/VKORC1 • June 2009: Plavix (Clopidogrel) 2C19 • July 2009: Erbitux (Cetuximab) KRAS • July 2009: Vectibix (Panitumumab) KRAS • March 2010: Plavix (Clopidogrel) – Black Box Warning 2C19

23. Drugs Prescribed with Genetic Tests

24. Drug-Drug Interactions ADRs Due to Drug-Drug Interactions: 50-60% 7% of hospitalizations ADRs Due to DDIs in Hospitals: 28% of Patients 17% of Children Adverse Drug Reactions due to Drug-Drug Interactions Can Be Avoided! Sources: Trescot, et. al. – American Society of Interventional Pain Physicians

25. PGx Drug Interactions Inducer Drug B CYP450 Extensive Metabolizer CYP450 Ultra Metabolizer CYP450 Poor Metabolizer Drug A Response A Response B Response C Substrate No Response Expected Drug C Inhibitor

26. PGx Drug Interactions Inducer Drug B CYP450 Poor Metabolizer CYP450 Extensive Metabolizer Response C Response A Overdosed Response Response Drug A Substrate

27. CYP450 2C19 Clinical Relevance: Clopidogrel Dosing

28. CYP2C19 - Overview Prominent Among: • 40 % of the African Americans • 30 % of the Caucasians • > 50 % of the Asians

29. Clinical Relevance Prescription Medicine • Esomeprazole (Nexium) No. 2 in prescriptions, 2008 • For Erosive Esophagitis • CYP2C19 Substrate • Escitalopram (Lexapro) No. 3 in prescriptions, 2008 • For Depression • CYP2C19 Substrate • Lansoprazole (Prevacid) No. 7 in prescriptions, 2008 • For Duodenal Ulcers • CYP2C19 Substrate Sources: Verispan/VONA “Top Drugs”; The Merck Index/Lexi-Comp

30. CYP2C19 Drugs • Nilutamide • Norethindrone • Oxcarbazepine • Pantoprazole • Phenobarbitone • Prednisone • Primidone • Probenicid • Propranolol • Rabeprazole • Ticlopidine • Topiramate • Carbamazepine • Cimetidine • Citalopram • Clomipramine • Diazepam • Escitalopram • Fluoxetine • Iansoprazole • Isoniazid • Omeprazole • Phenytoin • Rifampin • Sertraline • Amitriptyline • Carisoprodol • Chloramphenicol • Felbamate • Fluvoxamine • Hexobarbital • Indomethacin • Ketoconazole • R-mephobarbital • S-mephenytoin • Moclobemide • Modafinil • Nelfinavir

31. CYP2C19 A Necessity in Clopidogrel (Plavix) Therapy

32. Plavix 101 • Plavix also known as Clopidogrel Bisulfate • Anti-platelet (prevents blood from clotting) • Prescribed for heart disease, stroke, and heart attack • Most often indicated for prevention of stent thrombosis • 29.5 million prescriptions dispensed in the US in 2010 • 2nd highest selling drug in the world • Sales of $6.35 Billion in 2010 (2nd only to Lipitor 12.9B) • > 90 million patients worldwide • Now generic as of May 17, 2012

33. Adverse Events Associated with Plavix Common Adverse Events include: Respiratory Tract Infections – 8.7% Chest Pain = 8.3% Flu Symptoms – 7.5% Hypertension = 4.7% Diarrhea = 4.5% Major Bleeding Events – 3.7% Depression – 3.6% Gastrointestinal Bleeding - 2.0% Intracranial Hemorrhage = 0.4% Fatal Bleeding = 0.2% Inadequate platelet inhibition can lead to acute stent thrombosis and death.

34. Plavix Dosing In Post-Stent Placement: Stent Clotting (Stent Thrombosis) tends to occur within 3 to 12 months after placement. Inability to remove clots after stent placement is a major issue. Proper Dosing of Plavix will Prevent Clotting. Failure of Plavix therapy typically leads to death or Acute Stent Thrombosis.

35. Plavix Response Response to Plavix varies widely: • Up to 30% of patients do not respond to Plavix. • Up to 25% of patients treated with standard doses of Plavix will have sub-optimal response. These patients are at higher risk of recurrent ischemic events, especially stent thrombosis. • Explanations: • Under-dosing • Drug Interactions • Genetic Polymorphism effect • Genetic variations in CYP450 2C19 significantly affect response to Plavix and alter clinical outcomes.

36. Plavix Metabolism • Plavix is a pro-drug requiring several biotransformation steps (mediated by CYP450 enzymes) in order to generate an active metabolite. • Upon absorption 85% of the drug is degraded into inactive metabolites. • The remaining 15% of the drug is metabolized by CYP450 isoenzymes 3A4, 3A5 & 2C19 into its active form.

37. Publication in New England Journal Of Medicine – Jan’098 CYP450 Polymorphisms and Response to Clopidogrel – Mega, J., et. al – Published in NEJM – January 2009 • Tested the association between functional genetic variants in CYP genes in response to Clopidogrel. Also examined the association between genetic variants and cardiovascular outcomes. = 1447 Patients Conclusions: • Carriers of reduced-function 2C19 alleles had significantly lower levels of the active metabolite of Clopidogrel. • Subjects carrying at least one 2C19 reduced function allele were at significantly higher risk (50% greater) for death from cardiovascular causes, myocardial infarction, or stroke than were non-carriers. • The risk of stent thrombosis in carriers of a 2C19 reduced-function allele was three times that among non-carriers.

38. Publication in JACC: Cardiovascular Interventions – Nov ‘09 Pharmacogenomic Testing for Clopidogrel using the Rapid INFINITI® Analyzer – Gladding, P., et. al. Published in JACC: Cardiovascular Interventions – Nov. 2009 • To assess the antiplatelet effect of a higher Clopidogrel maintenance dose in CYP2C19*2 carriers relative to noncarriers in 40 patients. Conclusions: • CYP2C19 genotyping using the AmeriGene PGT showed that 33% of carriers had a *2 mutation and 46% of carriers had *17. • Patients who were CYP2C19*2 carriers showed significantly reduced antiplatelet (platelet inhibition) response compared to noncarriers (18% carriers vs. 59% noncarriers). • Rapid genotyping with extensive multiplexing abilities will be necessary for pharmacogenomics to be useful in clinical settings. This will improve personalized Clopidogrel therapy which can greatly optimize treatment.

39. Publication in JAMA – October, 2010 Reduced-Function CYP2C19 Genotype and Risk of Adverse Clinical Outcomes among Patients Treated with Clopidogrel Predominantly for PCI – Mega, J.L., et al. • To define the risk of major adverse cardiovascular outcomes among carriers of 1 and 2 reduced-function CYP2C19 genetic variants in patients treated with Clopidogrel. = 9,865 patients Conclusions: • Among 9,685 patients, 26.3% had 1 reduced-function CYP2C19 allele and 2.2% had 2 reduced-function CYP2C19 alleles. • The CYP2C19 gene is associated with almost 1 in 3 patients not receiving ideal protection from ischemic events when treated with standard dose of Clopidogrel for PCI. • Among patients treated with Clopidogrel, carriage of even 1 reduced-function CYP2C19 allele appears to be associated with a significantly increased risk of death, MI, stroke and particularly stent thrombosis. • Given how widely Clopidogrel is used to treat patients with Cardiovascular disease, determination of the optimal antiplatelet treatment doses or regiments for individual patients is needed to tailor therapy appropriately.

40. FDA Relabels Plavix FDA in an emergency relabeling, relabeled Plavix (Clopidogrel) with genetic information that could help identify best responders and help physicians make more informed prescribing decisions. Lawrence Lesko, FDA, Director of Office of Clinical Pharmacology: “A DNA test for 2C19 status would identify the subgroup of patients who would be resistant to Plavix. This would be actionable by giving a higher dose of the drug, or a different drug.”

41. Plavix – Black Box March 12, 2010 – In it’s most severe warning to date, the FDA issues a black box warning in the Plavix package insert indicating that the drug carries a significant risk of serious or even life-threatening adverse effects. WARNING: DIMINISHED EFFECTIVENESS IN POOR METABOLIZERS See full prescribing information for complete boxed warning. • Effectiveness of Plavix depends on activation to an active metabolite by the cytochrome P450 (CYP) system, principally CYP2C19. • Poor metabolizers treated with Plavix at recommended doses exhibit higher cardiovascular event rates following acute coronary syndrome (ACS) or percutaneous coronary intervention (PCI) than patients with normal CYP2C19 function. • Tests are available to identify a patient's CYP2C19 genotype and can be used as an aid in determining therapeutic strategy. • Consider alternative treatment or treatment strategies in patients identified as CYP2C19 poor metabolizers.

42. Comments from the Field A leader in the field, Dr. Eric Topol (Director, Scripps Translational Institute) States: “At this moment in time in cardiovascular medicine, this is perhaps the most striking, practical pharmacogenetic relationship demonstrated (2C19 & Plavix).  It has very significant practical implications for managing patients, especially since Plavix is the second-most-prescribed drug in the world.” “Genotyping could be incorporated into daily practice. If two million patients are getting stents annually, then some 700,000 individuals are at increased risk for stent thrombosis based on their genetics.”

43. Plavix – Timeline to Critical Mass January 26, 2009 Early Communication about an Ongoing Safety Review of clopidogrel bisulfate – MedWatch/FDA “Differences in effectiveness may be due to genetic differences in the way the body metabolizes clopidogrel,or that using certain other drugs with clopidogrel can interfere with how the body metabolizes clopidogrel.” - FDA Report May 5, 2009 FDA Updates Plavix Labeling to Include Pharmacogenetic Information – MedWatch/FDA “Patients with genetically reduced CYP2C19 function have lower systemic exposure to the active metabolite of clopidogrel and diminished antiplatelet responses, and generally exhibit higher cardiovascular event rates following myocardial infarction than do patients with normal CYP2C19 function.” - FDA Update to Label November 17, 2009 FDA Updates Plavix Labeling to Warn Against Co-Administration with Other CYP2C19 Inhibitors – GenomeWeb “Clopidogrel is metabolized to its active metabolite in part by CYP2C19. Concomitant use of drugs that inhibit the activity of this enzyme results in reduced plasma concentrations of the active metabolite of clopidogrel and a reduction in platelet inhibition. Avoid concomitant use of drugs that inhibit CYP2C19…” -FDA Update to Label MARCH 12, 2010 FDA Updates Plavix Labeling with Black Box Warning - MedWach/FDA “Effectiveness of Plavix depends on activation to an active metabolite by the cytochrome P450 (CYP) system, principally CYP2C19” - FDA Update to Label

44. Histopath Offerings • Polypharmacy • Cardiovascular • Pain Management • Psychological Drugs • Urology • 2C9 • 2C19 • 2D6

45. Importance of Variants on 2C19 Panel Common polymorphisms of the 2C19 gene are seen in approximately: • 30% of Caucasians • 40% of African-Americans • >55% of Asians 2C19 *2-*10 • Significantly reduces activity of Plavix • Shown in numerous studies as a major determinant for risk of recurrent events in MI survivors treated with Plavix 2C19 *17 • Increases 2C19 expression and enables rapid activity of Plavix

46. Aetna Reimbursement Policy

47. CYP450 2C19 – Multiple Uses Not only does CYP450 2C19 play a significant role in relation to Plavix, but it also plays a large role in metabolizing other groups of drugs (15% of all drugs): • Antidepressants (Prozac) • Antiepileptics (Valium) • Proton Pump Inhibitors (PPIs)

48. CYP2C19: Clopidogrel Update From the Black Box till Now…

49. Recent Studies on Utility of 2C19 Genotyping • CYP2C19 Genotype, Clopidogrel Metabolism, Platelet Function, and Cardiovascular Events, a Systematic Review and Meta-analysis • Journal of the American Medical Association, Dec., 2011 • Clinical, Angiographic, and Genetic Factors Associated with Early Coronary Stent Thrombosis • Journal of the American Medical Association, Oct., 2011 • Clinical Pharmacogenetics Implementation Consortium Guidelines for CYP450 2C19 Genotype and Clopidogrel • Clinical Pharmacology & Therapeutics, Jun. 2011

50. Utility of 2C19: Meta-analysis CYP2C19 Genotype, Clopidogrel Metabolism, Platelet Function, and Cardiovascular Events, a Systematic Review and Meta-analysis Objective: • To provide a consensus conclusion on the risk of various cardiovascular events in association with clopidogrel response and CYP2C19 genotype based on all evidence available to date. Conclusions: • 2C19 status was not associated with risk of stent thrombosis when considering total incidence over all 32 studies. • 2C19 status was not associated with risk of cardiovascular disease in those treated with clopidogrel when considering incidence over all 26 studies. Rebuttals: • Of the 32 studies, only 14 accounted for Stent Thrombosis. The conclusion of null association was thus drawn from a dilution of data. • Of the 26 studies, only 10 accounted for Stent Thrombosis. No data was presented on differentiation of 2C19 genotype by patient interaction.