PHARMACOGENETICS. Marshalling the human genome to individualize drug therapy

Transcription

1 PHARMACOGENETICS Marshalling the human genome to individualize drug therapy Prof. Ronen Loebstein Institute of Clinical Pharmacology & Toxicology Sheba Medical Center

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4 Some examples of drug response rates

5 How do we know if a patient will respond (or have an adverse response) to a drug? (we don t)

6 Dose determination Phase I clinical trials (safety) 20 to 80 healthy volunteers Determine max tolerable dose Phase II clinical trials (efficacy) Several hundred patients estimate therapeutic dose range Drugs approved with a specific indication and dose range Phase III monitoring. Dose that patients actually receive depends on Prior treatment history of patient (drug naïve?) Physicians experience with prescribing drug Empirical knowledge of appropriate dose In practice start low, ramp to ~80% of recommended dose before trying different drug Liability issues No clinical trials are done to study dose escalation Therapeutic drug monitoring

7 Potential causes of variability in drug effects: Pathogenesis and the severity of the disease being treated. Drug interactions from concomitant treatments (plasma protein binding, metabolism) Individual s age, gender, lifestyle (including environmental factors), behavior, nutritional state, renal and liver function, and concomitant illnesses. Genetic variation

8 A patient s response to a drug may depend on factors that can vary according to the alleles that an individual carries, including Pharmacokinetic factors Absorption Distribution Metabolism Elimination Pharmacodynamic factors target proteins downstream messengers

9 G6PD deficiency Story Hemolysis occurs in G6PD deficient individuals due to the consumption of certain foods or drugs. NADPH and GSH are necessary for the reduction of hydrogen peroxide. RBC membrane is unstable in the presence of free radicals and hydrogen peroxide.

10 G6PD Gene Location: q28 locus of X-chromosome. gene: 13 exons, 12 introns. 18,5 Kbp. 400 variant alleles point mutations, deletions and replacements. The G6PD enzyme is not fully active when it is mutated. Instability of the mutated enzymes due to its susceptibility to proteolytic enzymes.

11 Succinylcholine Story Mode of action: depolarizing muscle relaxant. Time of onset: minutes. Clinical duration: 5-8 minutes. Mode of elimination: hydrolysis by plasma ChEst. Prolonged muscle relaxation Due to Inherited deficiency of plasma cholinesterase

12 Pharmacogenetics of Plasma Cholinesterase Gene location: E1 locus on q of chromosome 3. Allelic variants: Eu (wt), Ea, Ef, Es. Ea: atipical dibucaine-resistant variant A209G Asp70Gly Polymorphism in phenotyping ED < 8 min 60 > ED > 8 min ED > 1 hr ED > 8 hr Polymorphism in genotyping Eu/Eu Eu/Ea; Eu/Ef; Eu/Es Ea/Ea; Ef/Ef Es/Es % of population :10 5 ED = effect duration

13 The isoniazid story Use: chemotherapy of tuberculosis Daily dose: 5mg/Kg PO Metabolism: acetylation by NAT; non active metabolite. Neurotoxicity (2% of population at 5mg/Kg and 10-20% at higher doses): peripheral neuritis, optic neuritis, dizzinesss, ataxia, paresthesias, muscle twitching, stupor, toxic encephalopathy.

14 300 pts. Isoniazide PO 5 mg/kg. serum levels after 3hrs.

15 Heterogeneity in the way individuals respond to medications in terms of: Efficacy Toxicity

16 Pharmacogenomics The ultimate goal of pharmacogenomics is to define the contribution of the genetics differences in drug disposition or drug targets to drug response, thereby to improve the safety and the efficacy of drug therapy through use of genetically guided, individualized treatment.

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25 Genotype vs Phenotype Polymorphism: drug deactivation

26 WARFARIN Oral anticoagulant. S-warfarin is hydroxylated to inactive 7-OH W by CYP2C9. CYP2C9*1:100%; *2(SNP in exon 3): 12%: * 3(SNP in exon 7): 5% activity

27 CYP2C9 mutations are associated with adverse clinical outcomes Higher incidence of serious bleeding: vs 4.89 pt-yr. Higher incidence of life-threatening bleeding: 1.56 vs 0.7 pt-yr. Increased risk of above-range INRs: (hazard ratio: 1.4). More time to achieve a stable warfarin dose (95 days; p<0.004). Increased risk of bleeding during initiation of therapy (hazard ratio 3.94). Increased risk of bleeding at any time during therapy (hazard ratio 2.39).

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29 Genotype vs Phenotype Polymorphism drug activation (1)

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34 Codeine and breastfeeding Pharmacogenetics of morphine poisoning in a breastfed neonate of a codeineprescribed mother Did the drug cause death? Codeine and breastfeeding

35 Codeine and breastfeeding

36 Clopidogrel

37 Clopidogrel

38 Clopidogrel and CYP2C19 polymorphism

39 Clopidogrel and CYP2C19 polymorphism

40 CYP 2C19 polymorphism and clopidogrel response

41 Tamoxifen

42 CYP 2D6 polymorphism and Tamoxifen

43 CYP2D6 effect of gene duplication: inadequate therapeutic response to standard doses of nortriptyline tricyclic antidepressant. Norepinephrine reuptake inhibitor. Undergo oxidation by CYP2D6 yielding inactive 10-OH metabolite. NEJM 6 feb 2003

44 Polymorphisms of the ß 2 - receptor (1) Gene: 1,239 nucleic acids (chromosome 5q) Protein: 413 AA Distribution: arterioles, veins, bronchi: vaso-/ bronchodilation Genotype: 9 single nucleotide polymorphisms, 5 silent 4 missense SNPs (at AA 16, 27, 34, 164)

45 ß 2 - Receptor-SNPs and Drug Response Bronchodilator responsiveness Among mild asthmatics, Arg16-homozygotes (WT) have a faster and greater bronchodilator response to oral albuterol (after a 12 hour drug holiday) when compared to Arg16Glyhomozygotes or heterozygotes PK PD CPT 1999

46 Gefitinib (Iressa)

47 Gefitinib Epidermal Growth Factor Receptor over-expressed in 40-80% of Non-Small-Cell Lung Carcinomas EGFR signaling triggered by binding of growth factors, resulting in dimerization of receptor, auto/trans phosphorylation via tyrosine kinase domain, recruitment of downstream effectors and activation of cell prolioferation/survival programs Gefitinib inhibits EGFR by blocking ATP cleft, thereby preventing activation by autophosphorylation Tumor responses seen in only ~10% of patients with chemotherapy resistant advanced NSCLC A subgroup of patients with NSCLC have specific mutations in the EGFR gene that constitutively activate the receptor, and confer sensitivity ot Gefitinib Lynch TJ et al., NEJM 350,2129; Paez JG et al., Science, 304,1497

48 Gefitinib 25/275 patients at Mass General identified as being responsive. EGFR sequenced in 9 responsive patients.

49 Gefitinib All mutations in kinase domain Matched tissue showed wild type sequence. No mutations seen in 7 nonresponsive cases analysed Heterozygous mutations seen in 2 cases. Dominant gain of function mutation Heterologous expression in Cos-7 cells shows mutant receptors (IC50 = 0.015uM) more sensitive to inhibition by gefitinib than wild type (IC50 = 0.1uM).

50 Gefitinib Conclusions Only a subgroup of NSCLC tumors harbour EGFR mutations. This subtype of NSCLC sensitive to genfitinib Patients should be screened for EGFR mutations, and if they have them, be given gefitinib as first line therapy.

51 P-Glycoprotein

52 P-glycoprotein genetic polymorphism

53 Polymorphisms in drug transporters

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55 Polygenic determinants of drug response NEJM 6 feb 2003 p

56 Treatment modifications and patient genotypes 8(6): ,2002

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58 Under-carboxylated (PIVKA) Coagulation factor II, VII, IX, X Glutamic acid Coagulation factor functional γ-carboxyglutamic acid carboxylase Vitamin K-H 2 (hydroquinone) Vitamin K epoxide Vit K reductase (VKOR) warfarin

59 Calumenin- a regulator of the vitamin K cycle Carboxylase Vitamin K 1 H C A L U M E N I N Vitamin K 2,3 epoxide VKOR

60 VKOR-a multicomponent enzyme complex Glutathione-S- Transferase VKORC1 Microsomal Epoxide Hydrolase High homology between GST A1 and purified microsomes that reconstituted VKOR activity An 18 kd warfarin sensitive protein. Missense mutations in its encoding gene associated with inherited warfarin resistance and VKDCF deficiencies Anchored to the ER membrane and harbors vitamin K 2,3 epoxide binding Sites.

61 Warfarin daily dose (mg) by genotype Warfarin daily dose (mg) wt/wt wt/mu CYP2C9 VKORC1 mu/mu CYP2C9 & VKORC1 genotype

62 Warfarin daily dose by genotype combinations Compound genotype N CYP2C9 VKORC1 CALU Warfarin daily dose (mg/day) mean ± SE I ± 0.8 II ± 2.1* III ± 0.5 IV ± 0.5 V ± 0.9 VI ± 1.1 VII ± 0.3** VIII ± 1.1

63 The relative effects of selective variables on warfarin dose requirements (mg/day) Variable β-coefficient Partial r 2 Significance Age Weight K 1 H CYP2C9 genotype VKORC1 genotype Total r 2 =0.714

64 A novel coding VKORC1 polymorphism Asp36tyr Resistant Patients (n=15) Sensitive Patients (n=8) Control patients (n=99) Asp36Tyr polymorphism 7/15 0/8 8/99 CYP2C9 alleles 2/30 8/16 47/198

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