1. Key Pharmacokinetic Concepts – Single Dose and Steady State Drug Administration Pankaj B. Desai. Ph.D. Professor of Pharmacokinetics and Biopharmaceutics Director, Drug Development Graduate Program

2. Morning Agenda: Wake Up and Smell the Coffee (Cytochrome P450 1A2 Substrate) • Overview of ADME principles • Important PK Parameters • First Pass Metabolism • Compartmental & Non- Compartmental Analyses • Single Dose Kinetics • Multiple Dose Kinetics • Drug-Drug Interactions • Inter-Subject Variability CYP1A2 Substrate

3. ADME ADME ADME ISSUES IN ANTIssues I-CANCER DRUG DEVELOPMENT

4. Clinical Pharmacology • First in Human -Pharmacokinetically Guided Dose Escalation/ Drug Tolerance Study • Pharmacokinetics-Pharmacodynamics • Drug Metabolism • Mass Balance with Radiolabeled Compounds • Bioequivalence:Generic compounds • Single and multiple doses • Conventional versus controlled release formulations • Bioavailability of metabolites • Drug-Drug/Drug Dietary Product Interactions • Special Populations

5. Drug Input & Different Routes of Administration • I.V. and I.A. injections: • Bolus dosing • Zero-Order Input (Infusions) • Extravascular Administration • First Order (mostly passive diffusion) • Zero Order (active transport and controlled release systems)

6. Factors Affecting Drug Distribution • Phyisco-chemical properties of the drug • Small vs. Large mol.wt. Compounds • Hydrophilic vs. Lipophilic compounds • pH of the milieu and pKa of the drug • Perfusion rate (blood flow/min/g tissue) • Protein binding • Anatomical restrictions • CNS- protected by the blood brain barrier • Transport across placenta • Salivary Drug Excretion (S/P ratios) • Excretion of the drug in milk (M/P ratios)

7. Drug L/Kg L/70 kg Sulfisoxazole 0.16 11.2 Phenytoin 0.63 44.1 Phenobarbital 0.55 38.5 Diazepam 2.4 168 Digoxin 7 490 Apparent Volume of Distribution • Mathematical term to correlate amount & concentration • Merely a tool to understand the EXTENT of drug distribution- not a real physiological volume • Compare to the volume of body waters • Best calculated from I.V. Dosing as I.V. Dose/Cpo

8. Conc = 0.2 mg/ml Vd = 500 ml 100 mg Beaker with Charcoal Apparent Volume of Distribution Conc = 2 mg/ml Vd = 50 ml Plasma Water-3.5 L, ~4.5 % body wt (w/w) 100 mg ECW Total extracellular water - 15 L, 20 % body wt (w/w) TBW Total Intracellular water –20 L, 30 % body wt (w/w) Beaker without Charcoal Total body Water 40 L, ~55 % body wt (w/w)

9. Biotransformation Excretion Major Drug Elimination Pathways(Coordinated defense mechanism) Extra-Hepatic HEPATIC Renal Biliary Phase I Phase II

12. Glomerular Filtration • Kidney receives 1.1 L of blood (20 – 25%) of cardiac output • 10 % is filtered at the glomerulus • Compounds with Mol.wt < 20,000 filtered • GFR = 120 ml/min • CLR of Inulin - a measure of GFR • Filtered freely into the tubule • Not influenced by protein binding and neither secreted nor reabsorbed • Rate of filtration = Fu. Cp.GFR • Not a very effective drug extraction process (maximal ~ 0.11 or 10 %)

13. Active Secretion • Detected when the overall rate of urinary drug excretion exceeds the rate of filtration • Secretory processes (proteins) located predominantly within the proximal tubules • Mechanisms exist for secreting acids (anions) and bases (cations) from plasma into the tubular lumen • Energy-dependent • Saturable processes • Subject to competitive inhibition • Effect of Protein-Binding • Depends upon secretion efficiency and contact time at the secretory sites • Restrictive (dependent on the Fub) vs. Non-Restrictive (perfusion-rate limited)

14. Reabsorption • Must occur when CLR < fu.GFR • Reabsorption occurs all long the nephron, associated with reabsorption of water; majority however occurring from the proximal tubules • Predominantly a passive diffusion process • Driven by concentration-gradient across the tubular lumen • Active secretion occurs for many endogenous compounds such as vitamins, electrolytes, glucose and amino acids • Urine-Plasma Ratio (U/P) based on Henderson-Hasselbalch equation • Influence of pKa and pH of urine

15. Major Tissues Involved in Drug Metabolism • Liver • Small intestines • Kidney • Lung • Other portals of entry into the body and protected organs. -e.g. nasal mucosa

16. Representation of drug metabolism and excretion by the hepatocyte

17. Biliary Excretion is Transporter Mediated

19. Phase I and Phase II Drug Metabolizing Enzymes Phase I enzymes: Predominantly cytochrome P450 (CYP)

20. Drug Metabolism by CYPs Theophylline, caffeine, Olanzapine CYP2A6 (Coumarin) CYP2E1 (Chlorzoxazone) CYP1A2 CYP2B6 bupropion, tamoxifen, efavirenz 5% CYP2C8 Paclitaxel Rosiglitazone cerivastatin CYP2C9 (15%) Includes: warfarin phenytoin tolbutamide Losartan CYP3A (50%) Includes: lovastatin cyclosporin nifedipine midazolam ethinylestradiol Ritonavir Midazolam testosterone CYP2D6 (25%) Includes: Tricyclic antidepressants, SSRI's, haliperidol, propanolol, atomoxetine Detxromethorphan,

21. Phase II Reactions • Also known as Synthetic (conjugation) reactions • Major reaction: Transfer of the conjugating moiety to the drug • Enzymes involved are “transferase” • Glucuronosyl transferase • Sulfotransferases • N-acetyltransferase • Methyltransferase • Glycine transferase • Glutathione-S-transferase

22. Drug Biotransformation Reactions • Active Drug to Inactive Metabolite • Amphetamine Phenylacetone • Phenobarbital Hydroxyphenobarbital • Taxol 6-hydroxytaxol • Active Drug to Active Metabolite • Codeine Morphine • Procainamide N-acetylprocainamide • tamoxifen 4-hydroxytamoxifen

23. Drug Biotransformation Reactions • Inactive Drug to Active Metabolite • Hetacillin Ampicillin • Sulfasalazine Sulfapyridine + 5 ASA • Cyclophosphamide Nitrogen mustard • Active Drug to Reactive Intermediates • Acetaminophen Reactive metabolites (hepatic necrosis) • Benzo(a)pyrene Reactive metabolite (carcinogenic)

26. Nomenclature • Basis: Amino acid sequence • Families: Less than 40 % a.a. sequence assigned to different gene families (gene families 1, 2, 3, 4 etc.) • Subfamilies: 40 – 55 % identical sequence (2A, 2B, 2C, 3A etc.) CYP3A4 Family Isoform Subfamily

27. CYP Nomenclature (Contd.) • Cytochrome P450 Nomenclature, e.g. for CYP2D6 • CYP = cytochrome P450 • 2 = genetic family • D = genetic sub-family • 6 = specific gene • NOTE that this nomenclature is genetically based: it has NO functional implication

29. Examples of CYP mediated Oxidative Metabolism Examples of reactions catalyzed by cytochrome P450: Hydroxylation of aliphatic carbon

30. Examples of CYP mediated Oxidative Metabolism Examples of reactions catalyzed by cytochrome P450: Heteroatom dealkylation

31. Clearance Concepts

33. Compartmental Modeling

34. One-Compartment Open Model I.V. bolus DB1 Cp1 Vd k10 K10 = overall Elimination Rate Constant

35. I.V. Bolus

36. Two-compartment Open model Central or Plasma Tissue k12 Dt Ct Vt I.V. bolus Cp1 VC Dp k21 1- hybrid rate constant (distribution) z- hybrid rate constant (terminal)

37. Two-compartment Open Model Elimination only

38. Blood flow to human tissues

39. Extravascular dose Dp Cp Vd e.v. dose ka Site of absorption k10

41. NCA Used to estimate • AUC • Bioavailability • Clearance • Volume of Distribution • Average Steady State Concentration

42. AUC Trapezoidal Rule AUC= ½(t3-t2)(C2+C3)

43. AUC

44. Example Cp(last)= 2.75/0.1419

45. Bioavailability • Absolute Bioavailability • Relative Bioavailability

46. Bioequivalence • Two products are considered to be bioequivalent if the concentration time profiles are so similar that they are likely to produce clinically relevant differences in either efficacy or toxicity. • Common measures used to assess differences are Tmax, Cmax and AUC.

47. Other Parameters • CL = Di.v/AUC • AUMC = ½(t2-t1)(C1t1 +C2t2) • MRT (Mean Residence Time) = AUMC/AUC or MRT = 1/K or CL/V • Vss = CL. MRT

48. Multiple Dosing –Overall Aims Key Concepts Principle of Superposition Drug Accumulation and Steady State Persistence Factor and Accumulation Factor Peak, Trough and Steady State Average Levels Applications Determination of drug concentrations and amounts following multiple i.v. and e.v. doses (Ka > > K10) max, min and during a dosing interval Determination of dosing regimens Doses (Maintenance and Loading) and Dosing Interval Cpmax consideration Cpmin consideration Cpmax and Cpmin consideration Practical Considerations in Decision Making

49. Drug Accumulation Depends on Frequency of Administration

50. Multiple I.V. Dosing The AUC within a dosing interval at steady state is equal to the total AUC of a single dose.