Phase 1 Bioavailability(BA)/Bioequivalence(BE) & Fed Studies Ruth E. Stevens, PhD, MBA Chief Scientific Officer, Executive Vice President Camargo Pharmaceutical Services ACPU: October 19, 2010

Approved Product Labeling Drug Substance Indication Safety Clinical Pharmacology DDI Pharmacokinetics (BA/BE/FED) Reproductive

Pharmacokinetics Therapeutics Concentration Pharmacokinetics Dose What the body does to the drug (Absorption, Distribution, Metabolism and Excretion (ADME)). PHARMACODYNAMICS: What the drug does to the body (Therapeutic Effects, Side Effects). Therapeutics Concentration Pharmacokinetics Dose Effect Pharmacodynamics

Bioavailability Cmax AUC For the purpose of this subsection: (A) The term “bioavailability” means the rate and extent to which the active ingredient or therapeutic ingredient is absorbed from a drug and becomes available at the site of drug action. AUC Note: AUC = AUC0-t and AUC0-inf Reference: Food, Drug and Cosmetic Act Section 505(j)(7) Bioequivalence

Comparative Bioavailability Study (Rate and Extent of Absorption) Y axis-Linear Scale Y axis-Log Scale A = Test; 1x/day B = Reference; 3x/day

Bioequivalence: Cmax AUC AUC (B) A drug shall be considered to be bioequivalent to a listed drug if: the rate and extent of absorption of the drug do not shown a significant difference from the rate and extent of absorption of the listed drug when administered at the same molar dose of the therapeutic ingredient under similar experimental conditions in either a single dose or multiple doses; or the extent of absorption of the drug does not show a significant difference from the extent of absorption of the listed drug when administered at the same molar dose of the therapeutic ingredient under similar experimental conditions in either a single dose or multiple doses and the difference from the listed drug in the rate of absorption of the drug in intentional, is reflected in its proposed labeling, is not essential to the attainment of effective body drug concentrations on chronic use, and is considered medically insignificant for the drug. AUC Note: AUC = AUC0-t and AUC0-inf Reference: Food, Drug and Cosmetic Act Section 505(j)(7) Bioequivalence

Bioequivalent Products - Concentration – Time Profiles Rate & Extent = “SAME” Y axis-Linear Scale Y axis-Log Scale

“Extent of Absorption” Area Under-the-Curve (AUC) “Extent of Absorption” LOQ = Limit of Quantitation or LQC = Last Quantifiable Concentration Concentration (ng/mL) Biological Matrix LOQ Time (hrs)

**Area Under-the-Curve (AUC0-t) “Extent of Absorption” ** = Pivotal Bioequivalence PK Parameter AUC 0-t or AUC0-last: Area under the plasma concentration-time curve from time zero to the last measurable time point. Concentration (ng/mL) Biological Matrix t Time (hrs)

**Area Under-the-Curve (AUC0-inf) ** = Pivotal Bioequivalence PK Parameter “Extent of Absorption” Concentration (ng/mL) Biological Matrix AUC 0-inf or AUC∞: Area under the plasma concentration-time curve from time zero to time infinity. 12 24 Time (hrs)

**Maximum Observed Concentration, Cmax “Rate” Value read off Y-axis  Concentration (ng/mL) Biological Matrix Cmax ** = Pivotal Bioequivalence PK Parameter 12 24 Time (hrs)

Time to Maximum Concentration, Tmax Concentration (ng/mL) Biological Matrix Tmax Value read off X-axis “Rate” 12 24 Time (hrs)

Terminal Elimination Rate Constant z (lambda, lambda z or Kel) Rate & Extent of Absorption z or Kel: slope ≥4 timepoints Concentration (ng/mL) Biological Matrix 12 24 Time (hrs)

Drug Elimination Half-Life, (t½) z = terminal slope Ln(2) = natural log of 2 Concentration (ng/mL) Biological Matrix t1/2 = ln(2)/z = 0.693 z ≥4 points on terminal slope to calculate z 12 24 Time (hrs)

Major Elements of Study Designs [BA, BE, Fed] (Example: Immediate-Release Products) Subject Selection Number of subjects: typically 24-36 Major objective: minimize intersubject variation conducted in healthy subjects, 18-50 years old, ± 10% (range 10%-20%) of ideal body weight. Populations traditionally excluded Elderly: stress, blood loss, chronic disease and polypharmacy, PK effects of altered organ function Patients: stress, blood loss, concurrent medications, special diets, PK effects of disease states Some Exceptions to When Patients are Enrolled Instead of Healthy Subjects: studies with pharmacodynamic or clinical end points, cytotoxic drugs Females are no longer excluded

Major Elements of Study Designs - Fed State (Example: Immediate-Release Products) 2. Meal – Office of Generic Drugs [Egg McMuffin] Thirty minutes before dosing, subjects are served a high-fat content meal consisting of: One fried egg One slice of American cheese One slice of Canadian bacon One buttered English muffin One serving of hash brown potatoes 180mL of orange juice 240mL of whole milk Grams No. of Calories Percent of Total Calories Protein 33 132 15.4% Fat 55 280 35.3% Carbohydrate 58 232 27%

Major Elements of Study Designs [BA, BE, Fed] (Example: Immediate-Release Products) 3. Exclusion Criteria Major organ, systemic, or mental disease, hypersensitivity to drug product or class recent participation in investigational drug studies, recent blood donation, recent exposure to enzyme-inducing or inhibiting agents abnormal diets or recent significant weight loss. 4. Restrictions No Rx medications within two weeks or OTC products within two days of study start. No alcohol for 48 hours prior to dosing and during sampling no xanthine-containing products for 48 hours prior to dosing and during sampling. No strenuous exercise or immobilization (except during sleeping times); normal activity for four hours post-dose. 5. Informed Consent, IRB approval 6. Additional subjects enrolled to replace dropouts

Major Elements of Study Designs [BA, BE, Fed] (Example: Immediate-Release Products) Overnight fast of at least 10 hours, fasting continued for four hours post dose, then standardized meals. Dose administered with 240 mL of water; fluids restricted within ± 1 hour of dosing. Two-way crossover: Subject randomization Washout period between treatments (7-10 drug half-lives) Potencies of test and reference products within ± 5%. Period 1 Period 2 Sequence 1 Treatment 1 Treatment 2 Sequence 2

Statistical Designs Reference Test Test Reference Crossover Studies: Subject receives each of the formulations one at a time in different time periods. Designed to eliminate individual differences. 2 x 2 Crossover Designs R A N D O M I Z T Period Subjects Sequence 1 I II W A S H O U T Reference Test Sequence 2 Test Reference

Statistical Designs: Crossover Study (Subject Acts as Own Control) 1 Test Period I Estimated Intra-subject Variability Reference Period II Intra-subject Variability (Estimated): variability within a subject

Major Elements of Study Designs [BA, BE, Fed] (Immediate-Release Products Example, Fasting Study) 8. Sampling (plasma, serum, whole blood) Sufficient sampling during absorption phase to define adequately to ascending portion of the curve; avoid first nonzero concentration as the Cmax. Intensive sampling around the time of the expected Cmax. Sampling duration of at least 3 to 6 drug half-lives (NDA) or 7-10 drug half-lives (ANDA) or longest half-life of any analyte.

Major Elements of Study Designs [BA, BE, Fed] (Immediate-Release Products Example, Fasting Study) 9. Pharmacokinetic Parameters Area Under the Curve (AUC) AUC0-t: Time of the last quantifiable concentration Calculated by the trapezoidal rule AUC0-inf: Extrapolated to infinity = AUC0-t + Ct/Kel Peak concentration (Cmax) and time to Cmax (Tmax) are obtained directly from the observed data Terminal elimination rate constant (Kel, λz) and half-life, t1/2 = ln(2)/Kel.

Pharmacokinetic Analysis BA/BE & FED Plasma/Serum and/or Whole Blood Drug level at sampling times Pharmacokinetic parameters *AUC0-t Last quantifiable concentration *AUC0-inf Infinity *Cmax Peak concentration Tmax Time to peak concentration Kel Terminal elimination rate constant t1/2 Elimination half-life CL/F Clearance Vd Volume of Distribution Urine Drug level at sampling intervals (Ae) Pharmacokinetic parameters Cumulative excretion (*Ae0-t) Maximum excretion rate (*Rmax) Time to maximum excretion rate (Tmax) The following information on urine potassium concentration data should be recorded for each subject: Amount excreted in each collection interval (Ae) Cumulative urinary excretion from 0 to 24 hours (Ae0-24h) Cumulative urinary excretion from 0 to 48 hours (Ae0-48h) Maximal rate of urinary excretion (Rmax) Time of maximal urinary excretion (Tmax) Excretion rate in each collection interval (R) Midpoint of each collection interval (t) All data should be calculated using baseline-adjusted and nonbaseline-adjusted data. Statistical analysis (p=0.05) should be done by ANOVA for baseline-adjusted parameters, and the 90 percent confidence intervals should be generated for natural log-transformed cumulative urinary excretion from 0 to 24 hours (Ae0-24h) and maximal rate of urinary excretion data (Rmax). The two one-sided tests procedure can be used to determine 90 percent confidence intervals. V. IN VITRO TESTING * = Pivotal Bioequivalence PK Parameter

Statistical Requirements: Bioequivalence Two one-sided tests procedure (also called the 90% confidence interval approach) The July 1, 1992 Statistical Procedures Guidance requires 90% confidence interval limits from 80% to 125% based upon log transformed AUC0-t, AUC0-inf and Cmax data. Result must be between Lower Bound 80% and Upper Bound 125% {Ln 80 – 125}.

BE Confidence Intervals For Bioequivalence, the 90% confidence interval of F’ must fall between 0.80 and 1.25. 0.80 1.00 1.25 BE SD X BE = Bioequivalent SD = Statistically Different

Statistical Designs - BE Subject-By-Formulation Interaction A statistical term, meaning that the difference between the subject-specific means for the test product and the reference product is not the same for all subjects in the population. The possibility of subject-by-formulation interaction is one of the main concerns in the discussions of “bioequivalence”.

Side-by-Side Spaghetti Plots: FASTED Test Product (n=108) Reference Product (n=108)

Phase 1: BA/BE (and/or FED) Concentration – Time Profiles Y axis-Linear Scale Y axis-Log Scale

Bioequivalence (BE) Excellent Concentration – Time Profile

Subject by Formulation Interaction? A=Test B=Reference

Cmax Plot: Text to Reference OK for the 22nd – not the 23rd Ratio: Cmax (Test)/Cmax (Reference) = 0.98

Pharmacokinetics Therapeutics Concentration Pharmacokinetics Dose What the body does to the drug (Absorption, Distribution, Metabolism and Excretion (ADME)). PHARMACODYNAMICS: What the drug does to the body (Therapeutic Effects, Side Effects). Therapeutics Concentration Pharmacokinetics Dose Effect Pharmacodynamics

Conclusion: What is in your Drug Product Labeling? Drug Substance Indication Clinical Pharmacology Safety DDI Pharmacokinetics (BA/BE/FED) Reproductive

Thank You ACPU Committee Dr. Charles Pierce Dr. Punkaj Desai Dr Thank You ACPU Committee Dr. Charles Pierce Dr. Punkaj Desai Dr. William Sietsema