Charlie Taylor, PhD CpTaylor Consulting Chelsea, MI, USA Dosage Determination from Preclinical to Proof-of-Concept Trials, (Including Toxicology) Charlie Taylor, PhD CpTaylor Consulting Chelsea, MI, USA

Choosing Doses for POC: Preclinical and early studies that enable dose selection Reasons for drug failure in clinical phase 2-3 Need to choose both low (ineffective) and high (maximum tolerated) doses within dose range Biomarkers (one endpoint: animal → human translation) PK/PD modeling – EC50 as a target for efficacy or AEs Toxicology/toxicokinetics – daily AUC(0-24) as a limit Putting it together – visualizing multiple datasets Human population PK modeling – determine which doses best fit the constraints

Sequence of Studies Needed Prior to Clinical Proof-of-Concept * Preclinical in vitro studies: action at drug target (pharmacology) Preclinical in vivo pain models: indicate treatment of disease Safety pharmacology: animal studies for adverse effects Preclinical (& human liver microsome) metabolism studies Animal toxicology & toxicokinetic studies to identify safety issues Clinical Phase I studies of pharmacokinetics and tolerance in healthy human volunteers (Optional) Biomarker studies with both animal models and humans to establish proof of pharmacology in vivo (apart from efficacy) * * * * * * Requires in vivo unbound plasma drug concentration or daily drug exposure to help choose human efficacy DOSES

Drug Development Failures – UK ’64-’85 Prentis et al. (1988) Brit J Clin Pharmacol 25:387-396

Determine Initial Phase 2 Dose Range ?? Clinical doses MUST encompass both low end (lack of efficacy) and high end (maximum tolerance) Data from animal efficacy, animal safety, biomarker and human tolerance ALL must be considered The peak unbound plasma drug concentration (animal studies), daily exposure (AUC0-24 - tox) and human multiple-dose PK each need consideration How to consider all these factors??

One Approach: Biomarkers – Surrogate HUMAN Endpoints for Efficacy Defines drug action in vivo Examples: Imaging, Adverse Event or Mechanism e.g. PET to measure receptor occupancy in CNS e.g. Nystagmus, dizziness, balance platform e.g. Experimental pain model w/ volunteers e.g. Electrographic response (EEG, retinogram, TMS) Biomarkers Allow no-go decision prior to proof of efficacy, for example: Poor oral drug absorption or lack of CNS penetration Lack of receptor occupancy at highest safe dose

Hypothetical Human Biomarker: Criterion: 75% drug receptor occupancy in human brain @ high dose This criterion met at animal effective dose (animal PET study) Drug displacement of PET ligand in human brain: 18F-x-drugamine given IV in tracer amount If greatest human volunteer dose of experimental drug reaches < 30% occupancy, NO-GO If greatest human volunteer dose > 75% occupancy, GO (further development) Caveat: Criteria must be selected based on results with a prior known compound – Otherwise, risk of poor validation

Toxicology Findings (non-pharmacology) are Based on Daily Drug Exposure (AUC0-24) Repeated-dose animal tox studies determine lowest toxic dose and greatest no-effect (daily) dose Toxicokinetics determine drug exposure (AUC0-24) in mg•hr/mL at greatest no-effect dose e.g. Drug X has 8 hr half-life; Cmax and AUC are determined from plasma drug samples taken 0, 1, 4, 7, 12, and 24 hr after single oral dose at steady-state Similar human pharmacokinetic data and PK modeling determine human drug exposure (AUC0-24) @ doses Analysis is adjusted for different drug binding of plasma proteins between species

Calculation of Animal Drug Exposure - Toxicokinetic AUC(0-24) Samples of drug in plasma of animal tox species Begin sampling after reaching repeated dose steady state Orange symbols are mean from n = 8 Mathematical fit to curves of oral absorption & elimination Measure area under curve for 0-24 hr = Drug Exposure Cmax = 10.8 mg/mL AUC(0-24) = 136 mg•hr/mL Dose = 50 mg/kg/day

Measured Drug Exposure in Rat Tox Studies Toxicokinetic Parameters in Multidose Oral Toxicity Studies Species Durat. Dose (mg/kg) Plasma Concentration (g/mL) AUC(0‑24) (g·hr/mL) Male Female Rat 2 Week Gavage 500 1250 2500 Diet 13.5 27.6 47.4 11.5 25.9 50.7 9.92 25.2 40.7 10.7 19.0 32.4 120.0 332.0 626.0 199.0 491.0 921.0 102.0 334.0 602.0 181.0 336.0 606.0

Animal Toxicology: Human Exposure Limits Are Set by Daily Drug Exposure (AUC0-24) RAT DOG MOUSE No-Effect Dose Limit: 200 mg•hr/mL – determines maximum permissible human exposure

PK Modeling of Drug in Human Plasma (daily dosing of 50 mg oral) Human Cmax Limit based on Animal Toxicology (Max no-effect dose AUC0-24) (mg/mL) AUC0-24

CYP2D6 Heterogeneity – Ca2+ Channel Blocker Smith & Jones (1999) Curr Opin Drug Discov Devl 2:33-41

Q: How to Predict Human Efficacious and Adverse Drug Doses Based on Animal Efficacy, Animal AEs and Human Pharmacokinetic Data?? A: Compare plasma drug Cmax obtained in animal pharmacology tests using a Napiergram to human Pharmacokinetic Cmax data

“Napiergram” Named for John Napier of Merchistoun (aka Marvelous Merchiston, Scotland) Inventor of Napier’s bones (slide rule), popularization of logarithms and the decimal point Also: used a pet black rooster to tell fortunes and devine truths Napiergram: graphic comparison of log10 unbound plasma drug concentrations associated with pharmacology and with safety concerns John Napier (1550-1617)

From Dose:Response experiments: Obtain ED5, ED50, ED95

Transform Pharmacology from ED50 to EC50

Napiergram: Many Pharmacology Datasets – Animal Cmax for doses with 5% 50% & 95% effect Cmax = 2,500 nM or 0.5 mg/mL (unbound) Cmin = 125 nM or 0.026 mg/mL (unbound)

PK Modeling of Drug in Human Plasma (daily dosing of 50 mg oral) (mg/mL) Animal Adverse Limit (EC50 for ataxia) Cmax (hi dose) Cmax (mid dose) Actual Human PK – mid dose Cmax (low dose) EC20 for Efficacy in Animal Model

Phase 2 Dose Selection (final chapter) Requires Deliberation from team of experts: Animal tox, Pharmacokinetics, PK/PD modeling, Clinical research, Preclinical pharmacology, (Biomarkers) Who pays the clinical trial bills? Clinical Research Despite planning, dosage and regimen often are readjusted during Phase 2 (toleration, efficacy or new safety findings) Dosages MUST continue to include both low (ineffective) and maximal tolerated dosages to provide basis for FDA approval Dose toleration may vary between healthy volunteers and patients with serious disease

SUMMARY: Preclinical Studies to Determine Phase 2 Dose Selection In vitro and in vivo animal pharmacology – target Cmax for therapy and adverse effects Animal toxicology & toxicokinetic studies – determines maximal human drug exposure (AUC0-24) Phase 1 Clinical trials determines human pharmacokinetics & drug exposure Napiergram – allows consideration of Cmax from multiple animal datasets & compare to human PK Phase 2 dose adjustment is common!

Example “Drug Killer” Problems Poor Oral Absorption (F < 25%) Poor Aqueous Solubility Poor Elimination Kinetics (t1/2 < 4 hr or t1/2 > 36 hr) Nonlinear Elimination Kinetics (e.g. blocked clearance at high doses) Extensive metabolism to active or toxic compound Excessive plasma protein binding (> 99%) Metabolism by variable CyP450 (CYP2D6, CYP2C19) Cardiac Q-T interval prolongation (hERG channel block) Genotoxic compound (Ames positive) Hepatic toxicity PK Tox