Support From Discovery to Registration

1

SUPPORT FROM DISCOVERY TO REGISTRATION

Successful NDACandidate Selection

to Early Clinical Trials

Hit to Candidate Selection

Late-

Stag

e Non

clinic

al

and C

linica

l Dev

elopm

ent

Summary of DMPK at Charles River

2

SUMMARY OF DMPK AT CHARLES RIVER

In Vitro Services• Metabolic stability

- Whole cells and subcellular fractions

- Human and preclinical species

• Metabolite profiling

- Accurate mass MS and nuclear magnetic resonance (NMR)*

• Drug absorption

- Intestinal and dermal

• Drug-drug interactions

- Induction and inhibition

• Plasma protein binding

• Assessment of reactive metabolite formation

In Vivo Services• PK assessment

- Naïve and non-naïve animals

- Rodents and large animals

- HPV cannulated models

• Mass balance

- Intact and bile duct

• Tissue distribution

- QWBA, microautoradiography, and dissect and burn

- Mass spectrometry imaging

• Metabolite profiling

- Accurate mass MS and NMR*

• Milk and placental transfer

*In partnership with a local university.

Program Considerations

3

What DMPK Activities Would You Recommend During Screening to Aid Candidate Selection?

0 4 8 12 16 20 24

350

300

250

200

150

100

50

0

PROGRAM CONSIDERATIONS

In vitro metabolic stability (microsomes and/or hepatocytes)

PK/formulation assessment (rodent and nonrodent)

Intestinal absorption (Caco-2 cell monolayers)

CYP inhibition (CYP3A4)

0.00 0.501.00 1.502.002.50 3.003.504.004.505.00

Metabolic Stability

Ln%

of

Par

ent

Time (min)

0 2 4 6 8 10 12 14 16

0

5

10

15

75

100

125

δAU

C

1+ (l/Ki) 0 25 50 75 100

Impact of fm on δAUC

fm=1

fm=0.9

fm=0.75fm=0.5fm=0.1

Transwellinsert

Cellmonolayer

Permeablemembrane

Apicalchamber

Basolateralchamber

Vectorial transport

A

B

Are Any ADME Studies Required Prior to FTIM?ICH M3 R2 Guidelines states, “plasma protein binding data for animals and humans… in the species used for repeated-dose toxicity studies generally should be evaluated before initiating human clinical trials.”

We can assess plasma protein binding by:

• Equilibrium dialysis

• Ultrafiltration

• Ultracentrifugation

• In humans and preclinical toxicology species

Any Other ADME Studies Recommended Prior to FTIM?ICH Guideline M3 (R2) additionally recommends that in vitro metabolic data from humans and intended repeat dose toxicity species should be available prior to first in man studies.

• Radiolabelled and nonradiolabelled test items can be incubated with isolated enzymes, subcellular fractions or hepatocytes.

• Metabolite profiling and identification is conducted by accurate mass MS (Waters Q-Tof, Shimadzu IT-Tof, Waters Synapt G2-S) with radiodetection.

PROGRAM CONSIDERATIONS

When Should We Consider Conducting In Vivo ADME Studies?ICH Guideline M3 (R2) recommends that ADME data are available in preclinical toxicology species before widespread exposure of human subjects to the drug (i.e., generally before Phase III clinical trials).

• An abbreviated study design (in rodents) can be used to provide an early understanding of rates and routes of metabolism and excretion and the potential for tissue accumulation.

• Full mass balance studies in both rodents and large animals can be undertaken to support later stage clinical trials.

168 h 10 min168 h, 1 albino & 1 pigmented

for QWBA4, 24 & 72 h

Oral Dosing

IV Dosing

PROGRAM CONSIDERATIONS

When Is It Appropriate to Address Metabolite Profiling? ICH Guideline M3 (R2)

• Nonclinical characterization of a human metabolite(s) is only warranted when that metabolite(s) is observed at exposures greater than 10% of total drug-related exposure and at significantly greater levels in humans than the maximum exposure seen in the toxicity studies.

FDA Guidance: Safety Testing of Drug Metabolites

• Generally, metabolites identified only in human plasma or metabolites present at disproportionately higher levels in humans than in any of the animal test species should be considered for safety assessment.

• Human metabolites that can raise a safety concern are those formed at greater than 10% of parent drug systemic exposure at steady state.

Metabolites in Safety Testing StrategyRadiolabelled SamplesNonradiolabelled Samples

Obtain early indication of biotransformation pathways and

metabolite quantities

Structural alerts Reactive metabolites

Radiolablled in vitro metabolism

Define biotransformation pathway and relative quantities

of metabolites in toxicology species and man

Active metabolitesUnique/disproportionate

metabolites

Measure key metabolites with validated methods and

reference standards

Early clinincal studies

Toxicology species radio ADME

Human radiolabelled study

Later phase clinical studies

Longer term toxicology studies

Discovery PK samples

Nonradiolabelled in vitro species comparison

Early toxicology samples

Establish route for production of reference standards for metabolites of concern. Develop and validate robust analytical methods for parent and key metabolites.

What Useful Information Would I Gain from a Tissue Distribution Study? • Tissue half-life

• Target organ penetration (e.g., brain)

• Dosimetry calculation for clinical mass balance study

• Understanding of toxicology findings

PROGRAM CONSIDERATIONS

Dissect and Quantify• Analysis by LC-MS/MS or

radioquantification

Quantitative Whole Body Autoradiography (QWBA)

Microautoradiography (MARG)• Both require radiolabelled

materials

Mass Spectrometry Imaging (MSI)• Can be conducted label

free (in collaboration with Imabiotech)

Additional Studies Required When Considering Women of Childbearing Age To support/validate pre- and postnatal toxicity studies:

• Placental transfer

- Conducted by QWBA or dissect and analyze

• Milk transfer

- Assess milk: plasma ratio of total radioactivity, prediction of exposure in suckling young to drug-related material

Assessment of Drug-Drug Interactions FDA (Draft Guidance 2012)

• Drug Interaction Studies — Study Design, Data Analysis, Implications for Dosing and Labelling Recommendations

EMEA (2013)

• Guideline on the Investigation of Drug Interactions

PROGRAM CONSIDERATIONS

• Activity and mRNA endpoints

Induction Assays

Inhibition Assays

• Phase I and Phase II metabolism

• Direct and time dependent inhibition (TDI)

• Transporters

• MDR1 and BCRP

Additional Program Considerations In support of toxicology or for mechanistic investigations:

• Ex vivo enzyme induction studies

• In vitro and in vivo investigation of reactive metabolite formation

• Bile duct and hepatic portal vein (HPV) cannulation studies in small and large animals

DMPK at Charles River

PROGRAM CONSIDERATIONS

Reno, Nevada

Discovery PK (biologics)

Wilmington, Massachusetts

Discovery PK (small molecule)

Edinburgh, Scotland

Development DMPK (small molecule & biologics)

Discovery DMPK (small molecule & biologics)