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)