2013南京国际药代会议—从临床前到临床的药代、药动

研究与实例探讨

6/9/2012 1

近年来药代研究新进展Recent Advances in Drug DMPK Research

Doing the Right Thing vs. Doing Things the Right Way

June 21-22, 2013

Seminar Outline

• Recent Advances in Instrumentations– All instrumentation companies have made newer and

advanced instrumentation for DMPK use in general

• Recent Advances in DMPK Regulations– US FDA issued Drug-Drug Interaction Guidance for Industry

• Need to know metabolic enzyme phenotyping, inhibition, and• Need to know metabolic enzyme phenotyping, inhibition, andinduction

• Need to know the interaction with market drugs in the clinical trial• Need to provide drug transporter information – substrate and/or

inhibitors

– CFDA is preparing for the updated preclinical DMPKguidance for industry

– EMA published other related guidance papers

• Recent Advances in DMPK Approaches

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40 年前的参考资料

6/9/2012 3

6/9/2012 6CEN February 4, 2013, Page 15-17

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Interesting Summary

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想想看：病患的选择在那里？

• 生，老，病，死？

• 万一不幸，病患的选择在那里？

• 医护人员，医院，政府的责任？

• 药厂有赚到钱了吗？

• 新药的开发有必要吗？

• 还有那些新药可开发？

• 人在天堂，钱在银行？

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Orphan (Rare) DiseasesCENews, May 13, 2013, page 10-23

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Elaprase forHunter

SyndromeOne of the

mostexpensivedrug in the

world

Drug Metabolism Studies in New DrugDiscovery and Development Process

Clinical developmentPreclinicaldeveopment

Phase I Phase II Phase IIILeadselection

Discovery

Optimization

NDAPreClinical Candidates

10,000 – 30,000 1250 5-10 2-5 2

IND NDA

Approval

In vitro speciescomparison

& in vitro-in vivoCorrelation

(labeled or unlabeled)

Soft-spot & reactivemetabolite analysis

Unlabeledcompounds

PreClinical Candidates

ADME in animalsusing radiolabel

Issues driven studies eg.polymorphic enzyme &

toxic metabolites

14C-Human AMEreaction phenotyping, &

issues-driven studies

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Discovery Screening WorkflowHighlighting DMPK Contributions

Biology in vitroefficacy & potency

Medicinal ChemistrySAR (Optimization)

Tier 1 ADME Assays

Rat IV/PO PK

Tier 2 ADME Assays

Single conc./time point screening•Metabolic stability•CYP inhibition•Gene regulation assays•Plasma protein binding assays

Full in vitro studies•Metabolic stability (Clint) for human PK/dose pred.•CYP inhibition (IC50, Ki, KI and Kinact) for DDI Pred.•Hepatocyte induction and reaction phenotyping

PK/F

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Tier 2 ADME Assays

In Vivo Efficacy

Rat ,Non-Rodent Tox

Multiple Species PK

In Vivo ADME

Preclinical Drug Candidate

IND Submission

•Hepatocyte induction and reaction phenotyping•Plasma protein binding•Permeability/transporter (Caco and recomb. Cells)•Covalent binding•Metabolite profiling and ID (microsomes/heptocytes)

PK and F (PD/SA species)Dose proportionalityPK/PD relationship

Mass balanceFraction of ClearanceDose proportionalityMetabolite ID and profiling

NOEL/NOAELSafetyMargin

Mechanisms of Drug Hepatotoxicity

Drug Metabolism

Elimination(Detoxification)

Stable Metabolites

ReactiveMetabolites

Covalent binding

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Covalent bindingEnzyme/protein inactivation

Altered Ca2+ homeostasisOxidative stress

Lipid peroxidationGSH depletion

Mitochondria toxicityATP depletion

Metabolic alteration

HepatotoxicityApoptosisNecrosis

Immune toxicity

Recent Advances in Instrumentation

• Mainly in mass spectrometry (MS):– Metabolite ID by MS: forty years of evolution:

Pat Wright, Xenobiotica, 2011; 41 (8), 670-686

– High Resolution Mass Spectrometer: LTQ Orbitrap– IonTraps, TOF/TOF, MALDI TOF/TOF– Others– Others

• Related software in DMPK MS applications– Mass defect: Zhang H, Zhang D, Ray K. J. Mass Spectrum. 2003; 38:

1110

– Background subtraction and noise reductionalgorithm (BgS-NoRA): Zhu Peijuan, S. Chowdhury; Rapid Commun.Mass Spectrum. 2009:23: 1563-1572

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Recent Advances in DMPK Regulations

• Recent Advances in DMPK Regulations– MIST Guidance – final version published in 2008; 10% of parent– ICH Guidance 2009; 10% of Total– US FDA Drug-Drug Interaction Guidance for Industry - 2012

• Need to know metabolic enzyme phenotyping, inhibition, and induction• Need to know the interaction with market drugs in the clinical trial• Need to provide drug transporter information – substrate and/or

inhibitors• FDA Guideline on drug-drug interaction studies (2012)• FDA Guideline on drug-drug interaction studies (2012)

– EMA Guidelines on the following:• investigations of Drug Interactions (2012)• Reflection paper on the data requirement for intravenous liposomal

products developed with reference to an innovator liposomal product(adopted by CHMP of Rev. 02, 21 February 2013)

– CFDA is preparing for the updated preclinical DMPK guidance forindustry

• 药物非临床药代动力学研究 – 技术指导原则• work in progress and will be published soon

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Preclinical and Clinical Applicationof Low Energy Radioisotopes

in Life Science R&Din Life Science R&D

A very useful tool for life science R&D

• Isotope-labeling (stable isotope, radio-labeling) is a very useful tool for various lifescience R&D:

– Pharmaceutical– Pharmaceutical

– Animal Health

– Agricultural

– Biotechnology

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Instrumentation for MeasuringRadioactivity

ß-RAM

vARC online ® System

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Biological Sample Oxidizer

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Organic compounds (containing C, H, O)can be oxidized to carbon dioxide andwater

CxHyOz + O2 → xCO2 + zH2O14Cx

3HyOz + O2 → x14CO2 + z3H2OHeteroatoms –

N, S, X (Cl, Br, F)Inorganic metals –

Pb, Cu, Fe, Mg, Mn, etc.

Radioactivity Used in ADME Studies

• Small animals – rat, mouse 10-40 µCi/animal

3-5 animals/gender

iv, po (usually 7 days)

Urine, bile, feces,blood/plasma, tissues,

• Human – [99%] 50 -125 µCi/subject

Usually male subjects only

3-6 subjects/study;

po or iv

Blood/plasma, urine, fecesblood/plasma, tissues,

• Larger animals – dog,monkey 10-20 µCi/Kg

3-5 animals/gender

IV, po (usually 7 days)

Urine, feces, blood/plasma

Blood/plasma, urine, feces

7-28 days collection based onthe release criteria

Microdosing (<1 µCi, 100 µg) Requires AMS detector

Special purpose

Limited information

<1% of study applied

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Metabolite Profiling and Identification

StopFlow® System

vARC Dynamic flow

Full Scan MS

0 5 10 15 20 25 30 35 40 45 50 55 60 65 70 75 80

LumaPlate TopCountQuantitative

B

COrbiTrap/Q-TOF

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Product Ion Scan –Accurate Mass

ß-RAM

96 Deep Well Plate

Nanospray LC/MS

A

HPLC Radio-chromatogramsDog and Rat Plasma – 3 hour

time pointDog

22

Rat

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0

10

20

30

0 5 10 15 20 25 30 35 40 45 50 55 60 65 70

0

300

600

900

0 5 10 15 20 25 30 35 40 45 50 55 60 65 70

Monkey Plasma and Urine RadioprofilesGender Comparisons

1 Hr – Male Plasma6 Hr – Male Urine

23

0 5 10 15 20 25 30 35 40 45 50 55 60 65 70

0

10

20

30

0 5 10 15 20 25 30 35 40 45 50 55 60 65 70

0 5 10 15 20 25 30 35 40 45 50 55 60 65 70

0

400

800

1,200

0 5 10 15 20 25 30 35 40 45 50 55 60 65 70

1 Hr – Female Plasma 6 Hr – Female Urine

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Recent Advances in DMPK Approaches

• Recent Advances in DMPK Instrumentations

• Recent Advances in DMPK Regulations

• Recent Advances in DMPK Approaches– With current instrumentation and regulations, any

different/new approaches?– With current instrumentation and regulations, any

different/new approaches?– Consider to apply to clinical and difficult to conduct cases– Combination of “New” and “Old” techniques to become new

approaches in DMPK– Using animal radiolabeled samples as radiocalibrator for human

metabolite searching and “quantitation”– HPLC- HRMS – AMS– MS as a surgical tool? (CEN, Feb 4, 2013) – molecular signature

during brain surgery

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MS as a Surgical Tool?

• MRI 3-D Brain Image

• Differentiate normal vs.cancer tissues

• Using Desorption Electrospray Desorption Electrospray

ionization (DESI) MS

Rapid Evaporationionization MS (REIMS)

• Acquire molecularsignatures to accuratelydiagnose tissues removed

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HPLC-HRMS-AMS in DMPK

• Early investigation inhuman?– Standard human AME

use 50-125 uCi/subject

– Micro-dosing,

• AMS (Accelerator massspectrometry)– Dosimetry to support

human studies notrequired– Micro-dosing,

radiotracer with <1 uCi[14C-Drug] – 100-400 nCi

• Microdosing, typically,plasma [14C] = 0.1-1dpm, how to measure?

• AMS cost? Benefits?

required

– Enables multiple dosesover time for steadystate metabolism

– GMP material notrequired

– Micro Tracer IV studiesfor absolute “F”

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HPLC-HRMS-AMS in DMPK

• Microdosing - Radiotracer– with <1 uCi [14C-Drug] – 100-

400 nCi, similar to our bodybackground value

– With preclinical ADME datafor clear metabolic profilesand animal metabolism data

• Excretion and mass balancein human– Very low exposure after PO

administration– Specific tissue level of 14C -

too low for LSC?

• Low absorptionand animal metabolism data

– With HRMS to obtain“complete” information formhuman

– Early understanding toxspecies vs. human profilesand metabolite information

– Efforts vs. Outcomes?

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• Low absorption– Intra-nasal administration– Dermatology products

• Metabolite profile– MIST – steady state AUC

• IV microdosing radiotracerPK with regular oral FIHstudy

Levels of Radio-Detection

10-9 g

10-12 g

nanograms

picograms

Liquid Scintillation Counting(β-decay of 14C atom)

Mass Spectrometry

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10-15 g

10-18 g

10-21 g

femtograms

attograms

zeptograms

Accelerator MassSpectrometry

(counting 12, 13, 14C atoms)

= 8.3 yr

1 DPM contains 7.2 fmol 14C or 4.35 billion 14Catoms – plenty for quantitation 29

Dose Normalized Mean Plasma Drug Concentration-Time ProfileFollowing Oral and IV Administration.

IV dose; 40 ug [14C]drug (normalized)Measured by LC-AMS15 min infusion of [14C]drug@ 1 h after oral dose

Abs BA (F) = 50%90% CI: 48 - 53%

Oral dose; 5 mg SaxagliptinMeasured by LC-MS/MS

90% CI: 48 - 53%%CV: 6.6%

Xu, Dueker et. Al. Bioanalysis Overcoming bioanalytical challenges in an Onglyza intravenous[14C]microdose absolute bioavailability study with accelerator MS. Bioanalysis. 4:1855-70. 2012 30

Disposition, Metabolismand Mass Balance of

[14C]Apremilast FollowingOral Administration

Clinical AME Case Studies

OCH3

O O

Absorption, metabolism andexcretion of [14C]pomalidomide

in human Following OralAdministration

N

NH

O

O

O

S

O

O

O

H

M. Hoffman (et al., from Celgene) andZ. Gu, D. Heller, Hao Feng, (from XBL)Xenobiotica, 2011; 41 (12): 1063-1075

N

NH2

O

O

NH

O

O

M. Hoffman (et al., from Celgene) andH. Kim-Kang (from XBL)

Cancer Chemother Pharmcol, Dec. 2012

Thank Youfor your participation