The XIII International Congress of Toxicology | July 2, 2013, Seoul, Korea 1

Predicting pyrethroids target tissue 

exposure across ages

Application of in vitro-in vivo extrapolation 

and physiologically based pharmacokinetic 

modeling

Miyoung Yoon and Harvey J. Clewell

The Hamner Institutes for Health Sciences

Research Triangle Park, NC USA

1

Exclusive Property of the Council for the Advancement of 

Pyrethroid Human Risk Assessment, LLC (CAPHRA)
Exclusive Property of the Council for the Advancement of 

Pyrethroid Human Risk Assessment, LLC (CAPHRA)

Hypothesis #2:

PBPK models can be used to predict 

target tissue levels in human adults 

and children 

2
The Hamner Institutes for Health Sciences | August 5, 2010 3

1yr 5 yr 10 yr Adult

Age

Brain Cmax (ng/ml) 

Provide a tool to predict target tissue exposure in children and 

adults 

Deltamethrin, 0.005 mg/kg

The objective

3

Exclusive Property of the Council for the Advancement of 

Pyrethroid Human Risk Assessment, LLC (CAPHRA) 
The Hamner Institutes for Health Sciences | August 5, 2010 44

Our approach

In vitro to in vivo extrapolation (IVIVE)

Predicting in vivo kinetics based on in 

vitro/in silico metabolism and other 

kinetic data

Exclusive Property of the Council for the Advancement of 

Pyrethroid Human Risk Assessment, LLC (CAPHRA) 
The Hamner Institutes for Health Sciences | August 5, 2010 5

The framework

Human Early Age PBPK model

(Hamner)

Human 

metabolism data 

in vitro (LFR)

Rat metabolism 

data in vitro 

(LFR)

Rat Early Age PBPK model

(Refinement of Tornero-Velez et 

al. model)

(Hamner)

IVIVE

In vivo PK data in 

growing rats

(UGa)

IVIVE approach 

validation

Human enzyme 

ontogeny 

(MCoW/Hamner

/Published data)

IVIVE

Prediction of target tissue dose

in the age of concern

at environmentally relevant 

exposure 

Application 

of the 

approach

Exclusive Property of the Council for the Advancement of 

Pyrethroid Human Risk Assessment, LLC (CAPHRA) 5
The XIII International Congress of Toxicology | July 2, 2013, Seoul, Korea 6

Target tissue dosimetry in AOP-based risk 

assessment

• Internal exposure at the target site of action is critical in 

defining accurate concentration-effect relationship for 

early cellular events in Adverse Outcome Pathway (AOP) 

framework

• Physiologically based pharmacokinetic (PBPK) modeling 

provides a tool to predict target tissue exposure and link 

it to estimated or observed effect concentrations from 

toxicity studies to inform human safe exposure

6

Exclusive Property of the Council for the Advancement of 

Pyrethroid Human Risk Assessment, LLC (CAPHRA)
Potential impacts of age-specific pharmacokinetics on 

tissue dosimetry and sensitivity

• Rapidly changing physiology and biochemical processes 

are expected to affect chemical kinetics in the growing 

body

• Resulting differences in target tissue exposure may lead 

to age-associated sensitivity to chemical effects during 

rapid growth period

7

Exclusive Property of the Council for the Advancement of 

Pyrethroid Human Risk Assessment, LLC (CAPHRA)
The XIII International Congress of Toxicology | July 2, 2013, Seoul, Korea 8

PBPK modeling to predict age-specific tissue dosimetry

PBPK models serve as a platform to coherently integrate diverse, 

independent data sets and provide predictive/extrapolation 

power

• Parameters (inputs) for PBPK models:

‒ Exposure (age-dependent; chemical-dependent)

‒ Physiology such as body weight, tissue volumes, blood 

flows etc. (age-dependent; chemical-independent)

‒ Biochemical processes such as metabolism, 

partitioning, protein binding etc. (age-dependent; 

chemical-dependent)

8

Exclusive Property of the Council for the Advancement of 

Pyrethroid Human Risk Assessment, LLC (CAPHRA)
IVIVE-PBPK modeling: Taking into account key 

determinants of early life dosimetry

Age-specific 

internal dose

Life stage PBPK MODEL

Age-dependent physiological changes

Age-dependent biochemical changes

Age-appropriate exposure 

(CARES or SHEDS)

Requires in vitro 

data-based 

parameterization

Exclusive Property of the Council for the Advancement of 

Pyrethroid Human Risk Assessment, LLC (CAPHRA)
The XIII International Congress of Toxicology | July 2, 2013, Seoul, Korea 10

• The bottom-up PBPK modeling concept has been used in 

environmental chemical risk assessment started with volatiles 

and later on for other compounds including pesticides 

(Clewell, 1993; Clewell and Andersen, 1996; Kedderis and 

Lipscomb, 2001; Hinderliter et al., 2011; Yoon et al., 2014) 

• The approach is widely accepted for drugs because of general 

PBPK modeling tools for drugs (Simcyp) and rapid advances in 

in vitro metabolism technologies

• Pediatric applications, initially used for pharmaceuticals, have 

successfully been adopted for environmental chemicals in in 

vitro-based risk assessment initiatives (Nong et al., 2005; Price 

et al., 2011; Wetmore et al., 2014)

10

The bottom-up (IVIVE) PBPK modeling approach is 

an already widely accepted concept

Exclusive Property of the Council for the Advancement of 

Pyrethroid Human Risk Assessment, LLC (CAPHRA)
The XIII International Congress of Toxicology | July 2, 2013, Seoul, Korea 11

IVIVE-PBPK application to pediatrics: A source 

to outcome modeling for chlolopyrifos

Copyright 2013 by The Hamner Institutes for Health Sciences. May not be reproduced without permission

-2 -1 0 1 2

10- 4

10- 3

10- 2

10- 1

100

101

Chlorpyrifos

Chlorpyrifos oxon

}3 Year Olds

}30 Year Olds

}3 Year Olds

}30 Year Olds

Z-Score (3-97th Percentiles)

Blood Concentration (ng/L)

-2 -1 0 1 2 10- 5

10- 4

10- 3

10- 2

}3 Year Olds

}30 Year Olds

Z-Score (3-97th Percentiles)

RBC AChE Inhibition (% basal)

-2 -1 0 1 2 10- 6

10- 5

10- 4

10- 3 }3 Year Olds

}30 Year Olds

Brain AChE Inhibition (% basal)

Z-Score (3-97th Percentiles)

CARES LifeLine Daily LifeLine Seasonal

Price et al., 2011, Regulatory Toxicology and Pharmacology Exclusive Property of the Council for the Advancement of 

Pyrethroid Human Risk Assessment, LLC (CAPHRA)
The XIII International Congress of Toxicology | July 2, 2013, Seoul, Korea 12

Chemical Median Css for 

Healthy

Population

Median Css

for Most 

Sensitive

Most 

Sensitive

Chemical-Specific 

Pharmacokinetic 

Adjustment 

Factors (PK-AF)

% Contribution of 

Metabolism to 

Average PK-AF

Acetochlor 0.026 0.063 Pediatric 2.5 80

Azoxystrobin 0.099 0.22 Pediatric 2.2 86

Bensulide 0.241 0.35 Pediatric 1.5 66

Carbaryl 0.043 0.17 Pediatric 4.0 88

Difenoconazole 0.201 0.30 Renal 

Insufficiency 1.5 58

Fludioxonil 0.38 1.45 Pediatric 3.0 86

Haloperidol 0.029 0.04 Pediatric 1.5 83

Lovastatin 0.001 0.003 Pediatric 2.1 92

Tebupirimfos 0.107 0.16 Renal 

Insufficiency 1.5 20

12

IVIVE-PBPK application to pediatrics: High-throughput 

IVIVE to address susceptible subpopulations

Copyright 2013 by The Hamner Institutes for Health Sciences. May not be reproduced without permission

Adapted from Table 4 in Wetmore et al., 2014, Toxicological Sciences

Exclusive Property of the Council for the Advancement of 

Pyrethroid Human Risk Assessment, LLC (CAPHRA)
Color coding used for this presentation

IVIVE-PBPK modeling – background

Human IVIVE-PBPK – general platform

Rat IVIVE-PBPK – case study

Human IVIVE-PBPK – case study

Concepts Application Proof of concept 

case study

Exclusive Property of the Council for the Advancement of Pyrethroid Human Risk Assessment, LLC (CAPHRA)
In vitro to in vivo extrapolation-based PBPK 

modeling for early life dosimetry

Human

in vitro metabolism 

data: age-specific

Rat

in vitro metabolism 

data: age-specific

Proof of concept case studies Application of the approach 14

Rat

Adult PBPK 

Model

Human

Adult PBPK 

Model

Human

Early Life PBPK 

Model

Rat

Early Life PBPK 

Model

Exclusive Property of the Council for the Advancement of Pyrethroid Human Risk Assessment, LLC (CAPHRA)
Human

Age-dependent 

PBPK Model

Human age-specific 

parameters in vitro

Human Growth Physiology

Human Age-relevant 

Exposure

(CARES or SHEDS)

Rat

Age-dependent 

PBPK Model

Rat Growth Physiology

Rat age-specific 

parameters in vitro

IVIVE

Step 1: Rat modeling for 

case compounds Validation 

of the concept

IVIVE

Step 2: Human modeling for case 

compounds Application of the 

approach to humans

General PBPK model 

for pyrethroids in 

humans across ages

Step 4: Exercising the general 

modeling platform for 

pyrethroid risk assessment in 

different ages of humans

Building a general model platform for 

pyrethroids for risk assessment

15

Step 3: Read across strategies for model parameterization built 

upon the experience with case studies

Targeted in vitro studies for 

compound specific key 

model parameters

Exclusive Property of the Council for the Advancement of Pyrethroid Human Risk Assessment, LLC (CAPHRA)
Targeted in vitro experiments for pyrethroid IVIVE

16

Types of 

studies Rat – case study Human - case study Human - general model Purpose*

Metabolism In vitro

Microsomes and 

cytosol; age-specific 

donors (LFR)

Microsomes and cytosol; agesepecific donors- DLM only;

Recombinant CYP/CES

enzymes (LFR)

Recombinant CYP/CES

enzymes (LFR) P

Enzyme 

ontogeny of 

expression

In vitro;

Published

database

n/a

Published database#;

CAPHRA data - CESs, CYP2C8, 

1A2 (MCoW; Hamner)

Published database#;

CAPHRA data - CESs, CYP2C8, 

1A2 (MCoW; Hamner)

P

Partition 

coefficient In vivo

Steady-state PK in 

young and adult rats 

(UGa)

Adopt rat parameters

Adopt rat parameters

Read-across based on 

underlying biology

P

Plasma 

protein 

binding

In vitro Plasma from agespecific donors (UGa)

Plasma from age-specific 

donors (UGa)

Plasma from adults;

Read-across by QSAR P

PK time 

course

In vivo PK studies in young 

and adult rats (UGa) n/a n/a P;V

* P indicates that the purpose of the study is for model parameterization and V for model validation

# The published database include those are used in Simcyp and those are available from a series of publications from 

Dr. Ron Hines lab. 

Exclusive Property of the Council for the Advancement of Pyrethroid Human Risk Assessment, LLC (CAPHRA)
Major challenges in IVIVE for human early ages 

for environmental chemicals

• Human in vivo data is necessarily limited (in contrast to 

pharmaceuticals), leading to a reliance on in vitro data. 

However:

– Pediatric tissues samples are very difficult to obtain 

and sample quality is generally uncertain

– Addressing human variability is challenging

• IVIVE with recombinant enzymes in conjunction with 

enzyme ontogeny of expression is selected as an 

alternative approach in CAPHRA

17

Exclusive Property of the Council for the Advancement of 

Pyrethroid Human Risk Assessment, LLC (CAPHRA)
In vitro metabolism studies for pyrethroid

IVIVE

18

Microsomes

Cytosol

Subcellular fractions

Age-specific liver donors

Human recombinant 

enzymes

Individual CYP or 

CES enzymes

CESs

CYPs + CESs

Recombinant enzymes

Rat Human

Exclusive Property of the Council for the Advancement of 

Pyrethroid Human Risk Assessment, LLC (CAPHRA)
The XIII International Congress of Toxicology | July 2, 2013, Seoul, Korea 19

Biological scaling is necessary in using in 

vitro data

• Extrapolation of in vitro metabolic parameters to in 

vivo systems requires:

– Relating the total amount of enzyme present in 

the system (capacity of metabolism, Vmax)

– Relating the available substrate concentration for 

enzyme reaction (affinity of metabolism, Km)

19

Exclusive Property of the Council for the Advancement of 

Pyrethroid Human Risk Assessment, LLC (CAPHRA)
Biological scaling for different in vitro systems

System Vmax or Clint in the 

System

Scaling factors

to whole liver

Microsomes nmol/min

/mg protein MPPGL x Liver weight

Cytosol nmol/min

/mg protein CPPGL x Liver weight

Expressed 

Enzyme

μl/min

/pmol enzyme

(ISEF xEnzabundance) x MPPGL x 

Liver weight

Liver nmol/min/g liver Liver weight

Whole Body nmol/min/whole liver

20

Exclusive Property of the Council for the Advancement of Yoon et al., 2012

Pyrethroid Human Risk Assessment, LLC (CAPHRA)
The XIII International Congress of Toxicology | July 2, 2013, Seoul, Korea 21

Human

Recombinant CYPs

and CESs with 

enzyme ontogeny 

data

Application and 

validation of the 

alternative IVIVE

approach

Human

Subcellular 

fractions from liver 

tissue bank with 

different ages

Application of the 

IVIVE approach to 

humans

Rat 

Subcellular 

fractions from 

young and adult 

rats 

Validation of IVIVE

approach for 

model 

parameterization

Deltamethrin modeling as a proof of concept

21

Exclusive Property of the Council for the Advancement of 

Pyrethroid Human Risk Assessment, LLC (CAPHRA)
The XIII International Congress of Toxicology | July 2, 2013, Seoul, Korea 22

Post-proof of concept IVIVE work 

flow (recombinant enzyme IVIVE)

22

Screening & 

Measuring

Metabolic rates for each 

active CYP or CES

(CAPHRA)

Biological scaling

Average adult Clints for 

each active enzyme

(CAPHRA + Published 

database)

Extrapolation 

across ages

Incorporating ontogeny 

of enzyme abundance

Distribution of agespecific Clints for each 

enzyme

(CAPHRA + Published 

database)

Integration with 

PBPK modeling

Distribution of agespecific total hepatic 

clearance for a given 

pyrethroid

(CAPHRA) 

Exclusive Property of the Council for the Advancement of 

Pyrethroid Human Risk Assessment, LLC (CAPHRA)
IVIVE with recombinant enzymes

A bottom-up modeling approach used by 

SimCyp for conducting population based 

PBPK modeling for diverse human 

populations including pediatrics

23

(Rostami-Hodjegan and Tucker, 2007)

Hepatic CYP Isozyme Abundance in Healthy 

Adults (% of Total)

Clintin vivo = ClintrEnz x EnzAbundance

x Scaling factor

ClintrEnz determined in recombinant 

CYPs and CESs

CYP and CES enzyme abundance in population

Screening and Measuring

Biological Scaling

Exclusive Property of the Council for the Advancement of Pyrethroid Human Risk Assessment, LLC (CAPHRA)
The XIII International Congress of Toxicology | July 2, 2013, Seoul, Korea 24

Recombinant enzyme IVIVE for pyrethroids

24

(Hines et al., submitted to Tox Sci)

• Collecting data for ontogeny 

and biological scaling factors 

for CESs and CYPs that are 

important for pyrethroids, 

but are currently not 

available in Simcyp

• Metabolism assays to 

determine pyrethroid

clearance rates for each 

enzyme that is active for the 

given pyrethroid

(Song et al., in preparation)

CES ontogeny

CYP2C8 ontogeny

Exclusive Property of the Council for the Advancement of 

Pyrethroid Human Risk Assessment, LLC (CAPHRA)
The XIII International Congress of Toxicology | July 2, 2013, Seoul, Korea 25

Characterizing human variability in metabolism 

based on in vitro

25

Inter-individual variations in expression levels (abundance) of each enzyme result in 

population variability in the given pyrethroid clearance

Individual 1

Individual 2

Individual 3

Individual 4

Individual i

Clint for each 

involved enzyme

Abundance of each involved 

enzyme in each individual

Clint_in vivo

20 year old

Clint_in vivo

1 year old

Extrapolation across Ages

Exclusive Property of the Council for the Advancement of 

Pyrethroid Human Risk Assessment, LLC (CAPHRA)
26

Subpopulation-based in vitro-to-in vivo extrapolation

CYP1A2

CYP2D6

CYP2C8

CYP2E1 

UGT1A4

CYP3A5

CYP3A4

CYP2C19

UGT1A1

CYP2C9

CYP2B6

UGT2B7

Primary Hepatocytes

-5

-4

-3

-2

-1

0

1

2

3

0 50 100 150

Ln Conc (uM)

Time (min)

 

 

Hepatic Clearance

Cl in vitro Plasma Css

General Population

CYP1A2

CYP...

CYP3A4

UGT...

ClrCYP1A2

ClrCYP...

ClrCYP3A4

ClrUGT...

Plasma Css for:

Neonates

Northern 

Europeans

Asians

Children

And so on...

Hepatic Intrinsic Clearance

rCYP1A2

rCYP3A4

rCYP...

rUGT ...

Wetmore et al., 2014, slide courtesy of Dr. Barbara Wetmore

An example for early life IVIVE for environmental chemicals

Exclusive Property of the Council for the 

Advancement of Pyrethroid Human Risk 

Assessment, LLC (CAPHRA) 26
The XIII International Congress of Toxicology | July 2, 2013, Seoul, Korea 27

0 20 40 60 80

-4

-3

-2

-1

0

1

 CYP3A4

 CYP3A5

 CYP2C19

CYP2C8

CYP2C9

CYP1A2

CYP2C18

Time (min)

Ln Concentration (uM)

0 20 40 60 80

-0.4

-0.2

0.0

0.2

0.4

0.6

UGT1A4

Time (min)

Ln Concentration (

μM)

0 20 40 60 80

-10

-8

-6

-4

-2

0

2

CYP2E1

CYP2C9

CYP1A2

CYP2C19

CYP2B6

Time (min)

Ln Concentration (

μM)

No UGT metabolism 

detected

Carbaryl Difenoconazole

Recombinant enzyme clearance rates

(Wetmore et al., 2014)

An example for early life IVIVE for environmental chemicals

27

Exclusive Property of the Council for the Advancement of 

Pyrethroid Human Risk Assessment, LLC (CAPHRA)
The XIII International Congress of Toxicology | July 2, 2013, Seoul, Korea 28

Comparison of css values derived across multiple 

subpopulations

Upper 95th percentile Css

Carbaryl

Gen

Jap

Ntrn Eur

Diabetic

Renal Insuff.

6-20

2-6

0.5-20-0.5

0.01

0.1

1

Css at 1 μM (

μM)

(5th-95th %ile)

Subpopulation

(Age (yr) or Ethnic)

Subpopulation

(Age (yr) or Ethnic)

(Wetmore et al., 2014)

An example for early life IVIVE for environmental chemicals

28

Exclusive Property of the Council for the Advancement of 

Pyrethroid Human Risk Assessment, LLC (CAPHRA)
29

Evaluation of IVIVE

approach for developing 

age-specific PBPK models 

Deltamethrin PBPK 

model for growing 

rats

Rat 

Subcellular 

fractions from 

young and adult 

rats 

Validation of IVIVE

approach for 

model 

parameterization

Exclusive Property of the Council for the Advancement of 

Pyrethroid Human Risk Assessment, LLC (CAPHRA) 
The XIII International Congress of Toxicology | July 2, 2013, Seoul, Korea 30

Deltamethrin in vivo kinetic studies for model 

evaluation

• Plasma time course in adult rats (PND 90)

– single oral bolus dose of 0.05-5.0 mg DLM/kg in 5 ml corn oil 

(CO)/kg

– serial micro-blood samples taken arterial cannula for up to 96 

hours post dose

– 4-5 animals per timepoint

• Tissue-distribution study

– single oral bolus dose of 0.1, 0.25 or 0.5 mg DLM/kg in 5 ml corn 

oil (CO)/kg

– plasma and tissues were collected at terminal sacrifice

– plasma, brain and liver analyzed

• Plasma and tissue samples were analyzed by GC-MS

• Time-course data analyzed to estimate standard PK parameters 

30

Exclusive Property of the Council for the Advancement of 

Pyrethroid Human Risk Assessment, LLC (CAPHRA)
The XIII International Congress of Toxicology | July 2, 2013, Seoul, Korea 31

Plasma kinetics in adult rats

single oral dose of DLM (in 5 ml/kg corn oil) 

31

Exclusive Property of the Council for the Advancement of 

Pyrethroid Human Risk Assessment, LLC (CAPHRA)
The XIII International Congress of Toxicology | July 2, 2013, Seoul, Korea 32

Pharmacokinetics in adult rats

Dose mg/kg 5 1 0.5 0.1 0.05

Tmax min 420 360 420 420 300

Cmax ng/ml 337±126 94.7±51.8 49.4±18.4 10.4±10.1 4.7±2.6 

Cmax

Dose 67 95 99 104 94

Effective 

Half-life hr 3.9 4.5 3.7 4.1 3.0

AUC∞ ng*hr/ml 3069 937 521 148 61

AUC∞

Dose 614 937 1042 1476 1225

• Single oral dose of DLM in 5 ml/kg corn oil to PND90 Rats with serial blood 

sampling

• Linear plasma kinetics over the 100-fold range of doses (0.05 to 5 mg DLM/kg).

• Parallel plasma absorption, distribution and elimination profiles

• Peak concentrations (Cmax) and area under the concentration–time curve 

(AUC) increased proportionately with dose

• Absorption rate-limited kinetics

• Broad peaks with Tmax values of 300-420 minutes

• Bioavailability orally in corn oil is low (5.5 to 9.9 %)

32 Exclusive Property of the Council for the Advancement of Pyrethroid Human Risk Assessment, LLC (CAPHRA)
The XIII International Congress of Toxicology | July 2, 2013, Seoul, Korea 33

Comparison of Cmax in plasma and brain across age

Adult PND15 Adult PND15

Plasma Cmax (ng/g)

Brain Cmax (ng/g)

33

Exclusive Property of the Council for the Advancement of 

Pyrethroid Human Risk Assessment, LLC (CAPHRA)
The XIII International Congress of Toxicology | July 2, 2013, Seoul, Korea 34

Pharmacokinetics in adult and juvenile Rats

Plasma Brain

Dose 0.5 mg/kg 0.25 mg/kg 0.1 mg/kg 0.5 mg/kg 0.25 mg/kg 0.1 mg/kg

Cmax

(ng/g)

Adult 50.4±24.3 36.4 + 12 14.6±11.9 7.63 + 1.59 2.52 + 0.31 <LOQ

PND15 284 + 52 128 + 36 21 + 6.6 16.3 + 5.7 10.7 + 1.4 3.1 + 0.8 

AUC

(ng-hr/mL)

Adult 258 327 96 113 29 <LOQ

PND15 1172 858 148 144 95 42

• Single oral dose of DLM in 5 ml/kg corn oil to PND90 Rats with terminal blood 

and brain sampling

• Plasma and brain levels higher at PND15 that PND 90 for both plasma and brain 

for all doe levels.

• Cmax 2 to 5-fold

• Note: At lowest dose of 0.1 mg/kg, no DLM was detected in PND90 rats, 

indicating rapid and complete elimination. 

34

Exclusive Property of the Council for the Advancement of 

Pyrethroid Human Risk Assessment, LLC (CAPHRA)
The XIII International Congress of Toxicology | July 2, 2013, Seoul, Korea 35

Deltamethrin in vitro metabolism studies for model 

parameterization

• Microsomal and cytosolic fractions were prepared from 

PND15, 21 and 90 SD rat liver samples

• Kinetic studies were performed to determine Vmax and Km 

in 

– Microsomes

• CYP enzymes

• CES enzymes

– Cytosol

• CES enzymes

• These in vitro-derived metabolic constants were 

extrapolated to in vivo according to biological scaling steps 

and used in the PBPK model

35

Exclusive Property of the Council for the Advancement of 

Pyrethroid Human Risk Assessment, LLC (CAPHRA)
The XIII International Congress of Toxicology | July 2, 2013, Seoul, Korea 36

In vitro measured age-specific metabolic capacity used 

for IVIVE

Age-specific metabolic capacity (Vmax) values in vitro

The data are collected by LFR in CAPHRA research efforts.

Km values range from 0.8 to 1 μM before adjusting for non-specific binding 

Exclusive Property of the Council for the Advancement of 

Pyrethroid Human Risk Assessment, LLC (CAPHRA)
The XIII International Congress of Toxicology | July 2, 2013, Seoul, Korea 37

Rat model parameterization supported by CAPHRA data

Parameters Values

Adult (PND15)

Liver cytochrome P450 (CYP)

Vmax_LCYP (umol/h/kg tissue) 786

(142)

Km_LCYP (umol/l) 0.739 (0.952)

Liver carboxylesterase (CaE)

Vmax_Lest_m (umol/h/kg tissue) 168

(56)

Km_Lest_m (umol/l) 1.007 (2.016)

Vmax_Lest_c (umol/h/kg tissue) 170

(33)

Km_Lest_c (umol/l) 0.51 (0.562)

Plasma carboxylesterase (CaEP)

Vmax_PLest (umol/h/kg tissue) 1264

(195)

Km_PLest (umol/l) 1

(1)

Parameters Values

Adult (PND15)

Partition coefficients 

Blood to plasma ratio (BPR) 1.15

Liver/plasma (DRL) 0.8047

Fat/plasma (DRF) 68.7

GI/plasma (DRGI) Same as Liver

Brain/plasma (DRBR) 0.22

(0.25)

Slowly perfused/plasma (DRS) 3.94 (1.02)

Rapidly perfused/plasma (DRR) Same as Liver

Plasma protein binding

Total unbound fraction (FFPlas) 0.2

Parameters Values

Adult (PND15)

Oral absorption related parameters

KA , uptake rate constant (h-1

) 0.35

K12 , rate constant, from corn 

oil to lumen (h-1

) 0.2

K21 , rate constant, from 

lumen to corn oil (h-1

) 0.000026

KFEC, Fecal excretion (h-1

) 0.025

Tissue permeability area-cross product 

(L/hr/tissue weight^0.75) 

Fat (PAFC) 1.8

Brain (PABRC) 0.05

Slowly perfused (PASC) 0.08

Note: The full parameter table can be found in the overview document

Exclusive Property of the Council for the Advancement of 

Pyrethroid Human Risk Assessment, LLC (CAPHRA)
IVIVE

Ageappropriate 

metabolism 

parameters 

- in vitro

Rat deltamethrin model 

38

Refined from Tornero-Velez et al, 2010 based on the new data collected 

so far in CHPHRA

Rapidly

Liver

QBR

QF

QS

QR

QH

GI tissue

QGI

Metabolism (CES)

Metabolism

(CES and CYP)

Plasma

Vehicle Oral dose

Fecal excretion

Lumen

Brain

Fat

Slowly

QGI

QL

QBR

QF

QS

QR

Lymphatic absorption

Rat Growth 

Physiology

Exclusive Property of the Council for the Advancement of 

Pyrethroid Human Risk Assessment, LLC (CAPHRA)
The Hamner Institutes for Health Sciences | August 5, 2010 39

Describing in vivo free concentration in the 

model 

DLM

chylomicron

DLM

chylomicron

DLM DLM

VLDL or HDL

DLM

VLDL or HDL

Plasma Tissue

Plasma 

proteins

/lipids

Partition 

to tissue

Lipolysis by 

lipoprotein 

lipases

39

Exclusive Property of the Council for the Advancement of 

Pyrethroid Human Risk Assessment, LLC (CAPHRA) 
The Hamner Institutes for Health Sciences | August 5, 2010 40

Recapitulating dose-dependent kinetic profiles of 

deltamethrin

PND90 (new CAPHRA data)

The data are collected by UGa in CAPHRA research efforts.

Serial sampling from plasma was performed ; 5 mlg/kg corn oil used as vehicle 

40 Exclusive Property of the Council for the Advancement of Pyrethroid Human Risk Assessment, LLC (CAPHRA) 
Exclusive Property of the Council for the Advancement of 

Pyrethroid Human Risk Assessment, LLC (CAPHRA)

PND90 (new CAPHRA data)

The data are collected by UGa in CAPHRA research efforts.

5 mlg/kg corn oil used as vehicle 

Recapitulating deltamethrin kinetics in adult rats

41
Exclusive Property of the Council for the Advancement of 

Pyrethroid Human Risk Assessment, LLC (CAPHRA)

Recapitulating deltamethrin kinetics in adult rats

PND90 (published data)

The data are from Kim et al., 2008

1 ml/kg glycerol formal used as vehicle

42
Exclusive Property of the Council for the Advancement of 

Pyrethroid Human Risk Assessment, LLC (CAPHRA)

Recapitulating deltamethrin kinetics in juvenile rats

PND15 (new CAPHRA data)

The data are collected by UGa in CAPHRA research efforts.

5 mlg/kg corn oil used as vehicle 

43
Exclusive Property of the Council for the Advancement of 

Pyrethroid Human Risk Assessment, LLC (CAPHRA)

Recapitulating deltamethrin kinetics in juvenile rats

PND10 (published data)

The data are from Kim et al., 2008

1 ml/kg glycerol formal used as vehicle

Note that PND15 metabolic constants were used here. 

44
45

Application of IVIVE

approach to develop 

age-specific PBPK 

models in humans 

Deltamethrin PBPK 

model for humans 

at different ages

Human

Subcellular 

fractions from 

adult liver with CES

ontogeny data

Application of the 

IVIVE approach to 

humans

Exclusive Property of the Council for the Advancement of 

Pyrethroid Human Risk Assessment, LLC (CAPHRA) 
Developing human early age models for 

deltamethrin

• Applies the in vitro-based parameterization approach 

validated in the rat modeling 

• Human in vitro metabolism data –for demonstration of the 

approach with deltamethrin, subcellular fractions from 

adults and CES enzyme ontogeny were used 

• Integration of information – human growth physiology 

(Clewell et al., 2004), in vitro-derived metabolic constants 

(LFR data in CAPHRA), and exposure levels are coherently 

integrated in PBPK models 

Copyright 2014 by The Hamner Institutes for Health Sciences. May not be reproduced without permission 46

Exclusive Property of the Council for the Advancement of 

Pyrethroid Human Risk Assessment, LLC (CAPHRA)
Human PBPK model for deltamethrin

47

Human relevant 

Exposure

IVIVE

Age-appropriate 

metabolism

parameters from 

in vitro data

Adult liver 

metabolism by 

CES (LFR)

CES ontogeny 

IVIVE (MCoW)

Rapidly

Liver

QBR

QF

QS

QR

QH

GI tissue

QGI

Metabolism

(CES and 

CYP)

Plasma

Oral dose

Fecal excretion

Lumen

Brain

Fat

Slowly

QGI

QL

QBR

QF

QS

QR

Lymphatic absorption

Human 

Growth 

Physiology

Exclusive Property of the Council for the Advancement of 

Pyrethroid Human Risk Assessment, LLC (CAPHRA)
IVIVE steps for human deltamethrin modeling

• The ultimate plan is to use recombinant enzyme-based 

data, but preliminary simulations were conducted using the 

average adult Clint from adult subcellular fractions data 

(CES-1 is the major contributor)

48

Measuring metabolic 

rates

Clint_microsomes = 24 

ml/min/mg 

Clint_cytosol = 6 ml/min/mg

(LFR, preliminary)

Average adult in vivo 

Clint = 86896 L/hr/kg liver

in adults

(mainly CES-1 mediated)

IVIVE

CES-1 ontogeny 

Mature at birth

(Hines et al., 

submitted)

Ontogeny on Liver weight and 

blood flow (Clewell et al., 2004)

Ontogeny on the scaling factor 

(MPPGL) (Barter et al., 2007;2008)

Copyright 2013 by The Hamner Institutes for Health Sciences. May not be reproduced without permission

Clint_in vivo

All ages

The estimated average Clints for 

each age are shown

PBPK modeling

Exclusive Property of the Council for the Advancement of Pyrethroid Human Risk Assessment, LLC (CAPHRA)
The XIII International Congress of Toxicology | July 2, 2013, Seoul, Korea 49

Human model parameterization based on IVIVE supported by 

CAPHRA data

Parameters Value

Metabolism

CLCAEC (L/h/kg liver)* 86896

CLCYPC (L/h/kg liver) 0

* Sum of microsomal and cytosolic CESs

Parameters Value

Partition coefficients

Blood to plasma ratio 

(BPR) 1.15

Liver/plasma (DRL) 0.8047

Fat/plasma (DRF) 68.7

GI/plasma (DRGI) Same as 

Liver/plasma

Brain/plasma (DRBR) 0.25

Slowly perfused/plasma 

(DRS) 3.94

Rapidly perfused/plasma 

(DRR)

Same as 

Liver/plasma

Erythrocyte/plasma 

(DRRBC) 1.34

Parameters Value

Oral absorption

KA , Uptake rate constants (h-1

) 0.35

KFEC, Fecal excretion (h-1

) 0

Protein binding

Total unbound fration (FFPlas) 0.1

Tissue permeability area-cross product 

(L/hr/tissue weight^0.75) 

Fat (PAFC) 1.8

Brain (PABRC) 0.05

Slowly perfused (PASC) 0.08

Note: The full parameter table can be found in the overview document

Exclusive Property of the Council for the Advancement of 

Pyrethroid Human Risk Assessment, LLC (CAPHRA)
The XIII International Congress of Toxicology | July 2, 2013, Seoul, Korea 50

Predicting deltamethrin internal exposure in 

humans across age

50

Simulation is performed using a single dose of deltamethrin at 0.005 mg/kg

Exclusive Property of the Council for the Advancement of 

Pyrethroid Human Risk Assessment, LLC (CAPHRA)
The XIII International Congress of Toxicology | July 2, 2013, Seoul, Korea 51

Which parameters have the most influence 

over the deltamethrin brain concentration? 

• The relative influence of each of the model parameters on 

brain deltamethrin concentrations was evaluated by a 

sensitivity analysis

• This information can determine key data requirements and 

identify studies needed for read-across for additional 

pyrethroids

51

Exclusive Property of the Council for the Advancement of 

Pyrethroid Human Risk Assessment, LLC (CAPHRA)
The XIII International Congress of Toxicology | July 2, 2013, Seoul, Korea 52

Sensitivity analysis of the human deltamethrin model

• The analysis was performed for 

– 8 hr post-exposure (around Cmax for all the ages simulated) to a single dose 

at 0.005 mg/kg

– two ages, adult (20 year old) and young (1 year old)

• The sensitivity coefficient (SC) is defined as the percent change in the dose metric 

(brain DLM concentration) resulting from a 1% change in the particular parameter. 

• Thus, an SC of 1 represents 1:1 relationship between the change in the parameter 

and the internal dose metric. A negative SC indicates and inverse relationship.

• Sensitivity coeffieicnts are grouped as high (absolute value greater than or equal 

to 0.5), medium (absolute value greater than or equal to 0.2 but less than 0.5) or 

low (absolute value greater than or equal to 0.1 but less than 0.2); parameters 

with sensitivities less than 0.2 are not listed (IPCS guideline).

• To determine the overall importance of a parameter, both sensitivity and 

uncertainty have to be considered together. 

Exclusive Property of the Council for the Advancement of 

Pyrethroid Human Risk Assessment, LLC (CAPHRA)
The XIII International Congress of Toxicology | July 2, 2013, Seoul, Korea 53

Model parameter sensitivity

Sensitivity coefficient

Adult

(20 year old)

Young

(1 year old)

Age-related data 

available?

8 hr post exposure (How uncertain we are in 

the parameter?)

parameter

SC 

designation SC value SC designation SC value

Chemical 

specific 

parameters

Maximum rate 

of metabolism CLCAECM* H 0 H 0 Age-specific in vitro data 

(Low uncertainty)

Brain 

permeability PABRC H 0.86 H 0.86

Estimated based on in 

vivo PK data/Lipoprotein 

lipase distribution

(Medium uncertainty)

Physiological 

parameters

Cardiac output QCC H -0.85 H -0.94

Published database 

available

(Low uncertainty)

Liver blood 

flow QLIVC H -0.8 H -0.89

Published database 

available 

(Low uncertainty)

Brain volume VBRNC M -0.22 M -0.22

Published database 

available 

(Low uncertainty)

Exclusive Property of the Council for the Advancement of 

Pyrethroid Human Risk Assessment, LLC (CAPHRA)
The XIII International Congress of Toxicology | July 2, 2013, Seoul, Korea 54

IPCS Sensitivity/Uncertainty matrix

UNCERTAINTY

High Medium Low

SENSITIVITY

High

— Brain permeability

Metabolic Clearance

Cardiac Output

Liver Blood Flow

Medium

— Brain volume

Low

—

Body weight,

Other tissue weights 

and blood flows

* From the International Programme on Chemical Safety (IPCS) Harmonization Project Document on 

‘Characterization and Application of Physiologically based Pharmacokinetic Models in Risk Assessment’

Exclusive Property of the Council for the Advancement of Pyrethroid Human Risk Assessment, LLC (CAPHRA)
The XIII International Congress of Toxicology | July 2, 2013, Seoul, Korea 55

• Predict brain and plasma deltamethrin

concentrations

• in different ages of humans 

• under different exposure scenarios – capability to connect to and use outputs 

from exposure models (CARES or SHEDS) 

• Use model-predicted brain dosimetry 

• to develop CSAF for early age subpopulation 

• to compare with effective concentrations 

observed in neurotoxicity studies to inform 

margin of safety

Copyright 2014 by The Hamner Institutes for Health Sciences. May not be reproduced without permission 55

Examples of model 

application

Exclusive Property of the Council for the Advancement of 

Pyrethroid Human Risk Assessment, LLC (CAPHRA)
The Hamner Institutes for Health Sciences | August 5, 2010 56

Simulation of dose-dependent brain Cmax profiles in 

vivo neurotoxicity studies

Note that the x-axis is in a log scale.

Exclusive Property of the Council for the Advancement of Pyrethroid Human Risk Assessment, LLC (CAPHRA) 56
The Hamner Institutes for Health Sciences | August 5, 2010 57

Reverse dosimetry to estimate human equivalent 

doses

RAT HUMAN

Age External 

Dose

Brain Cmax

(from model)

External Dose based on Brain 

Cmax Target

Juvenile PND15 1 mg/kg* 48 ng/mL 2.0 mg/kg Child

Adult PND90 1 mg/kg* 11 ng/mL 0.2 mg/kg Adult

* Rat NOAEL for both juvenile and adult rats

Exclusive Property of the Council for the Advancement of 

Pyrethroid Human Risk Assessment, LLC (CAPHRA) 57
The Hamner Institutes for Health Sciences | August 5, 2010 58

Distribution of simulated brain Cmax in early 

ages vs adults

Single dose of deltamethrin 0.005 mg/kg was used for Monte Carlo simulation

Horizontal line bisecting large rectangle, median; large rectangle, lower and upper quartiles; whiskers, 5th and 95th

percentiles

1yr 5 yr 10 yr Adult

Age

Brain Cmax (ng/ml) 

Exclusive Property of the Council for the Advancement of 

Pyrethroid Human Risk Assessment, LLC (CAPHRA) 58
Acknowledgement

Modeling

• The Hamner Institutes

• Huali Wu

• Gina Song

• Miyoung Yoon

• Harvey Clewell

• David Billings

• Alina Efremenko

Data collection

• LFR Molecular Sciences

• Brian G. Lake Lab

• University of Georgia

• James Bruckner Lab

• Medical College of Wisconsin

• Gail McCarver/Ron Hines Lab

59

CAPHRA discussion

• Thomas Osimitz (Science Strategies, 

LLC)

• Paul Hinderliter (Syngenta)

• Other members in the Consortium

Funding

The Council for Advancement of 

Pyrethroid Human Risk Assessment, 

LLC (CAPHRA)

Exclusive Property of the Council for the Advancement of 

Pyrethroid Human Risk Assessment, LLC (CAPHRA)