Frans Jongeneelen & Wil ten Berge

EPAA workshop, Ispra (I), 13-14 October 2011

A generic, cross-chemical predictive PBTK-model

PBTK-model in exposure/dose assessment

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Schematic from: Georgopoulos

What were the demands for our generic PBTK-model?

Simple and easy model• To be used for occupational and/or environmental exposures

(Variability of environmental exposure is higher than biological variability)

• 1st tier estimate accuracy (one order of magnitude)• Minimum of data input• Free open source model

Choices made• QSPR prediction of partitioning• metabolism data of in vitro metabolism kinetics• MS Excel software platform• Species: man (in rest or light activity ), rat, mouse

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Result: Overview of the PBTK-model IndusChemFate

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Exposure scenario Three routes of uptake:

Inhalation - concentrationDermal – dose rateOral - dose

Duration of exposure Personal Protective Equipment Species Physical activity level (rest/ light)

PBTK-model

Compound data Physical-chemical properties:

Density Molecular weight Vapour pressure Log(Kow) at pH 5.5 and 7.4 Water Solubility

Biochemical parameters : Metabolism (kM and Vmax) Renal tubulair resorption Enterohepatic circulation ratio

0,00E+00

5,00E-05

1,00E-04

1,50E-04

2,00E-04

2,50E-04

3,00E-04

3,50E-04

4,00E-04

4,50E-04

0,000 10,000 20,000 30,000 40,000 50,000 60,000 70,000 80,000

Hours

Pyrene and metabolites (Venous Blood)

VenBl C0 µmol/l

VenBl C1 µmol/l

VenBl C2 µmol/l

Scheme of the physiology of the PBTK-model

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Lungs

Excretion of parent compound in urine

InhalationExhalation

VENOUS

BLOOD

ARTERIAL

BLOOD

Oralintake

Dermalload

Heart

Brain

Dermis

Adipose

Muscle

Bone

Stomach + intestine

Liver

Kidney

Evaporation

Parent compound

Cyclus of 1st metabolite

To 2nd

metabolite cyclus

Bone marrow

Lungs

Excretion of 1st metabolitein urine

Exhalation

VENOUS

BLOOD

ARTERIAL

BLOOD

Heart

Brain

Dermis

Adipose

Muscle

Bone

Stomach + intestine

Liver

Kidney

Bone marrow

Routing of chemicals and metabolites in the PBTK-model

– Absorption– Inhalation– Oral uptake– Dermal uptake

– Distribution over the body– QSPR algorithm for estimate of blood:air partitioning– QSPR algorithm for estimate of tissue:blood partitioning

– Metabolism– Saturable metabolism according to Michaelis-Menten kinetics

– Excretion– Urine – Exhaled air

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Dermal absorption module of the model

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SubstanceStratum corneum

Viable epidermis

DepositionEvaporation

Absorption

To systemic circulation

Stagnant air layer

As liquid and/or solid As vapour/gas

Skin

Vapour of substance

= New model ofAIHA-EASCnamedIH SKINPERM

Distribution over compartments in the body

– Blood:air partition coefficient• QSPR Algorithm for estimation of blood:air partitioning

based on Henry coefficient and Koa

– Blood:tissue partition coefficient• QSPR Algorithm for estimation of blood:tissue

partitioning taken from De Jong et al (1997), based on lipid content and Kow

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Metabolism– Metabolism can be modeled in all tissues– Default is metabolism in liver only– Sequential metabolism

• Parent, 1st metabolite, 2nd metabolite and so on

– Stoichiometric yield in every step• Rate of loss of parent and appearance of metabolite• Difference gives stoichimetric yield of specific metabolite

– KM and Vmax from in experiments in microsomal liver fraction or hepatocytes

• Scaled to liver weight of individual/species

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Other aspects– Excretion in urine and alveolair air– With enterohepatic circulation

• fase 2 metabolites

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The PBTK-model is build as application in MS-Excel, called IndusChemFate

• The differential equations of the PBTK-model are written in speadsheet syntax (visual basic)

• The file IndusChemFate contains 4 sheets: 1. Tutorial with instructions in short2. Worksheet

– For data entry (exposure scenario, properties of chemical under study)

– For numerical output

3. Database of phys-chemical and biochemical properties of various chemicals

4. Graphical output sheet11

Simulation example 1 Average and individual variation in workers

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Paper: Inhaled dosimetry and biomonitoring of coal liquefaction workers (Quinlan et al, 1995)

• At coal liquefaction plants PAH occur as pollutants• 5 workers were followed during 4 shift on and 96h off work• Average pyrene in breathing zone was 1.3 µg/m3 pyrene.• 1-hydroxypyrene in urine was measured during 8 days = 192 h

Simulation example 1

Metabolism of pyrene

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Figure taken from: Luukkanen et al, 2001

Simulation example 1

Kinetics of in vitro metabolism of pyrene in human hepatic fractions

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Step Tissue Parameter and value ref

Pyrene to 1-OH-pyrene

Hepatic 9000*g fraction of 12 individuals

Vmax = 180 µmol/hr/kg tissue

KM = 4.4 µM

Jongeneelen (1987)

1-OH-Pyrene to

1-OH-pyrene- glucuronide

Hepatic microsomal fraction of 3 individuals

Vmax = 6,900 µmol/hr/kg tissue

KM = 7.7 µM

Luukkanen et al (2001)

Simulation example 1

Modeling: Data to be entered

1) Enter phys-chemical properties and biochemical parameters of parent compound and metabolites under study

2) Enter exposure scenario1) Inhalation: concentration and duration2) Dermal: dose rate and duration3) Oral: bolus dose

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Simulation example 1Entering properties & biochemdata of pyrene and metabolites

Pyrene

1-OH-Pyrene

1-OH-Pyrene-glucuronide

Parent Compound PyreneCAS 129-00-0Density (mg/cm3 or grams/litre) 1270Molecular weight 202,26Vapour Pressure (Pa) 0,0106Log(Kow) at skin pH 5.5 4,88Log(Kow) at blood pH 7.4 4,88Water solubility (mg/litre) 0,135Resorption tubuli (y/n/?) yEnterohepatic removal (relative to liver venous blood) 0Vmax Liver (parent[total] µmol/kg tissue/hr) 360Km Liver (parent[total] µmol/litre) 4,5Vmax Liver (parent[specif] µmol/kg tissue/hr) 180Km Liver (parent[specif] µmol/litre) 4,51st metabolite HydroxypyreneCAS 5315-79-7Density (mg/cm3 or grams/litre) 1000Molecular weight 218,28Vapour Pressure (Pa) 0,000022Log(Kow) at skin pH 5.5 Log(Kow) at blood pH 7.4 4,45Water solubility (mg/litre) 4Resorption tubuli (y/n/?) yEnterohepatic removal (relative to liver venous blood) 0Vmax Liver (1st metab[total] µmol/kg tissue/hr) 6900Km Liver (1st metab[total] µmol/litre) 7,7Vmax Liver (1st metab[specif] µmol/kg tissue/hr) 6900Km Liver (1st metab[specif] µmol/litre) 7,72nd metabolite Hydroxypyrene GlucuronideCAS 154717-05-2Density (mg/cm3 or grams/litre) 1000Molecular weight 394Vapour Pressure (Pa) 3,2E-17Log(Kow) at skin pH 5.5 Log(Kow) at blood pH 7.4 -2,12Water solubility (mg/litre) 40000Resorption tubuli (y/n/?) nEnterohepatic removal (relative to liver venous blood) 0,3Vmax Liver (2nd metab[total] µmol/kg tissue/hr)Km Liver (2nd metab[total] µmol/litre)Vmax Liver (2nd metab[specif] µmol/kg tissue/hr)Km Liver (2nd metab[specif] µmol/litre)

Simulation example 1

Entering exposure scenario of the coal liquefaction workers

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Airborne exposurescenario

Dermal exposurescenario

Oral intakescenario

Parameters Airborne ExposureµConcentration parent compound (mg/m3) 0,0013Start of airborne exposure (hours) 0Duration of airborne exposure (hours) 12Respiratory protection factor ( => 1) 1Dermal protection factor (air tight clothing => 1) 1

Parameters Dermal exposure to parent compoundSkin deposition pure substance (mg/cm2/hour) 0Start of skin exposure (hours) 0Duration of skin exposure (hours) 8Skin temperature (centigrade) 25Affected skin area (cm2) 5000

Parameters of oral absorptionBolusdose to stomach of parent compound (mg/kg bwt) 0Time of application (time in hours) 0Absorption rate into intestinal tissue (1/hour) 3

Simulation example 1

Run program - Results as table with levels and amounts in fluids and tissues

Simulation example 1

Run program- Results as graphs

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0,00E+00

2,00E-10

4,00E-10

6,00E-10

8,00E-10

1,00E-09

1,20E-09

1,40E-09

1,60E-09

1,80E-09

0,000 50,000 100,000 150,000 200,000 250,000

Hours

Pyrene and metabolites (Alveolar Air)

AlvAir C0 µmol/l AlvAir C1 µmol/l AlvAir C2 µmol/l

0,00E+00

5,00E-05

1,00E-04

1,50E-04

2,00E-04

2,50E-04

3,00E-04

3,50E-04

4,00E-04

4,50E-04

0,000 50,000 100,000 150,000 200,000 250,000

Hours

Pyrene and metabolites (Venous Blood)

VenBl C0 µmol/l VenBl C1 µmol/l VenBl C2 µmol/l

0,00E+00

5,00E-03

1,00E-02

1,50E-02

2,00E-02

2,50E-02

3,00E-02

3,50E-02

4,00E-02

4,50E-02

5,00E-02

0,000 50,000 100,000 150,000 200,000 250,000

Hours

Pyrene and metabolites (Urine)

UrinConc C0 µmol/l UrinConc C1 µmol/l UrinConc C2 µmol/l

Figure 3: Urine

Figure 2: Blood

Figure 1: Alveolair air

Simulation Example 1

Result: predicted concentration total 1-OHP in urine of the coal liquefaction workers (in umol/L)

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0,00

0,01

0,02

0,03

0,04

0,05

0,06

0,07

0,081 12 23 34 45 56 67 78 89 100

111

122

133

144

155

166

177

188

199

21Note: Dermal exposure was not measured and set at zero in simulation

Simulation example 1 Comparison measured 1-OHP in urine versus predicted

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Simulation Example 1

Result: Estimated fate of mass of pyrene + metabolites

0,00

0,05

0,10

0,15

0,20

0,25

0,30Mass of pyrene + metabolites (in µmol)

After 43 h After 102 h After 198 h

Simulation example 1

Conclusion

The average level of hydroxypyrene can be predicted when the exposure scenario is known

A rough predictive model does when we realise that differences of external exposure between workers are large

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Simulation example 2: dermal uptake after desinfection of hands with ethanol

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Nr. Compound Type of study

Exposure route

Exposure scenario Reference

1 Pyrene Observa-tional study of workers

Inhalation + dermal

4*12 on work, 96 hr off work

Quinlan, 1995

2 Ethanol Volunteer study

Application on skin, dermal

10 times disinfection of hands and arms. Rubbing during 80 min.

Kramer, 2007

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Dermal + inhalation of evaporated ethanol!

Solid line: dermal only exposure scenario

Simulation example 2Ethanol in blood after disinfecting of hands and arms (Kramer et al, 2007)

Simulation example 2

Conclusion

Dermal uptake of ethanol can be predicted

Evaporation of ethanol seems to be a significant extra route of uptake of ethanol after handrubbing

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What are the differences between this PBTK-model and other PBTK-models?

• GENERIC MODEL– Distribution of the chemical/metabolite in the body is

estimated by algorithms, thus model can easily can be used for many different volatile and semi-volatile chemicals

• RUNS IN WIDELY AVAILABLE SOFTWARE– The software application is running in MS EXCEL, a

software platform that is widely available

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Suggested application of this PBTK-model IndusChemFate

Chemical health risk assessors• Volatile and semi-volatile compounds• A priori (= 1st tier) estimation of concentration in body tisues

and/or body fluids at specific exposure scenarios• Screening of absorpion and fate of data-poor substances in

the human body• In vivo extrapolation of in vitro ADME data• Comparisons of toxicokinetics in different species (in-vivo to

in vivo extrapolation)• Assessment of contribution of dermal uptake to body burden

Student and non-experts• Educational tool to understand toxicokinetics of chemicals in

human body in relation to phys-chem properties

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Where to get more info?• Download EXCEL-application file and user manual:

– Website CEFIC LRI, webpage IndusChemFatehttp://www.cefic-lri.org/lri-toolbox/induschemfate

• Read Paper (in Annals Occupational Hygiene 2011) • See at our website: www.industox.nl• Ask us to do a live-demonstration

Acknowledgement

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Funding from CEFIC-LRI

Thank you

Any questions?