computational modeling of nonmonotonic dose … · nonmonotonic dose-response: insight into minimum...

Computational modeling of nonmonotonic dose-response:

Insight into minimum requirements

0

Rory Conolly

Low Dose Effects and Non-Monotonic Dose Responses for Endocrine Active Chemicals: Science to Practice

An International WorkshopSeptember 11-13, 2012

Charité Universitätsmedizin, Berlin, Germany

Disclaimer

This is a presentation of the opinions of Rory Conolly, and may opinions of Rory Conolly, and may not represent official policies of the US EPA.

Goal of presentation

To describe some biological mechanisms that can generate nonlinear and nonmonotonic dose-responses.that can generate nonlinear and nonmonotonic dose-responses.

2

Goal of presentation

� To describe some biological mechanisms that can generate nonlinear and nonmonotonic dose-responses.that can generate nonlinear and nonmonotonic dose-responses.

� To illustrate principles� won’t focus exclusively on endocrine

effects.

3

Outline

� Extrapolation, risk assessment, and uncertainty

� Review of determinants of dose-� Review of determinants of dose-response� Pharmacokinetics

� Tissue/cellular responses

� Nonmonotonicity

4

Historically, the data haven’t answered the question

Res

pons

eR

espo

nse

Dose

Interspecies

Extrapolation

� Extrapolation below the doses for which we have data is typical, no matter the actual shape of the dose-response curve.curve.� Some exceptions with epidemiological data

and newer tox. studies

� How much uncertainty we are willing to live with?

6

Key questions

� What is the shape of the curve?� More specifically, the dose-time response

surface.surface.

� What is the exposure?

7

Key questions

� What is the shape of the curve?� More specifically, the dose-time response

surface.surface.

� What is the exposure?

8

(Risk defined by intersection of exposurewith dose-time response surface)

Biological mechanismsdetermine dose-response

Exposure Tissuedose

Tissueinteraction

Pharmacokinetics

dose interaction

Tissueinteraction

Early tissueresponse

Irreversiblepathology

Pharmacodynamics

Vinyl chloride: Saturable bioactivation by CYP2E1

CYP2E1

13

CYP2E1

Nonmonotonicity and dose relevancerelevance

9/21/2012 15

Environmental Estrogen-induced luciferase expressionin vitro in MDA KBluc cells

80

100

Serum level inrats treated

High in vitro concentrations,unattainable in vivo,

causing a NMDRC

Per

cen

t of M

axim

um

Est

rog

enic

res

po

nse

-16-15-14-13-12-11-10 -9 -8 -7 -6 -5 -4 -30

20

40

60 rats treatedwith a very

high oral doseof a potentestrogen

Conc in waterblocking fishreproduction

Concentration (M)

Per

cen

t of M

axim

um

Est

rog

enic

res

po

nse

Earl Gray

Nonlinearity in PK,Dose relevance

� Not new concepts, but easy to ignore

17

� Not new concepts, but easy to ignore

� Important, especially considering the growing emphasis on in vitro studies

Biological mechanismsdetermine dose-response

Exposure Tissuedose

Tissueinteraction

Pharmacokinetics

dose interaction

Tissueinteraction

Early tissueresponse

Irreversiblepathology

Pharmacodynamics

“Normal”

Exposure

Uptake-Delivery to Target Tissues

Perturbation

Cellular response pathway

Homeostasis:Adaptation/Compensation

19

Biologic

inputs

“Normal”

Biological

Function

Adverse

Outcomes

(e.g., mortality,

Reproductive

Impairment)

Cell injury,

Inability to

regulate

Adaptive

Responses

Early cellular

changes

200

250

300

200

250

300L

abeling index

200

250

300

200

250

300L

abeling index

Adaptation:

Formaldehydecytotoxicity

Dose-time response surface for cytotoxicity of inhaled formaldehyde to the respiratory epithelium of the rat nose

20A

1A2A3A4A

5

B1B2B3B4B5

C1C2C3C4C5

D1D2D3D4D5E

1E2E3E4E5F

1F2F3F4F5

0. 1 4

0. 5 7

1 . 2 9

6 . 00

1 3 . 00

2 6 . 00

5 2 . 00

7 8 . 00

0

50

100

150

Duration of exposure

(weeks)

0.140.571.296.13.26.52.78.12

34

5

23

45

1

23

45

1

23

45

1

2345

1

234

5

1

control

0.7 ppm

2 ppm

6 ppm

10 ppm

15 ppm

0

50

100

150

Labeling index

A1A

2A3A4A

5

B1B2B3B4B5

C1C2C3C4C5

D1D2D3D4D5E

1E2E3E4E5F

1F2F3F4F5

0. 1 4

0. 5 7

1 . 2 9

6 . 00

1 3 . 00

2 6 . 00

5 2 . 00

7 8 . 00

0

50

100

150


(weeks)

0.140.571.296.13.26.52.78.12

34

5

23

45

1

23

45

1

23

45

1

2345

1

234

5

1

control

0.7 ppm

2 ppm

6 ppm

10 ppm

15 ppm

0

50

100

150

Labeling index

200

250

300

200

250

300L

abeling index

200

250

300

200

250

300L

abeling index

Adaptation:

Formaldehydecytotoxicity

Dose-time response surface for cytotoxicity of inhaled formaldehyde to the respiratory epithelium of the rat nose

21A

1A2A3A4A

5

B1B2B3B4B5

C1C2C3C4C5

D1D2D3D4D5E

1E2E3E4E5F

1F2F3F4F5

0. 1 4

0. 5 7

1 . 2 9

6 . 00

1 3 . 00

2 6 . 00

5 2 . 00

7 8 . 00

0

50

100

150


(weeks)

0.140.571.296.13.26.52.78.12

34

5

23

45

1

23

45

1

23

45

1

2345

1

234

5

1

control

0.7 ppm

2 ppm

6 ppm

10 ppm

15 ppm

0

50

100

150

Labeling index

A1A

2A3A4A

5

B1B2B3B4B5

C1C2C3C4C5

D1D2D3D4D5E

1E2E3E4E5F

1F2F3F4F5

0. 1 4

0. 5 7

1 . 2 9

6 . 00

1 3 . 00

2 6 . 00

5 2 . 00

7 8 . 00

0

50

100

150


(weeks)

0.140.571.296.13.26.52.78.12

34

5

23

45

1

23

45

1

23

45

1

2345

1

234

5

1

control

0.7 ppm

2 ppm

6 ppm

10 ppm

15 ppm

0

50

100

150

Labeling index

Adaptation: Plasma estradiol in fathead minnows exposed to fadrazole

0

1

2 CONFAD-3FAD-30*

Pla

sma

E2

(fol

d-ch

ange

rel

ativ

e to

con

trol

; lo

g 2)

22

0 2 4 6 8-4

-3

-2

-1

10 12 14 16

#

##

#

*

*Pla

sma

E2

(fol

d-ch

ange

rel

ativ

e to

con

trol

; lo

g 2)

direct effect

compensation recoveryVilleneuve et al. (2010) EHP

Adaptation: Induction of CYP19A

mRNA

23

mRNA

Nonlinear intracellular signaling

Bifurcation in a MAPK signaling pathway

24

Bifurcation in a MAPK signaling pathway due to a positive feedback loop

MAPK signaling cascades

25http://www.weizmann.ac.il/Biology/open_day/book/rony_seger.pdf

Growth factor

MAPKKKPKCPKC

26

MAPKK

MAPKPLA2

AA

PLA2

AA

MKPMKP

Bhalla et al., Science, 297, 1018-23, 2002

Growth factor

MAPKKKPKCPKC

27

MAPKK

MAPKPLA2

AA

PLA2

AA

MKPMKP

Bhalla et al., Science, 297, 1018-23, 2002

60

80

100A

ctiv

ated

MA

PK

(%

of m

ax)

Figure 4A

Nonlinearity in the PDGF – ERK dose-response

0 10 20 30 40 500

20

40

PDGF (ng/ml)

Act

ivat

ed M

AP

K (

% o

f max

)

Qiang Zhang

Inhibition of ERK reduces CYP1A1 induction by TCDD

30

Nonmonotonic dose-response

31

1717ββ--trenbolone effects trenbolone effects on on biochemical endpoints in fathead minnowsbiochemical endpoints in fathead minnows

0.00

1.50

3.00

4.50

6.00

T (

ng/m

l)

a

a,b

a,b

b

a,b

a,b

4.50

6.00

aa

a

*

*Testosterone

0.00

1.50

3.00

E2

(ng/

ml)

b

a,b

b

Control 0.005 0.05 0.5 5.0 500

6

12

18

24

30

Vite

lloge

nin

(mg/

ml)

aa

b

c c

b,c

Trenbolone (µg/l)

* *

*

*

* *

*

Ankley et al. (2003) Environ.Toxicol. Chem.

Estradiol

Vitellogenin

NMDR research collaboration

Dave Gaylor Werner Lutz

33

Rory Conolly

35

“Dose-response curves for the first interaction of a chemical with a biochemical target molecule are usu ally monotonic; i.e., they increase or decrease over the entire dose range.”

- Werner Lutz

DimerizationDimerization of the estrogen receptorof the estrogen receptor

Edwards & Boonvaratanakomkit, Molec. Interventions doi: 10.1124/mi.3.1.12 MI February 2003 vol. 3 no. 1 12-15 36

Maness et at., Toxicol. Appl. Pharmacol. 151, 135-142, 1998 37

Maness et at., Toxicol. Appl. Pharmacol. 151, 135-142, 1998 38

�Computational modeling assuming that homodimers promote gene transcription but heterodimers do not.

Receptor dimerization and Receptor dimerization and gene transcriptiongene transcription

39

�Homodimers�DHT�Hydroxyflutamide

�Heterodimers�DHT-hydroxyflutamide

Conolly & Lutz. Toxicol. Sci. 77, 151-157, 2004 40

�Phenylisopropyladenosine (PIA) effect on adenylyl cyclase activity in rat brain mediated by adenosine receptor binding

Adenosine receptors A1 & A2 Adenosine receptors A1 & A2 in rat brainin rat brain

41

receptor binding�NMDR curve for the formation of cAMP in the striatum (Ebersolt et al., 1983).

�Explained by the antagonistic action of two adenosine receptor subtypes that regulate adenyl cyclase in opposite directions

42Conolly & Lutz. Toxicol. Sci. 77, 151-157, 2004

Induction of DNA repairInduction of DNA repair

43

Induction

44

Conolly & Lutz. Toxicol. Sci. 77, 151-157, 2004 45

Monte Carlo modeling to mimic population variability

46

DNA damage, cell cycle checkpoint control, and mutation

47

Monte Carlo: DNA damage, cell cycle checkpoint control, and mutation

48

Sources of nonmonotonicity

� Feedback loops� And other kinds of regulatory modules

� When the chemical interacts � When the chemical interacts differently with different parts of the system

� When the target site dose elicits induction of a compensatory response

� Etc, etc…

49

Both qualitative and quantitative issues matter

� Examples show that quantitative factors (e.g., degree of induction of factors (e.g., degree of induction of DNA repair) can determine if the dose-response curve is monotonic, an (apparent) threshold, or nonmonotonic

50

Things to consider

� Dose relevance across the full extent of the nonmonotonic curve� Are all the exposures and associated doses

relevant in the real world?relevant in the real world?

� Level of biological organization� Are nonmonotonic responses as likely in vivo

as they are in vitro?

51

GnRHNeuronal

System

GABA Dopamine

Brain

Gonadotroph

Pituitary

D1 R

D2 R

?

?

Y2 R

NPY

Y1 R

GnRH R

?

GABAA

R

GABAB

RY2 R

D2 R

GnRH

PACAP

Follistatin

Activin PAC1 R

Activin R

FSHββββ LHββββ

GPαααα

Circulating LH, FSH

FSH RLH R

Circulating LDL, HDL

LDL R

HDL R

Outer mitochondrial membrane

StAR

Cholesterol

3

4

5

6

Blood

Compartment

Fipronil (-)

Muscimol (+)

Apomorphine (+)

Haloperidol (-)

Trilostane (-)

Ketoconazole (-)

2

1

Chemical ProbesChemical Probes

52

11βHSD

P45011β.

Activin

Inhibin

Circulating Sex Steroids / Steroid Hormone Binding Globlulin

Inner mitochondrial membrane

StAR

P450scc

pregnenolone 3ββββHSD

P450c17

androstenedione

testosterone

estradiol

Gonad(Generalized, gonadal,

steroidogenic cell)

Androgen / Estrogen Responsive Tissues

(e.g. liver, fatpad, gonads)

ARER

Blood

17βHSD

P450arom

11-ketotestosterone

progesterone17α-hydroxyprogesterone

17α,20β-P (MIS)

20βHSD

Ketoconazole (-)

Fadrozole (-)

Prochloraz (-,-)

Vinclozolin (-)

Flutamide (-)

17β-Trenbolone (+)

17α-Ethynylestradiol (+)

7

8

9

10

11

12

http://sbw.kgi.edu/

Acknowledgements

� Werner Lutz

� David Gaylor

� Gary Ankley� Gary Ankley

� Dan Villeneuve

� Earl Gray

� Miyuki Breen

� Qiang Zhang

54

EndEnd

55

computational modeling of nonmonotonic dose … · nonmonotonic dose-response: insight into minimum...

Documents