SUMMARY & NEXT STEPS 

ANNA LOWIT, PH. D.

SENIOR SCIENTIST

USEPA, OFFICE OF PESTICIDE 

PROGRAMS

FIFRA SAP April 10, 2012
Overview

2

 Follow text in Section 6.0 of the 2012 SAP

issue paper

 Summarize key points from previous 

presentations

 Discuss weight of the evidence concept

 Key science questions

 Qualitative & quantitative considerations

 Highlight aspects of the charge questions
NRC Reports

 NRC Reports

 Toxicity Testing in the 21st Century: A Vision and 

a Strategy (2007): 

 Adverse outcome pathways

3

 Integration across multiple lines of evidence

 Disruption of homeostasis leading to adverse 

outcomes

 Science & Decisions (2009):

 Improved technical analysis (including mode of action)

 Evaluation of alternative interpretations/approaches

 Better align risk assessment & risk management

 Use of improved problem formulation
Stepwise, Transparent Science 

Review

4

 2008, FIFRA SAP: Preliminary review 

of chlorpyrifos human health effects 

with focus on infants, children & 

pregnant women 

 2010, FIFRA SAP: Draft Framework 

for Incorporating Epidemiologic and 

Human Incidence Data in Human 

Health Risk Assessment 
Stepwise, Transparent Science 

Review

5

 2011, Preliminary risk assessment on chlorpyrifos: 

 Focus on AChE/ChE inhibition as the most sensitive 

dose-response data for deriving points of departure

 Plan to conduct a full weight of evidence (WOE) 

evaluation integrating the epidemiology studies with 

the experimental toxicology studies for the 

neurodevelopmental outcomes 

 2012, FIFRA SAP: New data & more in-depth 

analysis since 2008

 Charge questions on strengths & uncertainties of 

different lines of evidence

 Uncertainty analysis: qualitative & quantitative

 Next step: Agency will consider input from SAP in 

integrating data & developing the WOE
2007 NRC Toxicity Testing in the 21st

Century

Compounds Affected

Pathway

Dose Response

Analysis for 

Perturbations

Calibrating 

in vitro and human

Population Based

Studies

Exposure

Guideline

Mode of Action

Chemical

Characterization

Dose Response 

Assessment

Assess

Biological 

Perturbation

Measures of

dose in vitro

Perturbations

of Toxicity 

Pathways

in vitro and human

dosimetry

Human Exposure

Data 

Exposure

Guideline

Hazard Characterization

Risk Characterization

Exposure Assessment

NRC, 2007. Fig 3-7 Risk Assessment Compo

Metabolite(s)
Chlorpyrifos Dose Response

Analysis for 

Perturbations

Calibrating 

in vitro and human

Children’s Cohorts

Exposure 

Pathways: 

Food, water, 

Mode of Action

Chemical

Characterization

Dose Response 

Assessment

Affected

Pathway

• AChE inhibition

• Plausible hypotheses for 

neurodevelopmental effects

Assess

Biological 

Perturbation

Measures of

dose in vitro

Perturbations

of Toxicity 

Pathways

in vitro and human

dosimetry

Urinary biomonitoring of 

TCPy in various 

populations

Food, water, 

worker, 

air volatility, 

spray drift,

cross-placental

Hazard Characterization

Risk Characterization

Exposure Assessment

NRC, 2007. Fig 3-7 Risk Assessment Compo

Oxon, TCPy
Weight of Evidence (WoE)

 Process for characterizing the extent to which 

the available data support a hypothesis that an 

agent causes a particular effect 

 Informs Dose-Response Assessment

 Involves a number of steps 

 Starting with assembling the relevant data 

 Evaluating that data for quality and relevance 

 Followed by an integration of the different lines of 

evidence to support conclusions
Weight of Evidence (WoE)

 Transparency is important 

 explains the kinds of data available 

 how the data were selected and evaluated

 how the different lines of evidence fit together in 

drawing conclusions 

 significant issues, strengths, and limitations of the 

data and the uncertainties that deserve serious 

consideration are presented

 major points of interpretation highlighted

 Characterization of a range of probabilities and 

put in perspective (confidence/uncertainty)
Outline of Presentations

10

 Introduction (Anna Lowit)

 Adverse Outcome Pathways: Data for 

Chlorpyrifos at Varying Levels of Biological 

Organization (William Mundy)

 Chlorpyrifos Effects on the Developing Brain: 

Animal Studies (Virginia Moser)

 Review of Children’s Health Epidemiology 

Cohort Studies (Carol Christensen)

 Review of Chlorpyrifos Biomonitoring 

Research and Interpretive Approaches 

(Aaron Niman)

 Summary & Next Steps (Anna Lowit)
Summary

11

 2011 Preliminary Risk Assessment

 AChE/ChE inhibition remains the most sensitive 

dose-response data for deriving points of 

departure

 Consistent with 2008 SAP recommendations

 Route & lifestage specific 

 Robust BMD analysis

 Acute, oral: BMDL10 0.36 mg/kg for RBC AChE 

inhibition in PND11 rats

 Chronic, oral: BMDL10 0.03 mg/kg/day for RBC AChE 

inhibition from pregnant rats
Summary

12

 Compared to 2008, 2012 analysis incorporates 

new data available and/or provides more indepth analysis

 Key scientific questions 

 Strength/limitations of evidence that suggest 

chlorpyrifos exposure results in long-term 

neurodevelopmental effects from early life 

exposure 

 Strength/limitations of evidence that adverse 

effects may be attributed to doses lower than 

those which elicit 10% inhibition of AChE, i.e., the 

dose levels used for regulatory decision making
Summary-Qualitative lines of evidence: Long-term 

neurodevelopmental effects from early life 

exposure

13

 Strength/limitations of evidence that suggest 

chlorpyrifos exposure results in long-term 

neurodevelopmental effects from early life 

exposure-

 Mechanistic studies

 In vivo experimental toxicology studies

 Epidemiology
Summary - Qualitative lines of evidence: Longterm neurodevelopmental effects from early life 

14exposure

 Charge Question 2.1

 Mechanistic studies

 At this time, a mode of action/adverse 

outcome pathway has not been established 

for neurodevelopmental outcomes 

 Growing body of literature does demonstrate 

that chlorpyrifos and/or its oxon are 

biologically active on processes that affect the 

developing brain
Summary -Qualitative lines of evidence: Longterm neurodevelopmental effects from early life 

15exposure

 Charge Question 3.1

 In vivo experimental toxicology studies

 Multiple studies and laboratories reporting 

changes in aspects of neurobehavioral function in 

adult rodents following developmental exposure

 Effects seen long after chemical exposure and any 

ChE inhibition, if present, had recovered

 Behavioral alterations show some consistency in 

terms of broad domains of neurological function

 Most consistent effects on cognition, anxiety, social 

behavior
Summary - Qualitative lines of evidence: Longterm neurodevelopmental effects from early life 

16exposure

 Charge Question 4.1 & 4.2

 Epidemiology studies

 Current database strengthens conclusion from 2008 

“chlorpyrifos likely plays a role” in the 

neurodevelopmental outcomes specifically those 

reported by Columbia University

 Strength of observed associations high in some cases, but not 

all

 Non-differential exposure measurement error could reduce 

magnitude

 Some evidence of exposure-response in studies, when 

available

 Consistency in Brazelton (abnormal reflexes), ADHD-like 

behavior problems, Bayley (particularly MDI), and IQ

 Significant given differing exposure measures
Summary

17

 From a qualitative perspective---Agency 

preliminarily concludes: 

 Lines of evidence together support a 

conclusion that exposure to chlorpyrifos 

results in adverse neurodevelopmental 

outcomes in humans, at least under some 

(still unclear) conditions

 Mechanistic studies

 In vivo experimental toxicology studies

 Epidemiology
Summary

18

 Key science question

 Strength/limitations of evidence that adverse 

effects may be attributed to doses lower than 

those which elicit 10% inhibition of AChE, i.e., 

the dose levels previously used for regulatory 

decision making

 Quantitative considerations

 Mechanistic studies

 In vivo experimental toxicology studies

 Epidemiology
Summary - Quantitative Lines of Evidence: 

Potential for Effects < Doses Eliciting 10% AChE 

19 Inhibition

 Charge Question 2.2

 Mechanistic studies

 Multiple biologically plausible hypotheses are 

being pursued by researchers; no one pathway 

has sufficient data to be considered more credible 

than the others. 

 Cannot establish key events, causal linkages or doseresponse linkages

 Dose metric uncertain (peak exposures vs. AUC)

 There is some evidence of possible effects which 

are similarly sensitive or more sensitive than 

AChE inhibition. 

Thiif tit
Quantitative Lines of Evidence: Potential for 

Effects < Doses Eliciting 10% AChE 

20 Inhibition

 Charge Question 3.2

 In vivo experimental toxicology studies

 Many studies use only one, maybe two doses, and in 

several cases the lower but not higher dose level produced 

significant neurodevelopmental effects

 Most studies use of 1 mg/kg/day or higher

 Dose that produces some amount of brain ChE inhibition when 

given directly to the pups post-natally

 Limited or no dose response data on inhibition of fetal brain 

ChE activity below 1 mg/kg when given to the dams 

gestationally 

 One study (Braquenier et al., 2010) using lower doses, 

administered to the dam on GD15-LD14, reported a NOEL

of 0.2 mg/kg/d

 Similar to the acute, oral BMDL10 0.36 mg/kg

 About 10X higher than chronic, oral BMDL10 0.03 mg/kg/day
Quantitative Lines of Evidence: Potential for 

Effects < Doses Eliciting 10% AChE 

21 Inhibition

Effects seen 

6.17 pg/g 

upper-tertile

of >LOD 

(Rauh et al. 

2006)

Year N Mean CPF (pg/g)

1999 87 5.0 (4.1-6.2)

2000 83 1.9 (1.4-2.4)

2006) 2001 33 0.5 (0.3-0.6)

1998-2002 314 4.0 (sd 6.1) 

Range: ND-63 

2001-2004 65 <LOD

2.5 (95th)

Whyatt et al. 2003, 2004, 2009
Summary - Quantitative Lines of Evidence: 

Potential for Effects < Doses Eliciting 10% AChE 

Inhibition

22

Evidence suggesting that pg/g levels of 

chlorpyrifos unlikely to elicit AChE inhibition

 Nolan, 1984: Single dose, deliberate dosing study in adult males

 0.5 mg/kg in single oral dose

 No RBC AChE inhibition observed

 Plasma ChE w Plasma ChE was inhibited inhibited 83-89% relative to pre relative to pre-dose activities activities 

following the oral dose

 Peak blood levels of 0.01-0.03 ug/mL

 5 mg/kg, single dermal dose

 No RBC AChE or plasma ChE inhibition observed

 Mostly non-detects, maximum of 0.01 ug/mL

 Comparison to epidemiology studies needs to be interpreted with care:

 Males vs. Pregnant females

 Single Dose vs. Repeating exposure

 Controlled, clinical setting vs. “Real world”

 Oral route vs. Multiple routes/pathways
23

1000

10000

100000

1000000

10000000

100000000

P

e

a

k

Blo

o

d

C

o

n

c

e

n

t

r

a

tio

n (p

g/g)

Relationships between Single Oral Dose and Peak Blood 

Concentrations of Chlorpyrifos and Chlorpyrifos Oxon in Adults 

and Infants

Adult Chlorpyrifos

Adult Chlorpyrifos oxon

Child Chlorpyrifos

Child Chlorpyrifos oxon

Figure 40, Dow PBPK-CARES SAP Issue Paper, 

2010

0.1

1

10

100

0.01 0.1 1 10

P

e

a

k

Blo

o

d

C

o

n

c

e

n

t

r

a

tio

n (p

g/g)

Dose mg/kg

50 per. Mov. Avg. (Adult Chlorpyrifos)

50 per. Mov. Avg. (Adult Chlorpyrifos 

oxon)
Quantitative Lines of Evidence: Potential for 

Effects < Doses Eliciting 10% AChE 

24 Inhibition

 Exposure range cord blood 3-6 pg/g

 Remaining Uncertainties:

 Fetal exposure during critical window(s) of 

development

 Including blood levels or the AChE inhibition at the 

time of chlorpyrifos application

 Contribution of other ChE-inhibiting pesticides 

to OP exposure 

 Diazinon (ρ=0.63), propoxur (ρ=0.21) Whyatt et al. 2009
Summary - Quantitative Lines of Evidence: 

Potential for Effects < Doses Eliciting 10% AChE 

Inhibition

25

 Charge questions 5.3 & 5.4

 Given the uncertainties, area of 

where additional analyses may 

provide helpful characterization of 

biomonitoring data
Summary

26

 Charge Question 6.0

 Assessing and presenting the range 

of plausible responses at given 

doses, and the effect of the overall 

uncertainty and variability around that 

range. 
 Ensure scientific judgements are transparent

 Identify areas of uncertainty

 Practical steps towards further refinement of 

existing information with existing information

SAP Questions

existing information with existing information
28 Questions?