REVIEW OF CHILD

EPIDEMIOLOGY COH

CAROL H. CHRISTENSE

EP

USEPA, OFFICE OF PESTICID

FIFRA SAP 
Background and History 

 September 2008 EPA presented targ

three children’s health cohort studies

 FIFRA SAP Report:

 “Chlorpyrifos likely played a role”

 Cannot conclude “chlorpyrifos is the sole c

other co-exposures

 However, epidemiology studies "should 

considered ...basis for characterization 

 Lack biologically plausible MOA, exposur

 Supplemental analyses recommended

2
Background and History 

 April 2012 EPA updates and expands its ta

these data: 

 New etiologic papers

 Intelligence Quotient (IQ)

 New methodological papers

 Socioeconomic status (SES) Exposure characterization and validation

 Supplemental analyses (unpublished) (Append Draft Framework Analysis approach 

 Working toward WOE Evaluation

 Panel guidance regarding qualitative and/

use these studies in risk assessment

3
Overview of Cohorts

4

 Columbia cohort (1998-2002)

 NYC, Dominican and African -American Mothers-In Cord Blood Chlorpyrifos Validation studies (air, maternal blood, urine, meco

 Mt. Sinai cohort (1998-2001)

 NYC, multi-ethnic (African American, Hispanic, Whi Measured TPCy (fetal growth only) and DAPs PON1 genotype and phenotype

 U of CA Berkeley Cohort (CHAMACOS) (1999-2

 Mexican-American, agricultural worker population Measured TPCy and DAPs PON1 genotype and phenotype
Overview of Cohorts

5

 Three prospective birth cohorts to examin

exposures and adverse health outcomes 

 Chlorpyrifos/OP studies focus of this review

 Birth outcomes Neurodevelopment

 Enrolled women during pregnancy, follow c

time, multiple measures

 Children’s Environmental Health and Diseas

Research Centers (funded by EPA and NIEH
Population Characteristics

 Sought prenatal care (<20-

26 weeks)

 Residency requirement

 Deliver participating 

hospitals 

 No diabetes, hypertension, 

HIV, history birth defects

 No smoking, illegal drug 

use

 Multi-ethn

African-AmDominicanMexican-A

 Younger m

years 

 Low preva

behaviors, (criteria)

6

General Eligibility Cohort C
Exposure Measurement, Confoundin

Statistical Analyses

7 Methods and Analysis
Exposure Assessment

8

 Cohorts differing exposure profiles, pathw

 Inner-city, agricultural and both residential, oc

 Multiple biomarkers reflect different windo

 Chlorpyrifos in cord blood (Columbia) TCPy in urine (each cohort) DAPs in urine (CDC methods) (Mt. Sinai, CHAM TCPy meconium (validation study) (Columbia) AChE and BuChE in cord blood (CHAMACOS) PON1 genotype and phenotype Chlorpyrifos in air (validation study) (Columbi Self-report questionnaire (each cohort)
Exposure Assessment

9

 Ideal Measures

 Estimates exposure during critical window  Repeat samples

 periodicity, variability of exposure

 Reflective of true rank of actual exposure

 Timing of sample collection with application

 Actual Measures

 One-time, or average of two samples Third trimester (unknown window of effect Specific and non-specific biomarkers of ch

Adapted D. Savitz 2002
Exposure Measurement Er

10

 External exposure versus measured inte

 Reflects total exposure (all routes)

 Single measure, third trimester

 Temporal-spatial patterns of exposure unc

 Exposure during critical window of deve

 Likely third trimester important

 Missing exposure data

 Imputation methods, data

 Laboratory errors

 Extensive QA/QC measures, CDC laborat
Exposure Validation Study

11

 Questionnaire Data:

 85% report use pesticide, most repo

monthly use

 Evidence consistent use prior to 2000 

 External Data

 NYC Housing Authority reported use 

 Total usage in NYC housing greater than 

prior to voluntary cancellation
Exposure Validation Study

 Air monitoring

 Last 8-wks, stationary 

air 

 Personal (48-hr) and

 Intra<inter variability

 Biomarke

 Matern

 Last  At d

 Matern Mecon Neona

 Compare 

 Etiolo

12

Period of Rapid Decline of Use
Exposure Validation Study

13

First trimester

Meconium week 1

Second Trimester T

Week 3

weekly matern

Week 32

(pilot): 48 hr.air

Conception Week Week1414 Week 20 Week 24

Cord blood, 

meconium, 

maternal urine
Exposure Validation Study

14

Cord Blood Measures Correlate with Meconium (expos
Exposure Assessment

15

 Uncertainties Remain

 Limited window of exposure measured 

 One-time measure (Columbia, Mt. Sinai) CHAMACOS averages two exposures Validation studies suggest some degree o

 Unknown periodicity of exposure

 Peaks, constant

 Prevalence of pesticide exposure high

 Median concentration TCPy within range o

population (NHANES) 

 May indicate difference actual and meas
Methods to Control Confound

 Restriction (design)

 Through recruitment techniques, similar 

characteristics 

 Statistical Adjustment (individual-le

 Race, education, income, housing qualit Tested for statistical interaction

 Multilevel Model (SES, Columbia on

 Account group-level and individual-leve

16
Methods to Select Potentially 

Confounding Variables Methods

 Searching literature, other studies in area Tested univariable and multivariable mode

 Included if observed +/- 10% change in estim Evidence of statistical confounding

 Forward/Backward step-wise selection pro Include if known risk factor

 Evidence of biological confounding 

 Many known or suspected confounding v

measured and modeled

 Emphasis on Parsimony

17
Confounding Variables

 Demographic Health-related Environmental 

exposures PAH, ETS, DDT/DDE, PCB, OC/HCB

 Postnatal environment

 Other env

chemicalsDiazinon, 

 Socio-eco

 In gener

measure

18

General Factors Major Pot

Confoun
When Confounding Bias?

*P

ρ<0.20 – likely no effect0.20<

ρ<0.40 – little chance

0.40<

ρ<0.60- some effect

0.60<

ρ<0.80 – strong effect

ρ>0.80 – collinearity

Confounder 

19

Lead, Mercury* 

correlation <0.10
Confounding: Other OPs

20

Effects of Chlorpyrifos on the odds of Mental Development DeScales of Infant Development at age 36 months. (

β coefficient

Model 1 Model 2 Model 

Chlorpyrifos

2 2.3 (1.1, 5.2)

p=0.03

3.8 (1.5, 9.4)

p=0.004

2.0 (0.9, 

p=0.1

(ln)Diazinon 0.57 (0.35, 

0.95)

p = 0.03

(ln)2-isopropoxyphenol(propoxur)

1.5 (0.9, 

p=0.1

Appendix 4: Whyatt and R
SES: Multilevel Analysis

 The study of the effects of collective 

characteristics (as well as individual oindividual-level outcomes

 Not all associations explained by indiv

characteristics

 Group-level variables may affect indiv

and also constrain the choices that indiv

 Improve accuracy and precision of stat

include Group and Individual level 

21
SES: Multilevel Analysis (co

Group level variables

Individ

Lovasi et al. 2010

22
Statistical Analysis

23

 Statistical modeling decisions approp

research question, characteristics of d

 Missing data, <LOD

 (imputed, sensitivity analyses)

 Sample size adequate main effects

 Likely under-powered for effect modifi

 Sensitivity analyses to evaluate alter

modeling options
Fetal Growth, Neonatal Neurologic

Development, Infant Motor and MeDevelopment, Attention Disorders, IMeasures

24 Results
Fetal Growth and Charact

25

 Authors hypothesized link between pe

effects on fetal growth

 Hypothesized link to neurodevelopmen

 Medical record linkage

 Birth weight, length, ponderal index Gestational age, preterm births Head circumference

 Effect modification evaluated

 PON1 genotype and phenotype 
Fetal Growth and Charact

26

Columbia

Cohort

Mount Sinai

Cohort

U of 

Markers of

Exposure: Chlorpyrifos TCPy (fetal growth) TCP

Birth Length

Inverse

(Null post 2000) Null Positive 

Birth Weight

Inverse

(Null post 2000) Null Positive 

HeadCircumferenceat Birth

‐‐

Null

(Inverse (NS))* Nu

Gestational Age ‐‐ Null Nu

* PON1 genotype or phenotype
Neonatal Neurodevelopm

27

 Brazelton Neonatal Behavioral Asses

(BNBAS)

 Goal: provide information parents/car 28 behavioral items-habituation, range 

orientation

 18 primitive reflexes

 Assessed 2-5 days post-partum Trained neonatologists administered Hospital setting
Neonatal Neurodevelopm

28

Consistent, reflexesOutcome measure good qualityDAPs only, limitation

Columbia

Cohort

Moun

Coh

Exposure

Measure

CPF DA

BNBAS(Abnormalreflexes)

‐‐ Posit

BNBAS (OtherBehavioral)

‐‐ Nu

* PON1 genotype or phenotype
Infant Neurodevelopment

29

 Bayley Scales of Infant Development II

 Mental development index (MDI) Psychomotor development index (PDI) Measured 6-36 months High reliability and validity as screening tool for early i

 Child Behavior Checklist

 Designed to assess "social competence" and "behavior p

(Maternal report)

 Race/ethnicity small proportion of score variance 99-item questionnaire to ascertain behavior and/or atte

 Home Observation Measurement of the Environment (

 Stimulating environment for child development

 Effect modification evaluated

 PON1 genotype and phenotype
Infant Neurodevelopment

30

Columbia Cohort Mount Sinai Cohort

U of 

Markers of

Exposure: Chlorpyrifos DAPS TCPy 

Bayley Scores6 months

(MDI/PDI)

‐‐ ‐‐ Null/N

Bayley Scores12 months(MDI/PDI)

Null

/ Null Inverse(*/**)

/ null

(NS) Null/Nu

Bayley Scores24 months(MDI/PDI)

Null

/ Null Inverse(n.s.)/Null Null/ N

Bayley Scores36 months(MDI/PDI)

Inverse/ Inverse ‐‐ 

*PON1 genotype or phenotype; **Among blacks and Hispanics
Attention Problems

31

 Measurement Tools

 Combination Maternal report and dire

 Child Behavior Checklist (maternal report Direct Assessment of child (K-CPT) Hillside Psychometrician report

 DSVM-IV ADHD symptoms

 Assessed at 3, 3.5 and 5 years in two 
Attention Problems

32

Sensitivity of test at 3 yrs limitedConsistent results , two different exposuresPrenatal lead, lack postnatal

Columbia

Cohort

Mo

S

Co

Exposure: CPF D

(PDD) Positive (36 mo.) 

Odds of ADHD/

attentionproblems

Positive

(36 mo.)
Intelligence Measures 

33

 Wechslar Intelligence Scale for Child

 Verbal comprehension Perceptual Reasoning Working Memory (WM) Processing Speed Full Scale IQ (FSIQ)

 Standardized US population, norms E

Spanish speaking populations

 Cultural Bias has been noted

 Restricted sample population
Intelligence Measures

34

Consistent results across studies (WM)Large number missing tests, include/exclude comparable

Columbia

Cohort

Mount 

Coh

Exposure CPF DA

MentalDevelopment

(WISC

‐IV)

Inverse

(FSIQ, WM)

Invers

(FS

perce

reaso

WM
Draft Framework Analysis

35 Evaluation and Synthes
Strengths and Limitations

 Measurement outcome and 

exposure extensive, validated, reliable

 Important confounding 

variables measured and modeled

 Design and analysis decisions 

aid comparability between groups



R

educe man

y sources of bias



Peer-revie

wed publications 

 Unknown fac

with CPF 

 However, 

hypothesizpotential bias, lack observed 

 In general, d

SES in any e

 Restriction

level multimoderate

 Role of post

 Limited ins

measured

36

Strengths Lim
Study Design Issues

 A priori hypotheses  Statistical modeling 

appropriate

 Discussed varying 

degrees across studies

 Missing data imputed 

when possible

 Selection 

extensive

 Enhanc Reduce 

(genera

 High pre

exposureenhanced 

 Limited 

interactc

hemica

37
Statistical Analysis and Gr

 Several Methods Used across studies:

 Motivated by sample size, study hypoth Grouping straight-forward, fully expla Analysis issues not always discussed in f

 Several sensitivity analyses performe

 QA/QC measures Alternative hypotheses explored Model fitting exercises, shape of data

38
Exposure Measurement

39

 Chlorpyrifos (parent), non-specific (TCPy

 Lack specificity, direct comparability May explain heterogeneity 

 Unknown temporal, spatial pattern of e

 Constant, intermittent, and/or sporadic One-time measure correlated with longer-

 Meconium=maternal urine (ave.)=cord blood

 Cohorts’ exposures greater than, but co

population median

 Generalizable to US population
Outcome Ascertainment

40

 Outcome Measures

 Similar, aids comparison Clinical, research settings Spanish/English Quality Assurance and Control adequate

 Stated observers blinded in some, not all stud

 Limitations

 Neurodevelopmental assessments subjectiv Cultural bias –Wechslar tool

 Non-differential error likely

 Multiple exposure ameliorates potential d
Confounding Bias

41

 Multiple methods of control

 Restriction, individual- and group-level 

 Cannot exclude possibility positive co

of unknown factor

 Lead, mercury, other OP, OC, PCB, PAH SES

 Multi-dimensional, both +/- confounding b Hierarchal regression analysis utilized to 

 Lack examination of heterogeneity by 

environmental exposures
Threats to Validity

 Retention rate 

moderately high for birth cohorts (60-80% at 7 years)

 Included and excluded 

comparable

 Measured variables 

 Missing Data imputed

 Errors in 

exposure occurred

 Design 

differen

 Many e

 Non-diffe

have redsizes

42

Selection Bias Inform
Synthesis of Evidence

43

 Strength of observed associations hig

cases, but not all

 Non-differential exposure measuremen

reduce magnitude

 Some evidence of exposure-response  Consistency in Brazelton (abnormal re

ADHD-like behavior problems, Bayley MDI), and IQ (WM)

 Alternative explanations investigated

 Modeling, transformations, confounding 
Synthesis of Evidence

44

 Prospective studies

 Temporal relation Differential misclassification less likely 

 Biological plausibility

 See other sections, generally plausible, 

details MOA

 Influence of cessation of residential e

association

 Limited evidence (lo

w po

w

er), not desig
Conclusions

45

 Current database strengthens conclus

“chlorpyrifos likely plays a role” in obadverse effects on child neurodevelo

 Given study conduct, design, methods 

more likely under- than over-estimate 

 Integration across streams of informa

continue
Section 4.0

Appendices 4-6

46 Questions
Appendix 4: Supplementary 

Columbia U. Cohort Bounding

 See Hattis PBPK

 How many “LBW?”

 Not designed to 

address

 Bayley Scores

 Expanded two to four 

categories, highest category greatest risk

 Loss to f

 Non-d

exposu

 Pre-canc

clarifica

 Exposu

 Confoun

 Diazin

47