Aspirin Supplementation for Pregnancy Indicated Risk Reduction in Nulliparas (ASPIRIN): Neurodevelopmental Follow-up Trial
Conditions: Neurodevelopmental Abnormality
Sponsor: NICHD Global Network for Women's and Children's Health

 
 
ASPIRIN: 
NEURODEVELOPMENTAL 
FOLLOW-UP TRIAL 
NCT04888377 

Aspirin Follow-up Protocol V 1.1 
July 30, 2021 
 Aspirin Supplementation for Pregnancy Indicated Risk 
Reduction in Nulliparas (ASPIRIN): Neurodevelopmental 
Follow-up Trial 
 
ClinicalTrials.gov number: 
NCT04888377 
 
CTRI Trial Registration number: 2021/06/034432 
 
Protocol number: 
GN-LDA.FU 0.1 
 
Lead Study Investigator(s): 
Matthew Kendall Hoffman, MD, MPH-Christiana Care 
 
 
 
 
Richard J. Derman, MD, MPH-Thomas Jefferson University 
Shivaprasad S. Goudar, MD, MHPE, KLE Academy of Higher  
Education and Research, Jawaharlal Nehru Medical College 
 
 
       
Subcommittee Members:  
Tracy Nolen, DrPH, RTI International 
 
 
 
 
Elizabeth McClure, PhD, RTI International 
 
 
 
 
Norman Goco, MHS, RTI International 
 
Version Number: 
Version 1.1 
 
Version Date: 
July 30, 2021  
 
Funding:  
Thrasher Research Fund 
 
 
 
 
 
 

Aspirin ND Follow-Up Protocol V1.1 
 
 
July 30, 2021 
 
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Version Tracking 
Version 
Date 
Authors 
Comments 
1.0 
April 23, 2021 
MH 
First approved version 
1.1 
July 30, 2021 
AJW 
Updated NCT Number, 
Added CTRI Number, 
renamed IYCF index 
form. 

 
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Table of Contents 
ASPIRIN Neurodevelopmental Follow-up  Investigators……………………………………………………….……5 
ACRONYMS……………………………………………………………………………………………………………………………….8 
Abstract……………..…………………………………………………………………………………………………………………….9 
1. 
  Statement of Problem………….……………………………………….……………………….…………………..11 
1.2         Primary Hypothesis or Question…………………………………………………………………………..…….11 
1.2         Epidemiology……………………………………………………………………………………………….…………….11 
1.3         Antecedent of Neurodevelopment……….…………………………………………………..…..…………..11 
1.4         Implementation of Aspirin on Neurodevelopment…………………………..………….…………….12 
1.4.1      Biologic Rationale…………….…………………………………………………………………………..……….…..12 
1.4.2   Hypertensive Disorders of Pregnancy and Neurodevelopmental Delay………..…....………12 
1.4.3   Clinical Data of Neurodevelopment and Fetal Exposure to LDA…………..…..……..………….13 
1.4.3.1  Systematic Follow-up of Prospective RCT's of LDA……………………………….…………………….13 
1.4.3.2   Population-based Cohorts……….…………………………………………………………………………………13 
1.4.3.3  Summary of These Trials and the Potential to Improve Neurological Outcomes…………14 
1.4.3.4  General Safety of Aspirin during Pregnancy……………………………………………………………..…14 
1.5         Population Implications for the Changing Patters of Perinatal Aspirin Use………………….15 
1.6        Public Health Implications………………………………………..………………………………………………….15         
1.7 
   Summary………………………………………………………………………………………………………………….….15 
2.          Methods……………………………………………………………………………………………………..……………….16 
2.1 
   Study Design……………………………………………………………….………………………….……………………16 
2.2 
   Study Population…………………………………………………………………………………………………………16 
2.2.1     Inclusion Criiteria………………………………………………………………………………….…………………….16 
2.2.2     Exclusion Criteria…………………………………………………………………………………………………….…..16 
2.3 
   Sequence of Study Activities………………………………………………………………………………………..16 
2.4 
   Detailed Study Procedures…………………………………………………………………………………………..17 

 
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2.4.1     Initial Screening……………………………………………………………….………………………………………….17 
2.4.2      Blinding/Masking……………………………………………………………………………………………………….18 
2.4.3      Location of Participants .................................................................................................. 18 
2.4.4      
Consenting…………………………………………………………………………………………………………………188 
2.4.5      Screening for Medical Eligibility and Anthropometic Measurements………………………….19 
2.4.6      Performance of the BSID-III Assessment…………………………………………………………………….19 
2.4.7      Ages & Stages Questionnaire-3: ..................................................................................... 20 
2.4.8      Family Resources and Context Questionnaire…………………………………………………………….20    
2.4.9      Infant Feeding & Breastfeeding History: ........................................................................ 20 
2.4.10    Referral Process for Children Found to be At Risk. ........................................................ 20 
2.4.11    Study Governance and Logistics ..................................................................................... 20 
2.4.12    Role of Senior Foreign Investigators. .............................................................................. 21 
2.4.13    Institutional Review Boards and Trial Registration…………………………………….…………......21 
2.4.14    Data Monitoring Plan…………………………………………………………………………………………………21 
2.4.15    Training and implementation of Neurodevelopmental Testing:. ................................... 22 
2.4.16    Data Management Procedures…………………………………………………….………………….….…....22 
2.5          Primary Outcome………………………………………………………………………….……………………………23 
2.6 
    Safety Monitoring……………………………………………………..………………………………………..………23 
2.7 
    Site Preparation………………………………….……………………………………………………….………..……23 
2.8 
    Potential Risks and Benefits to Participants……………….…………………………………………..…..24 
3.            Analytical Plan………………………………………………………………………………………………….…….. . .25 
3.1 
    Statistical Analysis Plan………………….……………………………..…………………………………………….25 
3.1.1      Primary Analysis…………………………………………………………………………………..……………….…...25 
3.1.2      Secondary and Exploratory Analysis .............................................................................. .25 
3.2 
    Sample Size and Power Estimates………………………………………………..….…………………….…..25 
3.3 
    Available Population…………………………………………………………………………………………………..26 
3.4 
    Projected Recruitment Tme…………………………………………………………………………….………….27 

 
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3.5 
    Study Monitoring Plan…………………………………………………………………………………….………….27 
3.5.1      Reporting Serious Adverse Events .................................................................................. 27 
3.5.2      Method and Timing for Reporting Serious Adverse Events ............................................ 27 
3.5.3      Data Monitoring Plan………………………………………………………………………………………………...28 
3.8.4      Data Management Procedures.………………………………………………………………………………….28 
3.6 
    Quality Control……………………………………………………………………………………………………………28 
3.6.1      Training………………………………………………………………………………………………….…………………..28 
3.6.2      Study Monitoring ............................................................................................................ 29 
4.           Data Forms ...................................................................................................................... 30 
5. 
References ........................................................................................................................ 31 
Appendix 1. Sample Informed Consent........................................................................................ 35 
 
 

 
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ASPIRIN NEURODEVELOPMENT FOLLOW-UP TRIAL INVESTIGATORS 
Central Team: Global Network Site 08 (Belagavi, India) 
 
Richard Derman, MD, MPH 
Principal Investigator 
Thomas Jefferson University 
Philadelphia, PA 
Richard.Derman@jefferson.edu  
 
Shivaprasad S. Goudar, MD, MHPE 
Senior Foreign Investigator 
KLE Academy of Higher Education and Research, Jawaharlal Nehru Medical College 
Belagavi, India 
sgoudar@jnmc.edu 
 
Matthew K. Hoffman, MD, MPH 
Lead Investigator for Aspirin Protocol 
Christiana Care  
Newark, Delaware 
Mhoffman@christianacare.org 
Site Investigators 
 
Global Network Site 02 (Democratic Republic of Congo) 
Melissa Bauserman, M.D. 
 
 
 
 
. 
 
 
 
Principal Investigator 
University of North Carolina School of Medicine 
 
 
 
 
Chapel Hill, North Carolina 
 
 
 
 
 
mbauserman@med.unc.edu  
 
Antoinette Tshefu, M.D, Ph.D., M.P.H  
Senior Foreign Investigator 
Kinshasa School of Public Health 
antotshe@yahoo.com 
 
Global Network Site 03 (Zambia) 
Wally Carlo, MD 
Principal Investigator 
University of Alabama at Birmingham 
Birmingham, Alabama 
Wcarlo@PEDS.UAB.EDU 
 
Elwyn Chomba, MBChB, DCH, MRCP 
Senior Foreign Investigator 
University Teaching Hospital 
Lusaka, Zambia 
echomba@zamnet.zm 

 
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Network Site 06 (Guatemala)  
Nancy Krebs, MD 
Principal Investigator 
UCHSC 
Denver, Colorado 
Nancy.krebs@ucdenver.edu 
Manolo Mazariegos, MD  
Senior Foreign Investigator 
INCAP 
Guatemala City, Guatemala 
mmazariegos@incap.int  
 
Global Network Site 09 (Pakistan) 
Robert L. Goldenberg, MD 
Principal Investigator 
Columbia University 
New York, New York 
 
 
 
 
 
rlg88@columbia.edu 
 
 
 
Sarah Saleem, MD 
Senior Foreign Investigator 
Aga Khan University 
Karachi, Pakistan 
Sarah.saleem@aku.edu 
 
Global Network Site 11 (Nagpur, India) 
Patricia L. Hibberd, MD, PhD 
Principal Investigator 
Boston University 
Boston, Massachusetts  
plh0@bu.edu                                                 
 
Archana Patel, MD, DNB, MSCE 
Senior Foreign Investigator 
Lata Medical Research Foundation 
Nagpur, India 
Dr_apatel@yahoo.com 
RTI International 
 
Elizabeth McClure, PhD 
Principal Investigator, Data Coordinating Center 
mcclure@rti.org 
 
Tracy Nolen, DrPH 
Co-Principal Investigator, Data Coordinating Center 
Senior Statistician 
tnolen@rti.org 

 
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Janet Moore, BS 
Statistician, Data Coordinating Center 
Research Statistician, III 
jbartz@rti.org 
 
Norman Goco, MHS 
Protocol Manager for Aspirin Protocol 
ngoco@rti.org 
 
Alexis Williams, BS 
Project Coordinator, Data Coordinating Center 
Clinical Study Specialist 
arjones@rti.org 
 
Consultants  
Michele Lobo, PhD 
Expert in administration of Bayley examinations and oversight 
malobo@udel.edu  
 

 
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ACRONYMS 
ACOG  
American Congress of Obstetricians and Gynecologists  
ASA 
 
Acetylsalicylic acid 
ASPIRIN 
Aspirin Supplementation for Prevention of Inherent Risk in Nulliparas Trial 
ASQ 
 
Ages & Stages Questionnaire® Third Edition 
BSID-III  
Bayley Scales of Infant Development-III 
BSID-IV  
Bayley Scales of Infant Development-IV 
DCC 
 
Data Coordinating Center 
DMC 
 
Data monitoring committee 
DMS 
 
Data Management System 
DRC 
 
Democratic Republic of Congo 
ERC 
 
Ethical review Committee 
GA 
 
Gestational age 
GN 
 
Global Network for Women’s and Children’s Health Research 
HDP 
 
Hypertensive disorders of pregnancy 
JNMC  
Jawaharlal Nehru Medical College 
IRB 
 
Institutional Review Board 
IUGR 
 
Intrauterine growth restriction 
LBW 
 
Low birth weight   
LDA 
 
Low dose aspirin 
LMIC 
 
Low- and middle-income country 
LMP 
 
Last menstrual period 
MDI 
 
Mental Development Index 
MNH 
 
Maternal and Newborn Health 
NICHD  
Eunice Kennedy Shriver National Institute of Child Health and Human Development 
NIH 
 
National Institutes of Health 
NSAID   
Non-steroidal anti-inflammatory drug 
PIH 
 
Pregnancy induced hypertension 
PMNCH 
The Partnership for Maternal, Newborn, & Child Health 
PROM   
Premature rupture of membranes 
PTB 
 
Preterm birth 
RCT 
 
Randomized controlled trial 
RTI 
 
Research Triangle Institute International 
SAE 
 
Serious adverse event 
SFI 
 
Senior Foreign Investigator 
SGA 
 
Small for gestational age 
SPTB   
Spontaneous preterm birth 
VLBW  
Very low birth weight 
WHO 
 
World Health Organization 
 
 

 
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ABSTRACT 
Background: Over 52.9 million children under five years of age in low and middle-income countries 
(LMIC) are estimated to have neurodevelopmental delays1. Though neurodevelopmental delay is 
complex in its origins, preterm birth2, hypertensive disorders of pregnancy (HDP)3 and chronic 
inflammation4 are known primary drivers. Recently the ASPIRIN5 trial demonstrated that low dose 
aspirin (LDA) administered antenatally decreases the rate of preterm birth, preterm (HDP) and perinatal 
mortality. LDA has also long been known to decrease chronic inflammation. Therefore, antenatal LDA 
may prevent neurodevelopmental delay through its effect on preventing preterm birth & HDP and 
inhibiting chronic inflammation. Currently data of neurodevelopmental outcomes following antenatal 
LDA exposure can be summarized as: 1. multiple (37+) observation trials that have linked maternal 
inflammation with developmental delays/behavioral issues4, 2. Numerous studies showing that offspring 
of HDP pregnancies are at significant risk of cognitive delay3, 3. follow-up trials of children whose 
mothers were part of an antenatal LDA trial that the assessment occurred too early (e.g. 12 or 18 
months) or used outdated assessments (e.g. parental perception)  but show either no difference or 
improvement6–8 and 4. registry studies that have associated maternal LDA use with slight increases in 
cerebral palsy9 and hearing loss10. Despite the recent positive findings of the ASPIRIN trial, broad 
acceptance of low dose aspirin (LDA) may be tempered by concerns about effects on longitudinal 
neurodevelopment.  In contrast, if LDA is found to mitigate these risks, women would be further 
motivated to take LDA and professional societies to recommend LDA as a universal part of prenatal care.  
Hypothesis(es) and Aims:  
Primary Hypothesis:   Children exposed antenatally to low dose aspirin will have scores on the Bayley 
Scales of Infant Development-III (BSID-III) examination at 36 months of life(+/-3months) that are not 
inferior to their peers who were not exposed (i.e., by no more than a margin of 4 points). 
Sub-Hypothesis: Conditional on showing non-inferiority is that children exposed antenatally to low dose 
aspirin will have higher scores on the BSID-III examination compared to their peers who were not 
exposed.  
Secondary Hypothesis: The secondary hypothesis will be that children exposed antenatally to low 
dose aspirin will have similar scores on the Ages & Stage’s Questionnaire® Third Edition (ASQ-
3). 
Sub-Hypothesis: Conditional on showing non-inferiority is that children exposed antenatally to low dose 
aspirin will have higher scores on the ASQ-3 examination compared to their peers who were not 
exposed.  
Design: This will be a prospective masked matched cohort study of children between 33 and 39 months 
(mean 36 months) of age whose mothers were randomized in the ASPIRIN trial (1:1 Aspirin-Placebo), 
who will be evaluated using the BSID-III. Additionally, the Family Resources and Context questionnaire 
will be performed to adjust for the local context and the ASQ-3 will be administered as a secondary 
screen. Recognizing the significant role that preterm birth plays in neurodevelopment we will include 
100 (50 in each group) children who were delivered before 37 weeks. 

 
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Inclusion Criteria:  The following are the inclusion criteria: 1. Children whose mothers participated in the 
ASPIRIN trial whose parents/guardians are able to consent, 2. No evidence of  congenital anomalies that 
would affect development (e.g., blindness) and 3. Other conditions that would preclude them from 
completing a BSID-III. 
 
Exclusion Criteria: Children not meeting the inclusion criteria will be excluded.  Likewise, children may 
be excluded at the discretion of the site investigator out of concern for the child’s wellbeing (e.g. 
distress with testing). 
Power & Sample Size: Assuming the BSID-III score has a mean scaled score of 100 and standard 
deviation of 15, a non-inferiority margin of 4 points is clinically significant, and the true effect of LDA is 
not more than a 1-point decrease, a total sample size of 620 children with a 1:1 allocation (310 in each 
group) will have 80% power for a one-sided test for non-inferiority with α=0.05.  
Primary Outcome: The primary outcome will be the results of the BSID-III- specifically the motor, 
cognitive and language components. 
Potential Impact: The ASPIRIN trial results prove that LDA can improve maternal-child outcomes; 
nonetheless, women often underestimate their risk of preterm birth and are reticent to take medication 
during pregnancy. By demonstrating the safety and potential superiority of antenatal LDA on 
neurodevelopment, women will be more willing to accept LDA as part of routine care and obstetrical 
providers and countries to recommend its routine use.  
 

 
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1 STATEMENT OF PROBLEM 
1.1 PRIMARY HYPOTHESIS OR QUESTION: 
Children exposed antenatally to low dose aspirin will have scores on the Bayley Scales of Infant 
Development-III (BSID-III) examination at 36 months of life (+/-3months) that are not inferior to 
their peers who were not exposed (i.e., by no more than a margin of 4 points). 
1.2 EPIDEMIOLOGY:  
An estimated 250 million children under the age of 511 worldwide  are at risk for not achieving their 
developmental potential11; 52.9 million children under five years of age  in low- and middle-income 
country (LMIC) settings have neurodevelopmental delays1.  Compounding the issue is preterm birth 
(more common in LMICs 12) which  has consistently been identified as a cause of 
neurodevelopmental delay 2,13. A recent review reported that out of the estimated 13 million 
preterm infants who survive beyond the first month, 0.9 million will suffer long term 
neurodevelopmental impairment, with 345,000 moderately or severely affected 14.   This burden 
places a significant strain on the families, healthcare systems and societies that provide care for 
these children.   
1.3    ANTECEDENTS OF NEURODEVELOPMENT:   
Neurodevelopment of children is the aggregate outcome of complex exposures, both antenatally 
and post-delivery.  Antenatal factors such as maternal nutrition15, inflammation16, and stress may 
all play important roles in both cognitive development of children and attainment of optimal 
growth17,18.  As a single risk factor, preterm birth is strongly linked to abnormal growth, cognition 
and motor development 14,19 20. Inflammation, which is a common pathway of preterm birth21, has 
been further implicated in neurodevelopment. In a review of 37 papers, Nist and Pickler16 identified 
a series of inflammatory mediators (interleukin (IL)-6, IL-1β, IL-8, and tumor necrosis factor (TNF)-α) 
that were all associated with neurodevelopmental delay in children.   
Specific to LMIC settings,  Jiang and colleagues examined known markers of inflammation (C-
Reactive Protein, Soluble CD-14, IL-6 & Others) and found a negative correlation between elevated 
levels of inflammatory markers and outcomes of the Bayley exam22,23.  Additionally, they noted that 
elevated IL-4 levels (which help differentiate T Helper cells) were positively correlated with 
improved neurological outcomes.  
Poor nutrition and growth limitations in the fetus/infant have also been associated with 
impairments in neurodevelopment 15. In Guatemala, a cohort study found stunting, which is 
reflective of perinatal nutrition, to be prevalent in 47% of newborns; 53% at 3 months of age and 
56% at 6 months of age 24. This study concluded that growth failure in infants was predicted by 
growth failure in fetal life, thus indicating the importance of maternal interventions. Data from 
other Global Network participating sites (Guatemala, Democratic Republic of Congo, Zambia and 
Pakistan) also found strikingly high rates of stunting ranging from 44% to 66%, among infants and 
toddlers 25. Malnutrition, which also includes being overweight and obese, are known inflammatory 
processes that can potentially impact neurodevelopment.  These conditions are increasingly more 

 
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common to low and middle income countries 26.  Other newborn exposures including exclusive 
breastfeeding, stunting, as well as caregiver factors such as maternal/parental education, mental 
health, and home environment. Poverty additionally contributes to the attainment of optimal 
neurodevelopment.  As such, any study of neurodevelopment should at least document these 
potential confounders. 
1.4 IMPLICATIONS OF ASPIRIN ON NEURODEVELOPMENT:  
1.4.1 Biologic Rationale: 
In a review of 37 papers, Nist and Pickler16 identified a series of inflammatory mediators 
(interleukin (IL)-6, IL-1β, IL-8, and tumor necrosis factor (TNF)-α & others ) that were all associated 
with neurodevelopmental delay in children. Specifically both inflammatory mediators and 
hypertensive disorders have been shown to negatively affect neutrophins  and other neuroproteins 
that are critical in the neuronal programming and development27. Thus, there is direct biological 
plausibility of maternal inflammation hindering normal child neurodevelopment.  Of particular 
interest is that many of these same inflammatory mediators have been implicated as playing crucial 
roles in spontaneous preterm birth and maternal hypertensive disorders of pregnancy (HDP).    
Aspirin has been shown to predominantly affect both the COX-1 pathway which is involved in 
thrombosis and the COX-2 pathway, which affects inflammation through the production of Aspirin 
Triggered Lipoxins 28,29  Various Lipoxins have been shown to affect placental development and play 
roles in the development of preeclampsia and preterm birth30. Lipoxin A4 has been particularly 
singled out in playing a crucial role in placental inflammation and acts not only to inhibit new 
inflammation but to aid in the resolution of existing inflammation. More specifically, aspirin has 
been shown to inhibit the production of IL-6, IL-1B, CRP and TNF-α all of which have been shown to 
negatively affect child neurodevelopment and be involved in preeclampsia and preterm birth.  Of 
equal interest, Aspirin inhibition of COX-2 has been shown to increase the production of 
neuroprotection DI and  Resolvin D1 which have been suggested to lower the risk of Alzheimer’s 
disease and ischemic brain injury31.   
1.4.2 Hypertensive Disorders of Pregnancy and Neurodevelopmental Delay:  
The origins of prescribing LDA in pregnancy began with an attempt to decrease hypertensive 
disorders of pregnancy.  Through studies of over 50,000 pregnant women5,32, it has now become 
well established that LDA decreases the risk of both term and preterm HDP. The offspring of 
women whose pregnancies are complicated by preeclampsia/HDP have consistently shown both 
short and long term neurodevelopmental delays and behavioral issues regardless of the gestational 
age at birth3,33.  Nonetheless, studies showing that the prescription of LDA during pregnancy 
improves poor neurodevelopmental outcomes due to HDP are absent.  
 
 

 
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1.4.3  Clinical Data of Neurodevelopment and Fetal Exposure to LDA:  
1.4.3.1  Systematic Follow-up of Prospective RCT’s of LDA:  
Data regarding fetal exposure to LDA is noted to consist of the following: 
• The largest study to examine longitudinal neurological follow up of children born to women 
randomized to prenatal aspirin was a follow up trial of children whose mothers participated in 
the United Kingdom’s Collaborative Low-dose Aspirin Study in Pregnancy (CLASP) trial that 
included 9,364 women.  Amongst this cohort, the child’s medical practitioner provided 
information on development for 4,168 children at 12 months of life and 4,365 parents 
answered a standardized questionnaire at 18 months of age.  No differences in development, 
motor function or anthropometric measurements were identified 8. 
• Marret and colleagues examined 656 preterm children (125 LDA exposed) born to 584 women 
before 33 weeks who were enrolled in a French RCT of prophylactic LDA 6.  They examined 
children using the Kaufman Assessment Battery for Children at 5 years of age.   The percent of 
low simultaneous processing scores (<70) was lower in the group with LDA (7% vs 19% without 
LDA; P = .04). They also found that LDA exposure trended towards improvement in total 
behavior difficulties (aOR: 0.44 [95% CI: 0.19-1.02]) and hyperactivity (aOR: 0.43 [95% CI: 0.17-
1.05]).  
• In a letter to the Lancet, Parazzini et. al. provided information regarding neurodevelopment of  
788 children (427 aspirin exposed) whose mothers had participated in the Italian Aspirin in 
Pregnancy Study 34. They provided a questionnaire to parents of children at 18 months that 
addressed both neurodevelopment (Sonnander’s questionnaire) and fetal growth.  Though 
statistical analysis was not provided, the presented tables provide adequate information to 
perform simple univariate analysis.  Children exposed to aspirin had lower rates for less than  
the 10th  percentile for height (8.4% vs. 12.2%-P = 0.047) and they had significantly improved 
gross and fine motor function (3.5% vs. 7.2%-P=0.02).  Weight at less than the 10th percentile 
and language attainment were similar. These results must be viewed cautiously as the trial was 
not blinded, relied on maternal report and 27% of the children were lost to follow up. 
 
1.4.3.2  Population-based Cohorts:  
• The Extremely Low Gestational Age Newborns (ELGAN) study examined 877 extremely 
premature (<33 weeks) and surveyed mothers about antenatal exposures at the time of birth. 
Both neonatal ultrasound of the neonatal brain and neurological follow up at 24 months of 
corrected age was performed. The neurological follow up was standardized and blinded but did 
not use a widely recognized methodology.35  An association with maternal aspirin use was 
associated with a slightly increased risk of quadriparetic cerebral palsy following adjustment for 
multiple confounders (OR 3.0, 95% CI 1.3 to 6.9)36.   Data regarding the duration of exposure, 
dosage taken and indication for use was not presented.  The total number of women who took 
LDA was noted to be 49. There was no association with LDA and ultrasound findings that have 
been linked to cerebral palsy nor any association with diparetic cerebral palsy.  
• Petersen and colleagues examined both prenatal exposures to analgesics (acetaminophen, 
ibuprofen, and aspirin) and the risk of cerebral palsy by examining two large linked Dutch 
cohort studies (Norwegian Mother and Child Cohort Study and Danish National Birth Cohort)9.  
They examined the risk of cerebral palsy by exposure amongst 185,617 children.   They found 
that aspirin exposure was associated with an elevated risk of bilateral spastic cerebral palsy 
(aOR 2.4, 95% CI: 1.1–5.3). Nonetheless the data were limited by very low rates of antenatal 

 
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LDA exposure (3.1%).  In contrast, acetaminophen use occurred in 49% of women and was 
associated with a OR of 1.3 (95% CI:1.0–1.7) for unilateral cerebral palsy.  A significant 
limitation of this study is that these associations may be confounded by indication.  Both 
acetaminophen and aspirin are commonly utilized to treat fever. Infections in pregnancy have 
been consistently associated with cerebral palsy36. 
• Foch et al recently published a case control study examining maternal drug exposures during 
pregnancy comparing children who either had a normal hearing evaluation (N=28,046) or an 
abnormal hearing evaluation (N=1,245).  They found that children who had an abnormal 
hearing loss were more likely to have had an in utero exposure to LDA (OR 1.61, 95%CI 1.09-
2.37)10.      
 
1.4.3.3  Summary of These Trials and the Potential to Improve Neurological Outcomes:  
As detailed above, there is biologic plausibility and limited clinical evidence that by mediating 
inflammation and preterm birth during pregnancy, we may be able to improve neurodevelopmental 
outcomes.  However, there is conflicting information on the potential of harm versus benefit 
regarding motor function and hearing.  Though several studies have suggested harm, they are 
confounded by indication and report small increases of risk of rare conditions.   Nonetheless the 
aggregate data are sparse, utilize short duration of follow-up, are not based on well accepted 
instruments of development, do not account for potential confounding factors and rely 
predominantly on indirect observation.  None of these studies have utilized the BSID-III, the 
currently best validated and accepted method for evaluating neurodevelopment.  It is equally 
important to address that perinatal inflammation may be unique in LMIC communities, wherein the 
incidence and causes of inflammation may be different. Similarly, cultural variation may also have 
unique implications for differences in longitudinal development.   
The Global Network’s ASPIRIN study provides a unique opportunity to study the impact of LDA with 
earlier and validated exposure and infant outcomes at 33 to 39 months of age. This follow up study 
will likewise provide the infrastructure and a stable cohort to potentially examine children’s 
outcomes at 5 years of age, when issues like behavior and IQ can be more accurately assessed. In 
summary, the follow-up of infants born to participants in the ASPRIN trial will allow us to: a) analyze 
the safety of low dose aspirin during pregnancy relative to  neonatal health, b) study the impact of 
LDA on neurodevelopment, growth and morbidity, and  c) potentially establish a longitudinal cohort 
to allow meaningful exploration of issues related to child behavior in future investigations. 
1.4.3.4  General Safety of Aspirin During Pregnancy:  
To date, in excess of over 50,000 women have participated in research trials of aspirin37, 5.  Based on 
the available data, aspirin appears to have a favorable safety profile for both mothers 38 and their 
newborns 39.  Specifically in terms of the neonate, differences in anomalies 40, post-natal intracranial 
hemorrhage or complications in circumcision have not been detected despite robust numbers being 
reported 39. More recently, follow-up safety data from the ASPIRIN trial demonstrated comparable 
adverse events and usage of medical services among women who took aspirin compared to those 
who took placebo.  Finally, an analysis of the impact of aspirin randomization on hemoglobin 
revealed no impact amongst women enrolled in the ASPIRIN study41. 
 

 
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1.5  POPULATION IMPLICATIONS FOR THE CHANGING PATTERNS OF PRENATAL ASPIRIN USE: 
The necessity to examine the impact of LDA on longitudinal development is fueled by the fact that 
the utilization of LDA during pregnancy is poised to dramatically change. Low-dose aspirin has 
increasingly been recommended by professional societies (ACOG42 and Royal College of Obstetricians 
and Gynaecologists) and the World Health Organization (WHO) for women at risk for preeclampsia 
during pregnancy 43.  Following the recent publication of the ASPRE trial (Combined Multimarker 
Screening and Randomized Patient Treatment with Aspirin for Evidence-Based Preeclampsia 
Prevention)44 and the current Global Network’s ASPIRIN trial5, the number of potentially exposed 
newborns is anticipated to grow.  
1.6  PUBLIC HEALTH IMPLICATIONS:  
Although the benefits of LDA in preventing preeclampsia and now preterm birth have been proven, 
the ultimate public health impact will be decided by the willingness of pregnant women to choose 
and comply with this therapy.  Though complex and influenced by familial and cultural inputs, these 
decisions are largely based upon the individual patient’s perception of their risk/benefit ratio.  Prior 
research has demonstrated that mothers (particularly first-time mothers) underestimate or minimize 
their risks and have a strong reluctance to take ongoing medications during pregnancy45.  While both 
current recommendations and outcomes from the ASPRE and the ASPIRIN trial suggest significant 
fetal benefit, to fully inform individual patient decision making, systematic longitudinal follow-up 
studies are warranted. If LDA is found to lack detrimental effects for offspring or to have potential 
cognitive benefits, women will be much more likely to view low dose aspirin as an acceptable 
intervention. 
1.7 SUMMARY: 
Though the recent ASPRE and ASPIRIN trials have established low dose aspirin as a strong 
preventative measure in the prevention of the major causes of maternal and perinatal morbidity 
(preterm birth and preterm preeclampsia), the implications on longitudinal newborn outcome remain 
less clear.  
 

 
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2 METHODS 
2.1 STUDY DESIGN: 
We propose a matched (1:1) cohort study of children whose mothers were randomized to either low 
dose aspirin or placebo.  As preterm infants are at higher risk they will be oversampled (N=100) and 
matched for gestational age within one +week and equally distributed between mothers who 
received LDA and those who received placebo. 
2.2 STUDY POPULATION:   
The study population will consist of approximately 620 children, between 33 and 39 months of age, 
whose mothers were included in the ASPIRIN trial.   Among those, one hundred children who were 
born preterm (<37 weeks) will be selected as they have a higher risk of developmental delays.  They 
will be equally distributed between the LDA and placebo arms.  They will be matched by site and 
within 1 week of delivery.  
2.2.1 Inclusion Criteria:  
The following are the inclusion criteria:  
a. For a prior study sponsored by the Global Network for Women’s and Children’s Health 
Research, the child’s parents gave permission to recontact. 
b. The child’s parent/guardian is willing and able to provide consent for the child and willing also 
to participate in the child’s assessment (e.g., by providing answers to ASQ-3 and the Family 
Resources and Context questionnaire). 
c. The child will be between 33 and 39 months of age at time of assessment. 
d. The child does not have a significant congenital anomaly (e.g., blindness or deafness) that 
would affect development or preclude them from completing the BSID-III. This will be left at 
the discretion of the Senior Foreign Investigator (SFI) with the ability to consult with the 
Working Committee as needed. 
e. Other significant medical conditions that would preclude the child from completing a BSID III 
are absent. This will be left to the discretion of the Senior Foreign Investigator who will have 
the ability to consult with the Working Committee as needed. 
 
2.2.2 Exclusion Criteria: 
Children not meeting the inclusion criteria will be excluded.  Likewise, children may be excluded at 
the discretion of the site investigator out of concern for the child’s wellbeing (e.g., the child has a 
current respiratory infection or emotional distress during testing). 
2.3 SEQUENCE OF STUDY ACTIVITIES: 
The sequence of study activities is shown in Figure 1.  
 

 
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Figure 1. Flowchart of Study Activities
 
2.4 DETAILED STUDY PROCEDURES: 
2.4.1  Initial Screening: 
Initial screening will be done by the Data Coordinating Center (DCC).  ASPIRIN trial participants who 
agreed in their initial consent to be contacted for a future study will be identified by the DCC and 
this information will be supplied to the local sites.  Secondly, the DCC will review the delivery dates 
to identify children who are near 36-months of age.  Assuming that both criteria are met, the DCC 
will examine whether the child’s mother was in the aspirin group or the placebo arm.  Finally, the 
gestational ages will be reviewed.  The DCC will issue pairs of eligible children (1 Aspirin: 1 Placebo) 
which are matched within the site and are matched in gestational age within one week.  The DCC 
will track successful recruitment of pairs.  When successful recruitment of a pair cannot be 
obtained, a child meeting the gestational age, allocation and site requirements will be substituted 
to ensure that balance occurs across these groups.  As children will be older than 2 years of age, we 
will not correct for preterm birth.  

 
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2.4.2 Blinding/Masking: 
 
Throughout the study, research staff and local health providers will be blinded to treatment status 
of the mother.  Data regarding the subject’s allocation will be held and protected centrally and will 
not be made available to participants or their families.  
2.4.3 Location of Participants: 
As contact information was obtained during the initial study, this will be utilized to reach out to 
participants.  Local centers will determine the optimal method of contact.  This can include in 
person visits or phone calls.  If the parents/guardians decline participation, no further efforts will 
be made to encourage study participation.  To ensure that research is not coercive, no more than 
three attempts at contacting potential participants will be made. 
2.4.4 Consenting: 
Before a child participates in any research activity, the research staff must obtain the informed 
consent to voluntarily take part in the study from the child’s mother, father or guardian. Consent 
will be obtained from parents or guardians ≥ 18 years of age or minors 14-17 years of age in 
countries where married or pregnant minors (or their authorized representatives) are legally 
permitted to give consent.  When approaching guardians who are minors, we will follow the in-
country policies for human research protection and the guidelines approved by the local ethical 
review committees (ERCs). In the case of minors, this may require that written consent is obtained 
from a parent/guardian or husband, with written assent from the minor guardian.   
The research staff will give adequate opportunity to the individual providing consent for a child’s 
participation and/or immediate family members to read the consent form and ask questions. 
Recognizing that literacy levels will vary and may be a challenge, the consent process will include a 
verbal review of the consent form. If the individual requested to provide consent cannot read, the 
form will be read aloud to the individual by a person unaffiliated with the study. Alternatively, the 
Program Coordinator or a designate may read the consent, but in the presence of a witness who is 
unaffiliated with the research study. Potential consenters will be given an opportunity to discuss 
the study procedures and ask questions. Fair balance will be maintained while describing the risks 
and benefits of participation in the study.  No undue pressure will be placed on the potential 
consenter to enroll the child in the trial.  It will further be explained that lack of participation will 
not affect the usual and anticipated standard of care.  
If the individual providing consent (a parent/guardian) is unable to sign their name, he/she will be 
asked to use her/his thumbprint to indicate written approval. In both cases, the unaffiliated person 
will also sign the consent form.  Both the research staff and the study participant retain signed 
copies of the form.  The potential participant may also take the form home to discuss with the 
family before signing. 
The parent/guardian may refuse to agree to the child’s participation in the trial at the time of 
recruitment. This will be recorded in the Screening and Recruitment Form. He/she may also choose 
withdrawal from the study at any time after enrollment. This will be recorded on study forms.  
Similarly, if an enrolled child becomes ineligible prior to assessment, the child will be withdrawn 
from the study and this will be recorded. 

 
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All research staff responsible for obtaining consent will be trained and certified in the protection of 
human subjects and the study-specific consent procedures. A model written informed consent 
form, developed according to the requirements of the U.S. Office of Research Protections (OHRP), 
is found in Appendix 1.  Each site may modify the model consent to conform to local standards, but 
the OHRP required elements must be maintained. The research sites will also be responsible for 
translating the consent form into the appropriate language(s) for their local context. 
Global Network countries with legislation regarding the need to videotape consents will comply 
with the country regulations; however, this is not part of the consent form requirements. This will 
not be required by protocol but rather decided by each site so as to comply with country rules and 
regulations. 
2.4.5 Screening for Medical Eligibility & Anthropometric Measurements:  
Upon completion of the consenting process, an assessment of medical eligibility will be undertaken 
using the Medical History form.  This form contains a basic medical history and immediate 
assessment to ensure that the child has no significant medical problems that would preclude or 
interfere with the performance of the evaluation. Similarly, if the site coordinator is concerned 
about the child’s wellbeing on the date of assessment, she/he can either be rescheduled for a time 
that is deemed more appropriate or removed from the study.  Examples would include an acute 
rhinitis, asthma exacerbation or emotionally distressing event. If the child is scheduled more than 
two times but not assessed, the child will be excluded from the study. Decisions about medical 
eligibility will be made by the SFI and, if requested, in collaboration with the working committee. 
In addition, anthropometric measurements will be recorded on this form.  Measurements will be 
done by qualified staff who will be trained in accordance with prior protocols using standardized 
equipment.  Those who were train in the Women First: Preconception Nutrition protocol will be 
considered as certified. 
2.4.6 Performance of the BSID-III Assessment: 
The BSID-III assessment will be performed by only trained and qualified examiners.  To be 
considered trained, an examiner will have been qualified in a prior study that utilized the BSID-III 
assessment forms or a health professional experienced in use of the Bayley assessment tools.  If 
not previously trained, examiners must be willing to be trained and undergo a period of 
observation. Specifically, we will be performing the motor, cognitive and language components of 
the BSID-III. We will not include the social and emotional domains intentionally for the following 
reasons: 1. The social and emotional domains are lengthy and therefore result in significant 
participant burden and may negatively impact ascertainment of the other three domains,  2. The 
available qualified and trained staff have historically not performed these components and to 
introduce them now would be a significant challenge, and 3. The internal reliability of these 
sections are subject to greater social/cultural influences, particularly in a study involving different 
countries/culture. Results of the BSID-III will be recorded on the BSID-III assessment form. 
2.4.7 Ages & Stages Questionnaire-3: 
The Ages & Stages Questionnaire 3 (ASQ-3) is a broadly accepted and validated screening tool for 
neurodevelopment. Like the BSID-III, ASQ-3 has been utilized and validated in multiple languages 

 
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and cultural contexts.  The ASQ-3 will be administered by research staff to parents.  Where literacy 
issues may be present, staff will be trained on how to help administer the questionnaire to obtain 
accurate information without introducing bias.  Following completion of the questionnaire by the 
child’s parents, staff, will score the questionnaire and those results will be recorded on the ASQ 
form. 
2.4.8 Family Care Indicators Questionnaire:  
This questionnaire is based upon family care indicators developed in 2002 with UNICEF 
involvement for the purpose of assessing fundamental practices or qualities that support and 
encourage learning and development.  The Family Care Indicators questionnaire was developed to 
be implemented across diverse countries and different cultural backgrounds. Moreover, it has been 
explicitly field-tested and validated for this purpose46.  The questionnaire also has questions to 
judge psycho-social wellbeing developed by UNICEF working groups for a psychosocial evaluation 
project47. The results of this will be recorded on the Family Care Indicators form. 
2.4.9 Infant andYoung Child Feeding & Breastfeeding History:  
The Infant Feeding Index form will be administered by the assessment staff. This form will collect 
information on breastfeeding history, feeding practices, and food frequency, and recent dietary 
history. 
2.4.10 Referral Process for Children Found to be at Risk:  
Children who have a BSID-III score below <85% or an ASQ-3 below 2 standard deviations will be 
noted, and their parent(s) provided with this information.  Because complex factors can affect 
individual scores, parents will be encouraged to consider further evaluation of their child.  
2.4.11 Study Governance and Logistics: 
Oversight of the trial will be handled by two principal groups with different focuses: 
a. Composition and Role of the Steering Committee: The steering committee (SC) is comprised of 
the DCC, the BSID-III quality oversight individual, and Global Network Senior Foreign 
Investigators and US investigators associated with each of the participating sites (see pages 5-7). 
The steering committee will have primary responsibility for overall study design, approval of 
study materials and procedures, and oversight of study implementation. The SC will convene via 
conference call at least once per quarter and in person (pandemic permitting) in conjunction 
with a Global Network Steering Committee meeting to discuss the study and  implementation 
issues.  During the early phases of the study more frequent meetings may be needed.  If the 
budget allows, members of the group may conduct site visits to bolster enthusiasm, provide 
hands-on training and education to the participating staff, and address site-specific issues, if 
any. 
b. Composition of the Working Group:  The working group will consist of the lead investigators 
(Drs. Goudar, Hoffman, and Derman), the BSID-quality consultant (Dr. Michele Lobo), and 
representatives of RTI International, including the Data Coordinating Center.  They will meet via 
conference call at least monthly to monitor study progress and ensure proper implementation 
of the trial. They will have primary responsibility of the following tasks: 

 
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i. Protocol development and revision as needed 
ii. Manual of operations development and revision as needed 
iii. Oversight of data forms and database development 
iv. Tracking ethics approvals and renewals 
v. Monitoring recruitment and individual site progress 
vi. Monitoring of BSID-III examiners 
vii. Ongoing review of data quality 
viii. Review of serious adverse events and protocol violations 
ix. Budgetary oversight 
x. Review of findings and analysis 
xi. Manuscript drafting and submission 
xii. Coordination with the steering committee 
xiii. Handling of any and all operational concerns  
 
Members of the group may  also conduct site visits to research sites, as the budget allows, to 
bolster enthusiasm, provide hands-on training and education to the participating staff, and 
address site-specific issues, if any. 
2.4.12 Role of Senior Foreign Investigators: 
The Senior Foreign Investigators (SFIs) will be responsible for developing site-specific 
implementation plans, ensuring study staff are properly trained, and providing oversight of the 
study at the site level.  As members of the study SC, convene via conference call at least once per 
quarter and meet in person with other members of the study’s Steering Committee (if it is feasible 
to take advantage of an in-person Global Network Steering Committee meeting) to discuss study 
implementation issues. 
2.4.13 Institutional Review Boards and Trial Registration:   
Human subjects review will be undertaken by the IRB of Thomas Jefferson University and the 
ethical review committees of the foreign sites.  This effort will be coordinated across the varying 
groups by the Data Coordinating Center.  To ensure transparency, the study will be registered at 
Clinicaltrials.gov and at other required sites as stipulated by review committees of the foreign 
sites. 
2.4.14   Data Monitoring Plan: 
All participating foreign sites will transmit data to the Data Coordinating Center, located at RTI 
International. The data will be used to evaluate protocol adherence and site performance (e.g., 
recruitment, loss to follow-up, data quality). The DCC will provide standardized progress reports to 
Steering committee and the site investigators monthly to monitor outcome variables and data 
audits/edits. 

 
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 2.4.15  Training and Implementation of Neurodevelopmental Testing: 
Significant experience and trained staff already exist at a number of the sites due to participation 
in the BRAIN HIT trial48 and the Women First: Preconception Maternal Nutrition trial49.  Both trials 
had qualified individuals and the infrastructure to ensure training and ongoing validity of testing.    
Additionally, it is recognized that individuals may have been trained in the performance of these 
types of tests as part of clinical practice.  
BSID-III Training: To be qualified as a BSID-III examiner for the purpose of this study, the examiner 
must meet the following criteria: 
a. Demonstrate credentialling or consistent experience in administering Bayley Scales of 
Infant Development: 
• 
Prior successful administration of BSID-II or BSID-III in a Global Network study 
(BRAIN-HIT or Women First) or in another research trial using the Bayley Scales 
• 
Certification of training in administration of Bayley Scales by other approved means 
• 
Use of the BSID-III in association with clinical care  
b.  Attend either a virtual webinar on performance or watch a recording of a webinar on 
correct performance of the BSID-III featuring key metrics 
BSID-III Monitoring: Key to ensuring that high quality BSID-III testing is being done, the following 
provisions have been developed to ensure longitudinal follow up occurs.  This is noted to include 
the following: 
a. BSID-III examiners are asked to film one of their first 5 BSID-III examinations.  This is to 
be labeled with the participant number, examiner number and no other 
identifiers.  Feedback will be provided by the central BSID-III study credentialler.  The 
credentialler will provide feedback to the examiner in each of the domains used for this 
study that are designed to serve as positive coaching. 
b. Following each review, the credentialler will then determine the window to the next 
reevaluation.  The choices for duration can be either one within the next 5 exams or the 
next 10. She/he may also choose to have the next cycle of reevaluations performed by 
either the site lead for BSID-III training or by the central credentialler. 
c. If after 2 cycles significant concerns are raised by the central credentialler, the examiner 
may be asked not to continue in the study.  
Key to the success of this approach is to take a positive coaching approach as has been utilized in 
the past.  This process will use an IT infrastructure and central review process that has been 
utilized in a number of prior studies of both neurodevelopment48 and ultrasound50 in the recent 
past. 
2.4.16  Data Management Procedures: 
Data will be collected prospectively using hard copy forms or Android tablets. Regardless of data 
capture methodology, all data will be kept confidential. Each participant will be assigned a unique 
study ID which will be used to identify the participant.  Only the screening log will contain the 
name (which is not transmitted). If hard copy forms are used, they will be retained in a secure 
location for possible editing or queries at the central data entry site.  Data will be entered into 
computers using the Data Management System (DMS) developed by RTI and the assigned study 
number.  The DMS will also allow site staff to produce project reports and backup the study 

 
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database.  Electronic data will be transferred from each data management computer to a single 
Research Unit Data Center (RUDC) in each country, creating a complete data repository.  At least 
once a week, data will be transmitted from the RUDC to the DCC at RTI.  The data center will 
conduct training on the DMS system, as needed, and will maintain the central database for the 
study. Precision and accuracy of actual data collected will be checked by chart review (random 5%) 
and internal procedures using the computer program.  Monthly audits and incomplete data 
reports will be performed by a review team consisting of at least of the SFI and the country 
coordinator.  Data editing and error resolution will be performed monthly.  In addition, a sample 
of participants will be visited to confirm their participation, with procedures determined per site. 
These activities will be shared between the site and the data center.  
2.5  PRIMARY OUTCOME: 
The BSID-III provides systematic assessment of development in the three primary domains of 
cognition, language (receptive and expressive) and motor skills (fine & gross).  The BSID-III forms 
for these domains are validated tools which can be used for assessment of children at about 36 
months of age and are well accepted by the research community.   Equally important, this tool has 
been validated in a diverse group of languages and cultures including low/middle income 
countries 51. Specifically, Bayley forms have been validated in the following countries of relevance:  
Democratic Republic of Congo, Pakistan, Zambia, India and Guatemala. 
Foundational to utilizing the Bayley forms will be a requirement for both demonstration of 
competence in performing the examination and an ongoing assessment of correct implementation 
through periodic reassessment.  For this proposed trial we will utilize the existing staff, 
credentialing process and oversight mechanisms of the Women First: Preconception Maternal 
Nutrition trial.  
2.6 SAFETY MONITORING:   
The risk implicit in the BSID-III is noted to be minimal.  Little risk is anticipated.  Nonetheless, we 
will monitor non-completed studies, and case report forms will include any adverse events that 
have occurred during the visit.  This will include an open-ended question about issues that 
occurred during the visit. Review will occur on an ongoing basis.  Adverse events will be submitted 
on Serious Adverse Event forms. 
2.7 SITE PREPARATION: 
In preparation for study implementation, the sites will meet with health authorities and conduct 
community sensitization activities to ensure that study procedures are appropriate for the local 
context and to encourage commitment and engagement at the facility and community level. Site  
 
 

 
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preparation activities will focus on: 
• Identifying and hiring study staff; 
• Identifying potential implementation challenges and developing culturally-appropriate 
solutions;. 
• Educating health workers and community members on the purpose of this investigation. 
2.8   POTENTIAL RISKS AND BENEFITS TO PARTICIPANTS: 
No direct benefits to the current participants are noted beyond possible consideration for time 
and effort as allowed by local IRB standards.  The risks are noted to be nominal as no invasive 
procedures are planned.  
 
 

 
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3 ANALYTICAL PLAN 
3.1  STATISTICAL ANALYSIS PLAN: 
Prior to conducting formal analyses baseline demographic characteristics and key clinical measures 
will be compared between the mothers who participated in the two treatment arms using 
contingency table approaches for categorical variables and analysis of variance models and t-tests for 
continuous variables.  For all of these analyses, comparisons will be made within Global Network sites 
and overall across the sites controlling for site in the models.  
3.1.1  Primary Analysis:  
The primary outcome of BSID-III score is assumed to have a mean scaled score of 100 and standard 
deviation of 1547.  Assuming a non-inferiority margin of 4 points is clinically significant, and the true 
effect of LDA is not more than a 1 point decrease, a total sample size of 620 children with a 1:1 
allocation (310 in each group will have 80% power for a one-sided test for non-inferiority with 
α=0.05.  Conditional on non-inferiority being observed, this sample size will also provide over 90% 
power at α=0.05 to detect a different of 4 points between LDA exposed and non-exposed children 
based on a two-sided test for the secondary hypothesis. 
3.1.2  Secondary and Exploratory Analysis: 
In addition to the planned formal primary analysis, we also anticipate conducting exploratory 
outcomes for a number of other binary outcomes.  The analytic approach for these exploratory 
analyses, will be comparable to that planned for the primary and formal secondary analyses but will 
focus on estimation of effect sizes and generation of hypotheses.  Initially, contingency tables will be 
used to generate estimates of the risk of each outcome for the two treatment arms, both overall and 
separately by site and across all sites, and the relative risk associated with treatment will be 
estimated by site and across sites controlling for site. For each outcome a series of generalized linear 
models analogous to those described for the primary outcome to generate unadjusted estimates of 
risk, estimates of risk adjusted for potential confounders, and estimates of risk adjusted to account 
for any possible differences in risk between the arms. 
3.2  SAMPLE SIZE AND POWER ESTIMATES 
The primary outcome of BSID-III score is assumed to have a mean scaled score of 100 and standard 
deviation of 1547.  The second hypothesis of neurocognitive benefit of LDA will only be assessed 
conditional on a finding of non-inferiority for the primary hypothesis; thus, no further adjustment for 
multiplicity is required. Assuming a non-inferiority margin of 4 points is clinically significant, and the 
true effect of LDA is not more than a 1 point decrease, a  total sample size of 620 children with a 1:1 
allocation (310 in each group will have 80% power for a one-sided test for non-inferiority with 
α=0.05.  Conditional on non-inferiority being observed, this sample size will also provide over 90% 
power at α=0.05 to detect a different of 4 points between LDA exposed and non-exposed children 
based on a two-sided test for the secondary hypothesis. 

 
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3.3  AVAILABLE POPULATION: 
Children available across the sites will be identified as noted above.  The number of available children 
at different time periods is graphically displayed below (Figure 2). As the work will occur over time, 
the pool of children who will potentially age out will grow over the duration of the study.  Thus, the 
intent is to prioritize those children who are closer to aging out of the study parameters and those 
who were born prematurely. This will ensure the greatest chance of successful recruitment.  
Similarly, as a guiding principle, the attempt will be to try and identify children who are 36 months or 
greater to minimize variation in child age that may impact testing.  
 
 
Numbers of available children by site are displayed in Table 1 which follows. 
 
4,094
3,611
3,194
2,822
2,458
2,077
1,591
1,126
702
276
31
0
0
500
1,000
1,500
2,000
2,500
3,000
3,500
4,000
4,500
July-21 August-21
September-21
October-21
November-21
December-21
January-22
February-22March-22 April-22 May-22 June-22
Figure 2: 39 Month Children Available
1 
 Jul-21     Aug-21   Sep-21    Oct-21    Nov-21  Dec-21   Jan-22    Feb-22   Mar-22  Apr-22  May-22   Jun-21 

 
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3.4 PROJECTED RECRUITMENT TIME: 
Each site will begin recruitment after their site-specific ethical and regulatory approvals are in place, 
the site has been adequately prepared, and training is completed. This will require a staggered start, 
as the timeline for these activities will vary per site. The projected timeline for this study is two years 
with over 1 year dedicated for recruitment. This includes a 2-month preparatory phase for regulatory 
approvals, 2 months for BSID training, a 12-month recruitment period, time for data cleaning and 
analysis. A graphical representation of the timeline is found below: 
 
3.5 STUDY MONITORING PLAN: 
3.5.1 Reporting Serious Adverse Events: 
Serious Adverse events (SAEs) will be monitored continuously using a special form that will be 
required for any event that meets one or more of the following criteria: results in death, is life-
threatening, requires hospitalization or prolongs existing hospitalization, results in persistent or 
significant disability or incapacity, any other serious or unexpected adverse event that the study 
investigator(s) feels should be reported. 
3.5.2 Method and Timing for Reporting Serious Adverse Events 
The Senior Foreign Investigator (SFI) must report the following SAEs by transmitting the form to RTI 
as follows: 
Within 48 hours of SFI’s notification of the event: 
• All child deaths  
• All SAEs with a definite or suspected/probable relationship to the intervention 
 
Within 7 days of SFI’s notification of the event:  
• All life-threatening events 
• All SAEs considered to have a probable or possible relationship to the intervention. 
 

 
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All completed SAE forms should also be entered into the DMS and transmitted within 7 days as a 
back-up to ensure no SAE is missed. Additional reporting procedures include: 
• RTI will forward all SAEs to the US-based Principal Investigator (PI) for further assessment of 
relationship to study intervention.  
• The PI and SFI will be responsible for reporting to their respective IRB and other regulatory 
authorities per their institutional policy.  
• RTI will be responsible for reporting SAEs to the DMC bi-annually at a minimum. The frequency 
of reporting to the DMC may be increased if the reported events or interim data reviews by the 
DMC indicate that more frequent safety monitoring is needed. 
 
Any SAE considered unrelated to the intervention is not required to be reported in an expedited 
manner.  These events should be entered into the data management system and transmitted per 
routine procedures. 
3.5.3 Data Monitoring Plan: 
All participating sites will report data to the Data Coordinating Center, located at RTI International. 
The data will be used to evaluate protocol adherence and site performance (e.g., recruitment, and 
data quality). The DCC will provide standardized progress reports to the steering committee and the 
site investigators on a monthly basis to monitor outcome variables and adverse events. 
3.5.4 Data Management Procedures: 
Data will be collected prospectively using hard copy forms or Android tablets. Regardless of data 
capture methodology, all data will be kept confidential. Each participant will be assigned a unique 
study ID which will be used to identify the participant.  Only the screening log will contain the name 
(which is not transmitted). If hard copy forms are used, they will be retained in a secure location for 
possible editing or queries at the central data entry site.  Data will be entered into computers using 
the Data Management System (DMS) developed by RTI and the assigned study number.  The DMS 
will also allow site staff to produce project reports and backup the study database.  Electronic data 
will be transferred from each data management computer to a single Research Unit Data Center 
(RUDC) in each country, creating a complete data repository.  At least once a week, data will be 
transmitted from the RUDC to the DCC at RTI. 
 
The data center will conduct training on the DMS system, as needed, and will maintain the central 
database for the study. Precision and accuracy of actual data collected will be checked by chart 
review (random 5%) and internal procedures using the computer program.  Monthly audits and 
incomplete data reports will be performed by a review team consisting of at least the SFI and the 
country coordinator.  Data editing and error resolution will be performed monthly.  In addition, a 
sample of participants will be visited to confirm their participation, with procedures determined per 
site. These activities will be shared between the site and the data center. 
3.6 QUALITY CONTROL: 
3.6.1  Training: 
All study personnel must participate in training on the proper implementation of study procedures 
and the ethics of conducting research with human subjects before beginning any research activity. 
The SFI and project coordinator will ensure that all study personnel receive the appropriate training, 

 
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and obtain the required certification ensuring that they have met the training objectives. RTI will be 
responsible for developing a certification test and BSID-III proficiency. The SFI and project manager 
will be responsible for overseeing the certification process. 
3.6.2  Study Monitoring: 
Major monitoring responsibilities of the PI/SFI, assisted by the country coordinator, are (1) 
confirming IRB approval; (2) monitoring the recruitment of subjects; (3) assessing and evaluating the 
quality of study implementation;  (5) evaluating accuracy, precision, and completeness of data 
collected, entered, and transmitted (along with the DCC); (6) ensuring that all personnel are fulfilling 
their obligations; (7) maintaining morale and enthusiasm of the staff; (8) handling ad hoc problems 
and maintaining communication; (9) ensuring inter-site consistency; and (10) proposing 
improvements to the monitoring activities. 
 

 
Aspirin ND Follow-Up Protocol V1.1 
 
 
July 30, 2021 
4 DATA FORMS 
The following forms will be used for this study: 
Form Description 
Purpose 
Key Data Elements 
Data Source 
 
Screening Log 
To track screening and enrollment at 
facility or community level 
Contact information, screening and 
enrollment date and status 
 
Mother Report 
 
Screening 
To determine eligibility and record 
consent status 
 
Consent and inclusion/exclusion criterion 
 
Clinician Report 
 
Medical History  
To determine medical history and 
record anthropometric measures 
Limited medical history by parent report, 
height and weight measurement of child 
Parent or 
Guardian Report  
 
 
Infant and Young 
Child Feeding 
Index 
To determine the maternal 
breastfeeding experience and types 
of food the child receives 
 
Feeding practices, frequency of feedings, 
breastfeeding information 
 
Parent or 
Guardian Report 
 
BSID-III 
 
To record outcomes of BSID-III 
 
Scoring and elements of the BSID-III 
 
BSID-III 
 
ASQ-3 
To record outcomes of Ages and 
Stages Questionaire-3. 
 
Scoring and elements of the ASQ-3 
Parent or 
Guardian Report 
 
Family Care 
Indicators 
To record family care indicators and  
resources to understand the family 
context 
 
Parent report of family resources and 
context 
Parent or 
Guardian Report 
Serious Adverse 
Events  
To record and report serious adverse 
events 
 
Description of Serious Adverse Events 
Parent or 
Guardian Report 
Closeout Form 
To record participants completion 
status of study 
Final form that records if participant 
completed or withdrew from the study 
Study Staff 
 
 

 
Aspirin ND Follow-Up Protocol V1.1 
 
 
July 30, 2021 
5 REFERENCES 
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Moster D, Lie RT, Markestad T. Long-Term Medical and Social Consequences of Preterm Birth. N 
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Sun BZ, Moster D, Harmon QE, Wilcox AJ. Association of Preeclampsia in Term Births with 
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Nist MD, Pickler RH, Steward DK, Harrison TM, Shoben AB. Inflammatory Mediators of Stress 
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Hoffman MK, Goudar SS, Kodkany BS, et al. Low-Dose Aspirin for the Prevention of Preterm 
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Marret S, Marchand L, Kaminski M, et al. Prenatal low-dose aspirin and neurobehavioral 
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Parazzini F, Bortolus R, Chatenoud L, Restelli S, Benedetto C. Follow-up of children in the Italian 
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X 
8.  
Low dose aspirin in pregnancy and early childhood development: follow up of the collaborative 
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Petersen TG, Liew Z, Andersen A-MN, et al. Use of paracetamol, ibuprofen or aspirin in pregnancy 
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Foch C, Araujo M, Weckel A, et al. In utero drug exposure and hearing impairment in 2-year-old 
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11.  
Black MM, Walker SP, Fernald LCH, et al. Early childhood development coming of age: science 
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12.  
March of Dimes. Born too soon: The Global Action Report on Preterm Birth. CP Howson, MV 
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Reuner G, Hassenpflug A, Pietz J, Philippi H. Long-term development of low-risk low birth weight 
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14.  
Blencowe H, Lee AC, Cousens S, et al. Preterm birth–associated neurodevelopmental impairment 
estimates at regional and global levels for 2010. Pediatr Res. 2013;74(december):17-34. 
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15.  
Vohr BR, Poggi Davis E, Wanke CA, Krebs NF. Neurodevelopment: The Impact of Nutrition and 
Inflammation During Preconception and Pregnancy in Low-Resource Settings. Pediatrics. 2017. 
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16.  
Nist MD, Pickler RH. An Integrative Review of Cytokine/Chemokine Predictors of 
Neurodevelopment in Preterm Infants. Biol Res Nurs. May 2019:109980041985276. 
doi:10.1177/1099800419852766 
17.  
Krebs NF, Lozoff B, Georgieff MK. Neurodevelopment: The Impact of Nutrition and Inflammation 
During Infancy in Low-Resource Settings. Pediatrics. 2017;139(Supplement 1):S50-S58. 
doi:10.1542/peds.2016-2828G 
18.  
Marques AH, Bjørke-Monsen A-L, Teixeira AL, Silverman MN. Maternal stress, nutrition and 
physical activity: Impact on immune function, CNS development and psychopathology. Brain Res. 
2015;1617:28-46. doi:10.1016/j.brainres.2014.10.051 
19.  
Saigal S, Doyle LW. An overview of mortality and sequelae of preterm birth from infancy to 
adulthood. Lancet. 2008;371(9608):261-269. doi:10.1016/S0140-6736(08)60136-1 
20.  
Goldenberg RL, Rouse D. Prevention of preterm birth, Review Article. N Engl J Med. 
1998;339(5):313-320. 
21.  
Lockwood CJ, Kuczynski E. Risk stratification and pathological mechanisms in preterm delivery. 
Paediatr Perinat Epidemiol. 2001;15 Suppl 2:78-89. 
22.  
Ma JZ, Tofail F, Jiang NM, et al. Early Life Inflammation and Neurodevelopmental Outcome in 
Bangladeshi Infants Growing Up in Adversity. Am J Trop Med Hyg. 2017;97(3):974-979. 
doi:10.4269/ajtmh.17-0083 
23.  
Jiang NM, Tofail F, Moonah SN, et al. Febrile illness and pro-inflammatory cytokines are 
associated with lower neurodevelopmental scores in Bangladeshi infants living in poverty. BMC 
Pediatr. 2014;14(1):50. doi:10.1186/1471-2431-14-50 
24.  
Berngard SC, Berngard JB, Krebs NF, et al. Newborn length predicts early infant linear growth 
retardation and disproportionately high weight gain in a low-income population. Early Hum Dev. 
2013;89(12). doi:10.1016/j.earlhumdev.2013.09.008 
25.  
Krebs NF, Mazariegos M, Tshefu A, et al. Meat consumption is associated with less stunting 
among toddlers in four diverse low-income settings. Food Nutr Bull. 2011;32(3):185-191. 
doi:10.1177/156482651103200301 
26.  
Hambidge KM, Krebs NF, Garcés A, et al. Anthropometric indices for non-pregnant women of 
childbearing age differ widely among four low-middle income populations. BMC Public Health. 
2017;18(1):1-12. doi:10.1186/s12889-017-4509-z 
27.  
Sahay A, Kale A, Joshi S. Role of neurotrophins in pregnancy and offspring brain development. 
Neuropeptides. 2020;83. doi:10.1016/j.npep.2020.102075 
28.  
Cadavid AP. Aspirin: The mechanism of action revisited in the context of pregnancy 
complications. Front Immunol. 2017;8(MAR). doi:10.3389/fimmu.2017.00261 
29.  
Gil-Villa AM, Alvarez AM, Velásquez-Berrío M, Rojas-López M, Cadavid J AP. Role of aspirin-
triggered lipoxin A4, aspirin, and salicylic acid in the modulation of the oxidative and 
inflammatory responses induced by plasma from women with pre-eclampsia. Am J Reprod 
Immunol. 2020;83(2). doi:10.1111/aji.13207 
30.  
Szczuko M, Palma J, Kikut J, Komorniak N, Ziętek M. Changes of lipoxin levels during pregnancy 

 
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and the monthly-cycle, condition the normal course of pregnancy or pathology. Inflamm Res. 
2020;69(9):869-881. doi:10.1007/s00011-020-01358-6 
31.  
Miyazawa K, Fukunaga H, Tatewaki Y, et al. Alzheimer’s disease and specialized pro-resolving lipid 
mediators: Do MAR1, RVD1, and NPD1 show promise for prevention and treatment? Int J Mol Sci. 
2020;21(16):1-14. doi:10.3390/ijms21165783 
32.  
Duley L, Meher S, Hunter KE, Seidler AL, Askie LM. Antiplatelet agents for preventing pre-
eclampsia and its complications. Cochrane Database Syst Rev. 2019;2019(10). 
doi:10.1002/14651858.CD004659.pub3 
33.  
Zwertbroek EF, Zwertbroek J, Broekhuijsen K, et al. Neonatal developmental and behavioral 
outcomes of immediate delivery versus expectant monitoring in mild hypertensive disorders of 
pregnancy: 5-year outcomes of the HYPITAT II trial. Eur J Obstet Gynecol Reprod Biol. 
2020;244:172-179. doi:10.1016/j.ejogrb.2019.11.001 
34.  
Bortolus R, Chatenoud L, Restelli S, Di Cintio E, Parazzini F. [Follow-up of children included in the 
Italian Study on the Use of Low-Dose Aspirin During Pregnancy: development at 18 months of life 
and birth weight]. Pediatr Med Chir. 20(6):387-391. 
http://www.ncbi.nlm.nih.gov/pubmed/10335537. Accessed December 3, 2017. 
35. 
Tyler CP, Paneth N, Allred EN, Hintz D, Kuban K, McElrath T, O'Shea TM, Miller C, Leviton A, For 
the ELGAN Study Investigators. Brain damage in preterm newborns and maternal medication:the 
ELGAN Study.  Am J Obstet Gynecol. 2012;207:192.e1-9.  Published online 2012 Jul 
7. doi: 10.1016/j.ajog.2012.06.059   
36.  
Miller JE, Pedersen LH, Streja E, et al. Maternal Infections during Pregnancy and Cerebral Palsy: A 
Population-based Cohort Study. Paediatr Perinat Epidemiol. 2013;27(6):542-552. 
doi:10.1111/ppe.12082 
37.  
Knight M, Duley L, Henderson-Smart DJ, King JF. Antiplatelet agents for preventing and treating 
pre-eclampsia. Cochrane database Syst Rev. 2000;(2):CD000492. 
doi:10.1002/14651858.CD000492 
38.  
Ahrens KA, Silver RM, Mumford SL, et al. Complications and Safety of Preconception Low-Dose 
Aspirin Among Women With Prior Pregnancy Losses. Obstet Gynecol. 2016;127(4):689-698. 
doi:10.1097/AOG.0000000000001301 
39.  
LeFevre ML. Low-dose aspirin use for the prevention of morbidity and mortality from 
preeclampsia: U.S. Preventive Services Task Force recommendation statement. Ann Intern Med. 
2014;161(11):819-826. doi:10.7326/M14-1884 
40.  
Kozer E, Nikfar S, Costei A, Boskovic R, Nulman I, Koren G. Aspirin consumption during the first 
trimester of pregnancy and congenital anomalies: a meta-analysis. Am J Obstet Gynecol. 
2002;187(6):1623-1630. http://www.ncbi.nlm.nih.gov/pubmed/12501074. Accessed November 
28, 2013. 
41.  
Jessani S, Saleem S, Hoffman MK, et al. Association of hemoglobin levels in the first trimester and 
at 26 to 30 weeks with fetal and neonatal outcomes: A secondary analyses of the Global Network 
for Women’s and Children’s Health’s ASPIRIN Trial. BJOG. February 2021:1471-0528.16676. 
doi:10.1111/1471-0528.16676 
42.  
ACOG Committee Opinion No. 743: Low-Dose Aspirin Use During Pregnancy. Obstet Gynecol. 
2018;132(1):e44-e52. doi:10.1097/AOG.0000000000002708 
43.  
Bujold E, Roberge S, Lacasse Y, et al. Prevention of Preeclampsia and Intrauterine Growth 

 
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Restriction With Aspirin Started in Early Pregnancy. Obstet Gynecol. 2010;116(2, Part 1):402-414. 
doi:10.1097/AOG.0b013e3181e9322a 
44.  
Rolnik DL, Wright D, Poon LC, et al. Aspirin versus Placebo in Pregnancies at High Risk for Preterm 
Preeclampsia. N Engl J Med. June 2017:NEJMoa1704559. doi:10.1056/NEJMoa1704559 
45.  
D’Souza R, Shah PS, Sander B. Clinical decision analysis in perinatology. Acta Obstet Gynecol 
Scand. November 2017. doi:10.1111/aogs.13264 
46.  
Kariger P, Frongillo EA, Engle P, Britto PMR, Sywulka SM, Menon P. Indicators of family care for 
development for use in multicountry surveys. J Heal Popul Nutr. 2012;30(4):472-486. 
doi:10.3329/jhpn.v30i4.13417 
47. 
United Nations Children's Fund. Interagency Psychosocial Evaluation Report. 2011. United 
Nations Children's Fund: Shuffat, Jerusalem. 
48.  
Bann CM, Wallander JL, Do B, et al. Home-based early intervention and the influence of family 
resources on cognitive development. Pediatrics. 2016;137(4). doi:10.1542/peds.2015-3766 
49.  
Krebs NF, Hambidge KM, Westcott JL, et al. Growth from Birth through Six Months for Infants of 
Mothers in the “Women First” Preconception Maternal Nutrition Trial. J Pediatr. September 
2020. doi:10.1016/j.jpeds.2020.09.032 
50.  
Swanson JO, Plotner D, Franklin HL, et al. Web-based quality assurance process drives 
improvements in obstetric ultrasound in 5 low- and middle-income countries. Glob Heal Sci Pract. 
2016;4(4):675-683. doi:10.9745/GHSP-D-16-00156 
51.  
Semrud-Clikeman M, Romero RAA, Prado EL, Shapiro EG, Bangirana P, John CC. [Formula: see 
text]Selecting measures for the neurodevelopmental assessment of children in low- and middle-
income countries. Child Neuropsychol. 2017;23(7):761-802. doi:10.1080/09297049.2016.1216536 
 

 
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6 APPENDIX 1. SAMPLE INFORMED CONSENT 
Aspirin Supplementation for Pregnancy Indicated Risk Reduction in 
Nulliparas (ASPIRIN): Neurodevelopmental Follow-up Trial 
 
INVESTIGATORS:  
[LIST SITE INVESTIGATORS] 
 
 
SPONSOR: 
Thrasher Research Fund 
 
You are being asked to have your child participate in a research study. This project is funded by the 
Thrasher Research Fund. This form provides you with information about the study. A member of the 
research team will describe the study to you and answer all of your questions. Please read the 
information below and ask questions about anything you don’t understand before deciding whether or 
not to have your child take part. You may also request that the research staff read the form to you.  
 
What is the purpose of the study? 
The purpose of this study is to learn about your child’s development (motor skills, language skills, etc.) 
following the child’s mother having participated in a prior research trial of low dose aspirin during 
pregnancy. Prior trials of taking aspirin during pregnancy have been reassuring on development of 
children; however, the purpose of this study is to obtain information using different tests. 
 
Who will be in the study? 
A total of 620 children will be enrolled in this study from six sites in sub-Saharan Africa, South Asia, and 
Latin America. In [insert site name], no more than 248 children will be enrolled.  Half of the children’s 
mothers will have taken aspirin and half will have taken placebo, a pill that looks identical to aspirin but 
did not contain any medicine.  This will allow the researchers to compare results of the two groups of 
children and determine if aspirin benefitted or had no effect on children and their development. 
Your child qualifies for this study if he or she is a healthy child between 33 months of age of age to 39 
months of age and whose mother participated in the ASPIRIN trial.  
 
What will happen if you join this study? 
Before participating, you will be provided with information about the study procedures and given an 
opportunity to ask questions. If you agree to have your child participate, you will be asked to sign this 
form to indicate your consent. The research staff will ask you some questions about your child’s health, 
and your child’s ability to talk, play and follow commands.  They will also ask your child to do simple 
tasks and evaluate his or her ability to do them. Once these tests are done, this will complete your 
involvement in the study. 
What are the risks and discomforts by being in this study? 
No risks or discomforts are anticipated from this study. Your child will be asked to perform simple 

 
Aspirin ND Follow-Up Protocol V1.1 
 
 
July 30, 2021 
activities.  If your child is uncomfortable doing these activities or tires of them, they will not be required 
to continue.   
What are the benefits of participating? 
You will not receive any money from participating in this study, but your participation may provide 
important information that can be used in the future to prevent babies from being born too soon. Also, 
there is existing evidence that the use of aspirin in pregnancy can improve the infant’s neurological 
outcomes. 
If new information about the benefits or risks of aspirin use in pregnancy becomes available during this 
study, this information will be given to you by [Insert name of Senior Investigator] or his/her staff.  
Will I have to pay for anything? 
It will not cost you anything to be in the study. 
 
Is my participation voluntary? 
Taking part in this study is voluntary. You have the right to refuse to let your child participate or to 
withdraw approval for participation at any time.  If you refuse to allow participation or decide to 
withdraw, you will not lose any benefits or rights to which you are entitled. These actions will not have 
any negative effect on the health care you or the child receive from your local health providers. You and 
the child will have the right to receive normal medical care.   
 
Can I or my child be removed from this study?  
Your child will be withdrawn from the study if the research staff thinks that his or her participation may 
cause him or her harm. The research staff may also remove him or her from the study for other reasons 
at their discretion.  Also, the sponsor may stop the study at any time if there is good reason to do so. 
 
What will happen if you or the participating child are injured by this research? 
Although the risk of injury is expected to be very low, no research can provide an absolute guarantee 
that nothing bad will occur.  Despite safety measures, your child’s participation could result in a reaction 
or injury. If you or your child is injured as a result of study participation, emergency care will be provided 
and a referral made to a doctor for ongoing care, if needed. Ongoing care will not be paid for by the 
study. [Insert name of Research Institution] and the Thrasher Research Fund have not set aside funds to 
pay you for any such reactions, injuries or related medical care.  However, by signing this form, you do 
not give up any of your legal rights.  
What should you do if you have additional questions? 
If you have questions about this study or a project-related injury, you should contact [investigator 
contact]. If you have questions about your or your child’s rights as a project participant, please contact 
[insert ethics committee contact]. 
If you have any questions about the study, please call [insert senior investigator name/contact 
information]. 
Agreement to be in this study 
I have read this paper about the study or it was read to me.  I understand the possible risks and benefits 
of this study.  I know that being in this study is voluntary and I choose to be in this study. I understand I 
will get a copy of this consent form. 
 
Signature (or thumbprint): 
 
 
 
 
________ 
Date: 
 
______  
 
 
 
 
(Mother) 

 
Aspirin ND Follow-Up Protocol V1.1 
 
 
July 30, 2021 
 
Print Name: 
 
 
 
 
 
 
 
__ 
 
 
 
 
(Mother) 
 
Signature (or thumbprint): 
 
 
 
 
_________ 
Date: 
 
______  
 
 
 
 
(Parent/Guardian/Husband) 
 
Print Name: 
 
 
 
 
 
 
 
__ 
 
 
 
 
(Parent/Guardian/Husband) 

 Global Network:  Statistical Analysis Plan, ASPIRIN Follow-up Study  
 
 
   
Version 1.0 06/17/2022; Page i   
 
 
 
 
STATISTICAL ANALYSIS PLAN 
Aspirin Supplementation for Pregnancy Indicated Risk Reduction in 
Nulliparas (ASPIRIN): Neurodevelopmental Follow-up Trial 
 
SAP VERSION: 
Version 1 
 
SAP DATE: 
June 17, 2022 
 
SPONSOR: 
Thrasher Research Fund  
 
PREPARED BY: 
RTI International 
 
 
3040 Cornwallis Rd 
 
 
Research Triangle Park, NC 27709-2104 
 
AUTHOR (S): Tracy Nolen, Marissa Trotta, Janet Moore 
 
 
 
 
 
 
 
 
 
 
 

 Global Network:  Statistical Analysis Plan, ASPIRIN Follow-up Study  
 
 
   
Version 1.0 06/17/2022; Page 2   
 
Contents 
1. 
BACKGROUND AND PROTOCOL HISTORY 
5 
2. 
PURPOSE OF THE ANALYSES 
5 
3. 
STUDY OBJECTIVES AND OUTCOMES 
5 
3.1 
PRIMARY HYPOTHESIS AND ASSOCIATED OUTCOMES 
5 
3.2 
SECONDARY HYPOTHESIS AND ASSOCIATED OUTCOMES 
6 
3.3 
OTHER OUTCOMES OF INTEREST IN THIS STUDY 
6 
4. 
STUDY METHODS 
ERROR! BOOKMARK NOT DEFINED. 
4.1 
OVERALL STUDY DESIGN 
6 
4.2 
STUDY POPULATION 
7 
4.3 
PARTICIPANT SELECTION 
8 
4.4 
MASKING AND DATA LOCK 
8 
5. 
ANALYSIS POPULATIONS 
8 
6. 
SAMPLE SIZE DETERMINATION 
9 
7. 
STATISTICAL / ANALYTICAL ISSUES 
9 
7.1 
GENERAL RULES 
9 
7.2 
ADJUSTMENTS FOR COVARIATES  
10 
7.3 
MISSING DATA APPROACHES: 
10 
7.4 
MULTICENTER STUDIES 
10 
7.5 
MULTIPLE COMPARISONS AND MULTIPLICITY 
10 
7.6 
MASKED DATA REVIEW 
10 
8. 
STUDY PARTICIPANTS, TREATMENT EXPOSURE AND COMPLIANCE 
10 
8.1 
PARTICIPANT DISPOSITION 
10 
8.2 
PROTOCOL DEVIATIONS 
10 
8.3 
DEMOGRAPHIC AND BASELINE CHARACTERISTICS 
10 

 Global Network:  Statistical Analysis Plan, ASPIRIN Follow-up Study  
 
 
   
Version 1.0 06/17/2022; Page 3   
 
9. 
NON-INFERIORITY AND EFFICACY ANALYSES 
11 
9.1 
OVERVIEW OF EFFICACY ANALYSIS METHODS 
11 
9.2 
DEFINITION OF ANALYSIS VARIABLES 
11 
9.4 
PRIMARY ANALYSIS 
14 
9.5 
SECONDARY OUTCOME ANALYSES  
ERROR! BOOKMARK NOT DEFINED. 
9.6 
ANALYSES OF OTHER OUTCOMES OF INTEREST 
ERROR! BOOKMARK NOT DEFINED. 
10. 
SAFETY ANALYSIS: 
14 
11. 
CHANGES TO THE ANALYSES PLANNED IN THE PROTOCOL 
14 
12. 
REFERENCES 
14 
14. 
ATTACHMENTS  
15 
 
 
 

 Global Network:  Statistical Analysis Plan, ASPIRIN Follow-up Study  
 
 
   
Version 1.0 06/17/2022; Page 4   
 
LIST OF ABBREVIATIONS 
Acronym 
Definition 
DCC 
Data Coordinating Center 
DMC 
Data Monitoring Committee 
DRC 
Democratic Republic of the Congo 
EmONC 
Emergency Obstetric and Neonatal Care 
GA 
Gestational Age 
GEE 
Generalized Estimating Equation 
GN 
Global Network 
ITT 
Intention to Treat 
LMP 
Last menstrual period 
MDG 
Millennium Development Goals 
MNH 
Maternal and Neonatal Health 
PP 
Per protocol 
RCT 
Randomized Clinical Trial 
SAP 
Statistical Analysis Plan 
UN 
United Nations 
 
 
 
 

 Global Network:  Statistical Analysis Plan, ASPIRIN Follow-up Study  
 
 
   
Version 1.0 06/17/2022; Page 5   
 
1. BACKGROUND AND PROTOCOL HISTORY 
Background and Rationale 
Over 52.9 million children under five years of age in low and middle-income countries (LMIC) are 
estimated to have neurodevelopmental delays1. Though neurodevelopmental delay is complex in its 
origins, preterm birth2, hypertensive disorders of pregnancy (HDP)3 and chronic inflammation4 are 
known primary drivers. Recently the ASPIRIN5 trial demonstrated that low dose aspirin (LDA) 
administered antenatally decreases the rate of preterm birth, preterm (HDP) and perinatal mortality. 
LDA has also long been known to decrease chronic inflammation. Therefore, antenatal LDA may 
prevent neurodevelopmental delay through its effect on preventing preterm birth & HDP and 
inhibiting chronic inflammation. Currently data of neurodevelopmental outcomes following antenatal 
LDA exposure can be summarized as: 1. multiple (37+) observation trials that have linked maternal 
inflammation with developmental delays/behavioral issues4, 2. Numerous studies showing that 
offspring of HDP pregnancies are at significant risk of cognitive delay3, 3. follow-up trials of children 
whose mothers were part of an antenatal LDA trial that the assessment occurred too early (e.g. 12 or 
18 months) or used outdated assessments (e.g. parental perception)  but show either no difference or 
improvement6–8 and 4. registry studies that have associated maternal LDA use with slight increases in 
cerebral palsy9 and hearing loss10. Despite the recent positive findings of the ASPIRIN trial, broad 
acceptance of low dose aspirin (LDA) may be tempered by concerns about effects on longitudinal 
neurodevelopment.  In contrast, if LDA is found to mitigate these risks, women would be further 
motivated to take LDA and professional societies to recommend LDA as a universal part of prenatal 
care.  
Protocol History 
The protocol was initiated in April 2021.  
No formal interim analyses were planned nor completed for this study and as such, the first formal 
assessment for efficacy is planned for after completion of primary follow-up. 
2. PURPOSE OF THE ANALYSES 
This statistical analysis plan (SAP) contains detailed information about statistical analyses to be 
performed to assess the primary and secondary hypotheses and associated sub-hypotheses outlined in 
the protocol (i.e. children exposed antenatally to low dose aspirin will have a) scores on the Bayley 
Scales of Infant Development-III (BSID-III) examination at 36 months of life (+/-3months) that are 
not inferior to their peers who were not exposed (i.e., by no more than a margin of 4 points) and b) 
similar scores on the Ages & Stage’s Questionnaire® Third Edition (ASQ-3) with sub-hypotheses 
looking for superiority conditional on showing non-inferiority). The results of these analyses will be 
included in the study manuscript(s). Additional exploratory analyses may be performed to support 
further manuscript development.  These analyses will not require an update to the SAP, but 
abbreviated analysis plans will be prepared prior to conducting those analyses. 
 
3. STUDY OBJECTIVES AND OUTCOMES 
3.1 Primary Hypothesis and Associated Outcomes 
Our primary hypothesis is that children exposed antenatally to low dose aspirin will have scores on 
the Bayley Scales of Infant Development-III (BSID-III) examination at 36 months of life(+/-3months) 

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that are not inferior to their peers who were not exposed (i.e., by no more than a margin of 4 points). 
Conditional on showing non-inferiority is that children exposed antenatally to low dose aspirin will 
have higher scores on the BSID-III examination compared to their peers who were not exposed.   
The primary outcome of this study is scores on the BSID-III examination at 36 months of life(+/-
3months). The BSID-III provides systematic assessment of development in the three primary domains 
of cognition, language (receptive and expressive) and motor skills (fine & gross). The examination 
will be performed by only trained and qualified examiners. The cognitive score will be used as the 
primary endpoint with the other scores considered secondary. 
3.2 Secondary Hypothesis and Associated Outcomes 
Our secondary hypothesis is that children exposed antenatally to low dose aspirin will have similar 
scores on the Ages & Stage’s Questionnaire® Third Edition (ASQ-3). Conditional on showing non-
inferiority is that children exposed antenatally to low dose aspirin will have higher scores on the 
ASQ-3 examination compared to their peers who were not exposed.  
The secondary outcome of this study is the scores on the ASQ-3, a broadly accepted and validated 
screening tool for neurodevelopment. The ASQ-3 will be administered by research staff to parents.  
Where literacy issues may be present, staff will be trained on how to help administer the 
questionnaire to obtain accurate information without introducing bias.  Following completion of the 
questionnaire by the child’s parents, staff, will score the questionnaire and those results will be 
recorded on the ASQ form. All ASQ subscales will be analyzed. 
3.3 Other Outcomes of Interest in this Study 
Study analyses will examine a number of additional outcomes to assess whether use of LDA daily 
beginning between 6 0/7 weeks and 13 6/7 weeks has an effect on those outcomes.  Specific maternal 
and fetal/child outcomes may include: anthropomorphic measures, binary classifications for BSID 
and ASQ. 
4. STUDY METHODS 
4.1 Overall Study Design 
For the ASPIRIN Follow-up Study, we designed a cohort study of children whose mothers were 
randomized to either low dose aspirin or placebo in the ASPIRIN trial.  As preterm infants are at 
higher risk they will be over-sampled and the distribution of gestational age will be reviewed to 
ensure approximately equally distribution between mothers who received LDA and those who 
received placebo. Gestational age matching was originally proposed but ultimately determined to be 
operationally challenging given the limited population available for follow-up and short timeline until 
they aged out of ability to participate in this study. 
 
 
 
 

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Study Schematic 
 
4.2 Study Population 
The study population will consist of approximately 620 children, between 33 and 39 months of age, 
whose mothers were included in the ASPIRIN trial.   Among those, children who were born preterm 
(<37 weeks) will be prioritized for inclusion as they have a higher risk of developmental delays.  
Enrollment will be monitored to ensure approximately equal distribution between the LDA and 
placebo arms.   
Inclusion criteria 
• 
For a prior study sponsored by the Global Network for Women’s and Children’s Health 
Research, the child’s parents gave permission to recontact. 
• 
The child’s parent/guardian is willing and able to provide consent for the child and willing 
also to participate in the child’s assessment (e.g., by providing answers to ASQ-3 and the 
Family Resources and Context questionnaire). 
• 
The child will be between 33 and 39 months of age at time of assessment. 
• 
The child does not have a significant congenital anomaly (e.g., blindness or deafness) that 
would affect development or preclude them from completing the BSID-III. This will be left at 

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the discretion of the Senior Foreign Investigator (SFI) with the ability to consult with the 
Working Committee as needed. 
• 
Other significant medical conditions that would preclude the child from completing a BSID 
III are absent. This will be left to the discretion of the Senior Foreign Investigator who will 
have the ability to consult with the Working Committee as needed. 
 
Exclusion criteria 
Children not meeting the inclusion criteria will be excluded.  Likewise, children may be excluded at 
the discretion of the site investigator out of concern for the child’s wellbeing (e.g., the child has a 
current respiratory infection or emotional distress during testing). 
4.3  Participant Selection 
Participant selection was done by the study sites.  However, the Data Coordinating Center provided 
lists of children to prioritize based on gestational age.  Additionally, the Data Coordinating Center 
create and reviewed reports of enrollment by site, sex and gestational age to ensure balanced 
enrollment between treatment groups.  
 
4.4 Masking and Data Lock 
Masking was maintained from the larger ASPIRIN Trial through follow-up. As such, throughout the 
study, research staff and local health providers will be blinded to treatment status of the mother.  Data 
regarding the subject’s allocation will be held and protected centrally and will not be made available 
to participants or their families.  
Data Lock: Data will be locked at the site level with final analysis data sets generated and locked 
once data from all sites have been received and all queries processed.  For each site, data collection 
for the protocol will stop approximately 2 months after the last child was randomized into the study at 
that site. Site will transmit all data forms to the central site within one months of this date, and data 
will be locked two to three months post last follow-up visit. The study will not be unmasked until 
after data lock. 
5. ANALYSIS POPULATIONS 
Intention to Treat (ITT) Population  
The primary analysis population will be the intention to treat population, which includes all 
eligible and enrolled follow-up participants.  All participants will be assigned to the arm to which 
they were randomized irrespective of treatment received. Unless specified otherwise, all participants 
with data available will be included in analyses using this population. Missing data will be considered 
missing at random and not included.  This population will also be used for many of the secondary 
analyses. 
Per Protocol (PP) Population  
This population is based on the main study PP population and will be used to conduct sensitivity 
analyses for the primary outcome. This population will exclude all or part of the data obtained from 
any eligible, randomized participants that did not receive at least 90% of the full amount of intended 

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randomized study therapy or are considered to have substantially deviated from the protocol in a 
manner that may impact study outcome or treatment receipt.  The population will also exclude 
individuals who were randomized after 10 weeks, 6 days gestational age.  Participants will be 
grouped by actual treatment received. Treatment receipt reasons and substantial deviations that will 
lead to exclusion include:  
• 
Documented receipt of aspirin while on study outside of assigned study drug as identified by 
a reported protocol deviation 
• 
Not receiving treatment after randomization: All data from these participants will be 
excluded. 
• 
Receipt of less than 90% of planned total doses after randomization where the number of 
planned doses will be calculated as those expected between randomization and the earliest of 
end of pregnancy and 36 0/7 weeks GA irrespective of if study drug administration was 
prematurely discontinued for any reason prior to that point: All data from these participants 
will be excluded. 
Receiving more than the planned total doses of study drug will generally NOT be an exclusion 
criterion for the PP population. Additional exclusion reasons may be identified after completion of 
participant enrollment.  Any participant deviations that will exclude participants or their data from the 
PP analyses will be identified by the study team, prior to review of efficacy and safety data, as a part 
of a masked data review to reduce the opportunity for introducing of bias and will be documented in 
an addendum to this analysis plan.  
6. SAMPLE SIZE DETERMINATION 
The primary outcome of BSID-III score is assumed to have a mean scaled score of 100 and standard 
deviation of 15.47.  Assuming a non-inferiority margin of 4 points is clinically significant, and the 
true effect of LDA is not more than a 1 point decrease, a total sample size of 620 children with a 1:1 
allocation (310 in each group will have 80% power for a one-sided test for non-inferiority with 
α=0.05.  Conditional on non-inferiority being observed, this sample size will also provide over 90% 
power at α=0.05 to detect a different of 4 points between LDA exposed and non-exposed children 
based on a two-sided test for the secondary hypothesis. 
Analyses of secondary outcomes will be exploratory in nature and generally will focus on estimation 
of effect sizes and generation of hypotheses.   
7. STATISTICAL / ANALYTICAL ISSUES 
7.1 General Rules 
All statistical computations will be performed and data summaries will be created using SAS 9.3 or 
higher. If additional statistical packages are required, these will be discussed in the study report. For 
summaries of study data, categorical measures will be summarized in tables listing the frequency and 
the percentage of participants in each study arm; continuous data will be summarized by presenting 
mean, standard deviation, median, minimum, and maximum; and ordinal data will be summarized by 
only presenting median and the limits of the interquartile range. 

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7.2 Adjustments for Covariates  
While this is a cohort study conducted on a subset of participants from a randomized trial, covariates 
may be adjusted for in the analyses if treatment imbalance is present. Covariates for consideration 
include gender, breast feeding, stunting, weight, and family care indicators.  These factors will also be 
explored in secondary analyses for modifying/mediation effects. 
7.3 Missing Data Approaches:  
The primary reason for missing data will be participant withdrew consent or the child aging out.  All 
missing data will be excluded from primary analyses.     
7.4 Multicenter Studies 
For this multicenter study, randomization of study participants was stratified within site.  
Consequently, for all test-based analyses of treatment effect and model-based primary and secondary 
analyses, site will be included as a fixed effect in the models.  As exploratory analyses, analyses, 
particularly those providing estimate of treatment effect, will be conducted by region: Africa, Asia, 
and Guatemala.    
7.5 Multiple Comparisons and Multiplicity 
There is only one formal hypothesis test for this study. As such, a statistical test will be conducted at a 
5% type I error rate (two-sided) for the primary efficacy measure, and no adjustments for multiplicity 
will be made. All analyses of secondary outcomes are exploratory in nature; therefore, p-values and 
confidence intervals are provided for descriptive purposes only. All p-values provided for any 
baseline and demographic characteristics and safety parameters will be for descriptive purposes only. 
As such, unless otherwise specified, p-values presented will be on a per analysis basis, with no further 
control for multiple tests.  
7.6 Masked Data Review 
A masked data review is unlikely to be needed for this study.  Details of any masked data review 
conducted and resulting decisions will be described in a SAP addendum.  
8. STUDY PARTICIPANTS, TREATMENT EXPOSURE AND COMPLIANCE 
8.1 Participant Disposition   
Participant eligibility status will be summarized by study arm and overall disposition of study 
participants will be described using a standard CONSORT diagram.  
8.2 Protocol Deviations 
Any protocol deviations reported will be reviewed to determine if any data need to be excluded from 
analysis. 
8.3 Demographic and Baseline Characteristics 
Demographic and baseline clinical characteristics for the study participants will be summarized by 
treatment group using the general analysis rules describe above for the ITT and PP populations.  

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Variables of interest include the same characteristics as from the main trial: maternal age, maternal 
education, gravidity, maternal weight and BMI, previous pregnancy loss, number of antenatal care 
visits, delivery attendant, delivery location, and type of delivery.  
9. NON-INFERIORITY AND EFFICACY ANALYSES 
9.1 Overview of Efficacy Analysis Methods 
Efficacy analyses will be performed using the ITT population.  Sensitivity analyses will be performed 
using the per-protocol population as detailed below. The data will be summarized by treatment arm, 
overall and by region.  All model-based analyses and test-statistics examining the treatment effect 
will adjust for covariates as specified in sections 7.2 and 7.3. Additional details are presented in the 
sections below. 
9.2 Definition of Analysis Variables 
The following table defines each of the effectiveness and process analysis variables. 
 
 

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Variable 
Type 
Definition 
Primary Outcome  
BSID-III scores 
Continuous 
The unit of analysis is the child enrolled in the study. The three associated outcomes will 
be the mean scaled composite cognitive, language and motor BSID-III scores assessed 
and reported at 36 months of life (+/- 3 months).   The cognitive score will be considered 
primary. 
Raw scores will be summarized but not analyzed. 
Analysis Population: ITT  
Low BSID-III scores 
Binary 
The unit of analysis is the child enrolled in the study. The three associate outcomes will 
be whether the child scored less than an 85 on each of the composite cognitive, language 
and motor BSID-III scaled composite scores assessed at 36 months of life (+/- 3 months). 
Cognitive scores < 54 and motor scores < 45 will have missing associated binary 
outcomes. 
Analysis Population: ITT 
Secondary Outcome 
ASQ-3 scores 
Continuous 
The unit of analysis is the child enrolled in the study.  The 5 associated outcomes will be 
the scores of the components of the ASQ-3 assessment, including communication, gross 
motor, fine motor, problem solving and personal-social. 
 
Analysis Population: ITT 

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Variable 
Type 
Definition 
Low ASQ-3 scores 
Binary 
The unit of analysis is the child enrolled in the study. The outcome will be whether the 
child scored 2 standard deviations less than the mean on each of the communication, 
gross motor, fine motor, problem solving and personal-social ASQ-3 scores assessed at 
36 months of life (+/- 3 months). Established, published cut-offs using US standards will 
be used as there are no region specific cut-offs established for the study sites. 
 
Analysis Population: ITT 
 

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9.4 Primary Analysis 
The primary analysis will compare the BSID-III composite scores between the two treatment arms 
using a non-inferiority test.  For each of the three BSID-III composite scores, a mean difference 
between treatment arms and its associated 90% confidence interval will be calculated.  If the lower 
bound of the 90% confidence interval is greater than -4, then the treatment is assumed to be non-
inferior to placebo.  If non-inferiority is assumed, a one-sided t-test will be run to test the benefit of 
treatment compared to placebo.   All analyses will be modeled based controlling for site as the 
original randomization stratification factor.  For each outcome, the adjusted mean difference between 
treatment groups and the associated 90% confidence interval based on an ANCOVA model will be 
calculated and presented. Secondary models will include region and a region by treatment interaction 
in order to get region specific treatment effect estimates. 
Prior to conducting formal analyses baseline demographic characteristics and key clinical measures 
will be compared between the mothers who participated in the two treatment arms using contingency 
table approaches for categorical variables and analysis of variance models and t-tests for continuous 
variables.  For all of these analyses, comparisons will be made within Global Network sites and 
overall across the sites controlling for site in the models. 
9.5 Secondary Outcome Analyses  
For ASQ and any other continuous outcomes, modeled-based analyses similar to the primary analysis 
will be used to generate estimated treatment effects (i.e., ANCOVA models used to obtain adjusted 
mean differences controlling for site). For binary outcomes, robust Poisson regression models will be 
used controlling for site in order to obtain relative risk estimates and associated confidence intervals.  
All models will be repeated including terms for region and its interaction with treatment to get region 
specific estimates of treatment effect. 
10. SAFETY ANALYSIS:  
No formal safety analyses are planned as a part of this study, although SAEs will be summarized.  No 
formal hypothesis tests will be conducted, but descriptive p-values generated using Cochran-Mantel-
Haenszel statistics with stratification by GN site will be generated.  
11. CHANGES TO THE ANALYSES PLANNED IN THE PROTOCOL 
No changes.  
12. REFERENCES 
Abramovici A, Cantu J, Jenkins SM. Tocolytic Therapy for Acute Preterm Labor. Obstet Gynecol 
Clin North Am. 2012; 39:77-87. doi: 10.1016/j.ogc.2011.12.003.  
Anderson RN, Smith BL. Deaths: leading causes for 2001. Natl Vital Stat Rep. 2003;52:1-85.  
Beck S, Wojdyla D, Say L, et al. The worldwide incidence of preterm birth: A systematic review of 
maternal mortality and morbidity. Bull World Health Organ. 2010;88:31-38.  

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Blencowe H, Cousens S, Oestergaard MZ, et al. National, regional, and worldwide estimates of 
preterm birth rates in the year 2010 with time trends since 1990 for selected countries: a systematic 
analysis and implications. Lancet. 2012;379:2162-2172. doi:10.1016/S0140-6736(12)60820-4. 
Goldenberg R, Rouse D. Prevention of Premature Birth. N Engl J Med. 1998;339(5):313-320.  
Hogan JW, Lancaster T. Instrumental variables and inverse probability weighting for causal inference 
from longitudinal observational studies. Stat Methods Med Res. 2004;13:17-48. 
doi:10.1191/0962280204sm351ra. 
Liang KY and Zeger SL. Longitudinal Data Analysis Using Generalized Linear Models. Biometrika. 
1986:73(1): 13-22. 
March of Dimes, PMNCH, Save the Children, WHO. Born Too Soon: The Global Action Report on 
Preterm Birth. Geneva: World Health Organization; 2012. 
Mathews TJ, Menacker F, MacDorman MF. Infant mortality statistics from the 2002 period: linked 
birth/infant death data set. Natl Vital Stat Rep. 2004;53:1-29 
Maine D, Rosenfield A, McCarthy J, Kamara A, Lucas AO: Safe Motherhood Programs: options and 
issues. New-York: Columbia University; 1991. 
McCormick MC, Richardson DK. Premature infants grow up. N Engl J Med. 2002;346(3):197-198. 
doi:10.1056/NEJM200201173460310. 
McClure EM, Pasha O, Goudar SS, et al; Global Network Investigators. Epidemiology of stillbirth in 
low-middle income countries: a Global Network Study. Acta Obstet Gynecol Scand. 
2011;90(12):1379-85. 
Oestergaard MZ, Inoue M, Yoshida S, Mahanani WR, Gore FM, Cousens S, Lawn JE, Mathers CD; 
United Nations Inter-Agency Group for Child Mortality Estimation and the Child Health 
Epidemiology Reference Group. Neonatal mortality levels for 193 countries in 2009 with trends since 
1990: a systematic analysis of progress, projections, and priorities. PLoS Med. 2011;8(8):e1001080. 
Schisterman EF, Silver RM, Lesher LL, et al. Preconception low-dose aspirin and pregnancy 
outcomes: Results from the EAGeR randomised trial. Lancet. 2014;384:29-36. doi:10.1016/S0140-
6736(14)60157-4. 
Saigal S, Doyle LW. An overview of mortality and sequelae of preterm birth from infancy to 
adulthood. Lancet. 2008;371:261-269. doi:10.1016/S0140-6736(08)60136-1. 
Valero De Bernabé J, Soriano T, Albaladejo R, et al. Risk factors for low birth weight: A review. Eur 
J Obstet Gynecol Reprod Biol. 2004;116:3-15. doi:10.1016/j.ejogrb.2004.03.007. 
14. ATTACHMENTS 
 
Updates to these tables will not require an analysis plan amendment. 
Table 1  
Participant Distribution 

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Table 2  
Covariate Summaries 
Table 3  
BSID-III Outcome Descriptive Statistics 
Table 4  
ASQ Outcome Descriptive Statistics 
Table 5a 
BSID-III Adjusted Mean Estimates 
Table 5b 
BSID-III Adjusted Risk Ratio Estimates 
Table 6a 
ASQ Adjusted Mean Estimates  
Table 6b 
ASQ Adjusted Risk Ratio Estimates 
Figure 1 
Consort Diagram 
 

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Table 1. Participant Distribution  
Variable 
Overall 
DRC 
Zambia 
Belagavi, India 
Nagpur, India 
Pakistan 
Guatemala 
Aspirin 
Placebo 
Aspirin 
Placebo 
Aspirin 
Placebo 
Aspirin 
Placebo 
Aspirin 
Placebo 
Aspirin 
Placebo 
Aspirin 
Placebo 
Screened 
  
  
  
  
  
  
  
  
  
  
  
  
  
  
Ineligible 
  
  
  
  
  
  
  
  
  
  
  
  
  
  
Eligible 
  
  
  
  
  
  
  
  
  
  
  
  
  
  
Enrolled 
  
  
  
  
  
  
  
  
  
  
  
  
  
  
BSIDS III 
completed 
  
  
  
  
  
  
  
  
  
  
  
  
  
  
Within age 
required 
  
  
  
  
  
  
  
  
  
  
  
  
  
  
ASQ 
completed 
  
  
  
  
  
  
  
  
  
  
  
  
  
  
Within age 
required 
  
  
  
  
  
  
  
  
  
  
  
  
  
  
 
 
 

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Table 2: Covariate Summaries 
Variable 
Overall 
DRC 
Zambia 
Belagavi, India 
Nagpur, India 
Pakistan 
Guatemala 
Aspirin 
Placebo 
Aspir
in 
Placebo 
Aspirin 
Placebo 
Aspirin 
Placebo 
Aspirin 
Placebo 
Aspirin 
Placebo 
Aspirin 
Placebo 
Gender, n (%) 
  
  
  
  
  
  
  
  
  
  
  
  
  
  
Male 
  
  
  
  
  
  
  
  
  
  
  
  
  
  
Female 
  
  
  
  
  
  
  
  
  
  
  
  
  
  
Gestational age at 
delivery, n (%) 
  
  
  
  
  
  
  
  
  
  
  
  
  
  
< 37 
  
  
  
  
  
  
  
  
  
  
  
  
  
  
>= 37 
  
  
  
  
  
  
  
  
  
  
  
  
  
  
Breastfed after 
delivery 
  
  
  
  
  
  
  
  
  
  
  
  
  
  
Yes 
  
  
  
  
  
  
  
  
  
  
  
  
  
  
Months 
breastfed, mean 
(sd) 
  
  
  
  
  
  
  
  
  
  
  
  
  
  
No 
  
  
  
  
  
  
  
  
  
  
  
  
  
  
Stunting 
  
  
  
  
  
  
  
  
  
  
  
  
  
  
Yes 
  
  
  
  
  
  
  
  
  
  
  
  
  
  
No 
  
  
  
  
  
  
  
  
  
  
  
  
  
  
Family Care 
Indicators 
  
  
  
  
  
  
  
  
  
  
  
  
  
  
Books, mean (sd) 
  
  
  
  
  
  
  
  
  
  
  
  
  
  
Variety of play 
materials score  
(Q3-Q5), mean (sd) 
  
  
  
  
  
  
  
  
  
  
  
  
  
  
Sources of play 
materials score 
(Q7-Q13), mean 
(sd) 
  
  
  
  
  
  
  
  
  
  
  
  
  
  
Play activitites 
score (Q14-Q23), 
mean (sd) 
  
  
  
  
  
  
  
  
  
  
  
  
  
  
 
 

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Table 3: BSID-III Outcome Descriptive Statistics 
Variable 
Overall 
DRC 
Zambia 
Belagavi, India 
Nagpur, India 
Pakistan 
Aspirin 
Placebo 
Aspirin 
Placebo 
Aspirin 
Placebo 
Aspirin 
Placebo 
Aspirin 
Placebo 
Aspirin 
Placebo 
A
Evaluated on BSID III 
  
  
  
  
  
  
  
  
  
  
  
  
Age at Examination, N 
  
  
  
  
  
  
  
  
  
  
  
  
Mean (SD) 
  
  
  
  
  
  
  
  
  
  
  
  
Median (P25, P75) 
  
  
  
  
  
  
  
  
  
  
  
  
Minimum - Maximum 
  
  
  
  
  
  
  
  
  
  
  
  
Cognitive Test Results 
  
  
  
  
  
  
  
  
  
  
  
  
Language Test Results 
  
  
  
  
  
  
  
  
  
  
  
  
Motor Test Results 
  
  
  
  
  
  
  
  
  
  
  
  
Scaled Scores 
  
  
  
  
  
  
  
  
  
  
  
  
Cognitive Scaled Score, N 
  
  
  
  
  
  
  
  
  
  
  
  
Mean (SD) 
  
  
  
  
  
  
  
  
  
  
  
  
Median (P25, P75) 
  
  
  
  
  
  
  
  
  
  
  
  
Minimum - Maximum 
  
  
  
  
  
  
  
  
  
  
  
  
Receptive Language Scaled Score, N 
  
  
  
  
  
  
  
  
  
  
  
  
Mean (SD) 
  
  
  
  
  
  
  
  
  
  
  
  
Median (P25, P75) 
  
  
  
  
  
  
  
  
  
  
  
  
Minimum - Maximum 
  
  
  
  
  
  
  
  
  
  
  
  
Expressive Language Scaled Score, N 
  
  
  
  
  
  
  
  
  
  
  
  
Mean (SD) 
  
  
  
  
  
  
  
  
  
  
  
  
Median (P25, P75) 
  
  
  
  
  
  
  
  
  
  
  
  
Minimum - Maximum 
  
  
  
  
  
  
  
  
  
  
  
  
Summed Language Scaled Score, N 
  
  
  
  
    
  
  
  
  
  
  
  
Mean (SD) 
  
  
  
  
    
  
  
  
  
  
  
  
Median (P25, P75) 
  
  
  
  
    
  
  
  
  
  
  
  
Minimum - Maximum 
  
  
  
  
    
  
  
  
  
  
  
  
Fine Motor Scaled Score, N 
  
  
  
  
    
  
  
  
  
  
  
  

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Mean (SD) 
  
  
  
  
    
  
  
  
  
  
  
  
Median (P25, P75) 
  
  
  
  
    
  
  
  
  
  
  
  
Minimum - Maximum 
  
  
  
  
    
  
  
  
  
  
  
  
Gross Motor Scaled Score, N 
  
  
  
  
    
  
  
  
  
  
  
  
Mean (SD) 
  
  
  
  
    
  
  
  
  
  
  
  
Median (P25, P75) 
  
  
  
  
    
  
  
  
  
  
  
  
Minimum - Maximum 
  
  
  
  
    
  
  
  
  
  
  
  
Summed Motor Scaled Score, N 
  
  
  
  
    
  
  
  
  
  
  
  
Mean (SD) 
  
  
  
  
    
  
  
  
  
  
  
  
Median (P25, P75) 
  
  
  
  
    
  
  
  
  
  
  
  
Minimum - Maximum 
  
  
  
  
    
  
  
  
  
  
  
  
Composite Scores 
  
  
  
  
    
  
  
  
  
  
  
  
Cognitive Composite Score, N 
  
  
  
  
    
  
  
  
  
  
  
  
Mean (SD) 
  
  
  
  
    
  
  
  
  
  
  
  
Median (P25, P75) 
  
  
  
  
    
  
  
  
  
  
  
  
Minimum - Maximum 
  
  
  
  
    
  
  
  
  
  
  
  
Language Composite Score, N 
  
  
  
  
    
  
  
  
  
  
  
  
Mean (SD) 
  
  
  
  
    
  
  
  
  
  
  
  
Median (P25, P75) 
  
  
  
  
    
  
  
  
  
  
  
  
Minimum - Maximum 
  
  
  
  
    
  
  
  
  
  
  
  
Motor Composite Score, N 
  
  
  
  
    
  
  
  
  
  
  
  
Mean (SD) 
  
  
  
  
    
  
  
  
  
  
  
  
Median (P25, P75) 
  
  
  
  
    
  
  
  
  
  
  
  
Minimum - Maximum 
  
  
  
  
    
  
  
  
  
  
  
  
Low Composite Scores 
  
  
  
  
    
  
  
  
  
  
  
  
Cognitive Composite Score <85 
  
  
  
  
    
  
  
  
  
  
  
  
Language Composite Score <85 
  
  
  
  
    
  
  
  
  
  
  
  
Motor Composite Score <85 
  
  
  
  
    
  
  
  
  
  
  
  
 
 

 Global Network:  Statistical Analysis Plan, ASPIRIN Follow-up Study  
 
 
   
Version 1.0 06/17/2022; Page 22   
 
Table 4: ASQ Outcome Descriptive Statistics 
 
 

 Global Network:  Statistical Analysis Plan, ASPIRIN Follow-up Study  
 
 
   
Version 1.0 06/17/2022; Page 23   
 
Table 5a: BSID-III Adjusted Mean Estimates 
Variable 
Adjusted1 Mean (95% CI) 
Aspirin 
Placebo 
Difference 
BSID III Composite Scores 
  
  
  
Cognitive Composite Score 
  
  
  
Language Composite Score 
  
  
  
Motor Composite Score 
  
  
  
BSID III Scaled Scores 
  
  
  
Cognitive Scaled Score 
  
  
  
Receptive Language Scaled Score 
  
  
  
Expressive Language Scaled Score 
  
  
  
Summed Language Scaled Score 
  
  
  
Fine Motor Scaled Score 
  
  
  
Gross Motor Scaled Score 
  
  
  
Summed Motor Scaled Score 
  
  
  
 
Table 5b: BSID-III Adjusted Risk Ratio Estimates 
  
Aspirin 
n (%) 
Placebo  
n (%) 
RR (95% CI) 
BSID III Composite Scores 
  
  
  
Cognitive Composite Score <85 
  
  
  
Language Composite Score <85 
  
  
  
Motor Composite Score <85 
  
  
  
 
1. Mean estimates and 95% confidence intervals from a regression model adjusting for site, gender, breast feeding, stunting and family care indicators. 
 
 

 Global Network:  Statistical Analysis Plan, ASPIRIN Follow-up Study  
 
 
   
Version 1.0 06/17/2022; Page 24   
 
 
Table 6a: ASQ Adjusted Mean Estimates 
Variable 
Adjusted1 Mean (95% CI) 
Aspirin 
Placebo 
Difference 
ASQ Total Score 
  
  
  
Communication total score 
  
  
  
Gross motor total score 
  
  
  
Fine motor total score 
  
  
  
Problem solving total score 
  
  
  
Personal-social total score 
  
  
  
 
Table 6b: ASQ Adjusted Risk Ratio Estimates 
  
Aspirin 
n (%) 
Placebo  
n (%) 
RR (95% CI) 
ASQ Scores Below Cut Off 
  
  
  
Communication total score < 27.93 
  
  
  
Gross motor total score < 30.68 
  
  
  
Fine motor total score < 35.39 
  
  
  
Problem solving total score < 16.71 
  
  
  
Personal-social total score < 32.53 
  
  
  
1. Mean estimates and 95% confidence intervals from a regression model adjusting for site, gender, breast feeding, stunting and family care indicators. 

 Global Network:  Statistical Analysis Plan, ASPIRIN Follow-up Study  
 
 
   
Version 1.0 06/17/2022; Page 25   
 
Figure 1: Consort Diagram